If for some reason solid medium cannot be used for culturing the bacteria, viable counts can be carried out using liquid media, by means the Most Probably Number (MAN) Technique. The sample to be counted is diluted until small aliquots contain an estimated one viable cell each.
If the dilution is correct, some aliquots will contain a viable cell, and other will not, so that after incubation, growth will occur in some tubes and not in others. If the sample has not been diluted sufficiently, all aliquots will contain viable cells and all inoculated tubes will show growth on incubation. If the sample has been diluted too much, none of the inoculated tubes will show growth. MAN method can be carried out by a 3 tube or 5 tube methods. Three successive tenfold-diluted samples are inoculated. After incubation the inoculated bubs showing positive growth are recorded.
With reference to the MAN table the most probable numbers of bacterial sample analyses can be obtained. RESULTS Figure 1: Observation of the plate 1 labeled 10-8 Figure 2: Observation of the plate 2 labeled 10-8 Figure 3: Observation of the plate 1 labeled 10-7 Figure 4: Observation of the plate 2 labeled 10-7 Figure 5: Observation to the plate 1 labeled Figure 6: Observation of the plate 2 labeled 10-6 The most probable number (MAN) for 3 tubes method in this experiment are MAN 3 24. 00 Number of viable cells (/ml) =
DISCUSSION The spread plate method is a technique which used to grow and isolate colonies of bacteria. Nutrient was provided when bacteria is transferred to agar plate. The diluted liquid contain bacteria is applied to agar plate and the sterilized hockey stick will spread and dilutes the amount of bacteria in each section of the agar plate continuously, so that the isolated colonies can be easily studied. When the colonies develop on the agar medium plates, it is found that there are some plates in which well-isolated colonies grow. Most probable number method [MAN]
No plate viable count: The most probable number method is a way of determining approximate viable count by diluting cultures then growing the dilution cultures in broth tubes. At the dilution at which broth becomes turbid or not with similar likelihood, the culture has been dilution to the point that the broth tubes were inoculated with on the order of only a single microorganism (turbid ) or fewer (not turbid ). The concentration to the culture is then necessary to have reached this point. Taken to be equal to the amount to dilution Advantageous where broth is advantageous:
MAN is especially useful in situations where there is an advantage to using broth over solid medium . For example, many organisms are not good at forming colonies, such as highly motile organisms. “When samples contain too few organisms to give reliable measures of population size by the standard plate count method, as in food and water sanitation studies, or when organisms will not grow on agar, the most probable number method is used. ” (P. 143, Black, 1996) Black, J. G. (1996). Microbiology. Principles and Applications. Third Edition. Prentice Hall. Upper Saddle River, New Jersey. Up. 140-144.