Extractability of Cell Wall Polysaccharide from Lactobacilli and Streptococci by Autoclaving and by Dilute Acid

Abstract

Autoclaving cell wall of Streptococcus mutans Ingbritt for 15 min under the Rantz and Randall conditions released one-tenth of the total cell wall carbohydrate, whereas two-thirds was extracted after autoclaving for 180 min. The extract contained the serotype c -specific antigen but lacked the lipoteichoic acid component extracted when whole cells were autoclaved. Autoclaving cell wall preparations from other strains of S. mutans and also Streptococcus salivarius and Streptococcus mitis in 0.85% NaCl for 180 min released the major proportion of the wall polysaccharide fraction. Approximately 50 to 90% of wall carbohydrate of Lactobacillus fermentum and Lactobacillus casei was released when cell wall preparations were autoclaved in 0.85% NaCl for 180 min. For wall preparations from several strains of S. mutans , autoclaving for 60 min at pH 3.75 released only 39 to 62% of wall carbohydrate, whereas almost total release could be achieved with the lactobacilli. Heating S. mutans Ingbritt cell wall for 24 h at 60°C in 0.1 N H 2 SO 4 released only two-thirds of the wall carbohydrate; by comparison nearly all of the wall carbohydrate was released in 3 h from L. casei and L. fermentum. Autoclaving L. casei cell wall and purified soluble wall fractions hydrolyzed the phosphodiester bond between the polysaccharide and peptidoglycan. This was shown by the release of reactive N -acetylhexosamine in both cases and the presence of a phosphomonoester in the autoclaved soluble wall fractions. The results indicate that autoclaving can hydrolyze covalent linkages, and this must be considered when the Rantz and Randall procedure is used to obtain antigen preparations.