Maintenance of Quality Control Strains

With proper maintenance, many microorganisms produced by Microbiologics can be used in quality control tests for
up to a month after reconstitution. In this technical bulletin, Microbiologics provides two plans for maintaining qual-
ity control microorganisms. In the first plan there are fewer passages for the microorganism. In the second plan, the
working stock culture is prepared weekly instead of monthly. When choosing a plan, take into consideration regula-
tory guidelines and test manufacturer’s directions for maintaining microorganisms.

Tips for maintaining microorganisms can be found in the boxes below.

Getting Started
A KWIK-STIK™ or a LYFO DISK® should be started on a non-selective agar such as Tryptic Soy Agar or Sheep
Blood Agar. Do not use broth.

Tips For Best Performance

Do not test the original pellet plate. This is the plate on which the lyophilized pellet was started. The organisms
growing on this plate are not fully resuscitated.
Always use fresh microorganisms for testing. Many tests require that the microorganisms are not more than 24
hours old.
Select isolated colonies for the test.
It may be necessary to start microorganisms used for quality control of antibiotic susceptibility tests every two
weeks because some microorganisms lose resistance over time. Examples are: P. aeruginosa ATCC® 27853™*
(Catalog # 0353), E. faecalis ATCC® 51299™* (Catalog # 0959), and S. pneumoniae ATCC® 49619™* (Catalog #
0947).
Do not test colonies from a contaminated plate.

Storage of Microorganisms
Fastidious microorganisms have shorter survival periods than aerobic bacteria. They will need to be subcul-
tured every few days. For example, Streptococcus pneumoniae and Neisseria gonorrhoeae need to be subcul-
tured every third day.
Store aerobic bacteria at 2-8°C. Exceptions are a few species of Bacillus which remain viable for a longer period
when stored at room temperature. Store microorganisms which require CO2 at room temperature in a candle
jar or a container with a CO2 packet. Store yeast and fungus at room temperature. Store anaerobes in anaero-
bic conditions at 2-8°C. Store Campylobacter at 35°C in microaerophilic conditions.
If the resuscitated culture is stored in conditions other than the 2-8°C (e.g. frozen), Microbiologics cannot guar-
antee the stated characteristics of the product.

Wrapping Up
After the fourth week, dispose of plates and start the process over with a new lyophilized pellet.
A microorganism may be used beyond expiry date if (one) the lyophilized pellet is grown before expiry date and
(two) the microorganism is not used beyond week 4 of the maintenance program.

*The ATCC Licensed Derivative Emblem, the ATCC Licensed Derivative word mark and the ATCC catalog marks are trademarks of ATCC.
Microbiologics, Inc. is licensed to use these trademarks and to sell products derived from ATCC® cultures.

TIB.246 REVISION A Page 1 of 3

 Microorganism Maintenance Plan A 1

Reference Stock Culture

Reconstitute microorganism
(see product insert PI.194)

Start on non-selective agar or slant

(see TIB.081 “Recommended Growth
Requirements”)

Monthly Working Stock Culture “Pellet Plate”2

Subculture Weekly to Non-selective Agar

Week 1/Day 1 Week 2/Day 8 Week 3/Day 15 Week 4/Day 22

Weekly Working Culture Weekly Working Culture Weekly Working Culture Weekly Working Culture

Each day of Week 1, Each day of Week 2, Each day of Week 3, Each day of Week 4,
subculture colonies from subculture colonies from subculture colonies from subculture colonies from
this plate to a fresh plate3 this plate to a fresh plate3 this plate to a fresh plate3 this plate to a fresh plate3

Day 2 Day 9 Day 16 Day 23

Day 3 Day 10 Day 17 Day 24

Day 4 Day 11 Day 18 Day 25

Day 5 Day 12 Day 19 Day 26

Day 6 Day 13 Day 20 Day 27

Day 7 Day 14 Day 21 Day 28

1. Reference: CLSI M22-A3 Vol. 24 No.19, Quality Control for Commercially Prepared Microbiological Culture Media: Approved Standard - Seventh Edition, April 2004.
2. Do not test colonies from the pellet plate. If it is necessary to the test the microorganism on Day 1, start the reference stock culture one day earlier and subculture it for
the first day of testing.
3. Alternatively, subculture the day before testing.
TIB.246 REVISION A Page 2 of 3
 Microorganism Maintenance Plan B 1

Reference Stock Culture

Reconstitute microorganism
(see product insert PI.194)

Use Daily Plates to set up panels for QC.3

Start on non-selective agar or slant
(see TIB.081 “Recommended Growth
Requirements”)

Week 1/ Pellet Plate/Day 12

Day 2 Day 3 Day 4 Day 5 Day 6 Day 7

Sub to new slant or

plate every 7 days

Week 2/Day 8
Weekly Working Culture

Day 9 Day 10 Day 11 Day 12 Day 13 Day 14

Week 3/Day 15
Weekly Working Culture

Day 16 Day 17 Day 18 Day 19 Day 20 Day 21

Week 4/Day 22
Weekly Working Culture

Day 23 Day 24 Day 25 Day 26 Day 27 Day 28

1. CLSI M07-A8 Vol. 29 No. 2, Methods for Dilution Antimicrobial Susceptibility Tests for Bacteria that Grow Aerobically: Approved Standard-Eighth Edition, January 2009.
CLSI M02-A10 Vol. 29 No. 1, Performance Standard for Antimicrobial Disk Susceptibility Tests; Approved Standard-Tenth Edition, January 2009.
2. Do not test colonies from the pellet plate. If it is necessary to the test the microorganism on Day 1, start the reference stock culture one day earlier and subculture it for
the first day of testing.
3. Alternatively , subculture the day before testing.

TIB.246 REVISION A Page 3 of 3