The surface ultrastructure of three anaerobic Gram-positive cocci frequently encountered in oral infections, Peptostreptococcus micros, P. magnus and P. anaerobius, was studied. The type strains of P. micros (DSM 20468) and P. anaerobius (ATCC 27337), several clinical isolates of both species and the type strain of P. magnus (DSM 20470) were included. Thin-sectioned cells studied by electron microscopy revealed a homogeneous layer outside the peptidoglycan layer in P. anaerobius. In P. micros and P. magnus a more amorphous layer was present. No periodic structures were seen in negatively stained whole cells of these three species. However, in freeze-etched cells of P. anaerobius a crystalline surface protein layer (S-layer) was detected. No periodicity was seen in any of the P. micros strains or the P. magnus type strain by the methods used, but a periodic pattern was observed in negatively stained specimens of cell wall fragments of sonicated P. anaerobius cells. No capsular material was visible outside the S-layer in P. anaerobius. The cells of the Peptostreptococcus spp. were extracted for 30 min with detergents and urea. One per cent SDS and M urea both extracted a major 78 kDa protein from all strains of P anaerobius. Extraction of P. micros and P. magnus cells did not reveal any major protein bands comparable to that of P. anaerobius. Surface biotinylation of cells followed by Western blotting and detection by alkaline-phosphatase-conjugated extravidin showed strong staining of the 78 kDa band in P. anaerobius, further indicating that this molecule is located on the surface of the cell and is the S-protein.(ABSTRACT TRUNCATED AT 250 WORDS)