SEPTIC TANK STUDIES 9 



The procedures employed were those outlined in the Standard Methods of 

 Water Analysis. Difco dehydrated media were employed. Samples of the 

 effluent, diluted suitably to obtain optimum results, were inoculated into fer- 

 mentation tubes of lactose broth, five tubes for each dilution. After incubation 

 at 37° C. for 48 hours the numbers of tubes showing positive gas production were 

 recorded. Then plates of eosin-methylene blue agar were streaked from these 

 tubes to indicate the types of coliform bacteria present, and all cultures were 

 reinoculated from the plates into lactose-broth fermentation tubes to complete 

 the tests. Cultures were isolated from plates representing the highest dilutions 

 of the effluents giving positive gas tests, five cultures from each sample. These 

 cultures were subjected to differential tests to determine the distribution of 

 types within the coliform group of bacteria. This part of the study is discussed 

 later. 



It will be noted that the incubation temperature employed throughout for 

 the various coliform tests was 37° C, while that for the plate counts was 30° C. 

 The reason for the difference was that in isolating and differentiating the coli- 

 form bacteria the time required to complete the tests can be shortened con- 

 siderably by incubation at 37°, while the intent of the plate counts is to obtain 

 growth of as many types of bacteria as possible. Most types, including the 

 coliform bacteria, develop satisfactorily at 30°, and some types require the lower 

 incubation temperature for satisfactory development. 



The numbers of gas-positive fermentation tubes were calculated to determine 

 the M.P.N, (most probable number) of coliform bacteria by the "Standard 

 Methods" procedure, with the results summarized in table 3. One result for each 

 tank in each collecting period was much higher than any of the others for the 

 respective tank, so two sets of calculations were made: one set including all 

 results, and one with the extra high figures omitted. 



The results show that for the entire set of calculations of period I the middle 

 tank (12 hours retention) had the highest average M.P.N., the north tank 

 (8 hours retention) was second, and the south -tank (24 hours retention) had the 



Table 3. — Most Probable Nxbiber (M.P.N.) of Coliform 

 Bacteria per Milliliter of Septic Tank Effluent. 



* For each collection period, one M.P.N, value for each tank was much higher than other values 

 for that tank for the same period. Calculations were made with and without these extra high 

 values. 



