TESTS FOR B. COLI IN MILK 359 



of successful isolations came from the first presumptive test. 

 We are not at all sure what constitutes "attenuated" B. coli, 

 whether these are organisms that have temporarily lost their 

 ability to ferment lactose or whether they have lost only their 

 ability to ferment lactose aerobically but not anaerobically, 

 that is to say, under a reduced oxygen pressure. In water work 

 we and others have frequently isolated aerobic non acidifying 

 colonies from htmus lactose agar plates which form gas and 

 acid in the fermentation tube, though usually slowly; this seems 

 to point to the latter condition. In our milk studies no such 

 cases were encountered, due we believe to the fact that colon 

 baciUi recovered from milk on lactose agar are in full possession 

 of their lactolytic power because of the recent activity of that 

 function in the milk itself. 



Our main conclusion is that while we are not justified as yet 

 in claiming that the addition of gentian violet to lactose broth 

 will enable us to say in every case that B. coli is present when 

 gas is formed, yet its use certainly does not interfere with the 

 demonstration of B. coli; indeed it distinctly favors such demon- 

 stration. Further, the use of 1-100,000 gentian violet in the 

 litmus lactose agar plate does not interfere with the growth or 

 acid producing properties of the organism; rather it eliminates 

 to a large degree the masking of acid produced by B. coli, through 

 the inhibition of alkali forming, proteolytic, non-lactolytic. 

 Gram positive sporulating baciUi of the hay bacillus group, not 

 to mention the advantage of preventing spreading growths due 

 to these organisms. Moreover the inhibition of acidifying cocci 

 is of distinct advantage in the search for B. coli. 



We are frankly surprised not to have demonstrated sporulat- 

 ing anaerobes in milk heated for thirty minutes to 56 to 60°C. 

 It may well be that larger samples, say 10 cc, such as we have 

 used in our water examination would give positive results. 

 Another explanation may be mentioned tentatively, namely, 

 that the presence of lactose in the milk has caused lactolytic 

 sporulating anaerobes to vegetate so that they would be as 

 susceptible to heating as B. coli, since various investigators 

 such as Simonds (1915), Hibler (1908), and others, as well as our 



