Microorganisms require nutrition and artificial habitat to grow in a microbiological laboratory. Nutrition is provided in the form of culture media and other additives. In order to check the effectiveness of culture media, growth promotion test is used. Growth promotion test ensures that culture media is fit for microorganism’s growth or not.

Growth promotion test is performed on culture media only. In this article, I will explain how to perform this test in easy steps.

Types of Growth Promotion Test

Quantitative Analysis

Used for the media that is used for enumeration testing

• Tryptic Soy Agar
• Sabourad Dextrose Agar
• R2A Agar

Semi-Quantitative Analysis (Agar)

Used for selective media

• Mannitol Salt Phenol Red Agar
• Cetrimide Agar
• Triple Sugar Iron Agar
• Levine EMB Agar
• XLD Agar

Semi-Quantitative Analysis (Broth)

Used for media broths

• Lactose Broth
• Selenite Cestine Broth
• Mossel Broth
• Malachite Green Broth
• Peptone
• Tryptic Soy Broth

Procedure

Test Preparation

Prepare culture media as per labeled instructions, in order to check its ability to promote the growth of specific organisms.

Prepare culture suspension of specific organisms.

Innoculation

Inoculate portions of the test culture media in plates, tubes, and bottles with 0.1 ml of specific microbial culture suspension (containing 10-100 CFU).

For the selection of organisms and methods of inoculation, see the Table below.

Inoculation Method and Culture Media Used

Culture Media	Test Organism	Inoculation Method
Tryptic Soy Agar	S. aureus (ATCC 6538) B. subtilis (ATCC 6633) E. coli (ATCC 8739) C. albicans (ATCC 10231)	Pour Plate Method
Tryptic Soy Broth	P. aeruginosa (ATCC 9027) S. aureus (ATCC 6538) B. subtilis (ATCC 6633) C. albicans (ATCC 10231)	Pour Plate Method
Sabouraud Dextrose Agar	C. albicans (ATCC 10231)	Pour Plate Method
Levine EMB Agar	E. coli (ATCC 8739)	Streaking

Use one or more environmental isolates if available, preferably in the media used for environmental monitoring, enumeration test of water and products, etc.

Incubation

Incubate the inoculated media at recommended temperatures specified for the organism as per Table incubation temperature of test organisms.

Incubation Temperature of Test Organisms

Test Organism	Time	Temperature
C. albicans (ATCC 10231)	03 days	22.5 + 2.5 °C
S. typhi (ATCC 14028)	48 hours	32.5 + 2.5 °C
P. aeruginosa (ATCC 9027)	48 hours	32.5 + 2.5 °C
E. coli (ATCC 8739)	48 hours	32.5 + 2.5 °C
S. aureus (ATCC 6538)	48 hours	32.5 + 2.5 °C
B. subtilis (ATCC 6633)	48 hours	32.5 + 2.5 °C

Similarly, incubate the un-inoculated media as a negative control for sterility check at a specified temperature for a time specified as per Table above.

There should not be present any microbial contamination in the sterility check portions of media by the completion of the incubation period.

Result Interpretation

Observe and note the result in your relevant notebook.

Write the interpretation by comparing the acceptance criteria mentioned in Interpretation Table.

Record the details of the growth promotion test, sterility and inhibition tested for each autoclaved media lot as per Table.

Perform a growth promotion test on each received dehydrated lot of culture media.

Growth should be clear and visible in the form of turbidity in the case of liquid media and distinct colony formation in the case of agar medium.

Growth promotion results should be comparable to the previously approved batch of the media.

Interpretation Table

Media	Test Organism	Acceptance Criteria
Tryptic Soy Agar	S. aureus (ATCC 6538) B. subtilis (ATCC 6633) E. coli (ATCC 8739) C. albicans (ATCC 10231)	No. of colonies must be within a fracture of two of the no. of colonies on the previously approved medium > 70% recovery
Sabouraud Dextrose Agar	C. albicans (ATCC 10231)	> 70% recovery
Tryptic Soy Broth	P. aeruginosa (ATCC 9027) S. aureus (ATCC 6538) B. subtilis (ATCC 6633) C. albicans (ATCC 10231)	Visible growth should be observed
R2A Agar	S. typhi (ATCC 14028) E. coli (ATCC 8739)	> 70% recovery