48 MASS. EXPERIMENT STATION BULLETIN 319 



SEROLOGICAL STUDIES 



In the preceding sections it was shown that variation of 5. ptdlorum may 

 occur in colonial and cellular morphology, macroscopic appearance in broth 

 medium, stability in saline solution, and virulence. Similar observations have 

 been reported by other investigators for different species of bacteria. Arkwright 

 (1920, 1921), in a study of some of the members of the colon-typhoid-dysentery 

 group, observed that changes in colony form, salt-sensitiveness, and growth in 

 broth may be associated with profound alteration in antigenic structure. Specificity 

 in agglutination was observed for the normal strain, while non-specific agglutina- 

 tion occurred with the variant. The variant might be altered to the extent that it 

 would be agglutinated by immune serum homologous for variants of unrelated 

 species. Peculiar immunizing and agglutinogenic properties have been ascribed 

 to variants by such investigators as Schiitze (1921), De Kruif (1921), and Ark- 

 wright (1921.). 



According to the classification of the antigenic structure of the typhoid-para- 

 typhoid group by Kauffmann (1930) and White (1926), 5. pullorum contains 

 only a somatic antigen. White (1932, 1933), in his studies concerning different 

 antigenic fractions of the Salmonella group, observed that different surface anjti- 

 gens may be lost through bacterial variation. 



In these experiments several normal strains and variants were studied as to 

 their serological behavior. This phase of the investigation bears a direct relation- 

 ship to the routine serological diagnosis of pullorum disease because, as will be 

 pointed out, 5. pullorum strains can be so altered in their antigenic structure that 

 they lose their power to produce agglutinins in the host and fail to absorb the 

 specific agglutinins. 



Realizing that a salt-sensitive variant could not be employed as a satisfactory 

 antigen strain, an effort was made to stabilize it and yet maintain optimum condi- 

 tions for agglutination. From the literature one may gain the impression that the 

 problem of maintaining variants suspended in saline solution is of minor impor- 

 tance. It is possible that the variants reported by others were of a different charac- 

 ter from those dealt with here. Moreover, while variant types that are different 

 as to their salt-sensitiveness do occur, it is also possible that variation may exist 

 in the interpretation of the behavior of variants in salt solution. In the tests 

 conducted with the variants in this study, the author is inclined to believe that in 

 certain instances a combination of agglutination and settling of cells has occurred. 

 It has been reported by some that the phenomenon of self-clumping in salt solution 

 is identical with that of true agglutination which occurs in an agglutinating serum- 

 antigen-saline mixture. It is apparent that some variants will form perceptible 

 clumps in salt solution, but others may settle without perceptible clumping. 

 \\ ith either type of variant, when it is placed in contact with agglutinating serum, 

 the reaction may be quite complex and the end result baffling. The question is 

 to what extent the two phenomena are represented. 



White (1926) reported that variants can be made stable in salt solution by treat- 

 ing the culture with 95 per cent alcohol. He also stated that the alcoholic extract 

 contains an antigenic substance, which suggests that the variant itself has been 

 deprived of some of its chemical components by the alcohol treatment. To what 

 extent one can proceed in subjecting the organism to extracting agents without 

 depriving it of the essential constituents for true agglutination remains a question. 



Four pullorum-disease-free birds which were negative to the macroscopic 

 agglutination test with pullorum antigen were inoculated, one with normal 

 Strain YIII and the three others with Variants C, VHI-a, and IX-b, respectively. 



