22 



oysters which, had ,been in the sterile sea water for the 24 

 hours, in the following manner : a thick zinc wire, bent at 

 right angle at the last inch, is carefully inserted between the 

 two halves of the shell, the oysters in the sea water having 

 their shells spontaneously opened ; immediately as the wire 

 end is inserted the shell closes so tightly on it that the oyster 

 can now be lifted out of the water on to a glass plate ; the 

 end of the cannula of a hypodermic syringe filled with turbid 

 emulsion of B. typhosus is easily introduced close to the 

 wedged-in wire, and the desired amount (in our case 1 c.c.) 

 of culture slowly injected. When finished the cannula of 

 the syringe is withdrawn the cannula being thinner than 

 the wedge and by gentle action the wire is removed ; the 

 shell immediately closes again tightly. The whole pro- 

 ceeding need not and does not occupy more than a few 

 minutes, and, as subsequent observation showed, no harm had 

 thereby been done to any part of the oysters. In the above 

 manner 12 oysters were injected with the B. typhosus, each 

 with 1 c.c. of the emulsion. 



The emulsion injected was obtained by distributing in 

 sterile sea water the growth from the surface of a pure agar 

 culture 24-48 hours old of our laboratory B. typhosus. 

 Before using the emulsion for the injection, a Drigalski plate 

 was made with a definite amount of a definite dilution of it, 

 and after 48 hours' incubation at 37 C. the number of 

 colonies of B. typhosus was counted, and thereby their 

 number per 1 c.c. of emulsion ascertained. It was thus 

 found that the number of B. typhosus injected into each 

 oyster was 162,500,000. 



The determination was made thus : 1 c.c. of the typhoid 

 emulsion was added to 100 c.c. sterile sea water, well shaken ; 

 of this dilution 0*01 c.c. was carefully and well rubbed over 

 the surface of Drigalski plate. After incubation for 48 hours 

 the Drigalski plate showed numerous colonies of B. typhosus 

 and no others, all being of the same colour and aspect ; a 

 careful count showed 1625 colonies in the plate, that is to 

 say, 1625 x 100 x 1000 = 162,500,000 per 1 c.c. 



Although the amount actually injected within the cavity 



