220 WHITMORE. 



Using the four Shiga-Kruse strains^, I attempted to determine their 

 tjfpe according to Olmo's method. Two of my strains (P. S. I and P. 

 S. II) were isolated recently, while two (S. S. I and S. S.-II) had 

 been on artificial media for some time. All of the four belonged in 

 Ohno's type A. Eight months later I tested these same strains again. 

 One (P. S. II) had changed to Ohno's type B, while the other three 

 remained in his type A. 



I used rabbits for the preparation of specific sera and made all in- 

 jections intravenously. The animals were weighed once a week and 

 the injections were given as closely as possible within the same interval, 

 consideration being given to the weight and general conditions of the 

 animal. 



The first four or five injections were of organisms grown in agar for 

 18 hours and heated to 60° for one hour, Avhile the later ones were of 

 living organisms. Each animal usually received fourteen injections. 

 The first was always very small and later the dose gradually Avas in- 

 creased. I had great difBculty with the rabbits which were given the 

 Shiga-Kruse cultures, 1/50 of a loop of a killed culture of one of the 

 recently isolated strains killing rabbits when injected intravenously. I 

 lost animals from the use of these strains as late as the fourteenth 

 injection. The two strains received from Japan were not so virulent, 

 possibly because they had been on artificial media for some time. The 

 animals withstood larger doses of the latter and this may account for 

 the fact that I attained sera with stronger agglutination from them. 

 These facts appear in the agglutination tables given below. No especial 

 difficulty was encountered in giving the Plexner strains and non-dysentery 

 organisms in much larger doses than is possible with the Shiga-Kruse 

 strains.^ 



The animals were bled about ten days after the last injection. In 

 order to obtain the large amount of serum needed for this work and 

 still not to kill the rabbit, the following method was used : 



A large test tube was provided with a rubber stopper having two perforations. 

 A piece of glass tubing connected by a short piece of rubber tubing to a long, 

 slender aspirating needle passed through one perforation; a piece of glass tubing 

 through the other, the later connected with the vacuum apparatus by a long 

 piece of rubber tubing. The rabbit was placed on its back on an animal board, 



° At the same time I immunized a horse to both Shiga-Kruse and Flexner 

 types, by giving weekly intravenous injections of living organisms alternating 

 with the filtrate from an old culture in alkali bouillon. One horse died of an 

 intercurrent condition after it had received ten injections, but another received 

 seventeen injections and at the end of that time his blood agglutinated Shiga 

 at 1:800 and Flexner above 1:1000. The animal was bled ten days after the 

 last injection and the serum put up for use in the treatment of bacillary dysentery. 

 So far we have had very little opportunity to try it, as the epidemic dysentery 

 tmder discussion was oyer by the time the serum was ready for use. 



