BACTERIA AND RURAL WATER SUPPLIES 19 



If intestinal environment should be able to convert A. aerogenes into E. coli 

 or intermediates, it seemed that bile or bile salts, because of their concentration 

 in the intestine, could well be decisive factors in effecting such a conversion. 



Thirty strains of A. aerogenes were selected for the study. They had been 

 isolated from water and classified on the basis of the Imvic reactions and the 

 uric acid test. The cultures had been carried on nutrient agar slants for three 

 3ears, and had given constant dififerential reactions when retested periodically. 



The medium was 1 percent lactose broth buffered with K2HPO4. Bacto oxgall, 

 sodium glycocholate, and sodium taurocholate were added separately to portions 

 of the medium. One series of 1 percent and another of 5 percent oxgall or bile 

 salt were prepared. Cultures were inoculated into the media and incubated at 

 37°C. Once each week each culture was transferred to a fresh tube of the same 

 medium. 



At the end of five months of this treatment the cultures were tested for the 

 differential reactions already mentioned, as well as with lactose broth and Endo's 

 medium. Gram-stained smears were also examined. The cultures retained all 

 of their original differential reactions, which indicated that bile or bile salts 

 alone are not likely to convert A. aerogenes into other forms of coliform bacteria. 



J. Bacteriophage 



It is an accepted fact that bacteriophage is able to cause mutations among 

 'phage-susceptible bacteria. This is sum^marized by d'Herelle (18), who states 

 that bacteria, exposed to the action of bacteriophage, undergo mutations that 

 are frequently unstable but that may become fixed. 



It was the purpose of the present study (19) to investigate the possibility that 

 bacteriophage might so alter cultures of E. coli and A. aerogenes as to produce 

 mutants that would give differential reactions of other coliform bacteria, and 

 particularly^ of the intermediates. 



A culture of E. coli was isolated from sewage, and a homologous bacteriophage 

 was isolated from the same sewage. The technique of d'Herelle (18) was em- 

 ployed. The medium employed in isolating the bacteriophage was Rakieten's 

 (20) Savita broth. Other media were made and used as directed in the Standard 

 Methods of Water Analysis (9). 



Two-liter quantities of sterile tap water were inoculated with the bacteriophage. 

 Cultures of E. coli (including the homologous culture), and of A. aerogenes were 

 inoculated into the bottles and kept at room temperature for five months. Con- 

 trol inoculations were made in water with no bacteriophage added. From time 

 to time re-isolations were made from the bottles and tested, with results as 

 follows: 



The biochemical reactions of the several cultures of bacteria employed were 

 studied. Tests were lactose broth fermentation, Endo plates, Imvic tests, and 

 sodium malonate. Reactions were unaltered except that some cultures of the 

 homologous E. coli produced atypical colonies on Endo's agar. These same 

 re-isolated cultures displayed increased resistance to the bacteriophage. 



The cultures, both homologous and non-homologous, that were subjected to 

 the bacteriophage action were agglutinated by much higher dilutions of their 

 specific sera than were the parent cultures. 



No striking mutations were produced in the study. 



