Growth promotion test for culture media

Growth promotion test for culture media

1.0       OBJECTIVE

To lay down a standard procedure for growth promotion test of culture media.

2.0       SCOPE

This Standard Operating Procedure is applicable to the technical staff of Microbiology laboratory for growth promotion test of culture media.

3.0       RESPONSIBILITY

            Microbiologist shall be responsible to follow the procedure mention in this SOP.

4.0       ACCOUNTABILITY

Department Head & QA Head shall be accountable for effective implementation of this SOP.

5.0       ATTACHMENTS

List of Micro-organisms used for growth promotion test                          – Attachment-I

Format for the record of growth promotion test of culture media             – Attachment-II

6.0       PROCEDURE

6.1           Precautions

6.1.1        The dehydrated culture media as well as their ingredients are highly hygroscopic and must be stored

in a cool dry place away from bright light

6.1.2        Use fresh sterile pipette for each transfer.

6.1.3        If any spillage of cultures, immediately wash with 70% IPA Solution.

6.1.4        Always use dry spatula for weighing the dehydrated media. The weighing operation shall be

completed as quickly as possible to prevent absorption of moisture by the hygroscopic contents.

6.1.5        Always wear the face mask while weighing the dehydrated media to avoid inhalation of fine particles

of media.

6.2           Test for growth Promoting Properties of agar media

6.2.1        Label the plates with culture name, Media B.No., date of incubation on the base of petri plate.

6.2.2        Select the quantified microbial culture as per attachment-I for the growth promotion test.

6.2.3        Add 1.0ml suspension of specific culture containing 10 to 100 CFU/ml into two sterile petri plate.

6.2.4        Aseptically pour the cooled media at 40 to 45 ºC into both the labeled plates, mix the plates by gently

rotating clockwise and anticlockwise direction. Allow the plates to solidify at room temperature.

6.2.5        Simultaneously run a negative control to verify testing conditions, using the same procedure in place

of the test organism use diluents i.e. 1.0ml of saline solution.

6.2.6        Incubate the plates for 24 to 72 hours for the bacterial count 30-35ºC and 5 to 7 days for fungal count

at 20-25 ºC.

6.2.7        After incubation observe the plates for number of microbial colonies, and express the number of cfu

by following formula and record the average results in the attachment-II.

P1+P2

2

Where P1 and P2 are plate 1 and plate 2.

6.2.8         Calculate the microbial recovery in percentage by equation-

%Recovery = Mean cfu observed X100

Inoculated CFU/ml

6.2.9         Recovery should not less than 75%.

6.2.10       Frequency : for each batch of consignment.

6.3            Test for growth promoting properties of Broth Medium

6.3.1         Prepare required quantity of liquid culture media, dispense 100ml in test tube and sterilize as per

manufacturer’s instructions.

6.3.2         After sterilization transfer the media to LAF room and allow it to cool at room temperature.

6.3.3         Start the LAF as per SOP and proceed further work under LAF.

6.3.4         Add 1.0 ml of positive culture for growth promoting properties, containing 100 CFU cells into broth

tube and label with media B. No., Name of positive control and date of inoculation.

6.3.5         Incubate the tubes at 30-35ºC for 24-48 hours for bacterial count and at 20-25ºC 5-7 days for fungal

count.

6.3.6         Daily observe the tubes for growth for turbidity.

6.3.7         Satisfactory growth should be observed within 3 days of incubation in the test.

6.3.8        In case the media passes the growth promotion test, a approved label shall be affixed on the media

container, then the same should be used for analysis.

6.3.9        In case the media fails for growth promotion test then a rejected label shall be affixed on the

container and then the same shall be rejected and accordingly the rejection entry should be made in

the stock register.

6.3.10      The rejected media should be returned back to the supplier.

7.0       REFERENCES

In- house

8.0       ABBREVIATIONS

SOP: Standard operating procedure

°C     : Degree Celsius

Psi    :  Pound per square inch

I.P.A.: Isopropyl Alcohol

CFU: Colony forming unit

LAF: Laminar air flow

QA: Quality Assurance

QC: Quality Control

9.0       DISTRIBUTION LIST

Quality Assurance Department

Quality Control Department

10.0     HISTORY OF REVISION

Version Number Effective Date Reason for Revision

 

List of Micro-organisms used for growth promotion test                          – Attachment-I

S.No.

 

Media Micro-organisms used for growth promotion test
01

 

Soyabean Casein Digest Agar E.coli, p.aeruginosa Bacillus subtilis S.aureus, Salmonella spp. A.brasiliensis Candida albicans
02

 

Soyabean Casein Digest Medium E.coli Bacillus subtilis S.aureus A.brasiliensis Candida albicans
03

 

Sabouraud  Dextrose Agar A.brasiliensis Candida albicans N.A. N.A. N.A.
04

 

Cetrimide Agar P.aeruginosa N.A. N.A. N.A. N.A.
05

 

Mannitol Salt Agar S.aureus N.A. N.A. N.A. N.A.
06

 

MacConkey Agar E.coli N.A. N.A. N.A. N.A.
07

 

Xylose Lysine Deoxycolate Agar Salmonella sp. Shigella spp. N.A. N.A. N.A.
08

 

Baired  Parker Agar S.aureus N.A. N.A. N.A. N.A.
09

 

MacConkey broth E.coli N.A. N.A. N.A. N.A.
10

 

Bismuth sulfite agar Salmonella sp. N.A. N.A. N.A. N.A.
11

 

Nutrient agar E.coli Bacillus subtilis S.aureus Salmonella p.aeruginosa
12

 

Nutrient broth E.coli Bacillus subtilis S.aureus A.brasiliensis Candida albicans
13

 

Peptone water E.coli Salmonella spp. N.A. N.A. N.A.

Format for the record of growth promotion test of culture media             – Attachment-II

S. No  Date of analysis Name of Media Batch No. Use Before Container  No. issued No. of colonies Done By Checked by Remarks

 

                                                                   

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