Table of Contents

sop:growth promotion test

OBJECTIVE:The objective of this SOP is to provide guidelines for performing the growth promotion test of the media being used.

SCOPE:This procedure is applicable for growth promotion test in Microbiology Laboratory.

RESPONSIBILITY: Microbiologist: to follow the process given as per SOP.

Q.C Head: to ensure the proper execution of the SOP.

Q.A: to ensure that the SOP is being properly implemented.

PROCEDURE:

Prepare the media being tested as per specification or instructions given by manufacturer.
Sterilize the media at 121°C for 15 minutes or as per specifications.
Distribute the sterilized agar medium in sterile Petri-plate in duplicate and allow them to solidify.
In case of broth medium, distribute the medium in sterilized test tube in duplicate.
Streak a loop-full culture of test microorganisms on solid media or inoculate in case of liquid media.
Incubate the plates / tubes at 30-35°C in case of bacteria and at 20 – 25°C in case of fungi for 2 to 5 days.
After incubation observe the plate for appropriate colony characteristics of particular strain as shown in table given below and record your observation.

No.	Solid Media (Agar/ Broth)	Micro-organism	Property	Incubation	Colony Characteristic / Observation
1.	Soyabean Casein Digest agar	Escherichia coli	Growth promoting	30-35°C for 48 hours	Small, round regular raised white colonies.
1.	Soyabean Casein Digest agar	Staphylococcus aureus	Growth promoting	30-35°C for 48 hours	Small, regular raised smooth, golden yellow colonies.
2	Saboraud’s Chloramphenicol Agar	Aspergillus niger	Growth promoting	20-25°C for 5 days	Black, mycelia colonies.
2	Saboraud’s Chloramphenicol Agar	Candida albicans	Growth promoting	20-25°C for 5 days	Big, round, off-white raised colonies.
3	Cetrimide Agar	Pseudomonas aeruginosa	Growth promoting	30-35°C for 48 hours	Medium shows greenish colonies.
3	Cetrimide Agar	Escherichia coli	Inhibitory	30-35°C for 48 hours	Nil
4	Pseudomonas Agar for Fluorescein	Pseudomonas aeruginosa	Growth promoting	30-35°C for 48 hours	Colorless to yellowish colonies with yellowish fluorescence in Ultra Violet light.
5	Pseudomonas Agar for Pyocyanin	Pseudomonas aeruginosa	Growth promoting	30-35°C for 48 hours	Greenish colonies with blue fluorescence Ultra Violet light.
6	Bismuth Sulphite Agar	Salmonella typhi	Growth promoting	30-35°C for 48 hours	Small, round raised black or green colonies.
8	Vogel Johnson Agar	Staphylococcus aureus	Growth promoting	30-35°C for 48 hours	Black colonies surrounded by yellow zone.
9	Mannitol Salt Agar	Staphylococcus aureus	Growth promoting	30-35°C for 48 hours	Yellow colonies with yellow zone.
9	Mannitol Salt Agar	Escherichia coli	Inhibitory	30-35°C for 48 hours	Nil
10	Baird parker Agar	Staphylococcus aureus	Growth promoting	30-35°C for 48 hours	Black shiny colonies surrounded by clear zone
11	Soyabean Casein Digest Medium	Staphylococcus aureus	Growth promoting	20-25°C For 2 to 7 days.	Luxuriant growth must be observed within 48 hours.
		Pseudomonas aeruginosa			Luxuriant growth must be observed within 48 hours.
		Candida albicans			Luxuriant growth must be observed within 2 to 7 days.
		Aspergillus niger			Luxuriant growth must be observed within 2 to 7 days.
12	Triple Sugar Iron Agar	Salmonella typhi	Growth promoting	30-35°C for 24 hours	Slant-Red colour of Medium, Butt-Acidic reaction Yellowing of Medium, Gas and H₂S Produced.
13	MacConkey Broth	Escherichia coli	Growth promoting & Indicative	30-35° C For 48 hours.	Acid-Purple colour change to Yellow, Gas-Gas observed in Durham’s Tube.
13	MacConkey Broth	Staphylococcus aureus	Inhibitory	30-35° C For 48 hours.	Nil
14	Selenite F Broth	Salmonella typhi	Growth promoting	30-35° C For 48 hours.	Luxuriant growth must be observed within 48 hours.
15	Tetrathionate Brilliant Green Bile Broth	Salmonella typhi	Growth promoting	30-35° C For 48 hours.	Luxuriant growth must be observed within 48 hours.
16	Nutrient Broth	Escherichia coli	Growth promoting	30-35° C For 48 hours.	Luxuriant growth must be observed within 48 hours.
16	Nutrient Broth	Salmonella typhi	Growth promoting	30-35° C For 48 hours.	Luxuriant growth must be observed within 48 hours.
17	Urea Broth	Salmonella typhi	Growth promoting	30-35° C For 48 hours.	Luxuriant growth must be observed within 48 hours.
18	Peptone Water	Escherichia coli	Growth promoting	30-35° C For 24 hours.	Luxuriant growth must be observed within 24 hours. Red ring at interface of medium on addition of Kovac reagent.
19	R2A Hiveg Agar	Escherichia coli	Growth promoting	30-35°C for 48 hours.	White colored small colonies observed
		Pseudomonas aeruginosa			White colored small colonies observed
		Salmonella typhi			White colored small colonies observed
		Staphylococcus aureus			White colored small colonies observed
19	Nutrient Agar	Escherichia coli	Growth promoting	30-35°C for 48 hours.	Small, round regular raised white colonies.
19	Nutrient Agar	Pseudomonas aeruginosa	Growth promoting	30-35°C for 48 hours.	Small, round regular raised white colonies.
20	MacConkey’s Agar	Escherichia coli	Growth promoting	30-35°C for 48 hours.	Pink to red colonies with bile precipitate which shows haziness around the colony
		Salmonella abony	Growth promoting		Colorless colonies observed
		Staphylococcus aureus	Growth inhibitory		Nil
21	Rappaport vassiliadis salmonella enrichment broth	Salmonella abony	Growth promoting	30-35°C for 24 hours.	Luxuriant growth observed with turbidity in the broth.
		Escherichia coli	Growth inhibitory		Nil
		Staphylococcus aureus	Growth inhibitory		Nil
22	Xylose lysine deoxycholate agar	Salmonella abony	Growth promoting	30-35°C for 48 hours.	Red colonies with black centers
		Shigella boydii	Growth promoting		Red color colonies
		Staphylococcus aureus	Growth inhibitory		Nil
23	Hugh Leifson medium	Escherichia coli	Growth promoting	30-35°C for 18-24 hrs.	Acid and gas production
		Pseudomonas aeruginosa	Growth promoting		Acid and gas production with yellow coloration (anaerobic condition) & no color change (aerobic condition)
		Salmonella abony	Growth promoting		Acid and gas production
24	MYP agar	B. cereus	Growth promoting	30-35°C for 18-40 hrs	Red color colony with opaque zone around it.
24	MYP agar	Escherichia coli	Growth inhibitory	30-35°C for 18-40 hrs	Nil
25	Voges Proskauer Medium	B. cereus	Growth promoting	30-35°C for 18-40 hrs	Positive reaction, purple color
26	Reinforced medium for clostridia	Clostridium sporogenes	Growth promoting	30-35°C for 18-24 hrs.	Luxuriant growth observed
26	Reinforced medium for clostridia	Staphylococcus aureus	Growth inhibitory	30-35°C for 18-24 hrs.	Nil
27	Columbia agar	Clostridium sporogenes	Growth promoting	30-35°C for 18-48 hrs.	Luxuriant
27	Columbia agar	Staphylococcus aureus	Growth inhibitory	30-35°C for 18-48 hrs.	Nil
28	Gelatin phosphate salt agar	Vibrio cholerae	Growth promoting	30-35°C for 18-48 hrs	Luxuriant transparent colonies with a cloudy halo
28	Gelatin phosphate salt agar	Staphylococcus aureus	Growth inhibitory	30-35°C for 18-48 hrs	Nil
29	TCBS Agar	Vibrio cholerae	Growth promoting	30-35°C for 18-24 hrs	Luxuriant yellow colonies
29	TCBS Agar	Escherichia coli	Growth inhibitory	30-35°C for 18-24 hrs	Nil
30	Frazer broth base	Listeria monocytogenes	Growth promoting	30-35°C for 24-48 hrs	Positive reaction, blackening of medium
30	Frazer broth base	Escherichia coli	Growth inhibitory	30-35°C for 24-48 hrs	Nil
31	Listeria Oxford Agar Base	Listeria monocytogenes	Growth promoting	30-35°C for 24-48 hrs	Positive reaction, blackening of medium around the colony
31	Listeria Oxford Agar Base	Escherichia coli	Growth inhibitory	30-35°C for 24-48 hrs	Nil
32	GN Broth	Salmonella abony	Growth promoting	30-35°C for 24-48 hrs	Luxuriant growth
		Shigella boydii	Growth promoting		Luxuriant growth
		Staphylococcus aureus	Growth inhibitory		Nil
33	Nitrate broth	Bacillus cereus	Growth promoting	30-35°C for 18-24 hrs	Luxuriant growth observed
33	Nitrate broth	Staphylococcus aureus	Growth inhibitory	30-35°C for 18-24 hrs	Nil
34	Motility medium	E. coli	Growth promoting	30-35°C for 24-48 hrs	Positive growth away from the stabline as seen by turbidity
34	Motility medium	Staphylococcus aureus	Growth inhibtory	30-35°C for 24-48 hrs	Nil
35	Enterobacteria enrichment broth mossel	E. coli	Growth promoting	30-35°C for 24-48 hrs	Luxuriant growth with change in coloration to yellow
35	Enterobacteria enrichment broth mossel	Staphylococcus aureus	Growth inhibtory	30-35°C for 24-48 hrs	Nil
36	Violet red bile glucose agar	E. coli	Growth promoting	30-35°C for 24-48 hrs	Pink color colonies with bile precipitate
36	Violet red bile glucose agar	Staphylococcus aureus	Growth inhibtory	30-35°C for 24-48 hrs	Nil

FREQUENCY:

As and when a new media is received in the microbiology laboratory

ANNEXURE:

Log book of growth promotion test
Protocol for growth promotion test

SOP Growth promotion test