?] FATE OF TI-I;KKU.K 15. \ULLI OUTSIDE TIIK ANIMAL BODY 319 



DURATION OF LIFE OF BOVINE. TUBERCLE BACILLI IN Cow 



MANURE: 



PURPOSES OF The general purposes of these experiments was to 

 ME NTS determine how long bovine tubercle bacilli will 



live in cow manure when exposed to weather con- 

 ditions in a pasture field. Two series of tests have been made con- 

 cerning the resistance of these organisms when a! pure culture is 

 artificially mixed in cow manure exposed in the sunshine, and also 

 when it is exposed in a place protected from the sunshine. In a 

 similar way other tests have been made with naturally infected 

 manure from a tuberculous, cow. 



Series i 

 (Special Methods) 



PREPARATION Artificially Infected Manure. The culture used 

 OF SAMPLES in preparing this sample of artificially infected 

 manure was a strain of bovine tubercle bacilli ob- 

 tained from Dr. Theobald Smith of the Harvard Medical School. 

 It was received and has since been kept upon glycerine agar. The 

 strain readily produces an abundant growth and is quite virulent 

 to test animals. 



Four milligrams of the culture from two tubes of glycerine 

 agar were removed and emulsified as described on page 315. The 

 emulsion was diluted to 200 cc. with sterile salt solution. It was 

 expected that the large amount of solution used would give a 

 more uniform mixture of the tubercle bacilli in the cow manure. 



The sample of manure was obtained from two cows of the 

 dairy herd. It was taken by the method described in Bulletin 149 

 of this station; i. e., by injecting air into the rectum of the cow 

 until stimulated to defecate. The feces were caught in a sterile 

 pail and at once covered and brought to the laboratory. The 200 

 cc. emulsion of bovine tubercle bacilli was thoroly mixed with 

 1800 grams of the fresh cow manure. The; infected manure was 

 tested for virulence by inoculation of two guinea pigs. One gram 

 was rubbed up in 50 cc. of 0.8 percent salt solution, 40 cc. of the 

 emulsion was centrifuged and the sediment injected subcutaneously 

 into two guinea pigs, both of which became infected with gen- 

 eralized tuberculosis. 



Naturally Infected Manure. The sample of naturally infected 

 manure was obtained from a tuberculous cow (cow No. c;6 from 

 the dairy herd of this station) that had previously reacted to 

 tuberculin. The sample was taken by the method described in our 

 previous publication referred to above. Approximately three kilos 



