LABOKATORY AND GREENHOUSE STUDIES. 79 



Dr. W. G. Savage * studied the following points in identifying Bacil- 

 lus coli: Motility, liquefaction of gelatin, type of colony on gelatin, 

 indol production, acid production, milk coagulation, and fermentation 

 of dextrose and lactose. 



Dr. D. Rivas - states that the usual method for identifying Bacillus 

 coli is as follows: 



Plating out in Wurtz's litmus-agar plates with 1.5 to 2 per cent Parietti's solution. 

 Examined after 24 hours' incubation at 37° C. All the pink colonies are isolated and 

 transferred to sugar media for fermentation. 



Transfers are made to Dunham's solution to test for indol. Also to nitrate bouil- 

 lon to test for reduction of nitrates to nitrites. 



Further transfers are made to gelatin tubes for liquefaction or nonliquefaction. 



Dr. J. J. Kinyoun uses Endo's fuchsin agar for the determination 

 of Bacillus coli. His method of making up the medium is one that he 

 considers as furnishing a very distinctive test, as follows: 



Take 2 liters tap water, 40 grams Liebig's meat extract, 40 grams Witte's peptone, 20 

 grams sodium chlorid, 160 grams agar flour. Put into a tall beaker and steam for three 

 hours. Let settle over night. Cut off dirty part and throw away. Melt the remainder 



and neutralize to phenolphthalein. Add 4 c. c. of -- hydrochloric acid. Steam 



one hour. This forms the stock, which should be clear. The crux of the whole for- 

 mula lies in the following: Take 200 c. c. of this hot stock and add to it 2 grams of lac- 

 tose. Then add 2 c. c. of a solution of basic fuchsin (half-saturated solution) and 

 10 c. c. of fresh sodium-sulphite solution (5 grams to 100 c. c. of water). Divide into 

 eight lots of 25 c. c. each to form the trial lots. Make up a 10 per cent solution of sodium 

 carbonate, and add of this to the trial lots, in varying amounts, as follows: 0.01 c. c, 

 0.02 c. c, 0.03 c. c, 0.04 c. c, etc. Pour into large plates, cool, and streak for colon, 

 typhoid, etc. Incubate 24 hours at 37° C. The standard of alkalinity to be used on 

 the remainder of the stock is that of the plate which has given the most characteristic 

 results. Fill and set away the stock in 100 c. c. portions in bottles plugged with 

 cotton. As there is much water of condensation, the agar is hardened in the plates 

 uncovered in a clean place. Air germs (exclusive of molds) seldom grow on it. 



These points are stated to be the most essential in the identification 

 of Bacillus coli. 



The growth of the coconut organisms in various other media is de- 

 scribed on the following pages, 



dolt's synthetic medium no. 1. 



On slant cultures in Dolt's medium a good pink growth appeared 

 within 24 hours, and the agar became partly reddened. Evidence of 

 gas appeared in tubes of coconut No. 2 and Bacillus coli. Repetition 

 of this experiment gave exactly the same result with the exception of 

 no gas production. The growth along the streak was for the most 



1 Savage, W. G. The Characters of the Bacillus Coli as an Indicator of Excretal Contamination. Lan- 

 cet, London, vol. 168, Feb. 4, 1905, p. 284. 



2 Rivas, D. B. Coli Communis, "The Presumptive Test," and the Sewage Streptococci in Drinking 

 Water. Journal of Medical Research, vol. 16, 1907, pp. 85-98. 

 228 



