Cross-Reactions Among Group A Streptococci

Abstract

The strain-specific (Sp-M) and the cross-protective (X-M) antigens of type G1 cocci were isolated from RNase-treated cell walls and purified by ammonium sulfate fractionation, carboxymethylcellulose and Sephadex G-200 column chromatography and polyacrylamide gel electrophoresis. Acid hydrolysis resulted in fragmentation of the antigen molecules, but precipitin and bactericidal tests established that the major serologic activity characteristic of the native antigens, with minor exceptions, remained intact. The purified antigens absorbed their corresponding antibodies to yield specific antisera whose bactericidal activity correlated with bands of precipitate in agar gel diffusion tests. It was established that the X-M antigen has specific as well as cross-reactive activity, a determination which it was impossible to make before the purified material became available. The purified antigens with which antisera were absorbed had a ratio of O.D. 280 nm to O.D. 260 mn of 0.79 for the Sp-M and 1.00 for the X-M. The molecular size of the Sp-M was greater and the charge smaller than those of the X-M. Also, it contained twice as much glycine and half as much aspartic acid. The average ratio of lysine and arginine for the Sp-M and X-M was 2.3 and 1.5, respectively. The observation is of interest because it has been established in other work that the Sp-M requires a free ε-NH2 group of lysine and a free guanidino group of arginine for its serologic activity. The X-M antigen does not. The antigens were extractable as separate entities at pH values from 2 to 10 and at temperatures of 37°C and 100°C, thus demonstrating that they do not arise only by acid hydrolysis of a larger molecule with Sp-M and X-M serologic activity and may indeed exist as separate antigens on the cell wall. The molecules in alkaline extracts were larger than those in acid extracts. Acid hydrolysis of the alkaline preparation yielded products whose gel electrophoretic pattern was similar to that of the acid-extracted material. Therefore, it was possible to conclude that the smaller molecules of the latter were the result of acid hydrolysis and not the other procedures employed.