244 TRANSACTIONS LIVERPOOL BIOLOGICAL SOCIETY. 



viously poured into a Petri dish and allowed to set, and 

 is then spread evenly over the surface of this medium 

 by means of a sterile, wide loop of platinum wire. The 

 Petri dish is then inverted and put into the incubator. 

 At the same time a similar pipetteful of the liquid is 

 inoculated into a tube of sterile litmus milk, which has 

 previously been heated to 100° C. for half an hour, and 

 then cooled rapidly. The tubes of milk thus inoculated 

 are then heated to 75°-80° C. for twenty minutes, and are 

 packed into an ordinary litre gas- jar (which holds about 

 a dozen 5in. test-tubes). Pyrogallic acid has previously 

 been placed in the bottom of the jar and strong caustic 

 soda solution is then poured in so as to dissolve the acid. 

 The jar is then tightly stoppered by a rubber bung and 

 put into the incubator. At first I used a modification of 

 Bulloch's anaerobic culture apparatus and displaced the 

 air in the jar by hydrogen, but this treatment is 

 unnecessary, for anaerobic conditions are secured by the 

 method indicated above. 



The cultivation of the stomach liquid on the neutral- 

 red agar medium enables one to isolate colonies of B. coli. 

 The anaerobic cultivation in milk isolates and grows the 

 spores of a bacillus — in most cases B. enteritidis sporogenes. 



These cultures are incubated for 24 hours at 41°-5- 

 42° C. With some little experience one is then able to 

 pick out the colonies on the neutral-red agar plates, which 

 are probably those of B. coli. From each plate two or 

 more colonies, representative of those judged to be pro- 

 duced by this microbe, are then sub-cultured on the 

 surface of sloping nutrient agar in tubes. These 

 secondary cultures are then incubated for 24 hours at 

 42° C. Also at the end of 24 hours the milk tubes are 

 examined and the presence or absence of the well-known 

 " enteritidis reaction " recorded. 



