Table 1: Determination of coliform

Microorganism:
 Dilution

factor 10-1 10-2 10-3 10-4 10-5

Plate 1 + + + - -

Plate 2 + + - - -

Table 2: Observation of gas production inVRBA broth after 48 hours

Microorganism:

Positive/negative (+/-) of gas production

Dilution factor
10-1 10-2 10-3 10-4 10-5

Tube 1 + + - - -

Tube 2 + + - - -

Tube 3 + - - - -

Number of positive tube(s) 3 2 0 0 0

Refer 3-tubes MPN Table (200= 0.091)
Calculate the number of CFUs per ml of original sample as follows:

The number of CFUs per ml of sample =

The number of colonies (25-250 plate) X
The dilution factor of the plate counted X
1/ amount of sample in the plate

10-1 10-2 10-3

=0.091 X 101 X =0.091 X 102 X =0.091 X 103 X

=9.1x 10-1 CFu/ml =9.1 CFu/ml =91 CFu/ml

10-4 10-5 10-6

=0.091 X 104X =0.091 X 105 X =0.091 X 106 X

=9.1x 102 CFu/ml =9.1x 103 CFu/ml =9.1x 104 CFu/ml

References:

1. Feng, P., Weagant, S. D., Grant, M. A., Burkhardt, W., Shellfish, M., & Water, B. (2002).
BAM: Enumeration of Escherichia coli and the Coliform Bacteria. Bacteriological
analytical manual, 13-19.
2. Bell C., Neaves P. and Williams A.P. (2005). Food Microbiology and Laboratory Practice,
Wiley-Blackwell, Oxford.
3. Garg N. and Garg K.L. (2010). Laboratory Manual of Food Microbiology, IK
International Publishing House Pvt. Ltd.
4. Edberg, S. C., Allen, M. J., Smith, D. B., & Kriz, N. J. (1990). Enumeration of total
coliforms and Escherichia coli from source water by the defined substrate
technology. Applied and Environmental Microbiology, 56(2), 366-369.

5. Peter Feng & Stephen D. Weagant (2002 September) Enumeration of Escherichia

coli and the Coliform Bacteria [Online].

[Accessed 3th January, 2018]. Available from World Wide Web:

https://www.fda.gov/Food/FoodScienceResearch/LaboratoryMethods/ucm064948.htm
 APPENDIX:
3-tubes MPN Table

No. of Tubes Positive in MPN in the

inoculum of the 2 0 0 0.091
first middle last middle set of 2 0 1 0.14
set set set tubes 2 0 2 0.20
0 0 0 ‹0.03 2 0 3 0.26
0 0 1 0.03 2 1 0 0.15
0 0 2 0.06 2 1 1 0.20
0 0 3 0.09 2 1 2 0.27
0 1 0 0.03 2 1 3 0.34
0 1 1 0.061 2 2 0 0.21
0 1 2 0.092 2 2 1 0.28
0 1 3 0.12 2 2 2 0.35
0 2 0 0.062 2 2 3 0.42
0 2 1 0.093 2 3 0 0.29
0 2 2 0.12 2 3 1 0.36
0 2 3 0.16 2 3 2 0.44
0 3 0 0.094 2 3 3 0.53
0 3 1 0.13 3 0 0 0.23
0 3 2 0.16 3 0 1 0.39
0 3 3 0.19 3 0 2 0.64
1 0 0 0.036 3 0 3 0.95
1 0 1 0.072 3 1 0 0.43
1 0 2 0.11 3 1 1 0.75
1 0 3 0.15 3 1 2 1.2
1 1 0 0.073 3 1 3 1.6
1 1 1 0.11 3 2 0 0.93
1 1 2 0.15 3 2 1 1.5
1 1 3 0.19 3 2 2 2.1
1 2 0 0.11 3 2 3 2.9
1 2 1 0.15 3 3 0 2.4
1 2 2 0.20 3 3 1 4.6
1 2 3 0.24 3 3 2 11
1 3 0 0.16 3 3 3 ›24
1 3 1 0.20
1 3 2 0.24
1 3 3 0.29