60 



when dealing with untreated waters but it is not to depended upon when 

 testing chlorinated waters; 97.7 per cent of the former and only 44 per cent 

 of the chlorinated samples showing gas, were confirmed for Bact. coli 

 or its close allies. 



2. The doubtful presumptive test (less than 10 percent gas in 

 24 hours, or more than this quantity in 48 hours where 24 hours was nega- 

 tive) is only a fair index of the* probable presence of Bact. coli in un- 

 treated waters, while for chlorinated specimens it is practically negligible, 

 as 75.9 percent of gas tubes from untreated, and only 7.6 percent of those 

 from chlorinated samples were successfully confirmed. 



3. A small amount of gas in 24 hours is a more reliable index for 

 Bact. coli than 10 percent or more gas in 48 hours. The rate of gas pro- 

 duction seeems more significant than the total volume of gas formed as a 

 presumptive test. 



Similar results were obtained by Graff and Mote. Of 966 instances 

 of gas production in lactose peptone bile from settling basins (unchlorin- 

 ated) 41 percent were positive. Of 844 instances from reservoirs (after 

 chlorination) 12 percent were confirmed, and 1063 gas tubes from taps 

 only 5 percent were confirmed. They also noted that vigorous gas pro- 

 duction in 24 hours yielded positive tests for Bact. coli in a greater percent 

 of cases, from all sources, than was obtained if gas was not noted previous- 

 ly to the 48 hour period. 



It is thus apparent that gas production in either lactose broth or 

 lactose bile, particularly with treated, partially purified, or chlorinated 

 waters, cannot be relied upon as an index of the presence of the colon 

 groujj. Confirmation must be restored to, except possibly for water 

 known to be polluted. 



CONFIRMATORY TESTS 



it is merely necessary to streak out or smear on the surface of a differential 

 medium (litmus lactose agar, Endo agar, or Eosin-Methylene-Blue agar) 

 which is then included at 37 C. for 24 hours. If typical colonies develop, 

 the presence of the colon group in the presumptive test is considered con- 

 firmed. This is known as the partially confirmed test and for routine 

 work is sufficient, although it must be borne in mind that there are a few 

 spore forming lactose fermenters which are capable of growing aerobically. 



If the colonies developing on this differential medium are not typical 

 it becomes necessary to further study these colonies to determine that they 

 are Gram negative non-spore formers and that they are capable of pro- 

 ducing gas from lactose. The latter may be determined by merely fish- 

 ing a colony into lactose broth which is incubated for 24 hours. If gas 

 is formed and if the Gram stain showed the colony to be a non-spore form- 

 ing Gram negative short rod, the test constitutes what is spoken of in the 

 Standard Methods as the completed test for the colon group. 



If no growth at all develops on the confirmatory differential medium 

 the assumption is that the gas in the presumptive test tube was due to 

 strict anaerobes and the colon group is recorded as absent. It is con- 



