32 Part III. — Twenty-sixth Annual Report 



(3) Take solution (2) and add to solution (1) when cooled to 60° C. 

 Shake, render faintly alkaline. Then add 4 c.c. hot sterile solution of 

 10 per cent, water-free soda and 20 c.c. of freshly-prepared solution of 

 O'l gram crystal violet (Hochst) in 100 c.c. warm sterile distilled water. 



Particular media for subculture : — 



(1) Gelatine slope, stab and shake cultures. 



(2) Litmus milk. 



(3) Lactose-peptone litmus solution, 



(4) Peptone water. 



(5) Glucose neutral-red broth. 



In this way I followed Houston's "flaginac " basis of classification for 

 bacillus coli : — 



Fl. — Greenish fluorescence in neutral- red cultures. 

 A.G. — Acid and gas in lactose-peptone cultures. 

 In. — Indol formation in peptone cultures. 

 A.C. — Acid and clotting of litmus milk. 



A microbe which presents these characters in subcultaral tests is 

 indistinguishable, as regards the tests employed, from the typical bacillus 

 coli of the human intestine. 



Method of Procedure. — The fish examined were obtained at different 

 times and from different sources at the Aberdeen fish market during my 

 visitations — some line caught, some from trawlers. But the great 

 majority of the fish were not caught at any great distance from the shore, 

 although some were obtained from trawlers from the Iceland fishing 

 grounds. 



Each fish was washed in tap water, then in sterile water, and tacked 

 out on a board. The skin on the ventral aspect of the abdomen was 

 thoroughly seared with a red-hot cautery iron, and the stomach and 

 intestines dissected out by sterile instruments. Some of the intestinal 

 contents were then aspirated up into a pasteur pipette or removed by 

 sterile platinum wire to bile-salt glucose pe.ptone litmus in a Durham's 

 fermentation tube and incubated for 48 hours at 37° C, 1-5 tubes 

 inoculated for each fish. 



Sometimes the whole gut was removed, washed in tap water, then in 

 sterile water, and the whole contents well mixed with bile-salt glucose 

 broth. 



From this emulsion other bile salt glucose peptone tubes were inocu- 

 lated with varying quantities. None of the fish were cut up and mixed 

 with the intestinal contents. A positive reaction is shown by the pro- 

 duction of acid and gas. The acid turns the medium red, and gas is seen 

 in the inner small tube. But as this reaction is only presumptive evi- 

 dence of the presence of a member of the coli group, I always plated out 

 on one or other of the solid media — chiefly Digralski and Conradi— and 

 from the colonies on this media, when present, subcultures were made in 

 accordance with the " flaginac " basis. 



The following Tables exhibit my recorded results : — 



