Nonfermenting and

Miscellaneous
Gram-Negative Bacilli
Rochelle D. Darlucio, RMT, MPH
Our Lady of Fatima University
College of Medical Laboratory Science
General Characteristics of Nonfermenters
• fail to acidify an oxidative-fermentative (OF) medium when it is
overlaid with __________

• fail to acidify __________________ butts

• prefer and grow much better in an ____________

• Most are oxidase positive

Biochemical Characteristics and Identification
• ____________ reaction, although reaction can be weak and variable

• Nonreactivity in 24 hours in commercial multitest kit systems used

primarily for the identification of Enterobacteriaceae

• ________________in the slant or butt of TSIA or KIA

• Resistance to a variety of classes of antimicrobial agents, such as

aminoglycosides, third-generation cephalosporins, penicillins, and
fluoroquinolones
Clinically Significant Nonfermentative,
Gram-Negative Bacilli
• Pseudomonas
▫ Gram-negative bacillus or coccobacillus
▫ Strictly aerobic metabolism
▫ Motile usually with _________ or polar tuft of
flagella
▫ Oxidase positive (except ______and __________)
▫ Catalase positive
▫ Usually grows on MAC agar
▫ Usually an oxidizer of carbohydrates, but some species
are asaccharolytic
Pseudomonas Fluorescent Group
• Pseudomonas aeruginosa • Pseudomonas aeruginosa
▫ Causes the following infections:
 Severe wound infections in burn • Virulence Factors
patients
 UTI & Nosocomial Pneumonia
 Septicemia in immunosuppresed
patients and infants
 Chronic Lung Infections in Cystic
Fibrosis patients
 Septic arthritis in intravenous
drug abusers
 Destructive eye infection (keratitis
& corneal ulcers) in contact lens
wearers
 Swimmer’s ear – especially in
athletes
 Jacuzzi/Hot Tub Syndrome –
SHANGHAI FEVER
Pseudomonas Fluorescent Group
• Pseudomonas aeruginosa
▫ Identifying Characteristics

• P. aeruginosa
• P. fluorescens Produce pyoverdin
• P. putida •a yellow-green or
• P. veronii yellow-brown
• P. mosselii pigment.
• P. Monteilii

• P. aeruginosa- also produce pyocyanin

Pseudomonas Fluorescent Group
• Pseudomonas aeruginosa

▫ Large, mucoid, spreading colonies

▫ CETRIMIDE Agar – transparent selective culture media that aids in
pigment production visualization
▫ OXIDASE + to differentiate it from Family Enterobacteriaceae & other
enterics
▫ Oxidative
 Dextrose (Glucose) but NOT Maltose
▫ Fermentative
 TSI Reaction: K/K
▫ Growth @ 420C – to differentiate it from the rest of the pseudomonads
▫ Acetamide Utilization + (blue slant) after 7 days of standard incubation
Pseudomonas Non- Fluorescent Group
• Pseudomonas stutzeri
• wrinkled, leathery, adherent
colonies that may produce a _____
________________________
• ADH negative
• starch hydrolysis positive
• responsible for diseases that include
septicemia, meningitis in the human
immunodeficiency virus-infected patient,
pneumonia, endocarditis, postsurgical
wound infections, septic arthritis,
conjunctivitis, and UTIs.
• Pseudomonas mendocina
• nonwrinkled, flat colonies that may
appear with _____________
______________________
• ________________appearance.
• oxidase and ADH positive, and is
acetamide negative.
• motile by means of a single polar
flagellum
• oxidizes glucose and xylose
• nonproteolytic
• does not hydrolyze starch.
 Pseudomonas Non- Fluorescent Group
• Pseudomonas
pseudoalcaligenes and
Pseudomonas alcaligenes
• oxidase positive
• grows on MAC agar
• variable in the reduction of
nitrates to nitrites or nitrogen
gas.
• motile by means of a polar
flagellum.
• Pseudomonas luteola and
• Pseudomonas oryzihabitans
• oxidase-negative
• catalase positive
• Motile
• oxidize glucose
• grow on MAC agar
• often produce an intracellular
nondiffusible ____________.
• _____________________at
48 hours
 Acinetobacter
• Aerobic gram-negative coccobacilli
or even gram-negative cocci
• _____________and retain the
crystal violet stain, leading to
misidentification.
• appear as gram-positive cocci in
smears made from blood culture
bottles.
• oxidase negative,catalase positive,
and nonmotile.
• ______________is saccharolytic,
and ____________is
asaccharolytic.
Stenotrophomonas maltophilia
• oxidase-negative, nonfermentative,
gramnegative bacillus
• Colonies may appear _________ on MAC agar
• positive for catalase, DNase, esculin and gelatin
hydrolysis, and lysine decarboxylase
• Usually susceptible to SXT
Burkholderia
• Burkholderia cepacia Complex
• often produces a weak, slow, positive
oxidase reaction.
• Almost all strains oxidize glucose, and
many will oxidize maltose, lactose, and
mannitol.
• Most strains are lysine decarboxylase and
ONPG positive
• most strains are ornithine decarboxylase
negative and fail to reduce nitrate to
nitrite.
• motile by means of polar tufts of flagella
• Causes ___________in plants and
________________ in humans
• Burkholderia mallei
• Smooth and cream to white colonies in
BAP and weakly oxidase (+)
• The only non-motile among all
pseudomonads
• _____________________
• Causes ________________– infectious
disease of horses, goats, sheep, & donkey.
Rare cause of human infection acquired by
direct contact, trauma, or inhalation.
• _________– disseminated form of
glander’s disease
Burkholderia
• Burkholderia pseudomallei
• ___________________in BAP
• Motile via polar tuft flagella
• ADH (+)
• Highly oxidative – glucose, maltose,
lactose, mannitol
• Causes ______________ – glanderslike
disease which has a long latent period.
Manifestation of symptoms takes years
hence the synonym Vietnam Time Bomb
• Burkholderia gladioli
• Produces ____________after 48-72
hours of incubation
• motile by means of one or two polar
flagella
• Catalase and urease positive.
• grows on MAC agar
• Oxidizes glucose
• mannitol positive and decarboxylase
negative, and is negative for oxidase,
although some strains are weakly positive.
Less Commonly Encountered Nonfermentative,
Gram-Negative Bacilli
• Alcaligenes and Achromobacter • Brevundimonas

• B. diminuta
▫ motile and possess a single polar
flagellum
▫ oxidize glucose
▫ oxidase positive.
▫ Most strains grow on MAC agar.

• B. vesicularis
▫ slender rod, with polar flagella.
▫ Only about 25% grows on MAC agar.
▫ Most strains of produce an
________ intracellular pigment.
▫ oxidase positive and oxidizes glucose
and maltose.
Less Commonly Encountered Nonfermentative,
Gram-Negative Bacilli
• CDC Groups EO-3, EO-4, and
Paracoccus
• oxidase-positive
• nonmotile, saccharolytic coccobacilli
that grow weakly, if at all, on MAC
agar.
• All oxidize glucose and xylose
• EO-3 and many EO-4 isolates have a
________________________.
• Chromobacterium
• Chromobacterium violaceum
▫ Motile, facultative anaerobe,
OXIDASE (+)
▫ Rare cause of human infection.
▫ Found in soil and water
▫ Unique because of the violet pigment
that it produces – ___________

• P. yeei (EO-2) is further characterized

by the production of characteristic
______ or ___________on Gram
stain
Less Commonly Encountered Nonfermentative,
Gram-Negative Bacilli
• Comamonas and Delftia
• straight to slightly curved rods
• produce alkalinity in OF media
• catalase and oxidase positive,
• motile by multitrichous polar flagella
• reduce nitrate to nitrite.
• Comamonas testosteroni and Comamonas
terrigena
▫ have been reported to cause nosocomial bacteremia
• Delftia acidovorans
▫ associated with keratitis in soft contact lens wearers
and nosocomial infections including bacteremia
and endocarditis.
• Delftia tsuruhatensis
▫ Associated with catheter-related bacteremia
• Flavobacteriaceae
▫ Balaneatrix, Bergeyella,
Chryseobacterium, Elizabethkingia,
Empedobacter, Myroides, Weeksella,
Wautersiella, Sphingobacterium spp
• Elizabethkingia meningoseptica.
• meningitis or septicemia in a newborn,
especially in conjunction with prematurity
• Balneatrix alpaca
• First isolated in 1987 during an outbreak of
pneumonia and meningitis linked to
individuals attending a hot springs spa.
Less Commonly Encountered Nonfermentative,
Gram-Negative Bacilli
• Methylobacterium and • Ralstonia and Cupriavidus
Roseomonas ▫ linked to meningitis,
endocarditis, and osteomyelitis.
• ____________________ • Shewanella
▫ often mucoid and can produce a
______________ causing
________ discoloration of
SBA.
• Sphingomonas
▫ isolated from many water
sources, including swimming
pools, as well as from hospital
equipment and laboratory
supplies.
References:
• Study Guide on Diagnostic Bacteriology by Mr. Nathaniel Rañon
• Textbook of Diagnostic Microbiology (6th edition) by Connie Mahon