10th

A N N IV E R SA RY

Program & Abstracts Book

10th ASM Biodefense and Emerging Diseases Research Meeting
The Omni Shoreham Hotel | Washington, DC
February 26 - 29, 2012

www.asmbiodefense.org
 w w w.i ca ac .o rg
www.icaac.org

ICAAC, the premier conference on antimicrobial agents and infectious diseases, showcases the
latest-breaking science and lectures from top researchers from around the globe. Join 10,000 of the
world’s foremost physicians, clinical microbiologists, researchers, pharmacists, and other health care
professionals in San Francisco, California, for timely information that will improve health care and
the management of infectious diseases.

Abstracts
Abstract Submission Closes: May 7, 2012, 11:59 EST
Travel Grant Application Submission Closes: June 29, 2012

Meeting Highlights
Opening Keynote Session | Sunday, September 9, 2012
Emerging New Issues in the Management of Hepatitis C Infection
Charles M. Rice, PhD; The Rockefeller University, New York, NY
The Impact of Emerging New Diagnostic Laboratory Techniques on Clinical Infectious Disease
Franklin R. Cockerill, III, MD; Mayo Clinic, Rochester, MN
Prospects for New Antibiotics
Christopher T. Walsh; Harvard Medical School, Boston, MA

ICAAC Lecture | Monday, September 10, 2012

Fungal Disease in the 21st Century
Arturo Casadevall, MD, PhD; Albert Einstein College of Medicine of Yeshiva University, Bronx, NY
 W E LCO M E LE T T E R

Dear Colleagues,
Welcome to the 10th American Society for Microbiology’s Biodefense and Emerging Diseases Research
Meeting in Washington, DC. The ASM Biodefense Research Program Committee has assembled a pro-
gram that includes experts in multiple relevant fields presenting in several different session formats.
The 2012 program is intended to provide each attendee with the opportunity to participate in educa-
tional programming that has the most value and relevance to their individual professional needs. For
this year’s meeting we have brought together individuals who are carrying out research to defend
against the growing threats of bioterrorism and emerging infectious diseases and decision makers
Paul Keim, Ph.D. shaping the future of the biodefense research agenda.
The meeting opens Sunday at noon with four in-depth Focus Sessions followed by the Keynote Session
Sunday afternoon. This year’s Keynote Speaker, Paul Offit, M.D., from the Children’s Hospital of
Philadelphia, will present “Hard Knocks: Communicating Vaccine Science to the Public.” Dr. Offit is
one of the country’s foremost advocates for vaccines. We cordially invite all meeting attendees to
the Opening Reception in the Exhibit Hall immediately following the conclusion of Dr. Offit’s talk.
Beginning Monday morning, daily Plenary Sessions include “Outbreaks, Response, and Hindsight,”
“Targeting Virulence Factors in Acute Infection,” and “Immunity to Pulmonary Pathogens.” Diverse
concurrent symposia are also scheduled for Monday and Tuesday afternoon. In addition, for the first
time we have added new early morning sessions that will focus on current international security
challenges and policy issues. These will include a panel discussing the recent experiments modifying
Marshall Bloom, M.D.
the transmissibility of H5N1 avian influenza in mammals and the resulting implications for biosecurity
policy. High profile speakers have been recruited for these early morning sessions, including Dr. Bruce
Alberts, Dr. Michael Osterholm, Dr. Anthony Fauci, Dr. Ron A.M. Fouchier, Assistant Secretary Thomas
Countryman, and Dr. Alan Rudolph. Please check the details in this Program and Abstracts book to
identify the presentations that most appeal to you.
From over 180 accepted abstract submissions, the Program Committee has organized 2 Poster
Sessions and 6 Highlighted Oral Abstract Sessions. Poster presentations will be presented in the
Exhibit Hall, Monday and Tuesday afternoons. Poster presenters will be available from 1:00 – 3:00 PM
to answer your questions. The Exhibit Hall also features 36 exhibitors with the latest information on
new products. The Highlighted Oral Abstract Sessions occur Monday and Tuesday from 3:00 – 4:00 PM.
Please check the details in this Program and Abstracts book.
Again, the Program Committee welcomes you and looks forward to meeting you as we continue to
work together in addressing the challenges that lie ahead.

Sincerely,

Paul Keim, Ph.D. Marshall Bloom, M.D.

ASM Biodefense Research Meeting Co-Chair ASM Biodefense Research Meeting Co-Chair

BIODEFENSE PROGRAM & ABSTRACTS 1

 Table of Contents

1 Welcome Letter
2 Program Committee and ASM Officers
10th ASM Biodefense and Emerging 3 Program-at-a-Glance
Diseases Research Committee 4 Daily Schedule-at-a-Glance
5 Program Highlights
Paul Keim, Ph.D., Co-Chair 6 Student Postdoctoral Fellow Travel
Northern Arizona University, Flagstaff, AZ Grant Recipients
Marshall Bloom, M.D., Co-Chair 7 Biodefense Exhibits
NIH/NIAID, Hamilton, MT
8 Poster Presentation Hall Map—Monday
Catharine Bosio, Ph.D. 9 Poster Presentation Hall Map—Tuesday
NIH/NIAID, Hamilton, MT
10 Omni Shoreham Hotel Map
Elisabeth Carniel, M.D., Ph.D. 11 General Information
Institut Pasteur, Paris, France
13 Scientific Program
Miles Carroll, Ph.D.
33 Abstracts
Health Protection Agency Center for Emergency Preparedness & Response,
Wiltshire, United Kingdom 97 Keyword Index
Colleen Jonsson, Ph.D. 101 Author Index
Center for Predictive Medicine for Biodefense and Emerging Infectious
Disease Clinical & Transitional, Louisville, KY
Karl Klose, Ph.D.
University of Texas, San Antonio, TX
Carol Linden, Ph.D.
Office of the Biomedical Advanced Research & Development Authority,
Washington, DC
C. Rick Lyons, M.D.
Colorado State University, Fort Collins, CO ASM Officers
Petra Oyston, Ph.D. David C. Hooper, Ph.D., President
DSTL, Wiltshire, United Kingdom Massachusetts General Hospital, Boston, MA
Jean Patterson, Ph.D. Jeffery F. Miller, M.D., President-Elect
Texas Biomedical Research Institute, San Antonio, TX University of California, Los Angeles, CA
Theodore Pierson, Ph.D. Joseph Campos, Ph.D., Secretary
NIH/NIAID, Bethesda, MD Children's National Medical Center, Washington, DC 
Anne Rimoin, Ph.D. James Tiedje, Ph.D., Treasurer
UCLA School of Public Health, Los Angeles, CA Michigan State University, East Lansing, MI
Leonard Smith, Ph.D.
US Army Medical Research Institute, Fort Detrick, MD ASM Headquarters
American Society for Microbiology
Meetings and Exhibits Department
1752 N Street, NW
Washington, DC 20036-2904
Telephone: (202) 942-9248
Fax: (202) 942-9340
E-mail: biodefense@asmusa.org
Website: www.asmbiodefense.org

ISBN: 978-1-55581-840-1
©2012 American Society for Microbiology

2 10th ASM Biodefense and Emerging Diseases Research Meeting

 P R O G R A M - AT- A - G L A N C E

10th ASM Biodefense and Emerging Diseases Research Meeting

SUNDAY MONDAY TUESDAY WEDNESDAY

February 26 February 27 February 28 February 29

Registration 9:00 AM – 7:00 PM 6:30 AM – 5:00 PM 6:30 AM – 3:30 PM 6:30 AM – 8:30 AM

Focus Sessions 12:00 PM – 2:00 PM

2:15 PM – 4:15 PM

Keynote Session 4:30 PM – 6:00 PM

Global Threats, Collaborative Solutions 7:15 AM – 8:15 AM 7:15 AM – 8:15 AM

H5N1 Research Discussion 7:15 AM – 8:15 AM

Plenary Sessions 8:30 AM – 12:00 PM 8:30 AM – 12:00 PM 8:30 AM – 12:00 PM

Lunch 12:00 PM – 1:00 PM 12:00 PM – 1:00 PM

Exhibits & Poster Hall 6:00 PM – 8:00 PM 12:00 PM – 4:15 PM 12:00 PM – 4:15 PM
(Exhibits Only)

Opening Reception 6:00 PM – 8:00 PM

Poster Sessions 1:00 PM – 3:00 PM 1:00 PM – 3:00 PM

Highlighted Oral Abstract Presentations 3:00 PM – 4:00 PM 3:00 PM – 4:00 PM

Concurrent Symposia 4:15 PM – 6:15 PM 4:15 PM – 6:15 PM

Roundtable Discussion 6:30 PM – 7:30 PM 6:30 PM – 7:30 PM

BIODEFENSE PROGRAM & ABSTRACTS 3

 10th ASM Biodefense and Emerging Diseases Research Meeting
Daily Schedule-at-a-Glance
This section provides an at-a-glance look of the entire Biodefense program. Please reference the page numbers listed
under each session title to view the session’s description, moderators, and speakers.
SUNDAY, February 26
Focus Sessions
12:00 PM - 2:00 PM | See Page 13
001. Animal Models
12:00 PM - 2:00 PM | See Page 13
002. What’s Next? Pathogen
Discovery from the Field to
the Laboratory
2:15 PM - 4:15 PM | See Page 14
003. Burkholderia pseudomallei:
Understanding Pathogenesis
and Immunity to Guide the
Next Generation of Medical
Countermeasures
2:15 PM - 4:15 PM | See Page 14
004. Non-Coding RNAs and
the Regulation of Virulence
Keynote Session
4:30 PM - 6:00 PM | See Page 15
005. Hard Knocks:
Communicating Vaccine
Science to the Public
MONDAY, February 27
Global Threats, Collaborative
Solutions
7:15 AM - 8:15 AM | See Page 15
Plenary Session
8:30 AM - 12:00 PM | See Page 15
007. Outbreaks, Response, and
Hindsight
Poster Session
1:00 PM - 3:00 PM | See Page 16
008. Monday Poster Session
4 10th ASM Biodefense and Emerging Diseases Research Meeting
Highlighted Oral Abstract
Presentations
3:00 PM - 4:00 PM | See Page 20
009. Environmental Detection
010. Viral Agents
011. Bacterial Agents
Symposia
4:15 PM - 6:15 PM | See Page 21
012. Biodefense at the Dinner
Table: Emerging Foodborne
Illnesses
4:15 PM - 6:15 PM | See Page 21
013. Persistence and Inactivation
of Biothreat Agents and
Emerging Pathogens
4:15 PM - 6:15 PM | See Page 22
014. Taking the Bite Out of
Insect Bites: Biodefense
Implications of Vector-Borne
Infections
Roundtable Discussion
6:30 PM - 7:30 PM | See Page 22
015. Recapping the 2011
Biological Weapons Convention
Review Conference (BWC
RevCon)
TUESDAY, February 28
Global Threats, Collaborative Solutions
7:15 AM - 8:15 AM | See Page 23
Plenary Session
8:30 AM - 12:00 PM | See Page 23
017. Targeting Virulence Factors
in Acute Infection
Poster Session
1:00 PM - 3:00 PM | See Page 23
018. Tuesday Poster Session
Highlighted Oral Abstract
Presentations
3:00 PM - 4:00 PM | See Page 27
019. Immune Response
020. Therapeutics
021. Diagnostics and Vaccines
Symposia
4:15 PM - 6:15 PM | See Page 28
022. Host Genetic Resistance
to Infections
4:15 PM - 6:15 PM | See Page 29
023. Dynamic Pathogens Make
Difficult Targets
4:15 PM - 6:15 PM | See Page 29
024. Visual Discovery of
Microbe-Host Interactions
Roundtable Discussion
6:30 PM - 7:30 PM | See Page 30
025. Global Status of Strategies
for Addressing Intersection of
Science and Security
WEDNESDAY, February 29
H5N1 Research Discussion
7:15 AM – 8:15 AM | See Page 31
Plenary Session
8:30 AM - 12:00 PM | See Page 31
027. Immunity to Pulmonary
Pathogens
Program Highlights

Keynote Session
Sunday, February 26, 4:30 PM – 6:00 PM | Regency Ballroom
Hard Knocks: Communicating Vaccine Science to the Public

PAUL A. OFFIT, M.D., The Children’s Hosp. of Philadelphia, Philadelphia, PA

A major and critical goal in the area of biodefense and emerging diseases is the development
of safe and effective vaccines. However, due to often deliberate misinformation about vaccine
complications and poor news reportage, public acceptance of even currently available vaccines
is waning. If new vaccines are to be effectively deployed, it is critical that the biomedical and
research community be cognizant of the hurdles facing vaccination science. Dr. Offit is an
acknowledged expert in the area of vaccine science and one of the foremost advocates for
vaccines in the United States. He has written books and articles about vaccine science and
the controversies about vaccines, and has appeared numerous times in the public media. As a result, his keynote
lecture will provide timely and important information for anyone interested in the topic of vaccines.

Global Threats, Collaborative Solutions

Monday, February 27, 7:15 AM – 8:15 AM | Palladian Ballroom
Assistant Secretary Thomas Countryman, U.S. Department of State Bureau of International
Security and Nonproliferation, Washinton, DC
Assistant Secretary Thomas Countryman will share his vision for U.S. efforts to prevent the spread
of biological weapons, through cooperative engagement, international treaties and diplomacy.

Tuesday, February 28, 7:15 AM - 8:15 AM | Palladian Ballroom

Dr. Alan Rudolph, Director for Chemical and Biological Technologies Directorate, Research and
Development Enterprise, Defense Threat Reduction Agency (DTRA), Fort Belvoir,VA
Dr. Alan Rudolph will provide a unique look at DoD’s efforts to safeguard America and its allies
from weapons of mass destruction, drawing from his experiences in driving innovative technolo-
gies to reduce, eliminate, and counter biological threats.

H5N1 Research Discussion

Wednesday, February 29, 7:15 AM – 8:15 AM | Palladian Ballroom
Moderated by the Chair of the National Science Advisory Board for Biosecurity (NSABB),
Paul Keim, Ph.D., this newly added session at the ASM Biodefense and Emerging Diseases
Research Meeting will include discussions of the NSABB’s recommendations for the publication
of the controversial H5N1 research. This session will also provide an open forum for attendees to
give their feedback on such policy issues as the appropriate mechanisms to allay public concerns
about the safety of dual use research.
continued on next page...

BIODEFENSE PROGRAM & ABSTRACTS 5

PROGRAM H IG H LIG HTS
Presentations:
NSABB Recommendations
Michael T. Osterholm, Ph.D., MPH
University of Minnesota School of Public
Health, Minneapolis, MN
Director, Center for Infectious Disease
Research and Policy (CIDRAP)
Government Response to the
Recommendations
Anthony S. Fauci, M.D.
Director, National Institute of Allergy and
Infectious Diseases (NIAID)
Science’s Response to the Situation
Bruce Alberts, Ph.D.
Editor-in-Chief of Science
Perspective from an Investigator
Ron A.M. Fouchier, Ph.D.
Erasmus MC, Rotterdam, Netherlands
Student Postdoctoral Fellow
Travel Grant Recipients
Congratulations to our 10th ASM Biodefense and
Emerging Diseases Research Meeting Student
Postdoctoral Fellow Travel Grant Recipients!
The ASM Student Travel Grant Program was
established to support the travel of students to ASM
meetings. Recipients are selected by the Biodefense
Research Program Committee on the basis of the
scientific quality of the submitted abstracts.
6 10th ASM Biodefense and Emerging Diseases Research Meeting
Live Video Stream
H5N1 Research Discussion
Wednesday, February 29 | 7:15 AM – 8:15 AM
ASM will be hosting a free live video stream of the
H5N1 Research Discussion taking place on February 29,
2012 at that Biodefense Meeting. We encourage you
to share this information with your colleagues that
were unable to attend. The link will be available on
this meeting’s website the morning of the session.
If you or your colleagues are unable to participate
in the live event/video stream, you may access the
archived footage of the session that will be posted
to the website at 1:00 PM on Wednesday.
www.asmbiodefense.org
Andrew Clohessey
Rowan University, Glassboro, NJ
Session 008; Presentation 102 (J)
Susann Dupke
Robert Koch Institute, Berlin, Germany
Session 008; Presentation 042 (B)
Manoshi Gayen
Uniformed Services, University of Health Sciences, Bethesda, MD
Session 018; Presentation 199 (H)
Yanyan Li
University of Maryland, Baltimore, MD
Session 011; Presentation 116
Kevin McCormick
University of Pittsburgh, Pittsburgh, PA
Session 008; Presentation 031 (A)
Emilio Ortiz Riano
University of Rochester Medical Center, Rochester, NY
Session 018; Presentation 144 (A)
 E X H I B I T S I N F O R M AT I O N

Biodefense Exhibits Information

Exhibit Hall Information
Scientific exhibits are located in the Exhibit Hall, Level 1B,
co-located with poster sessions.
The Exhibit Hall will be open:
Sunday, February 26 ...................6:00 PM – 8:00 PM
Monday, February 27..................12:00 PM – 4:15 PM
Tuesday, February 28 .................12:00 PM – 4:15 PM
Additionally, lunch will be provided in the Exhibit Hall from
12:00 PM – 1:00 PM on Monday and Tuesday. Attendees can lis-
ten to poster presentations, then get the most current informa-
tion on scientific products and services relating to Biodefense.
Opening Reception in the Exhibit Hall
Sunday, February 26 | 6:00 PM – 8:00 PM
Network with colleagues, investigate the amazing variety of
exhibits, and visit with industry representatives at your leisure,
all while enjoying light hors d’oeuvres and drinks.
Name badges required.
Exhibits, posters, lunch and the Opening Reception
will be held in the Exhibit Hall.

The American Society for Microbiology welcomes the following exhibitors:

A2LA American Association for emka Technologies inc Reagent Proteins

Laboratory Accreditation
Booth 16 | www.A2LA.org
Advanced Instruments, Inc.
Booth 24 | www.aicompanies.com
ASM Press
Booth 21 | estore.asm.org
BEI Resources
Booth 40 | www.atcc.org
Bertin Technologies/Bertin Corp.
Booth 1 | www.bertin.fr
Biaera Technologies, LLC
Booth 42 | www.biaera.com
Bio-Synthesis, Inc.
Booth 37 | www.biosyn.com
Center for Predictive Medicine,
University of Louisville
Booth 19 | www.louisville.edu/predictivemedicine
Certek
Booth 28 | www.certekinc.com
CH Technologies (USA) Inc.
Booth 41 | www.inhalation.org
Data Sciences International
Booth 30 | www.datasci.com
Defense Threat Reduction Agency
Booth 18 | www.dtra.mil
Division of Select Agents & Toxins, CDC
Booth 14 | www.selectagents.gov
DynPort Vaccine Company LLC
Booth 38 | www.csc.com/dvc
Booth 43 | www.emkatech.com Booth 36 | www.reagentproteins.com
Germfree Labs, Inc. Southern Research
Booth 45 | www.germfree.com Booth 25 | www.southernresearch.org
Idaho Technology, Inc. Tetracore, Inc.
Booth 20 | www.bio-surveillance.com Booth 39 | www.tetracore.com
IIT Research Institute The Howard T. Ricketts Regional
Booth 29 | www.iitri.org Biocontainment Laboratory
Booth 26 | www.ntrl.uchicago.edu
InnovaPrep
Booth 35 | innovaprep.com UTMB-National Biocontainment
Training Center
(JPM-TMT) Joint Project Manager
Booth 22 | www.utmb.edu/nbtc
Transformational Medical Technologies
Booth 44 | www.jpmtmt.mil United States Army Healthcare Recruiting
Booth 17 | www.goarmy.com
List Biological Laboratories, Inc
Booth 31 | www.listlabs.com
Lovelace Respiratory Research Institute
Booth 32 | www.LRRI.org
Meso Scale Defense
Booth 46 | www.mesoscale.com
Nanologix
Booth 27 | www.nanologix.com
National BioSafety and Biocontainment
Training Program
Booth 2 | nbbtp.org
National Counterproliferation Center (NCP)
Booth 33 | www.phe.gov/s3
NIH Office of Biotechnology Activities
Booth 15 | www.oba.od.nih.gov
OpGen, Inc
Booth 47 | www.OpGen.com

BIODEFENSE PROGRAM & ABSTRACTS 7

E X H I B I T S I N F O R M AT I O N

Exhibits and Posters

Monday, February 27, 2012

Monday Poster Key

Category Presentation Number
A 027 (A) - 033 (A)
B 034 (B) - 061 (B)
D 062 (D) - 069 (D)
E 070 (E) - 075 (E)
F 076 (F) - 079 (F)
G 080 (G) - 083 (G)
H 084 (H) - 093 (H)
I 094 (I) - 098 (I)
J 099 (J) - 102 (J)
K 103 (K) - 107 (K)

8 10th ASM Biodefense and Emerging Diseases Research Meeting

 E X H I B I T S I N F O R M AT I O N

Exhibits and Posters

Tuesday, February 28, 2012

Exhibits and Posters

Tuesday, February 28, 2012

Tuesday Poster Key

Category Presentation Number
A 140 (A) - 146 (A)
B 147 (B) - 174 (B)
D 175 (D) - 180 (D)
E 181 (E) - 186 (E)
F 187 (F) - 191 (F)
G 192 (G) - 196 (G)
H 197 (H) - 209 (H)
I 210 (I) - 214 (I)
J 215 (J) - 216 (J)
K 217 (K) - 220 (K)

BIODEFENSE PROGRAM & ABSTRACTS 9

 LOWER LEVEL
2B
Health
Veranda Empire Club
Room

10th ASM Biodefense and Emerging Diseases Research Meeting

Robert’s
Private
Robert’s Restaurant Dining Palladian Diplomat
WEST LOBBY Room Room Room
Cabinet Room
Congressional A&B Room
Council Room Sales Conference
Empire Foyer
Executive Room Room
Forum Room
Sales Conference Room
Senate Room
West Registration Bird Cage Walk
Women’s Lounge ADA Lift to
(Lower Level)
BALLROOMS (West Lobby) Men’s
Ambassador and
Clothing Executive Regency Ball Rooms
Ambassador Ballroom ADA Elevator Store Room Telephones
Diplomat Room to Roberts Restaurant Committee Level 1B
Director’s Room
Empire Room and Palladian Room WEST LOBBY West Room
Elevators
Palladian Room Coat Check Stairs
Regency Room AM
BA
MAIN LOBBY Jewelry Men’s Business West BAL SSAD
Store
Restroom Center Registration LRO OR
Women’s Men’s
OM
Telephones Restroom Restroom
(Lower Level)
Front Desk Reg

Congressional
Council

Marquee Lounge
& Reception
Room Senate
Room
ency G

Room
alle
ry
Forum
Room
O M N I S H O R E H A M H OT E L

West
Conference
Cabinet Center
Room REG
ENC
Y BAL
LRO
OM
MAIN ENTRANCE
ADA Ramp For Access to Diplomat ballrooms
To Lobby Please use elevators on the West Side
and go to level 1B.
For Access to the Empire Ballroom
OMNI SHOREHAM HOTEL
and Health Club/Outdoor Pool
Please use elevators on the West Side
of the Hotel and go to level 2B.

10

General Information
Badge Policies
For security reasons, all attendees are required to wear a badge
to attend sessions or enter the Exhibit Hall. Please note that
badges are the property of ASM and must be relinquished at
any time at the request of ASM staff or security.
Attendee Badges
■ Access into all scientific sessions
■ Access to the Exhibit Hall and Poster Sessions during
operating hours
Guest Badges
■ Access to the Exhibit Hall during operating hours
■ Access to the Opening Reception and lunch on Monday
and Tuesday
Attendees wearing guest badges will not be permitted into scientific
session rooms.
Exhibitor Badges
■ Entrance into the Exhibit Hall during installation and
dismantling hours
■ Entrance into the Hall one hour before the Hall opens
and one hour after it closes
Attendees wearing exhibitor badges will not be permitted into
scientific session rooms.
Business Center
A full-service business center is located in the Lobby Level
of the Omni Shoreham Hotel, and is available 24 hours.
Cameras and Recording
Digital recorders, cameras (including camera phones), and video
cameras (including video phones) are prohibited in the poster
hall and session rooms. Anyone found photographing, videotap-
ing or recording in the prohibited areas will be asked to surren-
der their badge immediately and leave the conference. No
refund will be provided. This rule is strictly enforced.
G E N E R A L I N F O R M AT I O N
Certificate of Attendance
Certificates of Attendance are available at the Registration Desk
during registration hours beginning Tuesday, February 28th.
Certificates of Attendance do not list session information.
Child Safety
Children are not permitted into session rooms. Also, children
under the age of 16 are not permitted on the Exhibit Hall
floor.
Contact Information
To contact ASM staff on site at the conference, please call
the Omni Shoreham Hotel, at (202) 234-0700, and ask
to be transferred to ASM Headquarters located in the
Director’s Room.
Food and Beverage
There are many dining options within walking distance of the
Omni Shoreham Hotel. A list of restaurants can be obtained
from the hotel concierge.
Please Note: Lunch will be provided Monday, February 27th,
and Tuesday, February 28th from 12:00 PM – 1:00 PM in the
Exhibit Hall, Level 1B
Lost and Found
Unattended personal belongings will be removed and taken
to hotel security.
Meeting Announcements
A bulletin board will be available in the West Lobby
registration area for those attendees wishing to post one-page
flyers announcing upcoming meetings.
Messages
All mail and communications to meeting registrants should
be directed to the individual at the hotel where he or she is
staying. The phone number for the Omni Shoreham Hotel
is (202) 234-0700.
BIODEFENSE PROGRAM & ABSTRACTS 11
G E N E R A L I N F O R M AT I O N
ASM Press
ASM Press, the book publishing division of the
American Society for Microbiology, will be exhibiting
its broad selection of texts and references in molecular
biology and the microbiological sciences. Be sure to
shop during the meeting to receive special “meeting
only” discount.
Registration
Registration will be located at the West Registration Desk from
Sunday, February 26 – Wednesday, February 29.
Registration Hours:
Sunday, February 26 . . . . . . . . . . . 9:00 AM - 7:00 PM
Monday, February 27. . . . . . . . . . . 6:30 AM – 5:00 PM
Tuesday, February 28 . . . . . . . . . . . 6:30 AM – 3:30 PM
Wednesday, February 29. . . . . . . . . 6:30 AM – 8:30 AM
Smoking Policy
Smoking is prohibited in all areas of the Omni Shoreham Hotel.
Speaker Ready Room
The Speaker Ready Room will be located in the Director’s
Room on the Lobby Level.
Speaker Ready Room Hours:
Sunday, February 26 . . . . . . . . . . . 9:00 AM - 4:30 PM
Monday, February 27. . . . . . . . . . . 6:30 AM - 6:30 PM
Tuesday, February 28 . . . . . . . . . . . 6:30 AM - 6:30 PM
Wednesday, February 29. . . . . . . . . 6:30 AM -12:00 PM
Oral presenters are requested to bring their PowerPoint presen-
tations on a "thumb drive" or CD-ROM to the Speaker Ready
Room at least one hour prior to their session to be uploaded. A
technician will be available to help ensure your presentation is
properly uploaded.
Special Needs
Please contact an ASM staff member if you require special
accommodations while you attend the meeting.
12 10th ASM Biodefense and Emerging Diseases Research Meeting
Symposia
Symposia consist of 4-5 faculty, invited by the Biodefense
Research Program Committee, who present lectures on a
specific topic followed by a moderated Q&A period.
Poster Sessions
All accepted abstracts are scheduled for poster presentation
based on the December 2011 review of all abstract submis-
sions by the Biodefense Research Program Committee.
Posters presentations will take place in the Exhibit Hall
Monday, February 27th, and Tuesday, February 28th. Each
presenter will be available at his or her poster to answer
questions from 1:00 PM – 3:00 PM either Monday or
Tuesday as scheduled in this Program and Abstracts book.
Focus Sessions
Focus sessions are in-depth educational activities that highlight
a specific topic area. These sessions precede the Keynote session
and are an opportunity for attendees to explore new and
emerging issues with a group of faculty experts.
Plenary Sessions
Daily Plenary Sessions will be held Monday, Tuesday, and
Wednesday mornings from 8:30 AM to 12:00 PM. These
sessions provide all attendees an opportunity to collaborate on
presentations of science, policy, and public health issues. The
speakers were invited by the Biodefense Program Committee
based on their professional expertise and achievement. Plenary
Sessions may have a moderated Q&A period.
Highlighted Oral Abstract Sessions
These sessions highlight research submitted for the ASM
Biodefense and Emerging Diseases Research Meeting. Authors
are invited by topic to give a 10 to 15 minute presentation of
their research followed by a moderated Q&A period.
For specific presentation times and additional details, please
see the session information listed by date in this
Program and Abstracts book.
Roundtable Discussion Sessions
One hour sessions dedicated to providing attendees with infor-
mation about selected government programs to include how to
apply for grants and other financial support.
 SCIENTIFIC PROGRAM

Sunday, February 26, 2012 Focus Session

Focus Session
001. Animal Models
Sunday, February 26, 2012 | 12:00PM – 2:00PM
Diplomat Ballroom
Animal studies are the cornerstone of obtaining regulatory approval
for products to protect against biodefense agents and emerging
threats. To date, guidance from the FDA regarding these studies has
been minimal and sometimes contradictory. To bring a single product
to point where it can move forward for regulatory approval requires
the cooperation of a large community, to include government agen-
cies, contract research organizations and the biodefense pharma-
ceutical industry. Animal studies have to be designed and executed in
a manner that minimizes animal use and distress, while allowing for
studies to be scientifically valid and comparable across research sites.
In order for data to be comparable between laboratories, clear well-
defined ethical study endpoints have to be established for each in-
fectious agent in each appropriate animal model. This requires an
early understanding, During initial model development, of what re-
quirements will be needed to evaluate efficacy of medical counter-
measures.
MODERATORS:
Louise M. Pitt; USAMRIID, Frederick, MD.
Catharine Bosio; NIAID/Rocky Mountain Lab., Hamilton, MT.
PRESENTATIONS:
12:00PM
001 Introduction and Setting the Stage
Louise M. Pitt; USAMRIID, Frederick, MD.
12:20PM
002 Process and Implications of Animal Model Qualification
Rosemary Roberts; CDER, FDA, Silver Spring, MD.
002. What’s Next? Pathogen Discovery from the
Field to the Laboratory
Sunday, February 26, 2012 | 12:00PM – 2:00PM
Palladian Ballroom
Pathogen discovery is at the front line of predicting and identifying
emerging infections. Here we will cover pathogen discovery from sam-
pling and surveillance in the field, through development and applica-
tion of tools to interrogate samples, and descriptions of the discovery
of novel pathogens. The viewers will learn cutting edge technology of
viral discovery and where the field work is headed.
MODERATORS:
Jean Patterson; Texas Biomedical Res. Inst., San Antonio, TX.
Anthony Griffiths; Texas Biomedical Res. Inst., San Antonio, TX.
PRESENTATIONS:
12:00PM
007 Early Detection of Novel Biothreats Through International
Scientific Engagement and Medical Diplomacy
Joseph Fair; Global Viral Forecasting Initiative, San Francisco, CA.
12:30PM
008 Viral Discovery Using Pan-Pathogen Microarrays and Deep
Sequencing
Charles Chiu; Univ. of California Sch. of Med., San Francisco, CA.
1:00PM
009 Discovery and Characterization of Novel Viruses
David Wang; Washington Univ. Sch. of Med., St. Louis, MO.
1:30PM
010 Using Targeted Wildlife Surveillance to Predict Emerging
Zoonoses
Jonathan H. Epstein; EcoHlth. Alliance, New York, NY.

12:40PM
003 Issues in Animal Model Harmonization
Judith Hewitt; DMID/NIAID, Bethesda, MD.

1:00PM
004 Evaluating Study Endpoints and Euthanasia Considerations
for Animal Protocols
Estella Jones; FDA, Silver Spring, MD.

1:20PM
005 Exploring Approaches to Consistent Veterinary Care and
Euthanasia Criteria
Jim Swearengen; NBACC, Fort Detrick, MD.

1:40PM
006 Animal Rule Requirements vs. Expectations: How Can the
Developers of Medical Countermeasures Succeed?
Elizabeth Leffel; PharmAthene, Inc., Annapolis, MD.

BIODEFENSE PROGRAM & ABSTRACTS 13

S U N DAY S C I E N T I F I C P RO G R A M
Focus Session
003. Burkholderia pseudomallei: Understanding
Pathogenesis and Immunity to Guide the Next
Generation of Medical Countermeasures
Sunday, February 26, 2012 | 2:15 PM - 4:15 PM
Palladian Ballroom
This session will emphasize the tie from basic research into the biol-
ogy of a microbial pathogen to the development of medical counter-
measures against it. B. pseudomallei is particularly problematic in
that it is genetically diverse, there is a limited understanding of its
pathogenesis, the current therapeutic strategies are quite extensive,
and there are no viable vaccine candidates. Therefore, this session
will focus on the recent findings in Burkholderia genomics and how
this microbe interacts with its host, as well as how this information
could be used to develop novel countermeasures.
MODERATORS:
Daniel Wolfe; Defense Threat Reduction Agency, Fort Belvoir, VA.
Paul Keim; Northern Arizona Univ., Flagstaff, AZ.
PRESENTATIONS:
2:15PM
011 Melioidosis: Clinical and Therapeutic Issues
Bart J. Currie; Royal Darwin Hosp., Casuarina, Australia.
2:45PM
012 Population Genetics and Evolution of Burkholderia
pseudomallei
David Wagner; Northern Arizona Univ., Flagstaff, AZ.
3:15PM
013 Elucidation of Type 6 Secretion System as a Critical Virulence
Determinant of Burkholderia pseudomallei
David Deshazer; USAMRIID, Fort Detrick, MD.
3:45PM
014 Burkholderia pseudomallei Antibiotic Resistance
Mechanisms: Impact on Therapeutic Strategies
Herbert P. Schweizer; Colorado State Univ., Fort Collins, CO.
14 10th ASM Biodefense and Emerging Diseases Research Meeting
Focus Session
004. Non-Coding RNAs and the Regulation of
Virulence
Sunday, February 26, 2012 | 2:15 PM - 4:15 PM
Diplomat Ballroom
Non-coding RNAs are gaining increased attention as important mole-
cules involved in bacterial gene regulation, particularly of those genes
involved in virulence. Topics to be discussed in this session include
small RNA identification in bacterial pathogens, the regulatory mech-
anisms exerted by noncoding RNAs and their chaperones, the evolu-
tion of pathogens as mediated through changes in small RNA content
or expression, and the contributions to virulence of non-coding RNAs
in biodefense in emerging infectious diseases.
MODERATOR:
Wyndham Lathem; Northwestern Univ., Chicago, IL.
PRESENTATIONS:
2:15PM
015 Small RNAs and the Evolution of Yersinia Species
Wyndham W. Lathem; Northwestern Univ., Chicago, IL.
2:40PM
016 sRNA Regulation of EHEC Virulence
Vanessa Sperandio; Univ. of Texas Southwestern Med. Ctr., Dallas, TX.
3:05PM
017 Regulation of Francisella tularensis Virulence by Small Non-
Coding RNAs
John Gunn; Ohio State Univ., Columbus, OH.
3:30PM
018 Legionella pneumophila Small Regulatory RNAs and
Intracellular Multiplication
Sebastien P. Faucher; McGill Univ., Ste-Anne-de-Bellevue, Canada.
3:55PM
019 A Unique Intergenic RNA Thermosensor Controls Yersinia
Virulence
Petra Dersch; Helmholtz Ctr. for Infection Res., Braunschweig, Germany.
 S U N DAY / M O N DAY S C I E N T I F I C P RO G R A M
Keynote Session
005. Keynote Session
Sunday, February 26, 2012 | 4:30 PM – 6:00PM
Regency Ballroom
A major and critical goal in the area of biodefense and emerging dis-
eases is the development of safe and effective vaccines. However, due
to often deliberate misinformation about vaccine complications and
poor news reportage, public acceptance of even currently available
vaccines is waning. If new vaccines are to be effectively deployed, it
is critical that the biomedical and research community be cognizant of
the hurdles facing vaccination science. Dr. Offit is an acknowledged
expert in the area of vaccine science and one of the foremost advo-
cates for vaccines in the United States. He has written books and ar-
ticles about vaccine science and the controversies about vaccines,
and has appeared numerous times in the public media. As a result,
his keynote lecture will provide timely and important information for
anyone interested in the topic of vaccines.
MODERATORS:
Marshall E. Bloom; DIR/NIAID/NIH, Hamilton, MT.
Paul S. Keim; Northern Arizona Univ., Flagstaff, AZ.
PRESENTATION:
4:30PM
020 Hard Knocks: Communicating Vaccine Science to the Public
Paul A. Offit; The Children’s Hosp. of Philadelphia, Philadelphia, PA.
Monday, February 27, 2012
006. Global Threats, Collaborative
Solutions
Monday, February 27, 2012 | 7:15 AM – 8:15AM
Palladian Ballroom
This new program will provide insights into current interna-
tional security challenges from notable thought leaders, pol-
icy advisors, and senior government officials.
MODERATOR:
Dr. Jason Rao; ASM Director of International Affairs, Washington, DC.
PRESENTATION:
Assistant Secretary Thomas Countryman; U.S. Department of State
Bureau of International Security and Nonproliferation, Washington, DC.
Plenary Session
007. Outbreaks, Response, and Hindsight
Monday, February 27, 2012 | 8:30 AM – 12:00PM
Regency Ballroom
The session is designed to provide an insight into outbreaks of
infectious disease, how they occur, how agencies responded
and what we learned about them to prevent recurrence. By cov-
ering both viral and bacterial pathogens associated with recent
outbreaks of disease this will highlight how effective current
regulations and health frameworks are in different countries.
The session will also demonstrate the impact of new technol-
ogy in characterizing pathogens. Finally, since these are recent
outbreaks, this session will provide useful scientific informa-
tion from those with first-hand experience for the first time to
the audience and whether we are improving in our ability to
deal with such incidents.
MODERATORS:
Miles Carroll; Hlth. Protection Agency, Salisbury, United Kingdom.
David L. Heymann; Hlth. Protection Agency, London, United Kingdom.
8:30AM
021 Outbreaks: A Global Perspective
David L. Heymann; Hlth. Protection Agency, London, United Kingdom.
BIODEFENSE PROGRAM & ABSTRACTS 15
MO N DAY S C I E N T I F I C P RO G R A M

9:00AM 032 (A) Predicting the Timing of Peak Pandemic Influenza

022 Swine Flu Emergence in Mexico Activity: ED Volume, School Absenteeism, and Community
Reported ILI
Edgar E. Sevilla-Reyes; Natl. Inst. of Respiratory Diseases, Mexico City,
Mexico. T. Bell1, M. Richardson2; 1West Texas Influenza Ctr., TTUHSC, Amarillo, TX,
2
Amarillo Publ. Hlth. Dept., Amarillo, TX.
9:30AM
033 (A) The Role of Rare Variants in Coronavirus Cross-Species
023 Swine Flu in the United Kingdom Transmission Events
Gail Thomson; Hlth. Protection Agency, Salisbury, United Kingdom. M. Borucki, J. Allen, H. Harris-Chen; Lawrence Livermore Natl. Lab, Liver-
10:00AM more, CA.

Coffee Break 034 (B) Transient Bacterial Lipopolysaccharide-Induced

10:30AM Resistance to Inhalational Bacillus anthracis Infection in the New
Zealand White Rabbit Model
024 Overview of E. coli Outbreaks in the USA
S. B. Yee, D. N. Dyer, N. A. Twenhafel, M. L. M. Pitt; USAMRIID, Fort Detrick,
Robert Tauxe; CDC, Atlanta, GA. MD.
11:00AM
035 (B) Small Quinolone Resistance Plasmids: A Model for
025 E. coli O104:H4: The Outbreak Experience from Germany Evolution Mediated by Site-Specific Recombination (SSR) at oriT
Reinhard F. Burger; Robert Koch Inst., Berlin, Germany. and Xer Sites
T. Tran1,2, P. Andres3, A. Petroni3, A. Soler-Bistué4, E. Albornoz3,
11:30AM
A. Zorreguieta4, R. Reyes-Lamothe2, D. Sherratt2, A. Corso3, M. Tolmasky1,2;
026 Molecular Epidemiology and Characterization of the E. coli 1
California State Univ., Fullerton, CA, 2Univ. of Oxford, Oxford, United King-
O104:H4 Outbreak Strain dom, 3INEI-ANLIS, Buenos Aires, Argentina, 4Fundación Inst. Leloir, Buenos
Alexander Mellmann; Univ. Hosp. Muenster, Muenster, Germany. Aires, Argentina.

036 (B) Antibacterial Activity of Alpha-MSH Alone and in

Poster Session
008. Monday Poster Session
Monday, February 27, 2012 | 1:00PM – 3:00PM
Exhibit Hall
PRESENTATIONS:
027 (A) Viral Acute Respiratory Tract Infections in Non-Winter
Time
M. EL-BARRAWY; Alexandria Univ., Alexandria, Egypt.
028 (A) Laboratory Surveillance for Non-Influenza Respiratory
Viruses in Maryland
W. A. Murtaugh1,2, J. Tan1, H. Peters2, B. Healey2, M. Tran2, M. Ziese2, B. An-
derson2, M. P. Carlos1,2; 1Johns Hopkins Univ., Baltimore, MD, 2Lab. Admin.
Maryland Dept. of Hlth. and Mental Hygiene, Baltimore, MD.
029 (A) Natural History Studies for Ebola Virus Zaire Strain in
Hartley Guinea Pigs
A. Nalca, H. Bloomfield, S. Frederick, R. Baker, R. Williams, J. Yeager, E.
Zumbrun; USAMRIID, Frederick, MD.
030 (A) Experimental Respiratory Marburg Angola Virus Infection
in Hartley Guinea Pigs
A. Nalca, H. Bloomfield, S. Frederick, R. Baker, R. Williams, J. Yeager, E.
Zumbrun; USAMRIID, Frederick, MD.
031 (A) A Cytopathic Effect-Based High-Throughput Screening
Assay Identified Four Novel Compounds that Inhibit Dengue
Infection
K. D. McCormick; Univ. of Pittsburgh, Pittsburgh, PA.
Combination with Other Antibiotics against Methicillin-Resistant
Staphylococcuus aureus
M. Singh, K. Mukhopadhyay; Jawaharlal Nehru Univ., New Delhi, India.
037 (B) Enhanced Aminoglycoside Sensitivity of Ribonuclease
Mutants of Escherichia coli
A. D. Frazier, W. S. Champney; East Tennessee State Univ., Johnson City, TN.
038 (B) A Model of Inhalation Glanders in African Green Monkeys
J. Yeager, P. Facemire, D. Nyakiti, L. Pitt; USAMRIID, Frederick, MD.
039 (B) Genes Required for the Trafficking of S-Layer Proteins in
the Envelope of Bacillus anthracis
S-M. Nguyen-Mau1,2, D. Missiakas1,2, O. Schneewind1,2; 1Univ. of Chicago,
Chicago, IL, 2Howard Taylor Ricketts Lab., Argonne, IL.
040 (B) Development of a Selective Enrichment Media for
Brucella Species Isolation
M. Perry, A. Kidney, M. Peck, C. Kelly-Cirino, C. Egan; NYS Dept. of Hlth., Al-
bany, NY.
041 (B) Periplasmic YopR Promotes Type III Needle
Polymerization
B. Blaylock, O. Schneewind; Univ. of Chicago, Chicago, IL.
042 (B) Virulence Analysis of B. cereus biovar anthracis
S. Dupke1, T. Franz1, A. Lander1, R. Grunow1, P. L. Goossens2, C. Brezillon2,
S. R. Klee1; 1Robert Koch Inst., Berlin, Germany, 2Inst. Pasteur, Paris, France.
043 (B) Burkholderia thailandensis is a Better Surrogate for B.
pseudomallei Growth and Persistence than B. pseudomallei Bp82
M. A. Firmani, S. Lehman, S. Dieringer-Boyer, K. Bower, B. Dorsey, D. Strat-
ton, K. Delp, D. K. R. Karaolis; Natl. Biodefense Analysis and Countermea-
sures Ctr., Frederick, MD.

16 10th ASM Biodefense and Emerging Diseases Research Meeting


044 (B) WITHDRAWN
045 (B) Efficacy Testing of Orally Administered Antibiotics against
an Inhalational Bacillus anthracis Infection in BALB/CMice
G. J. Hatch, S. R. Bate, J. Fretwell, J. Vipond, S. G. P. Funnell, A. D. G.
Roberts; Hlth. Protection Agency, Salisbury, United Kingdom.
046 (B) Human Anthrax Outbreaks in Georgia in 2011
1 1 1 1
L. Malania , N. Tsertsvadze , N. Abazashvili , M. Ramishvili , M. Broladze ,
S. Tsanava1, N. Avaliani1, R. J. OBISO2, P. Imnadze1; 1The Natl. Ctr. for Dis-
ease Control and Publ. Hlth., Tbilisi, Georgia, 2The Microbe Company, Chris-
tiansburg, VA.
047 (B) Proteomic Analysis of Atypical Y. pestis Strains Isolated
from Natural Foci in Georgia
1 11 1 2
R. Solomonia , M. Nozadze , E. Mikeladze , E. Zhgenti , G. Chanturia , M.
kekeklidze2, R. J. Obiso3, S. Francesconi4, L. Bakanidze2; 1Life Sci. Res. Ctr.,
Ilia State Univ., Tbilisi, Georgia, 2The Natl. Ctr. for Disease Control and Publ.
Hlth., Tbilisi, Georgia, 3The Microbe Company, Christiansburg, VA, 4Naval
Med. Res. Ctr., Frederick, MD.
048 (B) Clinical and Epidemiologic Assessment of Brucellosis
in Georgia
T. Onashvili1, E. Mamisashvili11, R. J. Obiso2, M. Nikolaishvili, M. Za-
kareishvili, I. Beradze; M. Donduashvili, M. Kokhreidze, N. Vepkhvadze;
1 Arizona Univ., Flagstaff, AZ.
Lab. of Ministry of Agriculture (LMA) Tbilisi, Georgia, 2The Microbe
Company, Christiansburg, VA.
049 (B) Members of the FPT Subfamily of MFS Transporters are
Important for Pathogenesis of Francisella tularensis and
Contribute to Modulation of the Host Cytokine Response
1 2 1 1 1 1
M. Marohn , A. Santiago , K. Shirey , S. Vogel , E. Barry ; Univ. of Maryland
Sch. of Med., Baltimore, MD, 2Univ. of Virginia Sch. of Med., Charlottesville,
VA.
050 (B) Testing Diagnostic and Potential Therapeutic Values of
Bacteriophages Capable of Lysing Yersinia pestis and Yersinia
pseudotuberculosis
A. A. Filippov, K. V. Sergueev, Y. He, M. P. Nikolich; WRAIR, Silver Spring,
MD.
051 (B) Biosynthesis of Lipoteichoic Acid in Bacillus anthracis
G. Garufi1,2, A. Hendrickx1,2, S. G. Richter1,2, K. Beeri1, J. Kern1, O.
Schneewind1,2, D. Missiakas1,2; 1 Univ. of Chicago, Chicago, IL, 2Howard
Taylor Ricketts Lab., Chicago, IL.
052 (B) Resistance of Francisella to Fosmidomycin Associated
with Mutations in the Glycerol-3-Phosphate Transporter
R. S. Mackie1, E. S. McKenney2, M. L. van Hoek1,3; 1George Mason Univ.,
Manassas, VA, 2Univ. of Virginia Hlth. System, Charlottesville, VA, 3George
Mason Univ., Manassas, VA.
053 (B) Control of Pigment Production in a White Phenotypic
Variant of Staphylococcus aureus
1 2 1 1 1
V. Antonic , A. Stojadinovic , M. Izadjoo , M. R. Alavi ; Henry M. Jackson
Fndn. for the Advancement of Military Med., Gaithersburg, MD, 2Combat
Wound Initiative Program, Bethesda, MD.
M O N DAY S C I E N T I F I C P RO G R A M
054 (B) Interspecies Interaction of Francisella tularensis and
Pseudomonas aeruginosa
S. N. Dean, M. L. van Hoek; George Mason Univ., Manassas, VA.
055 (B) A Method for Detection of Bacillus anthracis Spores in
Complex Matrices
O. Stephansson1,2, P. Ågren3,2, S. Byfors3,2, S. Ehrs1,2, M. Klint1,2, M. Lavan-
1 der3,2, C. Nilsson3,2, A. Lundin Zumpe3,2; 1Natl. Vet. Inst., Uppsala, Sweden,
2
Swedish Joint Lab. for Food Safety and Biopreparedness, Uppsala, Swe-
den, 3Natl. Food Agency, Uppsala, Sweden.
056 (B) Comparative Genomic MGE Finding Process Streamlines
Mobile Element Search
S. Y. Choi1,2, N. A. Hasan1,2, J. Chun3, M. Hoq4, A. Huq1, T. A. Cebula5,2, R. R.
2 Colwell1,5,2; 1Univ. of Maryland, College Park, MD, 2CosmosID Inc., College
Park, MD, 3Sch. of Biological Sci. and Inst. of Microbiol., Seoul, Korea, Re-
public of, 4Univ. of Dhaka, Dhaka, Bangladesh, 5Johns Hopkins Univ., Balti-
more, MD.
057 (B) A Literature-Based System for Selecting Burkholderia
pseudomallei Strain Panels for Animal Model Testing Under the
FDA Animal Rule
K. E. Van Zandt1, A. Tuanyok2, H. C. Gelhaus, Jr.1, K. T. Brittingham1, S. A.
Sarrazine1, P. S. Keim2, R. L. Warren1; 1Battelle, Columbus, OH, 2Northern
058 (B) Paratransgenic and Transgenic Analysis of Cellular
Interactions Between Ixodes Ticks, Mammalian Cells, and
Rickettsiae: Lighting the Way with Fluorescent Proteins
T. J. Kurtti, J. D. Oliver, N. Y. Burkhardt, R. F. Felsheim, M. J. Herron, U. G.
Munderloh; Univ. of Minnesota, St. Paul, MN.
059 (B) Comparison of Challenge Outcomes with Three Different
Bacillus anthracis Isolates in New Zealand White Rabbits
A. Belle1, L. Schaefer1, A. E. Boyer2, M. Gallegos-Candela2, J. R. Barr2,
M. de la Garza3, J. Nunneley3, R. Carrion, Jr.3, K. Wycoff1; 1Planet Biotechnol-
ogy Inc., Hayward, CA, 2CDC, Atlanta, GA, 3Texas BioMed. Res. Inst., San
Antonio, TX.
060 (B) Genotyping of Mycobacterium tuberculosis Strains from
a Third Level Hospital in Lima, Peru
C. A. Tello, Sr., N. A. Borja, R. Garcia de la Guardia; Sand Marcos Univ.,
Lima, Peru.
061 (B) Evaluation of a Commercially Available Disposable
Nebulizer for Use in Nose-Only Pneumonic Plague, Tularensis,
and Melioidosis Murine Models
W. C. Lin, K. Siefkas, S-C. Hu, D. Sullivan, B. Gingras, L. Holland; IIT Res.
Inst., Chicago, IL.
062 (D) A PCR Platform Comparison Study to Support
Sustainability in Resource-Limited Settings
A. Ongarbaev1, A. Khodiev1, A. Smagin2, V. Garib2, G. Abdukhalilova2, G.
Ibadova2, R. Obiso3, S. Francesconi4, A. Baccetty5, J. Callahan6, D. Williams7;
1
Tech. Management Company, Tashkent, Uzbekistan, 2Tashkent Inst. of
Postgraduate Med. Ed., Tashkent, Uzbekistan, 3The Microbe Company,
Christiansburg, VA, 4Naval Med. Res. Ctr., Silver Spring, MD, 5Defense
Threat Reduction Agency, Ft. Belvoir, VA, 6Biotechnology and Diagnostic
Solutions Animal Hlth. Life Technologies, Austin, TX, 7Threat Reduction
Support Ctr., Alexandria, VA.
BIODEFENSE PROGRAM & ABSTRACTS 17
MO N DAY S C I E N T I F I C P RO G R A M
063 (D) Development of User-Defined Assays for the Genexpert
Platform
E. Felder, R. Wölfel; Bundeswehr Inst. of Microbiol., Munich, Germany.
064 (D) Production and Qualification of Unrestricted Reagents to
Support Anthrax Assays
K. H. Clement, T. L. Rudge, Jr., N. T. Huber, K. A. Newman, K. A. Sankovich,
R. H. Migliozzi, C. L. Sabourin; Battelle, Columbus, OH.
065 (D) Development of Rapid Diagnostic Platform for Detection
of Category A Biothreat Pathogens in the Field
P. Patel1, M. Weidmann2, K. Achazi1, S. Linke1, O. Strohmeier3, D. Mark4, T.
van Oordt4, J. Drexler5, M. Eberhard5, F. von Stetten4, M. Niedrig1; 1Robert
Koch Inst., Berlin, Germany, 2Univ. Med. Ctr., Göttingen, Germany, 3Dept. of
Microsystems Engineering (IMTEK), Freiburg, Germany, 4HSG-IMIT,
Freiburg, Germany, 5QIAGEN Lake Constance GmbH, Stockach, Germany.
066 (D) Indirect Phage-Based Detection of Yersinia pestis: Field
and Clinical Applications
K. V. Sergueev, M. P. Nikolich, A. A. Filippov; WRAIR, Silver Spring, MD.
067 (D) Quantification of Anthrax Lethal Factor by Mass
Spectrometry: Application to Clinical Anthrax
A. E. Boyer1, A. R. Hoffmaster1, C. P. Quinn1, M. Gallegos-Candela1, R. C.
Lins2, R. A. Stoddard1, E. Steward-Clark3, P. Maniatis1, C. Marston1, L. X.
Cronin1, D. Aranio1, H. Li1, V. Semenova3, K. Isbell2, S. Shadomy1, M.
Lehman1, D. Blaney1, N. Pesik1, T. L. Smith1, J. R. Barr1; 1CDC, Atlanta, GA,
2
Battelle Analytical Services, Atlanta, GA, 3CDC, Chamblee, GA.
068 (D) Applying Elastic Light Scatter Technology (BARDOT) to
Integrate Organism Classification
H. Motlagh1, A. Giordano1, C. Waters1, E. Bae2, J. G. Thomas1, J. Robinson2;
1
West Virginia Univ., Morgantown, WV, 2Purdue Univ., West Lafayette, IN.
069 (D) Negative Capture for Selective Enrichment of Pathogen-
Derived Nucleic Acids in Clinical Samples
Z. Bent, S. Langevin, V. VanderNoot, P. Lane, D. Curtis, M. Tran-Gyamfi, S.
Branda, T. Lane; Sandia Natl. Labs, Livermore, CA.
070 (E) Surveillance on Plague Natural Foci in Georgia
L. G. Bakanidze, P. G. Imnadze, S. A. Tsanava, N. S. Tsertsvadze; Natl. Ctr.
Disease Control and Publ. Hlth. of Georgia, Tbilisi, Georgia.
071 (E) Performance of Rapid Viability PCR Method for Detection
of Bacillus anthracis Ames Spores from Sponge-Stick, and Vacuum
Sock and Filter Sample Types
S. R. Shah1, S. Letant2, G. Murphy2, T. Alfaro2, S. Kane2; 1EPA, Washington,
DC, 2LLNL, Livermore, CA.
072 (E) An Operational Special Unit to Amend Biological Security
M. H. Richter, M-H. Lee, C. Herzog; Robert Koch Inst., Berlin, Germany.
073 (E) The Microfluidic Bioagent Autonomous Networked
Detector (M-BAND): An update
M. Sanchez, L. Probst, E. Blazevic, B. Nakao, M. A. Northurp; Microfluidic
Systems, Fremont, CA.
074 (E) Investigation on Drinking Water Quality of the Bogra
Sadar Upazila: Findings from a Rapid Assessment Lesson in the
Bogra District of Bangladesh
M. Mollik1,2; 1Peoples Integrated Alliance, Dhaka, Bangladesh, 2PreSci.
Trust Funds, Phoenixville, PA.
18 10th ASM Biodefense and Emerging Diseases Research Meeting
075 (E) Multi-User Validation of a Real-Time PCR System for
Detecting Bacillus anthracis Collected by Air Sampling Devices
in a Liquid Matrix
U. K. Spaulding1, K. M. Clemens1, T. Hadfield2, V. Ryan2, S. Brunelle3, I. M.
Ota1; 1Idaho Technology, Inc., Salt Lake City, UT, 2MRIGlobal, Inc., Palm Bay,
FL, 3AOAC Intl., Gaithersburg, MD.
076 (F) A Hemorrhagic Fever Virus Sequence Database and
Analysis Platform
C. Kuiken; Los Alamos Natl. Lab., Los Alamos, NM.
077 (F) Ethical & Social Issues Related to Synthetic Biology: What
We Are Doing as Academicians?
S. T. HAKIM, S. M. H. Tayyab, S. G. Nadeem; Jinnah Univ. for Women,
Karachi, Pakistan.
078 (F) Identification and Molecular Characterization of Novel
Group C Orthobunyaviruses Isolated from Mosquitoes Captured
in Peru
L. L. Lofts, M. J. Turell, J. R. Kugelman, G. I. Koroleva, G. F. Palacios, C. A.
Whitehouse; USAMRIID, Frederick, MD.
079 (F) Development of a High-Throughput Bioinformatics
Typing System for the Phylogenomic Analysis of Infectious
Category A and B Agents
M. Eppinger, S. Agrawal, S. Daugherty, K. Galens, A. Maroo, L. Tallon, H.
Tettelin, C. Fraser-Liggett, J. Ravel; Univ. of Maryland, Sch. of Med., Balti-
more, MD.
080 (G) Humanization of Anti-Ricin Neutralizing Monoclonal
Antibody
W. Hu, J. Yin, L. Negrych, D. Chau, J. Cherwonogrodzky; Defence R & D,
Medicine Hat, Canada.
081 (G) Inhibition of Lipoteichoic Acid Synthesis in
Staphylococcus aureus and Bacillus anthracis
S. Richter1,2, A. P. A. Hendrickx2, O. Schneewind2,1, D. Missiakas2,1; 1Howard
Taylor Ricketts Lab., Argonne, IL, 2Univ. of Chicago, Chicago, IL.
082 (G) Discovery of Targeted and Broad Spectrum Antiviral
Countermeasures for Alphaviruses
D. Chung1, B. Beitzel2, J. Arterburn3, J. O. Tret1, J. Smith2, C. Schmaljohn2, G.
Gene Olinger2, C. Scully2, C. B. Jonsson1; 1Univ. of Louisville, Louisville, KY,
2
US Army Med. Res. Inst. of Infectious Diseases, Fort Detrick, MD, 3New
Mexico State Univ., Las Cruces, NM.
083 (G) Generation of Biologically Active Nano-Aerosol by an
Electrospray-Neutralization Method
V. N. Morozov1, I. L. Kanev1, N. K. Balabaev1, A. V. Glyakina1, M. L. van
Hoek2; 1RAS, Moscow, Russian Federation, 2George Mason Univ.,
Manassas, VA.
084 (H) H5N1 Avian Influenza Specific Preventive Maintenance
in the Russia
D. Surnev, Yu. Kuznetsov, T. Okovytaya, V. Yelnikov; JSK Pokrov Biological
Plant, Pokrov, Russian Federation.

085 (H) Immunogenicity and Efficacy of a Recombinant Plague
Vaccine (rF1V) in Swiss Webster Mice
P. Fellows1, J. Price1, W. Lin2, A. Rom2, S. Marshall3, L. Holland2, L. Wolfraim1;
1
Dynport Vaccine Co., LLC, Frederick, MD, 2IIT Res. Inst., Chicago, IL, 3Bio-
Stat Solutions, Inc., Mount Airy, MD.
086 (H) Ribosomal Protein TgP0 in the Invasion of Host Cells by
Toxoplasma gondii Tachyzoites
S. Rajagopal, S. Sharma; Tata Inst. of Fundamental Res., Mumbai, India.
087 (H) Advancing Clinical Grade rV10-2 Plague Vaccine Toward
Licensure
L. E. F. Quenee, N. A. Ciletti, D. Elli, T. M. Hermanas, O. Schneewind; Univ. of
Chicago, Argonne, IL.
088 (H) Tularemia Vaccine Trials in Nonhuman Primates
R. Tempel1, R. Shirley2, A. Legasse1, F. Heffron1, M. Axthelm1; 1Oregon Hlth.
& Sci. Univ., Portland, OR, 2Virogenomics, Inc., Portland, OR.
089 (H) A Clinical Phase II Study Confirming the Safety and
Immunogenicity of One or Two Doses of IMVAMUNE (MVA-BN)
Smallpox Vaccine in Vaccinia-Experienced Elderly Subjects
R. N. Greenberg for the POX-MVA-024 Sites1, P. Chaplin2; 1Univ. of Kentucky
Sch. of Med., Lexington, KY, 2Bavarian Nordic GmbH, Martinsried, Germany.
090 (H) LC16M8, an Attenuated Smallpox Vaccine Will be a Useful
Countermeasure against Bioterrorism with Smallpox
H. Yokote1, Y. Shinmura1, C. Uemura1, T. Kanehara1, S. Maruno1, H. Matsui1,
K. Ohkuma1, K. Yokoi1, A. Funatsu1, S. Gordon2, G. Franchini2, S.
Hashizume3; 1Kaetsuken, Kumamoto, Japan, 2Natl. Cancer Inst., Bethesda,
MD, 3Univ. of Chiba, Chiba, Japan.
091 (H) Development of a Guinea Pig Passive Transfer Efficacy
Model Using Immunoglobulin Derived from Clinical Trial
Volunteers Vaccinated with Recombinant Botulinum Vaccine A/B
S. Martin1, W. Swiderski1, K. Metcalfe1, N. Niemuth2, M. Vassar2, J. Shearer1;
1
DynPort Vaccine Company, Frederick, MD, 2Battelle BioMed. Res. Ctr.,
Columbus, OH.
092 (H) Increased Dose of Recombinant F1/V Plague Vaccine
Results in Increased Survival in Plague-Challenged Cynomologus
Macaques
K. A. Overheim, S. Lemoine, K. Gallegos, F. Romero, I. Fisher, A. Monier, C.
Schneider, M. W. Valderas, J. Wilder, R. Russell, R. L. Sherwood, R. LeClaire;
Lovelace Respiratory Res. Inst., Albuquerque, NM.
093 (H) Generation of Protective Immunity against Fatal
Rickettsial Disease
S. P. Riley1,2, M. M. Cardwell1,2, R. Hillman1,2, L. Quenee1,2, J. J. Martinez1,2;
1
Univ. of Chicago, Chicago, IL, 2The Howard Taylor Ricketts Lab., Lemont, IL.
094 (I) Determination of Efficacious Praziquantel Dose in
Different Mouse Strains: BALB/C and Swiss Mice for Treatment of
Schistosoma mansoni
M. P. M. N. Na1, D. Y. S. N. Yole2; 1Mombasa Polytechnique Univ. Coll.,
Nairobi, Kenya, 2Kenya Polytechnique Univ. Coll., Nairobi, Kenya.
095 (I) A Novel Mechanism by Which Human Neutrophils Kill
Bacillus anthracis
G. Ramachandran1, L. Baillie2, P. Tsai3, G. Rosen3, A. Cross1; 1Univ. of Mary-
land Med. Sch., Baltimore, MD, 2Welsh Sch. of Pharmacy, Cardiff, United
Kingdom, 3Dept. of Pharmaceutical Sci., Baltimore, MD.
M O N DAY S C I E N T I F I C P RO G R A M
096 (I) Burkholderia pseudomallei Induces IL-1Beta and IL-6
Secretion Via Activation of MAPK and STAT3 in Differentiated THP-
1 Cells
M-H. Cho1, Y-W. Shin1,2, J-H. Chun1, K-J. Hong1, G-E. Rhie1; 1Korea CDC,
Chungbuk, Korea, Republic of, 2Yonsei Univ., Seoul, Korea, Republic of.
097 (I) Increased Protection of African Green Monkeys Vaccinated
with Increasing Doses of a Recombinant F1/V Plague Vaccine
K. A. Overheim1, S. Lemoine1, K. Gallegos1, K. Brooks1, I. Fisher1, A. Monier1,
C. Schneider1, M. W. Valderas1, J. Wilder1, R. Russell1, R. L. Sherwood1, R.
LeClaire2; 1Lovelace Respiratory Res. Inst., Los Lunas, NM, 2Lovelace Respi-
ratory Res. Inst., Albuquerque, NM.
098 (I) Ebola Virus Secreted Glycoprotein Can Function as a
Decoy for Antibodies
G. S. Mohan, Y. Chinglai, L. Ye, R. Compans; Emory Univ., Atlanta, GA.
099 (J) Bioactivity of an Indigenous Plant Used in the Treatment
of Malaria against Anopheles Mosquito Larvae in South West
Nigeria
F. O. Omoya, B. E. Boboye; Federal Univ. of Technology, Akure, Nigeria.
100 (J) Formaldehyde Disinfection Using Porcine parvovirus as
Indicator
E. Emmoth, U. A. Bengtsson, I. Dergel, C. Hultén; Natl. Vet. Inst., Uppsala,
Sweden.
101 (J) Bactericidal Activity of Antimicrobial Peptides Immobilized
on Planar Surfaces
S. Arcidiacono, J. Uzarski, L. Doherty, W. Muller, C. Mello; U.S. Army Natick
Soldier Ctr., Natick, MA.
102 (J) Investigating the Dispersion and Decontamination of
Anthrax Spores within Indoor Work Areas
A. Clohessey, S. Hoque; Rowan Univ., Glassboro, NJ.
103 (K) Preparing Biodefense Professionals: MS in Biotechnology
Concentration in Biodefense and Certificate in National Security
Studies
K. M. Obom1, A. I. Roth2, P. J. Cummings3; 1Johns Hopkins Univ., Rockville,
MD, 2Johns Hopkins Univ., Washington, DC, 3Johns Hopkins Univ., Balti-
more, MD.
104 (K) Recruiting First Responders from the Dental Profession
R. J. Boylan, D. L. Glotzer; NYU Coll. of Dentistry, New York, NY.
105 (K) Tungiasis (Jigger Infestation) in Rural Kenya: An Emerging
Infectious Disease
N. N. N. N. Ngomi, Jr.; Jomokenyatta Univ., Nairobi, Kenya.
106 (K) Essential Oils of Moroccan Thymus munbyanus as
Natural Product for Nosocomial Infection Prevention
M. Fatima Zohra, Mohammed Khatouf, Badiaa Lyoussi, Abdelfattah Abdel-
laoui; Faculty of Sci. Dhar Mhraz, Fes, Morocco.
107 (K) Biosafety and Biosecurity: The Case of a Third World
Country and the One Health Spectrum
D. B. Ndumu1, R. Heckert2, C. S. Rutebarika1, R. W. Kaboyo3, M. Busuulwa4,
P. Atimnedi5, S. Balinandi6, P. Delarosa7; 1Ministry of Agriculture, Animal In-
dustry and Fisheries, Entebbe, Uganda, 2Robert Heckert Consulting LLC,
Bowie, MD, 3Ministry of Hlth., Kampala, Uganda, 4AFENET, Kampala,
Uganda, 5Uganda Wildlife Authority, Kampala, Uganda, 6Uganda Virus Res.
Inst., Entebbe, Uganda, 7Booz Allen Hamilton, Middletown, DE.
BIODEFENSE PROGRAM & ABSTRACTS 19
MO N DAY S C I E N T I F I C P RO G R A M

Highlighted Oral Abstract Presentations 3:15PM

113 Distinct Differences in Disease Presentation after Aerosol
009. Environmental Detection Exposure to Rift Valley Fever Virus Dependent on Inbred Rat Strain
Monday, February 27, 2012 | 3:00PM – 4:00PM J. M. Bales, D. Powell, L. M. Bethel, D. Reed, I. Pandrea, A. L. Hartman; Univ.
of Pittsburgh, Pittsburgh, PA.
Diplomat Ballroom
MODERATOR: 3:30PM
Miles Carroll; Hlth. Protection Agency, Wiltshire, United Kingdom. 114 Interplay between RVFV and the Host NF-KappaB Signaling
Cascade
PRESENTATIONS:
A. Narayanan, K. Kehn-Hall, L. Hill, S. Senina, R. VanDuyne, I. Guendel, R.
3:00PM Das, A. Baer, C. Bailey, F. Kashanchi; George Mason Univ., Manassas, VA.
108 Case-Clusters of Melioidosis Associated with Climate 3:45PM
Conditions and Geographical Topography in Southern Taiwan
Y-L. Chen1, Y-S. Chen2, H-S. Lin3, P-J. Liu1; 1Natl. Kaoshiung Normal Univ.,
115 Interactions of Venezuelan Equine Encephalitis Virus with
Kaohsiung, Taiwan, 2Kaohsiung Veterans Gen. Hosp., Kaohsiung, Taiwan,
Host MicroRNA Processing Machinery
3
E-DA Hosp., Kaohsiung, Taiwan. C. A. King, N. Shafagati, I. Guendel, F. Kashanchi, K. Kehn-Hall; George
Mason Univ., Manassas, VA.
3:15PM
109 Environmental Contamination of Goat Farms with Coxiella
burnetii During a Q Fever Outbreak
Highlighted Oral Abstract Presentations
G. J. Kersh, K. A. Fitzpatrick, A. V. Kondas, A. J. Roche, J. S. Self, R. A. Priest- 011. Bacterial Agents
ley, A. Bjork, A. D. Anderson; CDC, Atlanta, GA.
Monday, February 27, 2012 | 3:00PM – 4:00PM
3:30PM Palladian Ballroom
110 Rapid and Sensitive On-Site Identification of Biothreat MODERATOR:
Agents Using Electrochemical Biochips and a Portable Detection
Platform Petra F. Oyston; DSTL, Salisbury, United Kingdom.
T. Elssner, C. Pöhlmann; Bruker Daltonik GmbH, Leipzig, Germany. PRESENTATIONS:

3:45PM 3:00PM
111 Detection of Botulinum Neurotoxin Serotypes A and B Using 116 Temperature-Regulated Membrane Remodeling is Essential
Chemiluminescence and Electrochemiluninescene Immunoassays for Francisella Pathogenesis
in Food and Serum Matrices Y. Li1, D. A. Powell1, S. A. Shaffer2, D. A. Rasko3, D. R. Goodlett2, C. R. H.
L. W. Cheng, L. H. Stanker; USDA, Albany, CA. Raetz4, X. Wang5, R. K. Ernst1; 1Univ. of Maryland, Baltimore, MD, 2Univ. of
Massachusetts Med. Sch., Worcester, MA, 3Univ. of Maryland Sch. of Med.,
Baltimore, MD, 4 Duke Univ. Med. Ctr., Durham, NC, 5State Key Lab. of Food
Highlighted Oral Abstract Presentations Sci. and Technology, Wuxi, China.

010. Viral Agents 3:15PM

Monday, February 27, 2012 | 3:00PM – 4:00PM 117 Development of a Luciferase Reporter System to Monitor
Plague Infection
Ambassador Ballroom
Y. Sun, J. Pennington, M. G. Connor, J. M. Warawa, M. B. Lawrenz; Univ. of
MODERATORS: Louisville, Louisville, KY.
Jean Patterson; Texas Biomedical Res. Inst., San Antonio, TX. 3:30PM
Colleen Jonsson; Univ. of Louisville, Louisville, KY.
118 A Variant of LcrV, the Plague Protective Antigen and Needle
PRESENTATIONS: Cap Protein of Yersinia pestis that Blocks Type III Secretion
3:00PM K. Given1, A. Mitchell2, N. Miller1, O. Schneewind1; 1Univ. of Chicago, Ar-
gonne, IL, 2Univ. of Chicago, Chicago, IL.
112 First Evidence of Hantavirus Circulation in Terrestrial Small
Mammals From Madagascar 3:45PM
J-M. Heraud1, N. Razafindralambo1, T. andrianaivo2, M-M. Olive1, V. L. Soari- 119 Hide and Seek: Changes in Francisella Surface Proteins
malala2, S. M. Goodman2, J-M. Reynes3; 1Inst. Pasteur de Madagascar, An- During Infection Reveal Ideal Vaccine Targets
tananarivo, Madagascar, 2Association Vahatra, Antananarivo, Madagascar,
3 J. Huntley; Univ. of Toledo Coll. of Med., Toledo, OH.
Inst. Pasteur, Lyon, France.

20 10th ASM Biodefense and Emerging Diseases Research Meeting


Symposium Session
012. Biodefense at the Dinner Table: Emerging
Foodborne Illnesses
Monday, February 27, 2012 | 4:15 PM - 6:15 PM
Diplomat Ballroom
This symposium will explore mechanisms of pathogenesis of a num-
ber of foodborne viral and bacterial pathogens. A key goal in under-
standing enteric infections is elucidating the mechanisms by which
epithelial barriers are evaded, and the impacts that systemic co-in-
fections can exert. Understanding these phenomena will inform
strategies to prevent and treat foodborne infections. The scope in the
symposium will be from clinical and molecular epidemiology studies
to molecular genetics, cell biology and ultrastructure.
MODERATORS:
Marshall E. Bloom; DIR/NIAID/NIH, Hamilton, MT.
Melita A. Gordon; Univ. of Liverpool, Liverpool, United Kingdom.
PRESENTATIONS:
4:15PM
120 Mechanisms of Murine Norovirus Interaction with the
Intestinal Epithelium
Christiane Wobus; Univ. of Michigan Med. Sch., Ann Arbor, MI.
4:45PM
121 Picking the Lock: Stealth Invasion of Epithelial Barriers by
Enteric Pathogens
Manuel R. Amieva; Stanford Univ. Sch. of Med., Stanford, CA.
5:15PM
122 Multi-Dimensional Analysis of E. coli Colonization of
Mammalian Cells and Green Leafy Vegetables
Jorge A. Giron; Univ. of Florida, Gainesville, FL.
5:45PM
123 Clinical and Molecular Epidemiology of Invasive Salmonella
Disease in Africa Adults and Children
Melita Gordon; Univ. of Liverpool, Liverpool, United Kingdom.
M O N DAY S C I E N T I F I C P RO G R A M
Symposium Session
013. Persistence and Inactivation of Biothreat
Agents and Emerging Pathogens
Monday, February 27, 2012 | 4:15 PM - 6:15 PM
Palladian Ballroom
Following the attacks of 2011, it was quickly realized that persistence
and inactivation data on biothreat agents was lacking. Very few stud-
ies in this area had been conducted since the 1960s. In addition to
summarizing the progress that has been made over the last decade,
the research presented will inform the research microbiologists of the
need for additional like studies to form risk assessments, remediation
efforts and biopreparedness planning. A better understanding of the
persistence and inactivation parameters necessary for remediation will
improve response timelines associated with contamination events.
MODERATORS:
Gene Rice; EPA, Cincinnati, OH.
Tonya Nichols; EPA, Washington, DC.
4:15PM
124 Persistence of Category A Select Agents
Chuck Gerba; Univ. of Arizona, Tucson, AZ.
4:45PM
125 Persistence and Inactivation of Vaccinia Virus
Joseph Wood; EPA, Research Triangle Park, NC.
5:15PM
126 Inactivation of Bacterial Biothreat Agents in Water
Laura Rose; CDC, Atlanta, GA.
5:45PM
127 Decontamination of Bacillus Spores from Water Using
Germinant
Jeff Szabo; USEPA/NHSRC, Cincinnati, OH.
BIODEFENSE PROGRAM & ABSTRACTS 21
MO N DAY S C I E N T I F I C P RO G R A M

Symposium Session Roundtable Discussion

014. Taking the Bite Out of Insect Bites: Biodefense
Implications of Vector-Borne Infections
Monday, February 27, 2012 | 4:15 PM - 6:15 PM
Ambassador Ballroom
The scope of this session is to focus on the impact of arthropod vec-
tors on the kinetics of bacterial, viral and parasitic infections affecting
global health and implications for emerging and biodefense-related
diseases. The topics will broadly explore changes in distribution of
vectors and disease prevalence; effect of ecological changes and its
impact on vector-borne infections; novel preventive strategies against
vector-borne infections; and biome to genome tweets: Vector-
pathogen interactions. These areas are at the cutting-edge of vector-
borne disease research. An understanding of the current trends in
these areas will help in developing strategies to combat several
emerging diseases with significant impact on global health. Strate-
gies to prevent vector-borne diseases using novel vector-specific
chemotherapeutics would add to the list of prevention strategies for
vector-borne infections.
MODERATORS:
Janakiram Seshu; Univ. of Texas, San Antonio, TX.
Karl E. Klose; Univ. of Texas, San Antonio, TX.
PRESENTATIONS:
4:15PM
128 Borrelia burgdorferi-Tick Interactions: New Insight into
Ixodes Proteins Modulating Pathogen Persistence in the Vector
Utpal Pal; Univ. of Maryland, College Park, MD.
015. Recapping the 2011 Biological Weapons
Convention Review Conference (BWC RevCon)
Monday, February 27, 2012 | 6:30 PM - 7:30 PM
Palladian Ballroom
Awareness of safety and security issues is fundamental to the re-
sponsible conduct of work in the life sciences. This session will con-
tribute to the overall effectiveness of the microbiologist who attends
this meeting by increasing awareness of the Biological Weapons Con-
vention, with particular attention to international efforts directly rel-
evant to the life of the microbiologist, including those related to
laboratory biosecurity and biosafety, regulation, and efforts to engage
the scientific community in discussions of education, awareness-rais-
ing, and self-governance.
MODERATOR:
TBD
PANELISTS:
132 Christopher Park; U.S. Dept. of State, Washington, DC.
133 Gregory Koblentz; George Mason Univ., Fairfax, VA.
134 Gerald L. Epstein; U.S. Dept. of Homeland Security, Washington, DC.

4:45PM
129 Dissecting the Mechanisms of Tick Transmission from
Anaplasma to Francisella
Susan M. Noh; USDA-ARS Animal Disease Res. Unit, Pullman, WA.

5:15PM
130 Characterizing 24-48h Tick Saliva Proteins to Understand
Early Stage Tick Feeding Biology
Albert Mulenga; Texas A&M Univ., College Station, TX.

5:45PM
131 Tweets from Ticks: Modulation of Vector-Specific Gene
Expression by BadR in the Agent of Lyme Disease
Janakiram Seshu; Univ. of Texas, San Antonio, TX.

22 10th ASM Biodefense and Emerging Diseases Research Meeting


Tuesday, February 28, 2012
016. Global Threats, Collaborative Solutions
Tuesday, February 28, 2012 | 7:15AM – 8:15AM
Palladian Ballroom
This new program will provide insights into current international se-
curity challenges from notable thought leaders, policy advisors, and
senior government officials.
MODERATOR:
Dr. Jason Rao; ASM Director of International Affairs, Washington, DC.
PRESENTATION:
Dr. Alan Rudolph; Director for Chemical and Biological Technologies Direc-
torate, Research and Development Enterprise, Defense Threat Reduction
Agency (DTRA), Fort Belvoir, VA.
Plenary Session
017. Targeting Virulence Factors in Acute Infection
Tuesday, February 28, 2012 | 8:30AM - 12:00PM
Regency Ballroom
Antibiotic resistance is a global crisis caused by antibiotic overuse,
misuse and a dearth of new targets under development. This session
will examine which approaches have been successful and brought to
the clinic (toxin inhibitors, type three secretion, for example) and
which have not yet succeeded and the underlying difficulties for these
relative failures (e.g. reaching intracellular microbial targets within
tissues, specificity, redundancy, etc.).
MODERATORS:
Petra F. Oyston; DSTL, Salisbury, United Kingdom.
Nancy Connell; UMDNJ-New Jersey Med. Sch., Newark, NJ.
PRESENTATIONS:
8:30AM
135 Antimicrobial Peptide Modulation of Chemokine and Pro-
Inflammatory Cytokine Responses: It’s all in the Timing
Kim A. Brogden; Univ. of Iowa, Iowa City, IA.
9:00AM
136 Anthrax Diagnostic Tools (Nonclinical and Clinical) for
Medical Countermeasure Development Under the Animal Rule
Elizabeth Leffel; PharmAthene, Inc., Annapolis, MD.
9:30AM
137
Christian Baron; Univ. de Montreal, Montreal, Canada.
10:00AM
Coffee Break
T U E S DAY S C I E N T I F I C P RO G R A M
10:30AM
138 Targeting Cyclic Di-GMP Signaling to Prevent Biofilm
Formation
Chris Waters; Michigan State Univ., East Lansing, MI.
11:00AM
139 Identification of Inhibitors of DNA Adenine Methylation as
Novel Therapeutics
Petra Oyston; DSTL, Salisbury, United Kingdom.
Poster Session
018. Tuesday Poster Session
Tuesday, February 28, 2012 | 1:00PM – 3:00PM
Exhibit Hall
PRESENTATIONS:
140 (A) Molecular Epizootiology of Street Rabies Virus Isolates
in Ukraine
I. Polupan, N. Ivanov, O. Deryabin, V. Skripnik; Inst. of Vet. Med., Kyiv,
Ukraine.
141 (A) Utility of a New Real-Time RT-PCR Assay for the Detection
of Crimean-Congo Haemorrhagic Fever Virus in Pakistan
B. Atkinson1, Z. Hasan2, A. Samreen2, B. Jamil2, T. Moatter2, J. Chamberlain1,
N. Cook1, C. Logue1, C. Bruce1, S. Dowall1, R. Hewson1; 1Hlth. Protection
Agency UK, Salisbury, United Kingdom, 2The Aga Khan Univ., Karachi, Pak-
istan.
142 (A) Development of Differential and Broad Spectrum
Immunological Reagents for Identification and Characterization
of Filovirus Sub-Types
C. Bruce, K. Richards, L. Easterbrook, H. Love, L. Hudson, J. Plank, H. Tolley,
R. Hewson, A. Robers; Hlth Protection Agency, Salisbury, United Kingdom.
143 (A) Fighting Fire with Fire: The Development of Antiviral AAVs
K. K. Wong, Jr.; City of Hope Natl. Med. Ctr., Duarte, CA.
144 (A) Arenavirus Reverse Genetics for Vaccine Development
E. Ortiz Riano1, B. Y. H. Cheng1, J. C. de la Torre2, L. Martinez-Sobrido1; 1
Univ. of Rochester Med. Ctr., Rochester, NY, 2The Scripps Res. Inst., La Jolla,
CA.
145 (A) New Select Agent Regulations
J. Garrett, V. Sutton, H. Carson; Texas Tech Univ. Sch. of Law, Lubbock, TX.
146 (A) Development of a Murine Model for Aerosolized Filovirus
Infection
E. E. Zumbrun, H. A. Bloomfield, T. B. Chance, D. K. Nichols, A. Nalca; US-
AMRIID, Fort Detrick, MD.
147 (B) Antibiotic Profile of Nosocomial Bacterial Pathogens
N. N. Eduok1, I. P. Udoh2, C. I. Eleazar2, B. O. Ogeneh2; 1Ministry of Sci.
and Technology, Uyo, Nigeria, 2Univ. of Nigeria, Enugu, Nigeria.
148 (B) Comparative Transcriptomics of Three Species of Fleas
(Siphonaptera: Pulicidae)
C. A. Whitehouse1, G. Koroleva1, J. R. Kugelman1, S. K. Williams2, S. W.
Bearden2, K. L. Gage2, G. F. Palacios1; 1USAMRIID, Frederick, MD, 2CDC,
Fort Collins, CO.
BIODEFENSE PROGRAM & ABSTRACTS 23
T U E S DAY S C I E N T I F I C P RO G R A M
149 (B) Development of a Coxiella burnetii Aerosol Infection
Model in Rodents
K. R. Bewley, J. K. Pitman, D. J. Atkinson, G. McLuckie, M. Charlwood, G.
Hatch, J. Vipond, S. G. P. Funnell, A. Roberts; Hlth. Protection Agency,
Salisbury, United Kingdom.
150 (B) Automated Extraction of Genomic DNA from Food-
Associated Pathogens and Environmental Samples
P. Williams, T. Parrish, M. Duggan, J. Chapman, W. Burke, J. Birkenholz;
Evogen, Lenexa, KS.
151 (B) Monoclonal Antibody against Capsule of Bacillus
anthracis Protects Mice from Enhanced Lethal Toxin Activity Due
to Capsule and Anthrax Spore Challenge
G-E. Rhie1, J. Jang1, M. Cho1, J-H. Chun1, J. Park2, K. Hong1; 1Korea NIH,
Chungbuk 363-951, Korea, Republic of, 2Hankuk Univ. of Foreign Studies, Yon-
gin, Korea, Republic of.
152 (B) The Cat Flea as a Laboratory Model for Understanding
Yersinia pestis Colonization of its Insect Vector
T. M. Hermanas, L. E. Quenee, O. Schneewind; Univ. of Chicago, Chicago,
IL.
153 (B) Bacillus anthracis Acetyl-Transferases PatA1 and PatA2
and Secondary Cell Wall Polysaccharide-Mediated Assembly of S-
Layer Proteins
J. M. Lunderberg1,2, S-M. Nguyen-Mau1,2, G. S. Richter1,2, Y-T. Wang1,2, J.
Dworkin3, D. M. Missiakas1,2, O. Schneewind1,2; 1Univ. of Chicago, Chicago,
IL, 2Howard Taylor Ricketts Lab., Argonne, IL, 3Columbia Univ., New York,
NY.
154 (B) Autotransported (Type V Secreted) Proteins of
Burkholderia pseudomallei and their Role in Biofilm Formation,
Serum Resistance and Pathogenesis
N. R. Lazar Adler1, R. E. Dean2, M. P. Stevens3, J. L. Prior2, T. P. Atkins2,4, E. E.
Galyov1; 1Univ. of Leicester, Leicester, United Kingdom, 2Defence Sci. and
Technology Lab., Salisbury, United Kingdom, 3Univ. of Edinburgh, Easter
Bush, United Kingdom, 4Univ. of Exeter, Exeter, United Kingdom.
155 (B) S-Layer Assembly in Bacillus cereus G9241 Requires csaB
Y-T. Wang1,2, S-Y. Oh1,2, A. P. Hendrickx1, O. Schneewind1,2; 1Univ. of
Chicago, Chicago, IL, 2Howard Taylor Ricketts Lab., Argonne, IL.
156 (B) Antibody Interactions with the Capsule of Bacillus
anthracis
M. A. Hubbard, P. Thorkildson, W. Welch, D. AuCoin, T. Kozel; Univ. of Ne-
vada Sch. of Med., Reno, NV.
157 (B) Salmonella Enteritidis Gene SEN1140 Contributes to
Accelerated Type III Secretion System-2 Dependent Early
Inflammation Kinetics in Mouse Colitis Model
V. Vishwakarma1, B. Periaswamy2, N. B. Pati1, W-D. Hardt2, M. Suar1; 1Sch.
of Biotech., Bhubaneswar, India, 2Inst. of Microbiol., Zürich, Switzerland.
158 (B) Activity of Two Novel Antibacterial Agents XF-70 & XF-73
against Multidrug-Resistant Acinetobacter baumannii
K. L. Lohman1, J. Coffin1, W. Rhys-Williams2, I. Hayter2, W. G. Love2; 1MRI
Global, Rockville, MD, 2Destiny Pharma Ltd., Brighton, United Kingdom.
24 10th ASM Biodefense and Emerging Diseases Research Meeting
159 (B) Anthrax Surveillance System and Canonical SNP Typing
of Bacillus anthracis Strains Isolated in the Country of Georgia
2009-2011
G. Chanturia1, E. Khmaladze1, E. Zhgenti1, L. Malania1, M. Zakalashvili1,
N. Tsertsvadze1, M. Kekelidze1, P. Imnadze1, S. Tsanava1, R. J. Obiso2, S.
Francesconi3; 1The Natl. Ctr. for Disease Control and Publ. Hlth., Tbilisi,
Georgia, 2The Microbe Company, Christiansburg, VA, 3Naval Med. Res. Ctr.,
Frederick, MD.
160 (B) Diversity of Bacteriophages Active to B. anthracis
Bacterial Strains Isolated in Georgia
S. Rigvava1, M. Natidze2, L. Gubeladze1, L. Kvachadze1, R. J. Obiso3,
M. Kutateladze1, R. Adamia1; 1G. Eliava Inst. of Bacteriophages, Microbiol.
and Virology, Tbilisi, Georgia, 2G, Tbilisi, Georgia, 3The Microbe Company,
Christiansburg, VA.
161 (B) Characterisation of Inhalational Infection with
Burkholderia pseudomallei in BALB/c mice
S. G. P. Funnell, G. Hatch, J. A. Kane, S. Bates, K. M. Tonks, C. H. Tsang,
J. Fretwell, H. Shuttleworth, J. Vipond, A. Roberts; Hlth. Protection Agency,
Salisbury, United Kingdom.
162 (B) Reaerosolization of Bacillus Spores During the Bio-
Response Operational Testing and Evaluation
S. C. Taft1, R. A. Lordo2, D. J. Chappie2, E. Silvestri1, T. L. Nichols1, J. A. Rosati
Rowe1; 1Natl. Homeland Security Res. Ctr., Cincinnati, OH, 2Battelle Mem.
Inst., Columbus, OH.
163 (B) Regulation of the Yersinia Type Three Secretion Pathway
K. Kopaskie1,2, J. McCool3, K. Aly1, B. Blaylock1,2, O. Schneewind1,2; 1Univ. of
Chicago, Chicago, IL, 2Howard Taylor Ricketts Lab., Argonne, IL, 3Boston
Univ., Boston, MA.
164 (B) Real-Time Monitoring of Bacillus thuringiensis as a
Surrogate for Bacillus anthracis in Potable Water Distribution
System
S. P. Sherchan, I. Pepper; Univ. of Arizona, Tucson, AZ.
165 (B) Achromobacter Xylosoxidans: An Emerging Pathogen
Carrying Different Elements Involved In Horizontal Genetic
Transfer
G. Traglia1, C. Adams2, M. Almuzara3, C. Vay3, D. Centrón1, M. Ramirez2,1;
1
Sch. of Med., Buenos Aires, Argentina, 2California State Univ., Fullerton,
CA, 3Hosp. de Clinicas, Buenos Aires, Argentina.
166 (B) Increase in Temperature of Cultivation Above 37°C
Influenced Production of Yersinia pestis Major Protective Antigens
F1 and LcrV
V. A. Feodorova1, T. I. Polyanina1, M. A. Khizhnyakova1, A. M. Lyapina1, O. V.
Ulianova2,1, V. L. Motin3; 1Saratov State Vet. Inst., Saratov, Russian Federa-
tion, 2Saratov State Univ., Saratov, Russian Federation, 3Univ. of Texas Med.
Branch, Galveston, TX.
167 (B) Infection and Dissemination, Francisella tularensis
holarctica in Mosquito Vector
S. Backman; CBRN Defence and Security, Umea, Sweden.

168 (B) Phylogeography of Bacillus anthracis Isolates from the
Country of Georgia
E. Khmaladze1, D. Birdsell2, E. Zhghenti1, G. Chanturia1, M. Kekelidze1,
M. Nikolich3, P. Imnadze1, S. Tsanava1, L. Malania1, N. Tsertsvadze1,
S. Beckstrom-Sternberg4, J. Beckstrom-Sternberg4, T. Pearson2, D. Wagner2,
P. Keim2; 1Natl. Ctr. for Disease Control and Publ. Hlth., Tbilisi, Georgia,
2
Northern Arizona Univ., Flagstaff, AZ, 3Walter Read Army Inst. of Res., Fort
Detrick, MD, 4Translational Genomics Res. Inst., Phoenix, AZ.
169 (B) Polymorphisms in the lcrv Gene of Yersinia enterocolitica
Do Not Provide for Escape from Plague Protective Immunity
N. C. Miller, L. Quenee, D. Elli, N. Ciletti, O. Schneewind; Univ. of Chicago,
Chicago, IL.
170 (B) Multiple-Locus Variable Number Tandem Repeat Analysis
of Brucella melitensis from Azerbaijan Using Eight Markers
X-Z. Huang, E. B. Zelazowska, M. P. Nikolich; Walter Reed Army Inst. of Res.,
Silver Spring, MD.
171 (B) Rapid Single Nucleotide Repeat Sequence-Based
Detection and Identification of B. anthracis Using Pyrosequencing
Technology
K. R. Hahn, E. Valle, M. Thomas, T. Janzen, M. Shields, N. Goji, K. Amoako;
Canadian Food Inspection Agency, Lethbridge, Canada.
172 (B) Beef and Chicken Contamination with Resistant
Escherichial coli O157 from City Abattoirs in Southwest Nigeria
O. Olatoye, A. E. Amosun; Univ. of Ibadan, Ibadan, Nigeria.
173 (B) Contribution of OxyR to Burkholderia mallei Survival and
Virulence In Vitro and In Vivo
D. A. Revelli, J. Boylan, F. Gherardini; Rocky Mountain Lab., NIAID/NIH,
Hamilton, MT.
174 (B) Growth Inhibition of Burkholderia pseudomallei by Onion
Isolates of Burkholderia cepacia
J. M. Inamine, T. M. Eckstein; Colorado State Univ., Fort Collins, CO.
175 (D) Quality Assurance for Select Agents: An Australian
Approach
T. Theis1, D. Liu1, J. Gray1, P. Roffey2, W. Rawlinson3,1; 1RCPA BioSecurity
QAP, Surry Hills, Australia, 2Australian Federal Police, Weston, Australia,
3
SEALS Microbiol. Prince of Wales Hosp., Randwick, Australia.
176 (D) Broad-Range Microbial DNA Isolation from Clinical
Specimens for Universal PCR Diagnosis
C. Disqué1, M. Linow1, N. Murphy2; 1Molzym GmbH & Co.KG, Bremen,
Germany, 2CaerusBio Inc., Downingtown, PA.
177 (D)
WITHDRAWN
178 (D) Laboratory Surveillance of Tick Born Causing Agents and
Other Relevant Pathogens in the State of Maryland
M. Dembele1, L. Trivedi2, T. Lawson2, S. Montgomery2, M. P. Carlos2,1; 1Johns
Hopkins Sch. of Publ. Hlth., Baltimore, MD, 2Maryland Dept. of Hlth. and
Mental Hygiene Lab. Admin., Baltimore, MD.
T U E S DAY S C I E N T I F I C P RO G R A M
179 (D) Development of Single Domain Antibodies for Bacillus
anthracis Spore Proteins
S. A. Walper1, P. Lee2, J. Liu3, G. Anderson3, E. Goldman3; 1Natl. Academy of
Sci., Washington, DC, 2Nova Res. Inc., Alexandria, VA, 3Naval Res. Lab.,
Washington, DC.
180 (D) Zeptomole/ML Quantification of Anthrax Edema Factor by
LC-ESI-MS/MS: Activity Enhancement by Monoclonal Antibodies
R. C. Lins1, Z. Kuklenyik2, C. P. Quinn2, K. Bedi2, J. Goldstein2, A. Lyons2,
M. Gallegos2, U. Ansari1, R. Pitts1, D. Bagarozzi1, K. Isbell1, C. E. Leysath3,
Z. Chen3, S. H. Leppla3, J. R. Barr2, A. Boyer2; 1Battelle CDC, Atlanta, GA,
2
CDC, Atlanta, GA, 3NIH, Bethesda, MD.
181 (E) Simultaneous Detection of Ricin and Abrin DNA by Real-
Time PCR
I. Mossbrugger, E. Felder, R. Wölfel; Bundeswehr Inst. of Microbiol.,
Munich, Germany.
182 (E) Low-Cost, Highly Efficient Airborne Pathogen Collection
and Detection System for Field Use
J. Gordon1,2, P. Gandhi1; 1Inspirotec LLC, Chicago, IL, 2Northwestern Univ.,
Evanston, IL.
183 (E) Evaluation of Automated and Manual Commercial Nucleic
Acid Extraction Methods for Detection of Ricinus communis DNA
by Real-Time PCR
A. S. Hutchins1, M. Astwood2, J. R. Saah1, L. A. Dauphin2; 1North Carolina
State Lab. of Publ. Hlth., Raleigh, NC, 2 CDC, Atlanta, GA.
184 (E) Development of Multiplex Rapid Tests Allowing Warfare
Agents Detection
T. Berthelot, J. Credou, H. Volland; CEA, Gif sur Yvette, France.
185 (E) Validation of Unique Signature Discovery in Select Agents
and an Emerging Pathogen
B. J. Haley1, E. Taviani1, A. Chen1, A. Phillippy2, A. Huq1,3, R. R. Colwell1,4,5,
I. Knight3,6; 1Maryland Pathogen Res. Inst., College Park, MD, 2Ctr. for Bioin-
formatics and Computational Biology, College Park, MD, 3Maryland Inst. for
Applied Environmental Hlth., College Park, MD, 4Univ. of Maryland Inst. for
Advanced Computer Studies, College Park, MD, 5CosmosID, College Park,
MD, 6Canon U.S. Life Sci., Inc., Rockville, MD.
186 (E) Novel Surface Chemistry For The Development of
Multiplex Immunochromatographic Tests
J. Credou, H. Volland, T. Berthelot; CEA, Gif sur Yvette, France.
187 (F) Virus Pathogen Resource (ViPR): An Open Bioinformatics
Database and Analysis Resource for Human Virus Pathogen
Research
Y. Zhang1, B. Pickett1, E. Sadat1, J. Noronha1, R. B. Squires1, V. Hunt1,
M. Liu2, S. Kumar3, S. Zaremba3, Z. Gu3, L. Zhou3, C. Larson4, J. Dietrich3, E.
Klem3, R. H. Scheuermann1; 1Univ. of Texas Southwestern Med. Ctr., Dallas,
TX, 2Southern Methodist Univ., Dallas, TX, 3Northrop Grumman Hlth. Solu-
tions, Rockville, MD, 4Vecna Technologies, Greenbelt, MD.
188 (F) Effects on Host Responses of Air-Borne and Aerosolized
Rabbitpox Infections in New Zealand Rabbits
L. DaSilva1, A. Porter1, A. Nalca1, R. Settlage2, C. L. Galindo3, L. McIver2,
K. McMahon2; 1USAMRIID, Frederick, MD, 2Virginia Bioinformatics Inst. at
Virginia Polytechnic Inst., Blacksburg, VA, 3Eagle Eye Data Analysis, Hous-
ton, TX.
BIODEFENSE PROGRAM & ABSTRACTS 25
T U E S DAY S C I E N T I F I C P RO G R A M
189 (F) Analysis of Next-Generation Sequencing for Detecting
Rare Variants in Bacillus anthracis
R. E. Colman1, J. Schupp1, J. Gillece1, A. Rawat1, D. M. Engelthaler1, J. Fos-
ter2, P. Keim2,1; 1Translational Genomics Res. Inst., Flagstaff, AZ, 2Northern
Arizona Univ., Flagstaff, AZ.
190 (F) Nested Amplification for Improved Genotyping of Coxiella
burnetii
R. Vipond, A. Pepler, G. Vincent, J. Latham; Hlth. Protection Agency, Salis-
bury, United Kingdom.
191 (F) Utility and Challenges Associated with Use of Next
Generation Sequencing in Ultra-Rare Variant Detection
H. Koshinsky1, V. Fofanov2, J. Liu2; 1Eureka Genomics, Hercules, CA, 2Eureka
Genomics, Houston, TX.
192 (G) PANACEA Broad-Spectrum Antiviral Therapeutics
T. H. Rider; Massachusetts Inst. of Technology, Cambridge, MA.
193 (G) Efficacy of a Doxycycline Treatment Regimen in Rhesus
Macaques Aerosol Challenged with Brucella melitensis
L. Henning, E. McGuinness, D. Pak, S. Miller, C. Briscoe, M. Anderson,
R. Warren; Battelle, Columbus, OH.
194 (G) Host-Directed Antimicrobial Therapeutics for Intracellular
Bacterial Pathogens
A. Levi1, D. Arvans2, P. Robins2, L. P. Potluri1, B. Heaton1,2, S. Riley1,2, S.
Crosson1,2, J. Martinez1,2, J. E. Gabay1, H. A. Shuman2,1; 1Univ. of Chicago,
Chicago, IL, 2Howard Taylor Ricketts Lab., Lemont, IL.
195 (G) Restoring Antibiotic Sensitivity with Antisense Inhibitors
of Gene Expression
P. L. Iversen, D. V. Mourich, F. Schnell, B. Geller; AVI BioPharma, Inc.,
Bothell, WA.
196 (G) Post-Exposure Administration of a Monoclonal Antibody
Cocktail is Protective in Orthopoxvirus-Infected Mice and
Cynomolgus Macaques
J. L. Nordstrom1, G. J. Rainey1, H. Li1, P. Silvera2, F. Koide2, M. Buller3, J.
Schriewer3, A. Dekonenko4, K. Shah1, L. Huang1, V. Ciccarone1, S. Burke1, M.
Bowe1, D. Farb1, M. Kadan1, E. Bonvini1, K. Stein1, S. Koenig1, S. Johnson1;
1
MacroGenics, Rockville, MD, 2Southern Res. Inst., Frederick, MD, 3Saint
Louis Univ., St. Louis, MO, 4Univ. of New Mexico, Albuquerque, NM.
197 (H) Clinical Development of a Plant-Produced Subunit
Vaccine against Bacillus anthracis
V. Yusibov, J. Chichester, S. Manceva, K. Musiychuk, M. Shamloul, A. Rhee,
J. Trefethen, M. Jones; Fraunhofer USA CMB, Newark, DE.
198 (H) Adoptive Transfer of Pulsed BMDC Enhances Specific
Immune Responses and Provides Protection against Lethal
Anthrax Challenge
I. J. Thompson1, M. G. Stokes1, B. Hallis2, A. J. Simpson1, E. D. Williamson1;
1
Defense Sci. and Technology Lab., Salisbury, United Kingdom, 2Hlth. Pro-
tection Agency, Salisbury, United Kingdom.
199 (H) Radioprotective Mn-DP-Pi Complexes Preserve
Immunogenicity of Inactivated VEEV During Gamma Irradiation:
A New Approach for Vaccine Development
M. Gayen1,2, E. K. Gaidamakova1, P. Gupta1,2, A. Sharma1, P. J. Glass3, M. J.
Daly1, R. K. Maheshwari1; 1Uniformed Services Univ. of Hlth. Sci., Bethesda,
MD, 2Birla Inst. of Tech. and Sci., Pilani, India, 3U.S. Army Med. Res. Inst. of
Infectious Diseases, Frederick, MD.
26 10th ASM Biodefense and Emerging Diseases Research Meeting
200 (H) Longevity of Vaccine Protection against Pneumonic
Plague in BALB/C Mice, Part 2
Z. N. Llewellyn1, F. Koide2, L. E. Bowen1, P. William2, S. Lin2, J. A. Boydston1,
N. Harman-Richardson3, P. M. Silvera2; 1Southern Res. Inst., Birmingham,
AL, 2Southern Res. Inst., Frederick, MD, 3Alpha StatConsult, LLC,
Damascus, MD.
201 (H) Dose Response of IMVAMUNE, a Third Generation
Smallpox Vaccine Candidate, in a Lethal Monkeypox Virus
Nonhuman Primate Inhalation Model
T. Brasel, K. Agans, K. Dearen, T. Bailey, A. Russo, S. Storch, R. Sherwood;
Lovelace Respiratory Res. Inst., Albuquerque, NM.
202 (H) Stabilization Technology for Biodefence Vaccines:
Exemplification Using a New Viral Vaccine and an Adjuvanted
Vaccine
J. Drew, C. Andrews, E. Abbott, S. Ward; Stabilitech Ltd., London, United
Kingdom.
203 (H) A Marmoset Model for Lethal Orthopox Virus Infection
Enables Study of Vaccines
C. Yue1, M. Kramski1,2, C. Stahl-Hennig3, K. Mätz-Rensing3, F-J. Kaup3, G.
Pauli1, H. Ellerbrok1; 1Robert Koch Inst., Berlin, Germany, 2Univ. of Mel-
bourne, Parkville, Australia, 3German Primate Ctr., Goettingen, Germany.
204 (H) New Adjuvants for Anthrax Vaccine
Y. Gauthier1, A. Borel2, Y. Trescos1, J. Mathieu1, B. Verrier2, J-N. Tournier1;
1
IRBA, La Tronche, France, 2CNRS-IBCP, Lyon, France.
205 (H) Effects of Signal Sequences on the Immunogenicity and
Efficacy of an MVA Vectored Plague Vaccine
J. Brewoo1, T. Powell2, G. Young1, C. Partidos1, D. Stinchcomb2, J. Osorio3;
1
Inviragen, Inc., Madison, WI, 2Inviragen, Inc., Fort Collins, CO, 3Univ. of
Wisconsin, Madison, WI.
206 (H) Second-Generation Inactivated Vaccines Protect Mice
from Lethal Venezuelan Equine Encephalitis Virus Infection
K. Spurgers1, S. Honnold1, R. Bakken1, J. Cohen1, L. Rowan1, C. Lind1, P.
Gupta2, A. Sharma2, R. Maheshwari2, P. Glass1; 1USAMRIID, Frederick, MD,
2
USUHS, Bethesda, MD.
207 (H) Evaluation of Inactivated and Live-Attenuated Bivalent
Vaccines that Confer Protection against Rabies and Ebola Viruses
in Mice
J. Blaney1, A. Papaneri1, C. Wirblich2, J. Cann1, M. Holbrook1,3, A. Freiberg3,
P. Jahrling1, M. Schnell2; 1NIAID/NIH, Fort Detrick, MD, 2Jefferson Med. Coll.,
Thomas Jefferson Univ., Philadelphia, PA, 3Univ. of Texas Med. Branch,
Galveston, TX.
208 (H) Improved Efficacy of a Multiagent Equine Encephalitis
Virus Vaccine Candidate
P. J. Glass1, R. R. Bakken1, J. Ennis2, K. B. Spurgers1, K. I. Kamrud3,
K. Alterson3, M. Custer3, J. Turner2; 1USAMRIID, Fort Detrick, MD, 2Defyrus
Inc., Toronto, Canada, 3Alphavax, Research Triangle Park, NC.
209 (H) Relationship between Vaccine Dosage, Immunogenicity
and Survival in rF1V-Vaccinated Cynomolgus Macaques
P. Fellows1, J. Price1, R. Krile2, H. Lockman2, L. Wolfraim1; 1Dynport Vaccine
Company LLC, Frederick, MD, 2Battelle, Columbus, OH.
 T U E S DAY S C I E N T I F I C P RO G R A M

210 (I) Peptide Mapping of Anthrax Toxin Protective Antigen
Using Anti-AVA Human Sera
V. A. Semenova, P. Svoboda, J. Pohl, S. Soroka, C. Quinn; CDC, Atlanta, GA.
211 (I) Serum Opsonization Promotes the Efficient Uptake and
Clearance of Burkholderia pseudomallei by Human Neutrophils
M. E. Woodman, R. G. Worth, R. M. Wooten; Univ. of Toledo Coll. of Med.,
Toledo, OH.
212 (I) Priming of Alveolar Macrophages Augments Inflammatory
Response on Anthrax Lethal Toxin Stimulation
J. Thomas, L. Xiao, J. W. Christman, J. L. Cook, I. Levitan; Univ. of Illinois
Med. Ctr., Chicago, IL.
213 (I) A Quantitative PCR-Based Method for the Evaluation of
Anti-Vaccinia Neutralization Titers
A. D. Garcia, C. A. Meseda, J. Weir; FDA/CBER, Bethesda, MD.
214 (I) The Chemokine Receptor CCR7 is Expressed in Cells of the
Human Lung in Association with Bacillus anthracis Spore Infection
M. Langer1, J. Booth1, E. Duggan1, V. Patel1, T. Veres2, F. Prenzler2, K. Sewald2,
K. Coggeshall3, A. Braun2, J. Metcalf1; 1Univ. of Oklahoma, Oklahoma City,
OK, 2Fraunhofer Inst. for Toxicology and Experimental Med., Hannover,
Germany, 3Oklahoma Med. Res. Fndn., Oklahoma City, OK.
215 (J) Forensic Entomology and Biodefense
A. Lindström; Natl. Vet. Inst., Uppsala, Sweden.
216 (J) Rapid Detection of Endospores by Capillary
Electrophoresis on a Hybrid Channel-to-Digital Microfluidic
Platform for Sterilization Validation
L. Li, W. Wu, W. Chan, H. Cheong, P. Yung; Chinese Univ., Shatin, Hong Kong.
Highlighted Oral Abstract Presentations
019. Immune Response
Tuesday, February 28, 2012 | 3:00PM – 4:00PM
Diplomat Ballroom
MODERATOR:
Catharine Basio; NIAID/Rocky Mountain Lab., Hamilton, MT.
PRESENTATIONS:
3:00PM
221 Ornithine Lipids as Therapeutic Targets for Melioidosis
M. Gonzalez-Juarrero1, R. Bowen1, W. Mwangi2, T. M. Eckstein1; 1Colorado
State Univ., Fort Collins, CO, 2Texas A&M Univ., College Station, TX.
3:15PM
222 Identification of Host Responses Contributing Towards
Neuroinvasion and Neurovirulence by Venezuelan Equine
Encephalitis Virus
P. Gupta1,2, A. Sharma1, M. Bhomia1,2, J. Han3, A. Yang3, R. K. Puri3,
R. K. Maheshwari1; 1Uniformed Services Univ. of Hlth. Sci., Bethesda, MD,
2
Birla Inst. of Technology and Sci., Pilani, India, 3Food and Drug Admin.,
Bethesda, MD.
3:30PM
223 Dynamic Analysis of Respiratory Immune System Provides
New Insights on Pulmonary Anthrax
D. Fiole1,2, A. Quesnel-Hellmann1, J. D. Mathieu1, J. Douady2, J-N. Tournier1;
1
Inst. de Recherche Biomédicale des Armées, Grenoble, France, 2Univ.
Joseph Fourier Grenoble, Saint Martin d’Hères, France.

217 (K) WMD Training Courses at Dugway Proving Ground, 3:45PM

Dugway, UT 224 Visualizing Early Immune Response: Bacterial Specific
P. B. Brown, J. Montague, J. Sorenson; U.S. Army, Dugway Proving Ground, Reorganization at the Nanoscale
Dugway, UT. J. S. Aaron, Q. B. Smith, B. D. Carson, J. A. Timlin; Sandia Natl. Lab.,
Albuquerque, NM.
218 (K)
WITHDRAWN

219 (K) Biological Security: Identifying Vulnerabilities,

Preventing Damage
C. Uhlenhaut; Robert Koch Inst., Berlin, Germany.

220 (K) Transitional Operations: Guiding Owners Through Start-

Up Operations and Regulatory Compliance
M. L. Weiss, D. C. Sharpe; The WorkingBuildings Companies, Atlanta, GA.

BIODEFENSE PROGRAM & ABSTRACTS 27

T U E S DAY S C I E N T I F I C P RO G R A M
Highlighted Oral Abstract Presentations
020. Therapeutics
Tuesday, February 28, 2012 | 3:00PM – 4:00PM
Ambassador Ballroom
MODERATOR:
Colleen B. Jansson; Univ. of Louisville, Louisville, KY.
PRESENTATIONS:
3:00PM
225 Novel Strategies to Enhance Current Antibiotic Therapies
R. V. D’Elia, T. R. Laws, A. Carter, R. A. Lukaszewsk, G. C. Clark; DSTL,
Salisbury, United Kingdom.
3:10PM
226 EV-035: A New Compound Series with Broad Spectrum
Antibiotic Activity against Burkholderia pseudomallei and Other
Select Agent Pathogens
S. Goldman1, D. Milligan1, H. Yao1, G. Liu1, G. Salerno2, P. Roussel2, J. Heim2;
1
Evolva, Inc., Palo Alto, CA, 2Evolva SA, Reinach, Switzerland.
3:20PM
227 A Single PMOplus Oligomer is Effective in a Cynomolgus
Macaque Marburg Virus Lethal Challenge Model
P. Iversen1, T. Warren2, A. Shrutleff2, J. Wells2, N. Garza2, K. Stuthman2, S.
Van Tongeren2, L. Dong2, D. Mourich3, L. Hauck1, J. Kugelman2, B. Beitzel2,
G. Palacios2, S. Bavari2; 1AVI BioPharma, Bothell, WV, 2USAMRIID, Fredrick,
MD, 3AVI BioPharma, Bothell, WA.
3:30PM
228 Therapeutic Efficacy of ST-246 in Cynomolgous Macaques
Challenged with Monkeypox Virus via Aerosol
A. T. Russo, K. Agans, T. Brasel, K. Dearen, T. Bailey, S. Storch, R. Baker;
LRRI, Albuquerque, NM.
3:40PM
229 Rationally Designed Broad-Spectrum Antifungals Agents for
the Treatment of Biodefense Pathogens
E. Garvey, S. Brand, W. Hoekstra, C. Yates, W. Moore, R. Schotzinger;
Viamet Pharmaceuticals, Morisville, NC.
28 10th ASM Biodefense and Emerging Diseases Research Meeting
Highlighted Oral Abstract Presentations
021. Diagnostics and Vaccines
Tuesday, February 28, 2012 | 3:00PM – 4:00PM
Palladian Ballroom
MODERATOR:
Elisabeth Carniel; Inst. Pasteur, Paris, France.
PRESENTATIONS:
3:00PM
232 Construction of a Lateral Flow Immunoassay for the
Diagnosis of Melioidosis
D. Reed1, N. Chantratita2, S. J. Peacock3, M. N. Burtnick4, P. J. Brett5, S. Ray-
chaudhuri6, R. Houghton6, T. R. Kozel1, D. P. AuCoin1; 1Univ. of Nevada Sch.
of Med., Reno, NV, 2Mahidol Univ., Bangkok, Thailand, 3Univ. of Oxford, Ox-
ford, United Kingdom, 4Univ. of South Alabama, Reno, NV, 5Univ. of South
Alabama, Mobile, AL, 6InBios Intl., Seattle, WA.
3:20PM
233 Immunization with Burkholderia pseudomallei Outer
Membrane Vesicles Protects against Pneumonic and Septicemic
Melioidosis
W. Nieves1, B. Judy2, A. Torres2, C. J. Roy3, L. A. Morici1; 1Tulane Univ. Sch. of
Med., New Orleans, LA, 2Univ. of Texas Med. Branch, Galveston, TX, 3Tulane
Natl. Primate Res. Ctr., Covington, LA.
3:40PM
234 Sortase-Conjugation Generates a Capsule Vaccine that
Protects Guinea Pigs against Bacillus anthracis
O. Schneewind, G. Garufi, Y-T. Wang, S-Y. Oh, H. Maier, D. Missiakas; Univ.
of Chicago, Chicago, IL.
Symposium Session
022. Host Genetic Resistance to Infections
Tuesday, February 28, 2012 | 4:15 PM - 6:15 PM
Diplomat Ballroom
It is now well established that the major factor accounting for the
lethality due to infectious diseases is not environmental, but genetic.
This session will provide state-of-the-art information on genetic de-
terminants that predispose individuals to resist to or to develop dif-
ferent types of infectious diseases. In this symposium, an example of
genetic determinisms to infectious disease caused by a parasite, a
fungus, a bacterium and a virus will be presented. In addition, the mo-
lecular and cellular bases for these susceptibility/resistance pheno-
types will be discussed.
MODERATORS:
Simon J. Foote; Macquarie Univ., Macquarie Park, Australia.
Elisabeth Carniel; Inst. Pasteur, Paris, France.

PRESENTATIONS:
4:15PM
235 Host Genetic Resistance to Parasitic Infections: The Case of
Malaria
Simon J. Foote; Macquarie Univ., Macquarie Park, Australia.
4:45PM
236 Host Genetic Resistance to Mycobacterial Infections: The
Case of Mycobacterium tuberculosis
Philippe Gros; McGill Univ., Montreal, Canada.
5:15PM
237 Host Genetic Resistance to Gram-Negative Bacterial
Infections: The Case of Yersinia pestis
Christian E. Demeure; Inst. Pasteur, Paris, France.
5:45PM
238 Host Genetic Resistance to Viral Infections: The Case of HIV
Florencia Pereyra; Brigham and Women’s Hosp. and the Ragon Inst. of
MGH, Boston, MA.
Symposium Session
023. Dynamic Pathogens Make Difficult Targets
Tuesday, February 28, 2012 | 4:15 PM - 6:15 PM
Ambassador Ballroom
The remarkable adaptability of viral and bacterial pathogens compli-
cates the development of diagnostics, therapeutics, and vaccines.
Considering how pathogens evolve in the face of changing environ-
mental pressures, the host response, or drug therapies is a critical
perspective. This session will highlight recent insights into the mo-
lecular mechanisms of pathogen evolution and how this informs the
design of novel more durable interventions.
MODERATORS:
Ted C. Pierson; NIAID, Bethesda, MD.
Jon Yewdell; NIAID, NIH, Bethesda, MD.
PRESENTATIONS:
4:15PM
239 Diversity and Evolution of Pathogenic Escherichia coli:
Lessons Learned from High-Throughput Genome Sequencing
David A. Rasko; Univ. of Maryland Sch. of Med., Baltimore, MD.
4:45PM
240 Adaptive Arbovirus Landscapes and Implications for Disease
Emergence
Scott C. Weaver; Univ. of Texas Med. Branch, Galveston, TX.
5:15PM
241 On the Molecular Evolution of Influenza A Surface Proteins
Joshua B. Plotkin; Univ. of Pennsylvania, Philadelphia, PA.
5:45PM
242 Burkholderia pseudomallei: A Bacterial Pathogen with a
Highly Adaptable Genome
Paul Keim; Northern Arizona Univ., Flagstaff, AZ.
T U E S DAY S C I E N T I F I C P RO G R A M
Symposium Session
024. Visual Discovery of Microbe-Host Interactions
Tuesday, February 28, 2012 | 4:15 PM - 6:15 PM
Palladian Ballroom
Pathogen-host interactions are being unraveled by a wide spectrum of
techniques ranging from high resolution imaging of pathogens, live
imaging of host response and interactions with specific cellular com-
ponents to whole animal imaging of pathogenesis. Within this vast
array of technical approaches, the direct visualization of pathogens
within cells, tissues and whole animals is revealing novel macro-level
interactions. Especially when combined with real-time imaging and 3-
dimensional reconstruction, the field is linking pieces of the patho-
genesis puzzle together and helping us understand pathogen biology
more thoroughly.
MODERATORS:
Colleen B. Jonsson; Univ. of Louisville, Louisville, KY.
Andrew Hayhurst; Texas Biomedical Res. Inst., San Antonio, TX.
PRESENTATIONS:
4:15PM
243 Viral Nesting: Rewiring the Host to Generate Organelle
Platforms for Replication
Nihal Altan-Bonnet; Rutgers Univ., Newark, NJ.
4:40PM
244 How Vaccinia Virus Manipulates Signaling and the
Cytoskeleton to Enhance its Spread
Michael Way; London Res. Inst., London, United Kingdom.
5:05PM
245 Lighting Up Paramyxovirus Pathogenesis: Is Virus Eradication
a Reality, a Redundant Concept, or a Risk?
Paul Duprex; Boston Univ. Sch. of Med., Boston, MA.
5:30PM
246 Insights into Respiratory Melioidosis through Optical
Diagnostic Imaging
Jonathan M. Warawa; Univ. of Louisville, Louisville, KY.
5:55PM
247 Computer-Assisted Quantitative Analysis of Infectious
Disease Imaging
Daniel J. Mollura; NIH, Bethesda, MD.
BIODEFENSE PROGRAM & ABSTRACTS 29
T U E S DAY S C I E N T I F I C P RO G R A M

Roundtable Discussion
025. Global Status of Strategies for Addressing
Intersection of Science and Security
Tuesday, February 28, 2012 | 6:30 PM - 7:30 PM
Palladian Ballroom
Life sciences research is a vital social undertaking that yields innu-
merable and immeasurably important benefits. At the same time, we
must also recognize that good science can be put to bad uses, and
that even a single misuse of certain information, knowledge, or tech-
nology could have devastating and far-reaching effects. Over the past
ten years, there has been a growing recognition that some information
resulting from life sciences research, in the wrong hands, can be mis-
used to pose a threat to public health and national security. Research
yielding new technologies or information with the potential for both
benevolent and malevolent applications is referred to as “dual use re-
search.” Because the life sciences are a global endeavor, it is impor-
tant to work together to develop effective approaches to minimizing
the risk of misuse, while still fostering the continued progress of the
science that underpins public health and safety. A critical step is en-
gaging microbiologists, who may not be aware of the issue. This ses-
sion provides an opportunity to learn about the dual use issue and
provides a forum for sharing international perspectives and activities
relating to dual use life sciences research. The presenters and partic-
ipants in this session will explore strategies for managing the over-
sight of dual use life sciences research, as well as strategies for
fostering international awareness and engagement on the issue. A
major focus of the session will be on lessons learned from the imple-
mentation of strategies in various countries as well as to hear from
those countries with strategies under development.

MODERATORS:
Stuart Levy; Tufts Univ. Sch. of Med., Boston, MA.
Paul Keim; Northern Arizona Univ., Flagstaff, AZ.
PANELISTS:
248 Stuart Levy; Tufts Univ. Sch. of Med., Boston, MA.
249 Koos Van der bruggen; Independent Researcher and Science Writer,
The Hague, Netherlands.
250 Catherine Jefferson; The Royal Society, London, United Kingdom.
251 Gerald L. Epstein; Biosecurity Analyst, Washington, DC.

30 10th ASM Biodefense and Emerging Diseases Research Meeting

 W E D N E S DAY S C I E N T I F I C P RO G R A M
Wednesday, February 29, 2012
Research Discussion
026. H5N1
Wednesday, February 29, 2012 | 7:15 AM – 8:15AM
Palladian Ballroom
This newly added session at the ASM Biodefense and Emerging Dis-
eases Research Meeting will include discussions of the NSABB’s rec-
ommendations for the publication of the controversial H5N1 research.
This session will also provide an open forum for attendees to give
their feedback on such policy issues as the appropriate mechanisms
to allay public concerns about the safety of dual use research.
MODERATOR:
Paul Keim; National Science Advisory Board for Biosecurity Chair,
Bethesda, MD.
PRESENTATIONS:
NSABB Recommendations
Michael T. Osterholm, PhD., MPH; University of Minnesota School of Public
Health, Minneapolis, MN; Director, Center for Infectious Disease Research
and Policy (CIDRAP).
Government Response to the Recommendations
Anthony S. Fauci, MD; Director, National Institute of Allergy and Infectious
Diseases (NIAID).
Science’s Response to the Situation
Bruce Alberts, PhD; Editor-in-Chief of Science.
Perspective from a Scientist
Ron A.M. Fouchier, PhD.; Elasmus Med. Ctr., Rotterdam, The Netherlands.
Plenary Session
027. Immunity to Pulmonary Pathogens
Wednesday, February 29, 2012 | 8:30AM - 12:00PM
Regency Ballroom
The lung is a dynamic and relatively immunosuppressed organ. Fur-
thermore, immune responses in the lung are often dissimilar to those
that occur following systemic infection. Many pathogens successfully
use these features of the pulmonary compartment to their advantage
in establishing disease. Thus, understanding how pathogens take ad-
vantage of the lung environment will increase our ability to design
novel therapeutics and vaccines that target this important tissue.
MODERATORS:
Catharine M. Bosio; NIAID/Rocky Mountain Lab., Hamilton, MT.
Daniel Voth; Univ. of Arkansas for Med. Sci., Little Rock, AR.
PRESENTATIONS:
8:30AM
252 The Host Immune Response to Pulmonary Virus Infection: It’s
About Self Control
Thomas J. Braciale; Univ. of Virginia, Charlottesville, VA.
9:15AM
253 Requirements for Pulmonary Immunity against Virulent
Francisella tularensis
Catharine Bosio; NIAID/Rocky Mountain Lab., Hamilton, MT.
10:00AM
Coffee Break
10:30AM
254 Infection of Human Alveolar Macrophages by the Q Fever
Agent, Coxiella burnetii
Daniel Voth; Univ. of Arkansas for Med. Sci., Little Rock, AR.
11:15AM
255 The Protective Role of Surfactant Protein-A During
Mycoplasma pneumoniae Infection in Allergic and Non-Allergic
Airways
Julie G. Ledford; Duke Univ., Durham, NC.
BIODEFENSE PROGRAM & ABSTRACTS 31
NOTE S

32 10th ASM Biodefense and Emerging Diseases Research Meeting

 ABSTRACTS

majority of all respiratory infections were male (n=116) and age ≤ 4 (n=111).
Poster Session Amongst those cases associated with mortality (n=11), 7 were male and 7
008. Monday Posters were age ≤ 4. Conclusions: This laboratory surveillance data emphasized the
temporal impact of non-influenza respiratory viruses during the 2009 H1N1
Monday, February 27, 2012 | 1:00 PM - 3:00 PM Influenza Pandemic. Characterizing disease severity, notably mortality, aids
Exhibit Hall in discerning the potential consequences of a pandemic. As the foundation
of respiratory viral surveillance, state laboratories describe the comprehen-
027 (A) sive viral etiology to rapidly implement the necessary control and preventive
measures in case of disease outbreaks and pandemics.
Viral Acute Respiratory Tract Infections in Non-Winter Time
M. EL-Barrawy; Alexandria Univ., Alexandria, Egypt.
Introduction: Acute respiratory tract infection (ARTI) is the most frequent ill- 029 (A)
ness globally and a leading cause of death in the developing world. The most Natural History Studies for Ebola Virus Zaire Strain in Hartley
common cause of an ARTI may be one of over 200 viruses. However, bacte-
Guinea Pigs
ria or fungi can be its cause as well. Although viral respiratory infections are
A. Nalca, H. Bloomfield, S. Frederick, R. Baker, R. Williams, J. Yeager, E. Zum-
relatively considered harmless, in developing countries they are an impor-
brun; USAMRIID, Frederick, MD.
tant cause of mortality, especially when followed by bacterial complications.
Background: Ebola virus (EBOV) is a negative-stranded, enveloped RNA virus
The viruses which are mostly implicated are: adenoviruses, influenza viruses,
and a member of the Filoviridae family. It causes severe viral hemorrhagic
RSV and HSV. Although Respiratory viral infection is commonly peaked in
fevers with possible case-fatality rates of more than 90% depending on the
winter, it still has its heavy impact upon human health rest of the year. Aim
virus and virus strain. The fear of a bioterrorism event involving filoviruses
of the work: was to determine the prevalence of the previous respiratory
has increased the efforts of the scientific community in the development of
viruses in non-winter period among patients with ARTIs. Subjects & Meth-
countermeasures against these viruses. Currently, there are no licensed med-
ods: This study was carried out through the period from March 2010 to No-
ical countermeasures against filoviruses. Therefore, it is important to have a
vember 2010. It included 477 patients attending Alexandria Fever Hospital
small animal model for these viruses especially for aerosol route of exposure
and showing ARTI symptoms. All patients were recruited after having their
which could be highly infectious. Methods: In this study, we exposed two
consents and proper history taking. Throat swabs were collected, transferred
groups of 10 naïve Hartley guinea pigs to target doses of 10 PFU (low dose)
to Naval American Medical Research Unit 3(NAMRU 3) in Cairo, and tested
and 1000 PFU (high dose) of aerosolized guinea pig-adapted Ebola virus Zaire
using PCR techniques. Results: Out of 477 patients, the tested viruses were
strain (ZEBOV). Blood was collected on day -7 and then on days 3, 6, and 9
detected in 158 (33.1%) patients. Single virus infection was detected in
PE to test complete blood count (CBC) and serum chemistries. Results:
144(30.2%). This was as follows: 63(13.2%) influenza-A virus, 54(11.3%)
Guinea pigs started to show clinical signs of the disease including weakness,
adenovirus, 26(5.5%) RSV and one (0.2%) HSV. Meanwhile, 14(2.9%) had
fever, anorexia, and weight loss on day 4 post exposure (PE). While the high-
combined viral infection: 7(1.5%) had a combination of adenovirus and RSV,
dose exposure group started to show increased body temperature on day 4
6(1.3%) had a influenza A virus and adenovirus, and one patient (0.2%) had
PE, the low-dose exposure group did not have increased body temperature
influenza A virus, adenovirus and RSV. Conclusion: Viral infections represent
until day 5 PE. Interestingly, as the guinea pigs white blood cells (WBC), lym-
a substantial cause of ARTIs , even in non-winter time. Influenza A virus was
phocytes, and platelets decreased, their monocytes, basophils and
the most prevalent respiratory virus (14.7%). The viral ARTIs were more
eosinophils increased throughout the study. Increases in liver enzymes such
prevalent in children <15 years. There was no association between the res-
as alkaline phosphatase, alanine transferase, and aspartate transferase, as
piratory viruses’ infections and: smoking, occupation, temperature, and/ or
well as total bilirubin, were observed on day 6 PE. Furthermore, blood urea
existence of any chronic disease.
nitrogen (BUN) and creatinine levels were high in moribund animals. Low-
dose and high-dose exposure groups succumbed to disease on average at
028 (A) days 9.0 ± 0.48 and 8.3 ±1.0 PE, respectively. Conclusion: These data very
closely reflect observations of ZEBOV-infected non-human primates. There-
Laboratory Surveillance for Non-Influenza Respiratory Viruses fore, the guinea pig model for aerosolized ZEBOV is a useful tool to screen
in Maryland medical countermeasures against filoviruses.
W. A. Murtaugh1,2, J. Tan1, H. Peters2, B. Healey2, M. Tran2, M. Ziese2, B. An-
derson2, M. P. Carlos1,2; 1Johns Hopkins Univ., Baltimore, MD, 2Lab. Admin.
Maryland Dept. of Hlth. and Mental Hygiene, Baltimore, MD. 030 (A)
Background: State laboratories routinely identify the viral causes of respira- Experimental Respiratory Marburg Angola Virus Infection in
tory diseases. We provide a summary of non-influenza respiratory viral test-
Hartley Guinea Pigs
ing from the Maryland State Laboratory during the period of the 2009 H1N1
A. Nalca, H. Bloomfield, S. Frederick, R. Baker, R. Williams, J. Yeager, E. Zum-
Influenza Pandemic, and describe the implications of continual monitoring
brun; USAMRIID, Frederick, MD.
of circulating respiratory viral strains. Methods: Between 2008 and 2011,
Background: Marburg virus (MARV) is a causative agent of severe viral hem-
5014 influenza virus negative respiratory specimens were tested for respira-
orrhagic fever and classified as a category A biological agent due to its po-
tory syncytial virus (RSV), adenovirus (AdV), and parainfluenza virus (PIV) 1-
tential use as a bioweapon. The objective of this study to develop small
3 by cell-culture. For three overlapping influenza periods (2008-2009
animal model and characterize natural history of aerosolized Marburg virus
influenza season, 2009 H1N1 Pandemic Influenza, 2010-2011 influenza sea-
infection in Hartley guinea pigs (GPs). Methods: Two groups of 10 GPs were
son), the number of specimens tested, virus identified, and positivity rates
challenged by aerosol route with target doses of 10 (low) and 1000 (high)
were summarized. We performed an analysis of the positive specimens strat-
PFU of GP-adapted Marburg Virus Angola strain on day 0. . Blood was col-
ified by respiratory disease severity (out-patient, hospitalized, deceased),
lected on day -7 and then on days 3, 6, 9, 12, 15 and 18 post exposure (PE)
age and sex. Results: A total of 199 respiratory viruses were detected: RSV
to test complete blood count (CBC) and serum chemistries. Results: It was de-
(56), AdV (66), PIV-1 (30), PIV-2 (21), PIV-3 (26). The testing volume peaked
termined that average presented doses were 55 PFU for low-target dose and
in October 2009 (n=613). RSV and PIV displayed their common seasonal pat-
4400 PFU for high-target dose. Although they were sick and showed the signs
terns in the Northern Hemisphere, while AdV was detected intermittently. The

BIODEFENSE PROGRAM & ABSTRACTS 33

M O N D A Y P O S T E R S E S S I O N | 1:00–3:00 p.m. | Exhibit Hall

of the disease, five GPs from low-dose group survived the challenge with
aerosolized GP-adapted Marburg virus and all high-dose exposed group GPs
except one succumbed to disease by day 12 PE. Last GP from high-dose
group was moribund and euthanized on day 17 PE. Guinea pigs started to
show clinical signs of the disease including weakness, fever, anorexia, and
weight loss on day 6 PE\. While the high-dose exposure group started to
show increased body temperature on day 5 PE, the low-dose exposure group
did not have increased body temperature until day 6 PE. The white blood cells
(WBC), lymphocytes, and platelets decreased throughout the study. Except
survivors, increases in liver enzymes such as alkaline phosphatase, alanine
transferase, and aspartate transferase, as well as total bilirubin, were ob-
served on day 9 PE. Conclusion: These results suggest that Hartley GPs are
susceptible to aerosolized MARV therefore this animal model might be a use-
ful tool to screen medical countermeasures against filoviruses.
on October 13th while school attendance dropped on October 9th, reaching
a nadir on October 16th. ILI reporting peaked during the same week although
data was unavailable until the following Monday (Figure). Although all
datasets showed similar activity, school absenteeism reports were available
on a “same day” basis, whereas reported ILI and rapid antigen test results
were collated weekly, delaying availability. During the 2009 H1N1 outbreak,
school absenteeism and reported ILI were temporally related to ED patient
visit volume. Health officials may be able to monitor influenza activity by fol-
lowing school attendance and predict strain on healthcare resources.

031 (A)
A Cytopathic Effect-Based High-Throughput Screening Assay
Identified Four Novel Compounds that Inhibit Dengue Infection
K. D. McCormick; Univ. of Pittsburgh, Pittsburgh, PA.
To date, no approved vaccine or antiviral therapy exists for dengue virus
(DENV). Based on the observation that DENV infection of hepatocytes in-
duces massive cell death, “cytopathic effect (CPE),” we developed a high-
throughput screening assay(HTS) whereby inhibitors of DENV prevent cells
from dying. In this assay, the viral induced CPE is quantitated using a lu-
ciferase system for monitoring cellular ATP levels, which positively correlates
with cellular viability. This assay simultaneously screens possible inhibitory
compounds while controlling for any cytotoxicity triggered by these com- 033 (A)
pounds. Moreover, it is highly reproducible yielding a screening window co-
efficient, Z-factor, of 0.78±0.12. Our initial screening of a 288 NCI The Role of Rare Variants in Coronavirus Cross-Species Trans-
small-compound library yielded a total of eleven hits that prevented the CPE mission Events
of DENV infection. Further evaluation with an immunofluorescence assay M. Borucki, J. Allen, H. Harris-Chen; Lawrence Livermore Natl. Lab, Livermore,
showed that four of these compounds, streptovitacin A, nagilactone C, bac- CA.
tobolin and trichodermin, are highly potent inhibitors of DENV infection. At ef- A vast majority of the recently discovered human pathogens are viruses that
fective inhibitory doses, they did not appear to be cytotoxic, and they have an animal reservoir. The genetic diversity of a viral population within a
inhibited all four DENV serotypes albeit with varied efficacies. Interestingly, host may allow the virus to adapt to a diverse array of selective pressures
these compounds displayed strong inhibitory effects on hepatitis c virus (also and enable cross-species transmission events. Understanding the role of ge-
belongs to Flaviviridae family) infection, but exerted no or only marginal ef- netic variants within a viral population is a necessary step toward predicting
fect on new castle diseases virus. Altogether, our data show that these com- and treating emerging infectious diseases. Deep genome sequencing was
pounds are possible DENV antivirals. combined with computational analysis to characterize genomic changes in
coronavirus quasispecies during simulated cross-species transmission
events. Three bovine nasal samples naturally infected with bovine coron-
032 (A) avirus were used to infect human and bovine macrophage and lung cell lines.
Virus present in each sample was able to reproduce relatively well in
Predicting the Timing of Peak Pandemic Influenza Activity: ED
macrophages but not in the lung cell lines. Approximately 12 kb of the
Volume, School Absenteeism, and Community Reported ILI
genome was amplified before and after passage and sequenced. The con-
T. Bell1, M. Richardson2; 1West Texas Influenza Ctr., TTUHSC, Amarillo, TX,
2 sensus sequence of viruses passaged in bovine macrophages changed more
Amarillo Publ. Hlth. Dept., Amarillo, TX.
rapidly than those passaged in human cell lines, and the consensus se-
Although influenza is usually a self-limited illness, the disease burden and ra-
quences of spike gene varied the most of any region tested. Many of the pas-
pidity of spread during a pandemic can strain healthcare resources. Rapid
saged samples had a 12 nucleotide insert in the consensus sequence of the
escalation of cases makes monitoring difficult. Monitoring systems should
spike gene, and multiple point mutations were associated with the presence
be 1) available in “real time” 2) utilize existing data or require minimal addi-
of the insert. Deep sequencing revealed that the insert was present but very
tional work and 3) reflect local disease activity. In 2009, a novel influenza
rare in the unpassaged samples but could quickly shift to dominate the pop-
pandemic occurred. Most cases were self-limited, and the greatest strain on
ulation when placed in a different environment. Analysis of the subconsen-
healthcare resources in our community was in Emergency Departments (ED).
sus (quasispecies) data indicated that two distinct genotypes circulated at
As part of our public health response, we asked providers to voluntarily re-
different frequency levels in each sample, and support the hypothesis that
port influenza-like-illness (ILI) to the public health department (PHD). We
the mutations present in passaged strains were “selected” from a pre-exist-
also monitored school absenteeism and influenza testing results from local
ing pool rather than through de novo mutation and subsequent population
providers. We compared these data sets to ED volume to determine which
fixation.
best predicts peak influenza activity. The PHD monitored school attendance
on a daily basis. ILI visits to physicians were reported on a voluntary basis
from sentinel sites. Community providers provided information on influenza
test results. We obtained a daily ED census from hospitals. ED volume peaked

34 10th ASM Biodefense and Emerging Diseases Research Meeting


034 (B)
Transient Bacterial Lipopolysaccharide-Induced Resistance to
Inhalational Bacillus anthracis Infection in the New Zealand
White Rabbit Model
S. B. Yee, D. N. Dyer, N. A. Twenhafel, M. L. M. Pitt; USAMRIID, Fort Detrick,
MD.
Recent studies have demonstrated prior bacterial infection induces resist-
ance to subsequent infections from gram-negative and gram-positive
pathogens. In the present study, we evaluated whether underlying inflam-
mation would enhance host resistance to inhalational Bacillus anthracis in-
fection in New Zealand white (NZW) rabbits. Accordingly, NZW rabbits
(specific pathogen-free, Bordetella-free) were either pretreated with the in-
flammagen, bacterial lipopolysaccharide (LPS), a component of the outer
membrane of gram-negative bacteria, or saline vehicle (VEH). Administration
of this noninjurious dose of LPS (60,000 EU/kg) resulted in brief pyrexia and
a significant increase in the pro-inflammatory cytokine tumor necrosis fac-
tor-alpha confirming LPS-induced inflammation. Approximately 24 h later,
rabbits were exposed to aerosolized B. anthracis spores (Ames strain; ~300
LD50). Blood samples collected at various time points postchallenge were
cultured for bacteremia. LPS-pretreated, B. anthracis-exposed rabbits com-
pared to VEH-pretreated, B. anthracis-exposed rabbits exhibited significantly
delayed biomarkers of B. anthracis infection, anthrax-induced pyrexia (66 h
vs. 25 h postchallenge, respectively) and bacteremia (63 h vs. 26 h), and sur-
vived significantly longer (90 h vs. 41 h). As with the control animals, all LPS-
pretreated, B. anthracis-exposed rabbits exhibited pathology consistent with
succumbing to inhalational anthrax. No differences in histologic lesions —
including acute necrotizing splenitis, lymphadenitis of the mediastinal lymph
node, mediastinitis, pneumonia, adenitis of the adrenal gland, and renal tu-
bular necrosis—were observed between the two groups. Taken together,
these results suggest that prior or underlying stimulation of the immune sys-
tem induces transient host resistance to and/or delayed B. anthracis infection
in NZW rabbits. In particular, these results emphasize the importance of using
animals free of underlying infections to prevent confounding data in inhala-
tional anthrax characterization and therapeutic studies.
035 (B)
Small Quinolone Resistance Plasmids: A Model for Evolution
Mediated by Site-Specific Recombination (SSR) at oriT and Xer
Sites
T. Tran1,2, P. Andres3, A. Petroni3, A. Soler-Bistué4, E. Albornoz3, A. Zorregui-
eta4, R. Reyes-Lamothe2, D. Sherratt2, A. Corso3, M. Tolmasky1,2; 1California
State Univ., Fullerton, CA, 2Univ. of Oxford, Oxford, United Kingdom, 3INEI-
ANLIS, Buenos Aires, Argentina, 4Fundación Inst. Leloir, Buenos Aires, Ar-
gentina.
Qnr are proteins that mediate resistance to quinolones by protecting DNA
gyrase. The qnrB19 gene was found in several Enterobacteriaceae isolated
from numerous geographical regions within large plasmids, in ISEcp1C -
based transposons, and in small ColE1-type plasmids (about 3 kbp) lacking
ISEcp1C or any other insertion sequence. In spite of being located in such
dissimilar elements, the qnrB19 genes share a conserved genetic environ-
ment. We found 4 small plasmids harboring qnrB19, pPAB19-1, pPAB19-2,
pPAB19-3 and pPAB19-4 in a collection of clinical enterobacteria with reduced
quinolone susceptibility. Nucleotide sequencing and analysis showed that
they share extensive homology among themselves and with other described
small qnrB19-harboring plasmids. The genetic environment of qnrB19 in all
four plasmids is identical to that in those other plasmids and in transposons
such as Tn2012, Tn5387, and Tn5387-like. These plasmids as well as those
previously described showed a variable region characterized by being flanked
by an oriT locus and a Xer recombination site (RS). Both loci were confirmed
to be functional. We propose that this arrangement could play a role in evo-
M O N DAY A B S T R AC TS
lution of plasmids and present a model for DNA swapping between plasmid
molecules mediated by successive SSR events at oriT and the Xer RS. The
first event involves SSR at oriT, which depends exclusively on oriT site and the
nickase activity, and leads to integration. The cointegrate includes two di-
rectly positioned non-identical Xer RSs that may serve as substrate for a sec-
ond SSR event mediated by Xer that leads to cointegrate resolution. The final
products are the original plasmids but they have exchanged the fragments
flanked by oriT and Xer. We propose that the combination oriT-Xer RS could
be considered an element that facilitates plasmid evolution by swaping DNA
regions.
036 (B)
Antibacterial Activity of Alpha-MSH Alone and in Combination
with Other Antibiotics against Methicillin-Resistant Staphylo-
coccuus aureus
M. Singh, K. Mukhopadhyay; Sch. of Environmental Sci., Jawaharlal Nehru
Univ., New Delhi, India.
Background: We recently demonstrated that melanocortin peptide alpha-
Melanocyte Stimulating Hormone (α-MSH), possesses antimicrobial activity
against Staphylococcus aureus (S. aureus) and causes bacterial membrane
permeabilization. The present study was performed to investigate the in vitro
antibacterial potential of α-MSH against the methicillin resistant S. aureus
(MRSA) and synergy between α-MSH and four other antibiotics (oxacillin,
ciprofloxacin, gentamicin and rifampicin) against MRSA strain resistant to all
selected antibiotics and α-MSH. The study had been extended to examine α-
MSH induced morphological and ultra structural changes in MRSA and he-
molytic effect of α-MSH was also evaluated. Method: An ATCC MRSA and a
multidrug resistant S. aureus were used in this study. MIC of selected antibi-
otics was determined using broth microdilution method. The staphylocidal
activities of α-MSH (10-6 to 10-9M) was assessed by quantitative culture
after 2h bacterial incubation with α-MSH and compared to peptide free con-
trols. Synergism was checked using the above staphylocidal assay for differ-
ent antibiotics alone and in combination with α-MSH. Scanning and
transmission electron microscopy was employed to visualize the morpho-
logical and ultrastructural changes. The hemolytic effect was evaluated by
determining hemoglobin release of fresh rat erythrocyte suspension treated
with α-MSH at 414nm. Results: α-MSH killed 80.57%±2.01 and 49±1.08%
ATCC MRSA strain in micromolar and nanomolar concentration respectively.
80%±1.2 (p <0.05) increase in killing was observed in case of combination
of ciprofloxacin and α-MSH in comparison of either of agent alone. Similar
observations were also obtained for oxacillin and gentamicin. SEM and TEM
images had shown remarkable alteration in bacterial cell envelope. <10%
hemolytic effect was obtained in case of 10-5M of α-MSH. Conclusion: α-
MSH is strongly active against the MRSA and its synergistic action with other
antibiotics is indeed encouraging and provides a hope for MRSA infection
treatment.
037 (B)
Enhanced Aminoglycoside Sensitivity of Ribonuclease Mutants
of Escherichia coli
A. D. Frazier, W. S. Champney; East Tennessee State Univ., Johnson City, TN.
Background: An important issue for global health is the emergence of bac-
terial resistance to many current antibiotics. This raises the possibility of
using antibiotic resistance for bioterrorism. Consequently, new cellular tar-
gets for the existing antibiotics are needed. Ribosome biosynthesis is a novel
antibiotic target in bacterial cells and ribonucleases are essential to the pro-
cessing and turnover of bacterial rRNA. Previous studies have shown E. coli
strains deficient for specific RNases displayed an enhanced sensitivity to
azithromycin. It was hypothesized the inhibition of specific RNases in bacte-
rial cells would lead to an increased sensitivity to aminoglycoside antibiotics.
BIODEFENSE PROGRAM & ABSTRACTS 35
M O N D A Y P O S T E R S E S S I O N | 1:00–3:00 p.m. | Exhibit Hall
Methods: Ten RNase deficient E. coli strains were grown in broth in the pres-
ence or absence of 5μg/mL of neomycin or paromomycin. Viable cell counts
were determined after two cell doublings. The synthesis of 30S and 50S ri-
bosomal subunits in aminoglycoside treated cells was determined by sucrose
gradient centrifugation of 3H uridine labeled cell lysates. RNA from these
cells was analyzed by Agilent chip analysis and Northern blotting. Results:
Mutant strains deficient for RNase E, G, or PH were found to display an in-
creased sensitivity to the inhibitory effects of neomycin and paromomycin. In
each of these mutant strains, cellular viability was significantly reduced in
the presence of the aminoglycosides. In the RNase PH deficient cells, growth
with the antibiotics present reduced the synthesis of the 30S and 50S sub-
units by more than 30%. Agilent chip analysis revealed an accumulation of
the degradation products of rRNA for each mutant strain. Northern blotting
analysis showed a large increase in the breakdown of 16S and 23S ribosomal
RNA in strains deficient for RNase E, G, or PH in comparison with a wild type
control. Conclusion: This work showed that E. coli mutants lacking RNases E,
G, or PH, revealed an enhanced sensitivity to aminoglycoside antibiotics. This
work also suggests the inhibition of essential RNases may be a new drug tar-
get for small molecules or RNAi sequences.
038 (B)
A Model of Inhalation Glanders in African Green Monkeys
J. Yeager, P. Facemire, D. Nyakiti, L. Pitt; USAMRIID, Frederick, MD.
Burkholderia mallei, the causative agent of glanders, has been nearly eradi-
cated in industrialized nations, but recently has had renewed interest due to
The Center for Disease Control classification of it as a Category B select agent.
There is a lack of modern clinical cases resulting in limited knowledge of sus-
ceptibility of B. mallei to antibiotics and of its mechanism of virulence. There-
fore an animal model(s) that represent human glanders is needed to evaluate
the natural history and evaluation medical countermeasures. The goal of this
study was to evaluate African green monkeys (AGM) effectiveness as a model
for glanders. Ten AGMs were challenges to 147±90 LD50 of aerosolized B.
mallei (ATCC 23344). Telemetry acquired temperature was collected hourly
and clinical symptoms were monitored daily for 45 days. Each AGM had com-
plete blood counts, bacteremia and cytokines measured daily for the first
seven days post challenge (p.c) and then reduced to weekly. Gross pathology
and histopathology analysis were performed on all animals. A total of 7/10
AGMs succumbed to glanders between 6 and 32 days p.c. Days 1 to 3 post
challenge a fever (≥38.5°C) or increase in the diurnal temperature was ob-
served in 10/10 AGMs. On day 2 p.c. 10/10 AGMs had an increase in leuko-
cytes with an increased percentage of neutrophils. Bacteremia was detected
in 5/10 AGMs ranging from 5 to 45 days p.c. Lesions and nasal discharge
were common and typically manifested in the presence of a sustained fever
rather than a shift in diurnal temperature. Common gross lesions included
abscesses in the lungs and skin. Histologically, the abscesses were composed
of necrotizing to necrosuppurative inflammation. Additionally, this inflam-
mation was often identified in the spleen, bone marrow and lymph nodes.
The three surviving AGMs developed limb weakness to partial paralysis. Here
we describe a model of inhalation glanders in AGMs. Much of the clinical in-
dicators of infection are those that have been described previously in human
glanders. This data suggests that AGMs are a reasonable model for studying
glanders, making them valuable in therapeutic treatment studies and future
vaccine studies.
36 10th ASM Biodefense and Emerging Diseases Research Meeting
039 (B)
Genes Required for the Trafficking of S-Layer Proteins in the
Envelope of Bacillus anthracis
S-M. Nguyen-Mau1,2, D. Missiakas1,2, O. Schneewind1,2; 1Univ. of Chicago,
Chicago, IL, 2Howard Taylor Ricketts Lab., Argonne, IL.
Many Gram-positive microbes, including Bacillus anthracis— the causative
agent of anthrax—assemble proteins into a crystalline lattice on their sur-
face. S-layer proteins contribute to the envelope assembly of B. anthracis S-
layer associated proteins (BSLs), which play important roles during the
pathogenesis of bacterial infections and are known to bind to host cell re-
ceptors or scavenge heme-iron from host hemoproteins. Precursors of S-layer
proteins and BSLs harbor N-terminal signal peptides followed by three S-
layer homology (SLH) domains. In mature polypeptides, the SLH domains are
responsible for the non-covalent association of these proteins with pyruvy-
lated secondary cell wall carbohydrate (SCWP), thereby enabling their S-layer
deposition on the bacterial surface. Comparably little is known about the
genes and molecular mechanisms whereby S-layer proteins BSLs are se-
creted and traffic to their site of envelope assembly. Here we report that the
efficient deposition of S-layer proteins depends on a homolog of the secA se-
cretion gene, designated secA2, and a gene encoding acytoplasmic protein,
designated slaP (S-layer assembly protein). Mutations in secA2 or slaP re-
duce the secretion of S-layer proteins Sap and EA1, without affecting the se-
cretion of two S-layer associated proteins, BslA and BslO, or of protective
antigen, a secreted polypeptide. SlaP associates with the Sap and EA1 pre-
cursors in the cytoplasm of B. anthracis, suggesting that it may function as a
chaperone for S-layer protein secretion via SecA2.
040 (B)
Development of a Selective Enrichment Media for Brucella
Species Isolation
M. Perry, A. Kidney, M. Peck, C. Kelly-Cirino, C. Egan; NYS Dept. of Hlth., Al-
bany, NY.
Brucellosis is a zoonotic disease transmittable to humans usually through
direct contact with infected animal tissues/fluids or by the consumption of
contaminated, unpasteurized dairy products and meat. Due to the fastidious
nature of Brucella and high background levels of competing organisms, con-
ventional microbiological methods can take days to weeks for isolation and
confirmation. To prevent the growth of competing organisms, isolation usu-
ally requires selective media. Current diagnostic isolation recommends Far-
rell’s medium (FM) and Modified Thayer-Martin medium (mTM), however they
are non-selective and can inhibit the growth of several Brucella species. In
order to facilitate isolation, the Wadsworth Center (WC) has developed a se-
lective enrichment media for solid plate agar and broth cultures incorporat-
ing both selectivity as well as color identification of Brucella species. The
Wadsworth Brucella Selective Media (WBSM) was compared to mTM, FM,
CITA, Brucella agar (BruA) and 5% sheep blood agar plates (SBAP) for speci-
ficity and selectivity. All Brucella spp. exhibited growth on the WBSM, how-
ever both Brucella canis and neotome colonies appeared smaller on WBSM
compared to CITA. In addition, growth was not optimal for Brucella abortus,
strain RB51, on either media but the concentration was 225 CFU/100 μL with
SBAP. CITA and WBSM outperformed all other media when Brucella was
spiked into food matrices. An added benefit of WBSM is that it incorporates
neutral red with erythritol, creating easily identifiable pinkish colonies. Fur-
thermore, growth of other organisms including E. coli and Salmonella were in-
hibited when plated on the WBSM, demonstrating its selectivity. The
capability of WBSM can be increased when combined with the WC enrich-
ment broth (mF). After overnight enrichment of Brucella in mF, there was a
10 fold increase from 18 to 310 CFU/100 μL in a 103 CFU/mL sample of feta
cheese. A similar increase post enrichment was seen for other matrices, in-
cluding infant formula. This study demonstrates the ability of a newly devel-
 M O N DAY A B S T R AC TS

oped media and enrichment, WBSM and mF, to selectively identify Brucella
spp. and allow for more rapid patient treatment.
043 (B)
Burkholderia thailandensis is a Better Surrogate for B. pseudo-
mallei Growth and Persistence than B. pseudomallei Bp82
041 (B) M. A. Firmani, S. Lehman, S. Dieringer-Boyer, K. Bower, B. Dorsey, D. Stratton,
Periplasmic YopR Promotes Type III Needle Polymerization K. Delp, D. K. R. Karaolis; Natl. Biodefense Analysis and Countermeasures
B. Blaylock, O. Schneewind; Univ. of Chicago, Chicago, IL. Ctr., Frederick, MD.
Yersinia pestis, the causative agent of plague, depends absolutely on type Burkholderia pseudomallei is a Category B select agent. B. thailandensis is
III secretion to cause disease. An essential component of the type III secre- closely related and coexists in the same environment. However, B. thailan-
tion machine is a hollow filament that extends from the outer membrane, the densis is not a human pathogen and is a commonly used surrogate for B.
“needle.” The proteins that regulate and assemble needle polymerization are pseudomallei studies. B. pseudomallei Bp82 is a recently developed B.
the earliest substrates of the type III pathway. In Yersinia species, YopR con- pseudomallei 1026b purM mutant that is avirulent and has been exempted
tributes to needle polymerization by a mechanism that is under investiga- as a select agent. In this study, we performed growth kinetics and evaluated
tion. Mutations in yopR result in the abundant secretion of the needle shaft the persistence of both organisms grown in three media conditions. Survival
protein, YscF, but abolish the assembly of this structure. Complementation of over time at four temperatures was evaluated, including survival of log vs
a yopR mutation with alleles that are tagged at the protein’s carboxy termi- stationary cells. In addition, phenotypic comparisons (i.e., antibiotic sus-
nus are secreted but non-functional. YopR variants where the tag is at the ceptibilities, biofilm formation, motility, and protease production) were per-
amino terminus would be expected to obscure the secretion signal; surpris- formed. Our results demonstrate that B. thailandensis biofilm production in
ingly, these alleles are secreted and functional. Sub-cellular fractionation re- stationary phase cells was significantly (P<0.05) higher than log cells. In con-
vealed that only functional YopR alleles were present in the periplasm. trast, Bp82 did not produce a biofilm in log or stationary phase. A key find-
Artificial delivery of YopR to the periplasm by means of the Sec translocation- ing was that Bp82 is not able to grow well in certain media. Using LB-Lennox
dependent PhoA signal sequence also resulted in the restoration of needle media, we were able to perform growth and persistence assays with Bp82. In-
polymerization and was surprisingly re-engaged by the type III pathway. Col- terestingly, persistence studies showed log phase Bp82 cells survive better
lectively, these data suggest a role for YopR at the earliest stages of type III than stationary phase cells, which is in contrast to B. thailandensis. Pheno-
machine assembly at a time when it is still open to the periplasmic contents. typic assays with Bp82 were difficult to perform due to the mutant’s strin-
gent growth requirements. Bp82 would not generate a sufficient lawn of
growth on Mueller Hinton agar for antimicrobial susceptibility testing nor
042 (B) would it grow on skim milk agar for protease assay production. Our results
Virulence Analysis of B. cereus Biovar Anthracis demonstrate that Bp82 has a growth deficiency and that B. thailandensis
S. Dupke1, T. Franz1, A. Lander1, R. Grunow1, P. L. Goossens2, C. Brezillon2, possesses characteristics which are more similar to B. pseudomallei. Conse-
S. R. Klee1; 1Robert Koch Inst., Berlin, Germany, 2Inst. Pasteur, Paris, France. quently, B. thailandensis may be a better B. pseudomallei surrogate than
During the last years, several great apes died of an anthrax-like disease in Bp82 for growth and persistence studies.
West- and Central Africa. Molecular analyses and genome sequencing re-
vealed a close relationship of the new strains to B. cereus rather than B. an-
thracis on chromosomal level, even though both characteristic virulence
044 (B)
plasmids pXO1 and pXO2 are present. Therefore the African isolates were WITHDRAWN
designated as B. cereus biovar anthracis. Virulence analysis showed that all
isolates tested are able to produce the pXO1-encoded toxin components as
well as the pXO2-encoded polyglutamic acid capsule. Also in animal experi-
ments with mice and guinea pigs, the LD50 was similar to wildtype strains of
B. anthracis. Furthermore, comparable regulation of virulence factors was
confirmed for B. cereus bv anthracis and B. anthracis using reverse tran-
scriptase quantitative PCR. Under host-mimicking culture conditions in
medium containing bicarbonate and CO2, increased expression of different
genes encoding virulence factors or regulators (e. g. capB, acpA and pagA)
could be detected. Levels of enhanced expression were comparable in B.
cereus bv anthracis and B. anthracis and led to increased production of cap-
sule and toxins. Although the B. anthracis-specific nonsense mutation in the
gene for the transcriptional regulator PlcR is absent in B. cereus bv anthracis,
the regulator is also inactive due to a frameshift mutation in the plcR gene,
resulting in non-hemolytic growth like B. anthracis. In contrast to B. anthracis,
spores of B. cereus bv anthracis displayed no intracellular outgrowth into
vegetative bacteria or replication after infection of macrophages. The reason
for this feature remains speculative. Diagnostics of these new African Bacil-
lus isolates as anthrax-causing bacteria is hindered by the fact that they dif-
fer from typical B. anthracis both in microbiological (e. g. motility, resistance
to gamma-phage) and molecular (e. g. lack of prophage regions) features.
The emergence of the isolates as well as the source of animal infection remain
unclear.

BIODEFENSE PROGRAM & ABSTRACTS 37

M O N D A Y P O S T E R S E S S I O N | 1:00–3:00 p.m. | Exhibit Hall
045 (B)
Efficacy Testing of Orally Administered Antibiotics against an
Inhalational Bacillus anthracis Infection in BALB/c Mice
G. J. Hatch, S. R. Bate, J. Fretwell, J. Vipond, S. G. P. Funnell, A. D. G. Roberts;
Hlth. Protection Agency, Salisbury, United Kingdom.
Introduction: The efficacy of therapies against inhalational anthrax cannot
be studied in human clinical trials and must be evaluated in animal models.
In a series of three studies conducted in BALB/c mice we evaluated the effi-
cacy of post-exposure, orally administered antibiotics in preventing disease
following inhalational exposure to Bacillus anthracis. Materials & Methods:
Groups of female BALB/c mice were challenged with an aerosol of approxi-
mately 20 LD50s of B. anthracis, Ames. From one day post-exposure, once
or twice daily oral gavage of 0.2ml of either the test antibiotic or carrier con-
trol commenced. Treatment continued for thirty days after which the animals
were observed for a further fourteen days. The test antibiotics were lev-
ofloxacin, amoxicillin, co-amoxiclav (amoxicillin plus clavulanic acid),
azithromycin and clarithromycin; the carrier control was sterile distilled water.
Six mice from each group were killed and their blood processed to confirm the
treatment peak and trough serum levels. Throughout the studies the mice
were observed for symptoms of disease and a clinical score assigned. Cause
of death was confirmed by the presence of bacteraemia. Results: A summary
of the protective efficacy of the test antibiotics is presented in Table 1.
Table 1
Treatment Treatment % Median time Log-rank statistic
dose survival to death (days) compared to control
levofloxacin 80 mg/Kg BID 97 – p < 0.001
amoxicillin 30 mg/Kg QD 95 – p < 0.001
co-amoxiclav 30 mg/Kg QD 79 – p < 0.001
azithromycin 12-6 mg/Kg QD 5 1.92 p = 0.198
clarithromycin 30 mg/Kg QD 0 1.67 p = 0.815
control (water) 0.2 ml BID 3 1.92 – we evaluated protein expression of these atypical isolates at different phys-
Conclusion: The BALB/c mouse appears to be an appropriate model for the
evaluation of post-exposure therapies for inhalational anthrax. Orally ad-
ministered post-exposure prophylaxis over a 30 day period with levofloxacin,
amoxicillin or co-amoxiclav conferred significant protection from inhalational
anthrax whilst azithromycin and clarithromycin failed to protect against mor-
bidity or mortality in this murine model.
046 (B)
Human Anthrax Outrbreaks in Georgia in 2011
L. Malania1, N. Tsertsvadze1, N. Abazashvili1, M. Ramishvili1, M. Broladze1,
S. Tsanava1, N. Avaliani1, R. J. Obiso2, P. Imnadze1; 1The Natl. Ctr. for Disease
Control and Publ. Hlth., Tbilisi, Georgia, 2The Microbe Company, Christians-
burg, VA.
Anthrax is a bacterial disease caused by a Gram positive, rod-shaped, spore-
forming bacterium; a zoonosis that affects wild and domestic mammals. An-
thrax is a widely spread endemic infection in Georgia, which remains a
permanent threat to humans due to active foci. Within last five years (2006-
2010) 28-62 human cases of anthrax are registered annually. In 2011, 10
small outbreaks and 43 sporadic cases were reported. Clinical, laboratory
and epidemiological aspects of all sporadic cases and outbreaks were stud-
ied. Field trips were conducted in each affected area. Samples, collected from
all potentially exposed individuals and suspected contaminated areas were
screened for the presence of B.anthracis. Human cases were clustered and
analyzed by age, sex, location, source, laboratory investigation and case def-
inition. Data were entered and analysed by the Electronic Integrated Disease
Surveillance System (EIDSS). To date, 79 human anthrax cases were regis-
38 10th ASM Biodefense and Emerging Diseases Research Meeting
tered (incidence - 1.77 per 100 000); In addition, 10 outbreaks were de-
scribed, affecting 36 cases in total. The most common exposure risk was con-
tact with animals or animal products (82.27%). The infection had seasonal
distribution; a majority of cases (93.67%) occured during the summer and
early fall. All 79 cases were investigated (bacteriology, real time-PCR);
75.94% (60/79) were confirmed by Real-Time PCR and 44.3% (35/79) by bac-
teriology. Environmental samples were also studied to determine epidemio-
logical links. Laboratory investigation at NCDC was performed according to a
DTRA recommeneded algorithm. Data indicate that in Georgia during 2011
the incidence rate of anthrax in humans had significantly increased in com-
parison to that in previous years. We suggest that a very hot weather condi-
tion and an incomplete registration system for cattle and subsequent lack of
sufficient vaccination of domestic animals in the country had been reflected
in the presented data.
047 (B)
Proteomic Analysis of Atypical Y. pestis Strains Isolated from
Natural Foci in Georgia
R. Solomonia1, M. Nozadze11, E. Mikeladze1, E. Zhgenti2, G. Chanturia2, M.
Kekeklidze2, R. J. Obiso3, S. Francesconi4, L. Bakanidze2; 1Life Sci. Res. Ctr., Ilia
State Univ., Tbilisi, Georgia, 2The Natl. Ctr. for Disease Control and Publ. Hlth.,
Tbilisi, Georgia, 3The Microbe Company, Christiansburg, VA, 4Naval Med. Res.
Ctr., Frederick, MD.
Yersinia pestis is the etiological agent of plague. The National Center for Dis-
ease Control has evaluated 40 strains of Y. pestis from two foci regions in
Georgia and from foci of neighbor countries. Genetic analysis of these strains
was obtained for these isolates of Y. pestis; the data indicates that several
Georgian isolates are atypical from Y. pestis type strains. Alterations in Y.
pestis virulence factor gene expression are characterized by a shift in tem-
perature and calcium ions. This property of Y. pestis is determined by low-
calcium response (LCR) elements and all Georgian isolates of Y. pestis contain
this DNA sequence in spite of lacking other virulence plasmids. In this study,
iological conditions. Y. pestis was studied using 2-Dimensional (2-D) gel elec-
trophoresis and Mass-Spectrometry (MS). We selected four genetically
distinct Y. pestis isolates, (2944, 8787, 1853 and 1390), which were grown at:
(i) 280C; (ii) 280C +Ca2+; (iii) 370C and (iv) 370C +Ca2. Gels were stained and
protein spot maps and intensities were compared. Proteins that show con-
sistent, 2-fold differences in expression were excised and analyzed by Mass
Spectrometry to determine identity. Selected isolates of Y. pestis show sev-
eral differentially expressed proteins including: virulence-related outer mem-
brane proteins, IV secretory pathway VirB4 components, Peptidyl-prolyl
cis-trans isomerases, trigger factor, the F1 capsule antigen and others. At-
tempts will be made to integrate genomic and proteomic information to the
molecular biology of these isolates. These studies contribute to our under-
standing of the uniqueness of Georgian and Caucus region Y. pestis strains.
In addition, this research may influence the development of new diagnostics,
which can be applied to the Georgian surveillance program.
048 (B)
Clinical and Epidemiologic Assessment of Brucellosis in Georgia
T. Onashvili1, E. Mamisashvili11, R. J. Obiso2, M. Nikolaishvili, M. Zakareishvili,
I. Beradze; M. Donduashvili, M. Kokhreidze, N. Vepkhvadze; 1Lab. of Ministry
of Agriculture (LMA) Tbilisi, Georgia, Tbilisi, Georgia, 2The Microbe Company,
Christiansburg, VA.
Brucellosis, a bacterial disease caused by the genus Brucella, is a zoonosis
and significant cause of reproductive losses in animals. Brucellosis is widely
spread among the humans and animals in Georgia. Human infection occurs
from exposure to infected animals or animal products. In this study, we ana-
lyzed blood, serum, and milk samples from animals collected in Georgia using

an algorithm of tests, including PCR and AMOS PCR to detection Brucella
species. From July 2008 through October 2011 we collected approximately
9000 animal specimens seasonally (spring, fall) in three regions of Georgia:
Kakheti, Kvemo Kartli and Imereti. To date, 32 strains (B. abortus and B.
melitensis) have been isolated and are being characterized genotypically. A
majority of positive cases come from the Kvemo Kartli (50%) region, whereas
25% come from Kakheti and 16% from the Imereti region. This study en-
hanced the capacity of the Georgian Ministry to identify and conduct sur-
veillance for brucellosis in animal populations in Georgia. An accurate and
efficient surveillance system is of economic value and impacts the health of
the Georgian population. This study provided improved laboratory diagnos-
tics and case definitions of Brucella infection in Georgia.
049 (B)
Members of the FPT Subfamily of MFS Transporters are Impor-
tant for Pathogenesis of Francisella tularensis and Contribute
to Modulation of the Host Cytokine Response
1 2 1 1 1 1
M. Marohn , A. Santiago , K. Shirey , S. Vogel , E. Barry ; Univ. of Maryland
Sch. of Med., Baltimore, MD, 2Univ. of Virginia Sch. of Med., Charlottesville,
VA.
Francisella tularensis (Ft) is the causative agent of tularemia. Due to its
aerosolizable nature and low infectious dose, Ft poses a significant threat as
a potential bioterror agent, is classified as a category A select agent by the
CDC and, therefore, is a priority for vaccine development. Survival and repli-
cation in macrophages and other cell types is critical to Ft pathogenesis, and
impaired intracellular survival has been linked to reduction in virulence. The
Ft genome is predicted to encode 31 Major Facilitator Superfamily (MFS)
transporters, and the 9 member Ft phagosomal transporter (Fpt) subfamily
possesses homology to virulence factors in other intracellular pathogens.
Our hypothesis is that these MFS transporters will play a role in the intracel-
lular survival and replication of Ft and in host response to infection and may
serve as viable targets for attenuation and vaccine development. We demon-
strated altered intracellular replication kinetics and attenuation of virulence
in mice compared to WT Ft LVS in 3 of the 9 Fpt mutant strains. Importantly,
we showed that vaccination of mice with these mutants was protective
against lethal i.p. challenge. Here we present studies that characterize the
cytokine response to infection with Fpt mutant strains in comparison to
parental LVS. While the cytokine responses to infection with these mutant
strains are largely unaffected in murine macrophages when compared to the
response to LVS, there are pronounced differences in cytokine profiles in the
livers of infected mice. We believe that these alterations in in vivo cytokine re-
sponse are a major contributor to the attenuation observed in these strains.
Studies are currently underway to investigate further the cellular and mo-
lecular mechanisms that underlie the observed alterations in cytokine re-
sponses. These results confirm that this subset of MFS transporters plays an
important role in the pathogenesis of Ft and suggest a strategy for attenua-
tion towards the development of an efficacious vaccine.
050 (B)
Testing Diagnostic and Potential Therapeutic Values of Bacte-
riophages Capable of Lysing Yersinia pestis and Yersinia
pseudotuberculosis
A. A. Filippov, K. V. Sergueev, Y. He, M. P. Nikolich; WRAIR, Silver Spring, MD.
Several bacteriophages (phages) have been used for the diagnosis of plague
and pseudotuberculosis. Some of them could also be utilized for antibacter-
ial therapy of corresponding drug-resistant infections. There are very limited
data on comparative efficiency of plating (EOP) of phages capable of lysing
Yersinia pestis and Yersinia pseudotuberculosis. Such tests performed at
both diagnostic (28°C) and body (37°C) temperature would help to better es-
timate the diagnostic value of phages and their potential impact on normal
M O N DAY A B S T R AC TS
microflora when used as antibacterial therapeutics. The purpose of this work
was to test the efficiencies of plating of 11 bacteriophages on Y. pestis, Y.
pseudotuberculosis, and Escherichia coli at 28 and 37°C. They included four
plague diagnostic phages (Pokrovskaya, φA1122, L-413C, and Y), three
pseudotuberculosis diagnostic phages (D’Herelle-m, PST, and R), two well-
known coliphages (T7 and P2 vir1), one new isolate from sewage (φJA1), and
a T7 host range mutant (T7Yp). Only two phages did not form plaques on E.
coli: Pokrovskaya and φJA1. The rest of the phages could plate on E. coli K-12
albeit with lower efficiencies at 28°C than on the corresponding Yersinia
species. The EOP assays allowed the identification of phages with both high
diagnostic value and therapeutic potential, which included φA1122, L-413C,
Pokrovskaya, φJA1, Y, T7Yp (for plague), and d’Herelle-m (for pseudotubercu-
losis).
051 (B)
Biosynthesis of Lipoteichoic Acid in Bacillus anthracis
G. Garufi1,2, A. Hendrickx1,2, S. G. Richter1,2, K. Beeri1, J. Kern1, O.
Schneewind1,2, D. Missiakas1,2; 1The Univ. of Chicago, Chicago, IL, 2Howard
Taylor Ricketts Lab., Chicago, IL.
Many Gram-positive bacteria synthesize lipoteichoic acids (LTA), zwitterionic
polymers that are covalently attached to a glycolipid anchor moiety. In
Staphylococcus aureus and Listeria monocytogenes, the membrane protein
LtaS or its homologues are responsible for the synthesis of polyglycerol-
phosphate LTA from phosphatidyl-glycerol substrate. Mutations in the ltaS
gene(s) of S. aureus or L. monocytogenes cause defects in cell division that
interfere with bacterial growth. Bacillus anthracis, a spore-forming Gram-pos-
itive microbe and the causative agent of anthrax, has previously been re-
ported to lack teichoic acids. However, we identified four ltaS homologues,
here designated ltaS1-4, in the genome of B. anthracis. Using LTA-specific
monoclonal antibodies, LTA synthesis was observed in the envelope of B. an-
thracis vegetative forms. B. anthracis mutants lacking ltaS1, ltaS2 or ltaS3
did not display defects in growth or in polyglycerol-phosphate synthesis.
Deletion of ltaS4 led to the production of LTA molecules with aberrant mass.
B. anthracis strains lacking both ltaS1 and ltaS2 were unable to synthesize
LTA and exhibited reduced viability, diminished growth, altered envelope mor-
phology and aberrant cell division. Expression of some, but not all, B. an-
thracis ltaS genes in S. aureus complemented the growth and LTA synthesis
defects of an ltaS mutant. Further, expression of some B. anthracis ltaS ho-
mologues was sufficient to promote LTA synthesis in the Gram-negative mi-
crobe Escherichia coli. Thus, similar to S. aureus and other Gram-positive
microbes, B. anthracis employs LtaS enzymes to synthesize polyglycerol-
phosphate LTA, which is an essential component of the envelope that pro-
motes bacterial growth and cell division.
052 (B)
Resistance of Francisella to Fosmidomycin Associated with
Mutations in the Glycerol-3-Phosphate Transporter
R. S. Mackie1, E. S. McKenney2, M. L. van Hoek1; 1George Mason Univ., Man-
assas, VA, 2Univ. of Virginia Hlth. System, Charlottesville, VA.
The methylerythritol phosphate pathway (MEP) is present and is absolutely
required in most prokaryotes and some lower eukaryotes but absent from
human cells, and thus is an attractive pathway for antimicrobial drug devel-
opment. The formation of MEP is catalyzed by 1-deoxy-D-xylulose 5-phos-
phate reductoisomerase. The MEP pathway is critical for the formation of
isoprenoids, used for cell wall synthesis and electron transport. MEP pathway
genes have been identified in many Category A and B biothreat agents, in-
cluding Francisella tularensis, Bacillus anthracis, and Yersinia pestis. Fos-
midomycin inhibits DXR from F. tularensis subsp. holarctica LVS. This
compound also inhibits the growth of Francisella tularensis NIH B38, F. novi-
cida and F. tularensis subsp. holarctica LVS bacteria. We observed some
BIODEFENSE PROGRAM & ABSTRACTS 39
M O N D A Y P O S T E R S E S S I O N | 1:00–3:00 p.m. | Exhibit Hall
breakthrough colonies of F. novicida growing in the presence of fos-
midomycin, suggesting spontaneous resistance. Sensitivity of these colonies
to a lipophilic pro-drug of FR900098 was not abated suggesting that spon-
taneous resistance is not due to mutation of DXR. We predicted that the Glyc-
erol-3-Phosphate transporter (GlpT) which allows entry of fosmidomycin into
the bacteria would be defective, as we have shown that transposon insertion
mutants at the glpT locus are resistant to fosmidomycin. We performed DNA
sequencing of glpT from the fosmidomycin-resistant colonies and demon-
strated multiple mutations including nucleotide deletions and additions in
this gene. The frequency of mutation at the glpT locus in F. novicida that leads
to fosmidomycin resistance was determined to be 6.3 x 10-8. Thus we con-
clude that one mechanism of resistance of Francisella novicida to fos-
midomycin is caused by mutations in glpT. This is the first description that we
are aware of for spontaneous mutation in Francisella leading to fosmidomycin
resistance.
053 (B)
Control of Pigment Production in a White Phenotypic Variant of
Staphylococcus aureus
V. Antonic1, A. Stojadinovic2, M. Izadjoo1, M. R. Alavi1; 1Henry M. Jackson
Fndn. for the Advancement of Military Med., Gaithersburg, MD, 2Combat
Wound Initiative Program, Bethesda, MD.
Background: Difficult to treat Traumatic wound infections are usually caused
by multiple species of bacteria colonizing the wound bed. We isolated two
pigment variants of Staphylococcus from a wound sample, which displayed
white and yellow phenotypes. Yellow pigment is a manifestation of staphy-
loxanthin biosynthesis carried out by crt operon enzymes. Staphyloxanthin is
a virulence factor, which protects the microorganism against the host oxygen
free radicals and antimicrobial peptides. We used biochemical and molecu-
lar biological methods to characterize the white and yellow pigment variants.
As quorum signaling by Pseudomonas aeruginosa is shown to affect Staphy-
lococcus gene expression, we explored the potential influence of P. aerugi-
nosa on Staphylococcal pigment production. Results: Biochemical tests by
Phoenix instrument and 16s Ribosomal DNA sequencing indicated that both
pigment variants were Staphylococcus aureus with similar antibiotic resist-
ance profiles. However, staphyloxanthin production in the yellow variant was
2-fold higher than the white as measured by aceton/methanol extraction and
spectrophotometry. We examined the crt operon from the two pigment vari-
ants by sequencing. The promoter region and several biosynthetic genes that
have been sequenced to date are identical. We tested the effect of
Pseudomonas signaling on pigment production. Interestingly, P. aeruginosa
induces pigment production in the white S. aureus variant when they are in-
cubated next to each other on a plate. Conclusion: The yellow and white S.
aureus variants are similar in both biochemical and antibiotic resistance pro-
files; however they differ in staphyloxanthin production. Interestingly, the
white variant is capable of pigment production in the of presence P. aerugi-
nosa. This observation and our sequencing data suggest that crt operon is
functional in the white S. aureus variant but production of the pigment is con-
trolled by P. aeruginosa signaling. We are currently investigating the mecha-
nism of this control.
054 (B)
Interspecies Interaction of Francisella tularensis and
Pseudomonas aeruginosa
S. N. Dean, M. L. van Hoek; George Mason Univ., Manassas, VA.
The objective of this study is to elucidate the interspecies interactions of Fran-
cisella tularensis within the polymicrobial natural environment and within the
host. These results will improve our knowledge of the persistence of this or-
ganism in its natural habitat, and may identify factors that can promote or
inhibit growth of this fastidious organism. Pseudomonas aeruginosa was se-
40 10th ASM Biodefense and Emerging Diseases Research Meeting
lected as the model bacterium to study for interaction with F. tularensis, as it
is one of the most prevalent organisms in the soil-microbiome, a common
opportunistic human pathogen, and a species that F. tularensis could likely
encounter in the soil. We have shown that Francisella and Pseudomonas can
influence each other’s growth in vitro. P. aeruginosa produces HQNO (2-hep-
tyl-4-hydroxy quinoline-N-oxide) that causes a significant promotion of Fran-
cisella tularensis novicida growth and biofilm formation of 13 and 26%,
respectively (p < 0.01). Our hypothesis is that P. aeruginosa HQNO and other
small soluble factors may have specific effects on F. tularensis growth and
biofilm formation mediated through specific changes in Francisella gene ex-
pression in response to this interspecies signal. Quantitative assessment of
the effect of HQNO on F. tularensis growth and biofilm formation was per-
formed, including a titration curve of HQNO, where significant effects on
growth and biofilm were observed at 385 nM. Static Francisella biofilms and
biofilms under flow conditions were also examined. Further quantitative
analysis of F. tularensis biofilm was performed with COMSTAT using CLSM im-
ages of Francisella biofilms following HQNO treatment. We have shown that
P. aeruginosa produces a small molecule that significantly affects F. tularen-
sis novicida growth and biofilm formation. In future work, we will assess the
effect of HQNO on Francisella virulence and gene expression. This study sug-
gests a mechanism of this previously unknown interspecies interaction be-
tween F. tularensis and a common soil bacterium P. aeruginosa and may lead
to a greater understanding of F. tularensis persistence in the natural envi-
ronment.
055 (B)
A Method for Detection of Bacillus anthracis Spores in Complex
Matrices
O. Stephansson1,2, P. Ågren3,2, S. Byfors3,2, S. Ehrs1,2, M. Klint1,2, M. Lavan-
der3,2, C. Nilsson3,2, A. Lundin Zumpe3,2; 1Natl. Vet. Insitute, Uppsala, Sweden,
2
Swedish Joint Lab. for Food Safety and Biopreparedness, Uppsala, Sweden,
3
Natl. Food Agency, Uppsala, Sweden.
In 2004 the Swedish Laboratory for Food Safety and Biopreparedness was
established in collaboration between the National Food Agency (NFA) and
the National Veterinary Institute (SVA). These agencies share responsibility to
ensure a secure food chain and have a mutual interest to develop and sustain
diagnostic preparedness in case of unusual contamination of the food chain.
In times of globalization, climate change and as a result of increased aware-
ness, the ability for reliable analysis of unusual/antagonistic contamination
of the food chain is emphasized. This joint effort focuses on establishing
methods for sample preparation and molecular detection of BSL 3 agents in
food, fodder, drinking water and livestock tissue samples. Work has been fo-
cused on developing a diagnostic capability for Bacillus anthracis, an agent
that has largely been considered extinct in Sweden. Hence, the reoccurrence
of anthrax at a farm in 2008 and a recent outbreak in cattle during 2011 il-
lustrated the importance of upholding biopreparedness also for unusual
agents. The current project concerns molecular detection of B. anthracis
spores in complex matrices. A protocol for germination followed by extraction
of nucleic acids and detection by real time PCR has been developed and is
presented here. Limits of detection (LOD) for twelve matrices was determined
and the results show that due to the highly varying properties of the sam-
ples LOD ranges from 9 to 16 000 spores/g. Further, in some cases there was
no detection of B. anthracis, why sample preparation and/or the PCR proto-
col has to be optimized to enable detection in these matrices.

056 (B)
Comparative Genomic MGE Finding Process Streamlines Mobile
Element Search
S. Y. Choi1,2, N. A. Hasan1,2, J. Chun3, M. Hoq4, A. Huq5, T. A. Cebula6,2, R. R.
Colwell1,6,2; 1Univ. of Maryland, College Park, MD, 2CosmosID Inc., College
Park, MD, 3Sch. of Biological Sci. and Inst. of Microbiol., Seoul, Korea, Re-
public of, 4Univ. of Dhaka, Dhaka, Bangladesh, 5Univ. of Maryland, College
Park, MD, 6Johns Hopkins Univ., Baltimore, MD.
Horizontal gene transfer (HGT) is a dynamic process that generates diversity
and drives microbial evolution. Moreover, these mobile genetic elements
(MGEs) with key features including location, size, sequence content, as well
as genomic architecture, are important attributes enabling pathogens in the
environment to be tracked to their ultimate source. A comparative genomic
approach was used to predict, locate, and characterize MGEs in Vibrio
cholerae utilizing both ORF-dependent and ORF-independent approaches.
With a GI-definition of five or more contiguous ORFs, 130 MGEs ranging from
2.9 to105 kb were identified among 46 V. cholerae genomes. While all known
pathogenicity islands and regions associated with variable serotypes were
identified by this method, most MGEs were novel, and BLASTP searches of
ORFs against the nucleotide sequence database yielded no extraneous hits.
Such a large number of MGEs indicates that although the current pandemic
of V. cholerae is caused by strains belonging to a single phyletic line, they
have diversified by lateral gene transfer from a Vibrio gene reservoir in the
natural environment. Notably, distribution of MGEs in non-O1/O139 V.
cholerae strains is much more diverse than in O1 strains. Whereas in O1
strains the number of MGEs appears to plateau at approximately 40 MGEs
(analysis of 25 toxigenic V. cholerae O1 genomes), the number of MGEs dra-
matically increased to approximately 140 when non-O1/O139 strains were
added. From an evolutionary standpoint, V. cholerae non-O1/O139 appears
to be the ‘genetic harbinger’ of MGEs destined to appear in V. cholerae O1
strains. Thus, by monitoring the ‘genetic cauldron’ of non-O1/O139 strains,
it becomes possible to forecast elements that endow toxigenic V. cholerae
O1 strains with survival advantages for new and future hostile environments.
057 (B)
A Literature-Based System for Selecting Burkholderia pseudo-
mallei Strain Panels for Animal Model Testing Under the FDA
Animal Rule
K. E. Van Zandt1, A. Tuanyok2, H. C. Gelhaus, Jr.1, K. T. Brittingham1, S. A. Sar-
razine1, P. S. Keim2, R. L. Warren1; 1Battelle, Columbus, OH, 2Northern Arizona
Univ., Flagstaff, AZ.
Burkholderia pseudomallei is a gram-negative bacterial pathogen responsi-
ble for the disease melioidosis and is considered a biodefense threat. Current
treatment of melioidosis involves 2-4 antibiotics administered for up to six
months and even with treatment there is a 15% relapse rate. Therefore, there
is significant interest in developing improved medical countermeasures
(MCMs) against melioidosis. A variety of animal models, bacterial strains,
and experiment conditions have been used to study melioidosis, resulting in
disparate data and difficulty comparing studies. We have intiated the devel-
opment of animal models of melioidosis suitable for licensure of MCMs under
the FDA Animal Rule which requires consistent, well characterized etiologic
agent. This is difficult in the case of B. pseudomallei because the large
genome of B. pseudomallei is highly diverse among various isolates. To bet-
ter meet the FDA Animal Rule guidance and to assist in standardizing research
conducted across laboratories, we are developing a panel of strains that will
be used to achieve consistency among laboratories developing and evaluat-
ing melioidosis MCMs. To this end, we have reviewed published and unpub-
lished data on various strains of B. pseudomallei. We ranked each strain
based upon a scoring matrix which had five categories: animal models, viru-
lence, source, passage history, and availability of the strain to determine
M O N DAY A B S T R AC TS
which strains to include in the reference panel for future animal studies. Our
review resulted in the identification of 109 strains that met one or more of the
parameters in the scoring matrix. Based on our scoring system, strains
K96243, MSHR305, MSHR730, 1106a, S191, and MSHR668 scored highest
and are suitable for inclusion in the reference panel. These strains were iso-
lated from melioidosis patients in Thailand or Australia, have either yersinia
or B. thuringiensis-like flagella, and can be obtained from a source with a
known passage history.
058 (B)
Paratransgenic and Transgenic Analysis of Cellular Interac-
tions between Ixodes Ticks, Mammalian Cells, and Rickettsiae:
Lighting the Way with Fluorescent Proteins
T. J. Kurtti, J. D. Oliver, N. Y. Burkhardt, R. F. Felsheim, M. J. Herron, U. G.
Munderloh; Univ. of Minnesota, St. Paul, MN.
Ixodes ticks, most notably Ixodes scapularis and I. ricinus, harbor diverse mi-
crobial communities. Members include the more prevalent endosymbionts
known as the Rickettsial Endosymbiont of I. scapularis (REIS) and C. Midichlo-
ria mitochondrii in I. ricinus. In addition, these ticks serve as vectors for a
wide range of protozoan, bacterial and viral pathogens of humans as well as
domestic and wild animals. These include special interest agents such as
Rickettsia rickettsii and Francisella tularensis. We are using paratransgenic
and transgenic approaches to examine the interactions between rickettsiae
and hosts at the cellular level. Rickettsiae are transformed using constructs
that incorporate portions of plasmids found in Rickettsia amblyommii,
pRAM18, pRAM23 and pRAM32, and backbones of commercially available E.
coli plasmids. These shuttle vectors are engineered to contain a multiple
cloning site for facile insertion of genes to be tested, and antibiotic resist-
ance and fluorescent protein markers. Diverse rickettsiae indigenous to North
America have been transformed using these shuttle vectors and will be use-
ful for tracking rickettsial growth and development in ticks or tick and mam-
malian cell cultures. In addition, we have transformed I. scapularis cells by
adapting a method used to transform vertebrate cells, the Sleeping Beauty
transposase system. We transformed I. scapularis cell line ISE6 and Vero cells
to express a red-fluorescent peptide that binds to f-actin to examine rick-
ettsial motility. Live imaging of transformed REIS during colonization of tick
organs and dispersal in ticks suggests that there is co-evolutionary adapta-
tion between tick species and their symbionts which serves to restrict ex-
change of rickettsiae to other tick species.
059 (B)
Comparison of Challenge Outcomes with Three Different Bacil-
lus anthracis Isolates in New Zealand White Rabbits
A. Belle1, L. Schaefer1, A. E. Boyer2, M. Gallegos-Candela2, J. R. Barr2, M. de
la Garza3, J. Nunneley3, R. Carrion, Jr.3, K. Wycoff1; 1Planet Biotechnology Inc.,
Hayward, CA, 2CDC, Atlanta, GA, 3Texas BioMed. Res. Inst., San Antonio, TX.
Bacillus anthracis is a gram-positive spore forming bacterium that causes in-
halational anthrax. While rabbits are an accepted model for studying anthrax,
every laboratory has its preferred strain/isolate of B. anthracis and values re-
ported in the literature for time to death, bacteremia levels, and other dis-
ease parameters vary widely. In this study, groups of four New Zealand white
rabbits were challenged with an equal number of spores (2.1 x 107 CFU), by
the same route (nasal instillation), with one of three B. anthracis isolates and
the time to death was determined. The isolates were NR-3838 from BEI Re-
sources (derived from Ames), Ames from USAMRIID and Vollum from USAM-
RIID. Bacteremia was quantified at various times post-challenge. Other key
virulence factors in the serum were quantified, including Protective Antigen
(PA), Lethal Factor (LF), Lethal Toxin (LeTx) and Capsular Antigen (PGA or
Polyglutamic Acid). There were striking differences between the three iso-
lates in the rabbit challenge model. Time to death with the USAMRIID Ames
BIODEFENSE PROGRAM & ABSTRACTS 41
M O N D A Y P O S T E R S E S S I O N | 1:00–3:00 p.m. | Exhibit Hall
isolate was 50 (± 4) hr; NR-3838 was 76 (± 28) hr, but only 2 of the 4 rabbits
died; Vollum was 78 (± 4) hr. There were also large differences in the meas-
ured virulence parameters, with Vollum>Ames (USAMRIID)>NR-3838. Taken
together, our preliminary data suggest that the therapeutic window of op-
portunity is variable between the different B. anthracis isolates and that the
kinetics of inhalation anthrax in rabbits is isolate specific.
060 (B)
Genotyping of Mycobacterium tuberculosis Strains from a
Third Level Hospital in Lima, Peru
C. A. Tello, Sr., N. A. Borja, R. Garcia de la Guarda; San Marcos Univ., Lima,
Peru.
Objective: To characterize the genotypes of Mycobacterium tuberculosis by
two PCR based methods and begin an epidemiology study. Methods: Twenty
one patients diagnosed with tuberculosis (TB) between June and October
was included in the study. The genotypes of M.tuberculosis isolates were es-
tablished by DRE PCR (Friedman et. al., 1995) and PCR based Genotyping
(Kotlowsky et. al., 2004). The susceptibility to isoniazid, rifampicin, and strep-
tomycin was studied. Were studied the risk factors for active transmission of
TB. Results: A total of 21 M. tuberculosis isolates from samples of new and
retreated TB cases were collected at Guillermo Almenara National Hospital.
We registered a 66.7% of primary resistance and 33.3% of secondary resist-
ance. The 9.5 % of the patients were VIH+. The 76.2 % were in the range of
the productive age. When considered the patients with MDR strains the 80%
reported previous TB. The epidemiology and demographic data were col-
lected by clinical records. Upon DRE and PCR-based genotyping analysis of
21 M. tuberculosis strains, we reported the presence of 18 and 15 different
fingerprints respectively, the number of bands for each fingerprint ranged
from 8 to 1 band(s). In the consensus tree we established 3 clusters. The MDR
isolates were non-included in a specific cluster. Conclusions: Lima has a high
incidence rate of TB MDR. Although many clusters don’t have an epidemiol-
ogy link, the patients of these groups registered a stance in the same hospi-
tal area. The isolates present a high variability of genotypes and a low level
of clonality, possibly due to a long stay in time of different strains in Peru that
could be creating different stages of microevolution.
061 (B)
Evaluation of a Commercially Available Disposable Nebulizer
for Use in Nose-Only Pneumonic Plague, Tularensis, and Me-
lioidosis Murine Models
W. C. Lin, K. Siefkas, S-C. Hu, D. Sullivan, B. Gingras, L. Holland; IIT Res. Inst.,
Chicago, IL.
Objectives: Our objective was to evaluate the PARI LC Plus nebulizer, for the
development of murine models of infection for pneumonic plague, tularemia,
and melioidosis using IITRI’s nose only bioaerosolization system to evaluate
efficacy of experimental therapeutics and vaccines. The causative agents for
these illnesses used for testing the PARI LC plus nebulizer were the highly
contagious Gram negative organisms Yersinia pestis CO92, Francisella tu-
larensis SCHU S4, and Burkholderia pseudomallei ATCC 23342, which are
HHS/USDA Select Agents and NIAID Category A and B Priority Pathogens.
Methods: 100mL cultures of Y. pestis, F. tularensis, and B. pseudomallei were
grown in Heart Infusion Broth with 1% xylose at 28°C, Brain Heart Infusion
Broth with IsoVitaleX at 37°C in 5% CO2, or Trypticase Soy Broth at 37°C, re-
spectively. The bioaerosol system utilized a 64-port Cannon-style nose only
exposure chamber and six PARI LC Plus nebulizers operating at 20psi for
10minutes. Aerosol sizing was measured with an Anderson 6-stage impactor
while aerosol titer was measured with dual all-glass impingers. Determina-
tion of spray factor and assessment of aerosol consistency were accom-
plished through independent aerosolization of the three organisms with
dilutions at concentrations that ranged between 1E3 to 1E8 CFU/mL. Results:
42 10th ASM Biodefense and Emerging Diseases Research Meeting
Spray factors for Y. pestis and F. tularensis were calculated over runs at dif-
ferent concentrations to be 1.4E-5 and 6.9E-6, respectively. Preliminarily, the
spray factor for B. pseudomallei was determined to be 8.6E-7, although op-
timization studies are still in progress. The particle size distribution was
found to be within the respirable range (~2μm) for all three organisms tested.
Conclusion: The PARI LC plus nebulizer is an effective nebulizer for use with
Y. pestis and F. tularensis and provides good consistency and efficient trans-
mission of the bacterial agents through the exposure chamber across a broad
range of bacterial concentrations. Preliminary results suggest this nebulizer
is also effective for use with B. pseudomallei.
062 (D)
A PCR Platform Comparison Study to Support Sustainability in
Resource-Limited Settings
A. Ongarbaev1, A. Khodiev1, A. Smagin2, V. Garib2, G. Abdukhalilova2, G.
Ibadova2, R. Obiso3, S. Francesconi4, A. Baccetty5, J. Callahan6, D. Williams7;
1
Technology Management Company, Tashkent, Uzbekistan, 2Tashkent Inst.
of Postgraduate Med. Ed., Tashkent, Uzbekistan, 3The Microbe Company,
Christiansburg, VA, 4Naval Med. Res. Ctr., Silver Spring, MD, 5Defense Threat
Reduction Agency, Ft. Belvoir, VA, 6Biotechnology and Diagnostic Solutions
Animal Hlth. Life Technologies, Austin, TX, 7Threat Reduction Support Ctr.,
Alexandria, VA.
Background: The US Defense Threat Reduction Agency through its Biological
Threat Reduction Program, equips diagnostic laboratories with real-time PCR
platforms for the surveillance of especially dangerous pathogens within part-
ner countries. To this end, the existing PCR platform (Roche LightCycler 2.0)
provided to Uzbekistan is being replaced with a locally accessible and hence
more sustainable system (Qiagen Rotor-Gene Q). Prior to this change, we as-
sessed the potential impact of switching PCR platforms. Assays intended for
the LightCycler were tested on the Rotor-Gene. We evaluated and compared
sensitivity (Se) and PCR efficiency (E) between the two platforms to ensure
equivalence. Methods: Eight commercial and “in-house” PCR assays for vet-
erinary diseases were run in parallel on the two PCR platforms. A ten-fold di-
lution series of RNA- or DNA-positive controls were prepared for real-time
PCR. Each control and control dilution was treated as an unknown sample
and tested four times on each platform. A standard curve was constructed
for each test. Real-time PCR efficiency was estimated using the formula:
E=10^[-1/ΔC] -1. Results: The PCR assays tested retained sensitivity and ef-
ficiency on the Rotor-Gene platform within the established limits: Se - 100%
(relative to the LightCycler); and 90%<E<110%. Conclusion: The equiva-
lence observed between the two PCR platforms indicates that replacing the
LightCycler will not reduce performance capability in Uzbekistan. We suggest
the transfer of assays between different platforms supported by a relevant
comparison study to ensure sustainability.
063 (D)
Development of User-Defined Assays for the Genexpert Plat-
form
E. Felder, R. Wölfel; Bundeswehr Inst. of Microbiol., Munich, Germany.
One of the major tasks of the Department for Medical Bio-Reconnaissance
and Verification at the Bundeswehr Institute of Microbiology in Munich is
rapid and reliable identification of biological agents and other dangerous
pathogens even under restrictive field conditions comprising varying climate
and limited cooling capacity at missions abroad. The GeneXpert system from
Cepheid, available in Germany since a few years, combines sample process-
ing and real-time PCR detection from e.g. human samples in one cartridge,
saving time consuming and labour intensive working steps from specifically
trained laboratory personal. At this time point kits for the detection of bio-
logical agents in humans are not commercially available. Nevertheless
Cepheid offers an open platform for in-house development of cartridges from

third parties. The open GeneXpert platform was used at the Institute of Mi-
crobiology to develop cartridges for the detection of Yersinia pestis, Fran-
cisella tularensis, Brucella abortus and Burkholderia mallei from bacterial
cultures as well as Crimean Congo hemorrhagic fever virus from viral sam-
ples. PCR chemistry for real time PCR detection from bacterial (qPCR) and
viral (RT-qPCR) samples was especially compiled for freeze drying. Complete
PCR mastermix pellets which can be placed into the cartridges in one piece
could be generated for bacterial PCR detection. Software programs for sam-
ple preparation and PCR of each biological agent were separately written and
linked with the GeneXpert cycler. Using those ready-to-use software pro-
grams together with our in-house freeze dried real time PCR mastermixes and
the automatic sample processing from the GeneXpert system, we could sig-
nificantly reduce hands-on time for the end user. The freeze-drying method
for the generation of complete real-time PCR mastermixes as well as for a
bacterial sample processing control was well established and provides a
basis for the development of more assays for the detection of bacterial
agents. In summary, the GeneXpert system with its in-house designed car-
tridges is a suitable platform for the detection of biological agents and could
be used even in a low-resource laboratory environment.
064 (D)
Production and Qualification of Unrestricted Reagents to Sup-
port Anthrax Assays
K. H. Clement, T. L. Rudge, Jr., N. T. Huber, K. A. Newman, K. A. Sankovich, R.
H. Migliozzi, C. L. Sabourin; Battelle, Columbus, OH.
Thousands of clinical and non-clinical samples are tested each year in various
in vitro assays to assess responses to anthrax vaccines and therapeutics.
Under a National Institute of Allergy and Infectious Diseases (NIAID) contract,
we validated (for several specific intended uses) lethal Toxin Neutralization
Assays (TNA), the anti-Protective Antigen Immunoglobulin G (anti-PA IgG) En-
zyme Linked Immunosorbent Assays (ELISA) and PA ELISAs for multiple
species. These assays were validated using reagents provided by NIAID’s
reagent repository, BEI Resources, which cannot be used for commercial pur-
poses. Therefore, new reagents were recently produced, characterized, and
then qualified using BEI Resources’ reagents as the reference standard. Crit-
ical reagents including cell banks, reference standards and quality controls
were produced in large, stable lots and have been well-characterized to make
certain that the assays using these reagents remain in control for future use.
To date, rare reagents have been produced and qualified for various species
for the TNA and ELISA assays. Another critical component to ensuring that
these assays remain in control longitudinally is to monitor reagent stability
and functionality. The reagents are tracked in a custom-made Sample Track-
ing Database, which inventories all aliquots using unique bar codes and is
linked to a specialized BioAssay Database for tracking assay results. These
databases cross-talk to follow reagent location, freeze/thaw cycles, and to
trend reagent performance in the assays to monitor stability and quality. In
summary, these new, unrestricted reagents have been successfully bridged
to BEI Resources reagents, and show excellent performance in the anthrax as-
says for future testing of clinical and non-clinical samples. These reagents
are available for unrestricted use at Battelle.
065 (D)
Development of Rapid Diagnostic Platform for Detection of Cat-
egory A Biothreat Pathogens in the Field
P. Patel1, M. Weidmann2, K. Achazi1, S. Linke1, O. Strohmeier3, D. Mark4, T.
van Oordt4, J. Drexler5, M. Eberhard5, F. von Stetten4, M. Niedrig1; 1Robert
Koch Inst., Berlin, Germany, 2Univ. Med. Ctr., Göttingen, Germany, 3Dept. of
Microsystems Engineering (IMTEK), Freiburg, Germany, 4HSG-IMIT, Freiburg,
Germany, 5QIAGEN Lake Constance GmbH, Stockach, Germany.
Increased threat of infectious diseases and bioterrorist incidents pose dan-
M O N DAY A B S T R AC TS
ger to the human population. As a preparedness of such incidents, a rapid de-
tection system for identification of biothreat agents is needed. The S.O.N.D.E.
project intends to develop a suitcase size platform for the detection of cate-
gory A biothreat pathogens by nucleic acid and antigen detection. One sub-
project is dealing with the development of molecular methods such as
enrichment of pathogens and nucleic acid isolation. These assays are de-
signed to be used in an integrated bio-analytical system being developed by
our S.O.N.D.E. project partners for rapid detection of a number of pathogens
and toxins. To concentrate enveloped and non-enveloped viruses from large
volume biological samples, an enrichment method based on polymer was
developed. Viruses captured by polymer were successfully analyzed for nu-
cleic acid detection, virus infectivity assay and electron microscopy. Also the
nucleic acid purification using paramagnetic beads was successfully devel-
oped and tested on different model pathogens (gram+ and gram- bacteria,
RNA and DNA viruses). Purified nucleic acid was detected by real-time PCR
and an isothermal amplification method for different pathogens in spiked
samples and the limit of detections was 10-100 genome copies/ml. Cen-
trifugal microfluidic platform and portable device have been developed by
our project partners for the detection of biothreat pathogens in automated
“sample to answer”-platform. Developed methods like nucleic acid isolation
and isothermal amplification were integrated into this platform. Furthermore,
this platform is going to be evaluated and compared with conventional lab-
oratory methods as well as commercial available rapid diagnostic platforms.
066 (D)
Indirect Phage-Based Detection of Yersinia pestis: Field and
Clinical Applications
K. V. Sergueev, M. P. Nikolich, A. A. Filippov; WRAIR, Silver Spring, MD.
Lysis with specific bacteriophages is often used for the identification of
Yersinia pestis and plague diagnosis, but the assay usually includes the iso-
lation of pure Y. pestis culture and takes about three days. We have recently
developed an alternative, rapid and sensitive test of indirect Y. pestis detec-
tion based on quantitative real-time PCR (qPCR) monitoring of the amplifica-
tion of reporter phages φA1122 and L-413C (Sergueev et al., 2010; PLoS ONE
5(6): e11337). In this work, we tested potential field application of this
method. Using JBAIDS platform, a portable field-deployable instrument
(Idaho Technology Inc., Salt Lake City, UT), the phage detection limit observed
(and thus the limit of Y. pestis indirect detection) was identical to that ob-
tained using a full-sized LightCycler 2.0 real-time PCR instrument (Roche Ap-
plied Science, Indianapolis, IN) and was equivalent to one live bacterial cell
in four hours. We also explored potential clinical and epizootiological appli-
cations of this indirect detection method using simulated bovine serum and
mouse tissue (spleen, liver) samples. Omitting the step of sample preparation
and DNA concentration, satisfactory levels of sensitivity were observed when
using diluted spleen suspensions. The most promising results were obtained
with diluted and whole serum. These results indicate that the method of in-
direct qPCR-mediated phage-based detection of Y. pestis can be used with
sample types that would be tested under field or clinical conditions.
067 (D)
Quantification of Anthrax Lethal Factor by Mass Spectrometry:
Application to Clinical Anthrax
A. E. Boyer1, A. R. Hoffmaster1, C. P. Quinn1, M. Gallegos-Candela1, R. C. Lins2,
R. A. Stoddard1, E. Steward-Clark3, P. Maniatis1, C. Marston1, L. X. Cronin1, D.
Aranio1, H. Li1, V. Semenova3, K. Isbell2, S. Shadomy1, M. Lehman1, D. Blaney1,
N. Pesik1, T. L. Smith1, J. R. Barr1; 1CDC, Atlanta, GA, 2Battelle Analytical Serv-
ices, Atlanta, GA, 3CDC, Chamblee, GA.
The toxins of B. anthracis are major virulence factors for anthrax and con-
tribute to rapid disease progression by disabling host immune defenses. We
have developed and validated mass spectrometry (MS) based methods that
BIODEFENSE PROGRAM & ABSTRACTS 43
M O N D A Y P O S T E R S E S S I O N | 1:00–3:00 p.m. | Exhibit Hall
can detect and quantify anthrax lethal factor (LF) to measure toxemia during
treatment and provide early diagnosis of anthrax. The MS based method for
LF includes three steps that contribute to sensitivity and specificity: 1) LF spe-
cific mAb purification, 2) LF specific enzymatic proteolysis of a selected sub-
strate, and 3) detection of the toxin-specific hydrolysis products by
MALDI/TOF/MS yielding a detection limit of 5 pg/mL. LF levels measured in
rhesus macaques and NZW rabbits have shown an association with survival
and a point of no return. The LF-MS method was applied to human inhala-
tion, cutaneous, gastrointestinal and injection anthrax cases to confirm its
ability to detect and quantify toxins in these forms of anthrax and during an-
timicrobial and anthrax immune globulin (AIG) treatment. For three inhala-
tion anthrax cases, anti-PA levels measured by ELISA were consistent
following AIG delivery (133.49±0.135 μg/ml). LF levels appeared to correlate
with the route of exposure and disease form. The LF levels were highest in two
inhalation anthrax cases (327.59 and 57.985 ng/ml), followed by the geo-
metric mean concentration±standard error for 12 injection anthrax cases
(13.83±0.49 ng/ml), three gastrointestinal cases (0.525±0.692 ng/ml), and
83 cutaneous cases (0.117±0. 221 ng/ml). LF levels during treatment of a
human inhalation anthrax case in 2011 declined gradually with antimicrobial
and rapidly with AIG treatment, similar to that reported previously (Walsh et
al, 2007). LF quantification by MS was shown to be effective for toxin detec-
tion in all forms of anthrax and for monitoring effectiveness of anti-microbial
and antitoxin treatments.
068 (D)
Applying Elastic Light Scatter Technology (BARDOT) to Inte-
grate Organism Classification
H. Motlagh1, A. Giordano1, C. Waters1, E. Bae2, J. G. Thomas1, J. Robinson2;
1
West Virginia Univ., Morgantown, WV, 2Purdue Univ., West Lafayette, IN.
Introduction: BARDOT (BActerial Rapid Detection using Optical scatter Tech-
nology) distinguishes Forward Scatter Phenotype (FSP), showing altered bac-
teria not identified by traditional methods. We expanded our international
library for “pattern matching,” assessing the system’s ability to triage com-
ponents of Bioterrorism Surveillance: Phenotype/Bioburden, Mechanism of
Resistance, Institutional Surveillance, and Origin. Materials/Methods: BAR-
DOT system is comprised of interactive components: red laser reader, com-
puter, and software. The FSP detected by the laser diode module, integrates
into ELS (Elastic Light Scatter), a mathematical function of culture colony re-
fractive index, texture, size and shape/symmetry. Comparison of ELS was cal-
culated on Circulatory Symmetric Features and Texture patterns, extracting
107 features from one image. Dilutions of Organisms yielding 25 to 50 iso-
lated colonies were grown on Mueller Hinton, analyzed at 12, 24 hours.
Global clinical isolates obtained from South Africa, focused on MRSA, MSSA,
and ESBL producing E. coli and K. pneumonia. Results: Laser scatter images
via FSP were clustered based on ELS for 75 WVUH clinical isolates and 40
global isolates enlarging our “library” for “pattern-matching.” Multiple-Drug
Resistance Organisms had unique, identifiable patterns corroborated by
Cross Matrix Validation. MRSA (98.6%) could be segregated from S. epider-
mis (99.1%), ESBL E. coli (88.4%) segregated from non-ESBL producing E.
coli (99.6%) and ESBL positive K. pneumonia (97.4%) from ESBL-negative
(97.3%). Globally, matrix analysis showed segregation of WVUH MRSA
(90.3%) and MSSA (69.8%) from South Africa MRSA (95.7%) in MSSA
(93.2%). Based on Scatter Plot Matrix, we extracted ELS “pattern matching”
to Four-tiered analysis: Potential Bioterrorism Phenotype and Bioburden
(CFU’s/ml), Mechanism of Resistance, Phenotype Location (Institutional Sur-
veillance), and Infection Control. Conclusion: These give validity to a four
component Discussion Tree obtained simultaneously for clinical action, lo-
cally and nationally.
44 10th ASM Biodefense and Emerging Diseases Research Meeting
069 (D)
Negative Capture for Selective Enrichment of Pathogen-Derived
Nucleic Acids in Clinical Samples
Z. Bent, S. Langevin, V. VanderNoot, P. Lane, D. Curtis, M. Tran-Gyamfi, S.
Branda, T. Lane; Sandia Natl. Labs, Livermore, CA.
Second generation sequencing (SGS) has enormous potential for use in de-
tecting and characterizing novel, emerging, and engineered pathogens in clin-
ical samples with no a priori knowledge of the pathogen required. However,
brute force sequencing of clinical samples has limited utility because the
overwhelming majority of sequences map to the host and not the pathogen
genome. We have developed a technique for affinity-mediated depletion of
host-derived nucleic acids (NA) for efficient SGS analysis of pathogen-de-
rived NA in clinical samples. In this technique, which we call “negative cap-
ture” the NA from a clinical sample are hybridized with tagged probe
sequences that are complimentary to the host. NA that bind to the probes
can then be removed from the solution using the probe’s tag, thereby leav-
ing behind a population that is enriched for pathogen sequences. Negative
capture can be carried out manually using standard laboratory equipment, or
with less hands-on labor using a meso-fluidic chromatography device that
we have developed. In each case, negative capture increases the proportion
of SGS reads mapping to the pathogen genome, as compared to that yielded
by brute-force SGS of the original sample. Importantly, the additional “hits”
on pathogen are well distributed across the genome. Thus, negative capture
serves to target SGS to pathogen NA, and thereby improves sequence cov-
erage of the pathogen’s genome. This added information will prove to be very
valuable in characterizing the pathogen and distinguishing it from close rel-
atives. Further, due to its flexible nature, negative capture can easily be ap-
plied to pathogen sequencing from clinical samples of virtually any host.
070 (E)
Surveillance on Plague Natural Foci in Georgia
L. G. Bakanidze, P. G. Imnadze, S. A. Tsanava, N. S. Tsertsvadze; Natl. Ctr.
Disease Control and Publ. Hlth. of Georgia, Tbilisi, Georgia.
Georgia is located on the crossroad of Europe and Asia, it was one of the
chains of the Great Silk Road. It always was very vivid transition point, thus
it was not protected from spread of different epidemics, and, among them,
plague. Plague in Georgian manuscripts first was mentioned in XI century. In
XV century, later in another Georgian manuscript - “Book of Medical Treat-
ment-Ustsoro Karabadini” (“Peerless Karabadini”). Only single cases of
plague were mentioned in XVI - XVII centuries, official registration of plague
cases started in the XIX century. Several plague epidemics were registered in
Georgia during XIX century, they mainly started in the south Georgia, and
later spread on east, north, west and in Tbilisi. The Transcaucasian Anti-
Plague Center was founded in Tbilisi In 1933. Later existence of two natural
foci of plague on the territory of Georgia (plain-foothills, high mountainous)
was established. Georgian Anti Plague Station for decades carried out active
surveillance on natural foci. Several epizootics were described in both foci.
Totally 122 strains of Yersinia pestis were isolated in Georgia, nowadays 46
strains are kept at NCDC Microbial Library. After collapse of Soviet Union
under difficult economic situation in independent Georgia active surveillance
on plague natural foci was quitted, and only within the scope of US DoD De-
fense Threat Reduction Agency (DTRA)-funded CBR projects reviving it still
became possible. In 2005-2009 the CBR project GG-1 “Ecology, Genetic Clus-
tering and Virulence of Yersinia pestis Strains Isolated from Natural Foci of
Plague in Georgia” enabled us to start surveillance on plague foci, and it was
continued in next project GG-18 “Active Surveillance on Especially Dangerous
Pathogens in the Southern Caucuses Region.” Though now new isolates were
obtained during last years, and the plague foci in Georgia are so close to pop-
ulated areas, that they must be under permanent control, in order to be able
to rapidly response to emergencies. Besides, it gives us possibility to study

Y. pestis isolates from NCDC collection with modern sophisticated methods
of investigation.
071 (E)
Performance of Rapid Viability PCR Method for Detection of
Bacillus anthracis Ames Spores from Sponge-Stick, and Vac-
uum Sock and Filter Sample Types
S. R. Shah1, S. Letant2, G. Murphy2, T. Alfaro2, S. Kane2; 1EPA, Washington,
DC, 2LLNL, Livermore, CA.
The U.S. Environmental Protection Agency (EPA) in collaboration with the
Lawrence Livermore National Laboratory (LLNL) has been developing rapid vi-
ability testing methods for detection of biothreat agents in environmental
samples. Following an intentional or accidental anthrax event, hundreds to
thousands of environmental samples of diverse types could need to be rapidly
processed and analyzed in order to both characterize the extent of contami-
nation and determine the efficacy of remediation activities. The Rapid Viabil-
ity Polymerase Chain Reaction (RV-PCR) method which integrates high
throughput sample processing with real-time PCR, has been developed to de-
tect Bacillus anthracis Ames spore levels of 10 CFU/sample in wipe, air filter,
and water samples. The method was recently evaluated and optimized for ap-
plication to other sample types including vacuum socks and filters, as well as
sponge-sticks. The results indicated that the spore recoveries from these sam-
ple types were lower than those from wipe samples. Upon further optimiza-
tion, the RV-PCR method consistently demonstrated a limit of detection at the
level of 10 CFU/sample in the presence of 250 mg of Arizona test dust with
these samples, following an algorithm based on positive results with PCR as-
says targeting one gene each on the B. anthracis pXO1 and pXO2 plasmids.
Additional challenges including high levels of dead B. anthracis Ames spores,
and high levels of combined live Bacillus atrophaeus subspecies globigii
spores and Pseudomonas aeruginosa cells were also tested with vacuum sam-
ples. This work was performed under the auspices of the U.S. Department of
Energy by LLNL under Contract DE-AC52-07NA27344. The National Homeland
Security Research Center of the U.S. EPA funded this research.
072 (E)
An Operational Special Unit to Amend Biological Security
M. H. Richter, M-H. Lee, C. Herzog; Robert Koch Inst., Berlin, Germany.
As part of a national program to timely and efficiently respond to severe and
extraordinary biological events a special operational unit has been estab-
lished at the Center of Biological Security at the Robert Koch-Institute located
in Berlin, Germany. Among the events that can lead to an activation of the
unit are highly unusual disease outbreaks, bio-accidents, or bio-terroristic
activities. The unit is also involved in preventive counter measures as in the
run up to large public events or state visits. Its unique modular concept al-
lows timely and most appropriate adjustments to mitigate severe biological
threats or events. Such actions can span from theoretical support of federal
units by phone up to active engagement at the site of the event, sample ac-
quisition and on site diagnostics. The unit is therefore assembled of experi-
enced scientists in the field who have extensive know-how in handling and
collecting highly pathogenic agents in laboratory and field settings. A pivotal
part is the integration of laboratories with reference character and close co-
operation with forensic and crime fighting units. Developing sampling strate-
gies that can increase the likelihood of acquiring a positive sample in
contaminated areas are also part of the scientific project to improve the unit’s
performance further. Also, developing and implementing of field diagnostics
is a central part within the unit’s framework. Moreover, frequent training and
consecutive skill, fit, and health tests of the unit leaders and staff assures
physical and psychological preparedness of the unit. Momentarily more top-
ics are subsequently added to complement the unit’s modular structure.
Upon completion of its modular character, the unit will be able to react to a
M O N DAY A B S T R AC TS
biological threat in a very precise manner by activating only the modules that
are useful to cope with a given situation in the most possible appropriate
way. This modular approach at the same time creates the most economic way
of response in a crisis or as a preventive measure.
073 (E)
The Microfluidic Bioagent Autonomous Networked Detector (M-
BAND): An Update
M. Sanchez, L. Probst, E. Blazevic, B. Nakao, M. A. Northurp; Microfluidic
Systems, Fremont, CA.
We will update the status of the M-BAND: a fully automated air-borne bio-
threat detection system for biosurveillance applications and its internal assay
capabilities. The system, including a multiplexed, nucleic-acid-based detec-
tion assay, was designed and built with DHS Science and Technology funding,
and shipped for field testing by Microfluidic Systems Inc (MFSI), a subsidiary
of PositiveID Corporation (PSID). Field testing was performed by third par-
ties in the US and Europe. The system and assay identifies pathogenic strains
of Bacillus anthracis, Yersinia pestis, Francisella tularensis, Burkholderia
mallei, Burkholderia pseudomalleiand Variola major virus. In order to assess
the assay’s ability to detect unknown samples, our team also challenged it
against a large and diverse series of blind samples provided by the Depart-
ment of Homeland Security (DHS). These samples included natural occurring
isolated strains, near-neighbor isolates, and environmental samples. Our re-
sults indicate that the multiplex assay was specific and produced no false
positives when challenged with in house gDNA collections and DHS provided
panels. The use of a multiplex assay format saves considerable time and cost,
while providing critical sensitivity and specificity. Herein, we present a viable
analytical tool for the rapid identification of six Centers for Disease Control
and Prevention category A and B biothreat organisms, either in an automated
field-based system, or in a laboratory setting.
074 (E)
Investigation on Drinking Water Quality of the Bogra Sadar Up-
azila: Findings from a Rapid Assessment Lesson in the Bogra
District of Bangladesh
M. Mollik1,2; 1Peoples Integrated Alliance, Dhaka, Bangladesh, 2PreSci. Trust
Funds, Phoenixville, PA.
Bangladesh is a low-lying, riparian country located in South Asia with a
largely marshy jungle coastline of 710 kilometers on the northern littoral of
the Bay of Bengal. Bangladesh’s alluvial soil is highly fertile but vulnerable
to flood and drought. Most of the people in Bangladesh are using ground
water as a source of safe drinking water. The present investigation assesses
the chemical and bacteriological quality of drinking water in several differ-
ent locations of the Bogra Sadar upazila (sub-district) in Bangladesh. The
samples were collected from supply water and tube-wells water, and ana-
lyzed for Arsenic (As), Chloride (Cl), Calcium (Ca), Iron (Fe), Lead (Pb), Mag-
nesium (Mg), pH, Phosphate (PO4), total coliform, total hardness, and total
viable bacteria. The Calcium (Ca), Magnesium (Mg), Phosphate (PO4), and
total hardness concentrations were found to be within the permissible limits.
In contrast, 13% of the water samples contained Arsenic (As), 8% of the sam-
ples contained Chloride (Cl), 18% of the water samples contained Iron (Fe),
and 5% of the water samples contained Lead (Pb) more than the permissible
limits. In addition, total coliform bacteria were found to be present in almost
all samples of water ranging from 60%-100%. Total viable bacteria were
found in 69% samples which exceeded the permissible limits recommended
by the World Health Organization. This investigation indicates that the water
used for drinking in many locations of the Bogra Sadar upazila (sub-district)
is unsafe for human consumption due to both bacterial and chemical con-
tamination. Tube-well water of this sub-district should be treated before using
for drinking purposes.
BIODEFENSE PROGRAM & ABSTRACTS 45
M O N D A Y P O S T E R S E S S I O N | 1:00–3:00 p.m. | Exhibit Hall
075 (E)
Multi-User Validation of a Real-Time PCR System for Detecting
Bacillus anthracis Collected by Air Sampling Devices in a Liq-
uid Matrix
U. K. Spaulding1, K. M. Clemens1, T. Hadfield2, V. Ryan2, S. Brunelle3, I. M.
Ota1; 1Idaho Technology, Inc., Salt Lake City, UT, 2MRIGlobal, Inc., Palm Bay,
FL, 3AOAC International, Gaithersburg, MD.
The RAZOR™ EX Anthrax Air Detection System, a real-time PCR system de-
veloped by Idaho Technology, Inc. (ITI), has undergone testing in accordance
with the harmonized Performance TestedSM Method (PTM) - Official Methods
of Analysis (OMA) validation process established by AOAC INTERNATIONAL
(AOAC). This system was developed for the US Department of Homeland Se-
curity Bioinformatics and Assay Development 2 program which is directed at
establishing standards and third-party validation protocols to evaluate bio-
threat-detection tools. The RAZOR Anthrax System detects and identifies vir-
ulent Bacillus anthracis spores present in samples from air collection devices
via the detection of three targets: 1) the pXO1 plasmid, 2) the pXO2 plasmid,
and 3) B. anthracis chromosomal DNA. An “Anthrax Detected” result is ob-
tained when all three targets are detected. The validation process included
evaluation by ITI, who is the method developer (MD), by MRIGlobal, an in-
dependent laboratory (IL) and by twelve independent collaborators. Based on
the results of the MD and IL validations, a PTM certificate was conferred in Oc-
tober, 2011. The work presented here describes the collaborative testing per-
formed according to the Biothreat Agent Method guidelines established by
AOAC. Twelve independent collaborators, four at each of the three partici-
pating MRIGlobal laboratories, participated in the study. A simulated liquid
air collection matrix was tested, spiked with B. anthracis at 2000 spores/mL
or Bacillus cereus, a near neighbor, at 20,000 spores/mL. Each collaborator
tested a total of 24 randomized blind-coded samples, including 12 B. an-
thracis-spiked samples and 12 B. cereus-spiked samples. Currently, the col-
laborative testing results are being reviewed. We expect that the RAZOR
Anthrax System will meet the requirements for OMA status as its perform-
ance has been robust through all previous testing.
076 (F)
A Hemorrhagic Fever Virus Sequence Database and Analysis
Platform
C. Kuiken; Los Alamos Natl. Lab., Los Alamos, NM.
Hemorrhagic fever viruses are a diverse set of over 80 viral species from nine
different genera and five viral families: arena-, bunya-, flavi-, filo-, and to-
gaviridae. All these viruses are highly variable and evolve rapidly, making
them elusive targets for the immune system and for vaccine and drug design.
Presently about 50,000 HFV sequences have been published. A central web-
site that provides annotated sequences and analysis tools will be helpful to
HCV scientists worldwide. The HFV sequence database collects and sequence
data, performs automated quality control and annotation, and provides them
to the public via a website that contains a user-friendly search interface and
a large number of sequence analysis tools, following the model of the highly
regarded and widely used Los Alamos HIV database. Offering similar capa-
bilities for viruses from so many different taxonomical classes presents ad-
ditional challenges. Much of the database’s infrastructure depends on
computer code that can place any sequence in the proper location relative to
a central reference sequence. Because of the diversity of these viruses, a ref-
erence sequence needs to be chosen for every species. The NCBI RefSeq
database is used for this purpose; for sequences that are not assigned to a
genus or species, a reference sequence is chosen using a similarity search.
The sequences are internally aligned to a genus-level Hidden Markov align-
ment model using HMMer; the location of the gaps is also stored, and upon
retrieval gaps are inserted where necessary to maintain the alignment. The
HFV website can be accessed via http://hfv.lanl.gov.
46 10th ASM Biodefense and Emerging Diseases Research Meeting
077 (F)
Ethical & Social Issues Related to Synthetic Biology: What We
are Doing as Academicians?
S. T. Hakim, S. M. H. Tayyab, S. G. Nadeem; Jinnah Univ. for Women, Karachi,
Pakistan.
Synthetic Biology (SB) is the design and construction of new biological parts,
devices, and systems, and the re-design of existing, natural biological sys-
tems for useful purposes. Finding the right balance between formal statutory
regulation and self-regulation of the scientific community remains a con-
tentious issue and will occur only after the risks of synthetic biology are more
widely understood and debated. A questionnaire based cross sectional sur-
vey. Total 25 young researchers of post graduate level in Karachi were sur-
veyed at two Universities of the city. Main outcome measures was
perceptions on the basic principles of Synthetic Biology. All participants were
females with mean age of 26+ 2 years, and enrolled for MS/ M.Phil. and Ph.D
dissertation. They responded that they were not aware of social and ethical
issues related to synthetic biology before coming into the research field; even
they never heard this term in this scenario before. A significant number of re-
spondents agreed that there is a need of creating awareness about benefits
and risks related to synthetic biology. It was also recommended that we have
to educate the community of students and teachers, especially those who
are involved in research to take guidance, and approval from their institu-
tional ethical review boards (if exist!) before start of any research study. Few
more issues were also analyzed including use of laboratory animals, using
clinical specimens without informing patients, sickness certification, human
cloning, and disclosure of information to different parties without prior per-
mission of the subject and publication of sensitive research. Conclusion: A
code of ethics and standards should emerge for biological engineering as it
has done for other engineering disciplines in both research and academic in-
stitutions of the country and all the concerned should forced to follow the
rules properly.
078 (F)
Identification and Molecular Characterization of Novel Group C
Orthobunyaviruses Isolated from Mosquitoes Captured in Peru
L. L. Lofts, M. J. Turell, J. R. Kugelman, G. I. Koroleva, G. F. Palacios, C. A.
Whitehouse; USAMRIID, Frederick, MD.
Members of the genus Orthobunyavirus (family Bunyaviridae) are seg-
mented, negative-sense, single-stranded RNA viruses that are responsible
for mild to severe disease in humans. As part of a long-term field study of ar-
bovirus ecology in the Amazon basin of Peru, more than 160 viral isolates
were identified in mosquitoes captured near Iquitos, Peru. Preliminary anti-
gen detection analysis classified some of these Peruvian viral isolates as ei-
ther Group C or Bunyamwera group orthobunyaviruses. Entire deep
sequence genome alignments confirmed the classification of the bun-
yamwera viruses; however, based on the S segment, the Peruvian viral iso-
lates classified as Group C orthobunyaviruses failed to group with the Group
C viruses isolated from mosquitoes captured in Brazil. To further character-
ize the Group C orthobunyaviruses, deep sequence genomes were deter-
mined for the Brazilian Group C mosquito viral isolates. The results provide
insights for the intra- and inter-reassortment among the orthobunyavirus
RNA segments (S, M, and L).

079 (F)
Development of a High-Throughput Bioinformatics Typing Sys-
tem for the Phylogenomic Analysis of Infectious Category A
and B Agents
M. Eppinger, S. Agrawal, S. Daugherty, K. Galens, A. Maroo, L. Tallon, H. Tet-
telin, C. Fraser-Liggett, J. Ravel; Inst. for Genome Sci. (IGS), Univ. of Mary-
land, Sch. of Med., Baltimore, MD.
Background: Infectious diseases outbreaks impose a significant public health
threat and the understanding of the genomic plasticity of the underlying bac-
terial agents is crucial to the epidemiology, surveillance, diagnostic and foren-
sic communities. We used the human pathogens Y. pestis and E. coli O157:H7
as model systems both of which present with severe and potentially life-
threatening complications in infected patients. Genomes were screened for
mutations and structural variants to assess the genome polymorphisms and
outbreak dynamics. Methods: Next generation whole-genome sequencing
followed by phylogenomic polymorphism discovery analyses in the con-
served genomic backbone as well as the dissected mobilome allowed track-
ing the pathogenome evolution. To investigate the phenotypic variations on
the level of individual polymorphisms, we have developed a high-through-
put bioinformatics pipeline for SNP discovery and validation taking into ac-
count the coverage and quality of underlying sequence reads. The SNP
pipeline is part of Ergatis, a web-based utility that is used to create, run, and
monitor reusable computational analysis pipelines. Results: Phylogenomic
analyses revealed numerous novel SNPs previously not available for the typ-
ing of these pathogen and other structural biomarkers many of which seem
to be intimately associated with their niche-adaptation, pathogenesis and
outbreak assignment. Conclusions: The detected polymorphisms in strains
acquired during routine laboratory cultivation and within strains derived from
a single outbreak suggest the need to study of outbreak populations rather
than rely on single outbreak-type strains. The established high-resolution
phylogenomic frameworks achieved refined a phylogenetic accuracy and res-
olution and aids current molecular assays used in public health laboratories
in order to better prepare for future outbreaks and better risk assessment in
response to novel emerging virulence phenotypes.
080 (G)
Humanization of Anti-Ricin Neutralizing Monoclonal Antibody
W. Hu, J. Yin, L. Negrych, D. Chau, J. Cherwonogrodzky; Defence R & D, Med-
icine Hat, Canada.
Ricin is regarded as a high terrorist risk for the public due to high toxicity
and ease of production. Unfortunately, there is currently no therapeutic or
vaccine available against ricin. The murine monoclonal antibody D9 can
strongly neutralize ricin and is a good candidate for the development of a
humanized antibody. Humanization of D9 variable domains was achieved by
a complementarity-determining region grafting approach. The humanized
D9 variable domains were further grafted onto human heavy and light chain
constant domains to assemble the whole antibody gene. Meanwhile, a foot-
and-mouth-disease virus-derived 2A self-processing sequence was intro-
duced between heavy and light chain DNA sequences to express a
full-length antibody from a single open reading frame driven by a single
promoter in an adenoviral vector. After expression in mammalian cells and
purification, the humanized antibody was demonstrated to have equimolar
expression of full-length antibody heavy and light chains and to retain anti-
gen-binding specificity and neutralizing capacity comparable to its parent
D9. This humanized D9 has potential use for prophylactic or therapeutic
purposes against ricin poisoning.
M O N DAY A B S T R AC TS
081 (G)
Inhibition of Lipoteichoic Acid Synthesis in Staphylococcus au-
reus and Bacillus anthracis
S. Richter1,2, A. P. A. Hendrickx2, O. Schneewind2,1, D. Missiakas2,1; 1Howard
Taylor Ricketts Lab., Argonne, IL, 2Univ. of Chicago, Chicago, IL.
Staphylococci and many other Gram-positive bacteria incorporate zwitteri-
onic lipoteichoic acid (LTA) into the cell wall. The polymer consists of a back-
bone of repeating glycerol-phosphate units decorated with D-alanine and a
glycolipid anchor. Membrane protein lipoteichoic acid synthase (LtaS) cat-
alyzes the biosynthesis of the polymer by transferring the polar head group
of phosphatidylglycerol to the growing chain of polyglycerophosphate. Mu-
tations in S. aureus ltaS cause severe cell division defects and trigger cell
death at 37°C. A two-step screening protocol was designed to identify small
molecule compounds that inhibit growth of staphylococci by disrupting LTA
biosynthesis. High-throughput screening for growth inhibition was conducted
with chemical libraries. Strong hit compounds were further tested for their ef-
fect on LTA biosynthesis at sub-lethal concentrations. One compound was
identified as LTA inhibitor for staphylococci. The inhibitory activity was con-
firmed with a heterologous system that utilizes S. aureus LtaS for synthesiz-
ing LTA in E. coli. We also demonstrate that the compound is active against
Bacillus anthracis and Enterococcus faecalis.
082 (G)
Discovery of Targeted and Broad Spectrum Antiviral Counter-
measures for Alphaviruses
D. Chung1, B. Beitzel2, J. Arterburn3, J. O. Tret1, J. Smith2, C. Schmaljohn2, G.
Gene Olinger2, C. Scully2, C. B. Jonsson1; 1Univ. of Louisville, Louisville, KY,
2
U.S. Army Med. Res. Inst. of Infectious Diseases, Fort Detrick, MD, 3New
Mexico State Univ., Las Cruces, NM.
Venezuelan (VEEV), eastern (EEEV), and western (WEEV) equine encephali-
tis viruses are positive strand RNA viruses of the genus Alphavirus, family To-
gaviridae, that are endemic throughout the Americas. Naturally transmitted
by mosquitos from bird and rodent hosts, alphaviruses cause severe disease
and life-threatening encephalitis in humans and equines. Alphaviruses rep-
resent significant potential biological defense threats and are classified as
Category B priority biodefense agents. There are limited therapeutics avail-
able to treat diseases that they cause. Our primary efforts focus on identify-
ing small molecules targeting the alphaviruses by leveraging high throughput
and virtual screening. In our current pilot screening for VEEV, 384-well cell-
based assays using a luciferase-reporter for V3526, and a CPE-based assay
for TC83 and/or VEEV 3000 were used to screen against the Prestwick Library
and a targeted library of 658 active compounds identified as actives in a
screen for bluetongue virus. Tranilast was identified as an active compound
and prioritized for further characterization of structure-activity. A set of 14
compounds based on an analysis of the functional groups and overall struc-
tural correlations to tranilast were made, but no improvement in potency was
observed. The drug diclofenac was also identified with a potent antiviral ac-
tivity (EC50= 37 μM, SI50 >3). While the EC50 is high, diclofenac is a nons-
teroidal anti-inflammatory drug (NSAID), and although its mechanism of
action is not confirmed, it has been shown to block voltage-dependent
sodium channels and acid sensing channels along with its activity as a COX
inhibitor. Current plans include testing this compound in a mouse model chal-
lenged with VEEV. In summary, we have screened more than 350,000 com-
pounds experimentally and over 1,300,000 compounds virtually, and are
continuing to analyze the efficacy of those showing promise.
BIODEFENSE PROGRAM & ABSTRACTS 47
M O N D A Y P O S T E R S E S S I O N | 1:00–3:00 p.m. | Exhibit Hall
083 (G)
Generation of Biologically Active Nano-Aerosol by an Electro-
spray-Neutralization Method
V. N. Morozov1, I. L. Kanev1, N. K. Balabaev1, A. V. Glyakina1, M. L. van Hoek2;
1
RAS, Moscow, Russian Federation, 2George Mason Univ., Manassas, VA.
Biological nano-aerosols promise tremendous opportunities in therapy of
pneumonic lung infections (eg tularemia), lung diseases and in vaccination.
However, generation of nano-aerosolized protein (e.g. antibodies or vaccines),
DNA, antibiotics and other drugs meets serious problems caused by their
fragility. We describe a simple method for manufacturing biological nano-
aerosols under mild conditions. It is based on electro-hydrodynamic atomiza-
tion of drug solutions followed by a gas-phase neutralization of a cloud of thus
generated highly charged nano-clusters and ions with a cloud of small oppo-
sitely charged ethanol ions also generated by electrospraying. We demon-
strated that enzymes aerosolized by this mild technique retained their specific
activity, which testifies that protein molecules are not notably damaged during
the process, despite the two highly energetic processes involved: electro-
spraying and gas-phase neutralization. To further evaluate possible damaging
factors in the technology we simulated process of gas-phase neutralization of
dry and hydrated multi-charged protein ions with small counter-ions (OH- and
NO3-, both dry and hydrated). Our computer simulations have revealed that
the hydration layer covering the surface of protein ion remarkably protects pro-
tein structure by channeling the impact energy to the water molecules. Thus,
while the impact of a small dehydrated single-charged counter-ion with a de-
hydrated lysozyme macroion with 8 uncompensated charges resulted in a short
local heating of 20-25 protein atoms by more than 700 K, only 5 atoms were
heated in a similar impact of the same ion with a hydrated lysozyme macroion.
These studies suggest that proteins retain their structure and function upon
nano-aerosol generation and may be used in therapy or as inhaled vaccines.
084 (H)
H5N1 Avian Influenza Specific Preventive Maintenance in the
Russia
D. Surnev, Yu. Kuznetsov, T. Okovytaya, V. Yelnikov; JSK Pokrov Biological
Plant, Pokrov, Russian Federation.
The analysis of epizootic situation indicates a real threat of a wide spread of
avian influenza on the territories of different states. The main carriers of avian
virus are wild migratory ducks. Avian influenza virus is reproduced in wild
ducks’s intestines and is excreted with excrements. As a result, the infection
spreads in water reservoirs, where from it can be contracted through domes-
tic ducks and geese to other poultry. With poultry the avian influenza virus can
progress as latent infection causing only decrease in productivity, as well as
devastating epizootic, accompanied by high death rate (up to 100 %). In the
Russian Federation the infection of avian influenza has not been registered for
a long time. Its outbreak occurred for the first time in Novosibirsk region in
July, 2005. The results of molecular-biological studies have shown the agent
belongs to high-virulent subtype H5N1. The disease was accompanied with
high death rate, up to 100% in poultry. At the end of 2005 on the instructions
of the Ministry of Agriculture of the Russian Federation the Russian scientists
developed the technology of vaccine production of H5N1subtype avian in-
fluenza vaccine. The preparations made on the basis of the virus strains, ho-
mologous to epizootic possess the highest protective potential. Thus, a virus
isolate isolated during the outbreak in Novosibirsk region as a vaccine mate-
rial was implemented. After successful carrying out of laboratory and field
tests, inactivated emulsified vaccine against H5N1avian influenza made on the
basis of strain homologous to the one spread in the Russian Federation, has
been introduced into veterinary practice. Since 2006 avian influenza vaccine
vaccine against H5N1avian influenza has been widely and successfully used in
the Russian Federation for specific preventive maintenance of the avian in-
fluenza in private farming and peasant household.
48 10th ASM Biodefense and Emerging Diseases Research Meeting
085 (H)
Immunogenicity and Efficacy of a Recombinant Plague Vaccine
(rF1V) in Swiss Webster Mice
P. Fellows1, J. Price1, W. Lin2, A. Rom2, S. Marshall3, L. Holland2, L. Wolfraim1;
1
Dynport Vaccine Co., LLC, Frederick, MD, 2IIT Res. Inst., Chicago, IL, 3BioStat
Solutions, Inc., Mount Airy, MD.
Objective: The objective of this study was to determine minimum protective
titers in Swiss Webster mice vaccinated with recombinant F1V (rF1V) and sub-
sequently challenged by aerosol exposure to Yersinia pestis CO92 to support
vaccine licensure strategy under the Animal Rule. Methods: Groups of Swiss
Webster mice were vaccinated by intramuscular injection on Days 0, 14, and
28 with dosages of the rF1V vaccine ranging from 0.0032 to 16 μg in a 100 µL
volume. Control mice received saline using the same schedule and route of
administration. Serum anti-rV, anti-rF1 and anti-rF1V titers were determined
on Day 43 by Bridge ELISA. Mice were challenged on Day 44 by aerosol ex-
posure to a lethal dosage of Y. pestis CO92. Results: A dose response curve
was obtained with respect to Bridge ELISA titers and probability of survival
as well as vaccine dosage and probability of survival. Logistic regression mod-
eling shows a statistically significant relationship between increased levels
of anti-rV, anti-rF1 and anti-rF1V Bridge ELISA titers and increased probabil-
ity of survival (p<0.0001) with anti-rF1V titers the most highly associated
with protection. Following challenge, the highest dosage of the vaccine
tested, 16 µg, elicited 87% protection. The 0.08 µg dosage of the rF1V vac-
cine was the lowest dosage providing protection (25%) following challenge.
All control mice died following challenge. There was also a statistically sig-
nificant relationship between vaccine dosage and probability of survival
(p<0.0001). The protective titer and protective vaccine dosage at 50% sur-
vival were estimated at 225 U/mL of anti-rF1V and 2.32 µg of rF1V vaccine,
respectively. Conclusions: The rF1V vaccine, when administered in a three-
dose schedule, provides protection in a dose-related manner to Swiss Web-
ster mice following aerosol exposure to Y. pestis. Minimum protective titers
were established and are an important component in bridging protective cor-
relates between species.
086 (H)
Ribosomal Protein TgP0 in the Invasion of Host Cells by Toxo-
plasma gondii Tachyzoites
S. Rajagopal, S. Sharma; Tata Inst. of Fundamental Res., Mumbai, India.
Apicomplexan parasites such as Plasmodium, Toxoplasma, Theileria,
Neospora and Eimeria are highly successful parasites with a wide range of host
specificities. A key event in the establishment of an infection is host cell inva-
sion. Proteins on the parasite surface mediate interactions to enable host cell
recognition and consequently, invasion. Earlier work from our lab, aimed at
identifying immunoprotective antigens of the malarial parasite, established
the ribosomal phosphoprotein PfP0 as a potential protective antigen. The can-
didature of the ribosomal P0 protein as a protective protein has been subse-
quently validated by passive immunization and in vitro growth inhibition
studies. Antibodies to P0 blocked invasion of both Plasmodium falciparum as
well as Toxoplasma gondii, and P0 protein was found to localize to the surface
of the merozoites and tachyzoites, respectively. The ribosomal P0 protein lacks
any classical secretory signals for localization to the surface or secretory or-
ganelles of the parasite. To identify such signals, we have generated truncated,
epitope-tagged constructs for transient transfection of Toxoplasma gondii. Ad-
ditionally, post-translational modifications of the protein might also act as trig-
gers for differential localization during different stages of growth. We are
characterizing post-translational modifications of parasite P0 by 2D gel elec-
trophoresis and Mass Spectrometry. Specific antibodies to Toxoplasma gondii
P0 have been raised and these are being used in suitable invasion inhibition as-
says to establish the point where P0 might mediate its role in the timeline of
invasive events. The results of these experiments will be discussed.

087 (H)
Advancing Clinical Grade RV10-2 Plague Vaccine Toward Licen-
sure
L. E. F. Quenee, N. A. Ciletti, D. Elli, T. M. Hermanas, O. Schneewind; Univ. of
Chicago, Argonne, IL.
Yersinia pestis causes plague, a zoonotic disease with high mortality rates
that can be transmitted by fleabite or aerosol. A US Food and Drug Adminis-
tration (FDA)-licensed plague vaccine is currently not available. Vaccine de-
velopers have focused on two subunits of Y. pestis: LcrV, a protein at the tip
of type III secretion needles, and F1, the fraction 1 pilus antigen. The rV10-2
vaccine is a non-toxigenic variant of LcrV lacking residues 271-300. We de-
veloped Current Good Manufacturing Practice (cGMP) protocols for rV10-2
and demonstrated its efficacy against lethal pneumonic plague challenge
with wild-type or F1 mutant Y. pestis in mice, rats, guinea pigs, cynomolgus
macaques and African Green monkeys. To further support FDA Investigational
New Drug (IND) authorization review and clinical testing for rV10-2, the vac-
cine was subjected to GLP long term stability testing. Clinical grade vaccine
vials were stored under controlled temperature conditions and analyzed for
stability over the course of 24 months. A GLP toxicology study in New Zealand
white rabbits was performed to determine preclinical safety in preparation
for a future Phase I clinical trial.
088 (H)
Tularemia Vaccine Trials in Nonhuman Primates
R. Tempel1, R. Shirley2, A. Legasse1, F. Heffron1, M. Axthelm1; 1Oregon Hlth.
& Sci. Univ., Portland, OR, 2Virogenomics, Inc., Portland, OR.
Francisella tularensis is the gram-negative bacterial pathogen that causes tu-
laremia, a debilitating and potentially fatal disease. Inhalation of as few as
ten bacteria can produce a severe pneumonic illness in humans. Despite its
high infectivity, classification as a category A select agent, and use as a bio-
logical weapon, there is currently no FDA-approved vaccine against tularemia.
Our previous studies identified several Francisella genes that, when mutated,
give rise to attenuated strains in cell culture and mouse models of infection.
Deletion derivatives of two of these strains, ΔdsbB and ΔpdpB, in Francisella
Schu S4 provided protection against wild-type challenge in mice, indicating
the potential of these genes to be included in a live attenuated vaccine. Here,
we describe testing these strains in the nonhuman primate (NHP), rhesus
macaque. Groups of three NHPs were inoculated subcutaneously with ΔdsbB
(108 CFU) or ΔpdpB (109 CFU). Four weeks after the primary inoculation, the
NHPs were boosted with 1010 CFU of the respective attenuated strain. Thirty-
three days post-boost, 104 CFU of wild-type Schu S4 were inoculated into the
lung. Two NHPs that received only vehicle during the primary and boost in-
oculations were concurrently challenged with 104 CFU of wild-type Schu S4.
The challenge dose was chosen based on our results that intrabronchial in-
oculation of 104 CFU resulted in tularemia-related death nine and 18 days
after infection. In contrast, naïve NHPs infected with 100 CFU survived until
the end of the study (day 23). During the study, blood, lymph node, bone
marrow, and bronchoalveolar lavage samples were obtained periodically to
determine the bacterial load, and O2 saturation, CBC panels, enzymatic func-
tion, and protein levels were monitored to assess the overall health of the
NHP. We will report these results, along with the gross pathology of the or-
gans after the NHP has succumbed to disease or upon necropsy at the end
of the study (28 days post-challenge). While there have been several studies
describing protective strains of Francisella in mice, these are the first such
studies in a NHP model of infection.
M O N DAY A B S T R AC TS
089 (H)
A Clinical Phase II Study Confirming the Safety and Immuno-
genicity of One or Two Doses of IMVAMUNE (MVA-BN) Small-
pox Vaccine in Vaccinia-Experienced Elderly Subjects
R. N. Greenberg for the POX-MVA-024 Sites1, P. Chaplin2; 1Univ. of Kentucky
Sch. of Med., Lexington, KY, 2Bavarian Nordic GmbH, Martinsried, Germany.
Background: IMVAMUNE® (MVA-BN®) is a live, highly attenuated Modified
Vaccinia Ankara virus vaccine formulated at a dose of 1 x 108 TCID50 / 0.5 ml.
IMVAMUNE® does not replicate in human cells and is in clinical development
as a 3rd generation smallpox vaccine. Methods: A Phase II study was con-
ducted to evaluate safety and immunogenicity (using both ELISA and PRNT)
of one and two doses of IMVAMUNE® smallpox vaccine in 56-80 year old vac-
cinia-experienced subjects (n= 120). Subjects received either two injections
of 0.5 ml IMVAMUNE® or one injection of 0.5 ml Placebo and one injection
of IMVAMUNE® four weeks apart. Results: Vaccinations were well tolerated
by all subjects. No serious adverse events related to IMVAMUNE® and no
cases of myopericarditis were reported. The overall incidence of unsolicited
AEs was similar in both groups. A second dose did not increase reactogenic-
ity. ELISA as well as PRNT results were comparable for the two groups
IMVAMUNE ELISA PRNT
Group N/Seroconversion Rate GMT/Seroconversion Rate GMT
(individual peak titers)
2 doses 61/90.2% 992.4/95.1% 257.6
1 dose 58/84.5% 645.2/77.6% 139.6
Response rates and seroconversion rates measured by PRNT increased after
a second dose. A second dose increases GMTs in the ELISA and to a higher ex-
tent in the PRNT. Conclusions: One or two doses of IMVAMUNE® were safe
and immunogenic in the 56-80 years old vaccinia-experienced population.
Safety, reactogenicity and immune responses are similar to that seen in the
younger (18-55 year old) healthy population as investigated in other trials.
The results indicate that in an emergency situation it is sufficient to vacci-
nate this population only once. Supported by Contract No. DMID-N01-AI-
40072.
090 (H)
LC16m8, an Attenuated Smallpox Vaccine Will Be a Useful
Countermeasure against Bio-Terrorism with Smallpox
H. Yokote1, Y. Shinmura1, C. Uemura1, T. Kanehara1, S. Maruno1, H. Matsui1,
K. Ohkuma1, K. Yokoi1, A. Funatsu1, S. Gordon2, G. Franchini2, S. Hashizume3;
1
Kaketsuken, Kumamoto, Japan, 2Natl. Cancer Inst., Bethesda, MD, 3Univ. of
Chiba, Chiba, Japan.
Background: During the current situation where numbers of patients with
some sort of immune impairment such as eczema, HIV-positive, etc. are in-
creasing, safer smallpox vaccines than the conventional ones are being de-
sired for a broad range of people including such immune impairment patients
in case of bioterror attacks with smallpox. LC16m8 is an attenuated licensed
smallpox vaccine and has been stockpiled in Japan. LC16m8 has demon-
strated a good protective efficacy in healthy animal models and has given so
far to approximately 100,000 healthy people without any severe adverse
events. We have further conducted a research into how LC16m8 behaves
under the immune impairment using immunocompromised animal models.
Methods: To investigate the immunologic basis of containment of vaccinia in
the vaccination site or dissemination from the skin lesions to distal sites,
macaques were made depleted systemically of T- or B-cells and vaccinated
with either LC16m8 or Dryvax. To evaluate the protective efficacy in the im-
munocompromised hosts, LC16m8 was vaccinated to MHC class I-KO and
MHC class II-KO mice. The mice were challenged with a lethal dose of vac-
cinia WR at 2 or 3 weeks post vaccination and observed for 14 days. Results:
B-cell depletion did not affect the size of skin lesions induced by either vac-
BIODEFENSE PROGRAM & ABSTRACTS 49
M O N D A Y P O S T E R S E S S I O N | 1:00–3:00 p.m. | Exhibit Hall
cine. However, while depletion of both CD4+ and CD8+ T-cells made no ad-
verse effects and systemic dissemination on LC16m8-vaccinated animals, it
caused disseminated vaccinia in the spleen or cerebrum of macaques im-
munized with Dryvax. No viral DNA was detected in the other major organs
(e.g. heart, genitals) of vaccinated animals. A single dose of LC16m8 protects
both MHC class I-KO and MHC class II-KO mice from a lethal WR challenge.
Conclusions: The study showed that LC16m8 is well-tolerated and effective
in the immunocompromised hosts. It is suggested that LC16m8 can be a po-
tent candidate smallpox vaccine for immune impairment patients.
091 (H)
Development of a Guinea Pig Passive Transfer Efficacy Model
Using Immunoglobulin Derived from Clinical Trial Volunteers
Vaccinated with Recombinant Botulinum Vaccine A/B
S. Martin1, W. Swiderski1, K. Metcalfe1, N. Niemuth2, M. Vassar2, J. Shearer1;
1 IgG levels of anti-F1 and V for protection against Y. pestis challenge.
DynPort Vaccine Company, Frederick, MD, 2Battelle BioMed. Res. Ctr.,
Columbus, OH.
Purpose: This study was designed to characterize a guinea pig model for eval-
uation of passive protection against inhalational exposure to botulinum neu-
rotoxin complex (BoNT) serotypes A1 (BoNT/A1) and B1 (BoNT/B1).
Methods: Naïve guinea pigs were exposed to a range of dosages of
aerosolized BoNT/A1 or BoNT/B1 to determine the dosage lethal to 50% of
animals (LD50) and to evaluate the progression of disease. Mortality, median
time-to-death (MTTD), onset and duration of clinical signs of illness associ-
ated with challenge for both BoNT serotypes were evaluated. For passive
transfer, immunoglobulin was purified from the sera of rBV A/B-vaccinated
clinical volunteers and the challenge dosage required to produce <80% sur-
vival for each BoNT serotype was determined using a stage-wise approach.
At 24 hours after passive transfer, guinea pigs were aerosol challenged with
BoNT/A1 or BoNT/B1 and observed for clinical signs of botulism and mor-
tality for 14 days. Results: The guinea pig aerosol LD50 for BoNT/Al was es-
timated to be 158 mouse intraperitoneal LD50 (MIPLD50) and estimated to be
<221 MIPLD50 for BoNT/B1. Increases in mortality and decreases in MTTD
following exposure to either BoNT were inversely related to dosage. Changes
in breathing was the first sign of disease observed following challenge. The
MTTD following onset of the first clinical sign was dosage related for both
BoNT. Passively immunized guinea pigs were protected against inhaled
dosages 700 times the guinea pig aerosol LD50 of BoNT/A1 and BoNT/B1.
Conclusions: Guinea pigs passively immunized with human antibodies were
protected against aerosol challenge levels several orders of magnitude above
the aerosol LD50 for each BoNT serotype. The guinea pig model is suitable
for evaluation of the effectiveness of antibodies induced by rBV A/B vacci-
nation.
092 (H)
Increased Dose of Recombinant F1/V Plague Vaccine Results in
Increased Survival in Plague-Challenged Cynomologus
Macaques
K. A. Overheim, S. Lemoine, K. Gallegos, F. Romero, I. Fisher, A. Monier, C.
Schneider, M. W. Valderas, J. Wilder, R. Russell, R. L. Sherwood, R. LeClaire;
Lovelace Respiratory Res. Inst., Albuquerque, NM.
Background: Yersinia pestis is the causative agent of plague and can mani-
fest as pneumonic or bubonic forms. Previously we tested a recombinant
plague protein F1/V vaccine in Cynomolgus macaques (CM) at 10 μg of vac-
cine vaccinated on Day 0 and 30. All vaccinated CM survived a challenge with
50 LD50 of Y. pestis CO92. In this study we chose to determine if reducing the
dose of the recombinant protein F1/V vaccine would provide adequate pro-
tection in CM, as well as to further determine the correlates of protection.
Methods: Groups of NHP (6 animals per group) were vaccinated intramus-
cularly with varying doses of the recombinant vaccine (0.125, 0.5, 2, or 8 μg)
50 10th ASM Biodefense and Emerging Diseases Research Meeting
on Days 0 and 30. Controls were given phosphate buffered saline only (PBS)
on the same vaccination regimen. All animals were aerosol challenged with
a target of 50 LD50 Y. pestis CO92 70 days after initial vaccination. Results:
Controls succumbed to Y. pestis within 9 days post exposure, as did all ani-
mals in the 0.5 μg group (by 5 days post exposure). 70% of animals in the 8
μg group survived, whereas 20% of animals in groups 2 μg and 0.125 μg sur-
vived. Y. pestis was cultured (controls and vaccinated) in the blood, lungs,
spleens, livers, brains, and lymph nodes from those animals that succumbed
to infection. Vaccinated animals that survived Y. pestis challenge had no de-
tectable tissue burden. Pathology showed minimal lung alterations in three
of four CM that survived challenge, indicating strong protection. The other
animal had localized residual fibrosis indicating vaccination was effective in
limiting infection. Survival correlated with a higher dose of vaccine as well
as higher serum IgG anti-F1 and -V levels measured by Protein G ELISA and
maintained lymphocyte and monocyte counts. Conclusions: These data sug-
gest that an optimal dosage of vaccine is required to induce optimal serum
093 (H)
Generation of Protective Immunity against Fatal Rickettsial
Disease
S. P. Riley1,2, M. M. Cardwell1,2, R. Hillman1,2, L. Quenee1,2, J. J. Martinez1,2;
1
Univ. of Chicago, Chicago, IL, 2The Howard Taylor Ricketts Lab., Lemont, IL.
Pathogenic rickettsiae are the causative agents of Rocky Mountain Spotted
Fever (RMSF), Mediterranean Spotted Fever (MSF), typhus and other human
diseases with high morbidity and mortality. Previous reports have demon-
strated that survivors of rickettsial disease are immune to subsequent in-
fections; however, the molecular basis of this immunity was not well defined.
We have recently demonstrated that purified recombinant outer membrane
protein B (rOmpB) is sufficient to elicit protective humoral immune re-
sponses against lethal disease in a murine model of MSF mediated by Rick-
ettsia conorii. This study also revealed that protective immunity was
correlated with the ability of generated antibodies to facilitate complement-
mediated elimination of rickettsiae from blood and plasma. The evolution-
ary and amino acid conservation of rOmpB among closely and distantly
related rickettsial species suggests that this protein may be sufficient to
serve as a protective antigen in other rickettsial disease models. To test this
hypothesis, we initially developed a C3H/HeN murine model of fatal Rocky
Mountain Spotted Fever (RMSF) mediated by the CDC-classified Select
Agent, Rickettsia rickettsii, and have demonstrated that the progression to
disease and pathologic injuries to the spleen, liver, heart and lungs closely
mimic those observed in fatal R. conorii infections. We are currently utilizing
this model to test the efficacy of recombinant purified R. rickettsii rOmpB
and other conserved rickettsial antigens in the generation of protective im-
munity against fatal disease. We will also determine whether complement-
mediated killing also plays a crucial role in protection against fatal R.
rickettsii infections. With the threat of the potential use of rickettsial species
as agents for bioterrorism, understanding the complex interplay between
pathogen and host is crucial for the development of novel antimicrobials
and more efficacious therapies against fatal diseases.

094 (I)
Determination of Efficacious Praziquantel Dose in Different
Mouse Strains: BALB/C and Swiss Mice for Treatment of Schis-
tosoma mansoni
M. P. M. N. NA1, D. Y. S. N. yole2; 1Mombasa Polytechnique Univ. Coll., Nairobi,
Kenya, 2Kenya Polytechnique Univ. Coll., Nairobi, Kenya.
Background: The drug Praziquantel has been in use for treatment of schis-
tosomiasis however the effective dose that can clear the worms completely
has not been determined. Mouse models have been used in this study to de-
termine effective Praziquantel dosage.This work therefore sought to deter-
mine the effective dose of praziquantel in different mouse strains, BALB/c
and Swiss mice. Methods: Six mice were divided into four groups namely
PZQ1350, PZQ900, PZQ450 and infected control. Mice were infected at week
0 and at week 4 all the experimental groups (ie PZQ1350, PZQ 900 and
PZQ450 mg/kg bw (Farah et al, 2000), respectively) were treated using vary-
ing praziquantel dosages. At week 6 all mice were perfused to recover adult
worms. Gross pathology and histopathology of the liver tissue were ob-
served. Results: More worms were recovered in infected control and there
was less reduction of worm burden was observed in this group p<0.001.
Gross pathology was severe and more granulomas were also recorded in this
group. In the experimental groups PZQ1350 had few numbers of worms re-
covered and a high worm reduction P<0.001. Also no granulomas were
recorded in this group due to the effect of drug and humoral responses in
clearing the worms. This was followed by PZQ900 but PZQ450 which is the
standard dosage used indicated more worm recovery and less reduction in
worm burden. Conclusion: The findings imply that the higher the dosage the
more the protection against Schistosoma mansoni infection; PZQ1350 indi-
cated better responses both in worm recovery, worm reduction, and pathol-
ogy compared to other groups. The findings also indicate that Swiss mouse
was a better permissive host than BALB/c as it allowed more parasites to ma-
ture instead of destroying them.Further experiments could be done,with de-
termination of humoral and cellular responses.
095 (I)
A Novel Mechanism by which Human Neutrophils Kill Bacillus
anthracis
G. Ramachandran1, L. Baillie2, P. Tsai3, G. Rosen3, A. Cross1; 1Univ. of Maryland
Med. Sch., Baltimore, MD, 2Welsh Sch. of Pharmacy, Cardiff, United Kingdom,
3
Dept. of Pharmaceutical Sci., Baltimore, MD.
Background: Bacillus anthracis (BA), the causative agent of anthrax, is ac-
quired by mammalian hosts from the environment as spores that are able to
survive harsh conditions. BA spores must germinate inside host cells to veg-
etative bacilli before they can express the virulence factors that enable them
to evade host defense and disseminate. While the role of macrophages and
dendritic cells in this initial interaction is well established, the role of poly-
morphonuclear leukocytes (PMNs) is not well defined. We now test if BA 34F2
Sterne spores and/or vegetative bacilli are killed by human PMNs in vitro and
also determine the mechanism of killing. Results: At an MOI of 1:1 PMNs are
unable to kill 34F2 Sterne or the germination deficient ΔgerH spores. How-
ever at the same MOI, PMNs killed over 80 % of vegetative bacilli within 20
min even in the absence of opsonins. In the absence of PMNs BA bacilli are
extremely susceptible to killing by superoxide generated by epigallocatechin
gallate. Human PMNs generate low levels of superoxide (or nitric oxide) only
in the presence of BA at a MOI of ≥ 1: 100. Lysates of human PMNs (106/ml)
were unable to kill 106 BA, but very high doses (10-100 ug/ml) of antimicro-
bial peptide, α-defensin, were required to kill BA bacilli. By using an inhibitor
of autophagy, 3MA, we identified autophagy as a mechanism by which PMNs
killed BA. In the presence of 3MA, there was over 50 % reduction in killing of
BA by PMNs. Moreover, formation of LC3 puncta, an autophagy marker, dra-
matically increased in the presence of BA within 20 min of incubation. Con-
M O N DAY A B S T R AC TS
clusion: PMNs are not effective killers of BA spores but they efficiently kill
vegetative bacilli. While oxidative mechanisms do not appear to play an im-
portant role in killing BA, PMN autophagy may be an important mechanism
to eliminate BA.
096 (I)
Burkholderia pseudomallei Induces IL-1beta and IL-6 Secretion
via Activation of MAPK and STAT3 in Differentiated THP-1 Cells
M-H. Cho1, Y-W. Shin1,2, J-H. Chun1, K-J. Hong1, G-e. Rhie1; 1Korea CDC, Chung-
buk, Korea, Republic of, 2Yonsei Univ., Seoul, Korea, Republic of.
Burkholderia pseudomallei is a bacterial pathogen that causes a broad spec-
trum of clinical symptoms collectively known as melioidosis. This study aims
to investigate the underlying mechanism of inflammatory responses elicited
by B. pseudomallei in the human monocytic cell line THP-1, which was dif-
ferentiated into macrophages with phorbol 12-myristate 13-acetate (PMA). B.
pseudomallei infection on differentiated THP-1 cells led to the specific ex-
tracellular release of interleukin (IL)-1β and IL-6 in a dose-dependent manner
as shown by ELISA. Also, we confirmed that B. pseudomallei-induced IL-1β
and IL-6 production occurs via activation of signal transducer and activator of
transcription-3 (STAT3) by Western blot analysis. Moreover, a STAT3 inhibitor
(static) inhibited B. pseudomallei mediated IL-1β and IL-6 production. p38,
ERK1/2 and JNK phosphorylation, modulate IL-1β and IL-6 secretion follow-
ing infection of B. pseudomallei on THP-1 cells. Also, B. pseudomallei induced
IL-1β and IL-6 secretion was reduced in the presence of the inhibitors of p38-
MAPK (SB203580), ERK1/2 (PD98059) and JNK (SP600125). These findings
provide the key role of STAT3 and MAPK kinase pathways in IL-1β and IL-6
production in response to inflammation induced by B. pseudomallei.
097 (I)
Increased Protection of African Green Monkeys Vaccinated with
Increasing Doses of a Recombinant F1/V Plague Vaccine
K. A. Overheim1, S. Lemoine1, K. Gallegos1, K. Brooks1, I. Fisher1, A. Monier1,
C. Schneider1, M. W. Valderas1, J. Wilder1, R. Russell1, R. L. Sherwood1, R.
LeClaire2; 1Lovelace Respiratory Res. Inst., Los Lunas, NM, 2Lovelace Respi-
ratory Res. Inst., Albuquerque, NM.
Background: Yersinia pestis is the causative agent of plague and can mani-
fest as pneumonic or bubonic forms. Previously we tested a recombinant pro-
tein vaccine (F1/V in alum) in African green monkeys (AGM) using 10 μg of
vaccine administered intramuscularly (i.m.) on Days 0 and 30. All vaccinated
AGMs, with the exception of one animal, succumbed to challenge with 50
LD50 of Y. pestis CO92. Therefore, we decided to determine a dose of vaccine
that would confer protection to AGM recombinant protein F1/V vaccine, as
well as determine the correlates of protection. Methods: Groups of NHP (6 an-
imals per group) were vaccinated i.m. with varying doses of the recombinant
vaccine (100 μg, 25 μg, 6 μg, 1.5 μg) on Day 0, 28, and Day 56. Controls were
given phosphate buffered saline only (PBS) on the same vaccination regi-
men. All animals were aerosol challenged with 25 LD50 Y. pestis CO92 70 days
after initial vaccination. Results: All but one control succumbed to Y. pestis
infection within 6 days post exposure. The low dose vaccinated group (1.5
μg) all succumbed by Day 5 post exposure. 70%, 50% and 33% of animals in
the 100 μg, 20 μg, and 6 μg groups survived to 21 days post exposure. Y.
pestis was cultured from control AGM and vaccinated AGM that succumbed
to disease from their blood, lungs, spleens, livers, brains, and lymph nodes.
Increased doses of vaccine roughly correlated with increased serum anti-F1
and anti- rV antibody titers. Conclusions: These data suggest that an opti-
mal serum IgG level to r-V and r-F1 are needed in AGM to confer protection
against Y. pestis challenge.
BIODEFENSE PROGRAM & ABSTRACTS 51
M O N D A Y P O S T E R S E S S I O N | 1:00–3:00 p.m. | Exhibit Hall
098 (I)
Ebola Virus Secreted Glycoprotein Can Function as a Decoy for
Antibodies
G. S. Mohan, Y. Chinglai, L. Ye, R. Compans; Emory Univ., Atlanta, GA.
Through an RNA editing mechanism, Ebola Virus makes two isoforms of its sur-
face glycoprotein (GP). The membrane-bound isoform (mGP) is the primary
viral mediator of host cell attachment and entry. However, ~80% of GP tran-
scripts encode a nonstructural secreted GP (sGP) whose function has been
widely debated. Here, we demonstrate that sGP can serve as a decoy for anti-
EBOV antibodies. In mice immunized with the wild-type GP construct, sGP was
able to compete with mGP for antibody binding. Surprisingly, in mice immu-
nized with mutant constructs expressing solely mGP, sGP was unable to com-
pete with mGP, even at 16-fold molar excess. In these mice, sGP also had no
effect on antibody-dependent pseudovirus neutralization, while sGP inhibited
neutralization in wild-type GP-immunized mice. Together, these data suggest
that during natural infection, an excess of sGP focuses the host immune re-
sponse against epitopes shared with mGP, allowing sGP to absorb anti-EBOV
antibodies. Interestingly, serum from mice primed against sGP and then
boosted with mGP still cross-reacts with sGP, suggesting that there may be a
component of original antigenic sin at play. This is particularly germane to Ebola
vaccinology, given the implication that vaccination of previously-exposed indi-
viduals may simply boost an already non-protective immune response.
099 (J)
Bioactivity of an Indigenous Plant Used in the Treatment of
Malaria against Anopheles Mosquito Larvae in South West
Nigeria
F. O. Omoya, B. E. Boboye; Federal Univ. of Technology, Akure, Nigeria.
Malaria has been one of the major sources of human death and it causes sig-
nificant economic losses in Africa. Even in developed countries, such as Eu-
rope and United States, re-emergency of this disease has been reported.
Plasmodium spp— the etiologic agent—has shown resistance to common
and cheapest drugs for the treatment and prevention of malaria. In other to
eradicate this disease, different control measures have since been exploited
by man. Control of Anopheles mosquito, the vector, is one of the ways our
research group is focusing on as a mean of controlling the disease. This re-
search work aimed at the use of different leaf extracts of the medicinal plant;
Vernonia amygdalina on the second and fourth instars larvae of Anopheles
arabiensis for a period of 24 hours in-vitro and the phytochemical screening
of the plant. Three different leaf extracts namely; water, ethanol and methanol
extracts showed different percentage mortality on the tested larvae. The high-
est mortality was recorded in ethanol treated larvae. The lethal concentra-
tion at LC50 and LC75 also varied with different extracts and instrars. In the
fourth instars larvae treatments; LC50 (15.43mg/mL) and LC75 (46.95mg/mL);
LC50 (12.54mg/mL) and LC75 (16.64mg/mL); and LC50 (2.51mg/mL) and LC75
(5.45mg/mL) were recorded in water, methanol and ethanol extracts respec-
tively. Phytochemical screening of this plant revealed the presence of some
chemicals that are insecticidal such as Tannins and Alkaloids. Summarily, our
finding revealed that Vernonia amygdalina might be considered as a larvici-
dal agent in the control of Anopheles mosquito larvae.
100 (J)
Formaldehyde Disinfection Using Porcine Parvovirus as Indicator
E. Emmoth, U. A. Bengtsson, I. Dergel, C. Hultén; Natl. Vet. Inst., Uppsala,
Sweden.
We used porcine parvovirus (PPV) to validate a formaldehyde fumigation
process of a biosafety level 3 laboratory of 190 m3 volume. PPV strain 893/76,
in Eagle’s minimal essential medium (EMEM) with 1% foetal bovine serum
(FBS) was dried in 50 μl volumes on stainless steel discs. The discs were
52 10th ASM Biodefense and Emerging Diseases Research Meeting
placed at nine locations in the lab, and subjected to formaldehyde of 1.7 g/m3
at >80% humidity at 22±2°C for 16h. Extraction media was EMEM with 10%
FBS. Titrations were by ten-fold dilutions and eight replicates per dilution,
using PK15 cells and immunoperoxidase staining, and controls and reduc-
tion factors according to (1). No PPV could be recovered after fumigation,
with a residual titre of ≤-0.27 log10 TCID50 disc-1 according to (2). A PPV re-
duction factor of ≥7.0±0.37 proved the fumigation process efficient, and the
controls showed that the assay was reliable (Table 1)
Table 1. Titres in TCID50a log10 ±SDbdisc-1 (recovery control) or 50 μl-1 (stop/in-
terference controls)
Controls Extracted EMEM 10% FBS EMEM Reduction
steel disc
Recovery controlc 6.7±0.37 n.a. n.a. n.a.
Stop control 1.5±0.45 1.4±0.25 n.a. -0.12±0.51
Interference control 3.1±0.44 n.a. 3.1±0.36 0±0.57
Cytotoxicity control No cytotoxicity n.a. No cytotoxicity n.a.
n.a. = not applicable; atissue culture infectious dose; bstandard deviation;
c
after 16h at 22±2°C
This work was part of a joint project between the Swedish Defence Research
Agency and the National Veterinary Institute, concerning various decontam-
ination strategies of high risk pathogens in the community, financed by the
Swedish Civil Contingencies Agency. Future experiments will include high
pathogenic avian influenza and spores of B. antracis. References: (1) Guide-
line of “Deutsche Vereinigung zur Bekämpfung der Viruskrankheiten e.V.“
and Robert Koch Institute for testing the virucidal efficacy of chemical disin-
fectants in the human medical area” (2) Blumel, J., I. Schmidt, H. Willkom-
men, and J. Lower. 2002. Inactivation of parvovirus B19 during pasteurization
of human serum albumin. Transfusion 42:1011-1018.
101 (J)
Bactericidal Activity of Antimicrobial Peptides Immobilized on
Planar Surfaces
S. Arcidiacono, J. Uzarski, L. Doherty, W. Muller, C. Mello; U.S. Army Natick
Soldier Ctr., Natick, MA.
Background: Substrates with antimicrobial functionality have utility in med-
ical applications, food safety, decontamination, and prevention of materials
degradation. Most surface activity measurements are made in solution (e.g.
growth medium or buffer). However, some applications occur in which the
antimicrobial compound needs to retain functionality in a non-solution state,
(i.e. in a range of water content or humidity). To examine the effect of hu-
midity on activity, bacteria were exposed to antimicrobial peptides (AMPs)
attached a planar surface and incubated under varying relative humidity.
Methods: Cysteine variants of Cecropin P1 and SMAP29 were attached to
gold via self-assembled monolayers consisting of mixed oligo(ethylene gly-
col) containing alkane thiols, functionalized with a sulfo-GMBS cross-linker.
Surface antimicrobial activity was determined against cells in the presence of
solution as well as cells dried on the surface and incubated at constant hu-
midity. Cells were also observed microscopically with LIVE-DEAD fluorescent
stain to assess activity. Results: Immobilized SMAP29 was active against the
Gram-negative bacteria Escherichia coli and Acinetobacter baumannii in the
presence of solution. Fluorescent microscopy confirmed that the surface was
active. However, no activity was observed against the Gram-positive bac-
terium Staphylococcus aureus. Surfaces containing immobilized Cecropin P1
were not active. Activity was evaluated against cells dried on a surface and
incubated under constant humidity. Suitable incubation conditions allowing
cells to retain viability during drying continues to be investigated. Conclu-
sions: Covalent attachment of SMAP29 imparted antimicrobial functionality
to gold surfaces. The research presented here lays the foundation for a new
generation of non-leaching antimicrobial treatments. As a potential alterna-

tive to antimicrobial agents currently in use, AMPs are not expected to pro-
mote emergence of resistant microorganisms.
102 (J)
Investigating the Dispersion and Decontamination of Anthrax
Spores Within Indoor Work Areas
A. Clohessey, S. Hoque; Rowan Univ., Glassboro, NJ.
The threat of biological agents used as weapons on the civilian population
has become a very real concern, especially after the anthrax-based attacks
between October 4 and November 2, 2001. This study is aimed at gathering
information on the dispersion of particles in an office space and the subse-
quent decontamination of that area. The goal of this investigation is to un-
cover answers to questions that first-responders would have to face in
situations where an infectious agent has been released in a building. Here,
furniture presence is investigated in the effects it has on particle dispersion.
Computational fluid dynamics software using LES method is used to simu-
lated particle releases in a modeled office space incorporating furniture.
100,000 Bacillus anthracis spores are released from 3 locations and the re-
sults are compared. Areas where flow is more stagnant allow particles to re-
main within the room longer and disperse throughout. The timeframe of this
investigation has spanned September, 2011 until the present. Future goals
aim at incorporating ClO2 disinfection agent into the room. The required con-
centration to decontaminate as well as time required to reach safe levels of
ClO2 will be investigated.
M O N DAY A B S T R AC TS
103 (K)
Preparing Biodefense Professionals: MS in Biotechnology
Concentration in Biodefense and Certificate in National
Security Studies
K. M. Obom1, A. I. Roth2, P. J. Cummings3; 1Johns Hopkins Univ., Rockville,
MD, 2Johns Hopkins Univ., Washington, DC, 3Johns Hopkins Univ., Baltimore,
MD.
To respond effectively to a biological threat, the nation will need a well trained
workforce. Currently, there is a shortage of biodefense professionals, partic-
ularly individuals that have a strong background in the science and policy re-
lated to biodefense issues. To prepare professionals who have this expertise,
students will be able to pursue a Certificate in National Security Studies and
a Master of Science in Biotechnology with a concentration in Biodefense at
Johns Hopkins University (JHU). This unique partnership between JHU’s Na-
tional Security Studies and the Center for Biotechnology Education programs
and the United States Army Medical and Research Institute of Infectious Dis-
ease (USAMRIID) will provide students with a curriculum that includes grad-
uate course offerings in Bioinformatics; Biothreat Response and Microbial
Forensics; Science, Medicine & Policy in Biodefense; Vaccinology; Radiation
Biology; and Biodefense Lab Methods, as well as Preserving American Se-
curity; Crisis Management and The Art and Practice of Intelligence. As part of
the curriculum, USAMRIID offers a competitive fellowship program for up to
five students each year to work at USAMRIID, while completing graduate
courses at JHU’s campuses in Baltimore or Rockville, MD and Washington
DC. Graduates of the degree program and the certificate will have an in-depth
knowledge of the policy and practical laboratory skills in the field of biode-
fense and will fill a growing need for trained professionals skilled in this es-
sential discipline within the science community and for homeland security.
104 (K)
Recruiting First Responders from the Dental Profession
R. J. Boylan, D. L. Glotzer; NYU Coll. of Dentistry, New York, NY.
Significant advances in our nation’s ability to detect environmentally-dis-
seminated bioweapons have been made over the past ten years. Manipula-
tion of these bioweapons in the laboratory has given us much information
about potential mutants that could be produced as well as more effective
ways to treat them. However, a recent report from the bipartisan WMD Ter-
rorism Research Center states that “the primary means of defending the
American Homeland against bioterrorism is the capability to effectively re-
spond after an attack has occurred.” Our communities’ first responders will
have the primary responsibility to respond to a major disaster. However, in
the event of a massive attack employing bioweapons, it may be necessary to
call upon other healthcare personnel to participate. Experience has shown
that one critical area that needs to be considered in catastrophe response is
the available numbers of trained individuals in the public health workforce
that can, and will respond when necessary. Healthcare workers who are
knowledgeable about the potential agents and their properties, modes of
transmission and means of personal protection, as well as the most effective
therapies, are more likely to engage effectively and safely in such a response
than those who are not. Since shortly after 9/11 NYU’s Dental School made
it a priority to educate its students in such matters throughout its 4-year cur-
riculum. They are provided with extensive information about the bioweapons
in the CDC’s categories A and B in a two hour conference in Infectious Dis-
eases; given a comprehensive 12 hour course in Bioterrorism Preparedness
in which the AMA’s Core Disaster Life Support program is presented; and en-
couraged to prepare a plan to protect themselves, their staff and patients in
the event of a disaster. If HR 570, The Dental Emergency Responder Act
(which passed the House this year) passes the Senate, then dentists by Fed-
eral regulation will be considered emergency responders. The model for ed-
ucating dental students in these areas, which will be described more
BIODEFENSE PROGRAM & ABSTRACTS 53
M O N D A Y P O S T E R S E S S I O N | 1:00–3:00 p.m. | Exhibit Hall
thoroughly at the meeting, will potentially provide a larger pool of health care
workers willing to be effective agents, contributing importantly to a
bioweapons attack.
105 (K)
Tungiasis (Jigger Infestation) in Rural Kenya: An Emerging In-
fectious Disease
N. N. N. N. Ngomi, Jr.; Jomokenyatta Univ. Kenya, Nairobi, Kenya.
Objective: To describe the prevalence of tungiasis (jigger flea infestation) and
associated risk factors in a sentinel group (children 5-12 years of age) in rural
Central Kenya. Methods: A cross-sectional study was carried out in Murang’a
South district during high transmission season (dry season, August - Sep-
tember 2009). A total of 385 randomly selected households were visited. Chil-
dren were examined for presence of tungiasis, and a questionnaire was
administered to collect demographic, behavioral and environmental data.
Results: Prevalence of tungiasis was 57% (218/385; 95% CI=51.7%-61.6%).
Itching (89.1%) was the most common associated symptom, followed by pain
upon pressure (67.3%), sleep disturbance (58.2%) and walking difficulties
(53%). In multivariate logistic regression analysis the following independent
factors were identified to be associated with tungiasis: living in houses with
an earthen floors (adjusted OR=3.84; 95%IC: 2.09-7.06), walking barefooted
(OR=3.28; 1.78-6.04), having a common resting place outside the house
(OR=2.36; 1.01-5.51) and presence of rats on the compound (OR=1.69; 1.03-
2.75). Conclusion: Tungiasis is an emerging neglected disease found in Africa.
It is highly endemic in rural Central Kenya and associated with considerable
morbidity. The disease is associated with poverty. Modifiable risk factors were
identified that should be the focus of sustainable and effective control meas-
ures.
106 (K)
Essential Oils of Moroccan Thymus munbyanus as Natural
Product for Nosocomial Infection Prevention
M. Fatima Zohra, Mohammed Khatouf; Badiaa Lyoussi and Abdelfattah Ab-
dellaoui; Faculty of Sci. Dhar Mhraz, Fes, Morocco.
Diseases caused by Gram-negative bacteria resistance to antibiotics are an
increasing problem in Nosocomial infections, and they are responsible of
morbidity in the hospital. In such cases, evaluation of antibacterial activity
must run tests against those bacteria to determine new natural product. The
purpose of this study was to compare the activity antibacterial of essential
oils of thymus to antibiotics used for treatment of Nosocomial infection in
Reanimation Polyvalent and Anesthesia service; CHU Hassan II Fez city. Thy-
mus species have been recommended in Moroccan traditional medicine as an
antispasmodic intestinal and antidiarrheal. The essential oils were isolated by
hydrodistillation, from the aerial parts of Thymus munbyanus possess an-
timicrobial properties important against bacteria tested: Escherichia coli;
Pseudomonas aeruginosa; Klebsiella pneumoniae. The results obtained sug-
gest that the essential oils of the plant could become a source of biologically
active compounds for pharmaceutical industry.
54 10th ASM Biodefense and Emerging Diseases Research Meeting
107 (K)
Biosafety and Biosecurity: The Case of a Third World Country
and the One Health Spectrum
D. B. Ndumu1, R. Heckert2, C. S. Rutebarika1, R. W. Kaboyo3, M. Busuulwa4, P.
Atimnedi5, S. Balinandi6, P. Delarosa7; 1Ministry of Agriculture, Animal In-
dustry and Fisheries, Entebbe, Uganda, 2Robert Heckert Consulting LLC,
Bowie, MD, 3Ministry of Hlth., Kampala, Uganda, 4AFENET, Kampala, Uganda,
5
Uganda Wildlife Authority, Kampala, Uganda, 6Uganda Virus Res. Inst., En-
tebbe, Uganda, 7Booz Allen Hamilton, Middletown, DE.
The silent ‘bioterror war’ on mankind in the Third World is attributed to epi-
demics and emerging infectious diseases. These developing countries are
the front line of defense against the devastation that would arise from pan-
demics or deliberate release of a pathogen. While disease outbreaks are also
linked to sociopolitical problems, integrated plans that prepare such coun-
tries to deal with pandemics and possible bioterror attacks have not been
devised. Recent efforts rely on a reporting system through the World Organ-
ization for Animal Health. Although important; this reporting alone is a frag-
ile early warning system, as it relies on a single response generated after
confirmation of an outbreak in as far as biodefence is concerned. Since many
human diseases have their source in animals as do 80% of the pathogens
that could potentially be used in bioterrorism, it is imperative to build mod-
els that incorporate ecological health into preparedness and response to
human disease outbreaks. Using the One Health paradigm, a robust biose-
curity system can be developed that adds a layer of support to international
disease reporting. This presentation examines a redefinition of traditional
biosafety and biosecurity within the context of One Health efforts in Uganda
with an emphasis on defining the elements of a One Health risk assessment
and application of biosafety and biosecurity efforts across a One Health plat-
form. The infrastructure and organization, or the lack of it, in Uganda is used
as an example. The article also portrays opportunities to strengthen world-
wide biosecurity by defining concentric layers of biosecurity and biosafety
using the One Health paradigm.

Poster Session
018. Tuesday Posters
Tuesday, February 28, 2012 | 1:00 PM - 3:00 PM
Exhibit Hall
140 (A)
Molecular Epizootiology of Street Rabies Virus Isolates in
Ukraine
I. Polupan, N. Ivanov, O. Deryabin, V. Skripnik; Inst. of Vet. Med., Kyiv,
Ukraine.
To develop and implement an effective program of rabies eradication in
Ukraine it is necessary to research the molecular genetic characteristics of
the causative agent of this disease in the country. To conduct this research in
2008 was founded the unique collection of samples of pathological materi-
als confirmed as positive in rabies at the regional veterinary laboratories of
Ukraine. The collection is constantly updated and to present moment it in-
cludes 1294 samples from all regions of Ukraine, selected from 16 animal
species and humans. Identification of the rabies virus in samples of patho-
logical material for their further selection was carried out using the test de-
veloped by us which based on RT-PCR with primers complementary to the
conservative fragments of the 5’-end of nucleoprotein gene of rabies virus.
For the study of the street rabies virus isolates from the collection we use RT-
PCR with the primers pair (509, 304) flanking the variable 3’-end part of nu-
cleoprotein gene of the reference strain of rabies virus CVS (fragment in 377
bp). With this pair of primers it was conducted amplification of 76 street iso-
lates, 3 vaccine strains and reference strain CVS of rabies virus. These primers
were complementary to the reference strain CVS and all of vaccine strains of
rabies virus. However from 76 street rabies virus isolates variable fragment
of 3’-end was amplificated at 32 (42%) but at 44 (58%) amplification didn’t
take place. Lack of amplification for the part of street rabies virus isolates is
indicative of polymorphism of the target nucleoprotein gene fragments. This
was used as a criterion for their genetic typing and makes it possible to divide
the researched isolates into two genetic clusters. The analysis of the results
allowed to draw conclusions about the absence of relation of genetic clusters
with the host species. But the territorial ranking of these genetic variants of
rabies virus showed their distinct geographical distribution limited by the
Dnipro river. Isolates belonging to cluster I circulate at the western side of
Ukraine and at the eastern side circulate isolates belonging to cluster II.
141 (A)
Utility of a New Real-Time RT-PCR Assay for the Detection of
Crimean-Congo Haemorrhagic Fever Virus in Pakistan
B. Atkinson1, Z. Hasan2, A. Samreen2, B. Jamil2, T. Moatter2, J. Chamberlain1,
N. Cook1, C. Logue1, C. Bruce1, S. Dowall1, R. Hewson1; 1Hlth. Protection
Agency UK, Salisbury, United Kingdom, 2The Aga Khan Univ., Karachi, Pak-
istan.
Introduction: Crimean-Congo Haemorrhagic Fever virus (CCHV) is a seg-
mented, negative stranded, RNA virus classified within the genus Nairovirus
family Bunyaviridae. It is serious human pathogen categorized at biosafety
level 4 (BSL4) mandating the use of maximum containment laboratory facil-
ities such as those at the HPA Porton Down in the UK. The virus is maintained
in nature in a zoonotic cycle involving a variety of vertebrate species, with
animal-to-animal transmission mediated by ticks. Both vertebrate hosts and
tick vectors act as reservoirs of virus. Transmission to humans commonly oc-
curs by bite from an infected tick. In addition to zoonotic transmission, CCHFV
can also be spread from person to person. Case fatality rates ranging from 10-
50% are common; however person to person transmission, especially in hos-
pital settings, often results in higher mortality. CCHF is endemic in Pakistan
T U E S DAY A B S T R AC TS
and sporadic outbreaks of disease associated with its propensity to cause
nosocomial infections are infamous. A rapid diagnosis makes useful contri-
butions to hospital and patient management and is especially pertinent dur-
ing simultaneous outbreaks of dengue fever which may also present with
haemorrhagic symptoms. Results: We have developed a novel and sensitive
real time RT-PCR assay for the detection of a broad range of CCHFV geno-
types. Based on a single probe the assay brings added benefits over other
similar tests. It has also been transitioned to colleagues in Karachi where it
has proved invaluable for rapid clinical diagnosis. Conclusions: In Pakistan
rapid diagnosis of patients with CCHF enables prompt admission to isolation
units and efficient patient management. Foreknowledge of the likely course
of disease and implementation of appropriate barrier nursing techniques
greatly reduces the risk of nosocomial transmission. The assay makes broad
contributions to differential diagnosis and supports the management of
haemorrhagic fever at Aga Khan University Hospital, Karachi.
142 (A)
Development of Differential and Broad Spectrum Immunologi-
cal Reagents for Identification and Characterization of Filovirus
Sub-Types
C. Bruce, K. Richards, L. Easterbrook, H. Love, L. Hudson, J. Plank, H. Tolley,
R. Hewson, A. Robers; Hlth.Protection Agency, Salisbury, United Kingdom.
Background: With the ever increasing levels of international travel, trade and
threat of bio-terrorism, the need to quickly and efficiently identify filovirus
infection is critical. Specialised high-containment laboratories required to
work with hazard group-4 (HG) agents are costly to run/maintain and can re-
strict experimental procedures. Alternate work strategies are therefore re-
quired to facilitate the swift and efficient development of new assays required
to support diagnostic and vaccine development programmes. Our work fo-
cuses on the development of differential immuno-assays for identification
and characterization of filovirus sub-types. Here we describe a strategy for
producing monoclonal antibodies (mAb) to HG-4 agents in Biological Safety
Level-2 (BSL) facilities using the baculovirus expression system (BES). Meth-
ods: Viruses Filoviruses were propagated in Vero E6 cells. The strains used
were Ebola Zaire Manyinga 1976, Sudan Boniface 1976, Ivory Coast 1994
and Bundibugyo 2007 and Marburg Ravn and Angola. Virus-Like Particles
Species-specific VLPs were produced using recombinant baculoviruses con-
taining glycoprotein (GP) and VP40. Monoclonal Antibodies Mice were im-
munized with EBOV or MARV VLPs. Monoclonal hybridoma populations were
assayed for antigen specificity by Enzyme-Linked Immuno-Sorbent Assay, na-
tive and denaturing Western blots. Results: EBOV and MARV recombinant
VLPs were shown to exhibit a similar morphology to wild-type virus, and pro-
duced an immune response in mice; producing both GP and VP40 antibodies.
Arrays of poly and mAbs were identified that displayed species-specificity, or
cross-reactivity in the assays used. Conclusions: We have successfully shown
that the BES can be used to produce EBOV and MARV VLPs, and that they
are immunogenic in mice, producing specific and cross-reactive Abs to dif-
ferent filoviruses. These reagents provide the basic building blocks for de-
veloping differential and broad spectrum immunological detection assays,
and potential therapeutics.
143 (A)
Fighting Fire with Fire: The Development of Antiviral AAVs
K. K. Wong, Jr.; City of Hope Natl. Med. Ctr., Duarte, CA.
Adeno-associated virus (AAV) is a 5 kb nonpathogenic, helper dependent
member of the Parvovirus Family. Helper viruses have included adenovirus
(Ad), all herpes viruses, human papillomavirus (HPV), and vaccinia virus, all
potential human pathogens. Classically, AAV has been utilized as a viral vec-
tor by removing endogenous AAV genes, substituting a gene of interest, and
encapsidating with AAV Rep/Cap genes and specific Ad or herpes simplex
BIODEFENSE PROGRAM & ABSTRACTS 55
T U E S D A Y P O S T E R S E S S I O N | 1:00–3:00 p.m. | Exhibit Hall
(HSV) genes. We have chosen another approach.In contrast to currently avail-
able rAAV vectors, we utilized full length wild type AAV (RC rAAV) and inserted
inhibitory siRNA cassettes targeting specific helper viruses. We were able to
significantly inhibit expression of the HPV31 E7 gene, which is essential for
HPV replication, by 4 different siRNAs. There are several novel and unique
features of this strategy. (1) When co-infected with a targeted helper virus
(THV), antiviral RC rAAVTHVi vectors replicate, amplify and potentiate their
antiviral effect(s) by increasing DNA template number and augmentation of
antiviral gene expression. In contrast, standard AAV Rep/Cap deficient (NR)
rAAVTHVi vectors cannot replicate, and require a larger amount of vector for
the same level of inhibition. (2) During active infection, RC rAAVTHVi package
and spread to surrounding cells to provide additional protection to both THV
infected and uninfected cells. (3) Antiviral RC rAAVTHVi express the AAV2 Rep
protein which possesses additional intrinsic antiviral activity against HPV,
and other viruses. (4) RC rAAVTHVi vectors would be easier to produce at
high titer using a different or irrelevant “helper” virus than the THV, because
they can generate a full infectious cycle (replication->packaging->infection
of new cells, etc.) within helper virus infected 293 cells, while NR rAAVTHVi
vectors cannot. (5) From a Biosafety standpoint, these vectors would be no
more toxic than wild type AAV2, which is considered a nonpathogen by the
CDC, assuming that the expressed siRNAs are themselves not toxic. Using
this approach, we have inhibited HSV (HSV UL29.2) and HPV (E7) genes, and
are examining inhibition of Ad genes for a potential human clinical trial.
144 (A)
Arenavirus Reverse Genetics for Vaccine Development
E. Ortiz Riano1, B. Y. H. Cheng1, J. C. de la Torre2, L. Martinez-Sobrido1; 1Univ.
of Rochester Med. Ctr., Rochester, NY, 2The Scripps Res. Inst., La Jolla, CA.
Arenaviruses are important human pathogens with no FDA-licensed vaccines
available and current antiviral therapy being limited to an off-label use of the
nucleoside analog ribavirin. Plasmid-based reverse genetics techniques for
the generation of recombinant arenavirus have proven valuable for virus re-
search; however, current approaches to generate recombinant arenaviruses
are based on transcription of genomic viral RNA by the murine RNA poly-
merase I promoter, allowing virus recovery only from murine cells. In order to
develop an efficient reverse genetic system in human cells, we first gener-
ated a minigenome assay based on the human RNA polymerase I promoter,
using Gaussia luciferase and the green fluorescent protein (GFP). Next, we
used this approach to successfully generate recombinant Lymphocytic Chori-
omeningitis Virus (rLCMV) from human cells and, more importantly, from FDA-
licensed Vero cells for the future development of potential vaccine seeds.
These novel reverse genetics techniques open new avenues for, both, the
identification of antivirals to treat arenavirus infections, and the generation
of recombinant arenavirus for the development of vaccines in FDA-approved
cell lines to prevent arenavirus infections in humans.
145 (A)
New Select Agent Regulations
J. Garrett, V. Sutton, H. Carson; Texas Tech Univ. Sch. of Law, Lubbock, TX.
Background: The U.S. Departments of Health and Human Services (HHS) and
Agriculture (USDA) were required to review and update the National Select
Agent Registry rules as promulgated in 42 C.F.R. 73, 9 C.F.R. 121, and 7 C.F.R.
331. The proposed final rules were published on October 3, 2011 in the Fed-
eral Register and contained several key changes important to researchers.
Methods: Prior to the publication of the new regulations, a thorough exami-
nation of the existing regulations, comments from the public, and review of
the recommendations of the Federal Experts Security Advisory Panel and
areas of probable change were identified. Results: Some, but not all, of the
recommendations were addressed in the new regulations, including those
from previous comment periods. The majority of the changes made were in
56 10th ASM Biodefense and Emerging Diseases Research Meeting
line with the areas of probable change identified during the regulation re-
view. Key changes have been identified and highlighted to aid in promotion
of the new regulations. Conclusion: The Center for Biodefense, Law & Public
Policy will conduct a comprehensive analysis and comparison of these new
regulations in relation to the old regulations.
146 (A)
Development of a Murine Model for Aerosolized Filovirus Infec-
tion
E. E. Zumbrun, H. A. Bloomfield, T. B. Chance, D. K. Nichols, A. Nalca; US-
AMRIID, Fort Detrick, MD.
Aerosol models of filovirus infection have been developed in guinea pig and
non-human primates; however, filovirus infection of wild-type mice by the
aerosol route has not been reported. A murine model of aerosolized filovirus
infection in mice will be useful for screening vaccine candidates and thera-
pies. In this study, various strains of wild-type and immunocompromised mice
were exposed to aerosolized wild-type (WT) or mouse-adapted (MA) Ebola
Zaire (EBOV). Upon exposure to aerosolized WT-EBOV, Balb/c, C57Bl6, and
DBA mice were unaffected, but 100% of SCID and 90% of Stat1 knock-out
(KO) mice succumbed to infection between 7-9 days post-exposure (dpe).
Balb/C mice lost approximately 15% of their body weight upon exposure to
MA-EBOV, but all mice recovered. In contrast, 10-30% lethality was observed
in C57Bl6 and DBA mice exposed to aerosolized MA-EBOV, and 100% of SCID,
Stat1 KO, Ifnγ -/- and perforin -/- mice succumbed to infection between 7-14
dpe. To identify one or more immunocompetent mouse strains in which ex-
posure to aerosolized MA-EBOV is uniformly lethal, 62 different BXD recom-
binant inbred (RI) mouse strains were tested for susceptibility or resistance
to infection. Aerosol exposure to a high dose of MA-EBOV produced a spec-
trum of disease severity, with most strains losing weight but recovering, and
a uniformly lethal disease within 7 to 12 dpe in five strains. Aerosol exposure
of these five BXD strains to 10-fold less MA-EBOV resulted in lethality rang-
ing from 0% in two strains to 100% lethality in only one strain. In the three
BXD strains with lethality at the lower dose of MA-EBOV, analysis of post-
mortem tissue showed liver damage as well as lung lesions in two of these
strains, but no lung lesions in the third strain. A computational genetic analy-
sis of the RI strains on the basis of outcome to aerosolized filovirus infection
may allow for identification of gene loci involved in susceptibility or resist-
ance to infection. Furthermore, the BXD strain for which even a low dose ex-
posure to MA-EBOV is uniformly lethal may be useful for testing vaccines and
therapies.
147 (B)
Antibiotic Profile of Nosocomial Bacterial Pathogens
N. N. Eduok1, I. P. Udoh2, C. I. Eleazar2, B. O. Ogeneh2; 1Ministry of Sci. and
Technology, Uyo, Nigeria, 2Univ. of Nigeria, Enugu, Nigeria.
Background: Antibiotic resistance among nosocomial bacteria is a global
problem. This is a concern to medical specialist and microbiologists espe-
cially as the choice of antimicrobial agents available for the treatment of such
infections become limited and inevitably more complicated. Methods: The
sensitivity pattern of nosocomial bacterial isolates from various clinical sam-
ples was investigated. A total of 450 samples were obtained from subjects
both gender of various ages. Isolates were identified using standard micro-
biological techniques. Antibiotic susceptibility test was carried out using disc
diffusion and modified agar dilution methods. The antibiotics tested against
the isolates were ampicillin (10μg), chloramphnicol (10μg), cloxacillin (5μg),
carbennicillin (30mcg), pefloxacin (5iu), ciprofloxacin (5iu), ofloxacillin (5iu),
gentamicin (10μg) and augmentin (5iu). Erythromycin (5μg), and strepto-
mycin (10μg) were tested against Gram positive organisms only, tetracycline
(10μg) and cotrimoxazole (25mcg) for Gram negative while nitrofurantoin
(200mcg), nalidixic acid (300mcg), were tested against the urine isolates

only.The MIC of ampicillin, gentamicin, ciprofloxacin used against some of
the clinical isolates were also carried out.Various strains were used as con-
trols. Result: Pathogens isolated were as follows: Staphylococcus aureus,
Klebsiella pneumoniae, Escherichia coli, Pseudomonas aeruginosa, Proteus
mirabilis, Enterococcus faecalis and Streptococcus pyogenes. Distribution of
infections in the various sites was as follows: urinary tract (44.5%), blood
(11.1%), genital tract (22.2%), upper/lower respiratory tract (11.1%), and
wound (11.1%). Ampicillin and gentamicin produced the highest MIC against
S. pyogenes, (3.75) and E. feacalis (1.875),while ciprofloxacin showed the
lowest MIC against E.coli (0.235) and greatest activity against the various
isolates. Conclusion: The high frequency of nosocomial infections places a
substantial burden on individual patients and on the health care system while
emergence of antibiotic resistance has become a medical catastrophe.
148 (B)
Comparative Transcriptomics of Three Species of Fleas
(Siphonaptera: Pulicidae)
C. A. Whitehouse1, G. Koroleva1, J. R. Kugelman1, S. K. Williams2, S. W. Bear-
den2, K. L. Gage2, G. F. Palacios1; 1USAMRIID, Frederick, MD, 2CDC, Fort
Collins, CO.
Background: Fleas serve as vectors of several important diseases. In partic-
ular, the oriental rat flea, Xenopsylla cheopis, is the primary vector of Yersinia
pestis, the causative agent of plague throughout much of the world. How-
ever, the ground squirrel flea, Oropsylla montana, is considered to be the pri-
mary vector of plague in the western U.S. The cat flea, Ctenocephalides felis,
can rarely transmit Y. pestis, but is a more important vector of bartonellosis,
murine typhus, and the dog tapeworm Dipylidium caninum. Despite their im-
portance, very little is known at the genomic level of these flea vectors, es-
pecially related to their transcriptome. Methods: Pools of 20 individual
colony-reared fleas from each of the three species were homogenized in 0.1
ml of TRIzol reagent, and total RNA was extracted. cDNA libraries were con-
structed using the SMART cDNA library construction kit to enhance se-
quencing. Libraries were processed for either 454 or Illumina sequencing.
Hybrid assemblies were produced using Lasergene nGen and BLASTx was
used to compare contigs to the GenBank non-redundant database. Results:
In total, we obtained 7.10 x 106 (Illumina) and 2.02 x 105 (454) reads for X.
cheopis, 9.90 x 106 and 1.88 x 105 reads for O. montana, and 8.90 x 106 and
1.69 x 105 reads for C. felis, which resulted in 16,257 contigs, 16,831 con-
tigs, and 14,307 contigs, respectively. BLAST searching of non-redundant
databases yielded ~15.4K significant matches to gene families potentially
encoding gene products for X. cheopis, ~15.6K for O. montana, and ~13.1K
for C. felis. Here we compare transcript identity and abundance between the
three flea species. Conclusions: This is the first study to elucidate the global
transcriptome of three species of fleas that are important disease vectors
and lays the foundation for future studies of the molecular mechanisms in-
volved in vector-pathogen interactions.
149 (B)
Development of a Coxiella burnetii Aerosol Infection Model in
Rodents
K. R. Bewley, J. K. Pitman, D. J. Atkinson, G. McLuckie, M. Charlwood, G.
Hatch, J. Vipond, S. G. P. Funnell, A. Roberts; Hlth. Protection Agency, Salis-
bury, United Kingdom.
Background: Coxiella burnetii is the causative agent of Q fever. Infection can
take several forms, although in man disease ranges from asymptomatic,
through acute, to life-threatening chronic illness. Chronic disease develops in
approximately 5% of those infected. Methods: In order to develop an aerosol
model of infection, we needed to ensure that the working stocks we produced
were virulent. As a result, we initially prepared a Master stock of C. burnetii
(Nine Mile strain, 1993) in guinea pigs then prepared a working stock from
T U E S DAY A B S T R AC TS
this in fertile hen’s eggs. The two stocks were compared before aerosol stud-
ies. As a virulent US strain was not possible to obtain, HPA recovered a vial
of yolk sac (YS) material containing C. burnetii (dated 1993) obtained origi-
nally from ATCC (RSA 423 9mi, 306GP/2EP, 4/26/71). This was used to infect
three male guinea pigs IP. Spleens were harvested and homogenised at peak
febrile response and used to inoculate 80 hen eggs. After 11 days, YSs were
harvested and homogenised. Results: Both C. burnetii stocks were inocu-
lated IP into guinea pigs at a dose adjusted to be equivalent to 1.5E+07
copies by qPCR (sod gene). Both groups of guinea pigs showed clinical signs
of infection with C. burnetii (body temperature increase and body weight re-
duction) but, unlike the spleen stock dosed group which all survived, 75%
of the group that received YS material had to be euthanised due to severity
of disease. These data suggest that the YS material may be more virulent.
The YS material was then delivered via the aerosol route to 3 groups of ani-
mals (guinea pigs and 2 strains of mice). The guinea pig group displayed clin-
ical signs of infection (increase in temperature and failure to gain body
weight) but the mice displayed no or minimal clinical signs. These data sug-
gest that PCR alone is inadequate in determining the virulence of an inocu-
lum of C. burnetii and guinea pigs are more susceptible than mice to aerosols
of this stock of C. burnetii. Conclusion: We have developed an aerosol model
of C. burnetii infection in guinea pigs which replicates the clinical signs seen
in acute human infection.
150 (B)
Automated Extraction of Genomic DNA from Food-Associated
Pathogens and Environmental Samples
P. Williams, T. Parrish, M. Duggan, J. Chapman, W. Burke, J. Birkenholz; Evo-
gen, Lenexa, KS.
Introduction: The extraction and purification of DNA from the bacteria Es-
cherichia coli O157:H7 and Salmonella enterica are essential first steps for
the successful amplification of gene targets by PCR. Automated sample prep
systems allow for less hands-on time, provide greater consistency of ex-
tracted sample, and use fewer consumables. We have developed a fully Au-
tomated Sample Preparation (ASP) instrument which was used to extract and
purify DNA from ground beef and turkey meat cultures and environmental
surface swab cultures. Method: 24 hour cultures of ground beef, turkey, pork,
and environmental swabs were established using standard USDA protocols.
200 μl of culture media from the meat and swab samples were placed in the
ASP cartridge and then extracted and purified using the following automated
protocol: 200 μl of Evogen ONE was added to the samples which were then
incubated at 95°C for 5 minutes. 250 μl of binding buffer was added to the
samples, followed by an incubation of 5 minutes. DNA bound to the magnetic
beads was washed and then incubated at room temperature for 5 minutes to
elute the purified DNA. 200 μl of purified DNA was then automatically trans-
ferred to an elution tube. DNA yields were measured with a NanoDrop sys-
tem. Validated primers and HyBeacons probes were targeted to the
Salmonella ttr gene and the E. coli eae (gama) gene. 3 μl of the purified DNA
was used in a standard PCR reaction using the following parameters: 95ºC for
15 seconds, 55ºC for 30 seconds, 72ºC for 30 seconds; for 40 cycles on a Bio-
Rad CFX 96 thermocycler. Results: DNA yields from ground beef and swab
cultures were 26 ng/μl and 17.2 ng/μl respectively. The mean DNA yields from
ground turkey and swab cultures were 21.9 ng/μl and 20.2 ng/μl respectively.
Complete analysis time was 3 hours. All gene targets were successfully am-
plified. Conclusion: The use of the ASP for genomic DNA extraction from S. en-
terica and E. coli O157:H7 proved to be a rapid and simple protocol which
provided high quality DNA for PCR amplification.
BIODEFENSE PROGRAM & ABSTRACTS 57
T U E S D A Y P O S T E R S E S S I O N | 1:00–3:00 p.m. | Exhibit Hall
151 (B)
Monoclonal Antibody against Capsule of Bacillus anthracis
Protects Mice from Enhanced Lethal Toxin Activity Due to Cap-
sule and Anthrax Spore Challenge
G-E. Rhie1, J. Jang1, M. Cho1, J-H. Chun1, J. Park2, K. Hong1; 1Korea NIH, Chung-
buk, Korea, Republic of, 2Hankuk Univ. of Foreign Studies, Yongin, Korea, Re-
public of.
The poly-γ-D-glutamic acid (PGA) capsule is one of the major virulence factors
of Bacillus anthracis, which causes a highly lethal infectious disease. The PGA
capsule has been known to disguise B. anthracis from immune surveillance
and allow its unimpeded growth in the host. Recently, the importance of PGA
in the pathogenesis of anthrax infection is being emphasized. The PGA cap-
sule is associated with lethal toxin (LT) in the blood of experimentally infected
animals and PGA was shown to produce enhancement of the cytotoxicity of
LT through increasing binding of protective antigen to its receptors resulting
more accumulation inside cells. In this study, we confirmed the enhancement
effect of PGA on LT-mediated cytotoxicity ex vivo using mouse bone marrow
derived macrophage (BMDM). Western blot analysis showed that the com-
bination of PGA and LT produced a greater degree of degradation of MAP ki-
nase kinases and an increased level of activation of the proform of caspase-1
to its processed form as compared to LT alone. For the evaluation antibody
against PGA on the prevention of progress of anthrax disease, mouse mono-
clonal antibodies (mAbs) against PGA were produced. mAbs bind encapsu-
lated B. anthracis H9401 specifically and showed opsonophagocytosis
activity against encapsulated B. anthracis H9401. Anti-PGA mAbs also
showed protective effect against both spore and PGA enhanced LT toxicity
challenges in mice. These results indicate that anti-PGA mAb protects the
host from the progression of anthrax not only by eliminating vegetative form
of encapsulated B. anthracis but also from by inhibiting the enhancing cyto-
toxic activity of LT by PGA and suggest a role of anti-PGA antibody in pre-
venting and treating anthrax infection.
152 (B)
The Cat Flea as a Laboratory Model for Understanding Yersinia
pestis Colonization of its Insect Vector
T. M. Hermanas, L. E. Quenee, O. Schneewind; Univ. of Chicago, Chicago, IL.
In North America, 28 species of flea have been confirmed as vectors capable
of transmitting Yerinsia pestis and maintaining the zoonotic/epizootic cycle.
Epizoonotic epidemics of plague are thought to initiate when fleas acquire
Yerinsia pestis by feeding on heavily septicemic rodents with subsequent
transmission to human hosts. Research on the rat flea (Xenopsylla cheopis)
suggests that roughly 50 percent of fleas that feed on a septicemic host are
colonized by Yersinia pestis. Colonization requires the formation of a biofilm
in the flea’s midgut, which eventually causes the infected flea to starve and
die. Colonization of the flea midgut has been linked to the production of n-
acetyl- glucosamine by the heamin storage (hms) locus and phospolipase D
activity of the Yersinia murine toxin gene (ymt). Our goal is to develop a model
for colonization of the cat flea (Ctenocephalides felis) to help identify the
Yersinia pestis requirements for flea colonization. Using an artificial feeding
system, we observe transmission of Yersinia pestis CO92 pCD1- from blood
to the flea with subsequent persistent colonization. The progress of the in-
fection is monitored using CFU enumeration and microscopy. Our results
show that Y. pestis persistence requires gradual growth in the digestive tract
for 21 days after feeding of the initial infectious dose. Experiments with wild-
type Y. pestis and isogenic mutants show that the hms and ymt genes are re-
quired for persistent infection of the flea digestive tract. Additionally, we
utilize the cat flea model to screen iron transport genes in an attempt to un-
derstand the role of iron acquisition during flea colonization.
58 10th ASM Biodefense and Emerging Diseases Research Meeting
153 (B)
Bacillus anthracis Acetyl-Transferases PatA1 and PatA2 and
Secondary Cell Wall Polysaccharide-Mediated Assembly of S-
Layer Proteins
J. M. Lunderberg1,2, S-M. Nguyen-Mau1,2, G. S. Richter1,2, Y-T. Wang1,2, J.
Dworkin3, D. M. Missiakas1,2, O. Schneewind1,2; 1Univ. of Chicago, Chicago,
IL, 2Howard Taylor Ricketts Lab., Argonne, IL, 3Columbia Univ., New York, NY.
The envelope of Bacillus anthracis, the causative agent of anthrax, is com-
prised of a poly-D-γ-glutamic acid (PDGA) capsule, a proteinaceous S-layer
with S-layer associated proteins and the cell wall peptidoglycan. Recent work
suggests that, in addition to PDGA, S-layer and S-layer associated proteins
contribute to the pathogenesis of anthrax. Protein assembly in the envelope
of B. anthracis requires S-layer homology domains (SLH) within S-layer and
S-layer associated proteins. SLH domains fold into a three-pronged spindle
structure that associates with the secondary cell wall polysaccharide (SCWP),
a carbohydrate that is tethered to the peptidoglycan of B. anthracis. Earlier
work described the structure of SCWP as [→6)-α-GlcNAc-(1→4)-β-ManNAc-
(1→4)-β-GlcNAc-(1→]n, where α-GlcNAc is substituted with α-Gal and β-Gal
at O3 and O4, respectively, and the β-GlcNAc is substituted with α-Gal at O3.
The SCWP is modified with pyruvyl and acetyl moieties. Pyruvylation involves
the csaB gene, which is essential for S-layer assembly and whose product is
thought to function as a pyruvyl-transferase. Here, the effects of two putative
acyl-transferases, PatA1 and PatA2, on SCWP acetylation are investigated
through MALDI-TOF mass spectrometry and fluorescent protein binding as-
says. Our data suggest PatA1 and PatA2 act on the SCWP and affect the dep-
osition of some, but not all, SLH-domain containing proteins into the S-layer
of B. anthracis.
154 (B)
Autotransported (Type V Secreted) Proteins of Burkholderia
pseudomallei and Their Role in Biofilm Formation, Serum Re-
sistance and Pathogenesis
N. R. Lazar Adler1, R. E. Dean2, M. P. Stevens3, J. L. Prior2, T. P. Atkins2,4, E. E.
Galyov1; 1Univ. of Leicester, Leicester, United Kingdom, 2Defence Sci. and
Technology Lab., Salisbury, United Kingdom, 3Univ. of Edinburgh, Easter
Bush, United Kingdom, 4Univ. of Exeter, Exeter, United Kingdom.
Burkholderia pseudomallei, a saprophyte endemic to the soil and water of
tropical regions, is the causative agent of Melioidosis, a potentially fatal
febrile human and veterinary disease. The molecular mechanisms of B.
pseudomallei pathogenesis remains to be fully characterised. Autotrans-
porters are a family of secreted or membrane-associated proteins which me-
diate their own secretion across the Gram-negative membranes via the type
V secretion pathway. Autotransporters from different bacterial species have
diverse functions, with many demonstrated to play a role in virulence. Eleven
genes encoding predicted autotransporters are present in the B. pseudoma-
llei K96243 genome (AT1-11). Most of these autotransporters remain un-
studied; hence, single cross-over insertional mutants were constructed and
subjected to a range of investigations to elucidate their role in pathogenesis.
All mutants were assessed for intracellular invasion and survival within the
A549 epithelial-like cell line with two mutants, AT7 and AT11, demonstrating
attenuation. Furthermore, AT11 also displayed attenuation for intracellular
survival in the J774 macrophage-like cell line. Biofilm assays were performed
for all mutants and AT5 demonstrated reduced biofilm formation at 37°C but
wild-type biofilm levels at 27°C. Evaluation of the survival of all mutants in
45% normal human sera identified that AT6 was sensitive to complement-
mediated killing. Eight mutants have been screened in the BALB/c Melioido-
sis mouse model, with five demonstrating attenuation for virulence (varying
from a 1 to a 1.6-log reduction). Taken together, our data indicate that several
putative Type V secreted proteins play a role in the virulence of B. pseudo-
mallei and warrant further study. (c) Crown Copyright. Dstl, 2011

155 (B)
S-Layer Assembly in Bacillus cereus G9241 Requires csaB
Y-T. Wang1,2, S-Y. Oh1,2, A. P. Hendrickx1, O. Schneewind1,2; 1Univ. of Chicago,
Chicago, IL, 2Howard Taylor Ricketts Lab., Argonne, IL.
The surface (S)-layer is a structure of crystalline protein arrays deposited on
top of the cell envelope of many bacterial species. We sought to identify and
characterize the S-layer in Bacillus cereus G9241, which causes an anthrax-
like respiratory illness in humans and immune-competent mice. Our results
suggest that B. cereus G9241 immobilize S-layer proteins in their envelope in
a csaB dependent manner. The product of the csaB gene is thought to pyru-
vylate the secondary cell wall polysaccharide of Bacillus anthracis and Ther-
mus thermophilus, thereby capturing secreted proteins with an N-terminal
S-layer homology domain (SLH). B. cereus csaB variants cannot retain the
main S-layer proteins, Sap and EA1, in the bacterial envelope. Further, com-
pared to the wild-type, the csaB variant displays altered colony morphology
and a long chain growth phenotype of its vegetative forms. When evaluated
in an intraperitoneal challenge model with C57Bl/6 mice, the csaB mutant
was attenuated as compared to wild-type B. cereus G9241. In addition to S-
layer proteins, B. cereus G9241 also elaborates two capsular polysaccha-
rides, which are encoded by plasmids pBCXO1 and pBC218. Our results
suggest that the assembly and function of the S-layer and the capsular poly-
saccharides is carefully coordinated to enable the pathogenesis of anthrax-
like disease.
156 (B)
Antibody Interactions with the Capsule of Bacillus anthracis
M. A. Hubbard, P. Thorkildson, W. Welch, D. AuCoin, T. Kozel; Univ. of Nevada
Sch. of Med., Reno, NV.
Bacillus anthracis is the etiologic agent of anthrax, a disease of large mam-
mals including humans. The bacterium synthesizes a defensive capsule en-
tirely composed of poly-gamma-D-glutamic acid (PGA). We previously
demonstrated that passive immunization with a murine monoclonal antibody
(mAb) reactive with PGA is protective in a pulmonary model of anthrax. These
results are part of a growing body of research that has demonstrated the
merit of capsule-targeted immunity, leading to considerable interest in the
incorporation of PGA into anthrax vaccines. In the present study we sought to
further understand the modality of antibody binding to this unique antigen.
The original mAb was subject to CDR3 randomization and phage display,
which lead to the recovery of a high affinity variant. Binding of both the par-
ent and mutant antibodies was analyzed using surface plasmon resonance.
The results of this study indicate that the association of mAb to PGA consists
of two kinetic classes: the faster is first order in mAb concentration, the
slower zero order in mAb concentration. Dissociation appears to occur as a
single kinetic class. A molecular model was constructed to provide an atom-
istic explanation. Molecular dynamics simulations in explicit solvent demon-
strate that the polyvalent nature of PGA is exploited by multiple interactions
with the surface of the antibody. Molecular contacts occur along a positively
charged groove. Although traditional CDRs were involved, surprisingly,
residues outside the region were large contributors to PGA binding. These
results may indicate an alternative method of antibody binding and selection
to combat polyanions by the immune system.
T U E S DAY A B S T R AC TS
157 (B)
Salmonella Enteritidis Gene SEN1140 Contributes to Acceler-
ated Type III Secretion System-2 Dependent Early Inflamma-
tion Kinetics in Mouse Colitis Model
V. VISHWAKARMA1, B. Periaswamy2, N. B. Pati1, W-D. Hardt2, M. Suar1; 1Sch.
of Biotechnology, Bhubaneswar, India, 2Inst. of Microbiol., Zürich, Switzer-
land.
Salmonella enterica subsp. I serovar Enteritidis (S. Enteritidis) exhibits faster
type III secretion system 2 (TTSS-2) dependent early colonization and in-
flammation kinetics compared to its closely related serovar Typhimurium. To
understand the molecular mechanistic differences, in particular to investi-
gate the TTSS-2 dependent accelerated pathogenic potential of S. Enteritidis,
we focused on genes unique to its genome. Previously published compara-
tive genomic findings revealed presence of mutually exclusive genes in both
serovars. As a pilot study we investigated the role of six unique genes of S.
Enteritidis in-vivo. Wild type isogenic tagged strains (WITS) of S. Enteritidis
with plasmid carrying gene specific putative promoter constructs with a
downstream GFP reporter were used to coinfect C57BL/6 mice and were later
interpreted by differential fluorescence induction (DFI), fluorescence acti-
vated cell sorting (FACS) followed by quantitative real-time polymerase chain
reaction (qRT-PCR) for WITS tags to identify differentially expressed promot-
ers in sorted fractions. Using this strategy, promoter construct for gene
SEN1140 was found to express GFP in gut lumen, lamina propria, mesenteric
lymph nodes, spleen and liver. To investigate the role of SEN1140, knockout
mutant of the same was compared with S. Typhimurium on a type III secre-
tion system 1 deficient (TTSS1-) background. Interestingly, the S. Enteritidis
mutant lacking SEN1140 did not show the unique early colonization and in-
flammation kinetics phenotype in comparison to S. Typhimurium. To sub-
stantiate it, complementation of SEN1140 restored the TTSS-2 dependent
accelerated inflammatory potential of S. Enteritidis. Furthermore, this pilot
study represents a suitable combination of DFI, FACS, qPCR and WITS tech-
niques to sensitively screen S. Enteritidis specific genes for their unique role
in bacterial pathogenesis.
158 (B)
Activity of Two Novel Antibacterial Agents XF-70 & XF-73
against Multidrug-Resistant Acinetobacter baumannii
K. L. Lohman1, J. Coffin1, W. Rhys-Williams2, I. Hayter2, W. G. Love2;
1
MRIGlobal, Rockville, MD, 2Destiny Pharma Ltd., Brighton, United Kingdom.
Background: XF-70 and XF-73 are members of a new class of membrane ac-
tive topical drugs which have broad spectrum activity against a range of bio-
threat agents, as well as other clinically relevant bacteria such as MRSA. This
intrinsic activity is, in certain cases, augmented by light activation to produce
an additional photodynamic effect. Here we report the results of an in vitro
screen against a panel of clinically relevant multidrug resistant Acinetobac-
ter baumannii isolates with and without light activation. Methods: Multidrug
resistant A. baumannii isolates from burn, bloodstream and upper and lower
respiratory tract infections were selected from the ATCC culture collection.
MICs were determined by broth microdilution according to standard CLSI
methodology (M100S21: Performance Standards for Antimicrobial Suscepti-
bility Testing, January 2011). Colistin and Imipenem were included as con-
trols. For light activation studies, the plates were illuminated with a light
source with a wavelength range of 380 - 480nm for 15 minutes. Staphylo-
coccus aureus and Pseudomonas aeruginosa were included as bacterial con-
trols. Results: The intrinsic A. baumannii MICs for XF-70 were in the range of
32 to 64 µg/ml, which reduced to <0.25 µg/ml after 15 minutes illumina-
tion. The intrinsic A. baumannii MICs for XF-73 were reduced from 64-128
µg/ml to <0.25 µg/ml after illumination. The antibiotic resistance profile of
the isolates had no effect on the activity of either XF-70 or XF-73. MICs for
the control drugs were as expected. Conclusions: The results reported here
BIODEFENSE PROGRAM & ABSTRACTS 59
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demonstrate that light activated XF-70 and XF-73 have potent activity against
all of the A.baumannii isolates tested, irrespective of their drug resistance
and infection source. The results suggest that XF-70 and XF-73 may be of util-
ity in the treatment of A.baumannii infections and is worthy of further inves-
tigation
159 (B)
Anthrax Surveillance System and Canonical SNP Typing of
Bacillus anthracis Strains Isolated in the Country of Georgia
2009-2011
G. Chanturia1, E. Khmaladze1, E. Zhgenti1, L. Malania1, M. Zakalashvili1, N.
Tsertsvadze1, M. Kekelidze1, P. Imnadze1, S. Tsanava1, R. J. Obiso2, S.
Francesconi3; 1The Natl. Ctr. for Disease Control and Publ. Hlth., Tbilisi, Geor-
gia, 2The Microbe Company, Christiansburg, VA, 3Naval Med. Res. Ctr., Fred-
erick, MD.
Bacillus anthracis, the causative agent of anthrax, due to its high virulence
and ability to persist as a spore, poses a great threat as an emerging bioter-
rorism agent. The epidemiology of this pathogen was studied in “Active Sur-
veillance of Especially Dangerous Pathogens in the Southern Caucuses
Region” project, focused on monitoring the prevalence and distribution of
several especially dangerous pathogens, natural foci of which are located on
the territory of the South Caucasus. Within the project framework 1989 en-
vironmental samples have been investigated in 2009-2011. Samples were
plated on PLET and SDA media. Suspicious for anthrax colonies were inves-
tigated according to SOPs provided by DTRA. The final confirmation was per-
formed using Real-time PCR. Totally 63 out of obtained 148 B. anthracis
strains were chosen for further investigation. These isolates in addition with
archival strains from National Center for Disease Control and Public Health
(NCDC) were characterized using canonical SNPs in order to place them in a
global phylogeographic context. From this analysis we discovered two very
distinct and relatively distant populations of B. anthracis. Majority of the
strains belonged to the A.Br.003 together with Australian 94 lineage and very
few of them went to the A.Br.008/009 (WNA lineage). Within the small geo-
graphic scale of Georgia, the identification of two distinct and genetically dis-
tant populations of B. anthracis suggests that Georgia is the site of multiple
waves of B. anthracis dispersals from different origins. Obtained results re-
vealed need for further in depth epidemiological study of the cases.
160 (B)
Diversity of Bacteriophages Active to B. anthracis Bacterial
Strains Isolated in Georgia
S. Rigvava1, M. Natidze2, L. Gubeladze1, L. Kvachadze1, R. J. Obiso3, M. Ku-
tateladze1, R. Adamia1; 1G. Eliava Inst. of Bacteriophages, Microbiol. and Vi-
rology, Tbilisi, Georgia, 2G, Tbilisi, Georgia, 3The Microbe Company,
Christiansburg, VA.
Bacteriophages are bacterial viruses that reveal specific lytic activity against
host bacterial strains. The Eliava Institute, Tbilisi, Georgia is one of the most
experienced institutes for phage isolation, research, and application. Several
bacteriophages active against B. anthracis have been characterized. The aim
of this research study is to isolate, characterize, and study new specific
phages against B. anthracis. Several lytic phage were isolated from two, ge-
ographically distinct loci: one from West Georgia (12 phage from the Kutaisi
region) and the second from Eastern Georgia (6 phage from the Gardabani re-
gion). The phages were isolated and then propagated on vaccine strains of B.
anthracis (34F2, 55 and STI1). Bacteriophage isolated in this study have the
same morphology as the internationally accepted B. anthracis bacteriophage
Gamma— the phages have hexagonal heads and long, non-contractile tails.
All phages are very specific and do not lyse any other Bacillus species other
than strains of Bacillus anthracis. Each of the phage isolated differ from each
other by DNA restriction and structural proteins. The phages were tested
60 10th ASM Biodefense and Emerging Diseases Research Meeting
against a virulent strain collection (A total of 37 strains were evaluated). All
phages revealed different lytic activity against virulent strains. Further re-
search will be focused on research of biological peculiarities of the genetically
different B. anthracis phages, which could lead to a more elaborate and im-
proved diagnostics capability of phage typing of B. anthracis.
161 (B)
Characterisation of Inhalational Infection with Burkholderia
pseudomallei in BALB/c mice
S. G. P. Funnell, G. Hatch, J. A. Kane, S. Bates, K. M. Tonks, C. H. Tsang, J.
Fretwell, H. Shuttleworth, J. Vipond, A. Roberts; Hlth. Protection Agency, Sal-
isbury, United Kingdom.
Background: Burkholderia pseudomallei is the aetiologic agent of melioido-
sis, a disease which is endemic in Southeast Asian and Northern Australia.
Natural infection can occur via a range of routes including contamination of
skin abrasions, inhalation of aerosols and ingestion of contaminated water.
The aim of this study was to characterise inhalational infection of BALB/c
mice with B. pseudomallei. Methods: Groups of BALB/c mice were challenged
by the aerosol route using a modified Henderson apparatus in conjunction
with the Biaera aerosol management platform with different doses ranging
from 3343 cfu to 3 cfu of B. pseudomallei strain K96243. Animals were
weighed and observed for clinical signs. In selected studies, at scheduled
time points, the lung, spleen, kidney, and liver tissues were collected for
histopathological profiling and also for enumeration of bacterial load using
Luria Bertani Agar. Ashdown’s agar was also used to assess morphological
variation. Enumeration was conducted using an SOP based on classical
colony counting, and morphology variation was recorded photographically.
Results: At high doses (above 500 cfu), all mice develop an acute disease
which resulted in death within ten days of infection. At low doses, the ma-
jority of mice survived beyond 14 days after infection. The disease in these
survivors appeared to take a more indolent course. The histopathology and
bacterial loads of the sampled tissues indicated that the disease progression
in survivors at day 30 would inevitably have led to a relapse. In addition, the
degree of B. pseudomallei morphological variation observed in tissue iso-
lates was greater than that seen in the challenge inoculum. Conclusion:
These results indicate that BALB/c mice can be used to model human infec-
tion with B. pseudomallei and that the challenge dose will determine whether
an acute or chronic manifestation of disease is observed.
162 (B)
Reaerosolization of Bacillus Spores During the Bio-Response
Operational Testing and Evaluation
S. C. Taft1, R. A. Lordo2, D. J. Chappie2, E. Silvestri1, T. L. Nichols1, J. A. Rosati
Rowe1; 1Natl. Homeland Security Res. Ctr., Cincinnati, OH, 2Battelle Mem.
Inst., Columbus, OH.
Background: The Federal Interagency Bio-Response Operational Testing and
Evaluation (BOTE) project was conducted to evaluate the efficacy of three de-
contamination technologies on the Bacillus anthracis spore surrogate, Bacillus
atrophaeus subspecies globigii (Bg), disseminated in a building. During BOTE,
complex air and surface sampling was also conducted to quantify potential ex-
posure from the aerosolized spores during the release, from the subsequent
re-aerosolized spores pre-decontamination, and from the re-aerosolized resid-
ual spores post-decontamination. Methods: SKC Bioaerosol samples were col-
lected at five different time stages (background, during dissemination, before
surface sampling, during pre-decontamination surface sampling, and during
post-decontamination surface sampling) for each of the three decontamination
technology test rounds (amended bleach, chlorine dioxide fumigation, and va-
porized hydrogen peroxide fumigation). Wipes, sponge sticks, and vacuum sur-
face samples were also collected in the same rooms during the last two time
stages in conjunction with the SKC Bioaerosol samples. The surface concen-

tration measurements were linked to the air concentration measurements to
parameterize potential reaerosolization of the spores due to human surface
sampling activity within a contaminated indoor setting. Results: The back-
ground and post-decontamination air and surface samples were primarily non-
detects. There were significant differences observed in the air sampling results
for the other the time stages (dissemination, before surface sampling, and dur-
ing pre-decontamination surface sampling). The surface to air concentration
measurement comparisons varied considerably among the different surface
sampling teams, surface sampling collection methods, and surface types. Con-
clusions: These data confirm that single, point reaerosolization rates or factors
are not sufficient for predicting potential reaerosolization from different sur-
face types or human activities.
163 (B)
Regulation of the Yersinia Type Three Secretion Pathway
K. Kopaskie1,2, J. McCool3, K. Aly1, B. Blaylock1,2, O. Schneewind1,2; 1Univ. of
Chicago, Chicago, IL, 2Howard Taylor Ricketts Lab., Argonne, IL, 3Boston Univ.,
Boston, MA.
Three pathogenic Yersinia species—Yersinia pestis, Yersinia pseudotubercu-
losis and Yersinia enterocolitica—use a virulence plasmid-encoded type III se-
cretion machine to inject effector proteins into host immune cells. The Yersinia
type III secretion pathway is essential for the pathogenesis of human or animal
disease and some of its subunits, which are currently being tested as purified
antigens for plague vaccine development. During host infection, Yersinia in-
sert their preassembled secretion needles into the plasma membrane of im-
mune cells. This encounter is perceived by the YscF needle protein as a drop in
calcium concentration, triggering the synthesis and secretion of effector pro-
teins (YopE, YopH, YopJ, YopM, YopO, and YopT). A hydrid protein, YopN-TyeA,
which is comprised of two other regulatory factors foe effector transport, com-
pletely blocked Yersinia type III secretion and rendered bacteria non-respon-
sive to calcium signaling. YopN-TyeA was localized to the bacterial membrane
fraction. Detergent solubilized membrane proteins were used for YopN-TyeA
affinity chromatography experiments to identify machine components that may
contribute to the shut-down of the Yersinia type III pathway.
164 (B)
Real-Time Monitoring of Bacillus thuringiensis as a Surrogate
for Bacillus anthracis in Potable Water Distribution System
S. P. Sherchan, I. Pepper; Univ. of Arizona, Tucson, AZ.
Following the events of September 11, 2001, and the subsequent anthrax
mail attack on the Eastern United States, the focus on bioterrorism in partic-
ular has become significant. Therefore, there is a need for integrated and in-
telligent sensors to operate in real-time,with the ability to recognize and
diagnose day-to-day and perhaps minute-to-minute water quality distur-
bances. Currently there is no available technology that will specifically iden-
tify a microorganism in real-time; however, there are commercial sensors
available that can detect microbial water quality changes in less than five
minutes with a sufficient sampling rate as to be considered “real-time.” The
objective of this study was to evaluate the potential for real-time monitoring
of B.thuringiensis as a surrogate for B. anthracis using a variety of water qual-
ity sensors including: the HACH Guardian Blue Monitoring Platform; the JMAR
BioSentry unit; and the S::CAN spectro::lyser technology. The minimum
threshold responses were examined by injecting B.thuringiensis either in
deionized (DI) or tap water with spore concentration ranging from 102- 105
colony forming units (cfu)/mL. Most sensors responded to increase in spore
concentration over the range of 102 - 105 cfu/ml and below this range; sen-
sors provided signals undistinguishable from noise. The data obtained thus
far suggest that select sensors can detect microbial water quality changes
and these advanced technologies can be integrated to monitor intrusion
events in water distribution systems.
T U E S DAY A B S T R AC TS
165 (B)
Achromobacter Xylosoxidans: An Emerging Pathogen Carrying
Different Elements Involved in Horizontal Genetic Transfer
G. Traglia1, C. Adams2, M. Almuzara3, C. Vay3, D. Centrón1, M. Ramirez2,1; 1Sch.
of Med., Buenos Aires, Argentina, 2California State Univ., Fullerton, CA, 3Hosp.
de Clinicas, Jose de San Martin, Buenos Aires, Argentina.
In the last few years numerous cases of multidrug resistant Achromobacter
Xylosoxidans infections have been documented in immunocompromised and
cystic fibrosis patients. To gain insights into the molecular mechanisms and
mobile elements contributing to multidrug resistance of this bacterium, we
studied 28 non-epidemiological A. xylosoxidans clinical isolates from Ar-
gentina. All isolates were identified using standard biochemical tests, API
20NE (Biomeriux), and 16S rRNA gene sequencing. Clonal relationships were
determined by macrorestriction mapping. Total DNA was used as template
for PCR amplification reactions. Specific primers for plasmids (IncP, IncW,
IncA/C, IncN, IncFII, repAci1), transposons (Tn1331, Tn3, Tn7), insertion se-
quences (IS) (IS26, IS1999, IS440, ISPa12, ISPa13, IS18, IS125, IS1008,
ISAba1), and the blaampC, intI1 and intI2 genes were used in PCR reactions.
The variable regions (vr) of integrons and their association with IS26 and
IS440 were analyzed by PCR cartography. IS26 and IS440 were found in 2
and 7 isolates, respectively. Wide host range IncP plasmids were identified in
10 isolates. The intI1 and intI2 genes were present in 10 and 3 isolates, re-
spectively. PCR cartography showed 4 different arrays in the vr of class 1 in-
tegrons. Relevant gene cassettes found in these vr include the
aminoglycoside resistance aac(6’)-Ib and aadA1, the trimethoprim resistance
dfrA1 and dfrA16, and the beta-lactamase blaOXA-2. In only one of the class 2
integrons a vr was amplified, which included dfrA1. No association of inte-
grons with IS26 and IS440 were found. The blaampC gene was found in all
isolates, confirming its ubiquitous nature. Our results show that A. xylosox-
idans clinical isolates contain a rich variety of genetic elements commonly
associated with resistance genes and their dissemination. This supports the
hypothesis that A. xylosoxidans is becoming a reservoir of resistance deter-
minants included in mobile elements.
166 (B)
Increase in Temperature of Cultivation Above 37°C Influenced
Production of Yersinia pestis Major Protective Antigens F1 and
LcrV
V. A. Feodorova1, T. I. Polyanina1, M. A. Khizhnyakova1, A. M. Lyapina1, O. V.
Ulianova2,1, V. L. Motin3; 1Saratov State Vet. Inst., Saratov, Russian Federa-
tion, 2Saratov State Univ., Saratov, Russian Federation, 3Univ. of Texas Med.
Branch, Galveston, TX.
Background: Human plague, which is an acute and often a fulminant disease,
is characterized by a rapid onset of high fever up to 40-42°C. The purpose of
this study was to investigate whether Y. pestis responds to elevated tem-
perature by changing its production of specific antigens. Methods: Y. pestis
EV NIIEG was grown in Heart Infusion Broth (HIB) supplemented with cal-
cium, magnesium, glucose, or a combination thereof at physiological con-
centrations, either at pH 7.2 or 5.5 and temperatures in the range of
26°C-42°C. Production of two major Y. pestis virulence factors and protective
antigens, such as capsular antigen F1 and LcrV, was evaluated in dot-ELISA,
SDS-PAGE and immunoblots by using monospecific polyclonal antibodies to
these proteins. In addition, the existence of a capsule was visualized by using
both light and electron microscopy. Results: Biosynthesis of both F1 and LcrV
by Y. pestis EV NIIEG at neutral pH in HIB with no supplements was signifi-
cantly enhanced at growth temperatures above 39°C with a maximum at 40-
42oC. Addition of either calcium or magnesium in combination with glucose
elevated the production of LcrV, but not of F1. The medium containing either
all three components or HIB with magnesium alone also stimulated the pro-
duction of both antigens at an elevated temperature with a peak at 38°C-
BIODEFENSE PROGRAM & ABSTRACTS 61
T U E S D A Y P O S T E R S E S S I O N | 1:00–3:00 p.m. | Exhibit Hall
39°C. When production of F1 and LcrV was tested at acidic pH, we observed
similar trends indicating that production of these antigens correlated with the
increase in cultivation temperature. Moreover, a 3.7-5.3-fold increase in the
bacterial yield, depending on the combination of supplements, was seen at a
growth temperature of 39°C-42°C. The largest capsule size was found at 39°C-
40°C. Conclusions: Elevation of temperature above 37°C influenced the pro-
duction of two major Y. pestis antigens F1 and LcrV that can be critical during
the infectious process and/or in eliciting protective immunity against plague.
167 (B)
Infection and Dissemination, Francisella tularensis holarctica
in Mosquito Vector
S. Backman; CBRN Defence and Security, Umea, Sweden.
Background: A number of different insect species are suggested to transmit
Francisella tularensis (F. t.), the etiological agent of the zoonosis tularemia,
to susceptible hosts. Mosquitoes are traditionally considered mechanical
vectors of F. t. holarctica and the main vector for transmission of tularemia to
humans in Sweden. It is however not clear how mosquitoes acquire and trans-
mits the bacterium. The aim of this study was to investigate the hypothesis
that the mosquito already as larvae acquire the bacterium and as adult trans-
mit the disease to susceptible hosts. Methods: In order to confirm presence
of F. t. holarctica in the Swedish mosquito population field sampled mosqui-
toes (sampled in an area with endemic tularemia, Örebro) were pooled (266
pools with 1-50 mosquitoes) and screened by PCR targeting F. t. in general
(lpnA2) and subsp. holarctica in specific (FtM19InDel). Further, a mosquito
model, Aedes aegypti, was established. Mosquitoes were infected in the lar-
val stage and the persistence of the bacterium was monitored through the de-
velopmental stages using PCR. To evaluate transmission, laboratory infected
mosquitoes were allowed to feed on mice. Results: Twenty out of 266 pools
of field sampled mosquitoes were positive for presence of F. t. specific se-
quences and 18 of those contained sequences specific for the clinically rele-
vant subsp. holarctica. Based on our model experiments F. t. holarctica persist
through mosquito development and 25 % of the flying adults (infected as lar-
vae) were PCR positive. None of the animals developed tularemia in the trans-
mission experiment. Conclusion: F. t. holarctica circulates in the mosquito
population in a Swedish area with endemic tularemia. Further, the bacteria
persist through the developmental stages of laboratory infected mosquitoes.
Taken together this supports the hypothesis of an aquatic reservoir for F. t.
holarctica between outbreaks. However, in order to establish vector compe-
tence and transmission rates further investigations are needed.
168 (B)
Phylogeography of Bacillus anthracis Isolates from the Coun-
try of Georgia
E. Khmaladze1, D. Birdsell2, E. Zhghenti1, G. Chanturia1, M. Kekelidze1, M.
Nikolich3, P. Imnadze1, S. Tsanava1, L. Malania1, N. Tsertsvadze1, S. Beck-
strom-Sternberg4, J. Beckstrom-Sternberg4, T. Pearson2, D. Wagner2, P. Keim2;
1
Natl. Ctr. for Disease Control and Publ. Hlth. of Georgia, Tbilisi, Georgia,
2
Northern Arizona Univ., Flagstaff, AZ, 3Walter Read Army Inst. of Res., Fort
Detrick, MD, 4Translational Genomics Res. Inst., Phoenix, AZ.
Background: Bacillus anthracis, the causative agent of anthrax, is a proven bi-
ological weapon. Very little is known about the genetic diversity of B. an-
thracis endemic to Georgia. To address this knowledge gap, we
phylogenetically characterized 92 Georgian B. anthracis isolates based on
Multiple-Locus Variable-Number Tandem Repeat Analysis (MLVA 15) and Sin-
gle Nucleotide Polymorphisms (SNP) to place them into a global phylogeo-
graphic context. Results: We discovered two very distinct and relatively
distant populations of B. anthracis: A.Br.008/009—Transeurasia and
A.Br.013/015 groups—Australian94 across the Georgian landscape. Se-
quenced Georgian B. anthracis isolate belonged to the A.Br.013/015 clade.
62 10th ASM Biodefense and Emerging Diseases Research Meeting
Novel SNPs were discovered and converted into molecular assays which were
used to screen the A.Br.013/015 Georgian isolates as well as other isolates
in our world-wide collection from this group. We identified a new sub-lineage
within the A.Br.013/015 clade that contained most of Georgian isolates and
twelve isolates from Turkey. Conclusion: This finding is consistent with the
idea that this new genetic group may be geographically restricted to border
regions of the European/Asian continents. Within this new sub-lineage, eight
new genetic subgroups were identified. This restricted geographic distribu-
tion, as well as the high levels of genetic diversity within the lineage, is con-
sistent with a relatively older origin and localized differentiation. Within the
small geographic scale of Georgia identification of two distinct and geneti-
cally distant populations of B. anthracis suggests that Georgia is the site of
multiple waves of B. anthracis dispersals from different origins.
169 (B)
Polymorphisms in the lcrv Gene of Yersinia enterocolitica Do
Not Provide for Escape from Plague Protective Immunity
N. C. Miller, L. Quenee, D. Elli, N. Ciletti, O. Schneewind; Univ. of Chicago,
Chicago, IL.
Current efforts to develop plague vaccines focus on LcrV, a polypeptide that
resides at the tip of type III secretion needles. LcrV-specific antibodies block
Y. pestis type III injection of Yop effectors into host immune cells, thereby en-
abling phagocytes to kill the invading pathogen. Earlier work reported that
antibodies against Y. pestis LcrV cannot block type III injection by Yersinia
enterocolitica strains and suggested that lcrV polymorphisms may provide
for escape from LcrV-mediated plague immunity. We show here that poly-
clonal or monoclonal antibodies raised against Y. pestis KIM D27 LcrV
(LcrVD27) bind LcrV from Y. enterocolitica O:9 strain W22703 (LcrVW22703) or
O:8 strain WA-314 (LcrVWA-314), but are otherwise unable to block type III in-
jection by Y. enterocolitica strains. Replacing the lcrV gene on the pCD1 viru-
lence plasmid of Y. pestis KIM D27 with either lcrVW22703 or lcrVWA-314 does
not affect the ability of plague bacteria to secrete proteins via the type III
pathway, to inject Yops into macrophages or to cause lethal plague infections
in mice. LcrVD27-specific antibodies blocked type III injection by Y. pestis ex-
pressing lcrVW22703 or lcrVWA-314 and protected mice against intravenous
lethal plague challenge with these strains. Thus, although antibodies raised
against LcrVD27 are unable to block the type III injection of Y. enterocolitica
strains, expression of lcrVW22703 or lcrVWA-314 in Y. pestis did not provide for
escape from LcrV-mediated plague protective immunity in the intravenous
challenge model.
170 (B)
Multiple-Locus Variable Number Tandem Repeat Analysis of
Brucella melitensis from Azerbaijan Using Eight Markers
X-Z. Huang, E. B. Zelazowska, M. P. Nikolich; Walter Reed Army Inst. of Res.,
Silver Spring, MD.
Background: Brucella melitensis is one of the pathogens that causes brucel-
losis, a common bacterial zoonosis with global spread that is endemic in For-
mer Soviet Union countries including in the South Caucasus and Central Asia.
To better understand the epidemiological and evolutionary relationships of B.
melitensis strains isolated in Azerbaijan, Multiple-Locus Variable number tan-
dem repeat Analysis (MVLA) was conducted using eight selected markers
(MLVA-8). Methods: 25 B. melitensis isolates collected in Azerbaijan and
three reference control strains were examined in this study. Genomic DNAs
were extracted and used as templates for a multiplex PCR to amplify eight
Variable Number Tandem Repeat (VNTR) loci. The sizes of PCR products for
each locus were determined using Beckman Coulter CEQ capillary sequencer.
BioNumerics 6.1 was used for cluster analysis of MVLA patterns. Results: In
most cases, B. melitensis strains from Azerbaijan clustered together upon
based on MLVA-8 when compared with previously determined genotypes

from Georgia and Kazakhstan (human and animal isolates), though some
strains from Azerbaijan shared identical MVLA-8 patterns with strains from
Kazakhstan. Strains from Georgia were genetically distinct from strains of the
other two countries. The Bruce21 locus was the most conserved locus across
the isolates compared, while the Bruce3, Bruce20 and Bruce28 loci showed
higher discriminatory power to differentiate strains from different geographic
locations and sources. Conclusions: This MVLA-8 subtyping approach proved
to be a generally useful tool to distinguish B. melitensis strains from differ-
ent geographic origins toward source attribution. However, with the excep-
tions already evident in this small data set, the possibility of distant
disseminations of B. melitensis subpopluations must be considered. Further
resolution may be obtained by including additional VNTR loci.
171 (B)
Rapid Single Nucleotide Repeat Sequence-Based Detection and
Identification of B. anthracis Using Pyrosequencing Technology
K. R. Hahn, E. Valle, M. Thomas, T. Janzen, M. Shields, N. Goji, K. Amoako;
Canadian Food Inspection Agency, Lethbridge, Canada.
Bacillus anthracis is a biothreat agent with potential use in foodborne bioter-
rorism and early detection and identification is paramount to a successful
mitigation strategy. Even though several technology platforms have been de-
veloped for the detection and strain identification of B. anthracis, these meth-
ods do not fully discriminate between outbreak strains due to the highly
clonal nature of B. anthracis. Single Nucleotide Repeats (SNRs) are stretches
of single nucleotides repeated anywhere from 6 and up to 30 times in the
genome of B. anthracis and can be used as genetic markers to genotype
strains in the case of a disease outbreak. SNRs are susceptible to homoplasy,
and thus are useful for detection and identification of anthrax. Pyrose-
quencing technology is a relatively rapid and inexpensive, easy-to-use
method for sequence based detection. Results are generated in a fraction of
the time it takes to produce standard sequence reads. Here we report a novel
application of pyrosequencing technology for the rapid analysis of SNR re-
gions as a detection and identification tool for B. anthracis. Five SNR targets
specific for B. anthracis with repeats of 7-10 bases were analysed using sev-
eral strains from disease outbreaks. The results indicate that this is a repro-
ducible method for confirmation of SNR number and can potentially be used
to rapidly detect and discriminate between B. anthracis isolates during dis-
ease outbreaks.
172 (B)
Beef and Chicken Contamination with Resistant Escherichial
coli O157 from City Abattoirs in Southwest Nigeria
O. Olatoye, A. E. Amosun; Univ. of Ibadan, Ibadan, Nigeria.
This study investigated the contamination of meat from cattle and chicken
slaughtered for human consumption with E. coli 0157:H7 at the metropolitan
abattoirs and slaughtered slabs of selected poultry farms in Lagos and
Ibadan, Nigeria. The aim was to compare the prevalence and antibiotic sus-
ceptibility patterns across the different locations and climatic seasons. The
organism was isolated by cultural method using selective media and con-
firmed serologically using latex agglutination kits (OxoidR UK). Antibiotic sus-
ceptibility to ten antimicrobial agents was performed by disc diffusion
method using commercial Gram negative discs (AbtekR). Out of 800 meat
samples collected, the overall prevalence of 17.1% (comprising of 19.8% and
14.5% of beef and chicken respectively) was obtained. The prevalence of E.
coli O157:H7 in beef from Ibadan and Lagos were 28.5% and 11.0%, while
those of chicken from Ibadan and Lagos markets were 13.0% and 14.0%, and
from Ibadan and Lagos farms were 18.0% and 13.0% respectively. The preva-
lence of E. coli O157 was significantly higher in beef compared to chicken
(p<0.05), while during wet season, contamination of beef was also higher
than in dry and significantly higher in beef from Ibadan than Lagos abattoir.
T U E S DAY A B S T R AC TS
All the isolates were resistant to one or multiple antibiotics, but the highest
resistance of 91.1% was to tetracycline and nine different resistance patterns
were observed among the isolates. Indiscriminate antibiotics usage in live-
stock predisposes meat consumers to risks of antibiotic resistant Escherichia
coli O157:H7 in southwest Nigeria. Regulatory control of antibiotics usage in
livestock production, meat hygiene and pharmaco-epidemiological surveil-
lance in food animals is hereby recommended to ensure consumer safety.
173 (B)
Contribution of OxyR to Burkholderia mallei Survival and Viru-
lence In Vitro and In Vivo
D. A. Revelli, J. Boylan, F. Gherardini; Rocky Mountain Lab./NIAID/NIH,
Hamilton, MT.
Background: Burkholderia mallei, the causative agent of glanders, is classi-
fied as a Category B Bioterrorism Agent. B. mallei is a facultative intracellu-
lar pathogen that replicates in a number of eukaryotic cell types, including
macrophage. Reactive oxygen and nitrogen species are important for the in-
nate immune response and contribute to the clearance of B. mallei from in-
fected macrophages. However, the regulatory mechanisms for protection
against reactive oxygen and nitrogen species in B. mallei are poorly under-
stood. The gene encoding the oxidative stress regulator, OxyR, is involved in
protecting a number of organisms against oxidative challenge. B. mallei en-
codes an oxyR homologue that may play an important role in protection
against oxidative stress. To assess the role of OxyR in the survival of B. mallei
intracellularly, we constructed an oxyR mutant. Methods: Mutants in B. mallei
ATCC23344 were generated through insertional inactivation by transfer of
the plasmid pZSV. Sensitivity to reactive oxygen species was performed by
disc diffusion assay using 100 μM hydrogen peroxide. Intracellular survival in
murine J774.1 and human macrophage was assessed using a kanamycin pro-
tection assay. For in vivo experiments, 6-8 weeks old, BALB/c female mice
were anaesthetized and inoculated intratracheally. An inoculum of 6x105 cfu
was used to infect groups of 5 mice with wild-type or ΔoxyR B. mallei strains.
PBS was used as a mock infection control. Results: The ΔoxyR mutant was
sensitive to hydrogen peroxide compared to the wild-type B. mallei at a con-
centration of 100 μM. OxyR-deficient B. mallei did not replicate in either
J774.1 or human macrophages. All wild-type infected mice presented with
late stage disease by day 5; in contrast 4 out of 5 mice infected with the
ΔoxyR mutant survived. Conclusion: Inactivation of oxyR increased sensitiv-
ity of B. mallei to hydrogen peroxide in vitro, demonstrating its protective
role against reactive oxygen species. The oxyR mutant was also attenuated
for virulence in macrophages and in mice using an acute model of infection.
Collectively, these data reveal an important role for OxyR in B. mallei patho-
genesis.
174 (B)
Growth Inhibition of Burkholderia pseudomallei by Onion Iso-
lates of Burkholderia cepacia
J. M. Inamine, T. M. Eckstein; Colorado State Univ., Fort Collins, CO.
Burkholderia pseudomallei (BP) is the causative agent of melioidosis, an
emerging infectious disease that is endemic to Southeast Asia and Northern
Australia. It is a major biodefense concern because of its potential use as a
bioterror agent along with its inherent antibiotic resistance. Although the
ecological niche of BP hasn’t been clearly identified, it is usually described as
an environmental organism. Given that Burkholderia species isolated from
soil and plants can produce bacteriocins, bacteriophages and anti-biologics,
we addressed the hypothesis that plant isolates of Burkholderia can inhibit
the growth of BP. Such inhibitors could provide a new approach to the de-
velopment of anti-BP agents. In this study, plant isolates of Burkholderia (pro-
vided by Dr. Jan Leach, Colorado State University) were screened by agar
diffusion for their ability to inhibit the growth of B. pseudomallei strain Bp82
BIODEFENSE PROGRAM & ABSTRACTS 63
T U E S D A Y P O S T E R S E S S I O N | 1:00–3:00 p.m. | Exhibit Hall
(provided by Dr. Herbert Schweizer, Colorado State University). In brief, eight
plant isolates were individually cross-streaked with Bp82 on LB and Nutrient
Agar plates (supplemented with 0.6 mM adenine and thiamine because Bp82
is a delta-pur M mutant), and incubated at room temperature for two days.
None of the four Burkholderia andropogonis or two Burkholderia gladioli
strains showed any inhibitory activity against Bp82. However, the two Burk-
holderia cepacia strains completely inhibited the growth of Bp82 as evi-
denced by clear zones of inhibition. Both of these strains were isolated from
onion, and both produce a bright yellow, diffusible pigment when grown on
LB or Nutrient Agar plates. Work is in progress to see if the pigment is a key
inhibitor, as well as determine the chemical nature of the inhibitor(s). Inves-
tigators have recently shown that human isolates of B. cepacia can produce
bacteriocin-like inhibitors active against other clinical B. cepacia complex
strains, while another group reported that Burkholderia multivorans strain
NKI379 antagonizes the growth of BP in soil; however, to our knowledge this
is the first report of BP inhibition by B. cepacia plant isolates.
175 (D)
Quality Assurance for Select Agents: An Australian Approach
T. Theis1, D. Liu1, J. Gray1, P. Roffey2, W. Rawlinson3,1; 1RCPA BioSecurity QAP,
Surry Hills, Australia, 2Australian Federal Police, Weston, Australia, 3SEALS
Microbiol. Prince of Wales Hosp., Randwick, Australia.
In 2008 the Australian Government established a two tiered list of Security
Sensitive Biological Agents (SSBA). Tier 1 agents include Bacillus anthracis,
Yersinia pestis, botulinum toxin and ricin, and pose the highest security risk
to Australia, while Tier 2 agents such as Clostridium botulinum, Salmonella
typhi, and Francisella tularensis, pose a high security risk. The RCPA BioSe-
curity QAP has been established to provide a quality assurance and profi-
ciency testing program (PTP) for Tier 1 and Tier 2 SSBAs. Participants in this
PTP require a regular supply of testing material from the RCPA BioSecurity
QAP to assess their laboratory competency in detecting SSBAs. The dispatch
of virulent material has significant issues for regulation, potentially for na-
tional security. Therefore, RCPA BioSecurity QAP developed a set of surro-
gate agents that can be shipped and handled safely, while displaying testing
characteristics identical to Tier 1 and Tier 2 SSBAs. The RCPA BioSecurity QAP
has developed proficiency testing surveys for the SSBAs B. anthracis, Y.
pestis, C. botulinum, botulinum toxin and ricin. The non-virulent surrogates
include carefully selected and evaluated bacterial strains that are similar to
B. anthracis, Y. pestis, and C. botulinum. We have established a comprehen-
sive plasmid library which includes more than 20 of the most common gene
targets used in nucleic acid testing of SSBAs. The proficiency testing surveys
for botulinum toxin and ricin were developed using inactivated material, or
non-toxic protein domains. We were able to show that immunological tests
performed by reference laboratories yielded results comparable to those ob-
tained with holotoxins, making them ideal surrogates to be used in a profi-
ciency test. All specimens developed for these proficiency testing surveys
can be handled at Biosafety level 2. This allows for training and education of
laboratory staff with minimal risk of infection. The educational component of
this proficiency testing program is further strengthened by specimen-free on-
line modules.
176 (D)
Broad-Range Microbial DNA Isolation from Clinical Specimens
for Universal PCR Diagnosis
C. Disqué1, M. Linow1, N. Murphy2; 1Molzym GmbH & Co. KG, Bremen, Ger-
many, 2CaerusBio Inc., Downingtown, PA.
Objective: Molecular analysis has proven to be a valuable tool in the timely
diagnosis of the etiological agents of infectious diseases such as sepsis, en-
docarditis and meningitis. DNA extraction of a broad range of microorgan-
isms is a key issue for sensitive detection and precise sequence-based
64 10th ASM Biodefense and Emerging Diseases Research Meeting
identification. The aim was to test the broad-range microbial DNA extraction
capability of the molecular analytic kit, Universal Microbe Detection™, using
a variety of clinical material coming in for routine molecular analysis. Meth-
ods: Specimens originated from patients under suspect of an infection. Sam-
ples were processed as advised (Molzym, Bremen, Germany). Microbial DNA
was detected using universal rRNA gene PCR assays. Amplicons were se-
quenced and BLAST-analyzed to identify the organisms. Results: In total, 430
clinical specimens were analyzed, including blood, synovia, swabs (wounds,
prostheses, pus, bones), CSF, and heart valve/aortal tissues. Among 186 pos-
itive samples, 95 showed typical pathogens (e.g., S. aureus, E. faecalis/fae-
cium, E. coli, C. albicans) and 91 opportunistic strains (e.g., CoNS, viridans
streptococci, Moraxella osloensis). Rare etiologies comprised bacterial
pathogens, among them Bacteroides fragilis, Bartonella quintana, Borrelia
graninii, Coxiella burnetii, Listeria monocytogenes, Providencia stuartii, and
Tropheryma whippleii, and fungal pathogens (non-albicans Candida, black
yeast). One blood sample from a malaria patient was positive in a specific
assay for the etiological agent, Plasmodium falciparum. Conclusion: The pre-
analytical processing comprises the removal of human DNA, concentration
and enzymatic lysis of microbes, and DNA extraction. The data from randomly
collected, diverse clinical material indicated that a broad range of microor-
ganisms was lysed, including Gram-positive and Gram-negative bacteria,
yeasts and other fungi, as well as a protist. The supply of DNA from diverse
microbial taxons to the PCR analysis greatly supports the detection and pre-
cise molecular identification of common and rare etiologies.
177 (D)
WITHDRAWN
178 (D)
Laboratory Surveillance of Tick Born Causing Agents and Other
Relevant Pathogens in the State of Maryland
M. Dembele1, L. Trivedi2, T. Lawson2, S. Montgomery2, M. P. Carlos2,1; 1Johns
Hopkins Sch. of Publ. Hlth., Baltimore, MD, 2Maryland Dept. of Hlth. and Men-
tal Hygiene Lab. Admin., Baltimore, MD.
Background: The purpose of study is to analyze the immunological test re-
sults from suspected Lyme disease (LD), Ehrlichiosis (HME and HGE), Tu-
laremia, Rocky Mountain Spotted Fever (RMSF), Babesiosis, Coxiella burnetii,
and Yersinia pestis specimens submitted to the Maryland Department of
Health and Mental Hygiene (DHMH) Laboratories Administration Division of
Virology and Immunology. This analysis will provide a better understanding
of the occurrence of emerging and re-emerging tick and rodent born diseases
in Maryland. Methods: From 2008 to 2011, de-identified data (N= 3578) were
obtained from the MD DHMH Laboratories Administration Laboratories In-
formation Management System. Tests were performed at Division of Virology
and Immunology, however tests for Babesiosis, C. burnetii and Y. pestis were
sent out to CDC. Immunological methods included: immunofluorescent assay
for LD, HME, HGE, RMSF, Babesiosis and C. burnetii; western blot for LD con-
firmation; agglutination test for Tularemia; and fluorescent antibody test for
Y. pestis. Results: Lyme disease was the most prevalent disease with male
cases accounting for 71.4 % and the 25-59 years old age group for 45%.
RMSF was the second most predominant disease, with male cases and the
25-59 years old age group accounting for the majority of the detected cases
with 58.6 % and 69%, respectively. There were 3 positive cases of C. burnetii
and Babesiosis. Ehrlichiosis was diagnosed in 47.6% of the female popula-
tion, and 38.1% in the 25-59 and > 60 years old age groups. No case of Tu-
laremia was detected. Conclusions: Laboratory diagnosis can alleviate the
burden of an outbreak by the immediate identification of early exposure
cases, and confirmation allows appropriate treatment. Moreover, a compre-
hensive laboratory surveillance of emerging and re-emerging pathogens in
the most susceptible populations is a valuable tool in public health preven-
tion and control.

179 (D)
Development of Single Domain Antibodies for Bacillus an-
thracis Spore Proteins
S. A. Walper1, P. Lee2, J. Liu3, G. Anderson3, E. Goldman3; 1Natl. Academy of
Sci., Washington, DC, 2Nova Res. Inc, Alexandria, VA, 3Naval Res. Lab., Wash-
ington, DC.
Background: The spores of Bacillus anthracis are considered a potential
bioweapon of high concern. Assays are needed to rapidly analyze samples at
the point of concern which is often far removed from a laboratory facility. The
high affinity and stability exhibited by single domain antibodies (sdAb) from
camelids make these elements suitable for integration into assays for use in
such environments. Targeting proteins specific to B. anthracis spores will
allow for the development of highly specific assays capable of rapidly de-
tecting the pathogen. Methods: Two llamas were immunized with six protein
constituents of the B. anthracis spore. A sdAb library was constructed and
enrichment of antigen binders accomplished using phage display. Unique
clones were characterized for binding kinetics to the antigen and ability to
detect spore material from B. anthracis. The melting temperature and ability
to refold following thermal denaturation was assessed using circular dichro-
ism and ELISAs. Results: Representative sdAbs showed nanomolar or better
affinities for target antigens as determined by SPR. SdAb targeting BclA and
gerQ antigens were able to detect spore material from B. anthracis in ELISAs
and did not recognize vegetative cell samples. Those sdAbs targeting BclA
exhibited excellent specificity when tested with spores of other Bacillus
species. SdAb directed towards several other spore proteins however, were
not able to detect spore material, potentially due to a low abundance of the
target antigen. Melting points ranged from 58 - 72 °C, and many of the sdAbs
were successful in antigen detection following iterations of heating and cool-
ing. Conclusions: We successfully isolated sdAbs to six proteins of the B. an-
thracis spore, all with affinities in the nanomolar range. Only sdAbs targeting
the BclA and gerQ antigens were able to directly detect spore material in
ELISAs . These sdAbs may potentially serve in combination with other de-
tection elements for the development of fieldable assays for the detection
and differentiation of B. anthracis spores.
180 (D)
Zeptomole/ML Quantification of Anthrax Edema Factor by LC-
ESI-MS/MS: Activity Enhancement by Monoclonal Antibodies
R. C. Lins1, Z. Kuklenyik2, C. P. Quinn2, K. Bedi2, J. Goldstein2, A. Lyons2, M. Gal-
legos2, U. Ansari3, R. Pitts3, D. Bagarozzi3, K. Isbell1, C. E. Leysath4, Z. Chen4,
S. H. Leppla4, J. R. Barr2, A. Boyer2; 1Battelle CDC, Atlanta, GA, 2Ctr. for Disease
Control and Prevention, Atlanta, GA, 3Ctr. for Disease Control and Prevention
Battelle, Atlanta, GA, 4NIH, Bethesda, MD.
Anthrax is caused by the pathogenic bacterium Bacillus anthracis. It secretes
two major exoproteins: lethal factor and edema factor (EF) that combined
with protective antigen form lethal toxin and edema toxin respectively. EF is
a highly active calmodulin-dependent adenylyl cyclase (AC) which converts
ATP to cyclic AMP (cAMP) causing edema and immune dysfunction in anthrax.
We have developed a high sensitivity AC activity-based method for quantify-
ing EF. Analytic sensitivity is in part dependent on prior purification of EF with
monoclonal antibodies (mAbs). Here we report screening of 12 murine anti-
EF mAbs for their ability to bind and modulate EF. The mAbs were tested for
1) reactivity with EF by western blot (WB), 2) interference/enhancement of EF
catalytic activity, 3) immunoprecipitation (IP) of EF and 4) combined EF IP/ac-
tivity. For EF-MS mAbs were immobilized on magnetic protein-G or tosyl-ac-
tivated beads to purify and concentrate bound EF which was then reacted
with ATP/calcium/calmodulin to generate cAMP, subsequently quantified by
LC-ESI-MS/MS. All 12 mAbs were reactive for EF in WB. Three mAbs enhanced
and two highly neutralized EF activity. Combined capture/activity was almost
doubled for two mAbs. In summary, two of the anti-EF mAbs identified were
T U E S DAY A B S T R AC TS
good candidates for improved EF detection by MS. Performance of the EF
method for these new mAb’s were compared to anti-EF mAb’s described pre-
viously. The EF method is specific, fast (≤4h) and sensitive with a lower limit
of detection of 16 fg/mL (180 zmoles/mL) for 200 µl sample. The EF-MS
method is the most sensitive reported for quantitative detection of any an-
thrax toxin. Combined with our MALDI-TOF-MS method for LF, the EF method
expands our capability to detect and measure both toxins in early asympto-
matic infection and throughout treatment.
181 (E)
Simultaneous Detection of Ricin and Abrin DNA by Real-Time PCR
I. Mossbrugger, E. Felder, R. Wölfel; Bundeswehr Inst. of Microbiol., Munich,
Germany.
Ricin is one of the most potent plant toxins known, and the castor plant from
which it is derived, Ricinus. The harvesting of castor beans exceeds one mil-
lion tons annually, and the toxin may be easily obtained in high yield from
the castor seeds using rather simple technology. As a result, ricin is a potent
and available toxin for criminal or terrorist acts. Abrin is a small toxic protein
similar to ricin, found in the beans of the Jequirity plant (Abrus precatorius),
and shares structural and functional principles similar to those of ricin. As
the production of highly purified ricin or abrin toxin is labour intensive and re-
quires sophisticated lab equipment, it is more likely that crude, rather than
purified, extracts would be used by non-governmental perpetrators. In such
crude extracts remaining nucleic acids provide appropriate surrogate ana-
lytes for the toxins themselves and allow the PCR detection and identifica-
tion of abrin or ricin. As part of an effort to provide protocols for the detection
of various biological toxins we developed a real-time PCR assay for simulta-
neous detection of ricin and abrin nucleic acid. Oligonucleotide primers were
designed manually based on an alignment of R. communis, A. precatorius
and other lectine-producing plant genome information. Two primer pairs were
identified to be efficient for amplification of lectine-specific target sequences.
The differentiation between both ricin and abrin was enabled by toxin-spe-
cific, dye-labelled 5’exonuclase hybridizations probes. The resulting assay
reliably amplified the targeted region of ricin and abrin DNA. To determine
the analytical sensitivity of the assay, cloned and quantified fragments of ricin
and abrin DNA were amplified in limiting log10 dilution series. The 95% de-
tection limit of the real-time PCR was 3.0 genome copies per reaction for the
detection of ricin and 1.2 genome copies per reaction for the detection of
abrin. In summary, the novel ricin and abrin DNA real-time PCR assay pro-
vides a rapid and sensitive method for the detection and identification of ricin
and abrin nucleic acid. It may prove useful for the investigation of deliberate
castor or Jequirity bean contaminations.
182 (E)
Low-Cost, Highly Efficient Airborne Pathogen Collection and
Detection System for Field Use
J. Gordon1,2, P. Gandhi1; 1Inspirotec LLC, Chicago, IL, 2Northwestern Univ.,
Evanston, IL.
Efficient and cost effective collection of airborne pathogens, and subsequent
detection, is a critical need for biodefense. We describe use of an ionic
propulsion device as a platform for air sampling. A plasma is generated at a
high voltage wire. The ions generated are propelled by a geometric arrange-
ment of electrodes, creating an airflow. Particulates acquire an induced
charge and are deposited on plate electrodes. From electrical field theory,
non-conductive elements will not affect the electrostatic field. We evaluated
the use of non-conductive textile envelopes on the electrodes, for collection
of samples for analysis. We used cat allergen Fel d1 and ELISA assay as a
model system. Possible effects of the textile on the air-flow pattern and ve-
locity were investigated with a smoke gun and an anemometer. Captured al-
lergen was measured with an ELISA. The recovery of allergen extracted from
BIODEFENSE PROGRAM & ABSTRACTS 65
T U E S D A Y P O S T E R S E S S I O N | 1:00–3:00 p.m. | Exhibit Hall
textile was as efficient as with a published swabbing procedure. Of materials
evaluated, silk gave the most efficient capture of the allergen. Diluent from
the ELISA was effective as an extraction medium. Extractables from the silk
had no effect on the ELISA performance. The detection limit was less than 5
pg/M3. We are further developing the system to include a specialized capture
electrode, which will functional as a potential well and trap the sample on a
small area. The small area will permit collection of the sample in a small vol-
ume with consequent increase in sensitivity. The sample collection means
may be optimized for delivering analyte to any relevant rapid test. The tech-
nology will be developed into a cost-effective, handheld device for use in the
field by first responders and military personnel.
183 (E)
Evaluation of Automated and Manual Commercial Nucleic Acid
Extraction Methods for Detection of Ricinus communis DNA by
Real-Time PCR
A. S. Hutchins1, M. Astwood2, J. R. Saah1, L. A. Dauphin2; 1North Carolina State
Lab. of Publ. Hlth., Raleigh, NC, 2CDC, Atlanta, GA.
Background: The castor bean plant, Ricinus communis, produces ricin toxin,
which has been used as a biological weapon in the past. Ricin is classified by
the CDC as a Category B critical biological agent. Currently, diagnostic labo-
ratories use real-time PCR assays for rapid detection of R. communis DNA in
environmental samples, which is an indicator that ricin toxin could be pres-
ent. DNA extraction methods can impact the sensitivity of real-time PCR as-
says; therefore, determining the optimal extraction method is important for
effective detection of R. communis DNA. Methods: This study evaluated com-
mercial nucleic acid extraction methods for their efficiency in isolating R. com-
munis DNA from castor bean preparations. Four extraction methods,
including two automated methods, the MagNA Pure Compact and MagNA
Pure LC, and two manual methods, the QIAamp DNA Blood Mini Kit and the
Masterpure Complete DNA and RNA Purification Kit, were compared. Castor
beans were homogenized in phosphate buffered saline, and the extraction
methods were performed in triplicate upon dilutions of the castor bean ho-
mogenates. The effectiveness of nucleic acid extraction with the four meth-
ods was determined by comparisons of cycle-threshold (CT) values with a R.
communis-specific TaqMan® real-time PCR assay with an internal positive
control (IPC), as well as comparisons of DNA yields and DNA purity. Re-
sults/Conclusion: Although the four extraction methods yielded DNA that re-
sulted in differences in real-time PCR CT values; the limit of detection was
equivalent for all extracts. There was no evidence of PCR inhibition for any of
the extraction methods as measured by the IPC assay. The results from this
study indicate that diagnostic laboratories have the option to select from au-
tomated or manual extraction methods without loss of sensitivity when using
real-time PCR for detection of R. communis DNA.
66 10th ASM Biodefense and Emerging Diseases Research Meeting
184 (E)
Development of Multiplex Rapid Tests Allowing Warfare Agents
Detection
T. Berthelot, J. Credou, H. Volland; CEA, Gif sur Yvette, France.
Background: Biological warfare agents are inconspicuous, their detection is
difficult and, within the limit of our current knowledge, far from being in-
stantaneous. This point is even more crucial because the effects of these
agents, dramatically effective, are insidious. The development of powerful
systems of detection and allowing identification of these agents, as early as
their dispersion, is therefore crucial to be efficient against those. These sys-
tems will have to be easy to use, because they are susceptible to be used by
non specialists. Methods: Among the various methods of detection devel-
oped, immunochromatography, while preserving the property of specificity of
classical immunological assays, presents several advantages: fast, ease of
use, adapted to measurements on the field and very low cost. Unfortunately
classical lateral flow immunoassays could not detect several agents at the
same time in the same sample. Based on the Whitesides research group work
we have developed immunochromatographic tests which could allow the de-
tection and identification of up to 16 different agents simultaneously in the
same sample. This project implies the development of chemistry which en-
ables both the binding molecule on paper and the conservation of their bio-
logical activity. We also compare different kind of colorimetric labels linked to
monoclonal antibodies. Results: The developed fluidic system constituted
by paper allows the fraction of one sample in 16 isolate spots. We are also
able to bind one different biological molecule, like monoclonal antibody, or
chemical component to each spot. Moreover specific signals visually detected
were associated to the presence of molecules in the sample. Conclusions:
Our device is suitable for a field use and authorizes a multi detection/iden-
tification of up to 16 agents. Currently we work on different parameters (flow
rate, label, monoclonal antibody density on the spot) in order to increase the
limit of detection.
185 (E)
Validation of Unique Signature Discovery in Select Agents and
an Emerging Pathogen
B. J. Haley1, E. Taviani1, A. Chen1, A. Phillippy2, A. Huq1,3, R. R. Colwell1,4,5, I.
Knight3,6; 1Maryland Pathogen Res. Inst., College Park, MD, 2Ctr. for Bioin-
formatics and Computational Biology, College Park, MD, 3Maryland Inst. for
Applied Environmental Hlth., College Park, MD, 4Univ. of Maryland Inst. for
Advanced Computer Studies, College Park, MD, 5CosmosID, College Park,
MD, 6Canon U.S. Life Sci., Inc., Rockville, MD.
The Insignia informatics pipeline (http://insignia.cbcb.umd.edu/), which rap-
idly detects unique DNA sequences of bacteria, viruses, and bacteriophage
against a background of RefSeq sequences, was used to determine unique
genomic signatures of three select bacterial agents (Yersinia pestis, Brucella
abortus, and Clostridium botulinum) and one emerging bacterial pathogen
(Campylobacter jejuni). These species-specific signatures were validated by
TaqMan Real-Time PCR assays. For Y. pestis, C. botulinum, and C. jejuni; 43,
54, and 82 unique sequences of at least 100 nucleotides in length were dis-
covered. B. abortus signatures were too short for primer/probe generation,
so at most 100 bp of non-unique sequence was permitted between the
shorter signature sequences, from which the primers and probes were se-
lected. When validated against a DNA from Y. pestis and its near-neighbors,
42 primer and probe sets were discriminatory between Y. pestis and near-
neighbors, while 1 set showed late amplification. All primer and probe sets
were unique for Clostridium dificile when compared with the near neighbors
C. glycolicum, C. thermocellum, and C. proteoclastium. All primer and probe
sets generated amplification for all Campylobacter jejuni strains except for 2
which did not show amplification for a strain whose genome had yet to be
sequenced. Eleven other primer and probe sets did amplify regions of four

near neighbors whose genomes were not yet deposited in RefSeq. Seven
primer and probe sets from non-overlapping regions of one gene sequence
were able to discriminate between B. abortus and the near neighbors B. suis
and B. melitensis. Results of this study validate the ability of the Insignia in-
formatics pipeline to generate a suite of unique genomic DNA targets for the
detection of bacteria, including select agents.
186 (E)
Novel Surface Chemistry for the Development of Multiplex
Immunochromatographic Tests
J. Credou, H. Volland, T. Berthelot; CEA, Gif sur Yvette, France.
Background: The development of new chromatographic format as lab-on-
paper by the Withesides group [1] involves the replacement of conventional
material (nitrocellulose) used for lateral flow immunoassays by paper. This
technological breakthrough imposes to develop novel surface chemistry to
functionalize paper with biomolecules without altering their biological activ-
ity. Methods: Our project implies the development of chemistry which en-
ables both the biomolecules grafting on paper and the conservation of their
biological activity. Moreover, our process must be operated in soft conditions
(aqueous systems, compounds and methods compatible with biological ma-
terial) but also must be compatible with paper sheet format. In fact, it is well
known that cellulose is not water-soluble which implies that its chemical
modification is performed under harsh conditions in organic solvents. To over-
come these drawbacks, we chose two synthetic pathways to functionalize
paper sheets based (i) on an innovative surface chemistry, named GraftFast™
[2, 3], and (ii) on a photochemical process. Results: By our approaches, paper
sheets have been successfully modified and bear different functionalizable
groups, as carboxyl, amino or thiol groups for example, to graft biomolecules
by classical bioconjugate techniques. These paper modifications were per-
formed in aqueous environment and at room temperature. Conclusions: The
chemical ways developed here are suitable for new immunochromatographic
format as lab-on-paper. Currently, our work is focused on different parame-
ters (chemical groups, linker density onto the surface…) which can play a role
in the detection limit. [1]. A.W.Martinez, et al. Analytical Chemistry, 2010, 82
[2]. Mévellec, V. et al. Chem. Mater. 2007, 19, 6323-6330. [3]. Mevellec, V. et
al. Patent n° WO2008/078052; 2006.
187 (F)
Virus Pathogen Resource (ViPR): An Open Bioinformatics
Database and Analysis Resource for Human Virus Pathogen
Research
Y. Zhang1, B. Pickett1, E. Sadat1, J. Noronha1, R. B. Squires1, V. Hunt1, M. Liu2,
S. Kumar3, S. Zaremba3, Z. Gu3, L. Zhou3, C. Larson4, J. Dietrich3, E. Klem3, R.
H. Scheuermann1; 1Univ. of Texas Southwestern Med. Ctr., Dallas, TX, 2South-
ern Methodist Univ., Dallas, TX, 3Northrop Grumman Hlth. Solutions,
Rockville, MD, 4Vecna Technologies, Greenbelt, MD.
The Virus Pathogen Resource (ViPR, www.viprbrc.org) is a freely available,
NIAID-funded resource providing integrated datasets and analysis tools for
fourteen different virus families: Arenaviridae, Bunyaviridae, Caliciviridae,
Coronaviridae, Flaviviridae, Filoviridae, Hepeviridae, Herpesviridae, Paramyx-
oviridae, Picornaviridae, Poxviridae, Reoviridae, Rhabdoviridae and Togaviri-
dae. ViPR includes information from: GenBank sequence records, gene
annotations, strain metadata, Gene Ontology classifications, UniProtKB pro-
tein annotations, relevant 3D protein structures from the Protein Databank
(PDB), immune epitopes from the Immune Epitope Database (IEDB), clinical
and surveillance metadata and novel data derived from comparative ge-
nomics analysis. ViPR data can be searched and analyzed through user-
friendly web interfaces for calculating: BLAST, multiple sequence alignments,
phylogenetic trees, single nucleotide polymorphisms, sequence feature vari-
ations, and metadata-driven comparative analyses. Tools also exist to: view
T U E S DAY A B S T R AC TS
phylogenetic trees, assign annotations to new genomes, and store results
from any search or analysis within a personal Workbench space on the ViPR
server for future use. We expect our current (and future) suite of tools to fur-
ther assist viral researchers in developing diagnostics, prophylactics, and/or
therapeutics for viruses that are pathogenic to humans.
188 (F)
Effects on Host Responses of Air-Borne and Aerosolized Rab-
bitpox Infections in New Zealand Rabbits
L. DaSilva1, A. Porter1, A. Nalca1, R. Settlage2, C. L. Galindo3, L. McIver2, K.
McMahon2; 1USAMRIID, Frederick, MD, 2Virginia Bioinformatics Inst. at Vir-
ginia Polytechnic Inst., Blacksburg, VA, 3Eagle Eye Data Analysis, Houston,
TX.
Rabbits are commonly used research animals. The understanding of the ge-
netic structure of the rabbit genome represents therefore an important step
towards the full characterization of the rabbit animal model in biodefense re-
search. We have developed a custom microarray covering 35,058 probe set
identifiers representing the leporine genome to determine the effects of an
aerosol exposure to two doses of the rabbitpox virus in the host’s lungs. Se-
quence information was obtained from the 2X coverage of the Oryctolagus cu-
niculus genome provided by the Broad Institute. Microarrays were performed
using RNA isolated from lungs of rabbits challenged with aerosolized rabbit-
pox virus (RPXV) at a low dose (165 PFU) (n=10) or at a high dose (4,171 PFU)
(n=10), and compared to environmental control rabbits (n=4), yielding sta-
tistically significant (p<0.05) changes in gene expression in the low- and
high-dose exposure groups. Hierarchical clustering and phylogenic analyses
of the rabbit genome array data confirmed the existence of at least three dif-
ferent groups: non-responders, mild responders, and high responders among
the significant transcripts. The high responders group consisted mostly of
the animals receiving the lower dose of the aerosolized RPXV. Host responses
in the low- and high-RPXV-dose groups revealed the involvement of genes of
the cell cycle, growth and development and carbohydrate metabolism. In ad-
dition, rabbitpox produced a transcriptional response resembling humoral
responses and more so than the classical antiviral responses. Interestingly,
the data have also suggested an effect on RNA post-transcriptional modifi-
cation genes to account for viral survival. Potential biomarkers of rabbitpox
infection were also identified, including transcripts for FLRT2, TPM3, CD14
and HP. The present work should contribute to a more comprehensive eval-
uation of the rabbit when used as an animal model of orthopox viral disease
and of other human infectious diseases as well.
189 (F)
Analysis of Next-Generation Sequencing for Detecting Rare
Variants in Bacillus anthracis
R. E. Colman1, J. Schupp1, J. Gillece1, A. Rawat1, D. M. Engelthaler1, J. Foster2,
P. Keim2,1; 1Translational Genomics Res. Inst., Flagstaff, AZ, 2Northern Arizona
Univ., Flagstaff, AZ.
Next-generation sequencing is rapidly advancing capabilities for bioforen-
sics. Bacterial populations, whether grown in the lab or found in the envi-
ronment, will represent a mixture of genotypes even if the population arose
from a single cell. Detection and use of rare variants as a forensic diagnostic
tool for identifying particular preparations is an important capability, e.g. the
Amerithrax investigation, and a potentially powerful application of next-gen-
eration sequencing. We have developed and synthesized plasmid constructs
with engineered SNP and INDEL mutations for use in characterizing se-
quencing error and as an internal sequencing control (ISC) in forensic analy-
sis. We Illumina sequenced mixtures of these different plasmid constructs to
extremely high coverage (~100,000X), as well as DNA from the well charac-
terized Ames and Sterne strains of Bacillus anthracis (~1,000X). We then cal-
culated average baseline error rates of 0.41% and 0.55% for plasmids and B.
BIODEFENSE PROGRAM & ABSTRACTS 67
T U E S D A Y P O S T E R S E S S I O N | 1:00–3:00 p.m. | Exhibit Hall
anthracis, respectively. Using the plasmid standards, we characterized three
factors that contribute to error: library preparation variation, multiple PCR
steps on a single preparation variation, and lane to lane variation. We found
very little variation in the average baseline error rate ranging from 0.36-
0.44%. We determined that the reference base differentially contributes with
C to A, A to C, G to T, and T to G errors being 2-3 times more likely to occur
than other errors. In our sequence analysis of the Ames strain, the distribu-
tion of error is not Poisson distributed, indicating that error is non-random.
The majority of positions have a low error rate, 87% of positions with error
have an error of 0.5% or less; however there are notable error hotspots. We
have also sequenced a known artificial mixture of B. anthracis with our syn-
thesized plasmid spiked in to serve as ISC. This enabled us to characterize
within-lane error rate and true variant limits of detection, increasing the con-
fidence in minor variant detection.
190 (F)
Nested Amplification for Improved Genotyping of Coxiella
burnetii
R. Vipond, A. Pepler, G. Vincent, J. Latham; Hlth. Protection Agency, Salis-
bury, United Kingdom.
Coxiella burnetii is one of the few biothreat agents naturally present in the UK
and has been responsible for large outbreaks of disease as well as sporadic
cases. Diagnosis is often by serological screening for an immune response to
the bacterium, though direct detection by PCR early during infection is also
possible. Because this is a zoonotic infection, linking human cases to out-
breaks in animals or reservoirs of infection is important for disease manage-
ment and risk analysis. The highly infectious nature of the pathogen and
intracellular lifestyle mean that genotypic analysis, in our hands at least, is
often directly on extracted clinical or veterinary samples rather than cultured
material. Using published Multi-spacer typing (MST) and Multiple Locus Vari-
able Number Tandem Repeat Analysis (MLVA) methods we found direct analy-
sis frequently failed even with samples which were known to have pathogen
nucleic acid present by real time Polymerase Chain Reaction (PCR). Thus we
designed sets of flanking primers for nested PCR reactions and adapted the
method to improve the sensitivity of the assay and permit genotyping of ex-
tracted samples without enrichment or culture of PCR-positive samples. The
nested MST and MVLA approach was applied to a range of clinical, veterinary
and environmental samples. Results showed a significant increase in the
number of samples for which complete genotype profiles were obtained (from
50% up to 85%, n= 22 samples) using the nested methods compared to the
original direct PCR approach. The limit of analysis, not surprisingly, correlated
to the abundance of template as determined by real time PCR, whilst there
was variance between loci, a 15-100 fold increase in the lowest level of tem-
plate which yielded a result was observed. In summary, we have shown a
nested approach improves the sensitivity of existing typing methods for C.
burnetii which, given the difficulty of working with this particular organism,
is of great utility in genotypic analysis of a variety of sample types which may
be encountered during outbreak investigations or routine surveillance.
191 (F)
Utility and Challenges Associated with Use of Next Generation
Sequencing in Ultra-Rare Variant Detection
H. Koshinsky1, V. Fofanov2, J. Liu2; 1Eureka Genomics, Hercules, CA, 2Eureka
Genomics, Houston, TX.
Presence of rare variants in bacterial samples could act as a sample’s fin-
gerprint and be important in investigation and prosecution of bioterrorism
attacks or attempts. Existing PCR-based approaches cannot detect unknown
variants if present in less than 10% of the sample. The recent advances in
Next Generation Sequencing technologies may have sufficient throughput to
make feasible the cost-acceptable generation of the amount of sequence
68 10th ASM Biodefense and Emerging Diseases Research Meeting
data required for the detection SNPs that are present in 0.1% of a sample
(ultra-rare variants). We have conducted computational simulations and deep
sequencing experiments on Illumina’s GAIIx using Enterobacteria phage
phiX174 (NC_001422), Pseudomonas aeruginosa (NC_008463), and PCR am-
plified regions from Mycobacterium tuberculosis (mixed at 1:1,000 mutant
to wildtype ratios). Results suggest that Next Generation Sequencing ap-
proaches can, in a cost acceptable manner, produce sufficient sequence data
to successfully detect ultra-rare variants. However, distinguishing true ultra-
rare variants from false positive SNPs is a major limiting factor when the vari-
ant SNP is present at less than 1%. We have performed a detailed analysis of
factors contributing to false positive SNP identification. Both systematic and
random sources of errors have been identified. The systematic sources of
error include: machine sequencing error, alignment algorithm errors, and bias
induced by features of the reference sequence. Random sources of errors
identified as related to multiple hypotheses testing during SNP detection.
Corrections for systematic and random errors have been designed and found
to increase the specificity (reduce the number of false positive SNPs identi-
fied) of ultra-rare variant detection.
192 (G)
PANACEA Broad-Spectrum Antiviral Therapeutics
T. H. Rider; Massachusetts Inst. of Technology, Cambridge, MA.
Background: Although there is great concern over emerging and category A-
C priority viruses, there are relatively few prophylactics or therapeutics for
these viruses, and most which do exist are highly pathogen-specific or have
undesirable side effects or other disadvantages. We have developed a radi-
cally new and very broad-spectrum antiviral therapeutic/prophylactic that
has the potential to revolutionize the treatment of viral infections, including
those due to emerging, category A-C, and common clinical pathogens. Our
Double-stranded RNA (dsRNA) Activated Caspase Oligomerizer (DRACO) ap-
proach selectively induces apoptosis in cells containing any viral dsRNA, rap-
idly killing infected cells without harming uninfected cells. Methods: We have
produced preliminary DRACO designs and evaluated their toxicity, antiviral
efficacy, and other properties in quantitative cell assays and in mouse trials.
Results: We have demonstrated that DRACOs are nontoxic in all 11 cell types
tested thus far, including human, monkey, and mouse cells representing nu-
merous tissue types. Using these cells, we have shown that DRACOS are ef-
fective against all 15 viruses tested thus far, including DNA and RNA viruses,
enveloped and nonenveloped viruses, human and rodent viruses, and viruses
that use a variety of receptors. Among the viruses against which DRACOs
have proven effective in vitro are dengue hemorrhagic fever virus, multiple
arenaviruses, and multiple bunyaviruses. In mice, we have demonstrated that
DRACOs rapidly penetrate into all organs tested, persist for over 24 hours
after each dose, and are nontoxic. We have shown that DRACOs rescue mice
from lethal H1N1 influenza and Amapari arenavirus challenges, and we are
conducting efficacy trials against dengue in mice. Conclusions: We have
demonstrated that DRACOs are nontoxic and effective against a broad spec-
trum of viruses. We hope to optimize our DRACO designs and demonstrate
them against additional viruses and in additional animal models. This work
should greatly advance DRACOs toward ultimate utility as safe, broad-spec-
trum therapeutics/prophylactics for priority and emerging viral pathogens,
filling a large gap in existing therapeutics.

193 (G)
Efficacy of a Doxycycline Treatment Regimen in Rhesus
Macaques Aerosol Challenged with Brucella melitensis
L. Henning, E. McGuinness, D. Pak, S. Miller, C. Briscoe, M. Anderson, R. War-
ren; Battelle, Columbus, OH.
Background: The non-human primate inhalational brucellosis model was pre-
viously characterized at Battelle. This large animal model was used to assess
the efficacy of a doxycycline treatment regimen when administered to rhe-
sus macaques (RMs) therapeutically after aerosol challenge with B. meliten-
sis. Methods: Fourteen RMs were aerosol challenged with a targeted dose
of 2.50E+05 cfu B. melitensis. Beginning 21 days post-challenge, doxycycline
was administered orally to 10 RMs. Treatment was continued for 14 consec-
utive days (BID). Body temperature, blood culture, splenomegaly and tissue
burden were assessed. In addition, biomarkers were evaluated by the mi-
croagglutination assay and ELISAs (IgG, IgM, and neopterin) throughout the
in-life period. Results: Aerosol challenged RMs developed fevers and sero-
converted (≥ 1:80) by Day 21 post-challenge. Ninety-three percent (13/14) of
challenged animals were bacteremic by enriched blood culture on Day 21.
On Day 28, while 9/10 treated animals were negative, all untreated animals
remained bacteremic. While treated animals had either no tissue burden
(6/10) or below the limit of quantitation (4/10) by Day 37 post-challenge, all
untreated animals had a tissue burden in the lung, liver, spleen and/or kid-
ney. In addition, splenomegaly resolved for most treated animals one to two
weeks after treatment was initiated, while it was still visualized by ultrasound
in untreated animals. Conclusion: The oral doxycycline treatment regimen
appears to be effective against B. melitensis infection in RMs. The results
from this study support the use of the inhalational brucellosis model for test-
ing therapeutics. This work was supported by the Transformational Medical
Technologies program through CBRNIAC contract number SP0700-00-D-
3180, Task CB-09-0018 Task 769 from the Department of Defense Chemical
and Biological Defense program through the Defense Threat Reduction
Agency (DTRA).
194 (G)
Host-Directed Antimicrobial Therapeutics for Intracellular
Bacterial Pathogens
A. Levi1, D. Arvans2, P. Robins2, L. P. Potluri1, B. Heaton1,2, S. Riley1,2, S.
Crosson1,2, J. Martinez1,2, J. E. Gabay1, H. A. Shuman2,1; 1Univ. of Chicago,
Chicago, IL, 2Howard Taylor Ricketts Lab., Lemont, IL.
Although antibiotic therapy is often successful, it is becoming increasingly
clear that alternatives to conventional pathogen-directed antibiotic therapy
must be developed in the face of increased antibiotic resistance. We aim to
identify human genes that encode functions required for successful intracel-
lular infection by a variety of bacterial pathogens. We hypothesize that novel
host-directed antimicrobial therapies can be developed by targeting specific
host pathways used by individual or groups of pathogens. We have identified
shRNA libraries as the method with the highest potential to discover human
genes required for a successful infection. As a proof of principle experiment,
we have prepared 18 cell lines containing candidate shRNA constructs. The
candidate genes were chosen based on preliminary data and literature re-
ports that certain receptors or regulators were important for successful in-
fection by specific pathogens. Additionally, we have also transferred an
shRNA library that consists of several shRNA constructs per human gene and
covers the entire human genome, to HEK293T cells in which several differ-
ent intracellular pathogens are able to grow. We have begun to test the abil-
ity of phase II Coxiella burnetii and Rickettsia conorii to replicate in the cell
lines in which the candidate genes have been knocked down. In addition, we
have constructed a Brucella abortus strain that expresses mCherry, a red flu-
orescent protein that facilitates the measurement of intracellular growth. In
order to discover additional potential host targets we screened a collection
T U E S DAY A B S T R AC TS
of annotated small molecule inhibitors for their ability to block intracellular
growth of C. burnetii, B. abortus and R. conorii. Using this inhibitor screening
assay, we have identified host functions that likely play a role in regulating in-
tracellular growth of C. burnetii and rickettsial species. Experiments are un-
derway to elucidate the molecular mechanisms underlying this phenotype.
195 (G)
Restoring Antibiotic Sensitivity with Antisense Inhibitors of
Gene Expression
P. L. Iversen1,2, D. V. Mourich1,2, F. Schnell1,2, B. Geller1,2; 1AVI BioPharma, Inc.,
Bothell, WA, 2AVI BioPharma, Bothell, WA.
The objective of the current study was to demonstrate a reduction in the min-
imum inhibitory concentration (MIC) of aminoglycosides, beta-lactams, chlo-
ramphenicol, tetracycline and sulfonamides by inhibiting bacterial genes
linked to their drug resistance phenotype. Phosphorodiamidate morpholino
oligomers (PMOs) are short synthetic DNA mimics that inhibit gene expres-
sion by binding specifically to mRNA with complementary base sequences.
Previously, we synthesized peptide conjugated PMOs (PPMO) targeted to es-
sential genes of bacteria, and found that these PPMOs can be potent bacte-
ricidal compounds. Escherichia coli SMS-3-5 represents an excellent
organism to evaluate PPMO inhibitors of gene expression linked to antibac-
terial resistance because it is resistant to a record number of antimicrobial
compounds from all major classes. We evaluated 11 different PPMOs in liq-
uid cultures with a range of dilutions of the selected antibiotics to establish
the minimum inhibitory concentration (MIC indicated as µg/mL) over a range
of concentrations of PPMO from 0.5 up to 32 uM. A scrambled PPMO se-
quence had no influence on the MIC value for at least three different antibi-
otics. Antibiotic sensitivity was nearly completely restored for streptomycin
by PPMOs targeting strA and strB. Less effective restoration of sensitivity was
observed for streptomycin with PPMOs targeting aadA and aph genes. These
observations further indicate the PPMO alone is not an effective inhibitor of
bacterial growth. The observations may suggest that both aadA and aph con-
tribute less to streptomycin resistance than strA or strB, or that these two
target genes may not have been effectively inhibited. indicating a need for
continued effort to identify improved targets. The restoration of sensitivity
to ampicillin and chloramphenicol was also substantial while restoration of
sensitivity to tetracycline and trimethoprim was minimal. This results show
that sensitivity to antibiotics can be modulated and restored by PPMOs, and
that variations in PPMO design can greatly affect the results.
196 (G)
Post-Exposure Administration of a Monoclonal Antibody Cock-
tail Is Protective in Orthopoxvirus-Infected Mice and Cynomol-
gus Macaques
J. L. Nordstrom1, G. J. Rainey1, H. Li1, P. Silvera2, F. Koide2, M. Buller3, J.
Schriewer3, A. Dekonenko4, K. Shah1, L. Huang1, V. Ciccarone1, S. Burke1, M.
Bowe1, D. Farb1, M. Kadan1, E. Bonvini1, K. Stein1, S. Koenig1, S. Johnson1;
1
MacroGenics, Rockville, MD, 2Southern Res. Inst., Frederick, MD, 3Saint
Louis Univ., St Louis, MO, 4Univ. of New Mexico, Albuquerque, NM.
An orthopoxvirus-neutralizing immunotherapeutic with post-exposure effi-
cacy would be valuable in case of a bioterrorism event. One candidate is a
monoclonal antibody (mAb) cocktail consisting of h7D11, a humanized ver-
sion of 7D11 (Hooper Virol 266:329) which targets L1 protein of intracellular
mature virions, and ch12F, a human/chimpanzee chimera of 12F (Chen J Virol
81:8989) which targets A33 protein of extracellular enveloped virions. Re-
search lots and a GMP lot of the mAbs were produced; the GMP product was
formulated as a 1:1 cocktail and has shown good stability. Individual mAbs
and the mixture were administered as single i.p. injections at doses ranging
from 10 to 100 mg/kg to vaccinia-infected Balb/c mice (5e5 PFU/intratra-
cheal) at 3 days post infection (dpi), ectromelia-infected A/NCR mice (15
BIODEFENSE PROGRAM & ABSTRACTS 69
T U E S D A Y P O S T E R S E S S I O N | 1:00–3:00 p.m. | Exhibit Hall
PFU/intranasal) at 5 dpi, or monkeypox (MPXV)-infected CASTEiJ mice (4400
PFU/intranasal) at 3 dpi. The cocktail significantly improved survival and/or
reduced disease compared to vehicle-treated animals, with h7D11 as the key
contributor in the vaccinia model and ch12F in the ectromelia model. A phar-
macokinetic study of the cocktail in naïve cynomolgus monkeys showed ter-
minal t values of ~9 and 13 days for h7D11 and ch12F, respectively.
Cynomolgus macaques challenged i.v. with 5e7 PFU of MPXV and treated i.v.
at 1 dpi with 100 mg/kg mAb cocktail, 1000 mg/kg vaccinia immune globu-
lin (VIG) or vehicle demonstrated 100, 67 and 0% survival, respectively. The
mAb cocktail reduced viremia, the number of pox lesions and body weight
loss to a greater extent than VIG. In summary, the data show the mAb cock-
tail to be an effective poxvirus-neutralizing immunotherapeutic that is more
potent than VIG when administered post-exposure. (Support: NIAID grant
5U01AI070504-04 and NIAID contract N01AI30063 Task Order C24)
197 (H)
Clinical Development of a Plant-Produced Subunit Vaccine
against Bacillus anthracis
V. Yusibov, J. Chichester, S. Manceva, K. Musiychuk, M. Shamloul, A. Rhee, J.
Trefethen, M. Jones; Fraunhofer USA CMB, Newark, DE.
The potential use of biological agents or biowarfare threatens the security of
populations globally. Biological weapons can be easily concealed, trans-
ported and released with devastating consequences, making rapid response
capabilities a task of paramount importance. A quick response to a potential
bioterrorist attack requires the immediate availability of reagents for mass
therapeutic treatment or for mass vaccination. Bacillus anthracis is one of
the most-often cited biological threats. Extensive research efforts have been
directed towards the development of recombinant subunit vaccines based
on protective antigen (PA) of anthrax. Among the emerging technologies for
the production of these vaccine antigens is our “launch vector” system. Plant
virus-based transient expression systems are suitable for rapid engineering,
ease of scale-up and cost-effective production of target antigens. In addition,
this technology is particularly well suited for producing large quantities of
vaccine antigens in a short period of time, which is particularly important
when faced with natural outbreaks or intended release of bio-treat agents
such as anthrax. To this point, we have successfully engineered and ex-
pressed a PA antigen in Nicotiana benthamiana plants using agroinfiltration.
This plant-produced antigen elicited high toxin neutralizing antibody titers
in mice and rabbits after two vaccine administrations with Alhydrogel. In ad-
dition, immunization with this vaccine candidate protected 100% of rabbits
from a lethal aerosolized B. anthracis challenge. These results support the
testing of this vaccine candidate in human volunteers and the utility of our
plant expression system for the production of a recombinant anthrax vaccine.
The availability of safe, efficient and easily delivered vaccines against this
pathogen is of paramount importance to protect civilian populations and mil-
itary personnel.
198 (H)
Adoptive Transfer of Pulsed BMDC Enhances Specific Immune
Responses and Provides Protection against Lethal Anthrax
Challenge
I. J. Thompson1, M. G. Stokes1, B. Hallis2, A. J. Simpson1, E. D. Williamson1;
1
Defense Sci. and Technology Lab., Salisbury, United Kingdom, 2Hlth. Pro-
tection Agency, Salisbury, United Kingdom.
Dendritic cells are potent activators of the immune system and have a key
role in linking innate and adaptive immune responses. In the current study we
have used Bacillus anthracis as a model to develop an adoptive transfer strat-
egy, using pulsed Bone Marrow Dendritic Cells, to provide protection against
infectious agents. Recombinant Protective Antigen (rPA) formulated with
alum provides a high titre antibody response to rPA, a detectable T-cell re-
70 10th ASM Biodefense and Emerging Diseases Research Meeting
sponse and protection against lethal anthrax challenge. BMDC pre-pulsed
with heat-killed B. anthracis, rPA and CpG have upregulated expression of
CD80, CD86 and MHC-II, and by seven days post transfusion have induced a
specific T cell response in recipient mice. When rPA in alum is co-delivered
with pulsed BMDC we see enhanced T cell and antibody responses compared
to rPA and alum alone. Mice vaccinated ona a single occasion with a sub-op-
timal rPA and alum dose together with pulsed BMDC have 100% survival fol-
lowing lethal B. anthracis challenge, compared to 80% for rPA and alum
vaccinated control mice and 40% for mice receiving pulsed BMDC only.
199 (H)
Radioprotective Mn-DP-Pi Complexes Preserve Immunogenic-
ity of Inactivated VEEV During Gamma Irradiation: A New Ap-
proach for Vaccine Development
M. Gayen1,2, E. K. Gaidamakova1, P. Gupta1,2, A. Sharma1, P. J. Glass3, M. J.
Daly1, R. K. Maheshwari1; 1Uniformed Services Univ. of Hlth. Sci., Bethesda,
MD, 2Birla Inst. of Tehnology and Sci., Pilani, India, 3U.S. Army Med. Res. Inst.
of Infectious Diseases, Frederick, MD.
Venezuelan equine encephalitis virus (VEEV) is a mosquito borne New World
alphavirus in the family Togaviridae. VEEV causes biphasic disease with ini-
tial replication in lymphoid and myeloid tissues followed by infiltration of the
central nervous system initiating neuropathology and sometimes fatal en-
cephalitis. Currently there are no FDA approved vaccines for VEEV prophy-
laxis. A live attenuated strain of VEEV, V3526, was shown to provide excellent
immunogenicity in Phase I clinical trial however caused adverse effects in
vaccinees. Therefore, attempts to generate an inactivated VEEV vaccine are
ongoing. A high dose of gamma irradiation ensures complete virus inactiva-
tion but also leads to loss of immunogenicity due to oxidative damage of epi-
topes. Here, we describe a novel approach of gamma inactivation of V3526
while preserving the viral immunogenicity. Use of the attenuated V3526 over
the virulent TrD strain provides an additional layer of protection, the ability to
work in BSL-2 conditions and an additional protective epitope on the surface
of the virus. In previous work, we identified radioprotective Mn2+-peptide-Pi
complexes in highly radioresistant bacteria, Deinococcus radiodurans. In this
study we show that reconstituted Mn2+-DP-Pi complexes protect V3526 epi-
topes from high doses of gamma radiation, which obliterated its genome.
Viral envelope protein immunogenicity was examined using PE2 monoclonal
antibody. RNA infectivity was examined by transfection into sensitive cell
lines. Our study demonstrates that Mn2+-peptide-Pi complexes should be
further explored for generation of highly immunogenic gamma irradiated
VEEV vaccine. This work was supported by funding from the DTRA and Air
Force Office of Scientific Research. Opinions presented here are of the au-
thors and should not be construed as that of USUHS, DoD, DTRA, and US-
AMRIID.
200 (H)
Longevity of Vaccine Protection against Pneumonic Plague in
BALB/C Mice: Part 2
Z. N. Llewellyn1, F. Koide2, L. E. Bowen1, P. William2, S. Lin2, J. A. Boydston1, N.
Harman-Richardson3, P. M. Silvera2; 1Southern Res. Inst., Birmingham, AL,
2
Southern Res. Inst., Frederick, MD, 3Alpha StatConsult, LLC, Damascus, MD.
Background: Yersinia pestis is the etiological agent of bubonic and pneu-
monic plague. Subunit vaccine approaches comprising recombinant forms of
the fraction 1 (F1) and V (virulence) proteins have demonstrated protection
against pneumonic plague in mice. At last year’s meeting, we reported on
the protection conferred by rYP vaccine at early times post-vaccination and
showed that F1 and V antibody levels correlated with protection against chal-
lenge with Y. pestis CO92 [Abstract number 207 (H)]. Here, we have extended
our studies to further investigate the longevity of protection induced by rYP
vaccine. Methods: On day 0 and 21, three sets of Balb/c mice (N=110/set)

in Groups 1-6, were vaccinated with 2.50, 0.63, 0.16, 0.04, 0.01 and 0μg vac-
cine plus Alhydrogel, respectively. Blood collected on various times post-vac-
cination was analyzed for F1 and V antibodies by ELISA. On day 79, 109 and
139, the 3 sets of mice were exposed to 1.1x106, 1.4x106 and 2.0x106 CFU/L
Y. pestis CO92, respectively, via nose-only inhalation exposure and monitored
for 14 days. Results: On Day 21, mean F1 and V antibody titers in Groups 1-
5 were 2.72, 2.61, 2.34, 1.74, 1.32 and 1.56, 1.55, 1.55, 1.56, 1.56 log10
assay units, respectively. F1 and V antibodies continued to rise in Groups 1-
5 following the 2nd vaccination on Day 21. Notably, V-specific antibodies were
significantly boosted in Groups 1-4 by Day 29 (p<0.01). As expected, F1 and
V antibodies were not detected in Group 6. Following challenge with Y. pestis
CO92 on day 79 (set A), day 109 (set B) and day 139 (set C), survival rates in
Groups 1-6 were 95, 90, 85, 35, 0, 0, 90, 60, 85, 55, 5, 0, and 90, 90, 75, 45,
15, and 0%, respectively. Moreover, survival probability correlated with F1
and V antibody titers (p<0.001). Conclusions: These results corroborate our
previous findings demonstrating that F1 and V antibodies induced by an F1/V-
based vaccine correlated with protection against pneumonic plague. This
work was funded by NIAID contract N01-AI-30063
201 (H)
Dose Response of IMVAMUNE, a Third Generation Smallpox
Vaccine Candidate, in a Lethal Monkeypox Virus Nonhuman Pri-
mate Inhalation Model
T. Brasel, K. Agans, K. Dearen, T. Bailey, A. Russo, S. Storch, R. Sherwood;
Lovelace Respiratory Res. Inst., Albuquerque, NM.
First and second generation smallpox (SPX) vaccines composed of replicating
vaccinia are associated with adverse health risks. Because of this, testing of
new, safer vaccines is desirable. IMVAMUNE®, a formulation of live Modified
Vaccinia Ankara, is a promising candidate due to its inability to replicate in
mammalian cells. We have previously shown that a prime-boost regimen of
IMVAMUNE® is fully protective in a lethal aerosol monkeypox virus (MPXV)
nonhuman primate (NHP) challenge model. The objective of this study was to
determine the minimum effective dose of the vaccine using the same model.
NHP were vaccinated with IMVAMUNE® at 1 x 108, 1 x 107, or 1 x 106 TCID50
or vehicle on Study Days -67 and -39. MPXV neutralizing antibody (Ab) pro-
duction was assessed at key time points post vaccination. Animals were chal-
lenged with aerosolized monkeypox virus (1 x 105 pfu) on Study Day 0.
Clinical observations were performed daily for up to 28 days post-challenge.
Blood and pharyngeal swab analysis, lesion counts, and clinical pathology
were conducted at routine intervals. Upon necropsy, tissues from each animal
were collected and analyzed for MPXV burden and histopathological changes.
A dose response in neutralizing Ab titers was measured in non-control ani-
mals following vaccination. Alterations, predominantly associated with res-
piratory distress, were noted in all groups post-exposure, though were most
severe in controls. Among the 24 vaccinated animals, four succumbed to
MPXV-associated complications. Alterations in these monkeys were compa-
rable to those observed in controls. Death was attributed to severe fi-
bronecrotic pneumonia. Surviving animals demonstrated minimal alterations,
though severity was inversely proportional to the IMVAMUNE® dose. Cu-
mulatively, results demonstrated that the protective efficacy of IMVAMUNE®
was directly proportional to the administered dose. Though a definitive min-
imum effective dose was not determined, results demonstrated a waning ef-
ficacy as the vaccine dose decreased.
T U E S DAY A B S T R AC TS
202 (H)
Stabilization Technology for Biodefence Vaccines: Exemplifica-
tion Using a New Viral Vaccine and an Adjuvanted Vaccine
J. Drew, C. Andrews, E. Abbott, S. Ward; Stabilitech Ltd., London, United King-
dom.
Improved stability against temperature excursions for vaccines and biophar-
maceuticals would have a significant and obvious impact, particularly with re-
spect to the logistics of distribution in challenging conditions or where
temperature control variances are prone to occur. Stabilitech Ltd is a UK-
based company which has overcome thermostability issues for a wide range
of viruses, vaccines, biopharmaceuticals and Alhydrogel adjuvant using our
proprietary stabilization technology. We have developed proprietary excipi-
ent systems which, when mixed with suspensions of target vaccine or pro-
tein dramatically improve stability when stored at elevated temperatures (in
dry or liquid presentations) or when taken through multiple freeze thaw cy-
cles. The protein-free technology can be readily incorporated into cGMP man-
ufacturing processes using standard equipment. In order to illustrate the
application of this technology, results will be presented showing enhanced
stability of a novel recombinant adenovirus-based vaccine, at a range of tem-
peratures and timepoints, both in a iquid and lyophilized form. Data for an ad-
juvanted subunit protein vaccine of biodefense significance will also be
shown, showing long-term retention of both Alhydrogel and protein-antigen
structure. Stabilitech’s technology has overcome several major hurdles af-
fecting vaccine and biopharmaceutical storage and distribution. This propri-
etary technology has the potential to facilitate significantly the deployment
of vaccines in harsh conditions in the field.
203 (H)
A Marmoset Model for Lethal Orthopox Virus Infection Enables
Study of Vaccines
C. Yue1, M. Kramski1,2, C. Stahl-Hennig3, K. Mätz-Rensing3, F-J. Kaup3, G.
Pauli1, H. Ellerbrok1; 1Robert Koch Inst., Berlin, Germany, 2Univ. of Mel-
bourne, Parkville, Australia, 3German Primate Ctr., Goettingen, Germany.
Variola virus is under continuous discussion as a biological threat agent. This
discussion is paralleled by increasing numbers of zoonotic infections in hu-
mans with other orthopox viruses like monkeypox or cowpox viruses. While
the development of safer vaccines and antiviral therapies is desirable the
question remains how to test their efficacy. Recently, we have developed a
lethal non-human primate infection model using calpox virus and marmosets
to study the pathogenesis of orthopox virus infections (Kramski et al. 2010,
PLoS one 5, e10412). We have applied this model to test vaccines and vacci-
nation strategies using Vaccinia Virus Lister-Elstree (LE) and Modified Vac-
cinia virus Ankara (MVA) as vaccines. Marmosets were immunized with LE or
MVA and challenged with a lethal dose of calpox virus either four or ten weeks
after immunization (p.im.). Animals were routinely checked for clinical symp-
toms. Viral load in blood and in saliva were analyzed by real-time PCR p.im.
and post challenge. Virus-specific antibody titers were determined by im-
munofluorescence test and by plaque-reduction neutralization test. All four
marmosets immunized with LE and challenged ten weeks p.im. survived with-
out any clinical symptoms although calpox virus was transiently detectable
in blood. Only one out of four animals challenged four weeks p.im. survived
the lethal calpox virus challenge. In contrast, none of the animals (n=4) vac-
cinated with MVA and challenged 10 weeks p.im. survived and only 2 out of
7 animals challenged 4 weeks p.im. survived. In some of the 5 animals suc-
cumbing to disease the onset of symptoms was only slightly delayed. In con-
clusion, the non-human primate model is suitable for the study of orthopox
virus pathogenesis as well as for different vaccination strategies but exten-
sion of the observed results to humans needs to be analyzed in more detail.
BIODEFENSE PROGRAM & ABSTRACTS 71
T U E S D A Y P O S T E R S E S S I O N | 1:00–3:00 p.m. | Exhibit Hall
204 (H)
New Adjuvants for Anthrax Vaccine
Y. Gauthier1, A. Borel2, Y. Trescos1, J. Mathieu1, B. Verrier2, J-N. Tournier1;
1
IRBA, La Tronche, France, 2CNRS-IBCP, Lyon, France.
We developed a vaccine based on PA and spore (FIS) with two adjuvants. Alu-
minum hydroxide (Al) has side effects. The mucosal adjuvant cholera toxin
(CT) is not usable in humans. These limitations led to the search of new ad-
juvants. Particular synthetic oligodeoxynucleotides (CpG ODN) are potent ad-
juvants. The safe and biodegradable PLA-based (Poly (D,L)-Lactic Acid)
nanoparticles (NPs) have adjuvant properties. The synthetic RNA polyri-
boinosinic polyribocytidylic acid (poly(I:C)) also has adjuvant effects. Our aim
was to assess the efficiency of these potent adjuvants in a mouse model of
inhalational anthrax. Mice were immunized with rPA. Animals were IN infected
with spores of either the 7702 Sterne strain or the 9602 strain of B. anthracis.
PA-PLA was as efficient as both PA-Al and PA+CT for SC and IN immunizations
respectively, and elicited 100% protection against an IN lethal challenge of
DBA/2 mice with 10.LD50 of the Sterne strain. PA+Poly(I:C) elicited full pro-
tection as well, through either the SC or the IN route of immunization. Inter-
estingly, PA alone was fully protective when administered SC, and elicited no
protection through the IN route. PLA+Poly(I:C), without PA, elicited partial
protection when administered IN. In SWISS mice immunized with PA+FIS and
challenged IN with 10.LD50 of the fully virulent 9602 strain, SC immunization
with CpG did not elicit better protection than Al. Total anti-PA and anti-spore
IgG titers, both in the sera and BALs from CpG immunized mice, were not
modified compared with titers obtained with Al. Nevertheless, when PA+FIS
were delivered both SC and IN, anti-PA titers were decreased in an average
range of 10 times in mice immunized with CpG compared with Al. TNA titers
were decreased 4 times as well after CpG immunization. PA+Poly(I:C) was as
efficient as PA-Al for inducing 100% protection against an IN lethal infection
with the Sterne strain. Immunization of DBA/2 mice with biocompatible 200
nm PA-PLA NPs was as fully protective as PA-Al. CpG-ODN 1826 used as sin-
gle adjuvant for vaccination with (PA+FIS) did not improve the immune re-
sponse and protective activity against lethal inhalational anthrax compared
with Al.
205 (H)
Effects of Signal Sequences on the Immunogenicity and Effi-
cacy of an MVA Vectored Plague Vaccine
J. Brewoo1, T. Powell2, G. Young1, C. Partidos1, D. Stinchcomb2, J. Osorio3; 1In-
viragen, Inc., Madison, WI, 2Inviragen, Inc., Fort Collins, CO, 3Univ. of Wis-
consin, Madison, WI.
Background: Signal sequences have been employed in vaccine development
to facilitate the expression of target antigens and significantly enhance their
immunogenicity. We report on the effects of different post-transcriptional reg-
ulatory signals on the immunogenicity and protective efficacy of a recombi-
nant modified vaccine Ankara (MVA) vector expressing a truncated version
of the low-calcium virulence antigen (V307) from Yersinia pestis. Methods:
MVA recombinants were constructed expressing antigen from Y. pestis (V307)
under the translational control of the encephalomyocarditis virus (EMCV) in-
ternal ribosomal entry site (IRES) and/or fused to the tissue plasminogen ac-
tivator (tPA) or a putative vaccinia virus secretory signal (C13L). The candidate
vaccines were tested for antigen expression in vitro, immunogenicity and pro-
tection in a mouse plague challenge model. Results: Western blot analysis
showed increased antigen expression from vaccine constructs co-expressing
V307 and tPA (MVA/tPA/V307) or C13L (MVA/C13L/V307) compared to con-
structs expressing only the V307 (MVA/V307). Antigen expression from the
construct co-expressing V307, IRES and tPA (MVA/IRES/tPA/V307) was signif-
icantly lower than the other constructs. Interestingly, immunogenicity of the
candidate vaccines was superior with constructs co-expressing signal se-
quences. Following prime and boost (28 days apart) vaccination, MVA/V307
72 10th ASM Biodefense and Emerging Diseases Research Meeting
elicited antibody titer of 1000 GMT compared to 43000 - 66000 GMT from
constructs co-expressing signal sequences. Similarly, MVA/V307 conferred
25% protection against challenge with 50MLD of Y. pestis (CO92) compared
to 100% protection conferred by constructs co-expressing signal sequences.
Conclusions: The adoption of MVA as a second-generation smallpox vaccine
makes our recombinant MVA candidate vaccines attractive alternatives to
protect against smallpox and plague.
206 (H)
Second-Generation Inactivated Vaccines Protect Mice from
Lethal Venezuelan Equine Encephalitis Virus Infection
K. Spurgers1, S. Honnold1, R. Bakken1, J. Cohen1, L. Rowan1, C. Lind1, P.
Gupta2, A. Sharma2, R. Maheshwari2, P. Glass1; 1USAMRIID, Frederick, MD,
2
USUHS, Bethesda, MD.
Venezuelan (VEEV), eastern (EEEV), and western equine encephalitis (WEEV)
viruses, members of the genus Alphavirus, are causative agents of debilita-
tive and sometimes fatal encephalitis in North, Central, and South America.
Natural human cases are rare, and occur from the bite of an infected mos-
quito. However, these viruses pose a threat to military personnel, and to pub-
lic health, because of their potential use as bioweapons. There are currently
no licensed vaccines to prevent alphavirus infections. To address this need,
our laboratory group derived and tested an attenuated, PE2 cleavage site
mutant of VEEV, termed V3526. V3526 was a safe and effective vaccine plat-
form in rodents and primates, and advanced to phase I clinical trials. How-
ever, human recipients of V3526 exhibited an unacceptably high frequency of
adverse events requiring cessation of the study. To increase the safety of this
vaccine candidate, several inactivated V3526 formulations were tested for
efficacy against VEEV infection. Inactivation methods included formalin,
gamma irradiation, and 1,5 iodonaphthyl azide (INA). In initial studies, small
doses of vaccine were administered, and afforded only partial protection
against aerosol VEEV challenge. The current study tests higher doses of three
inactivated V3526 formulations without adjuvant, administered by three dif-
ferent routes (intranasal, intramuscular, and subcutaneous). Cohorts of mice
received either one dose, or two doses of vaccine (28 d apart). Mice were
then challenged with aerosolized VEEV (Trinidad). Complete protection was
observed when two doses of gamma irradiated-V3526 or INA-V3526 were
administered i.m. For each cohort of mice, total serum anti-VEEV antibody,
and neutralizing antibody levels are reported. Best performing vaccine for-
mulations, route, and dose will be down-selected for further testing with ad-
juvants.
207 (H)
Evaluation of Inactivated and Live-Attenuated Bivalent Vac-
cines That Confer Protection against Rabies and Ebola Viruses
in Mice
J. Blaney1, A. Papaneri1, C. Wirblich2, J. Cann3, M. Holbrook3,4, A. Freiberg4, P.
Jahrling1,3, M. Schnell2; 1 NIAID, NIH, Fort Detrick, MD, 2 Jefferson Med. Coll.,
Thomas Jefferson Univ., Philadelphia, PA, 3Integrated Res. Facility, NIAID,
NIH, Fort Detrick, MD, 4Univ. of Texas Med. Branch, Galveston, TX.
Our objective is to generate an Ebola virus vaccine with maximum likelihood
of licensure based on considerations of safety, manufacture, and cost. To this
end, we developed (A) replication-competent, (B) replication-deficient, and
(c) chemically inactivated rabies virus (RABV) vaccines expressing Zaire
ebolavirus glycoprotein (GP) using a reverse genetics system based on the
SAD B19 RABV wildlife vaccine. Each vaccine candidate is apathogenic upon
peripheral administration in mice, induces anti-RABV and anti-Ebola humoral
immunity, and confers protection from lethal mouse-adapted Ebola and RABV
challenge. We have further characterized the safety and immunogenicity of
these live and inactivated vaccine candidates. Replication-deficient vaccine
candidates expressing GP with a deletion in the RABV glycoprotein were

found to be avirulent after intracerebral inoculation of adult and suckling
mice and peripheral administration in SCID mice. Analysis of replication-de-
ficient viruses in the brain by quantitative PCR, determination of virus titer,
and immunohistochemistry indicated greatly restricted virus replication. Fur-
ther examination of the immune response to these RABV vectored vaccines
by ELISPOT assay indicated that a memory T cell response is induced by both
live and killed vaccines. Finally, we demonstrated that a multivalent antibody
response can be induced by co-administration of the RABV/Ebola vaccine
and another RABV vectored vaccine expressing the botulinum neurotoxin A
HC50 fragment. Based on the strong safety and immunogenicity profiles of
these bivalent vaccines directed against RABV and Ebola in mice, we are cur-
rently planning studies in nonhuman primates.
208 (H)
Improved Efficacy of a Multiagent Equine Encephalitis Virus
Vaccine Candidate
P. J. Glass1, R. R. Bakken1, J. Ennis2, K. B. Spurgers1, K. I. Kamrud3, K. Alter-
son3, M. Custer3, J. Turner2; 1USAMRIID, Fort Detrick, MD, 2Defyrus Inc.,
Toronto, Canada, 3Alphavax, Research Triangle Park, NC.
Background: Virus-like replicon particles (VRPs) were produced that express
the Venezuelan (VEEV), eastern (EEEV) or western (WEEV) equine en-
cephalitis virus E1 and E2 glycoproteins. Previous studies showed that a mul-
tiagent equine encephalitis virus vaccine consisting of an equal mixture the
VEEV, EEEV and WEEV VRPs (1x107 IU each) administered subcutaneously
protected mice from lethal aerosol challenge with VEEV, EEEV and WEEV.
Methods: In this study, we examined the efficacy of a lower dose of the mul-
tiagent VRP (maVRP) vaccine (1x105 IU each) administered by intramuscular
(IM) or intranasal (IN) routes, with or without DEF201. DEF201 consists of a
proprietary adenovirus type 5 (Ad5) vector expressing interferon (IFN)-α, that
is administered IN. BALB/c mice were given one or two doses of the maVRP
vaccine or PBS, with or without DEF201 (primary vaccination only) and chal-
lenged 28 days post-final vaccination with VEEV, EEEV, or WEEV. Mice were
monitored daily for mortality and clinical signs of disease for 21 days
postchallenge. Results: Mice given one or two doses of the maVRP vaccine
alone by the IN or a single dose by the IM route were not protected against
lethal aerosol challenge with VEEV, EEEV or WEEV. DEF201 administered in
conjunction with one or two IN doses of the maVRP vaccine provided en-
hanced protection (80-100%) against lethal EEEV and WEEV aerosol chal-
lenge. Two IM doses of the maVRP vaccine alone resulted in 70-90%
protection. Administration of DEF201 in conjunction with two IM doses of the
maVRP vaccine completely protected mice against lethal VEEV, EEEV, or WEEV
challenge. Conclusions: IM vaccination with a lower dose of the maVRP vac-
cine effectively protected mice from lethal challenge with VEEV, EEEV, or
WEEV when administered as a single dose regimen with DEF201 or a two-
dose regimen with or without DEF201. Future studies will examine onset and
duration of the maVRP vaccine efficacy when administered in conjunction
with DEF201.
209 (H)
Relationship between Vaccine Dosage, Immunogenicity and
Survival in RF1V-Vaccinated Cynomolgus Macaques
P. Fellows1, J. Price1, R. Krile2, H. Lockman2, L. Wolfraim1; 1Dynport Vaccine
Company Llc, Frederick, MD, 2Battelle, Columbus, OH.
Objective: The objective of these studies was to determine minimum protec-
tive titers in cynomolgus macaques (CM) vaccinated with a recombinant
plague vaccine (rF1V) and subsequently challenged by aerosol exposure to
Yersinia pestis CO92. Methods: Two dosage ranging studies were performed
to establish a full survival curve (0 to 100% survival) associated with rF1V
vaccination. CMs were vaccinated with rF1V on Days 0, 56 and 121 by intra-
muscular injection with dosages ranging from 0.016 to 20 µg and were chal-
T U E S DAY A B S T R AC TS
lenged on Day 149 by aerosol exposure to Y. pestis CO92 at a target dosage
of 5.5 x 103 cfu. Animals were observed for signs of disease and changes in
clinical pathology, body temperature and activity. Samples for evaluation of
humoral immune responses were taken throughout the study and on the day
of challenge and were tested in a Bridge ELISA for presence of anti-rF1, anti-
rV and anti-rF1V antibodies. The data generated from these studies were sta-
tistically analyzed to provide a more precise estimate of the relationship
between vaccine dosage, Bridge ELISA titers and survival. Histopathology
and residual Y. pestis were evaluated in the lungs, liver, kidney and spleen in
all animals that succumbed to disease. Results: A full survival curve was ob-
tained over the course of both studies. The protective titer and protective
vaccine dosage at 50% survival were estimated to be 878 U/mL of anti-rF1V
and 2.05 µg rF1V vaccine, respectively. Conclusions: The results of these
studies indicate a significant relationship between vaccine dosage and sur-
vival (p<0.001) and between Bridge ELISA titer and survival (p<0.001) for all
analytes (rF1, rV and rF1V).
210 (I)
Peptide Mapping of Anthrax Toxin Protective Antigen Using
Anti-AVA Human Sera
V. A. Semenova, P. Svoboda, J. Pohl, S. Soroka, C. Quinn; CDC, Atlanta, GA.
Anthrax toxin protective antigen (PA) is a major protective immunogen in the
anthrax vaccine adsorbed (AVA, BioThrax). Identification of antigenic se-
quences of PA is important for future vaccine designs and immunotherapeu-
tics. We characterized PA linear epitopes using sera with anti-PA IgG
concentrations ≥ 300 μg/ml from human volunteers in the CDC Anthrax Vac-
cine Research Program (AVRP). Fmoc synthesis was used to prepare 37 N-
terminally biotinylated peptides of 30 amino acid (AA) residues, overlapping
by 10 AA. Except for the C-terminal peptide made as the free acid, sequences
were Fmoc synthesized as C-terminal amides, HPLC-purified and prepared as
trifluoracetic acid salts. Peptides were captured on streptavidin coated (2
μg/ml) microtiter plates. Seroreactivity was evaluated colorimetrically using
sera of 60 volunteers who had received 8 subcutaneous (SQ) or at least 3 in-
tramuscular (IM) injections of AVA with the licensed regimen. A peptide was
considered antigenic if its OD value was greater than the mean OD+3SD of
control group sera (n=24). Sera from all vaccinees reacted with peptide se-
quences corresponding to all 4 domains of PA. There were four predominant
seroreactive PA peptide regions. These peptides corresponded to amino acid
(AA) sequences in the chymotrypsin sensitive loop 2ß2-2ß3 (AA 301-330),
2ß3- 2α1(AA321-350), 2ß10-12(AA421-450) in domain 2, and 4ß5-8(AA641-
670) in domain 4 of mature protein sequence Bacillus anthracis V770-NP1-R
(BioThrax Vaccine Strain). We demonstrated that serum antibodies from
human AVA recipients exhibit reactivity to a range of different sequences in
PA. The major antigenic regions were located in domains 2 and 4. These data
in humans concur with the previous findings in non-human primates that pro-
tective epitopes can be attributed to specific PA domains (Flick-Smith et al.,
2002; Williamson et al., 2005).
211 (I)
Serum Opsonization Promotes the Efficient Uptake and Clear-
ance of Burkholderia pseudomallei by Human Neutrophils
M. E. Woodman, R. G. Worth, R. M. Wooten; Univ. of Toledo Coll. of Med.,
Toledo, OH.
Burkholderia pseudomallei is the causative agent of melioidosis and is a
major mediator of sepsis in its endemic areas. Because of the low LD50 via
aerosols and resistance to multiple antibiotics, it is considered a Category B
bioterrorism agent by the CDC. B. pseudomallei is an encapsulated bacterium
that can infect, multiply, and persist within a variety of host cell types. Burk-
holderia thailandensis is closely-related to B. pseudomallei, but lacks a poly-
saccharide capsule and is relatively avirulent in infection models. In vivo
BIODEFENSE PROGRAM & ABSTRACTS 73
T U E S D A Y P O S T E R S E S S I O N | 1:00–3:00 p.m. | Exhibit Hall
studies suggest that macrophages and neutrophils are important for con-
trolling B. pseudomallei infections, however few details are known regard-
ing how neutrophils respond to these bacteria. Our goal is to describe the
capacity of human neutrophils to control highly virulent B. pseudomallei com-
pared to the relatively avirulent B. thailandensis using in vitro analyses. B.
thailandensis was more readily phagocytosed than B. pseudomallei, but both
displayed similar rates of persistence within neutrophils, indicating they pos-
sess similar inherent abilities to escape neutrophil clearance. Serum op-
sonization studies showed that both were resistant to direct killing by
complement, although B. thailandensis acquired significantly more C3 on its
surface than B. pseudomallei. Both Burkholderia spp. showed significantly
enhanced uptake and killing by neutrophils after serum opsonization. Killing
of serum-opsonized B. pseudomallei and B. thailandensis was prevented with
inhibition of the NADPH-oxidase indicating the importance of reactive oxy-
gen species in this process. Serum-opsonized Burkholderia induced a sig-
nificant respiratory burst by neutrophils compared to unopsonized bacteria.
In summary, while complement opsonization is important for efficient uptake
and killing of both strains, the absolute levels of surface C3 does not corre-
late with increased killing of B. pseudomallei and B. thailandensis. Based on
these findings, neutrophils appear able to play a significant role in clearing
B. pseudomallei infection.
212 (I)
Priming of Alveolar Macrophages Augments Inflammatory Re-
sponse on Anthrax Lethal Toxin Stimulation
J. Thomas, L. Xiao, J. W. Christman, J. L. Cook, I. Levitan; Univ. of Illinois Med.
Ctr. at Chicago, Chicago, IL.
Background: The cause of rapid death in anthrax infection both in animals
and humans are unknown and concern over the use for biological warfare
has renewed interest in elucidating the mechanisms of anthrax induced in-
flammation. Interleukin-1β (IL-1β) secretion is an important inflammatory re-
sponse against anthrax lethal toxin (LeTx) a virulence factor of Bacillus
anthracis and Lipopolysaccharides (LPS), an endotoxin of all gram-negative
bacteria. Recently, we reported that in mouse macrophages LeTx activates, in-
wardly-rectifying K+ (Kir) and voltage-gated K+ channels (Kv), the 2 types of K+
channels in mouse macrophages. Blocking these channels inhibits IL-1β se-
cretion. Susceptibility to LeTx is augmented by a two-fold increase in IL-1β se-
cretion primed overnight with LPS compared to unprimed cells challenged
with LeTx alone. We hypothesize that reactive oxygen species (ROS) gener-
ated by LPS activate K+ channels and may prime cells for increased activation
upon LeTx challenge. Methods: Whole-cell patch clamp was used to deter-
mine if toxin and ROS activates K+ channels in macrophages. ROS activation
of K+ channels was measured by a brief exposure to H2O2 or a ROS scavenger
MnTmPyP. Release of IL-1β was determined by ELISA. Results: LPS primed
LeTx stimulation yielded a higher IL-1β secretion and Kir current compared to
unprimed LeTx stimulation suggesting priming affect of LPS on K+ channels
mediated inflammatory response. As a first report we show that LPS alone is
sufficient to illicit K+ channel activation and IL-1β secretion. LPS stimulate K+
channels in a dose and time depended manner. Lastly, H2O2 alone can acti-
vate K+ channels and inhibition of LPS-generated ROS by MnTmPyP inhibits
K+ channel activation , suggesting the role of ROS in K+ channel activation.
Conclusion: In macrophages LeTx or LPS alone is sufficient to induce both IL-
1β secretion and activation of K+ channels. LPS-generated ROS may be a prim-
ing mechanism of K+ channel activation and IL-1β secretion. This model may
be useful to design new therapeutic strategies to block endotoxic-shock re-
sponse to anthrax infection.
74 10th ASM Biodefense and Emerging Diseases Research Meeting
213 (I)
A Quantitative PCR-Based Method for the Evaluation of Anti-
Vaccinia Neutralization Titers
A. D. Garcia, C. A. Meseda, J. Weir; FDA/CBER, Bethesda, MD.
The plaque reduction neutralization test (PRNT) is a standard method used
for measuring poxvirus neutralizing antibodies in vaccinees. However, the
PRNT is labor- and time consuming and can require relatively large amounts
of immune serum, making this neutralization test impractical for use in large
scale studies and in the mouse challenge model. Therefore, the goal of this
work was to develop a rapid and efficient method to measure vaccinia virus
neutralization based on viral DNA sampling performed in a multi-well format.
Here, we have applied a SYBR green endpoint quantitative PCR methodology
to quantitate the changes or reduction in vaccinia genome copy numbers, as
the assay readout, in response to neutralization of virus infectivity by vari-
ous immune sera. To evaluate the feasibility of a vaccinia qPCR assay method,
we have designed and tested PCR primer sets specific to 3 vaccinia virus
genes: J2R, G9R, and E9L. Our preliminary studies have determined that
primers to the J2R and G9R genes were suitable for further development of a
vaccinia qPCR method. Next, the qPCR assay was optimized for both primer
sets in which the PCR reactions contained ~102 genomic equivalents of the
respective control reference plasmid, pCR-vvJ or pCR-vvG. The assay quanti-
tative range was determined to be ~102 to >108 genomic equivalents. In ad-
ditional feasibility studies, we have demonstrated that the qPCR assay can
reproducibly measure virus genomic copies in total cell extracts prepared
from Vero cell monolayers (in 48-well plates) that were infected with ~100
pfu of vaccinia virus (WR strain or NYCBOH strain). Based on neutralization
experiments done with rabbit anti-vaccinia virus polyclonal antibodies, our re-
sults indicate that the qPCR method represents a viable approach for the de-
tection and measurement of vaccinia-specific neutralizing antibodies in
immune sera.
214 (I)
The Chemokine Receptor CCR7 is Expressed in Cells of the
Human Lung in Association with Bacillus anthracis Spore In-
fection
M. Langer1, J. Booth1, E. Duggan1, V. Patel1, T. Veres2, F. Prenzler2, K. Sewald2,
K. Coggeshall3, A. Braun2, J. Metcalf1; 1Univ. of Oklahoma, Oklahoma City, OK,
2
Fraunhofer Inst. for Toxicology and Experimental Med., Hannover, Germany,
3
Oklahoma Med. Res. Fndn., Oklahoma City, OK.
Background: The lung is the entry site for Bacillus anthracis in inhalation an-
thrax, the most deadly form of the disease. B. anthracis does not cause dis-
ease in the lung. Instead, spores are internalized by resident lung cells and
disseminate to cause systemic disease as the vegetative form of the organ-
ism. Three cell types resident in the lung have been previously shown to in-
ternalize spores: alveolar macrophages (AM), dendritic cells (DC) and alveolar
epithelial cells (AEC). One of these cells may contribute to dissemination by
migrating to the lymphatic system after uptake of spores. In particular, we
hypothesized that DC may responsible for carrying spores to the lymph
nodes. If this occurs, the chemokine receptor CCR7, a receptor involved in
DC chemotaxis, may be upregulated after spore uptake. Methods: We used
fluorescent immunohistochemistry and confocal microscopy, in addition to
immunofluorescent staining and flow cytometry to investigate spore uptake
and CCR7 expression. Our models were human lung organ culture and iso-
lated human AM obtained by bronchial-alveolar lavage. Results: We found
that the majority of spores were ingested by AM and DC, while AEC ingested
significantly fewer spores. Spore ingestion was associated with increased ex-
pression of CCR7 in cells that take up numerous spores, either AM or DC.
However, the relevant cell type is not likely AM because isolated AM do not
express CCR7 after taking up spores. Conclusions: Our results suggest that
DC may increase expression of CCR7 after internalization of spores and may

migrate to the lymph nodes carrying spores. Our results further suggest that
CCR7 is a potential therapeutic target in preventing dissemination of B. an-
thracis from the lung, and thereby preventing systemic disease.
215 (J)
Forensic Entomology and Biodefense
A. Lindström; Natl. Vet. Inst., Uppsala, Sweden.
Forensic entomology is the application of the science of entomology to any
legal case. Traditionally it is used for postmortem interval estimates in homi-
cide investigations where the age of insects on the corpse is used as an ap-
proximation of the time that have elapsed since the person died. For
biodefense purposes the science of forensic entomology is more involved in
predicting scenarios where insects are used to inflict economic or bodily harm
to humans, livestock or crops. If such an attack was discovered the forensic
entomologist would also have an important task in collecting and interpeting
the evidence so that the offenders can be brought to justice. Unfortunately
from a biodefense viewpoint insects are a problem since they present a rel-
atively cheap low tech, high impact, low risk way for terrorists to spread
pathogens or at least the fear of pathogens. Instead of obtaining pathogens
by stealing them from high security facilities, which is a common scenario,
they can be found naturally infecting wild animals and livestock in several
nations of concern in for example North Africa and the Middle East as well as
USA. Diseases such as bubonic plague and Rift Valley Fever could easily be
obtained together with appropriate insect vectors. Propagation of the vector
and the pathogen would then be straightforward utilizing information read-
ily available on the internet. For the medical entomology community as well
as for medical and veterinary practitioners it is important to have forensic
awareness when it comes to vector borne diseases in order to discover in-
tentional release of disease vectors.
216 (J)
Rapid Detection of Endospores by Capillary Electrophoresis on
a Hybrid Channel-to-Digital Microfluidic Platform for Steriliza-
tion Validation
L. Li, W. Wu, W. Chan, H. Cheong, P. Yung; The Chinese Univ. of Hong Kong,
Shatin, Hong Kong.
We present the design, implementation and demonstration of a microfluidic
device for the rapid detection of endospores by time-gated fluoresecnce and
bioluminescence. Endospore formation is a very successful survival and dis-
persal strategy. They are found in nearly every conceivable natural habitat
on Earth and persist for long times due to their extreme resistance. There-
fore, endospores have been fascinating subjects of investigation for a num-
ber of important applications, from medical sterilization validation, food
safety, biodefense to astrobiology. Detection of low levels of dormant en-
dospores under various extreme environments presents great challenges to
researchers. Here we propose a rapid and automated device to determine
endospore viability from different perspectives, namely germination propen-
sity, metabolic activity, protein degradation and membrane integrity. En-
dospores are detected based on a series of unique physiological changes
during germination, exemplified by dipicolinic acid release, ATP synthesis,
protein degradation and membrane stainability. Total endospore population
is determined by monitoring dipicolinic acid release upon electroporation.
Sample handling, processing and analyses are incorporated into a hybrid mi-
crofluidic chip highlighting capillary electrophoresis and channel-to-digital
conversion for smaller reagent volumes, shorter reaction times, and a better
separation efficiency. This lab-on-a-chip design enables portability and on-
the-spot analysis to probe the in situ viability state of endospores in their na-
tive environments, providing us valuable information in medical sterilization
validation and food safety.
T U E S DAY A B S T R AC TS
217 (K)
WMD Training Courses at Dugway Proving Ground, Dugway UT
P. B. Brown, J. Montague, J. Sorenson; U.S. Army, Dugway Proving Ground,
Dugway, UT.
How do you collect a powder sample aseptically and limit the spread of the
contamination? What does Bacillus anthracis look like on an agar plate? Do
all Bacillus spores look the same under the microscope? What are the signa-
tures for a bacterial production lab? Where is the best sample point when
sampling a clandestine biological laboratory? All of these questions and more
will be answered during courses offered at US Army, Dugway Proving Ground.
Courses are taught by microbiologists that emphasize hands on training. Stu-
dents will gain experience in aseptically collecting powder and liquid sam-
ples. They will practice collecting samples from powder disseminations; they
will learn to recon and sample a rogue laboratory, and work with attenuated
strains of Bacillus anthracis, Yersinia pestis and Francisella tularensis. Stu-
dents will gain knowledge and experience using Biological Detection equip-
ment such as Hand Held Assays, RAZOR, RAPID, HazMat ID, and Microscopes.
Customers include First Responders, Civil Support Teams and other Military
units.
218 (K)
WITHDRAWN
219 (K)
Biological Security: Identifying Vulnerabilities, Preventing
Damage
C. Uhlenhaut; Robert Koch Inst., Berlin, Germany.
Our goal is to identify vulnerabilities within the biosecurity context in Ger-
many by combining different approaches in an innovative way. One main pil-
lar of this program is developing generic scenarios. Being build from blocks,
these should ideally work for several settings or several pathogens, e.g., de-
pending on the infectious route. These scenarios will then be used to work on
risk assessments, involving scientists but also specialists from other fields.
Since the scenarios are scalable, several theoretic outcomes can be modeled
and evaluated for each pathogen or setting. The scenarios will be tested by
entering data and information from previous outbreaks and incidents. These
can then be modified by changing the setting, climate conditions, mode of
transmission etc. The information obtained will be used to identify knowl-
edge gaps, prioritize resources (e.g. financial, research) and to provide im-
proved information. This information will be custom-tailored, e.g., for policy
makers, first responders or the general public. To achieve this, we identify
the risk communication needs and networks for which we then produce our
own videos, print material and other online resources. One of these will be a
new databank which will contain all relevant information about biological
materials, not only biothreat agents or highly pathogenic organisms. We
chose this broad approach in order to provide one format for all potential bi-
ological hazards, not only intentional releases but also accidental releases or
naturally occurring infections. We will develop different output formats for
different users, e.g., first responders will have all the relevant information at
hand while someone trying to assess the potential economic impact of a cer-
tain biothreat agent will get a different set of information. However, this will
be done to facilitate working with the database and if necessary more or all
information will be available. We are working on a biothreat safety net, com-
bining known facts with generic scenarios and expertise from a broad range
of experts. In the end we will have a set of tools, such as generic scenarios
and an intelligent database that will allow assessing risks and identifying vul-
nerabilities.
BIODEFENSE PROGRAM & ABSTRACTS 75
T U E S D A Y P O S T E R S E S S I O N | 1:00–3:00 p.m. | Exhibit Hall

220 (K)
Transitional Operations: Guiding Owners through Start-Up Op-
erations & Regulatory Compliance
M. L. Weiss, D. C. Sharpe; The WorkingBuildings Companies, Atlanta, GA.
Objective: To introduce a new process in construction project delivery that
incorporates SOPs, Regulatory Compliance, Safety and Maintenance as the
drivers for successful design, commissioning and operations. The focus is on
the transition period between acceptance of a new building and full opera-
tions. Methods: This topic will be explored through a case study that includes
various testing protocols in BSL-2 and BSL-3 spaces. The presentation will
outline the importance of the Owner’s responsibility for information gather-
ing, organization and documentation on the successful transition between
acceptance and operations. Discussion: Laboratories are becoming more
complex to design, build and operate, and in many cases have been deliv-
ered but never become fully operational. In response to this, commissioning
has become an accepted practice to ensuring that the complex systems are
aligned with the Design Intent. Conclusion: Transitional Operations is a
process utilizing a team of knowledgeable facility operators and regulatory
compliance experts that is on-site full-time during the transitional period to
ensure the Design Intent is met and to guide Owners through the task of op-
erating and maintaining their new laboratory. It is a process that builds upon
traditional commissioning activities and extends into through the first year of
operations allowing Users and Facilities Management time to focus on the
ongoing operations of their existing facility and coordinating the move and
assistance with the laboratory regulatory compliance and certification
process. This time also allows for facility and regulatory compliance training
and shortening the users learning curve on the operations and management
of the laboratory’s new building systems.

76 10th ASM Biodefense and Emerging Diseases Research Meeting

 M O N D AY O R A L A B S T R A C T S

dicated that C. burnetii spreads into the environment in the direction of the
Highlighted Oral Abstract Presentations prevailing wind, but at 200-400 meters from the breeding areas very limited
009. Environmental Detection amounts of C. burnetii could be detected. These results demonstrate that al-
though large amounts of C. burnetii were released into the environment at
Monday, February 27, 2012 | 3:00 PM - 4:00 PM these farms during the birthing of infected goats, conditions at these farms
Diplomat Ballroom resulted in limited spreading to locations outside of the birthing area.

108 110
Case-Clusters of Melioidosis Associated with Climate Condi- Rapid and Sensitive On-Site Identification of Biothreat Agents
tions and Geographical Topography in Southern Taiwan Using Electrochemical Biochips and a Portable Detection Plat-
Y-L. Chen1, Y-S. Chen2, H-S. Lin3, P-J. Liu1; 1Natl. Kaoshiung Normal Univ.,
form
Kaohsiung, Taiwan, 2Kaohsiung Veterans Gen. Hosp., Kaohsiung, Taiwan, 3E-
T. Elssner, C. Pöhlmann; Bruker Daltonik GmbH, Leipzig, Germany.
DA Hosp., Kaohsiung, Taiwan.
Rapid detection of toxins and pathogenic bacteria is a prerequisite for a huge
As an emerging disease, melioidosis case clusters have been occurred in
number of applications, including bio-security and defense, medical diag-
2009 (n=8) and 2011 (n=12), in a small area (13 km2), in southern Taiwan
nostics, environmental and food analysis. Over the last decade, the anthrax
after violent typhoons go through. The geographical topography of this cer-
or ricin findings in the US postal system clarified the urgent need to develop
tain area appears the hills (50-400 m) are surrounding on north, east and
rapid, sensitive and specific detection systems for biothreat agents. Currently,
south as well as seashore on west. By culture-confirmed and PCR-based de-
the most widely used test to detect the presence of toxins is ELISA. However,
tection using soil (n=80) or water (n=36) specimens, the positivity to Burk-
these techniques cannot be performed by untrained personnel in the field
holderia pseudomallei was found among specimens that collected from the
and they are time-consuming. Remarkable advantages of electrochemical
north of this certain area but negativity in that collected from south of this
biosensing compared to common used optical methods are the high sensi-
area or out of surrounding by hills. By PFGE (pulsed-field gel electrophoresis)
tivity, simple operation, and the possibility of being used in portable instru-
analysis, the molecular linking of isolates between environments and pa-
ments for on-site testing. Here, we present an electrochemical detection
tients was demonstrated. In following as typhoon history, at least, Taiwan
technology applying biochips with immobilized antibodies against BoNT/A,
has been occurred 20 typhoon invasion from 2008 to 2010. If the typhoon
BoNT/B, BoNT/F, ricin and SEB. The detection platform allows simultaneous
appeared the maximum wind velocity of 40 m/sec and wind direction from
identification of these toxins within less than 25 minutes in an automated
northwest to southeast, the soil or water contaminated with B. pseudomallei
process. Detection limits for BoNTs, ricin and SEB are in the pg/mL to low
could be brought by wind vector from north and it associated with the oc-
ng/mL-range. We demonstrate identification of ricin from different Ricinus
currence of melioidosis case clusters in south of this certain area after ty-
communis varieties, whereas closely related lectins result in no signal. Fur-
phoon go through. However, out of this area (appeared hill barriers), no
thermore, we show specific detection of BoNTs and SEB directly from various
melioidosis case was occurred. We suggested that occurrence of melioidosis
Clostridium botulinum or Staphylococcus aureus strains, respectively. Finally,
case clusters was associated with the certain climate condition and geo-
we present the detection of different toxins in various sample matrices inoc-
graphical topography in southern Taiwan.
ulated with toxins applying minimal sample preparation. Furthermore, we in-
troduce the application of the electrochemical detection platform for rapid
109 identification of bacteria possessing high potential for use in biological war-
fare. After rapid PCR amplification labeled amplicons of different Bacillus sp.
Environmental Contamination of Goat Farms with Coxiella bur- are detected within ten minutes using DNA/DNA hybridization on biochips.
netii During a Q Fever Outbreak Limits of detection and specificity using several Bacillus sp. strains are pre-
G. J. Kersh, K. A. Fitzpatrick, A. V. Kondas, A. J. Roche, J. S. Self, R. A. Priest- sented. The presented results demonstrate the potential of electrochemical
ley, A. Bjork, A. D. Anderson; CDC, Atlanta, GA. biochips for sensitive and universal on-site detection of biothreat agents.
Coxiella burnetii is an obligate intracellular bacterium that cause4s the
zoonotic disease Q fever. It is considered a potential bioweapon due to its
stability, low infectious dose, and ease of transmission by inhalation. A con- 111
cern for C. burnetii is its ability to maintain infectivity and spread easily Detection of Botulinum Neurotoxin Serotypes A and B Using
through the environment carried by the wind or on animals. However, a quan-
Chemiluminescence and Electrochemiluninescene Immunoas-
titative evaluation of C. burnetii environmental spreading from a known
says in Food and Serum Matrices
source has not been reported. In the summer of 2011, an outbreak of Q fever
L. W. Cheng, L. H. Stanker; USDA, Albany, CA.
on multiple goat farms in Montana and Washington took place. Large num-
Botulinum neurotoxins (BoNT) are some of the most potent b4iological tox-
bers of organisms in the placentas of goats resulted in goat abortions and at
ins with BoNT serotypes A and B being most prevalent in foodborne contam-
least 17 cases of Q fever in humans. Here we report quantitative detection of
inations. BoNTs are also likely targets for use in intentional adulteration of
C. burnetii DNA on farms involved in the outbreak. Sponge, vacuum, and bulk
food or animal feeds and are thus classified as Select Agents. In our labora-
samples were taken, and inhibitor-free DNA isolated. Quantitative PCR tar-
tories, high-affinity monoclonal antibodies (mAbs) have been developed for
geting the IS1111 insertion sequence was performed to determine genome
the detection of BoNT/A and BoNT/B in traditional sandwich type ELISA as-
equivalents (GE) of C. burnetii in the environmental samples. Seven farms
says. To improve toxin detection sensitivity in complex matrices of food and
were included and between 14 and 138 samples were collected on each farm.
sera, we tested the use of electrochemiluminescence (ECL) type immunoas-
C. burnetii DNA could be detected at all seven of these farms, and of 390 total
says using a Meso Scale Diagnostic instrument. Both ELISA and ECL im-
samples, 274 (70.3%) were positive. Of the 274 positives, 202 (73.7% of pos-
munoassay formats detected BoNT/A and BoNT/B with better detection
itives) contained less than 100 GE per gram, but on the two farms that were
sensitivities than the “gold standard” mouse bioassays. Both immunoassays
most closely linked to human Q fever cases, 20 samples contained greater
had limits of detection of 5 pg/ml for BoNT/A and 20 pg/ml for BoNT/B in
than 10,000 GE per gram, with 5 samples from goat breeding areas having
buffer matrices, below mouse lethal dose levels. Although detection sensi-
greater than 1 million GE per gram. Further sampling on these two farms in-
tivities of ECL and ELISA platforms were similar in buffer matrices, ECL as-

BIODEFENSE PROGRAM & ABSTRACTS 77

M O N D A Y O R A L A B S T R A C T S | 3:00–4:00 p.m.
says outperformed ELISA in detection sensitivity in most liquid food matrices.
ELISA and ECL immunoassay platforms remain fast and simple alternatives to
the mouse bioassay for use in the routine detection of BoNTs.
Highlighted Oral Abstract Presentations
010. Viral Agents
Monday, February 27, 2012 | 3:00 PM - 4:00 PM
Ambassador Ballroom
112
First Evidence of Hantavirus Circulation in Terrestrial Small
Mammals from Madagascar
J-M. Heraud1, N. Razafindralambo1, T. Andrianaivo2, M-M. Olive1, V. L. Soari-
malala2, S. M. Goodman2, J-M. Reynes3; 1Inst. Pasteur, Antananarivo, Mada-
gascar, 2Association Vahatra, Antananarivo, Madagascar, 3Inst. Pasteur, Lyon,
France.
Background: Hantaviruses infect terrestrial small mammals and occasionally
humans. In human, the symptoms can be severe hemorrhagic fevers or acute
pulmonary syndromes. Previous study have shown the presence of antibod-
ies directed against Hantaan virus in R. rattus and R. norvegicus specimens.
Nevertheless, no study have shown viral circulation in small terrestrial mam-
mals of Madagascar. Material and Method: During two years, we collected
serum and organs from animals obtained in the forest of Anorana. We de-
tected hantavirus RNA using RT-PCR performed with degenerated primers.
The products of amplification of the expected size were sequences and the
sequences obtained were analyzed. Results: 586 small terrestrial mammals
from 17 different species were captured between October 2008 and March
2009. About half of the animals (n = 283) represents endemic taxa, while the
rest (n = 303) was R. rattus. We detected RNA of hantavirus and phyloge-
netic analysis of the partial sequences obtained suggested the presence of
two new hantavirus species. One was associated to the introduced species
R. rattus and an endemic rodent Eliurus majori of the family Nesomyidae.
These viral sequences were different from the sequence of Seoul virus asso-
ciated with R. rattus. The other hantavirus detected was associated to the
Tenrecidae species Oryzorictes hova and was close to sequences of viral
species associated to Soricomorpha from elsewhere in the world. Raw preva-
lence was different between endemic and introduced species (2 and 50% re-
spectively). Conclusion: Studying arenaviruses and hantaviruses are of
interest in Madagascar due to high rate of endemism (100%) among native
small mammal species. Our study allowed us to detect for the first time, two
different hantaviruses in rodents and tenrecs. These results will open new
research projects, which will address the question of infection of these
viruses in the malagasy population.
113
Distinct Differences in Disease Presentation after Aerosol
Exposure to Rift Valley Fever Virus Dependent on Inbred Rat
Strain
J. M. Bales, D. Powell, L. M. Bethel, D. Reed, I. Pandrea, A. L. Hartman; Univ.
of Pittsburgh, Pittsburgh, PA.
Background: Humans infected with RVFV generally have a self-limiting febrile
illness, but a small percentage of infected people develop complications in-
cluding acute hepatitis, fatal hemorrhagic fever, and delayed-onset en-
cephalitis. Inbred rat strains differ in their susceptibility to disease after
parenteral infection with RVFV. The objective of our study is to determine the
disease course and outcome of inbred rat strains after aerosol exposure to
Rift Valley Fever virus (RVFV) with the goal of using different inbred strains to
model the spectrum of clinical outcomes in humans. Methods: Female rats of
78 10th ASM Biodefense and Emerging Diseases Research Meeting
each strain were exposed inside a whole-body exposure chamber to aerosols
of RVFV generated with a Collison 3-jet nebulizer controlled by the AeroMP ex-
posure system. Results: After aerosol infection, Wistar-Furth rats died of a
severe hepatic disease within 4 - 6 days post-infection (LD50 = 2 pfu). High
viral loads were seen in the liver, lung, and other peripheral organs. The LD50
for RVFV in ACI rats was 124 pfu, with a survival time between 8 and 14 days
post-infection. ACI rats succumbed to neurological disease which included
signs such as rolling in cage, head tilt, circling, and abnormal behavior. High
levels of virus were detected in brain tissue but not in peripheral tissues. Un-
like what has been reported for parenteral inoculation with RVFV, Lewis rats
succumbed to neurological disease after inhalation of RVFV; the LD50 was
112 pfu and the time to death ranged from 7 - 9 days. Conclusions: WF and
ACI rats appear to be potential models for the hepatic/hemorrhagic and neu-
rological disease outcomes in humans, respectively. However, Lewis rats were
susceptible to neurological disease, which contradicts historical data for par-
enteral inoculation of RVFV. It is not clear whether this difference is a result
of the route of exposure or other factors. Additional studies are ongoing to
further characterize the disease outcome after aerosol exposure to RVFV in
all 3 strains of rat.
114
Interplay between RVFV and the Host NF-KappaB Signaling
Cascade
A. Narayanan, K. Kehn-Hall, L. Hill, S. Senina, R. VanDuyne, I. Guendel, R.
Das, A. Baer, C. Bailey, F. Kashanchi; George Mason Univ., Manassas, VA.
Background: Rift Valley fever virus (RVFV) is a category A select agent, an
emerging infectious agent and an agricultural pathogen. There are currently
no FDA approved therapeutics targeted against RVFV and understanding the
virus-host interactions will be critical to the development of novel thera-
peutics. Our studies of host phospho-signaling responses to RVFV infection
have revealed that infection causes activation of multiple signaling events
including the NFkB cascade and that inhibitors of host signaling responses
down regulate virus. Methods: We analyzed p65 phosphorylation following
RVFV infection in cells by western blots. We utilized size exclusion chro-
matography to study the nature of IKK complexes in infected cells. Using Ki-
nase assays, we demonstrate the enzymatic activity of the IKK complex. We
utilized multiple inhibitors of the NFkB pathway to determine whether they
can inhibit viral replication. We evaluated inhibitor toxicity using Cell Titer
Glo. We utilized nanoparticles with immobilized inhibitors to identify the
host targets. We performed LC-MS/MS to understand the nature of the IKK
complex in infected cells. Finally, we carried out animal experiments to
demonstrate in vivo efficacy of NFkB inhibitors. Results: Our results suggest
that the MP12 strain of RVFV causes an increase in the phosphorylation of
p65 by the classical pathway in different human cells. Our experiments point
to the existence of a novel low molecular weight IKK complex in MP12 in-
fected cells which retains kinase activity. Among the multiple NFkB inhibitors
tested, we have identified inhibitors of the IKK complex that effectively down
regulate virus and the mechanism may involve binding to IKK-β. Finally, we
show that INFAR-/- mice, when treated with NFkB inhibitors, have up to 96%
lower viral load in the liver. Conclusions: Our data suggests that RVFV in-
fection induces formation of a novel IKK complex in infected cells that is crit-
ical to RVFV replication. Inhibition of IKK down regulates virus in both in vitro
and in vivo systems.

115
Interactions of Venezuelan Equine Encephalitis Virus with Host
MicroRNA Processing Machinery
C. A. King, N. Shafagati, I. Guendel, F. Kashanchi, K. Kehn-Hall; George
Mason Univ., Manassas, VA.
Venezuelan equine encephalitis virus (VEEV) is a new-world alphavirus which
has been classified as a Category B select agent and potential bioterror
weapon, both for its severe disease course in humans and equines, and for
its ability to be transmitted by aerosol exposure. There are no current FDA li-
censed vaccines or specific therapies against VEEV, making identification of
potential therapeutic targets a priority. With this aim, our research focuses on
the interactions of VEEV with the host cell microRNA (miRNA) machinery.
MiRNA are small non-coding RNA which act as master regulators of the cell
by downregulating or degrading messenger RNA, thus suppressing produc-
tion of the resultant proteins. Recent publications implicate miRNA interac-
tion in the pathogenesis of HIV, rabies virus, HCV, and other viral diseases,
leading to the hypothesis that interactions with host miRNA machinery would
be important in a human cell model of VEEV infection. These interactions
were tested by infecting cells in which specific miRNA machinery was inacti-
vated through null mutation or siRNA knockdown. Inactivation of cytoplasmic
components Dicer and TRBP caused little to no decrease in viral replication
when tested in null-mutant cells or by siRNA knockdown, whereas siRNA
knockdown of cytoplasmic Ago-2 caused a decrease of approximately 2 log
in viral genomic copies as measured by PCR, and 1.5 log in viral replication
as measured by plaques. Inactivation of nuclear components Drosha, DGCR8
(Pasha), or Exportin-5 also resulted in significant decreases, of 3.5-5 log in
viral genomic copies as measured by PCR, and a 2-3 log decrease in viral repli-
cation as measured in plaque assay. Taken together, these findings indicate
that the nuclear processing and mRNA binding steps are the most critical for
VEEV infection, and suggest that the host cell may use redundant or alter-
nate means of cytoplasmic miRNA processing during VEEV infection. This
question presents ample opportunity for further study.
Highlighted Oral Abstract Presentations
011. Bacterial Agents
Monday, February 27, 2012 | 3:00 PM - 4:00 PM
Palladian Ballroom
116
Temperature-Regulated Membrane Remodeling is Essential for
Francisella Pathogenesis
Y. Li1, D. A. Powell1, S. A. Shaffer2, D. A. Rasko3, D. R. Goodlett2, C. R. H. Raetz4,
X. Wang5, R. K. Ernst1; 1Univ. of Maryland, Baltimore, MD, 2Univ. of Massa-
chusetts Med. Sch., Worcester, MA, 3Univ. of Maryland Sch. of Med., Balti-
more, MD, 4 Duke Univ. Med. Ctr., Durham, NC, 5State Key Lab. of Food Sci.
and Technology, Wuxi, China.
Maintenance of membrane function is essential for bacterial pathogenesis.
Adaptive mechanisms through which the Francisella membrane is remod-
eled, in response to transmission from the environment to a host requires al-
terations in the structure of the lipid A component of LPS. Francisella
synthesize lipid A with an altered distribution of fatty acids in response to
growth at environmental (21ºC) versus host (37ºC) temperatures. Mass
spectrometric analysis shows that this is due to alterations mainly at the N-
linked hydroxy fatty acids of lipid A by the acyltransferase, LpxD. In this
study, we indentified two N-acyltransferase enzymes, designed as LpxD1
and LpxD2 present in the genomes of all Francisella subspecies. To deter-
mine the role of the individual enzymes in membrane remodeling, LpxD1
and LpxD2 deletion mutants were made by allelic exchange. Complete lipid
M O N D AY O R A L A B S T R A C T S
A structural analysis showed that the LpxD1 preferentially added a 3-OH-
C18 at 37ºC, whereas the LpxD2 added a 3-OH-C16 at 21ºC. Transcriptional
analysis using microarray confirmed that the lpxD1 is up regulated at 37ºC
and lpxD2 is up regulated at 21ºC. Post-transcriptional analysis using puri-
fied enzymes in an in vitro assay showed that these enzymes have different
optimal temperature (LpxD1 at 37ºC, and LpxD2 at 21ºC). Finally, to evalu-
ate the potential role of these enzymes in pathogenesis, we tested both
deletion mutants in mice using a subcutaneous route of infection. The
LpxD1-null mutant was completely attenuated in mice and protected mice
from lethal wild-type challenge, whereas the LpxD2 was as virulent as the
wild-type organism. Thus, by restricting the bacteria’s ability to remodel its
membrane in response to temperature may represent a novel pathway for in-
tervention or vaccine development.
117
Development of a Luciferase Reporter System to Monitor
Plague Infection
Y. Sun, J. Pennington, M. G. Connor, J. M. Warawa, M. B. Lawrenz; Univ. of
Louisville, Louisville, KY.
Background: No effective plague vaccine is currently approved for use in the
US, and concerns over antibiotic resistant strains of Yersinia pestis have em-
phasized a need to develop new vaccine and therapeutic candidates. A
rapid, high-throughput plague model that allows for real time monitoring of
infection, while minimizing group sizes, would improve the early screening
steps of new anti-plague drug discovery. Methods: The lux operon from P. lu-
minescens was integrated into the Y. pestis chromosome. Three promoters
were cloned to drive the lux operon, and light output was monitored using
a luminescent plate reader (Biotek) or CCD camera (IVIS Spectrum). As con-
trols, reporters were integrated into a αphoPQ mutant (attenuated for sur-
vival in macrophages) and a αpla mutant (attenuated during animal
infection). Results: We tested three reporters during in vitro culture and in-
tracellular infection of macrophages. We observed differences in light pro-
duction for each promoter. We were also able to differentiate survival
phenotypes in the macrophage model. Using one reporter, we compared in-
fection by WT Y. pestis or an attenuated mutant in both the bubonic and
pneumonic plague models. The reporter allowed us to visualize differences
in bacterial dissemination in real time. We also used lux reporters to show
differences in colonization phenotypes between Y. pestis and B. pseudo-
mallei intranasal infections. Conclusions: Conventional animal models re-
quire groups of animals to be sacrificed at various time points to monitor
progression of disease. Here we demonstrate a lux -reporter strain can be
used to rapidly visualize differences in the progression of Y. pestis infection
in real time. Advantages of this system are: 1) smaller number of animals
per group, 2) less time required at each time point, 3) ability to follow the
course of the disease in the same animal over time, and 4) identification of
unexpected differences in disease progression. We expect that this reporter
system will allow for rapid screening of drugs during early steps of drug dis-
covery. Furthermore, this system can also be applied to virulence studies of
different Y. pestis mutants.
BIODEFENSE PROGRAM & ABSTRACTS 79
M O N D A Y O R A L A B S T R A C T S | 3:00–4:00 p.m.
118
A Variant of LcrV, the Plague Protective Antigen and Needle Cap
Protein of Yersinia pestis that Blocks Type III Secretion
K. Given1, A. Mitchell2, N. Miller1, O. Schneewind1; 1Univ. of Chicago, Argonne,
IL, 2Univ. of Chicago, Chicago, IL.
Yersinia pestis, the causative agent of plague, employs a type III secretion
machine to infect animal and human hosts. A key feature of its pCD1 viru-
lence plasmid-encoded type III machine is the transport of effector Yops into
immune cells, thereby blocking phagocytosis and with it the ability of infected
hosts to clear the invading pathogen. Type III needle complexes are com-
prised of machine components in the bacterial inner and outer membrane
that enable the transport and assembly of the YscF needle protein. Each hol-
low needle structure is capped by a ring-shaped molecule comprised of five
subunits of LcrV, the plague protective antigen. LcrV is thought to fulfill sev-
eral different dynamic functions, which include the regulation of Yop effector
secretion and the translocation of these proteins via the YopB/YopD effector
pore across the plasma membrane of host immune cells. We discovered that
the addition of more than three amino acid residues to the C-terminus of LcrV
abrogates type III secretion in response to calcium depletion even when mu-
tant lcrV is expressed in Y. pestis strains with wild-type lcrV. Using an LcrV
variant with a C-terminal Strep-tag, which is comprised of eight residues, we
observed binding to LcrG, the cytoplasmic chaperone for LcrV. A protocol was
developed that allowed the visualization of type III secretion needles from Y.
pestis. Strep-tagged LcrV was deposited at the tip of YscF needles in a man-
ner similar to wild-type LcrV. We propose that the extended variant of LcrV oc-
cludes the hollow needle, possibly by inhibiting the dynamic opening of the
LcrV lid, and abolishes its calcium sensing via the YscF protein, a prerequisite
for the transport of effector Yops into host immune cells.
80 10th ASM Biodefense and Emerging Diseases Research Meeting
119
Hide and Seek: Changes in Francisella Surface Proteins during
Infection Reveal Ideal Vaccine Targets
J. Huntley; Univ. of Toledo Coll. of Med., Toledo, OH.
Francisella tularensis is one of the most dangerous bacterial pathogens
known, due to its ease of aerosolization, low infectious dose, and ability to
cause rapid, fatal disease in humans. Identification and characterization of F.
tularensis virulence factors has been a central focus of many investigators,
yet little is actually known about how the bacterium binds to, invades, or ma-
nipulates the host cell despite years of research. We have focused our efforts
on studying F. tularensis outer membrane proteins (OMPs), given their pe-
ripheral localization and surface-exposed domains. Indeed, OMPs have been
shown to be key virulence factors for many disease-causing bacteria. Con-
versely, OMPs also have been used as potent immunogens to stimulate pro-
tection against bacterial infections. As proof of principle, we previously
demonstrated that F. tularensis outer membrane vesicles (including OMPs)
were capable of protecting animal models from Type A F. tularensis pulmonary
infection. In order to identify the most relevant OMPs to include in a future
subunit vaccine, we have performed a series of transcriptional and proteomic
analyses to identify OMPs that are specifically up-regulated during in vivo in-
fection. Surprisingly, F. tularensis dramatically modifies its OMP profile dur-
ing infection, with different OMP subsets being expressed in particular
tissues (lung, liver, or spleen) and a large number of OMPs being down-reg-
ulated during mouse infection. Further, our results demonstrated that OMP
expression in vivo varies dramatically from in vitro macrophage infections,
highlighting the fact that mammalian infections are complex processes that
cannot be recapitulated in vitro. Finally, we compared immunoproteomic
analyses with in vivo OMP expression data and found that the most im-
munogenic OMPs (those which stimulate the strongest antibody responses)
were typically down-regulated immediately following infection. Taken to-
gether, these studies provide novel information about pathogen “decoy” pro-
teins and offer a rationale strategy to develop safe and effective vaccines
against intracellular bacteria.

Highlighted Oral Abstract Presentations
019. Immune Response
Tuesday, February 28, 2012 | 3:00 PM - 4:00 PM
Diplomat Ballroom
221
Ornithine Lipids as Therapeutic Targets for Melioidosis
M. Gonzalez-Juarrero1, R. Bowen1, W. Mwangi2, T. M. Eckstein1; 1Colorado
State Univ., Fort Collins, CO, 2Texas A&M Univ., College Station, TX.
Burkholderia pseudomallei (BPS) is the etiological agent of melioidosis.
There is an increased interest in the pathogenesis of BPS following its clas-
sification as category B priority pathogen. Although antibiotics have been
proven to be effective tools in treating melioidosis, for unknown reasons they
lack efficacy during the chronic and/or recurring stages of melioidosis. In-
terestingly, BPS does not develop a rapid drug resistance that could explain
the treatment failure and it seems that BPS is capable of modifying the im-
mune system to allow its persistence. BPS contains large amounts lipids on
the surface that could serve as target or tool for additional therapeutic op-
tions. Lipids were extracted and purified from BPS by standard methods.
Polar lipids were used to stimulate caprine dendritic cells and autologous T
cells. Intracellular staining for cytokines was analyzed by Flow Cytometry.
Bacteria were also grown in U937 for various times. Lipids of intracellular
grown bacteria were extracted by standard methods. Lipids were analyzed
by mass spectrometry. We identified several polar lipids involved in the reg-
ulation of the immune response. The immune response for the in-vitro polar
lipids seems to be balanced between stimulating and repressing the immune
response. There are four major polar lipid groups and two of them (rhamno-
lipids, ornithine lipids) are strong inducer of IFNγ while the other two groups
(phospholipids, unknown group) stimulate IL-10 production. However, this
balanced immune response dramatically changes when the pathogen enters
cells. During the first hours of infection the lipid profile of BPS changes to a
more IL-10 stimulatory profile with reduced amounts of rhamnolipids and or-
nithine lipids. Since ornithine lipids are the strongest inducer of IFNγ among
polar lipids they would be excellent tools to stimulate the host immune re-
sponse during infection. Further investigation with chemically synthesized
ornithine lipids will allow for additional analyses.
222
Identification of Host Responses Contributing Towards Neu-
roinvasion and Neurovirulence by Venezuelan Equine En-
cephalitis Virus
1,2 1 1,2 3 3 3
P. Gupta , A. Sharma , M. Bhomia , J. Han , A. Yang , R. K. Puri , R. K. Ma-
heshwari1; 1Uniformed Services Univ. of Hlth.Sci., Bethesda, MD, 2Birla Inst.
of Technology and Sci., Pilani, India, 3Food and Drug Admin., Bethesda, MD.
Venezuelan equine encephalitis virus (VEEV) is an alphavirus which may
cause a bi-phasic illness in humans where primary replication in lymphoid
organs is followed by entry into the central nervous system (CNS). The CNS
phase of infection causes encephalitis and neuronal death ultimately result-
ing in mortality. Host factors which play an important role in VEEV neuroin-
vasion are poorly understood. Since the host-pathogen interactions are
reflected as specific changes in the gene-expression pattern of the host, we
compared the host responses during pathogenic and non-pathogenic VEEV
infection to identify factors involved in neuroinvasion and/or neurovirulence.
CD-1 mice were infected with either neuroinvasive V3000 or partially neu-
roinvasive V3034 or non-neuroinvasive V3014 strain of VEEV. Whole genome
microarrays were performed to obtain the differential gene expression ki-
netics in the blood and brain of infected mice. A signature gene expression
pattern specific to VEEV infection independent of the type of strains in the
T U E S D AY O R A L A B S T R A C T S
blood and brain was identified. Similarly, a gene expression signature specific
to neuroinvasive and the non-neuroinvasive strains of VEEV were identified.
Computational analysis identified pathways such as Tight junction signaling
and Granzyme-A signaling to be exclusively modulated during infection by
the neuroinvasive strains of VEEV. These gene signatures can be further ex-
plored to develop a biomarker signature against VEEV infection, help in un-
derstanding the mechanisms involved in VEEV neuroinvasion and identifying
host derived drug development targets against VEEV infection. Further stud-
ies correlating the host responses in the brain and blood using the gene ex-
pression data are in progress. This work was supported by funding from
DTRA. Opinions expressed herewith are those of the authors and do not re-
flect those of USUHS, BITS or FDA.
223
Dynamic Analysis of Respiratory Immune System Provides New
Insights on Pulmonary Anthrax
D. Fiole1,2, A. Quesnel-Hellmann1, J. D. Mathieu1, J. Douady2, J-N. Tournier1;
1
Inst. de Recherche Biomédicale des Armées, Grenoble, France, 2Univ. Joseph
Fourier Grenoble, Saint Martin d’Hères, France.
Lung efficiency as gas exchanger organ is based on the delicate balance of its
associated mucosal immune system between inflammation and sterility. Ex-
ploration of lung immune system dynamic under infection has not been per-
formed in detail so far, mainly because ventilation mechanic and chest
movement impair microscope focus. We addressed this issue by synchroniz-
ing ventilation and imaging, allowing the collection of important information
about the early stages of pulmonary anthrax pathophysiology, a severe dis-
ease due to inhalation of Bacillus anthracis spores. Mice are anesthetized
and placed under a ventilator linked to the LSM 710 Zeiss™ microscope al-
lowing two-photon imaging. Alexa chemically labelled Sterne’s strain spores
are instilled in CX3CR1+/gfp mice, which express green fluorescent protein
(GFP) mainly in dendritic cells (DCs) and monocytes. Our study shows for the
first time the infection dynamic in the lungs after inhalation of B. anthracis
spores, demonstrating that subepithelia DCs take up intraluminal spores
through trans-epithelial extensions. To our knowledge, the use of this novel
in vivo lung imaging protocol led to the first visualization of pathogen uptake
by immune cells in the lungs, demonstrating the key role of dendritic cells
acting as Trojan horses to penetrate the host defence system.
224
Visualizing Early Immune Response: Bacterial Specific Reor-
ganization at the Nanoscale
J. S. Aaron, Q. B. Smith, B. D. Carson, J. A. Timlin; Sandia Natl. Lab., Albu-
querque, NM.
One of the early events in cellular innate immune response to gram(-) bacte-
ria is the spatial reorganization of Toll-like Receptor 4 (TLR4) at the plasma
membrane following binding of bacteria-specific lipopolysaccharides (LPS)
that coat the bacterial surface. Spatial reorganization of receptors is often a
necessary component of downstream cell signaling and alhough bulk assays
have indicated that TLR4 receptors cluster upon stimulation by LPS, the
TLR4:LPS distribution has remained largely uncharacterized in intact cells
due to the mismatch between the optical diffraction limit and the nanoscale
at which reorganizations occur. Over the past decade several optical ap-
proaches have been introduced that effectively break the optical diffraction
barrier. Of these, Stochastic Optical Reconstruction Microscopy (STORM) rep-
resents an attractive method for studying early immune response because it
can provide nanometer scale resolution with specificity to the plasma mem-
brane. Using a simultaneous dual-color, STORM imaging system we imaged
the organization of TLR4 at <30 nm resolution. We will show that challenge
with LPS produces a significant increase in TLR4 clusters on the cell mem-
brane of mouse macrophages, presumably due to recruitment of receptors to
BIODEFENSE PROGRAM & ABSTRACTS 81
T U E S D A Y O R A L A B S T R A C T S | 3:00–4:00 p.m.
lipid rafts. We also exploit dual-color imaging capabilities to visualize both
the receptor and LPS simultaneously permitting TLR4:LPS complexes to be
distinguished from TLR4 without LPS. We show for the first time that LPS de-
rived from E. coli, an immuno-stimulatory bacteria, result in significantly
higher LPS:TLR4 cluster sizes in the cell membrane, as well as a nearly two-
fold greater ligand/receptor colocalization as compared to immuno-evading
LPS from Y. pestis. Additionally, preliminary results studying the effect of the
actin cytoskeleton has on TLR4 receptor reorganization will be presented.
Collectively, these results provide novel insight into the nanoscale spatial dy-
namics of TLR4 in response to bacterial-specific LPS and are important to elu-
cidate possible routes by which pathogens such as Y. pestis are able to
circumvent the innate immune response.
Highlighted Oral Abstract Presentations
020. Therapeutics
Tuesday, February 28, 2012 | 3:00 PM - 4:00 PM
Ambassador Ballroom
225
Novel Strategies to Enhance Current Antibiotic Therapies
R. V. D’Elia, T. R. Laws, A. Carter, R. A. Lukaszewsk, G. C. Clark; DSTL, Salis-
bury, United Kingdom.
Current treatment options for personnel exposed to bacterial BW agents pre-
dominantly involve the use of licensed antibiotics. These treatments work
very effectively when used shortly after exposure and, ideally, before the
symptoms of disease associated with infection begin to emerge. The timing
of the decision for military personnel to take these treatments in theatre is
therefore important to the overall effectiveness of these medical counter-
measures (MedCMs). This decision is primarily informed by a “triggering
event” such as a detector signalling the presence of a BW agent. However, if
this information is not available the first “triggering event” may be the symp-
toms associated with infection being displayed in personnel exposed to the
BW agent. Initiating treatment at this point (i.e. post-symptomatically) is
likely to be less effective, putting personnel at risk of developing a life-threat-
ening disease. We are evaluating a novel therapeutic strategy which aims to
make existing MedCMs more effective and which may make it possible to ex-
tend the timeframe (“window of opportunity”) within which antibiotics can be
administered. This project aims to establish whether the use of specific anti-
inflammatory compounds, that target fundamental immune pathways, can
be used to enhance the overall effectiveness of antibiotics by controlling the
unbridled (and ultimately futile) immune response typically found during the
course of infection with the bacterium Francisella tularensis. Anti-HMGB1 an-
tibody, which targets the nuclear protein released by cells following injury or
infection, has been assessed during in vitro and in vivo models of infection.
Preliminary data suggests that this compound may be beneficial in enhanc-
ing the effectiveness of current MedCMs in treating the disease tularaemia
caused by F. tularensis. Strategies, such as this, will be essential for the de-
velopment of a post-exposure MedCM system.
82 10th ASM Biodefense and Emerging Diseases Research Meeting
226
EV-035: A New Compound Series with Broad Spectrum Antibi-
otic Activity against Burkholderia pseudomallei and Other Se-
lect Agent Pathogens
S. Goldman1, D. Milligan1, H. Yao1, G. Liu1, G. Salerno2, P. Roussel2, J. Heim2;
1
Evolva, Inc., Palo Alto, CA, 2Evolva SA, Reinach, Switzerland.
The need for new antimicrobial compounds is increasing due to the devel-
opment of resistance in bacterial pathogens and the pathogens used as
weapons of mass destruction. The compound class described here are novel,
small synthetic molecules with antimicrobial activity against the intracellular
pathogen, Burkholderia pseudomallei, as well as several other select agents.
Evolva has developed this antibiotic series which acts against the clinically
validated target, topoisomerase IIa, with broad-spectrum antibacterial ac-
tivity and favorable drug-like properties. The EV-035 series exhibits antibac-
terial activity via specific inhibition of DNA gyrase and topoisomerase IV.
Moreover, the EV-035 series is active against many bacterial pathogen strains
that are resistant to quinolones and other bacterial topoisomerase inhibitors.
We have tested the EV-035 series for potency against B. pseudomallei in
mammalian cell intracellular infection models using B. pseudomallei and
Burkholderia thailandensis. Along with activity against Bps and Bth, we have
determined the activity of selected EV-035 molecules against other select
agents including: B. anthracis, F. tularensis, B. abortus, Y. pestis, F. tularen-
sis and B. mallei. Potency was determined by broth microdilution assays per-
formed according to CLSI guidelines. Bacterial and human topoisomerase
inhibition assays were used to quantify on and off target activities and mam-
malian cell cytotoxicity assays were used to identify compounds with suit-
able toxicity profiles. Compounds were further characterized for bacterial
cidality and spontaneous resistance rates and for drug-like properties such
as solubility, serum effects and hERG channel inhibition. The potency, selec-
tivity and excellent preclinical toxicity profile of the EV-035 series suggest
that further development of these antibiotic molecules could help fill the
need for novel, effective drugs for treatment of infections caused by biothreat
agents as well as drug-resistant pathogens in the public health arena.
227
A Single PMOplus Oligomer is Effective in a Cynomolgus
Macaque Marburg Virus Lethal Challenge Model
P. Iversen1, T. Warren2, A. Shrutleff2, J. Wells2, N. Garza2, K. Stuthman2, S. Van
Tongeren2, L. Dong2, D. Mourich3, L. Hauck1, J. Kugelman2, B. Beitzel2, G. Pala-
cios2, S. Bavari2; 1AVI BioPharma, Bothell, WV, 2USAMRIID, Fredrick, MD, 3AVI
BioPharma, Bothell, WA.
Marburgvirus (MARV) is a highly virulent RNA virus of the family Filoviridae
and a causative agent of viral hemorrhagic fever (VHF). PMOplus oligomers
targeting NP and VP24 in the Ravn virus mouse lethal challenge model re-
sulted in 65% (13/20) and 90% (18/20) survival while the combination of NP
and VP24 provided 95% (19/20) survival. The MARV Musoke guinea pig lethal
challenge model resulted in 55% (6/11) and 0% (0/11) survival for NP and
VP24, but the combination provided for 73% (8/11) survival. The current
study was to determine the contributions of oligomers targeting NP and VP24
individually versus the combination of the two in a cynomolgus
macaque(NHP) MARV lethal challenge model. Two independent and blinded
studies were conducted in which NHP were infected with a target dose of
1000 pfu of MARV Musoke administered subcutaneously. The first study in-
volved 4 groups; group 1 = 5 NHP with VP24, group 2 = 5 NHP with NP, group
3 = 5 NHP with VP24 and NP with all oligmers dosed at 7.5 mg/kg adminis-
tered daily by the iv route, and group 4 was a single NHP administered saline.
The second study involved 3 groups; group 1 = 10 NHP with NP and VP24,
group 2 = 10 NHP with NP only with all oligomers dosed at 15 mg/kg ad-
ministered daily by the iv route, and group 3 = 5 NHP administered saline.
Survival was determined at 36 (study 1) and 41 (study 2) days post infection.
 T U E S D AY O R A L A B S T R A C T S

Survival observations in study 1 included 0/5 (0%) in group 1 targeting VP24, collection of more than 700 CYP51 antifungal inhibitors that show differing
3/5 (60%) in group 2 targeting NP, 3/5 (60%) in group 3 targeting both VP24 potencies against diverse fungi including yeast, dermatophytes, molds, and
and NP, and 0/1 (0%) in group 4, the saline control. Survival observations in plant pathogens. The molecules exemplified in the table below illustrate po-
study 2 included 9/10 (90%) in group 1 targeting NP and VP24, 10/10 (100%) tential new and safer treatments for a wide range of fungal infections.
in group 2, and 0/5 (0%) in group 3, the saline control group. We conclude the Compound C. albicans T. rubrum A. fumigatus CYP3A4 Selectivity
single PMOplus oligomer (AVI-6003) targeting NP provides for dose depend- MIC1 MIC1 MIC1 IC503 Ratio4
ent protection against a MARV lethal challenge in NHP.
VT-1148 ≤0.0001 ≤0.008 0.52 18 160000
VT-1161 ≤0.0001 0.016 8 65 720000
228 VT-1546 0.008 0.031 42 >200 9125
Therapeutic Efficacy of ST-246 in Cynomolgous Macaques Itraconazole 0.016 0.062 0.52 0.07 4.3
Challenged with Monkeypox Virus via Aerosol
Posaconazole ≤0.001 ≤0.001 ≤0.0622 0.05 50
A. T. Russo, K. Agans, T. Brasel, K. Dearen, T. Bailey, S. Storch, R. Baker; LRRI,
Albuquerque, NM. 1. MICs in µg/mL. 2. Complete inhibition of growth. 3. IC50s in µM. 4. CYP3A4
Monkeypox virus (MPXV) is a biological agent that could be used as a IC50 (µM)/C. albicans MIC (µg/mL). VT-1161 is also active against Coccid-
bioweapon. It is also a useful surrogate for smallpox in studies of potential ioides immitis. VT-1161 and related molecules have excellent pharmacoki-
medical countermeasures against this agent, which is regarded as one of the netic and CNS-penetration properties. Theability for these agents to treat
most devastating pathogens of human history. The deliberate release of pathogens in neural tissues provides for a clear path to rationally design ad-
smallpox is regarded as a possible threat by the US government. Smallpox is ditional drug candiates for the treatment of the Biodefense pathogen, Coc-
highly contagious and can cause severe disease and death in humans. cidiodes. Data related to these findings will be presented.
Human trials on candidate antiviral drugs cannot be done for either of these
agents so well characterized animal models of severe orthopoxvirus disease
are needed. NHPs were treated orally once daily for 14 days with ST-246 start-
Highlighted Oral Abstract Presentations
ing 1, 2, 3, or 4 days post MPXV aerosol challenge with 1x105 PFU. Clinical ob- 021. Diagnostics and Vaccines
servations were made twice daily for up to 45 days post challenge.
Pharyngeal swab and blood analysis, clinical pathology, chest x-rays, and le- Tuesday, February 28, 2012 | 3:00 PM - 4:00 PM
sion counts were conducted at routine intervals. Tissues were collected from Palladian Ballroom
each animal at necropsy for determination of viral load and histopathologi-
cal analysis. All treated NHPs (26) survived to the end of the study while only
1 of the 4 controls survived. Severity of illness in the treated NHPs correlated
231
with the delay of treatment post challenge with NHPs starting treatment 1 WITHDRAWN
day post-challenge showing virtually no signs of infection and those starting
treatment 4 days post challenge becoming nearly as sick as controls. These
animals exhibited changes in hematology and clinical chemistry consistent
232
with but less severe than those observed in control NHPs and also showed Construction of a Lateral Flow Immunoassay for the Diagnosis
moderate weight loss and elevated respiration rates. NHPs starting treat- of Melioidosis
ment two or three days post-challenge presented signs of illness intermedi- D. Reed1, N. Chantratita2, S. J. Peacock3, M. N. Burtnick4, P. J. Brett5, S. Ray-
ate between the extremes. Delay of treatment also correlated with increased chaudhuri6, R. Houghton6, T. R. Kozel1, D. P. AuCoin1; 1Univ. of Nevada Sch. of
presence of virus in pharyngeal swabs. Results indicate that significant sur- Med., Reno, NV, 2Mahidol Univ., Bangkok, Thailand, 3Univ. of Oxford, Oxford,
vival efficacy with ST-246 administration has been demonstrated in this United Kingdom, 4Univ. of South Alabama, Reno, NV, 5Univ. of South Alabama,
aerosol MPXV model up to 4 days post challenge and that delay of treatment Mobile, AL, 6InBios Intl., Seattle, WA.
is correlated with an increase in severity and incidence of signs of disease. The goal of our research is to develop a point-of-care (POC) immunoassay
that detects secreted Burkholderia pseudomallei bacterial antigens in pa-
tient samples for early diagnosis of melioidosis. A POC immunoassay could
229 greatly impact patient outcome because a high percentage of melioidosis pa-
Rationally Designed Broad-Spectrum Antifungals Agents for tients with acute septicemia die before culture results are available. Also, the
the Treatment of Biodefense Pathogens antibiotics used for empirical treatment of septicemia are not effective for B.
E. Garvey, S. Brand, W. Hoekstra, C. Yates, W. Moore, R. Schotzinger; Viamet pseudomallei. In order to develop a POC diagnostic our first objective was to
Pharmaceuticals, Morisville, NC. identify B. pseudomallei antigens that are secreted/shed during infection.
There is an urgent unmet biodefense need to develop therapeutic agents for We employed a novel approach termed In vivo Microbial Antigen Discovery
the treatment of the deadly infection coccidioidal meningitis (CM) caused by (InMAD) to identify secreted/shed B. pseudomallei antigens. By using this
Coccidioides, a fungus that can be weaponized and used in a bioterrorist at- method we have identified a candidate polysaccharide and numerous pro-
tack. CM is 100% lethal if untreated and early intervention with current ther- tein antigens that could be targeted for diagnosis. The B. pseudomallei cap-
apy results in a 40% mortality rate. Currently available antifungals agents sular polysaccharide (CPS) was one antigen identified by InMAD and a
also are limited by inconvenient routes of administration, toxicities, and the CPS-specific mAb was produced. The mAb was able to directly detect anti-
potential for drug-drug interactions. Viamet Pharmaceuticals, Inc, discovers gen in urine from melioidosis patients by Western blot and antigen-capture
and develops ‘best-in-class’ metallo-enzyme inhibitor drugs, including those ELISA. A prototype lateral flow immunoassay (LFI) was developed in collab-
that target the validated anti-fungal target, lanosterol demethylase (CYP51). oration with our private-sector partner InBios International (Seattle, WA). The
To our knowledge, the oral clinical candidate VT-1161 is the most potent and LFI has a sensitivity of 0.2 ng/ml when purified CPS is used as an antigen
selective yeast CYP51 antifungal compound described to date. It is currently source. Preliminary studies indicate that the LFI can detect CPS in a variety of
in Phase I clinical studies in the U.S. as a treatment for either invasive can- patient samples including serum, urine, pus and respiratory secretions.
didiasis or oncychomycosis. In addition, Viamet has a proprietary chemical

BIODEFENSE PROGRAM & ABSTRACTS 83

T U E S D A Y O R A L A B S T R A C T S | 3:00–4:00 p.m.

233
Immunization with Burkholderia pseudomallei Outer Mem-
brane Vesicles Protects against Pneumonic and Septicemic Me-
lioidosis
W. Nieves1, B. Judy2, A. Torres2, C. J. Roy3, L. A. Morici1; 1Tulane Univ. Sch. of
Med., New Orleans, LA, 2Univ. of Texas Med. Branch, Galveston, TX, 3Tulane
Natl. Primate Res. Ctr., Covington, LA.
B. pseudomallei (Bps), the causative agent of melioidosis, can be naturally
acquired through inhalation, ingestion, or inoculation. Due to its threat as a
biological warfare agent, vaccine development against Bps remains a high
priority. We previously demonstrated that immunization of mice with natu-
rally-derived outer membrane vesicles (OMVs) provided significant protec-
tion against lethal aerosol challenge with Bps strain 1026b. OMVs are
constitutively shed from the surface of Gram-negative bacteria and are en-
riched with lipids, polysaccharides, and proteins. We postulated that the
multi-antigen nature of OMVs could provide protection against additional
Bps strains and routes of infection. In this work, we examined the protective
efficacy of OMVs against systemic challenge with a heterologous Bps strain.
BALB/c mice (n=10) were immunized subcutaneously with 5 μg of Bps OMVs
+/- 10 μg CpG ODN 2395 on day 0 and boosted on days 21 and 42. Control
groups received CpG only or were untreated. Serial bleeds were performed
over the course of immunization and at the time of challenge for measure-
ment of serum antibody. One month after the last immunization, mice were
challenged intraperitoneally with 40 LD50 Bps strain K96243. Survival was
monitored for 3 weeks. OMV immunization provided significant protection
against high dose systemic challenge with Bps K96243 and protection was
significantly enhanced by the addition of CpG. Control groups displayed
100% mortality by day 3, while mice that received OMV/CpG demonstrated
80% survival over the 3 week study. OMV immunization induced high titers
of OMV-specific serum IgG1 and IgG3. Addition of CpG stimulated production
of IgG2a, indicative of Th1-type immune response which could account for
the increased protective efficacy of the OMV/CpG formula. In conclusion,
OMVs represent a promising vaccine against pneumonic and septicemic Bps
infection.

234
Sortase-Conjugation Generates a Capsule Vaccine That Pro-
tects Guinea Pigs against Bacillus anthracis
O. Schneewind, G. Garufi, Y-T. Wang, S-Y. Oh, H. Maier, D. Missiakas; Univ. of
Chicago, Chicago, IL.
Capsules protect bacteria against phagocytic clearance. Capsular polysac-
charides or polyglutamates have evolved also to resist antigen presentation
by immune cells, thereby interfering with the production of opsonophago-
cytic antibodies. Linking capsular material to a carrier protein stimulates its
presentation to the immune system. For many conjugate vaccines this is
achieved by a process of random chemical cross-linking. Here we describe a
new technology, designated sortase-conjugation, which generates a single
amide bond between the C-terminal end of a carrier protein and the capsu-
lar material. Sortase-conjugation was used to link the poly-D-γ-glutamic acid
(PDGA) capsule of Bacillus anthracis to the receptor binding domain (D4) of
protective antigen (PagA). When used as a vaccine, PDGA-D4 conjugate
elicited robust antibody responses against both capsule and D4. Immuniza-
tion with PDGA-D4 afforded guinea pigs complete protection against anthrax
challenge with wild-type B. anthracis Ames.

84 10th ASM Biodefense and Emerging Diseases Research Meeting

 S U N D AY I N V I T E D S P E A K E R A B S T R A C T S
Focus Session
001. Animal Models
Sunday, February 26, 2012 | 12:00 PM - 2:00 PM
Diplomat Ballroom
001 Introduction and Setting the Stage
Louise Pitt; Center for Aerobiological Sciences, USAMRIID, Frederick, MD.
Animal studies are the cornerstone of obtaining regulatory approval for prod-
ucts to protect against biodefense agents and emerging threats. To date,
guidance from the FDA regarding these studies has been minimal and some-
times contradictory. To bring a single product to the point where it can move
forward for regulatory approval requires the cooperation of a large commu-
nity, to include government agencies, contract research organizations and
the biodefense pharmaceutical industry. Animal studies have to be designed
and executed in a manner that minimizes animal use and distress, while al-
lowing for studies to be scientifically valid and comparable across research
sites. In order for data to be comparable between laboratories, clear well-de-
fined ethical study endpoints have to be established for each infectious agent
in each appropriate animal model. This requires an early understanding, dur-
ing initial model development, of what requirements will be needed to eval-
uate efficacy of medical countermeasures.
002 Process and Implications of Animal Model Qualification
Rosemary Roberts; CDER, FDA, Silver Spring, MD.
Background: The final rule, “New Drug and Biological Drug Products; Evi-
dence needed to demonstrate effectiveness of new drugs when human effi-
cacy studies are not ethical or feasible” (21 CFR 314.600 for drugs; 21 CFR
601.90 for biological products), was published in May 2002. This regulation
is referred to as the “Animal Rule,” and provides a regulatory pathway for
the development of products for which human efficacy studies cannot be con-
ducted for reasons of ethics or feasibility. In these situations, the Food and
Drug Administration (FDA) will rely on evidence from studies in animals to
provide substantial evidence of effectiveness of these products. Methods:
Under the Animal Rule, FDA may grant marketing approval based on ade-
quate and well-controlled animal efficacy studies when the results of those
studies establish that the product is reasonably likely to produce clinical ben-
efit in humans. Developing animal models that will yield efficacy results that
can be expected to be predictive for human response is challenging. To help
animal model developers with this challenge, the Agency is developing an
Animal Model Qualification Program. Results: Product-independent animal
models can be evaluated and qualified as part of the qualification program
outlined for Drug Development Tools ( Draft Guidance for Industry: Qualifi-
cation Process for Drug Development Tools, October 2010). The initial ani-
mal model to be qualified should demonstrate the natural history of the
disease caused by the threat agent and will have a defined context of use.
After the natural history model is qualified, the model may be used for drug
development for efficacy indications. Qualified animal models will be pub-
lished in the public domain. Conclusion: The sponsor of an investigational
product can use a qualified animal model within its defined context of use
and the model will not require another review by the FDA. The use of quali-
fied models will provide economies of time, animals, and resources in a drug
development program under the Animal Rule.
005 Exploring Approaches to Consistent Veterinary Care and
Euthanasia Criteria
Jim Swearengen; NBACC, Fort Detrick, MD.
With the initial implementation of the Animal Rule in 2002, the language re-
garding marketing approval criteria for a product developed and submitted
under the Animal rule was quite stark with regard to study endpoints. The
guidance stated that animal study endpoints must be clearly related to the
desired benefit in humans, generally the enhancement of survival or preven-
tion of morbidity. Many considered the guidance to imply that most studies
conducted under the auspices of the Animal Rule would require animals to
reach a moribund state or die as a result of the agent or toxin, with no inter-
vention. In the 2009 nonbinding FDA document “Guidance for Industry: Ani-
mal Models - Essential Elements to Address Efficacy Under the Animal Rule,”
some consideration was given to a refinement in this approach where it states
that in some instances, supportive care should be administered to animals as
part of the study design. More recent discussions in regulatory circles have
begun to acknowledge that animal efficacy studies are clinical trials in which
the animal patients should be provided some basic level of supportive care
that would be expected for a human patient. While this issue can be become
quite complicated based on the type of countermeasure being developed
(e.g., vaccine versus therapeutic) and the model being used, this presenta-
tion will explore how we might consider and develop approaches that pro-
vide consistent veterinary supportive care and early endpoint (i.e.,
euthanasia) criteria under different scenarios.
006 Animal Rule Requirements vs. Expectations: How Can the
Developers of Medical Countermeasures Succeed
Elizabeth Leffel; PharmAthene, Inc., Annapolis, MD.
In 2002 the FDA finalized the “Animal Rule” which allowed for the approval
of medical products whose efficacy cannot be demonstrated in humans for
ethical reasons. To date, no new molecular entity has been licensed under
the Animal Rule despite large investments by the US government and indus-
try. Central to product approval via the Animal Rule is the development of re-
liable animal models that can be predictive of human benefit. Issues
regarding the development of animal models to support product develop-
ment will be discussed including the need for realistic understanding of the
challenges associated with heavy reliance upon government and private lab-
oratories (i.e. inter-lab variability with the models, methods, and challenge
agents) and conducting trials in high containment facilities (i.e. animal mon-
itoring and observation). Also discussed will be the need for a compelling ra-
tionale from the FDA regarding topics like added benefit and supportive care
of animals in light of these challenges; the rationale for conducting studies
of these products in special populations that will derive no benefit also needs
clarification. Finally, an attempt to identify some steps needed for the indus-
try to have a better understanding of the regulatory strategy to licensure
using the Animal Rule, or other avenues making these products available for
use will be considered (e.g. EUA).
Focus Session
002. What’s Next? Pathogen Discovery From the
Field to the Laboratory
Sunday, February 26, 2012 | 12:00 PM - 2:00 PM
Palladian Ballroom
007 Early Detection of Novel Biothreats Through International
Scientific Engagement and Medical Diplomacy
Joseph Fair; Global Viral Forecasting Initiative, San Francisco, CA.
Imperative to the future of global health is the early detection, identification,
and characterization of novel biological threats, followed by the implemen-
tation of public health measures to mitigate the pandemic potential of such
events. Historically, the global health community has distinctly segregated
human, animal and plant disease surveillance systems; however, given the
current state of their interconnectedness it is imperative that those systems
be linked. We face a unique moment in history in that recent advances in tech-
nology, including molecular diagnostics, rapid sequencing, smartphone tech-
nology, and the internet, has made it possible to create a virtual immune
system that detects, identifies, and responds to biological challenges in all
sectors. For such a system to function, there must be international buy-in
from wealthy and impoverished nations and their populations, as well as non-
governmental stakeholders - all with varying agendas, societal challenges,
BIODEFENSE PROGRAM & ABSTRACTS 85
S U N DAY I N V I T E D S P E A K E R A B ST R AC TS
and differences in health laws, use of biological materials and national de-
fense policies. While the challenges are great, they can be overcome through
human relationships and understanding that is developed by international
scientific engagement and medical diplomacy.
008 Viral Discovery Using Pan-Pathogen Microarrays and Deep
Sequencing
Charles Chiu; Univ. of California Sch. of Med., San Francisco, CA.
Traditional approaches to identify novel pathogens are limited in sensitivity
and scope. We still fail to diagnose approximately 25% of cases of respiratory
infections and more than 50% of diarrheal disease and encephalitis, and
threats from novel viruses such as the SARS coronavirus and 2009 pandemic
influenza H1N1 are continually on the horizon. New strategies are urgently
needed to increase the breadth, speed, and “throughput” of pathogen de-
tection. Here we will describe the utility of the ViroChip, a pan-viral microar-
ray, in combination with deep sequencing, in rapidly screening for the causes
of unknown outbreaks in the field. We will highlight two recent applications
of the technology— (1) identification of a novel adenovirus responsible for a
deadly pneumonia outbreak in a New World monkey colony with cross-
species transmission to a human researcher, and (2) discovery of a novel
human hemorrhagic fever virus from the Congo, Africa.
Focus Session
003. Burkholderia pseudomallei: Understanding
Pathogenesis and Immunity to Guide the Next
Generation of Medical Countermeasures
Sunday, February 26, 2012 | 2:15 PM - 4:15 PM
Palladian Ballroom
011 Melioidosis: Clinical and Therapeutic Issues
Bart J. Currie; Menzies Sch. of Hlth. Res. and Dept. of Infectious Diseases,
Royal Darwin Hosp., Casuarina, Australia.
The clinical course of melioidosis following infection is likely to be determined
by a combination of: 1. Host risk factors for disease; 2. Mode of infection; 3.
Infecting bacterial load and 4. Putative Burkholderia pseudomallei strain dif-
ferences in virulence. Diabetes is the most important host risk factor and se-
vere disease and death are very uncommon in a normal host if there is timely
diagnosis with appropriate treatment. Therefore B. pseudomallei should be
seen more as an opportunistic pathogen. It has been proposed that
aerosolization of B. pseudomallei during severe weather events results in a
shift from percutaneous inoculation to inhalation, thus accounting for more
severe disease and higher mortality. Nevertheless the occurrence, frequency
and magnitude of such aerosolization in endemic locations remain entirely
undetermined. Despite the recognition that melioidosis can be such a dra-
matically overwhelming infection, the specific virulence factors responsible
for severe disease remain surprisingly poorly elucidated, as does whether all
or only a subset of environmental B. pseudomallei are capable of causing
melioidosis. In addition to early diagnosis, the use of IV ceftazidime or car-
bapenems is the most important therapeutic factor, with longer durations of
IV therapy potentially curative without the need for a prolonged eradication
course of oral antibiotics.
86 10th ASM Biodefense and Emerging Diseases Research Meeting
012 Population Genetics and Evolution of Burkholderia
pseudomallei
David Wagner; Northern Arizona Univ., Flagstaff, AZ.
Burkholderia pseudomallei is a significant cause of mortality and morbidity
in endemic regions where it is widespread in the environment. We have found
that recombination rates in B. pseudomallei, are very high, with a recombi-
nation to mutation ratio that is higher than any other bacterial species yet
reported. The high rates of recombination (i.e. genetic plasticity) observed
within B. pseudomallei is a consequence of its lifestyle. It is a saprophytic
organism that lives in soil where it comes into contact with numerous other
species, as well as conspecifics. As a result, it exhibits incredibly high-levels
of genetic/genomic diversity at even very small spatial scales. For example,
several recent studies using multi-locus sequence typing have demonstrated
that multiple B. pseudomallei genotypes are present in individual soil sam-
ples from sites in Northeast Thailand; this high genetic diversity is known to
be associated with lateral gene transfers. This is in contrast to B. mallei, which
is actually a clone of B. pseudomallei that exhibits a clonal evolutionary struc-
ture because it has evolved to live within equine hosts and therefore rarely
comes into contact with conspecifics with which it could exchange genomic
material. Due to its high recombination rates, B. pseudomallei has what is
referred to as an “open genome”—it readily incorporates genomic material
from conspecifics and other species. As we recently reported, 10-15% of the
genome of a given strain of B. pseudomallei can be novel genomic compo-
nents not previously observed in other whole genome sequences from this
species. These novel components, found in some B. pseudomallei strains but
not others, comprise the accessory genome of this species and can be con-
trasted with the core genome, which is comprised of genomic components
found in all B. pseudomallei strains. Together, the accessory genome and the
core genome make up the pan genome— the full complement of genomic
material within this species.
013 Elucidation of Type 6 Secretion System as a Critical Viru-
lence Determinant of Burkholderia pseudomallei
David Deshazer; USAMRIID, Fort Detrick, MD.
The Burkholderia pseudomallei K96243 genome encodes six type VI secre-
tion systems (T6SSs), but little is known about the role of these systems in
the biology of B. pseudomallei. We constructed six hcp deletion mutants
(Δhcp1 through Δhcp6) and tested them for virulence in the Syrian hamster
model of infection. The 50% lethal doses (LD50s) for the Δhcp2 through Δhcp6
mutants were indistinguishable from K96243 (<10 bacteria), but the LD50
for the Δhcp1 mutant was >103 bacteria. Interestingly, similar results were
obtained using the Madagascar Hissing Cockroach as a novel alternative
host. The hcp1 deletion mutant also exhibited a growth defect in RAW 264.7
macrophages and was unable to form multinucleated giant cells in this cell
line. Unlike K96243, the Δhcp1 mutant was only weakly cytotoxic to RAW
264.7 macrophages 18 h after infection. The results suggest that the cluster
1 T6SS is essential for virulence and plays an important role in the intracel-
lular lifestyle of B. pseudomallei.
 S U N D AY I N V I T E D S P E A K E R A B S T R A C T S
014 Burkholderia pseudomallei Antibiotic Resistance Mecha-
nisms: Impact on Therapeutic Strategies
Herbert P. Schweizer; Colorado State Univ., Fort Collins, CO.
Burkholderia pseudomallei is the etiologic agent of melioidosis. The bac-
terium is an emerging pathogen and potential biothreat agent. B. pseudo-
mallei infections are difficult to treat because of the bacterium’s intrinsic
resistance and biofilm and intracellular lifestyle. Antimicrobial therapy is
biphasic and prolonged to address life-threatening acute infections, as well
as the organism’s propensity to enter into prolonged latency. This presenta-
tion will address the major antibiotic resistance mechanisms impacting ther-
apeutic strategies for the treatment of B. pseudomallei infections. Special
emphasis will be on beta-lactam resistance mechanisms and efflux pump-
mediated multidrug resistance. The discussion will address how knowledge
gained from studies of clinical isolates and laboratory-generated mutants
can be exploited to rapidly detect resistance and to redirect (clinical settings)
or initiate (intentional release) proper melioidosis therapies.
Focus Session
004. Non-Coding RNAs and the Regulation of
Virulence
Sunday, February 26, 2012 | 2:15 PM - 4:15 PM
Diplomat Ballroom
015 Small RNAs and the Evolution of Yersinia Species
Wyndham W. Lathem; Northwestern Univ., Chicago, IL.
Small, noncoding RNAs (sRNAs) are major regulators of gene expression in
bacteria, affecting protein synthesis at the post-transcriptional level. In this
session we will discuss the discovery and contribution to virulence of sRNAs
in Yersinia pseudotuberculosis, a mild gastrointestinal pathogen, and Yersinia
pestis, the causative agent of the devastating disease plague. By taking a
deep sequencing approach, we have identified 216 previously unannotated
sRNAs expressed in vitro by either Y. pseudotuberculosis or Y. pestis. While
most sRNAs are conserved between these two closely related species, the
plague pathogen lacks 6 sRNAs that are present in Y. pseudotuberculosis and
expresses 5 sRNAs that are absent from Y. pseudotuberculosis. In addition,
the expression of many sRNAs conserved between Y. pseudotuberculosis and
Y. pestis differs in both timing and dependence on Hfq, suggesting evolu-
tionary changes in post-transcriptional regulation. We will discuss how these
changes may have contributed to the dramatic difference in virulence po-
tential between the species.
016 sRNA Regulation of EHEC Virulence
Vanessa Sperandio; Univ. of Texas Southwestern Med. Ctr., Dallas, TX.
Background: Enterohemorrhagic Escherichia coli O157:H7 is a major cause of
hemorrhagic colitis and hemorrhagic uremic syndrome throughout the world.
One major virulence factor of this organism is a type three secretion system
(TTSS) encoded by the five operons of the locus of enterocyte effacement
(LEE). Two of these operons, LEE4 and LEE5, are known to be post-transcrip-
tional processed. The first gene in these operons, sepL and tir respectively,
are cleaved off from the rest of the operon. The regulation of this post-tran-
scriptional modification is largely unknown. The QseCE system is intimately
involved in the regulation of virulence in EHEC and this system regulates two
sRNAs. The role these sRNAs play in the post-transcriptional regulation of
the LEE will be investigated. Methods: Overexpression constructs for the
sRNAs were constructed. Knockouts were constructed through the use of
lambda red. Bacterial strains were grown in DMEM to an OD of 1.0 and RNA
was extracted. Northern blots were probed with RNA probes against sepL
and espA or tir and eae. rtPCR was done to measure the expression levels of
ler and stx2a. FAS was performed by infected HeLa cells with EHEC. Cells were
infected for 6hrs and stained with FITC-phalloidin and propidium iodide and
visualized with fluorescent microscopy. Results: Northern blots show that
overexpression of both sRNAs result in a decrease in RNA levels of the 3’seg-
ment of both transcripts while the first gene is unaffected. FAS revealed that
knockouts of these RNAs attach to and from pedestals on HeLa cells at lev-
els far higher than the wild type strain. rtPCR also showed that these sRNAs
have no effect on the transcript levels of ler, the master regulator of the LEE
as well as shiga toxin. Conclusion: These two sRNAs play a vital role in the
post-transcriptional regulation of LEE4 and LEE5. This level of regulation may
be important for the timing of infection.
017 Regulation of Francisella tularensis Virulence by Small
Non-Coding RNAs
John Gunn; Ohio State Univ., Columbus, OH.
Background: Francisella tularensis is a gram-negative non-motile intracellu-
lar pathogen and the causative agent of the zoonotic human disease tu-
laremia. It can survive in the environment and in a wide range of hosts,
making the ability of this pathogen to sense its environment and respond ac-
cordingly key to pathogenesis. Most of the virulence genes of F. tularensis
identified localize to the Francisella Pathogenicity Island (FPI). FPI gene ex-
pression is affected by various environmental cues and transcriptional viru-
lence regulators. The published literature suggests that MglA, SspA, PmrA,
MigR, Hfq and FevR/PigR co-operate to regulate the Francisella virulence net-
work. Methods: Because of limited protein transcriptional regulatory factors
in Francisella, we have focused on sRNA where a genome wide in silico search
identified 23 intergenic loci that encode putative sRNAs. Results: Two of the
sRNAs were predicted to target Francisella virulence regulators or FPI genes.
rprA is 55 nucleotides, located between FTT1564 and FTT1565 and predicted
to bind to 42 targeted mRNAs including pmrA and FPI genes. rfrA is 88 nt, lo-
cated between FTT0618 and FTT0619 and predicted to bind to 51 targeted
mRNAs including FevR/PigR. Over expression of these sRNAs in the human
virulent F. tularensis subsp. tularensis demonstrated a 400-500 fold growth
defect in J774.1, THP-1 and human monocyte-derived macrophages com-
pared to the wild type strain. The quantitative RT-PCR results demonstrated
that over expression of rprA down regulates several target genes including
pmrA, fevR and FPI genes while over expression of rfrA down regulated tar-
get genes including fevR. rfrA and rprA regulation of target genes is primarily
independent of the RNA chaperone, Hfq. Deletion of the sRNAs confirm the
regulation and phenotypes observed in overexpression experiments. Con-
clusion: These data suggest that rprA and rfrA regulate Francisella virulence
factors, though the exact mechanism behind this regulation is still unknown.
Future studies will focus on all 23 sRNAs and their importance in Francisella
pathogenesis.
018 Legionella pneumophila Small Regulatory RNAs and Intra-
cellular Multiplication
Sebastien P. Faucher; McGill Univ., Ste-Anne-de-Bellevue, Canada.
Background: Legionella pneumophila is a gram-negative human oppor-
tunistic pathogen and the causative agent of Legionnaires’ disease, a pul-
monary infection acquired by inhaling aerosols of contaminated water. It is
able to infect and multiply in a broad range of protozoan and macrophages.
Several regulatory proteins are known to be involved in the regulation of its
virulence factors. However, as it has become evident that small regulatory
RNAs (sRNA) play an important role in the regulation of genes involved in
pathogenesis, we wanted to test the hypothesis that sRNA also control viru-
lence-related functions in L. pneumophila. Methods: Potential sRNA genes
were predicted by searching intergenic regions for Rho-independent termi-
nators and requiring evolutionary conservation of the sequences upstream
them. A microarray was then constructed for the detection of 101 predicted
sRNAs from bacteria grown under a variety of conditions. Results: Twenty-
two predicted sRNAs were shown to be actively transcribed in at least one
condition. Two of those sRNAs, called RsmY and RsmZ, are homologs of the
CsrB sRNA that inhibits the action of the CsrA RNA-binding protein in other
bacterial species. By using complementation assays we were able to show
that RsmY and RsmZ functionally link two major response regulators of L.
BIODEFENSE PROGRAM & ABSTRACTS 87
S U N DAY I N V I T E D S P E A K E R A B ST R AC TS

pneumophila virulence factors, the sigma factor RpoS and the RNA-binding
protein CsrA. Another identified sRNA is a homolog of 6S RNA, a widely dis-
tributed prokaryotic sRNA. Deletion of 6S RNA significantly reduced L. pneu-
mophila intracellular multiplication but had no effect on growth in rich media.
By using a whole genome microarray, 6S RNA was shown to positively regu-
late expression, in stationary phase, of Icm-Dot effectors, stress response
genes such as groES as well as many genes with unknown functions. Con-
clusion: Computational prediction and microarrays were successfully used
to identify several sRNAs. RsmY and RsmZ were shown to be the missing link
between major regulators of virulence in L. pneumophila. In addition, the L.
pneumophila 6S RNA homolog was identified and shown to be involved in
the regulation of genes related to intracellular growth.

019 A Unique Intergenic RNA Thermosensor Controls Yersinia

Virulence
Petra Dersch; Helmholtz Ctr. for Infection Res., Braunschweig, Germany.
Background: Expression of all Yersinia pathogenicity factors encoded on the
virulence plasmid, including the yop effector and the ysc type III secretion
genes, is controlled by the transcriptional activator LcrF in response to tem-
perature. Methods: Gene expression analysis, structure probing and toeprint-
ing analysis were performed to characterize temperature regulation of lcrF
on the transcriptional and post-transcriptional level. Results: We found that
a protein- and RNA-dependent hierarchy of thermosensors induce LcrF syn-
thesis at body temperature. Thermally regulated transcription of lcrF is mod-
est and mediated by the thermo-sensitive modulator YmoA, which represses
transcription from a single promoter located far upstream of the yscW-lcrF
operon at moderate temperatures. The transcriptional response is comple-
mented by a second layer of temperature-control induced by a unique cis-
acting RNA element located within the intergenic region of the yscW-lcrF
transcript. This region forms a secondary structure composed of two stem-
loops at 25°C of which the second hairpin sequesters the lcrF ribosomal bind-
ing site by a stretch of four uracils. Opening of this structure was favored at
37°C and permitted ribosome binding at host body temperature. Following
oral infections in mice, we found that Y. pseudotuberculosis patient isolates
expressing stabilized thermometer variants were strongly reduced in their
ability to disseminate into the Peyer’s patches, liver and spleen and have fully
lost their lethality. Intriguingly, Yersinia strains with a destabilized version of
the thermosensor were attenuated or exhibited a similar, but not a higher
mortality. Conclusion: The LcrF RNA thermometer is relevant during infection
and is the decisive control element providing just the appropriate amounts of
LcrF protein for optimal infection efficiency.

88 10th ASM Biodefense and Emerging Diseases Research Meeting

 M O N D AY I N V I T E D S P E A K E R A B S T R A C T S
Plenary Session
007. Outbreaks, Response, and Hindsight
Monday, February 27, 2012 | 8:30 AM - 12:00 PM
Regency Ballroom
022 Swine Flu Emergence in Mexico
Edgar E. Sevilla-Reyes; Natl. Inst. of Respiratory Diseases, Mexico City, Mex-
ico.
Background: Infectious diseases have always been a threat to human popu-
lations. Followed by evidence of a number of influenza pandemics in recent
centuries, the threat caused by the emergence of the highly pathogenic H5N1
viruses generated a number of actions around the world: development of pre-
vention, control and recovery programs. Situation: The surprising 2009 out-
break in Mexico and the pandemic that followed was closely followed and
documented by the media, the scientific community and health systems. In
this presentation I will review a number of actions and programs already in
place by the time of the H1N1 outbreak and the difficulties found during im-
plementation at multiple levels: in hospitals and small clinics, education cen-
ters and public places, but also at the epidemiological surveillance level. The
Mexican population was receptive and responded positively to most of these
actions. Outcome: The H1N1 health response in Mexico followed Interna-
tional Health Regulations along other preparedness and response programs.
A number of useful lessons were learned during the pandemic, and further-
more, multiple economic, human and material resources were put in place
all around the country that can be reapplied in the future. One of the main
challenges today is not to forget those lessons and to continue building
stronger prevention, detection and control networks despite economic and
political swings.
023 Swine Flu in the United Kingdom
Gail Thomson; Hlth. Protection Agency, Salisbury, United Kingdom.
During the 2009 pandemic the clinical response took on an additional mode
of communication and coordination - the virtual network. This framework
been utilised during the SARS outbreak and had served the frontline well in
2003. The Health Protection Agency’s (HPA) network of Intensivists caring for
severe cases of H1N1 ran for six months and then again in winter 2010. With
weekly calls the UK intensive care community had an open line of communi-
cation and a forum to discuss with world influenza experts, public health and
to discuss challenging case management issues. Clinical colleagues from
across the globe contributed to the discussions. The outputs included guid-
ance notes for the care of critically ill adults and children. During the pres-
entation consideration will be given to how this model could be built upon for
future events.
024 Overview of E. coli Outbreaks in the USA
Robert Tauxe; CDC, Atlanta, GA.
Since 1982, when Escherichia coli O157:H7 first emerged as a foodborne
pathogen, the Shiga toxin producing E. coli (STEC) have driven major changes
in public health surveillance and food safety. Pulsenet, the national molecu-
lar subtyping network housed at CDC, includes public health and food regu-
latory laboratories across the country, and has greatly improved detection,
investigation and control of bacterial enteric outbreaks by CDC, state and
local public health authorities. They have been linked to contaminated foods,
and to water, contact with animals and spread in daycare-centers. The dom-
inant pathway from the ruminant reservoir has been through beef. While the
number of outbreaks reported has increased, their size has dropped, and the
incidence of STEC O157 infections had dropped by 44%, as prevention meas-
ures driven by outbreak findings have been implemented, particularly for
ground beef. In recent years, outbreaks related to produce (including apple
cider, sprouts, and leafy greens) have also driven prevention efforts; further
measures are needed to reach the national target of 0.6 per 100,000 for STEC
O157. Use of Shiga toxin-based screening tests facilitates the diagnosis of in-
fection with STEC serotypes other than O157, now reported more frequently
than O157, but about which less is known. Such culture independent tests
need to be coupled to reflex culture and subtyping in PulseNet labs, or ulti-
mately to advanced methods that provide the same subtyping and virulence
information critical to public health, or our ability to detect, investigate and
ultimately to prevent these outbreaks and infections will rapidly erode.
025 E.coli O104:H4: The Outbreak Experience from Germany
Reinhard F. Burger; Robert Koch Inst., Berlin, Germany.
Germany experienced in 2011 one of the largest reported EHEC/HUS out-
breaks worldwide with almost 3,000 EHEC and 855 HUS cases and 53 deaths.
Evidence for the role of raw vegetables as vehicle of the infection was rapidly
obtained. The pathogen was identified as EHEC O104:H4. Over 30 cohorts
including a recipe-based restaurant cohort study revealed sprouts as vehicle.
The food safety agencies (BfR, BVL) applied forward- and backward-tracing
strategies. 40 outbreak clusters were linked to a farm producing sprouts in
Lower Saxony using fenugreek seed from Egypt. The rare serotype O104:H4
produced Shigatoxin 2 and had virulence characteristics of enteroaggregative
E.coli indicating a virulence combination of two different pathogens. Sec-
ondary infections in household partners, hospitals, and via food distribution
chain occurred, emphasising the importance of hygiene.
026 Molecular Epidemiology and Characterization of the E.coli
O104:H4 Outbreak Strain
Alexander Mellmann; Univ. Hosp. Muenster, Muenster, Germany.
Background: In the past early summer, we were confronted with the largest
outbreak of hemolytic uremic syndrome ever caused by an exceptionally vir-
ulent Shiga toxin (Stx)-producing Escherichia coli O104:H4. Previously, this
serotype has rarely been associated with HUS. Methods: This presentation
will focus on the unique characteristics of this strain by comparing the out-
break isolates with other O104:H4 strains on the genome level. Results: In-
terestingly, the current outbreak strain and the historical isolates associated
with HUS carried genes typically found in two types of pathogenic E. coli, en-
teroaggregative E. coli (EAEC) and enterohemorrhagic E. coli (EHEC). Phylo-
genetic analyses of the core genome genes indicated that the HUS-causing
O104:H4 strains and the previously published sequence of the EAEC strain
55989 showed a close relationship but were only distantly related to com-
mon EHEC serotypes like O157:H7 or O26:H11. Though closely related, the
outbreak strain differed from the historical strains in plasmid content and
fimbrial genes. Conclusion: The data determined enabled an evolutionary
model in which EAEC 55989 and EHEC O104:H4 strains evolved from a com-
mon EHEC O104:H4 progenitor, and suggest that by stepwise gain and loss
of chromosomal and plasmid-encoded virulence factors, a highly pathogenic
hybrid of EAEC and EHEC emerged as the current outbreak clone.
Symposium
012. Biodefense at the Dinner Table: Emerging
Foodborne Illnesses
Monday, February 27, 2012 | 4:15 PM - 6:15 PM
Diplomat Ballroom
120 Mechanisms of Murine Norovirus Interaction with the In-
testinal Epithelium
Christiane Wobus; Univ. of Michigan Med. Sch., Ann Arbor, MI.
Murine norovirus (MNV) is an enteric virus that replicates in the intestine of
its host. It has a tropism for murine macrophages and dendritic cells both in
culture and in mice. However, it does not infect intestinal epithelial cells, rais-
ing the question how MNV crosses the epithelial barrier to reach its target
cells. To investigate the initial steps of MNV penetration of the intestinal mu-
cosa, we are using two approaches; an in vitro M cell model of differentiated
epithelial monolayers and in vivo studies in mice. For in vitro studies, mICcl2
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MO N DAY I N V I T E D S P E A K E R A B ST R AC TS
cells are plated into transwells for differentiation and some are also co-cul-
tured with murine B cells to promote M cell development. MNV was able to
translocate across the epithelial layer without disrupting tight junctions or
virus replication. Consistent with a role for M cells in viral transcytosis, signif-
icantly more MNV reached the basolateral compartment under co-culture con-
ditions, and MNV-1 was localized in M-like cells by confocal microscopy. To
study early time points during MNV pathogenesis and determine whether de-
tected virus is replicating or input virus, we have adapted the use of light-sen-
sitive, neutral red (NR)-containing virus for in vivo studies. The basic premise
of this technology is that NR-labeled virions are inactivated upon exposure to
light, whereas virions that have undergone replication are no longer labeled
with NR and remain viable. Following oral infection of Balb/c mice with either
MNV-1 or CR3, only input virus was detected in the small or large intestine at
3 hrs postinfection (hpi), while by 24 hpi only replicated virus was detected.
To determine whether M cells are critical for MNV infection in vivo, we condi-
tionally depleted mice of M cells using an antibody against RANKL (receptor
activator of NF-kB ligand), which is essential for the production of M cells, and
orally infected mice with neutral red containing MNV-1 and CR3. MNV-1 and
CR-3 replication in the intestine was significantly reduced in RANKL-depleted
compared to isotype-depleted or untreated mice. Taken together our data
demonstrate that M cells mediate MNV transport across the intestinal ep-
ithelium and are required for MNV pathogenesis in mice.
121 Picking the Lock: Stealth Invasion of Epithelial Barriers by
Enteric Pathogens
Manuel R. Amieva; Stanford Univ. Sch. of Med., Stanford, CA.
A number of bacterial pathogens can cross the gastrointestinal epithelium
to cause inflammatory gastroenteritis, enteric fever or sepsis. The receptors
for epithelial cell invasion have been defined for several of these pathogens,
and include adhesion molecules like E-cadherin for Listeria monocytogenes,
integrins for Yersinia enterocolitica and Shigella sp. Of note is that many of
these receptors for entry are not found on the luminal side of the intestine,
but are instead abundant components of the basolateral membranes of ep-
ithelial cells. This implies that pathogens first breach the epithelial tight junc-
tions, or find alternative mechanisms to cross the epithelial barrier before
reaching their receptors for epithelial invasion. We will discuss a novel model
of intestinal invasion by bacterial pathogens. We discovered that the tips of
intestinal villi are a specialized site of microbial invasion where the normal
process of cell senescence and cell extrusion allows basolateral receptors to
be exposed at the luminal surface.
122 Multi-Dimensional Analysis of E. coli Colonization of Mam-
malian Cells and Green Leafy Vegetables
Jorge A. Giron; Univ. of Florida, Gainesville, FL.
The several classes of diarrheagenic Escherichia coli continue to represent a
significant health burden in developed and developing countries, particularly
in the pediatric population. In the United States and other industrialized
countries, Enterohemorrhagic or Shiga-toxigenic E. coli (EHEC or STEC)
strains of several serotypes are associated with serious intestinal and kid-
ney illness due to the consumption of tainted beef and agricultural products.
The alarming outbreaks of bloody diarrhea and hemolytic uremic syndrome
caused in the Summer of 2011 in Northern Europe by a new enteroaggrega-
tive E. coli (EAEC) strain of O104:H4 serotype producing the Shiga toxin high-
light, undoubtedly, the significance of pathogenic E. coli and the plasticity of
their genomes that allows the emergence of variants with novel and high in-
fective and disease causing capabilities, as well as the need to better un-
derstand the function of putative virulence factors present in the E. coli
pangenome against the possibility that similar outbreaks may occur anytime
elsewhere in the world. The strategies that EHEC O157:H7 utilizes to colo-
nize human, animal and epithelial will be discussed. The dissection of the
mechanisms that these bacteria employ to interact with their host cells will
help better understand the role of the several pili and other products in the
different E. coli lifestyles, outside and within the host, and establish a ground
90 10th ASM Biodefense and Emerging Diseases Research Meeting
for proposing novel strategies towards prevention and treatment of diarrheal
illnesses that overwhelmingly inflict the health of the world’s population.
Symposium
013. Persistence and Inactivation of Biothreat
Agents and Emerging Pathogens
Monday, February 27, 2012 | 4:15 PM - 6:15 PM
Palladian Ballroom
124 Persistence of Category A Select Agents
Chuck Gerba; Univ. of Arizona, Tucson, AZ.
Fomites (intimate objects) contaminated with biothreat agents can play an
important role in the transmission of respiratory, enteric and dermal infec-
tions that is often not recognized. Biothreat agents can survive for days to
months on common fomites depending on the type of organism (bacteria,
virus, spore), fomite (stainless steel, paper, ceramic, glass, cloth), suspend-
ing media (water, mucus, feces, soluble organics) and relative humidity. Die-
off is usually biphasic with greater die-off during drying of suspending media.
Information on inactivation of biothreat agents is essential in the develop-
ment of models for predicting exposure and the potential success of inter-
ventions in indoor environments.
125 Persistence and Inactivation of Vaccinia Virus
Joseph Wood; EPA, Research Triangle Park, NC.
Background: Variola major is a CDC Category A biological agent, and is the
etiological microorganism for smallpox disease. Since the variola virus is un-
available for research, we used the vaccinia virus (lyophilized) as a surrogate.
Methods: Tests were conducted to determine how long the vaccinia virus
would remain infectious in the environment, as a function of material, tem-
perature, and relative humidity. Tests were also conducted to assess the ef-
ficacy of two common disinfectant chemicals (1% citric acid and quaternary
ammonium) in inactivating the virus. Approximately 107 plaque forming units
(using Vero cells) per coupon material were used in the persistence and dis-
infection tests. Results: The virus generally remained viable longer under low
RH conditions, and after 56 days at low temperature and humidity, more than
104 PFU were detected on 3 of the 4 materials tested. In the inactivation tests,
neither of the disinfectants reduced the virus to non-detectable levels. Con-
clusion: The data presented elucidates some conditions when natural atten-
uation or common disinfectants may inactivate vaccinia and related viral
agents.
126 Inactivation of Bacterial Biothreat Agents in Water
Laura Rose; CDC, Atlanta, GA.
Concern for the security of drinking water supplies has prompted questions
about the survival of biothreat bacteria in water and the efficacy of current
disinfection practices to control contamination. We investigated the survival
of six species of non-spore forming biothreat bacteria (Burkholderia pseudo-
mallei, Burkholderia mallei, Brucella suis, Brucella melitensis, Yersinia pestis,
and Francisella tularensis) in water without disinfectants. B. suis and Y. pestis
were culturable for up to 21 days, and B. pseudomallei was culturable up to
30 days. All were metabolically active, as determined by esterase activity,
after 30 days. The non-spore forming bacteria and Bacillus anthracis spores
were tested for susceptibility to free available chlorine (FAC, ≤ 1 - 4.8 mg/L),
monochloramine (1 or 2 mg/L), chlorine dioxide (0.25 or 2.0 mg/L) and ul-
traviolet light (254 nm). The non-spore forming bacteria were susceptible to
all disinfectants tested within reasonable contact times, with some disinfec-
tants more effective than others. B. anthracis spores were more resistant than
the non-spore forming bacteria to all disinfectants tested, with chlorine diox-
ide more efficacious for biofilm associated spores and FAC more efficacious
for planktonic spores.
 M O N D AY I N V I T E D S P E A K E R A B S T R A C T S
127 Decontamination of Bacillus Spores from Water
Using Germinant
Jeff Szabo; USEPA/NHSRC, Cincinnati, OH.
Background: Bacterial spores are persistent on drinking water infrastructure
and common decontamination methods such as flushing and chlorination
have had limited success. Germination was evaluated as an enhancement to
disinfection of Bacillus spores from drinking water infrastructure with free
chlorine and flushing. Methods: A pilot scale pipe loop was outfitted with
iron (corroded) and cement-mortar coupons, which were conditioned in tap
water for one month. Bacillus globigii spores were injected into the loop and
allowed to adhere for 2 hours. Germinant was added after the adhesion
phase, and allowed to contact the spores for an additional 2 hours. Germinant
was flushed out of the loop, and chlorination followed by flushing was per-
formed. Experiments only using chlorination and flushing were also per-
formed to determine the effectiveness of the germinant. Results:
Decontamination with free chlorine at 5 mg/L was ineffective (~0.2 log re-
moval) on iron and achieved a 1.8-log reduction on cement mortar. Increas-
ing free chlorine concentration to 25 mg/L resulted in 1.2 and 2.2-log
reductions of spores on iron and cement-mortar, respectively. Flushing after
disinfection provided additional reduction, but spores persisted in each case
except cement-mortar decontaminated with 25 mg/L, where they dropped
to undetectable levels. Adding a germinant (trypic soy broth) alone decreased
the number of spores adhered cement-mortar and iron by 1.1 and 1.4 log,
respectively. Chlorination after germination at 5 mg/L further reduced spores
attached to cement-mortar to undetectable levels. Spores were reduced to
undetectable levels on iron coupons by chlorinating at 5 mg/L and then flush-
ing (increasing shear) after germination. Application of chlorine dioxide may
also be an effective method of decontamiantion. Conclusion: This study
shows that germinating spores before application of disinfectant or flushing
is an effective way to decontaminate drinking water infrastructure.
Symposium
014. Taking the Bite Out of Insect Bites: Biodefense
Implications of Vector-Borne Infections
Monday, February 27, 2012 | 4:15 PM - 6:15 PM
Ambassador Ballroom
129 Dissecting the Mechanisms of Tick Transmission from
Anaplasma to Francisella
Susan M. Noh; USDA-ARS Animal Disease Res. Unit, Pullman, WA.
For ongoing transmission, vector-borne pathogens must make the remark-
able transition from the mammalian host to the vector. In the case of intra-
cellular pathogens, little is known about the molecular requirements of this
transition. To identify these mechanisms, the transcriptional and proteomic
profiles of Ehrlichia spp. and Anaplasma spp. have been studied and shown
to be host cell-type dependent. For example, in A. marginale many surface
proteins are down-regulated in tick cells, while approximately 15 proteins,
including one surface protein are specifically up-regulated. In A. marginale
and A. phagocytophilum a majority of genes or gene products which are up-
regulated in the tick are hypothetical, thus function cannot be inferred. Ad-
ditionally genetic manipulation of these pathogens is difficult, further limiting
functional characterization. Because Francisella sp. are tick transmissible,
we used Francisella novicida, which is amenable to genetic manipulation, to
develop a tick transmission model to directly interrogate gene function at the
vector-pathogen interface with the long-term goal of identifying convergent
and divergent mechanisms of tick transmission.
130 Characterizing 24-48h Tick Saliva Proteins to Understand
Early Stage Tick Feeding Biology
Albert Mulenga; Texas A&M Univ., College Station, TX.
Through out the feeding process ticks differentially inject proteins to regulate
feeding, acquisition and transmission of disease agents. In our research we
are interested in discovery and characterization of tick saliva that are injected
into the host during the first 24-48h of the tick feeding process. To do this
we have generated antibodies to 24-48h tick saliva proteins. We are using
these antibodies to mine for cDNAs that encode tick saliva proteins. Here we
have described characterization of an apparent universally conserved tick
anticoagulant protein present in Amblyomma americanum and Ixodes scapu-
laris. The role of these proteins and others in tick feeding regulation and con-
sequently tick borne disease agent transmission is discussed
131 Tweets from Ticks: Modulation of Vector-Specific Gene Ex-
pression by BadR in the Agent of Lyme Disease
Janakiram Seshu; Univ. of Texas, San Antonio, TX.
Borrelia burgdorferi, the agent of Lyme disease, is intimately dependent on
its arthropod and vertebrate hosts for a variety of nutrients as it lacks several
metabolic pathways. While there is a large body of information on how B.
burgdorferi adapts to vertebrate host-specific conditions following the bite of
an infected tick, there is a dearth of information on how it adapts to arthro-
pod host when uninfected ticks feed on infected vertebrate hosts. Since reg-
ulators of carbon and nutrient utilization facilitate virulence and adaptive
gene expression, we analyzed the role of BB0693, annotated as xylose
operon regulatory protein (XylR1) in the patho-physiology of B. burgdorferi.
XylR1 shares homology with two other ROK family repressors, NagC involved
in regulating N-acetylglucosamine (GlcNAc) utilization and Mlc, a global reg-
ulator of glucose metabolism. Immunoblot analysis indicated increased ex-
pression of XylR1 in B. burgdorferi lysates grown under conditions mimicking
arthropod hosts compared to that of vertebrate hosts. A xylR1 deficient mu-
tant generated in the infectious B31A3 strain of B. burgdorferi exhibited in-
creased levels of pathogenesis related proteins synthesized under vertebrae
host-specific conditions, such as RpoS and OspC, suggesting that increased
levels of this regulator under tick-specific conditions could repress vertebrate
host-specific gene expression. Microarray analysis and quantitative real-time
reverse transcription-PCR analysis further confirmed drastic increases in the
expression of these vertebrate host-specific determinants in the xylR1-defi-
cient strain. Based on these findings, we have designated this regulator as
Borrelia host-adaptation Regulator (BadR). Collectively, the badR mutant has
uncovered another layer of regulation in this spirochetal pathogen, and has
demonstrated BadR’s role as a putative carbon regulatory protein involved in
sensing the host-specific environment as well as modulating gene expres-
sion necessary for survival in the ticks or vertebrate hosts. Further analysis
of BadR mutant strain will facilitate identification potential targets to reduce
the incidence of Lyme disease.
BIODEFENSE PROGRAM & ABSTRACTS 91
T U E S DAY I N V I T E D S P E A K E R A B ST R AC TS
Plenary Session
017. Targeting Virulence Factors in Acute Infection
Tuesday, February 28, 2012 | 8:30 AM - 12:00 PM
Regency Ballroom
135 Antimicrobial Peptide Modulation of Chemokine and Pro-
Inflammatory Cytokine Responses: It’s All in the Timing
Kim A. Brogden; Univ. of Iowa, Iowa City, IA.
Background: B. anthracis produces lethal toxin (LeTx) composed of lethal fac-
tor, a metalloprotease that cleaves mitogen activated protein kinase (MAPK)
kinases (MKKs) and protective antigen, a carrier. Through cleavage of MKKs,
LeTx suppresses early cytokine responses by dendritic cells. However, recent
studies show that suppression of cytokine production is not uniformly ob-
served suggesting that methods to enhance cytokine production may reverse
the early events of immune suppression. Methods: Recently, we assessed
the chemokine and cytokine response of human myeloid dendritic cells to
Porphyromonas gingivalis hemagglutinin B (HagB) and human beta defensin
3 (HBD3) mixtures. Results: HagB administered alone induces a robust
chemokine and cytokine response. HagB administered with HBD3 does not.
Interestingly, HBD3 administered 1 hour prior or 1 hour after HagB, results in
an early, robust, and sustained chemokine and cytokine response to HagB
(Figure 1 example). Conclusion: It is tempting to speculate that a mechanism
involving HBD3 could ‘arrest’ the early events of LeTx immune suppression
and ‘normalize’ the host immune response to the presence of B. anthracis
toxins thus facilitating normal clearance and subsequent adaptive immune
responses to B. anthracis components.
136 Anthrax Diagnostic Tools (Nonclinical and Clinical) for
Medical Countermeasure Development Under the Animal Rule
Elizabeth Leffel; PharmAthene, Inc., Annapolis, MD.
The process of identifying a virulence factor or simply a marker of infection,
followed by developing an assay to detect such a biomarker can be difficult.
When developing a therapeutic drug under the FDA Animal Rule, efficacy has
to be tested in animals infected with the pathogen of interest. Care must be
taken to ensure that animals have active infection so that treatment is not
prematurely initiated. For this reason, the successful identification of bio-
markers and the ability to test for them becomes critical. This discussion will
focus on an assay to detect a biomarker of Bacillus anthracis. A historical
overview of development of the electrochemiluminesence (ECL) assay to de-
tect protective antigen (PA) and how it was transitioned from the bench to
use as a diagnostic assay in regulated (21CFR58) nonclinical studies will be
presented. The ECL-PA was used to confirm infection in appropriate animal
models designed to evaluate the efficacy of an anti-toxin monoclonal anti-
92 10th ASM Biodefense and Emerging Diseases Research Meeting
body (mAb) intended for treatment of inhalation anthrax. The mAb is highly
efficacious and resulted in no adverse effects in aerosol challenge or toxi-
cology studies. The ability to correlate the predictive value of nonclinical mod-
els to help ascertain potential safety risks in clinical research will be
discussed. It is anticipated that because the ECL-PA assay has been devel-
oped and tested in numerous nonclinical studies, it could be appropriately
applied in a clinical setting in the event of an ‘anthrax attack’ and this theory
will be discussed.
138 Targeting Cyclic Di-GMP Signaling to Prevent Biofilm
Formation
Chris Waters; Michigan State Univ., East Lansing, MI.
Background: Biofilms are multicellular communities of bacteria encased in a
network of protective extracellular polymers. In this state, bacteria are re-
sistant to environmental stress, predation, immune clearance, and anti-mi-
crobial treatment. Cyclic di-GMP is a nearly ubiquitous second messenger in
bacteria that induces biofilm formation and represents an attractive target
for the development of new anti-biofilm strategies. Methods: A high-through-
put small molecule screen utilizing a cyclic di-GMP inducible luciferase tran-
scriptional reporter construct in Vibrio cholerae was developed. 66,000
compounds/extracts from the University of Michigan Center for Chemical Ge-
nomics chemical library were screened, and 184 hits possessing IC50 values
less than 10 micromolar were identified. The ability of these compounds to
inhibit biofilm formation and c-di-GMP synthesis both in vivo and in vitro was
determined. Results: Two classes of compounds that possess broad-spec-
trum anti-biofilm activity were identified. One class of compounds, repre-
sented by the molecule we have designated anti-biofilm compound 1 (ABC-1)
inhibits biofilms of multiple bacterial pathogens including, but not limited to,
Vibrio cholerae, a cystic fibrosis isolate of Pseudomonas aeruginosa, and
MRSA Staphylococcus aureus. The mechanism of action of ABC-1 is currently
under investigation. Moreover, we have identified 7 lead compounds that di-
rectly inhibit in vitro c-di-GMP synthesis by GGDEF enzymes. These com-
pounds also inhibit biofilm formation of V. cholerae. Two of these compounds
reduce total intracellular c-di-GMP. Conclusions: This research has identified
multiple small molecules that disrupt biofilm formation in a broad spectrum
manner, including the molecules that directly inhibit c-di-GMP synthesis by
GGDEF enzymes. These molecules have potential for the treatment of biofilm
based infections, and we are currently examining their in vivo efficacy in rel-
evant biofilm based disease models.
139 Identification of Inhibitors of DNA Adenine Methylation as
Novel Therapeutics
Petra Oyston; DSTL, Salisbury, United Kingdom.
Multiply antibiotic resistant strains of plague are emerging, driving a need
for the development of novel antibiotics effective against Yersinia pestis. DNA
adenine methylation (Dam) regulates numerous fundamental processes in
bacterial cells and alteration of Dam expression is attenuating for several
pathogens, including Y. pestis. The lack of a functionally similar enzyme in
humans makes Dam a suitable target for development of novel therapeutics
for plague. We have identified a novel class of potent Dam inhibitors. The
most active compound was shown to have a competitive mode of inhibition
with respect to DNA and a Ki of 6.46 nM. Moving from activity in vitro against
purified protein to activity against the bacterium requires inhibitors capable
of penetrating into cells, accumulating to a sufficient level intracellularly, and
interacting with the target proteins to achieve an effect on the physiology of
the cell. One compound was found to meet these criteria, as indicated by in-
hibition of DNA methylation in Yersinia cultures. The effects of this inhibition
on the physiology of the cell were widespread, and included altered expres-
sion of known virulence traits, including iron acquisition and Type III secre-
tion. suggesting these inhibitors may attenuate bacterial virulence and
function as antibiotics.
 T U E S D AY I N V I T E D S P E A K E R A B S T R A C T S

Symposium 237 Host Genetic Resistance to Gram-Negative Bacterial Infec-

tions: The Case of Yersinia pestis
022. Host Genetic Resistance to Infections Christian E. Demeure; Inst. Pasteur, Paris, France.
While laboratory mice are usually highly susceptible to Yersinia pestis, the
Tuesday, February 28, 2012 | 4:15 PM - 6:15 PM
SEG mouse exhibit an exceptional capacity to resist bubonic plague. Genetic
Diplomat Ballroom and physiopathologic comparisons with C57BL/6 mice were performed to
235 Host Genetic Resistance to Parasitic Infections: The Case identify genetic loci and immune mechanisms associated with resistance.
Strikingly, bacterial invasion in SEG occurred earlier and transiently compared
of Malaria
to C57BL/6 mice, which bacterial load was delayed but continuous until
Simon J. Foote; Macquarie Univ., Sydney, Australia.
death. Characterisation of cellular populations and cytokine production dy-
Malaria is a parasitic disease that kills around 1 million women and children
namics in various tissues revealed that the production of five cytokines (G-
annually. There is increasing resistance to current antimalarials with the time
CSF, KC, MCP-1, IL-1α and IL-6) involved in monocyte and neutrophil
between introduction of a new drug and the development of resistance de-
recruitment was significantly correlated with the early bacterial load in all or-
clining with each new addition. However there are natural, genetic resistance
gans. Indeed, an early massive macrophage recruitment in SEG spleen was
mechanisms to malaria. These usually focus on the red cell and are ubiqui-
observed. SEG and B6 macrophages were further compared to define the role
tous in areas of malaria endemicity. We are studying the host response to
of such cells in the early and effective innate immune response necessary for
disease in order to identify host targets for therapeutic intervention. We an-
surviving plague.
ticipate that host targets will provide antimalarials that are steadfast against
the development of resistance. We use several approaches to identify host 238 Host Genetic Resistance to Viral Infections: The Case of HIV
targets, these include a bioinformatics approach, an ENU mutagenesis screen Florencia Pereyra; Brigham and Women’s Hosp. and the Ragon Inst. of MGH,
and the development of a program looking at the role platelets play in a Boston, MA.
malarial infection. The results of some of these programs will be presented Genetic studies of infectious and inflammatory diseases have repeatedly
in this talk. Our ENU mutagenesis program screens offspring of ENU-treated shown strong associations within the MHC region of human chromosome 6.
malaria-susceptible mice. Mice surviving a murine model infection are prog- However, the basis for these associations remains elusive, in part because ex-
eny-tested and exons sequenced looking for segregating mutations. We have tensive linkage disequilibrium precludes assignment of causal variants. To
identified a number of mutations and several mutation affecting red cell cy- further define host genetic effects on the outcome of a chronic viral infec-
toskeletal proteins will be discussed. Bioinformatic analysis has identified tion, we performed genome-wide association analysis in a multiethnic co-
the heme-synthetic pathway as providing potential targets. We will present hort of HIV-1 infected persons, and coupled this to a novel imputation
data examining the effect of inhibition on two enzymes within this pathway. method to analyze the effects of individual amino acids within the classical
HLA proteins. Comparing 974 HIV-1 controllers, who maintain low virus loads
236 Host Genetic Resistance to Mycobacterial Infections: The
in the absence of therapy, to 2,648 persons with high virus loads and pro-
Case of Mycobacterium tuberculosis
gressive disease, we identified &gt;300 variants within the HLA region of
Philippe Gros; McGill Univ., Montreal, Canada.
chromosome 6 that reach genome-wide significance (P &lt; 5 x 10-8), and
Early response to infectious agents involves the concerted action of various
none elsewhere. Subsequent association analysis of amino acids within the
cell surface receptors and associated signaling pathways, and the rapid re-
HLA proteins pinpointed six independently significant residues in class I, five
sponse of different cell types such as NK cells, macrophages, dendritic cells,
lining the peptide binding groove, that explain the SNP associations in the
and T lymphocytes. These cells secrete regulatory cytokines and chemokines
GWAS and reconcile both protective and risk HLA associations with control of
that initiate a very rapid “inflammatory” response, direct early Th1 polariza-
HIV-1. These results implicate the nature of the peptide-HLA class I interac-
tion of the immune response, ultimately leading to destruction of the invad-
tion as the major genetic factor modulating control of HIV infection.
ing agent and establishment of protective immunity. IRF8 is a transcription
factor that plays a key role in all aspects of this response, both with respect
to ontogeny of different myeloid lineages, activation of host defenses in these
cells, including production of IL-12. Mice bearing mutations in IRF8 or its het-
erodimerization partner IRF1 are extremely susceptible to mycobacterial in-
fections, including pulmonary tuberculosis. Mutation in human IRF8 cause a
novel type of immunodeficiency associated with partial or complete absence
of dendritic cells, and susceptibility to mycobacterial infections. On the other
hand, IRF8 mutant mice are completely protected against cerebral malaria, a
disease in which excessive host-driven inflammatory response has lethal
pathological consequences. A genome-wide ENU mutagenesis screen has re-
cently been implemented to identify mutations that systematically protect
against cerebral malaria in vivo. The corresponding genes may play a critical
role in early host defenses against mycobacterial infections, including the
ontogeny and function of myeloid cells.

BIODEFENSE PROGRAM & ABSTRACTS 93

T U E S DAY I N V I T E D S P E A K E R A B ST R AC TS
Symposium
023. Dynamic Pathogens Make Difficult Targets
Tuesday, February 28, 2012 | 4:15 PM - 6:15 PM
Ambassador Ballroom
239 Diversity and Evolution of Pathogenic Escherichia coli:
Lessons Learned from High-Throughput Genome Sequencing
David A. Rasko; Univ. of Maryland Sch. of Med., Baltimore, MD.
Background: The identification of the E. coli O104:H4 outbreak in Germany
in the summer of 2011 was a tragic reminder that bacterial pathogens are
dynamic entities. High-throughput genome sequencing has become a feasi-
ble option for outbreak investigation at the diagnostic level as well as having
potential in the routine diagnostic laboratory. Methods: Through high-
throughput genome sequencing and comparative genomic analyses we have
been able to identify regions of genomes that are shared or are unique in iso-
lates or groups of isolates that could be used as diagnostic markers. In the
case of the German E. coli O104:H4 outbreak we examined the phylogeny of
the organism at the whole genome level as well as identified unique viru-
lence factor profiles among the outbreak isolates. Results: The isolates from
the E. coli O104:H4 outbreak in Germany were analyzed using a whole
genome phylogeny and demonstrated that these isolates were most similar
to other enteroaggregative E. coli (EAEC), but had also acquired a phage con-
taining the Shiga toxin genes. The phage was inserted into a genomic loca-
tion that is common in O157:H7 E. coli isolates. Additionally, we
demonstrated that subclinical doses of ciprofloxacin activated the expres-
sion of the Shiga toxin genes by greater than 80 fold. Virulence factor pro-
files of the outbreak isolates also revealed the carriage of an increased
number of serine protease autotransporters (SPATEs) compared to other
EAEC isolates. Conclusion: High-throughput rapid genomics was a central
tool utilized in the identification of the phylogeny and virulence factors as-
sociated with the isolates responsible for the E. coli outbreak in Germany in
2011. This methodology also provided information of a unique assemblage
of virulence factors in the EAEC background of the isolate. With the expansion
of genomic sequencing projects the variants that do not fit our current typing
schema will be identified more often and we must aware of these rapidly
evolving pathogens.
240 Adaptive Arbovirus Landscapes and Implications for Dis-
ease Emergence
Scott C. Weaver; Univ. of Texas Med. Branch, Galveston, TX.
Arboviruses, like other RNA viruses, exhibit high rates of mutation and there-
fore exist as genetically diverse populations within their hosts and vectors.
Many infect people via direct spillover from enzootic cycles, but some can
also undergo secondary transmission cycles in domesticated animals to in-
crease human exposure in agricultural settings. Moreover, several of the most
important arboviral pathogens such as dengue and chikungunya viruses can
emerge from primate-mosquito enzootic cycles to circulate exclusively among
humans, transmitted by peridomestic mosquito vectors in urban habitats.
These animal- or human-amplified secondary transmission cycles are some-
times but not always associated with adaptation to the amplification hosts or
bridge vectors, and single amino acid substitutions often mediate these host
range changes. Using the alphaviruses chikungunya and Venezuelan equine
encephalitis as examples, I will review several examples where host range
changes, with or without adaptive mutations, have led to major epidemics
and epizootics. I will also present evidence that, despite some having dra-
matic affects on fitness for infection and transmission, these host- and vec-
tor-adaptive mutations appear to be selected very inefficiently in nature, even
during major epidemics with large amounts of virus circulation. Finally, I will
review experimental studies designed to test mechanistic evolutionary hy-
potheses to explain restrictions on the Darwinian selection of adaptive al-
phavirus mutations in nature.
94 10th ASM Biodefense and Emerging Diseases Research Meeting
241 On the Molecular Evolution of Influenza A Surface Proteins
Joshua B. Plotkin; Univ. of Pennsylvania, Philadelphia, PA.
Influenza viruses evolve rapidly. Their evolution reflects not only high muta-
tion rates, but also strong pressures for mutations that evade host immunity
or anti-viral drugs. A detailed understanding of which mutations are of adap-
tive value to the virus, and in what combinations, can improve our ability to
make rational decisions about control measures, including the composition
and deployment of seasonal vaccines and drugs. I will discuss several recent
developments, arising from computational studies, in our understanding of
how hemagglutinin and neuraminidase evolve. I will also discuss ongoing
studies into the relationship between the structure and stability of influenza
surface proteins, and their molecular evolution.
242 Burkholderia pseudomallei: A Bacterial Pathogen with a
Highly Adaptable Genome
Paul Keim; Northern Arizona Univ., Flagstaff, AZ.
The great biological and genomic diversity of the B. pseudomallei are
doubtlessly related, as this organism occupies multiple niches that require a
wide repertoire of genetic options. Environmentally, B. pseudomallei can be
commonly found in water and soil of certain geographic regions, with some
evidence of plant associations. Its human pathogenic niche seems to be only
opportunistic and infrequent, but related to particular strain genotype.
Among strain genetic variation includes single nucleotide polymorphisms
(SNPs), variable number tandem repeats (VNTR) and genomic islands (GI).
Genomic islands are by definition due to horizontal gene transfer from some-
what distantly related bacteria. But the core genome is also affected by this
process; we estimate that recombination is 20 to 30 times more important
than mutation in generating new MLST alleles. VNTR loci are numbers and
novel allele generation here is probably due to slip strain mismatch repair.
Most genomic diversify is generated in the environment prior to infection,
but the processes continue in vivo and change the phenotype during a single
host event. Some of these events are clearly adaptive in providing antibiotic
resistance or evasion of the immune responsible. Others such as VNTR mu-
tations may be merely stochastic events and observable due to extraordi-
narily high rates.
Symposium
024. Visual Discovery of Microbe-Host Interactions
Tuesday, February 28, 2012 | 4:15 PM - 6:15 PM
Palladian Ballroom
243 Viral Nesting: Rewiring the Host to Generate Organelle
Platforms for Replication
Nihal Altan-Bonnet; Rutgers Univ., Newark, NJ.
Many RNA viruses upon infecting the host, generate membrane-bound repli-
cation organelles derived from the host’s intracellular membranes. The mem-
brane surface of these organelles are then used as a platform on which viral
replication machinery is assembled and viral RNA is synthesized.
We have been investigating the lipid microenvironment of these replication
platforms to identify panviral features that facilitate viral RNA synthesis. We
have recently discovered that diverse set of RNA viruses all rely on host phos-
phatidylinositol 4- kinases for replication. Specific viral proteins selectively hi-
jack these host enzymes, recruiting them to the replication membranes, in
order to generate a PI4P lipid enriched microenvironment which is critical for
viral RNA synthesis. In addition we have recently discovered a role for cho-
lesterol in organizing the replication complexes and a pathway by which RNA
viruses route host cholesterol lipids to these replication platforms. We will
discuss roles for PI4P lipids and cholesterol in regulating viral replication; de-
signing antiviral therapeutics targeting the production of PI4P lipids; and how
studying viral biology can provide insight into cellular biology.
 W E D N E S D AY I N V I T E D S P E A K E R A B S T R A C T S
245 Lighting Up Paramyxovirus Pathogenesis: Is Virus Eradica-
tion a Reality, a Redundant Concept, or a Risk?
Paul Duprex; Boston Univ. Sch. of Med., Boston, MA.
Understanding the myriad of interactions which occur when a pathogen in-
fects a primary target cell, spreads within an organism, establishes a sys-
temic infection and positions itself for transmission to the next susceptible
host is critical for the development of novel therapeutics. Such studies de-
mand access to unpassaged “real world” clinical isolates which are grown in
disease-relevant cells. This ensures that no untoward laboratory-adapted
mutations find their way into the genome as such changes can lead to erro-
neous conclusions when in vivo pathogenesis experiments are performed,
particularly if the model of disease uses an outbred species where animal to
animal variations are greater. Recognizing the importance of these issues we
developed a range of reverse genetics systems based on unpassaged
paramyxovirus isolates which can be used to produce recombinant viruses
that illuminate pathogenesis both macroscopically and microscopically.
These viruses express fluorescent proteins from an additional transcription
unit which allows infected cells to be detected with unprecedented levels of
sensitivity. Measles virus (MV) is currently being considered for global erad-
ication, although there is no real consensus as to whether this is a reality.
Following the elimination of smallpox, MV is one of the most infectious
human pathogens with an R0 of between 12 and 18. This high level of trans-
missibility is evident by the frequent importations into the US from endemic
regions and although vaccine uptake remains high the virus targets suscep-
tible hosts who have either not been vaccinated or have not generated a pro-
tective immune response post-vaccination. If MV were to be eradicated
globally discussions would ensue as to whether or not to terminate vaccina-
tion and destroy all laboratory stocks. Two critical factors would need to be
considered. First, how straightforward would it be to use synthetic biology
to recreate a fully pathogenic, highly transmissible biosafety level 2 agent?
If that is the case then eradication is a redundant concept. Second, what is
the potential for closely related animal viruses to jump the species barrier
and fill the empty post-eradication niche? If this happened one might predict
it would be associated with greater levels of morbidity and mortality and
therefore be a risk.
246 Insights into Respiratory Melioidosis through Optical Di-
agnostic Imaging
Jonathan M. Warawa; Univ. of Louisville, Louisville, KY.
Background: Diagnostic imaging approaches have the potential to facilitate
a better basic understanding of the disease process in real time as well as the
potential to translate into rapid clinical diagnostics. In these studies, we char-
acterize the respiratory disease progression of Burkholderia pseudomallei in
several mouse models using optical imaging technologies to evaluate the re-
latedness to descriptions of clinical melioidosis. Methods: B. pseudomallei
was engineered to constitutively express light from the chromosome and the
resultant lux+ strain was used to study respiratory melioidosis in a variety of
mouse models including BALB/c, SKH1, and C57BL/6J mice. Results: In-
tranasal infections of mice with B. pseudomallei result in significant colo-
nization of both the upper and lower respiratory tracts (URT/LRT), where
colonization of the lung can be detected by imaging at 24 h post infection
and earlier. In BALB/c and SKH1 mice, the lung represents the primary site of
infection. Interestingly, a capsule mutant was found to colonize the URT at
higher levels than the parent strain. In contrast, a novel intratracheal delivery
of bacteria results in LRT infection without involvement of the URT, with a
lower infectious dose than intranasal infections. Additionally, we observe an
extended course of disease which progresses into a systemic disease more
closely resembling clinical melioidosis. Conclusion: B. pseudomallei is in-
fectious by several routes of entry, including the biodefense-related respira-
tory tract infection. Mice are excellent surrogate models for the study of
melioidosis, however infections involving the URT do not accurately model
clinical melioidosis given the non-clinical propensity of B. pseudomallei to
colonize the URT in mice leading to rapid death. Intratracheal infections allow
for lung-specific delivery of organisms resulting in a disease model that more
closely resembles the clinically observed duration of disease culminating in
a systemic disease. The ability to monitor pulmonary-specific disease dis-
semination in real time will support future studies investigating therapeutic
interventions of melioidosis.
247 Computer-Assisted Quantitative Analysis of Infectious
Disease Imaging
Daniel J. Mollura; NIH, Bethesda, MD.
Background: Quantitative and molecular methods can greatly enhance de-
tection and study of preclinical and clinical infectious disease models. Meth-
ods: Computer-based quantitative measurements of CT and PET were used in
animal models and human patients to measure pulmonary infectious dis-
ease. Results: FDG-PET provided in vivo measurement of pulmonary infec-
tion in Ferret models with strong correlations between histology and FDG
uptake in H1N1 infected subjects. Computer-based measurement of respira-
tory infection accurately detected and quantified small airway inflammation
on computed tomography (CT). Computer-assisted detection (CAD) models
achieved higher diagnostic accuracy than comparison models for character-
izing pulmonary disease. Conclusion: Molecular imaging using FDG-PET and
quantitative computer-based assessments of CT can be effective tools for
studying infectious pulmonary disease.
Plenary Session
027. Immunity to Pulmonary Pathogens
Wednesday, February 29, 2012 | 8:30 AM - 12:00 PM
Regency Ballroom
252 The Host Immune Response to Pulmonary Virus Infection:
It’s About Self Control
Thomas J. Braciale; Univ. of Virginia, Charlottesville, VA.
The lower respiratory tract is the site of oxygen uptake for the body. The com-
promise of this process resulting from severe respiratory virus infection is in-
compatible with life. Effective recovery from respiratory virus infection
necessitates that the infecting virus be eliminated from the respiratory tract
with minimal inflammation and injury to the surrounding tissue. During ex-
perimental murine infection with type A influenza virus (IAV), CD8+ CTL infil-
trating the IAV infected lungs produce high levels of both proinflammatory
mediators (cytokines/chemokines) and the anti-inflammatory/regulatory cy-
tokine IL-10. We report on recent studies examining the interaction of CD8+
CTL with viral antigen expressing target cells in the IAV infected lungs, and the
consequences of these interactions for virus clearance, the regulation of IL-
10 production and the control of excess inflammation during IAV infection
(supported by USPHS grants AI-15608, HL-33391, and U19-AI-083024).
253 Requirements for Pulmonary Immunity Against Virulent
Francisella tularensis
Catharine Bosio; NIAID/Rocky Mountain Lab., Hamilton, MT.
Background: Francisella tularensis is a Gram negative, facultative intracellu-
lar bacterial pathogen. Infections mediated by F. tularensis, i.e. tularemia,
can result in lethal disease within 5-7 days following inhalation of as few as
10 bacteria. Although several animal models are available to study infection
mediated by virulent F. tularensis, identification of the specific immune com-
ponents that contribute to pathogenesis or resolution of infection have been
hampered due to the rapid mean time to death. Methods: To resolve this
problem we developed a convalescent model of tularemia in C57BL/6 mice
that relies upon the host response for clearance and survival of infection. Re-
sults: Using this model we found that, consistent with reports using more at-
tenuated strains of Francisella, alpha beta TCR+ cells and B cells were
required for survival. Although gamma delta TCR+ cells contributed to sur-
vival they were not as critical as alpha beta TCR+ cells for resolution of F. tu-
BIODEFENSE PROGRAM & ABSTRACTS 95
T U E S DAY I N V I T E D S P E A K E R A B ST R AC TS

larensis SchuS4 infection. We also found that both IL-12p40 and IL-12p35
were essential for survival of SchuS4. Similarly, IFN-gamma was also required
since animals lacking IFN-gamma receptors failed to survive antibiotic ther-
apy following SchuS4 infection. Interestingly, unlike infections mediated by
attenuated strains of F. tularensis, SchuS4 provoked both CD4+ and CD8+
IFN-gamma producing cells in the lungs, whereas CD8+ cells dominated the
IFN-gamma response in the spleen. NK and gamma delta TCR+ cells made lit-
tle contribution toward IFN-gamma production after day 7 of infection. Con-
clusion: Together this model provides a means to identify key components of
the host response required for survival or exacerbation of tularemia and will
be useful in development of novel therapeutics and vaccines.

254 Infection of Human Alveolar Macrophages by the Q Fever

Agent, Coxiella burnetii
Daniel Voth; Univ. of Arkansas for Med. Sci., Little Rock, AR.
The intracellular bacterial pathogen Coxiella burnetii is a category B select
agent that causes human Q fever. In vivo, C. burnetii targets alveolar
macrophages in which the pathogen generates a lysosome-like para-
sitophorous vacuole (PV) for replication. C. burnetii infects a variety of cul-
tured cell lines that have collectively been used to model host cell
interactions. To obtain a clearer picture of an in vivo scenario, we are estab-
lishing primary human alveolar macrophages (hAMs) as an in vitro model of
C. burnetii-host cell interactions. Virulent C. burnetii pathotypes infect and
replicate in hAMs, albeit with unique PV formation properties. The pathogen
also modulates hAM signaling required for PV formation, including the pro-
tein kinase C and cAMP-dependent protein kinase cascades. We are currently
assessing the hAM cytokine response to infection and the role of the C. bur-
netii type IV secretion system to better define the host response to this highly
infectious pathogen. The hAM model represents a disease relevant platform
for identifying potential therapeutic targets to combat Q fever.

255 The Protective Role of Surfactant Protein-A During My-

coplasma pneumoniae Infection in Allergic and Non-Allergic
Airways
Julie G. Ledford; Duke Univ., Durham, NC.
Mycoplasma pneumoniae (Mp) frequently colonizes the airways of chronic
asthmatics and is thought to contribute to exacerbations of asthma. Surfac-
tant protein A (SP-A) has well-established functions in reducing bacterial in-
fections but its role in chronic lung diseases, such as asthma, is less well
defined. Our previous work shows that mice lacking SP-A have increased air-
way constriction during Mp infection compared to WT mice and that inhibition
of TNF-α transcription reduces their responses. Additionally, mice deficient in
SP-A have enhanced inflammation and airway constriction in an allergic/in-
fection model (Ova+Mp) and inhibition of TNF-α transcription prior to Mp in-
fection can also attenuate airway reactivity in SP-A-/- allergic mice to levels
measured in WT allergic mice. Our current research provides evidence that
Mp interacts with pulmonary mast cells in non-allergic airways and causes
activation/degranulation and that SP-A plays a role in protecting from Mp-
stimulation, thereby protecting the airways from damage due to the poten-
tial release of harmful products. Additionally, in Mp-infected allergic airways
SP-A interacts with eosinophils and mediates EPO-driven clearance mecha-
nisms and protects against Mp-induced exacerbations. Therefore, SP-A plays
a protective role in non-allergic and allergic airways via regulation of mast
cell and eosinophil cellular activation during Mp infection.

96 10th ASM Biodefense and Emerging Diseases Research Meeting


Keyword........................................Presentation Number
16S/18S PCR ...................................176 (D)
A Bacillus anthracis toxins.........................042 (B)
abrin ..................................................181 (E)
Achromobacter xylosoxidans....................165 (B)
Acinetobacter baumannii.............101 (J), 158 (B)
acute respiratory infection ..................027 (A)
adeno-associated virus ........................143 (A)
Adenylate cyclase ..................................180 (D)
adjuvant................................204 (K), 208 (K)
adoptive transfer .................................198 (K)
aerosol.......029 (A), 030 (A), 038 (B), 146 (A),
.............................................149 (B), 161 (B)
African green monkeys ........................097 (I)
alhydrogel ..........................................202 (K)
allergen ..............................................182 (E)
alphavirus...........................................208 (K)
aminoglycosides..................................037 (B)
animal model development.................034 (B)
animal model...........029 (A), 030 (A), 038 (B)
Anopheles mosquito ............................099 (J)
anthracis ..................046 (B), 159 (B), 160 (B)
anthrax lethal toxin..............................212 (I)
anthrax toxin .......................................210 (I)
anthrax...................064 (D), 067 (D), 071 (E),
.................075 (E), 156 (B), 162 (B), 180 (D),
........................197 (K), 198 (K), 204 (K), 223
antibacterial activity ...........................106 (K)
antibiotic resistance..035 (B), 172 (B), 195 (G)
antibiotic .......................045 (B), 147 (B), 225
antibody ............................................080 (G)
antifungal .................................................229
antimicrobial agents............................165 (B)
antimicrobial peptide ...........................101 (J)
anti-ricin ...........................................080 (G)
antisense .....................................195 (G), 227
antiviral ...........082 (G), 143 (A), 192 (G), 228
arenavirus...........................................144 (A)
assay...................................................064 (D)
automation.........................................150 (B)
autonomous .......................................073 (E)
autotransporter ...................................154 (B)
avian influenza, inactivated vaccine.....084 (H)
Keyword........................................Presentation Number
B C
Bacillus anthracis capsule .........042 (B), 151 (B)
Bacillus anthracis .......034 (B), 039 (B), 045 (B),
.......059 (B), 75 (E), 081 (G), 095 (I), 153 (B),
.................162 (B), 168 (B), 171 (B), 179 (D),
...............................................189 (F), 214 (I)
Bacillus cereus G9241 ...........................155 (B)
Bacillus thuringiensis .............................164 (B)
bacteria.................................147 (B), 177 (D)
bacterial agents ..................................178 (D)
bacterial pathogenesis .........................049 (B)
bacteriophage-based qPCR................066 (D)
bacteriophages....................................050 (B)
baculovirus expression system.............142 (A)
Bangladesh .........................................074 (E)
beef chicken .......................................172 (B)
benefits and risks.................................077 (F)
bioaerosol...........................................061 (B)
biochip.....................................................110
biodefense concentration....................103 (K)
biodefense pathogen.................................229
biofilm...............................................068 (D)
bioinformatics resource .......................187 (F)
biological Security..............................072 (E)
biological toxins .......................................110
biosecurity ..............107 (K), 177 (D), 219 (K)
bioterror..............................................215 (J)
bioterrorism .......................................164 (B)
biothreat pathogens............................065 (D)
botulinum neurotoxins .............................111
botulism ............................................091 (H)
broad-spectrum..................................192 (G)
Brucella melitensis..................170 (B), 193 (G)
Brucella ..................................040 (B), 049 (B)
BSL-2, BSL-3 ....................................220 (K)
Burkholderia cepacia ..............................174 (B)
Burkholderia mallei ..................038 (B), 173 (B)
Burkholderia pseudomallei.........043 (B), 096 (I),
.................174 (B), 211 (I), 221, 226, 232, 233
Burkholderia thailandensis......................043 (B)
Burkholderia ........................................154 (B)
BIODEFENSE PROGRAM & ABSTRACTS
KEYWORD I N DE X
Keyword........................................Presentation Number
capillary electrophoresis .......................216 (J)
capsule ........................................156 (B), 234
capture...............................................069 (D)
cat fleas ..............................................152 (B)
cell division ........................................051 (B)
cell envelope ......................................039 (B)
centrifugal microfluidic platform........065 (D)
challenge ............................................059 (B)
chemical library .................................194 (G)
chemokine receptor CCR7 .................214 (I)
climate condition .....................................108
clonality .............................................060 (B)
Coccidioides ............................................229
complex matrices ...............................055 (B)
compound screen ...............................031 (A)
computational fluid dynamics ..............102 (J)
coronavirus ........................................033 (A)
correlates of protection.......................200 (K)
Coxiella burnetii ...................................190 (F)
Coxiella ..............................................149 (B)
Crimean-Congo Haemorrhagic
Fever Virus .......................................141 (A)
csaB ...................................................155 (B)
cytopathic-effect ................................031 (A)
D
decontamination..................................102 (J)
dendritic cells...........................................223
Topic Categories:
Below is a list of descriptions of topic cate-
gories (letters in parentheses by each pres-
entation number refer to this chart).
A. Viral Agents
B. Bacterial Agents
C. Fungal Agents
D. Diagnostics
E. Environmental Detection
F. Informatics and Genomics
G. Therapeutics
H. Vaccines
I. Immune Responses
J. Decontamination, Biosafety,
and Containment
K. Other and Information Only
97
KEYWORD I NDE X
Keyword........................................Presentation Number
dengue ...............................................031 (A)
dentistry.............................................104 (K)
detection ..............................073 (E), 177 (D)
diagnosis ............................................178 (D)
diagnostic and therapeutic potential ....050 (B)
diagnostics ..................................067 (D), 232
DNA extraction.................................176 (D)
DNA..................................................150 (B)
doxycycline........................................193 (G)
drinking water quality ........................074 (E)
DTRA....................046 (B), 047 (B), 048 (B),
.............................................159 (B), 160 (B)
E lung dendritic cells ..............................214 (I)
Ebola Virus ...........................029 (A), 146 (A)
Ebola .....................................098 (I), 207 (K)
Edema Factor.....................................180 (D)
education .............................104 (K), 175 (D)
elderly ...............................................089 (H)
electrochemiluminescence........................111
Electrospray-Neutralization Method ..083 (G)
emerging pathogen.............................165 (B)
endospore............................................216 (J)
enteritidis ...........................................157 (B)
environmental samples ........................071 (E)
epidemiology study ............................060 (B)
epidemiology .....................................032 (A)
Escherichia coli O157:H7.........079 (F), 172 (B)
evolution............................................033 (A)
export control ....................................218 (K)
F immunoassay...............................182 (E), 111
F1 and LcrV antigens..........................166 (B)
F1 and V Antiboides ...........................200 (K)
field and clinical applications ..............066 (D)
filovirus .............098 (I), 142 (A), 146 (A), 231
first responders ...................................104 (K)
forensic awareness ................................215 (J)
forensic entomology ............................215 (J)
formaldehyde.......................................100 (J)
fosmidomycin.....................................052 (B)
Francisella tularensis ................049 (B), 054 (B),
...................................................167 (B), 225
98 10th ASM Biodefense and Emerging Diseases Research Meeting
Keyword........................................Presentation Number
Francisella ...............052 (B), 088 (H), 116, 119
G
gamma irradiation ..............................199 (K)
genetic sequence analysis.....................076 (F)
GeneXpert ........................................063 (D)
genomics ............................................078 (F)
genotyping .........................................190 (F)
geographical topography ..........................108
Georgia .....046 (B), 047 (B), 048 (B), 159 (B),
.............................................160 (B), 168 (B)
growth temperature............................166 (B)
guinea pig.............................091 (H), 149 (B)
gyrA...................................................044 (B)
H lymphocytic choriomeningitis virus ...144 (A)
hantavirus.................................................112
hemorrhagic fever virus...............076 (F), 227
hepatic disease..........................................113
high-throughput screen......................082 (G)
HMGB-1.................................................225
horizontal gene transfer ......................056 (B)
host responses .....................................188 (F)
humanization .....................................080 (G)
I medicinal plant ....................................099 (J)
identification......................................068 (D)
IL-1beta and IL-6................................096 (I)
immobilized ........................................101 (J)
iImmune response ........................094 (I), 221
immunity ...........................................207 (K)
immunochromatographic test 184 (E), 186 (E)
Immunocompromised model.............090 (H)
imvamune..........................................089 (H)
in vivo imaging ........................................117
inactivated..........................................206 (K)
indirect Yersinia pestis detection...........066 (D)
inflammation ......................................157 (B)
influenza-A virus................................027 (A)
influenza ............................................032 (A)
inhibitors .................................................114
insect..................................................148 (B)
Keyword........................................Presentation Number
invasion .............................................086 (H)
Ionic propulsion .................................182 (E)
L
lab-on-paper .........................184 (E), 186 (E)
larvicidal agent.....................................099 (J)
LC16m8 ............................................090 (H)
LcrV ........................................................118
lipid A......................................................116
lipopolysaccharide ..............................034 (B)
lipoteichoic acid ...................051 (B), 081 (G)
LtaS....................................................051 (B)
lung .........................................................223
lyophilisation .....................................063 (D)
M
macaque .....................................201 (K), 228
macrophages........................................212 (I)
Madagascar...............................................112
MAPK and STAT3..............................096 (I)
Marburg Virus.............................030 (A), 227
master’s degree...................................103 (K)
melioidosis........057 (B), 108, 161 (B), 211 (I),
.........................................................226, 232
microbial infectious diseases ................079 (F)
microfluidics........................................216 (J)
microRNA expression profiling................231
microRNA ..............................................115
Mn-DP-Pi Complexes .......................199 (K)
mobile genetic element ......................056 (B)
modifiable risk factors ........................105 (K)
monkeypox.................................201 (K), 228
monoclonal antibodies ..........142 (A), 151 (B)
morbidity...........................................105 (K)
mosquito............................................167 (B)
MRSA ...............................................036 (B)
multiplex...............................073 (E), 184 (E)
murine..................................045 (B), 161 (B)
MVA..................................................205 (K)

Keyword........................................Presentation Number
MVLA................................................190 (F)
MVLA-8............................................170 (B)
N plague vaccine ......................087 (H), 200 (K)
nano-aerosol ......................................083 (G)
national security studies......................103 (K)
natural foci .........................................070 (E)
neuroinvasion...........................................222
neurovirulence .........................................222
neutrophil ...............................095 (I), 211 (I)
new antibiotic ..........................................226
new....................................................145 (A)
next generation sequencing....189 (F), 191 (F)
NF-kappB signaling cascade .....................114
non-human primate model ................203 (K)
nonhuman primate .............088 (H), 092 (H),
............................................196 (G), 209 (K)
nosocomial infection ..........................106 (K)
O ROS....................................................212 (I)
on site collection and detection ..........072 (E)
One Health........................................107 (K)
on-site detection ......................................110
ornithine lipids.........................................221
orthobunyavirus..................................078 (F)
orthopox virus ...................................203 (K)
outbreaks ...........................................067 (D)
outer membrane protein...........................119
outer membrane vesicle............................233
oxyR..................................................173 (B)
P Topic Categories:
Pakistan..............................................141 (A)
pandemic ...........................................028 (A)
parvovirus............................................100 (J)
passive transfer ...................................091 (H)
pathogen detection ............................069 (D)
pathogen ............................................218 (K)
pathology ............................................094 (I)
PCR platform comparison.................062 (D)
peptide mapping..................................210 (I)
periplasm............................................041 (B)
persistence..........................................043 (B)
Keyword........................................Presentation Number
personal safety.....................................077 (F)
phosphorodiamidate morpholion oligomers
195 (G)..........................................................
plague ........061 (B), 070 (E), 092 (H), 097 (I),
........................117, 148 (B), 152 (B), 205 (K)
plant-produced vaccine ......................197 (K)
preparedness .......................................028 (A)
priorization........................................219 (K)
protective antigen.........................210 (I), 234
protective immunity ...........................169 (B)
protein therapeutic.............................083 (G)
Pseudomallei ........................................154 (B)
Pseudomonas aeruginosa ...........053 (B), 054 (B)
pyrosequencing ..................................171 (B)
Q
Q fever ....................................................109
qPCR .................................................213 (I)
quality assurance ................................175 (D)
quasispecies ...........................033 (A), 076 (F)
quinolone...........................................035 (B)
quorum sensing..................................054 (B)
R salmonella .............................044 (B), 157 (B)
rabbit genome array ............................188 (F)
rabbitpox ............................................188 (F)
rabbits ................................................059 (B)
rabies virus .........................................140 (A)
rapid detection ...................................185 (E)
rapid diagnostic platform....................065 (D)
rare pathogens....................................176 (D)
rare variant .........................................189 (F)
reaerosolization...................................162 (B)
reagent...............................................064 (D)
real-time PCR........055 (B), 141 (A), 181 (E),
..........................................................185 (E)
reassortment .......................................078 (F)
receptor cluster formation ........................224
regulation .............................145 (A), 218 (K)
regulatory compliance, safety and
maintenance.....................................220 (K)
respiratory viruses...............................027 (A)
BIODEFENSE PROGRAM & ABSTRACTS
KEYWORD I N DE X
Keyword........................................Presentation Number
replicon platform ...............................208 (K)
resistance .................044 (B), 052 (B), 068 (D)
respiratory viral pathogens ..................028 (A)
reverse genetics ..................................144 (A)
rhesus macaque ..................................193 (G)
ribonuclease mutants ..........................037 (B)
ribosome assembly..............................037 (B)
ribosome ...........................................086 (H)
ricin ...................................................181 (E)
Ricinus communis .................................183 (E)
Rickettsia ...........................................058 (B)
Rickettsiae.........................................093 (H)
Rift Valley Fever Virus, aerosol,
enchephalitis ..........................................113
Rift Valley Fever Virus ..............................114
risk factors..........................................060 (B)
rodent................................................196 (G)
rOmpB..............................................093 (H)
rPA vaccine........................................197 (K)
RV-PCR ............................................071 (E)
S
sample preparation ................055 (B), 150 (B)
sampling.............................................217 (K)
scenario .............................................219 (K)
Schistosom mansoni ................................094 (I)
school absenteeism .............................032 (A)
Below is a list of descriptions of topic cate-
gories (letters in parentheses by each pres-
entation number refer to this chart).
A. Viral Agents
B. Bacterial Agents
C. Fungal Agents
D. Diagnostics
E. Environmental Detection
F. Informatics and Genomics
G. Therapeutics
H. Vaccines
I. Immune Responses
J. Decontamination, Biosafety,
and Containment
K. Other and Information Only
99
KEYWORD I NDE X
Keyword........................................Presentation Number
second generation sequencing ............069 (D)
secondary cell wall polysaccharide ......153 (B)
select agent...........................145 (A), 175 (D)
sGP .....................................................098 (I)
shRNA..............................................194 (G)
single domain antibodies ....................179 (D)
single nucleotide repeats .....................171 (B)
siRNA ...............................................143 (A)
site specific recombination..................035 (B)
S-layer...................................153 (B), 155 (B)
small mammals .........................................112
smallpox vaccine ....................090 (H), 213 (I)
smallpox.............................................196 (G)
SNP, MLVA........................................168 (B)
sortase ......................................................234
special unit .........................................072 (E)
spores ..................................................102 (J)
stabilization ........................................202 (K)
Staphylocidal ......................................036 (B)
Staphylococcus aureus ...............053 (B), 081 (G)
Staphyloxanthin..................................053 (B)
super resolution microscopy .....................224
surface chemistry ................................186 (E)
surveillance.........................................070 (E)
Swiss Webster mice ............................085 (H)
synergistic ..........................................036 (B)
synthetic biology (SB).........................077 (F)
T
temperature regulation .............................116
therapeutic.........................................192 (G)
therapeutics ........................................156 (B)
third world.........................................107 (K)
Thymus munbyanus ..............................106 (K)
tick born diseases ...............................178 (D)
ticks ...................................................058 (B)
toll-like receptor ......................................224
Toxoplasma gondii................................086 (H)
training courses ..................................217 (K)
transformation....................................058 (B)
transitional operations ........................220 (K)
transmission........................................167 (B)
100 10th ASM Biodefense and Emerging Diseases Research Meeting
Keyword........................................Presentation Number
Tularemia............................................088 (H)
Tularensis ............................................061 (B)
tungiasis .............................................105 (K)
type III secretion041 (B), 118, 163 (B), 169
(B) .................................................................
typing.................................................170 (B)
U
ultra-rare variant detection..................191 (F)
V
vaccine efficacy......................085 (H), 097 (I)
vaccine...........092 (H), 093 (H), 119, 201 (K),
.................202 (K), 204 (K), 205 (K), 206 (K),
...................................................207 (K), 233
vaccinia neutralization assay .................213 (I)
vaccinia virus .....................................203 (K)
vaccinia-experienced .........................089 (H)
validated PCR system.........................075 (E)
vector....................................148 (B), 152 (B)
VEEV ......................................................222
Venezuelan Equine Encephalitis virus
........................082 (G), 115, 199 (K), 206 (K)
vibrio cholerae ...................................056 (B)
ViPR..................................................187 (F)
virulence analysis................................042 (B)
virulence ............................................173 (B)
virus pathogen resource ......................187 (F)
W
water quality sensors...........................164 (B)
weapons mass destruction...................217 (K)
World Health Organization ................074 (E)
X
XF Drugs ...........................................158 (B)
Y
Yersinia pestis, Clostridium botulinum ......185 (E)
Yersinia pestis,Y. pseudotuberculosis ..........050 (B)
Yersinia pestis...........085 (H), 079 (F), 087 (H),
..............................117, 118, 166 (B), 209 (K)
Yersinia .....................041 (B), 047 (B), 169 (B)

Listed by Author Name, Presentation Number, Abstract Category. *Presenting author is in Bold Face Type.
Aaron, J. S. ...............................................224
Abazashvili, N. ...................................046 (B)
Abbott, E. .........................................202 (K)
Abdukhalilova, G. ..............................062 (D)
Achazi, K. .........................................065 (D)
Adamia, R. ........................................160 (B)
Adams, C. .........................................165 (B)
Agans, K. ...................................201 (K), 228
Agrawal, S. .........................................079 (F)
Ågren, P. ............................................055 (B)
Alavi, M. R. ......................................053 (B)
Albornoz, E. ......................................035 (B)
Alfaro, T. ............................................071 (E)
Allen, J. .............................................033 (A)
Almuzara, M. .....................................165 (B)
Altan-Bonnet, N. ....................................243
Alterson, K. .......................................208 (K)
Aly, K. ...............................................163 (B)
Amieva, M. R. .........................................121
Amoako, K. .......................................171 (B)
Amosun, A. E. ....................................172 (B)
Anderson, A. D. .......................................109
Anderson, B. ......................................028 (A)
Anderson, G. .....................................179 (D)
Anderson, M. ....................................193 (G)
Andres, P. ...........................................035 (B)
Andrews, C. ......................................202 (K)
Andrianaivo, T. ........................................112
Ansari, U. ..........................................180 (D)
Antonic,V. .........................................053 (B)
Aranio, D. .........................................067 (D)
Arcidiacono, S. ...................................101 (J)
Arterburn, J. ......................................082 (G)
Arvans, D. .........................................194 (G)
Astwood, M. ......................................183 (E)
Atimnedi, P. .......................................107 (K)
Atkins, T. P. ........................................154 (B)
Atkinson, B. ......................................141 (A)
Atkinson, D. J. ....................................149 (B)
AuCoin, D. ........................................156 (B)
AuCoin, D. P. ..........................................232
Avaliani, N. ........................................046 (B)
Axthelm, M. .....................................088 (H)
Baccetty, A. .......................................062 (D)
Backman, S. ......................................167 (B)
Bae, E. ..............................................068 (D)
Baer, A. ....................................................114
Bagarozzi, D. .....................................180 (D)
Bailey, C. .................................................114
Bailey, T. .....................................201 (K), 228
Baillie, L. ............................................095 (I)
Bakanidze, L. .....................................047 (B)
Bakanidze, L. G. ................................070 (E)
Baker, R. ..............................029 (A), 030 (A)
Baker, R. .................................................228
Bakken, R. ........................................206 (K)
Bakken, R. R. ...................................208 (K)
Balabaev, N. K. ..................................083 (G)
Bales, J. M. ..............................................113
Balinandi, S. ......................................107 (K)
Baron, C.. ................................................137
Barr, J. R. ...............059 (B), 067 (D), 180 (D)
Barry, E. ............................................049 (B)
Bate, S. R. ..........................................045 (B)
Bates, S. .............................................161 (B)
Bavari, S. .................................................227
Bearden, S. W. ....................................148 (B)
Beckstrom-Sternberg, J. .....................168 (B)
Beckstrom-Sternberg, S. ....................168 (B)
Bedi, K. .............................................180 (D)
Beeri, K. ............................................051 (B)
Beitzel, B. ..................................082 (G), 227
Bell, T. ..............................................032 (A)
Belle, A. ............................................059 (B)
Bengtsson, U. A. ..................................100 (J)
Bent, Z. ...........................................069 (D)
Berthelot, T. ..........................184 (E), 186 (E)
Bethel, L. M. ...........................................113
Bewley, K. R. ....................................149 (B)
Bhomia, M. .............................................222
Birdsell, D. .........................................168 (B)
Birkenholz, J. .....................................150 (B)
Bjork, A. ..................................................109
Blaney, D. ..........................................067 (D)
BIODEFENSE PROGRAM & ABSTRACTS
AUTHOR INDEX
Blaney, J. ...........................................207 (K)
Blaylock, B. .........................041 (B), 163 (B)
Blazevic, E. ........................................073 (E)
Bloomfield, H. .....................029 (A), 030 (A)
Bloomfield, H. A. ..............................146 (A)
Boboye, B. E. ......................................099 (J)
Bonvini, E. ........................................196 (G)
Booth, J. .............................................214 (I)
Borel, A. ............................................204 (K)
Borja, N. A. ........................................060 (B)
Borucki, M. ......................................033 (A)
Bosio, C. .................................................253
Bowe, M. ..........................................196 (G)
Bowen, L. E. ......................................200 (K)
Bowen, R. ...............................................221
Bower, K. ..........................................043 (B)
Boydston, J. A. ...................................200 (K)
Boyer, A. ...........................................180 (D)
Boyer, A. E. .........................059 (B), 067 (D)
Boylan, J. ...........................................173 (B)
Boylan, R. J. .....................................104 (K)
Braciale, T. J. ............................................252
Brand, S. ..................................................229
Branda, S. ..........................................069 (D)
Brasel, T. ....................................201 (K), 228
Braun, A. ............................................214 (I)
Brett, P. J. .................................................232
Brewoo, J. .........................................205 (K)
Brezillon, C. ......................................042 (B)
Topic Categories:
Below is a list of descriptions of topic cate-
gories (letters in parentheses by each pres-
entation number refer to this chart).
A. Viral Agents
B. Bacterial Agents
C. Fungal Agents
D. Diagnostics
E. Environmental Detection
F. Informatics and Genomics
G. Therapeutics
H. Vaccines
I. Immune Responses
J. Decontamination, Biosafety,
and Containment
K. Other and Information Only
101
AUTHOR I N DE X
Listed by Author Name, Presentation Number, Abstract Category. *Presenting author is in Bold Face Type.
Briscoe, C. ........................................193 (G)
Brittingham, K. T. ..............................057 (B)
Brogden, K. A. .........................................135
Broladze, M. ......................................046 (B)
Brooks, K. ..........................................097 (I)
Brown, P. B. ......................................217 (K)
Bruce, C. ............................141 (A), 142 (A)
Brunelle, S. ........................................075 (E)
Buller, M. ..........................................196 (G)
Burger, R. F. ............................................025
Burke, S. ...........................................196 (G)
Burke, W. ...........................................150 (B)
Burkhardt, N.Y. .................................058 (B)
Burtnick, M. N. .......................................232
Busuulwa, M. ....................................107 (K)
Byfors, S. ..........................................055 (B)
Callahan, J. ........................................062 (D)
Cann, J. .............................................207 (K)
Cardwell, M. M. ................................093 (H)
Carlos, M. P. ......................................028 (A)
Carlos, M. P. ......................................178 (D)
Carrion, R. .......................................059 (B)
Carson, B. D. ...........................................224
Carson, H. .........................................145 (A)
Carter, A. .................................................225
Cebula, T. A. ......................................056 (B)
Centrón, D. .......................................165 (B)
Chamberlain, J. ..................................141 (A)
Champney, W. S. ................................037 (B)
Chan, W. .............................................216 (J)
Chance, T. B. .....................................146 (A)
Chantratita, N. .........................................232
Chanturia, G. ......................047 (B), 159 (B)
Chanturia, G. .....................................168 (B)
Chaplin, P. ........................................089 (H)
Chapman, J. .......................................150 (B)
Chappie, D. J. .....................................162 (B)
Charlwood, M. ..................................149 (B)
Chau, D. ...........................................080 (G)
Chen, A. ............................................185 (E)
Chen,Y.-L. ..............................................108
Chen,Y.-S. ..............................................108
102 10th ASM Biodefense and Emerging Diseases Research Meeting
Chen, Z. ...........................................180 (D)
Cheng, B.Y. H. ..................................144 (A)
Cheng, L. W. ............................................111
Cheong, H. .........................................216 (J)
Cherwonogrodzky, J. .........................080 (G)
Chichester, J. .....................................197 (K)
Chinglai,Y. .........................................098 (I)
Chiu, C. ..................................................008
Cho, M.-H. .......................................096 (I)
Cho, M. .............................................151 (B)
Choi, S.Y. ..........................................056 (B)
Christman, J. W. ..................................212 (I)
Chun, J. .............................................056 (B)
Chun, J.-H. ...........................096 (I), 151 (B)
Chung, D. ........................................082 (G)
Ciccarone,V. ......................................196 (G)
Ciletti, N. ..........................................169 (B)
Ciletti, N. A. .....................................087 (H)
Clark, G. C. .............................................225
Clemens, K. M. ..................................075 (E)
Clement, K. H. .................................064 (D)
Clohessey, A. ......................................102 (J)
Coffin, J. ............................................158 (B)
Coggeshall, K. .....................................214 (I)
Cohen, J. ...........................................206 (K)
Colman, R. E. ...................................189 (F)
Colwell, R. R. ......................056 (B), 185 (E)
Compans, R. ......................................098 (I)
Connor, M. G. .........................................117
Cook, J. L. ..........................................212 (I)
Cook, N. ...........................................141 (A)
Corso, A. ...........................................035 (B)
Credou, J. ............................184 (E), 186 (E)
Cronin, L. X. .....................................067 (D)
Cross, A. .............................................095 (I)
Crosson, S. ........................................194 (G)
Cummings, P. J. .................................103 (K)
Currie, B. J. .............................................011
Curtis, D. ..........................................069 (D)
Custer, M. .........................................208 (K)
D’Elia, R.V. .............................................225
Daly, M. J. ..........................................199 (K)
Das, R. ....................................................114
DaSilva, L. .........................................188 (F)
Daugherty, S. ......................................079 (F)
Dauphin, L. A. ...................................183 (E)
de la Garza, M. ..................................059 (B)
de la Torre, J. C. .................................144 (A)
Dean, R. E. ........................................154 (B)
Dean, S. N. .......................................054 (B)
Dearen, K. .................................201 (K), 228
Dekonenko, A. ..................................196 (G)
Delarosa, P. ........................................107 (K)
Delp, K. .............................................043 (B)
Dembele, M. ....................................178 (D)
Demeure, C. E. ........................................237
Dergel, I. ............................................100 (J)
Dersch, P. ................................................019
DeShazer, D. ............................................013
Dieringer-Boyer, S. ............................043 (B)
Dietrich, J. .........................................187 (F)
Disqué, C. .........................................176 (D)
Doherty, L. .........................................101 (J)
Dong, L. ..................................................227
Dorsey, B. ..........................................043 (B)
Douady, J. ................................................223
Dowall, S. ..........................................141 (A)
Drew, J. ............................................202 (K)
Drexler, J. ..........................................065 (D)
Duggan, E. ..........................................214 (I)
Duggan, M. .......................................150 (B)
Dupke, S. ..........................................042 (B)
Duprex, P. ...............................................245
Dworkin, J. ........................................153 (B)
Dyer, D. N. ........................................034 (B)
Easterbrook, L. ..................................142 (A)
Eberhard, M. .....................................065 (D)
Eckstein, T. M. ............................174 (B), 221
Eduok, N. N. ....................................147 (B)
Egan, C. ............................................040 (B)
Ehrs, S. ..............................................055 (B)
EL-BARRAWY, M. .........................027 (A)
Eleazar, C. I. ......................................147 (B)
Ellerbrok, H. ....................................203 (K)

Listed by Author Name, Presentation Number, Abstract Category. *Presenting author is in Bold Face Type.
Elli, D. .................................087 (H), 169 (B)
Elssner, T. ................................................110
Emmoth, E. .......................................100 (J)
Engelthaler, D. M. ..............................189 (F)
Ennis, J. .............................................208 (K)
Eppinger, M. .....................................079 (F)
Epstein, G. L. ...................................134, 251
Epstein, J. H. ............................................010
Ernst, R. K. .............................................116
Facemire, P. .......................................038 (B)
Fair, J. ......................................................007
Farb, D. .............................................196 (G)
Fatima zohra, M. ..............................106 (K)
Faucher, S. P. ............................................018
Felder, E. ..............................063 (D), 181 (E)
Fellows, P. ...........................085 (H), 209 (K)
Felsheim, R. F. ...................................058 (B)
Feodorova,V. A. .................................166 (B)
Filippov, A. A. ......................050 (B), 066 (D)
Fiole, D. ...................................................223
Firmani, M. A. ...................................043 (B)
Fisher, I. ................................092 (H), 097 (I)
Fitzpatrick, K. A. ......................................109
Fofanov,V. ..........................................191 (F)
Foote, S. J. ...............................................235
Foster, J. .............................................189 (F)
Francesconi, S. ...................................047 (B)
Francesconi, S. ..................................062 (D)
Francesconi, S. ...................................159 (B)
Franchini, G. .....................................090 (H)
Franz, T. .............................................042 (B)
Fraser-Liggett, C. ...............................079 (F)
Frazier, A. D. .....................................037 (B)
Frederick, S. .........................029 (A), 030 (A)
Freiberg, A. ........................................207 (K)
Fretwell, J. ............................045 (B), 161 (B)
Funatsu, A. ........................................090 (H)
Funnell, S. G. P. .....045 (B), 149 (B), 161 (B)
Gabay, J. E. ........................................194 (G)
Gage, K. L. ........................................148 (B)
Gaidamakova, E. K. ...........................199 (K)
Galens, K. ..........................................079 (F)
Galindo, C. L. .....................................188 (F)
Gallegos, K. ..........................092 (H), 097 (I)
Gallegos, M. ......................................180 (D)
Gallegos-Candela, M. ...........059 (B), 067 (D)
Galyov, E. E. ......................................154 (B)
Gandhi, P. ..........................................182 (E)
Garcia, A. D. .......................................213 (I)
Garcia de la Guardia, R. .....................060 (B)
Garib,V. ............................................062 (D)
Garrett, J. ..........................................145 (A)
Garufi, G. ..................................051 (B), 234
Garvey, E. ................................................229
Garza, N. .................................................227
Gauthier,Y. .......................................204 (K)
Gayen, M. ........................................199 (K)
Gelhaus, H. C. ...................................057 (B)
Geller, B. ...........................................195 (G)
Gene Olinger, G. ...............................082 (G)
Gerba, C. ................................................124
Gherardini, F. .....................................173 (B)
Gillece, J. ............................................189 (F)
Gingras, B. .........................................061 (B)
Giordano, A. ......................................068 (D)
Giron, J. A. ..............................................122
Given, K. .................................................118
Glass, P. ............................................206 (K)
Glass, P. J. ............................199 (K), 208 (K)
Glotzer, D. L. .....................................104 (K)
Glyakina, A.V. ....................................083 (G)
Goji, N. .............................................171 (B)
Goldman, E. ......................................179 (D)
Goldman, S. ............................................226
Goldstein, J. .......................................180 (D)
Gonzalez-Juarrero, M. .............................221
Goodlett, D. R. ........................................116
Goodman, S. M. ......................................112
Goossens, P. L. ...................................042 (B)
Gordon, J. .........................................182 (E)
Gordon, M. .............................................123
Gordon, S. ........................................090 (H)
Gray, J. ..............................................175 (D)
Greenberg for the POX-MVA-024
BIODEFENSE PROGRAM & ABSTRACTS
AUTHOR INDEX
Sites, R. N. ....................................089 (H)
Gros, P. ....................................................236
Grunow, R. .......................................042 (B)
Gu, Z. ................................................187 (F)
Gubeladze, L. .....................................160 (B)
Guendel, I. .......................................114, 115
Gunn, J. ...................................................017
Gupta, P. .......................199 (K), 206 (K), 222
Hadfield, T. ........................................075 (E)
Hahn, K. R. ......................................171 (B)
Hakim, S. T. .......................................077 (F)
Haley, B. J. .........................................185 (E)
Hallis, B. ............................................198 (K)
Han, J. .....................................................222
Hardt, W.-D. ......................................157 (B)
Harman-Richardson, N. ....................200 (K)
Harris-Chen, H. ................................033 (A)
Hartman, A. L. .........................................113
Hasan, N. A. ......................................056 (B)
Hasan, Z. ...........................................141 (A)
Hashizume, S. ...................................090 (H)
Hatch, G. .............................149 (B), 161 (B)
Hatch, G. J. .......................................045 (B)
Hauck, L. ................................................227
Hayter, I. ...........................................158 (B)
He,Y. .................................................050 (B)
Healey, B. ..........................................028 (A)
Heaton, B. .........................................194 (G)
Heckert, R. .......................................107 (K)
Topic Categories:
Below is a list of descriptions of topic cate-
gories (letters in parentheses by each pres-
entation number refer to this chart).
A. Viral Agents
B. Bacterial Agents
C. Fungal Agents
D. Diagnostics
E. Environmental Detection
F. Informatics and Genomics
G. Therapeutics
H. Vaccines
I. Immune Responses
J. Decontamination, Biosafety,
and Containment
K. Other and Information Only
103
AUTHOR I N DE X
Listed by Author Name, Presentation Number, Abstract Category. *Presenting author is in Bold Face Type.
Heffron, F. .........................................088 (H)
Heim, J. ...................................................226
Hendrickx, A. ....................................051 (B)
Hendrickx, A. P. .................................155 (B)
Hendrickx, A. P. A. ............................081 (G)
Henning, L. ......................................193 (G)
Heraud, J.-M. ..........................................112
Hermanas, T. M. ..................087 (H), 152 (B)
Herron, M. J. .....................................058 (B)
Herzog, C. .........................................072 (E)
Hewitt, J. .................................................003
Hewson, R. ........................141 (A), 142 (A)
Heymann, D. L. .......................................021
Hill, L. .....................................................114
Hillman, R. .......................................093 (H)
Hoekstra, W. ............................................229
Hoffmaster, A. R. ..............................067 (D)
Holbrook, M. ....................................207 (K)
Holland, L. ........................................061 (B)
Holland, L. ........................................085 (H)
Hong, K.-J. .........................................096 (I)
Hong, K. ...........................................151 (B)
Honnold, S. .......................................206 (K)
Hoq, M. ............................................056 (B)
Hoque, S. ............................................102 (J)
Houghton, R. .........................................232
Hu, S.-C. ...........................................061 (B)
Hu, W. ..............................................080 (G)
Huang, L. ..........................................196 (G)
Huang, X.-Z. ...................................170 (B)
Hubbard, M. A. .................................156 (B)
Huber, N. T. ......................................064 (D)
Hudson, L. ........................................142 (A)
Hultén, C. ...........................................100 (J)
Hunt,V. ..............................................187 (F)
Huntley, J. ...............................................119
Huq, A. ................................056 (B), 185 (E)
Hutchins, A. S. ..................................183 (E)
Ibadova, G. ........................................062 (D)
Imnadze, P. ...........................046 (B), 159 (B)
Imnadze, P. ........................................168 (B)
Imnadze, P. G. ....................................070 (E)
104 10th ASM Biodefense and Emerging Diseases Research Meeting
Inamine, J. M. ...................................174 (B)
Isbell, K. ..............................067 (D), 180 (D)
Iversen, P. ................................................227
Iversen, P. L. .....................................195 (G)
Izadjoo, M. ........................................053 (B)
Jahrling, P. .........................................207 (K)
Jamil, B. .............................................141 (A)
Jang, J. ...............................................151 (B)
Janzen, T. ...........................................171 (B)
Jefferson, C. .............................................250
Johnson, S. ........................................196 (G)
Jones, E. ..................................................004
Jones, M. ...........................................197 (K)
Jonsson, C. B. ....................................082 (G)
Judy, B. ....................................................233
Kaboyo, R. W. ....................................107 (K)
Kadan, M. .........................................196 (G)
Kamrud, K. I. ....................................208 (K)
Kane, J. A. ..........................................161 (B)
Kane, S. .............................................071 (E)
Kanehara, T. ......................................090 (H)
Kanev, I. L. ........................................083 (G)
Karaolis, D. K. R. ..............................043 (B)
Kashanchi, F. ....................................114, 115
Kaup, F.-J. ..........................................203 (K)
Kehn-Hall, K. ..................................114, 115
Keim, P. .........................168 (B), 189 (F), 242
Keim, P. S. ..........................................057 (B)
Kekeklidze, M. ...................................047 (B)
Kekelidze, M. ....................................159 (B)
Kekelidze, M. ....................................168 (B)
Kelly-Cirino, C. .................................040 (B)
Kern, J. ..............................................051 (B)
Kersh, G. J. ..............................................109
Khizhnyakova, M. A. ..........................166 (B)
Khmaladze, E. ....................................159 (B)
Khmaladze, E. ...................................168 (B)
Khodiev, A. .......................................062 (D)
Kidney, A. ..........................................040 (B)
King, C. A. ..............................................115
Klee, S. R. .........................................042 (B)
Klem, E. ............................................187 (F)
Klint, M. ...........................................055 (B)
Knight, I. ...........................................185 (E)
Koblentz, G. ............................................133
Koenig, S. ..........................................196 (G)
Koide, F. ..............................196 (G), 200 (K)
Kondas, A.V. ............................................109
Kopaskie, K. .....................................163 (B)
Koroleva, G. ......................................148 (B)
Koroleva, G. I. ....................................078 (F)
Koshinsky, H. ....................................191 (F)
Kozel, T. ............................................156 (B)
Kozel, T. R. .............................................232
Kramski, M. ......................................203 (K)
Krile, R. ............................................209 (K)
Kugelman, J. ............................................227
Kugelman, J. R. .....................078 (F), 148 (B)
Kuiken, C. ........................................076 (F)
Kuklenyik, Z. ....................................180 (D)
Kumar, S. ...........................................187 (F)
Kurtti, T. J. .........................................058 (B)
Kutateladze, M. ..................................160 (B)
Kvachadze, L. ....................................160 (B)
Lander, A. ..........................................042 (B)
Lane, P. ..............................................069 (D)
Lane, T. .............................................069 (D)
Langer, M. .........................................214 (I)
Langevin, S. .......................................069 (D)
Larson, C. ..........................................187 (F)
Latham, J. ...........................................190 (F)
Lathem, W. W. ..........................................015
Lavander, M. ......................................055 (B)
Lawrenz, M. B. ........................................117
Laws, T. R. ...............................................225
Lawson, T. .........................................178 (D)
Lazar Adler, N. R. .............................154 (B)
LeClaire, R. ..........................092 (H), 097 (I)
Ledford, J. G. ...........................................255
Lee, M.-H. ........................................072 (E)
Lee, P. ...............................................179 (D)
Leffel, E. ..........................................006, 136
Legasse, A. .........................................088 (H)
Lehman, M. ......................................067 (D)

Listed by Author Name, Presentation Number, Abstract Category. *Presenting author is in Bold Face Type.
Lehman, S. .........................................043 (B)
Lemoine, S. ...........................092 (H), 097 (I)
Leppla, S. H. ......................................180 (D)
Letant, S. ............................................071 (E)
Levi, A. .............................................194 (G)
Levitan, I. ...........................................212 (I)
Levy, S. ....................................................248
Leysath, C. E. ....................................180 (D)
Li, H. ................................................067 (D)
Li, H. ................................................196 (G)
Li, L. ...................................................216 (J)
Li,Y. ........................................................116
Lin, H.-S. ................................................108
Lin, S. ................................................200 (K)
Lin, W. ..............................................085 (H)
Lin, W. C. ..........................................061 (B)
Lind, C. .............................................206 (K)
Lindström, A. .....................................215 (J)
Linke, S. ............................................065 (D)
Linow, M. .........................................176 (D)
Lins, R. C. ..........................067 (D), 180 (D)
Liu, D. ...............................................175 (D)
Liu, G. .....................................................226
Liu, J. ................................................179 (D)
Liu, J. .................................................191 (F)
Liu, M. ..............................................187 (F)
Liu, P.-J. ..................................................108
Llewellyn, Z. N. ................................200 (K)
Lockman, H. .....................................209 (K)
Lofts, L. L. .........................................078 (F)
Logue, C. ..........................................141 (A)
Lohman, K. L. ....................................158 (B)
Lordo, R. A. .......................................162 (B)
Love, H. ............................................142 (A)
Love, W. G. .......................................158 (B)
Lukaszewsk, R. A. ....................................225
Lunderberg, J. M. ..............................153 (B)
Lundin Zumpe, A. .............................055 (B)
Lyapina, A. M. ....................................166 (B)
Lyons, A. ...........................................180 (D)
M. Nikolaishvili, M. Zakareishvili,
I. Beradz, e. ......................................048 (B)
Mackie, R. S. ....................................052 (B)
Maheshwari, R. .................................206 (K)
Maheshwari, R. K. .....................199 (K), 222
Maier, H. .................................................234
Malania, L. .............046 (B), 159 (B), 168 (B)
Mamisashvili1, E. ...............................048 (B)
Manceva, S. .......................................197 (K)
Maniatis, P. ........................................067 (D)
Mark, D. ...........................................065 (D)
Marohn, M. ......................................049 (B)
Maroo, A. ...........................................079 (F)
Marshall, S. .......................................085 (H)
Marston, C. .......................................067 (D)
Martin, S. .........................................091 (H)
Martinez, J. .......................................194 (G)
Martinez, J. J. ....................................093 (H)
Martinez-Sobrido, L. .........................144 (A)
Maruno, S. ........................................090 (H)
Mathieu, J. .........................................204 (K)
Mathieu, J. D. ..........................................223
Matsui, H. .........................................090 (H)
Mätz-Rensing, K. ..............................203 (K)
McCool, J. .........................................163 (B)
McCormick, K. D. ............................031 (A)
McGuinness, E. .................................193 (G)
McIver, L. ..........................................188 (F)
McKenney, E. S. .................................052 (B)
McLuckie, G. .....................................149 (B)
McMahon, K. ....................................188 (F)
Mellmann, A. ..........................................026
Mello, C. ............................................101 (J)
Meseda, C. A. ......................................213 (I)
Metcalf, J. ...........................................214 (I)
Metcalfe, K. ......................................091 (H)
Migliozzi, R. H. ................................064 (D)
Mikeladze, E. .....................................047 (B)
Miller, N. ................................................118
Miller, N. C. .....................................169 (B)
Miller, S. ...........................................193 (G)
Milligan, D. .............................................226
Missiakas, D. ....039 (B), 051 (B), 081 (G), 234
Missiakas, D. M. .................................153 (B)
BIODEFENSE PROGRAM & ABSTRACTS
AUTHOR INDEX
Mitchell, A. .............................................118
Moatter, T. .........................................141 (A)
Mohan, G. S. ......................................098 (I)
Mollik, M. ........................................074 (E)
Mollura, D. J. ...........................................247
Monier, A. ............................092 (H), 097 (I)
Montague, J. ......................................217 (K)
Montgomery, S. .................................178 (D)
Moore, W. ................................................229
Morici, L. A. ............................................233
Morozov,V. N. ..................................083 (G)
Mossbrugger, I. ..................................181 (E)
Motin,V. L. ........................................166 (B)
Motlagh, H. ......................................068 (D)
Mourich, D. ............................................227
Mourich, D.V. ...................................195 (G)
Mukhopadhyay, K. .............................036 (B)
Mulenga, A. .............................................130
Muller, W. ...........................................101 (J)
Munderloh, U. G. ..............................058 (B)
Murphy, G. ........................................071 (E)
Murphy, N. ......................................176 (D)
Murtaugh, W. A. ................................028 (A)
Musiychuk, K. ...................................197 (K)
Mwangi, W. .............................................221
N. Ivanov, O. Deryabin,V. Skripnik, ....140 (A)
Na, M. P. M. N. ..................................094 (I)
Nadeem, S. G. ....................................077 (F)
Nakao, B. ...........................................073 (E)
Topic Categories:
Below is a list of descriptions of topic cate-
gories (letters in parentheses by each pres-
entation number refer to this chart).
A. Viral Agents
B. Bacterial Agents
C. Fungal Agents
D. Diagnostics
E. Environmental Detection
F. Informatics and Genomics
G. Therapeutics
H. Vaccines
I. Immune Responses
J. Decontamination, Biosafety,
and Containment
K. Other and Information Only
105
AUTHOR I N DE X
Listed by Author Name, Presentation Number, Abstract Category. *Presenting author is in Bold Face Type.
Nalca, A. ..029 (A), 030 (A), 146 (A), 188 (F)
Narayanan, A. ..........................................114
Natidze, M. .......................................160 (B)
Ndumu, D. B. ...................................107 (K)
Negrych, L. .......................................080 (G)
Newman, K. A. .................................064 (D)
Ngomi, N. N. N. N. ..........................105 (K)
Nguyen-Mau, S.-M. ............039 (B), 153 (B)
Nichols, D. K. ....................................146 (A)
Nichols, T. L. .....................................162 (B)
Niedrig, M. .......................................065 (D)
Niemuth, N. .....................................091 (H)
Nieves, W. ................................................233
Nikolich, M. ......................................168 (B)
Nikolich, M. P. .....................050 (B), 066 (D)
Nikolich, M. P. ..................................170 (B)
Nilsson, C. .........................................055 (B)
Noh, S. M. ..............................................129
Nordstrom, J. L. ................................196 (G)
Noronha, J. ........................................187 (F)
Northurp, M. A. .................................073 (E)
Nozadze1, M. ....................................047 (B)
Nunneley, J. .......................................059 (B)
Nyakiti, D. .........................................038 (B)
Obiso, R. ..........................................062 (D)
Obiso, R. J. ............046 (B), 047 (B), 048 (B),
.............................................159 (B), 160 (B)
Obom, K. M. ....................................103 (K)
Offit, P. A. ................................................020
Ogeneh, B. O. ....................................147 (B)
Oh, S.-Y. ....................................155 (B), 234
Ohkuma, K. ......................................090 (H)
Olatoye, O. .......................................172 (B)
Olive, M.-M. ...........................................112
Oliver, J. D. .......................................058 (B)
Omoya, F. O. ......................................099 (J)
Onashvili, T. .....................................048 (B)
Ongarbaev, A. ...................................062 (D)
Ortiz Riano, E. .................................144 (A)
Osorio, J. ...........................................205 (K)
Ota, I. M. ...........................................075 (E)
Overheim, K. A. ..................092 (H), 097 (I)
106 10th ASM Biodefense and Emerging Diseases Research Meeting
Oyston, P. ................................................139
Pak, D. ..............................................193 (G)
Pal, U. .....................................................128
Palacios, G. ..............................................227
Palacios, G. F. ......................................078 (F)
Palacios, G. F. .....................................148 (B)
Pandrea, I. ...............................................113
Papaneri, A. .......................................207 (K)
Park, C. ...................................................132
Park, J. ...............................................151 (B)
Parrish, T. ..........................................150 (B)
Partidos, C. ........................................205 (K)
Patel, P. ............................................065 (D)
Patel,V. ...............................................214 (I)
Pati, N. B. ..........................................157 (B)
Pauli, G. ............................................203 (K)
Peacock, S. J. ............................................232
Pearson, T. .........................................168 (B)
Peck, M. ............................................040 (B)
Pennington, J. ..........................................117
Pepler, A. ...........................................190 (F)
Pepper, I. ...........................................164 (B)
Pereyra, F. ................................................238
Periaswamy, B. ...................................157 (B)
Perry, M. ..........................................040 (B)
Pesik, N. ............................................067 (D)
Peters, H. ...........................................028 (A)
Petroni, A. .........................................035 (B)
Phillippy, A. .......................................185 (E)
Pickett, B. ..........................................187 (F)
Pitman, J. K. ......................................149 (B)
Pitt, L. .......................................001, 038 (B)
Pitt, M. L. M. .....................................034 (B)
Pitts, R. .............................................180 (D)
Plank, J. .............................................142 (A)
Plotkin, J. B. ............................................241
Pohl, J. ................................................210 (I)
Pöhlmann, C. ..........................................110
Polupan, I. ........................................140 (A)
Polyanina, T. I. ...................................166 (B)
Porter, A. ............................................188 (F)
Potluri, L. P. .......................................194 (G)
Powell, D. ................................................113
Powell, D. A. ............................................116
Powell, T. ...........................................205 (K)
Power, A. .................................................231
Prenzler, F. ..........................................214 (I)
Price, J. ..............................085 (H), 209 (K)
Priestley, R. A. .........................................109
Prior, J. L. ..........................................154 (B)
Probst, L. ..........................................073 (E)
Puri, R. K. ..............................................222
Quenee, L. ........................................093 (H)
Quenee, L. ........................................169 (B)
Quenee, L. E. ....................................152 (B)
Quenee, L. E. F. ................................087 (H)
Quesnel-Hellmann, A. .............................223
Quinn, C. ...........................................210 (I)
Quinn, C. P. ........................067 (D), 180 (D)
Raetz, C. R. H. .......................................116
Rainey, G. J. ......................................196 (G)
Rajagopal, S. ....................................086 (H)
Ramachandran, G. .............................095 (I)
Ramirez, M. ......................................165 (B)
Ramishvili, M. ...................................046 (B)
Rasko, D. A. .....................................116, 239
Ravel, J. .............................................079 (F)
Rawat, A. ...........................................189 (F)
Rawlinson, W. ...................................175 (D)
Raychaudhuri, S. .....................................232
Razafindralambo, N. ................................112
Reed, D. ..................................................113
Reed, D. ..................................................232
Revelli, D. A. ....................................173 (B)
Reyes-Lamothe, R. ...........................035 (B)
Reynes, J.-M. ..........................................112
Rhee, A. ............................................197 (K)
Rhie, G.-E. ...........................096 (I), 151 (B)
Rhys-Williams, W. .............................158 (B)
Richards, K. ......................................142 (A)
Richardson, M. .................................032 (A)
Richter, G. S. .....................................153 (B)
Richter, M. H. ..................................072 (E)
Richter, S. ........................................081 (G)

Listed by Author Name, Presentation Number, Abstract Category. *Presenting author is in Bold Face Type.
Richter, S. G. .....................................051 (B)
Rider, T. H. ......................................192 (G)
Rigvava, S. ........................................160 (B)
Riley, S. .............................................194 (G)
Riley, S. P. .........................................093 (H)
Robers, A. .........................................142 (A)
Roberts, A. ...........................149 (B), 161 (B)
Roberts, A. D. G. ...............................045 (B)
Roberts, R. .............................................002
Robins, P. ..........................................194 (G)
Robinson, J. ......................................068 (D)
Roche, A. J. .............................................109
Roffey, P. ...........................................175 (D)
Rom, A. ............................................085 (H)
Romero, F. ........................................092 (H)
Rosati Rowe, J. A. ..............................162 (B)
Rose, L. ...................................................126
Rosen, G. ...........................................095 (I)
Roth, A. I. .........................................103 (K)
Roussel, P. ...............................................226
Rowan, L. .........................................206 (K)
Roy, C. J. .................................................233
Rudge, T. L. ......................................064 (D)
Russell, R. ............................092 (H), 097 (I)
Russo, A. ...........................................201 (K)
Russo, A. T. ..............................................228
Rutebarika, C. S. ...............................107 (K)
Ryan,V. .............................................075 (E)
Saah, J. R. ..........................................183 (E)
Sabourin, C. L. ..................................064 (D)
Sadat, E. .............................................187 (F)
Salerno, G. ...............................................226
Samreen, A. .......................................141 (A)
Sanchez, M. .......................................073 (E)
Sankovich, K. A. ................................064 (D)
Santiago, A. ........................................049 (B)
Sarrazine, S. A. ...................................057 (B)
Schaefer, L. ........................................059 (B)
Scheuermann, R. H. ..........................187 (F)
Schmaljohn, C. ..................................082 (G)
Schneewind, O. ...................039 (B), 041 (B),
.........051 (B), 081 (G), 087 (H), 118, 152 (B),
...........153 (B), 155 (B), 163 (B), 169 (B), 234
Schneider, C. ........................092 (H), 097 (I)
Schnell, F. ..........................................195 (G)
Schnell, M. ........................................207 (K)
Schotzinger, R. ........................................229
Schriewer, J. ......................................196 (G)
Schupp, J. ...........................................189 (F)
Schweizer, H. P. .......................................014
Scully, C. ...........................................082 (G)
Self, J. S. ..................................................109
Semenova,V. ......................................067 (D)
Semenova,V. A. ..................................210 (I)
Senina, S. .................................................114
Sergueev, K.V. ......................050 (B), 066 (D)
Seshu, J. ...................................................131
Settlage, R. ........................................188 (F)
Sevilla-Reyes, E. E. ..................................022
Sewald, K. ...........................................214 (I)
Shadomy, S. .......................................067 (D)
Shafagati, N. ............................................115
Shaffer, S. A. .............................................116
Shah, K. ............................................196 (G)
Shah, S. R. ........................................071 (E)
Shamloul, M. .....................................197 (K)
Sharma, A. ....................199 (K), 206 (K), 222
Sharma, S. .........................................086 (H)
Sharpe, D. C. ....................................220 (K)
Shearer, J. ..........................................091 (H)
Sherchan, S. P. ...................................164 (B)
Sherratt, D. ........................................035 (B)
Sherwood, R. ....................................201 (K)
Sherwood, R. L. ....................092 (H), 097 (I)
Shields, M. .........................................171 (B)
Shin,Y.-W. ..........................................096 (I)
Shinmura,Y. .....................................090 (H)
Shirey, K. ...........................................049 (B)
Shirley, R. .........................................088 (H)
Shrutleff, A. .............................................227
Shuman, H. A. ..................................194 (G)
Shuttleworth, H. ................................161 (B)
Siefkas, K. ..........................................061 (B)
Silvera, P. ...........................................196 (G)
BIODEFENSE PROGRAM & ABSTRACTS
AUTHOR INDEX
Silvera, P. M. .....................................200 (K)
Silvestri, E. .........................................162 (B)
Simpson, A. J. ....................................198 (K)
Singh, M. ..........................................036 (B)
Smagin, A. .........................................062 (D)
Smith, J. ............................................082 (G)
Smith, Q. B. .............................................224
Smith, T. L. ........................................067 (D)
Soarimalala,V. L. ......................................112
Soler-Bistué, A. ..................................035 (B)
Solomonia, R. ..................................047 (B)
Sorenson, J. ........................................217 (K)
Soroka, S. ............................................210 (I)
Spaulding, U. K. ................................075 (E)
Sperandio,V. ............................................016
Spurgers, K. ......................................206 (K)
Spurgers, K. B. ...................................208 (K)
Squires, R. B. .....................................187 (F)
Stahl-Hennig, C. ...............................203 (K)
Stanker, L. H. ...........................................111
Stein, K. ............................................196 (G)
Stephansson, O. .................................055 (B)
Stevens, M. P. .....................................154 (B)
Steward-Clark, E. ..............................067 (D)
Stinchcomb, D. ..................................205 (K)
Stoddard, R. A. ..................................067 (D)
Stojadinovic, A. ..................................053 (B)
Stokes, M. G. .....................................198 (K)
Storch, S. ....................................201 (K), 228
Topic Categories:
Below is a list of descriptions of topic cate-
gories (letters in parentheses by each pres-
entation number refer to this chart).
A. Viral Agents
B. Bacterial Agents
C. Fungal Agents
D. Diagnostics
E. Environmental Detection
F. Informatics and Genomics
G. Therapeutics
H. Vaccines
I. Immune Responses
J. Decontamination, Biosafety,
and Containment
K. Other and Information Only
107
AUTHOR I N DE X
Listed by Author Name, Presentation Number, Abstract Category. *Presenting author is in Bold Face Type.
Stratton, D. ........................................043 (B)
Strohmeier, O. ...................................065 (D)
Stuthman, K. ...........................................227
Suar, M. .............................................157 (B)
Sullivan, D. ........................................061 (B)
Sun,Y. .....................................................117
Surnev, D. ........................................084 (H)
Sutton,V. ...........................................145 (A)
Svoboda, P. .........................................210 (I)
Swearengen, J. .........................................005
Swiderski, W. .....................................091 (H)
Szabo, J. ...................................................127
Taft, S. C. ..........................................162 (B)
Tallon, L. ...........................................079 (F)
Tan, J. ................................................028 (A)
Tauxe, R. ................................................024
Taviani, E. ..........................................185 (E)
Tayyab, S. M. H. .................................077 (F)
Tello, C. A. ........................................060 (B)
Tempel, R. .......................................088 (H)
Tettelin, H. ........................................079 (F)
Theis, T. ...........................................175 (D)
Thomas, J. ..........................................212 (I)
Thomas, J. G. ....................................068 (D)
Thomas, M. .......................................171 (B)
Thompson, I. J. .................................198 (K)
Thomson, G. ...........................................023
Thorkildson, P. ..................................156 (B)
Timlin, J. A. .............................................224
Tolley, H. ...........................................142 (A)
Tolmasky, M. ....................................035 (B)
Tonks, K. M. ......................................161 (B)
Torres, A. .................................................233
Tournier, J.-N. ...........................204 (K), 223
Traglia, G. ..........................................165 (B)
Tran, M. ............................................028 (A)
Tran, T. ..............................................035 (B)
Tran-Gyamfi, M. ...............................069 (D)
Trefethen, J. .......................................197 (K)
Trescos,Y. ..........................................204 (K)
Tret, J. O. ...........................................082 (G)
Trivedi, L. .........................................178 (D)
108 10th ASM Biodefense and Emerging Diseases Research Meeting
Tsai, P. ................................................095 (I)
Tsanava, S. ............................046 (B), 159 (B)
Tsanava, S. .........................................168 (B)
Tsanava, S. A. .....................................070 (E)
Tsang, C. H. ......................................161 (B)
Tsertsvadze, N. .....................046 (B), 159 (B)
Tsertsvadze, N. ..................................168 (B)
Tsertsvadze, N. S. ...............................070 (E)
Tuanyok, A. .......................................057 (B)
Turell, M. J. ........................................078 (F)
Turner, J. ...........................................208 (K)
Twenhafel, N. A. ................................034 (B)
Udoh, I. P. .........................................147 (B)
Uemura, C. .......................................090 (H)
Uhlenhaut, C. ...................................219 (K)
Ulianova, O.V. ...................................166 (B)
Uzarski, J. ...........................................101 (J)
Valderas, M. W. ......................092 (H), 097 (I)
Valle, E. .............................................171 (B)
van der Bruggen, K. ................................249
van Hoek, M. L. ......052 (B), 054 (B), 083 (G)
van Oordt, T. .....................................065 (D)
Van Tongeren, S. ......................................227
Van Zandt, K. E. ...............................057 (B)
VanderNoot,V. ..................................069 (D)
VanDuyne, R. .........................................114
Vassar, M. ..........................................091 (H)
Vay, C. ...............................................165 (B)
Veres, T. ..............................................214 (I)
Verrier, B. ..........................................204 (K)
Vincent, G. ........................................190 (F)
Vipond, J. ................045 (B), 149 (B), 161 (B)
Vipond, R. ........................................190 (F)
Vishwakarma,V. ................................157 (B)
Vogel, S. ............................................049 (B)
Volland, H. .......................................184 (E)
Volland, H. ........................................186 (E)
von Stetten, F. ...................................065 (D)
Voth, D. ...................................................254
Wagner, D. ..............................................012
Wagner, D. .........................................168 (B)
Walper, S. A. .....................................179 (D)
Wang, D. .................................................009
Wang, X. .................................................116
Wang,Y.-T. ..................153 (B), 155 (B), 234
Warawa, J. M. ...................................117, 246
Ward, S. .............................................202 (K)
Warren, R. ........................................193 (G)
Warren, R. L. .....................................057 (B)
Warren, T. ................................................227
Waters, C. .........................................068 (D)
Waters, C. ................................................138
Way, M. ...................................................244
Weaver, S. C. ...........................................240
Weidmann, M. ..................................065 (D)
Weir, J. ................................................213 (I)
Weiss, M. L. .......................................220 (K)
Welch, W. ..........................................156 (B)
Wells, J. ...................................................227
Whitehouse, C. A. ................078 (F), 148 (B)
Wilder, J. ...............................092 (H), 097 (I)
William, P. .........................................200 (K)
Williams, D. ......................................062 (D)
Williams, P. .......................................150 (B)
Williams, R. .........................029 (A), 030 (A)
Williams, S. K. ...................................148 (B)
Williamson, E. D. ...............................198 (K)
Wirblich, C. ......................................207 (K)
Wobus, C. ...............................................120
Wölfel, R. .........................063 (D), 181 (E)
Wolfraim, L. ......................................085 (H)
Wolfraim, L. ......................................209 (K)
Wong, K. K. .....................................143 (A)
Wood, J. ..................................................125
Woodman, M. E. ................................211 (I)
Wooten, R. M. ...................................211 (I)
Worth, R. G. .......................................211 (I)
Wu, W. ................................................216 (J)
Wycoff, K. .........................................059 (B)
Xiao, L. ...............................................212 (I)
Yang, A. ...................................................222
Yao, H. ....................................................226
Yates, C. ..................................................229
Ye, L. ..................................................098 (I)
 AUTHOR INDEX

Listed by Author Name, Presentation Number, Abstract Category. *Presenting author is in Bold Face Type.

Yeager, J. ................029 (A), 030 (A), 038 (B)

Yee, S. B. ...........................................034 (B)
Yin, J. ................................................080 (G)
Yokoi, K. ..........................................090 (H)
Yokote, H. ........................................090 (H)
yole, D.Y. S. NA. .................................094 (I)
Young, G. ..........................................205 (K)
Yu, Kuznetsov ...................................084 (H)
Yue, C. ..............................................203 (K)
Yung, P. ..............................................216 (J)
Yusibov,V. .........................................197 (K)
Zakalashvili, M. .................................159 (B)
Zaremba, S. .......................................187 (F)
Zelazowska, E. B. ...............................170 (B)
Zhang,Y. ............................................187 (F)
Zhgenti, E. ...........................047 (B), 159 (B)
Zhghenti, E. ......................................168 (B)
Zhou, L. ............................................187 (F)
Ziese, M. ...........................................028 (A)
Zorreguieta, A. ..................................035 (B)
Zumbrun, E. ........................029 (A), 030 (A)
Zumbrun, E. E. ................................146 (A)

Topic Categories:
Below is a list of descriptions of topic cate-
gories (letters in parentheses by each pres-
entation number refer to this chart).

A. Viral Agents
B. Bacterial Agents
C. Fungal Agents
D. Diagnostics
E. Environmental Detection
F. Informatics and Genomics
G. Therapeutics
H. Vaccines
I. Immune Responses
J. Decontamination, Biosafety,
and Containment
K. Other and Information Only

BIODEFENSE PROGRAM & ABSTRACTS 109

NOTE S

110 10th ASM Biodefense and Emerging Diseases Research Meeting

NOTE S

BIODEFENSE PROGRAM & ABSTRACTS 111

NOTE S

112 10th ASM Biodefense and Emerging Diseases Research Meeting

 A M E R I C A N S O C I E T Y F O R M I C R O B I O L O G Y

ASM TELECONFERENCES
March 7, 2012
Optimizing Specimen Management
J. Michael Miller

March 28, 2012

Medical Entomology: Arthropod Pests of Public
Health Importance
Chris Evans

April 11, 2012

CLIA: Past, Present, and a Glimpse of the Future ASM CONFERENCES
Nancy Anderson

May 16, 2012

March 29–April 2, 2012
What Does A Culture of Safety Mean in a Clinical
11th ASM Conference on Candida and Candidiasis
Microbiology Lab?
San Francisco, California
Rebecca T. Horvat
May 6–10, 2012
August 29, 2012
4th ASM Conference on Prokaryotic Cell Biology
Tools for Identification of Mycobacteria in 2012
and Development
Susan M. Harrington
Montreal, Canada
September 26, 2012
June 26–29, 2012
HIV Laboratory Diagnosis: Current and
3rd ASM Conference on Antimicrobial Resistance in
Future Methodology
Zoonotic Bacteria and Foodborne Pathogens
Michele Owen
Aix–en–Provence, France
October 24, 2012
September 29–October 4, 2012
Emergency Preparedness
6th ASM Conference on Biofilms
Thomas Williams
(ASM Biofilms 2012)
Miami, Florida
www.asmaudio.org
October 22–26, 2012
4th ASM Conference on Beneficial Microbes
San Antonio, Texas

www.asm.org/conferences