VARIATION OF SALMONELLA PULLORUM 13 



per cent NaCl solution which contained 0.5 per cent phenol. These antigens were 

 tested with negative and positive pullorum sera in dilutions of 1:10 and higher. 

 The results of the test were recorded after 48 hours incubation. 



All strains were stained by the Gram method. 



The cultures were obtained from several different sources, including eggs, 

 chicks, and mature birds. A large number of cultures was received from my 

 associates in the Department of Veterinary Science. The original cultures re- 

 ceived from Amherst were isolated on chicken meat infusion agar plates. The 

 age of the culture at the time of the examination varied from three to seven days. 

 In the majority of cases plain meat extract agar plates were streaked with the 

 primary cultures, in order to determine the character of the growth on meat 

 extract agar during the various incubation periods. 



Primary isolations of S. pullorum from chicks and adult birds were also received 

 from various members of the Yale Department of Bacteriology. 



The Department of Veterinary Science also furnished fresh eggs laid by hens 

 whose sera reacted with pullorum antigen. Among 176 eggs cultured, 39 yielded 

 S. pullorum. As a matter of interest concerning the bacteriological examinations 

 of the eggs, it may be stated that the percentage of isolation is markedly raised 

 by increasing the incubation period from two to six days. No definite explanation 

 is offered as to why the presence of the organism is not detected by streaking 

 on agar plate with egg yolk-broth mixture which has been incubated only 48 

 hours. In every instance the eggs were incubated in the shell for a period of seven 

 days before being cultured. It was also observed that occasionally eggs contain 

 particles of necrotic tissue which have been picked up in the abdominal cavity 

 by the oviduct and expelled as part of the egg. Such bodies may be found in the 

 albuminous portion. By careful technique the yolk and the foreign body may 

 be cultured separately. In such cases it was observed that the yolk may be sterile, 

 while the necrotic tissue yields 5. pullorum. Hence, it appears important, in the 

 bacteriological examination of fresh eggs, to recognize the fact that the organism 

 may exist in other portions of the egg aside from the yolk. Also this observation 

 suggests that the disease may be transmitted from adult to chick by means of 

 such infective foreign bodies, instead of the yolk in every instance, as is the 

 common belief. 



A total of 163 strains was examined as to colonial morphology, Gram stain 

 reactions, carbohydrate fermentations, and agglutinability. During a period 

 of approximately fourteen months, 69 strains were isolated from maturing and 

 adult birds, 55 strains from chicks and 39 strains from fresh eggs. The organism 

 was recovered from various organs, tissues and diseased processes. 



Thirteen of the 163 strains did not resemble typical 5. pullorum in every respect. 

 Table 1 shows that the majority of these 13 strains were salt sensitive, exhibiting 

 a tendency to settle out in normal NaCl solution. 



In the original isolation the colonial and cellular morphology, carbohydrate 

 reaction and Gram-staining properties resembled those of 5. pullorum, except in 

 two strains, 3 and 12. The latter exhibited a rough type of colony, but on sub- 

 sequent transfers to meat extract agar and beef and liver infusion agar the growth 

 was smooth and typical. No explanation is offered for this marked temporary 

 roughness as manifested in the primary culture. 



The different atypical strains were tested on several occasions with positive 

 and negative pullorum sera. The results of a few of the tests are shown in Tables 

 2 and 3. 



The antigens listed in Table 2 were prepared from meat extract agar cultures 

 which had been incubated at 37° C. for 48 hours. The cells were suspended in 



