Labeling of binding sites for beta 2-microglobulin (beta 2m) on nonfibrillar surface structures of mutans streptococci by immunogold and beta 2m-gold electron microscopy.

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As little detail is known about the surface structure of streptococci in the mutans group and the relationship of surface structure to host ligand-binding functions, the twofold purpose of this investigation was to examine in detail, by a range of electron microscopic techniques, the surface structures of streptococci in the different species of the mutans group and to investigate the distribution of beta 2-microglobulin (beta 2m)-binding sites on such structures. Strains representing Streptococcus mutans, S. cricetus, S. rattus, S. sobrinus, and four fresh isolates were studied by shadowcasting and histochemical staining of whole-mounted cells as well as by ultrathin and thick sectioning of embedded specimens. beta 2m-binding site distribution was visualized by indirect immunogold electron microscopy and by direct bacterial binding of beta 2m-conjugated gold probes. Shadowcast preparations revealed binding of gold probes to the cell surface of known beta 2m-binding strains but not to their polar fibrillar appendages. These long fibrils, common to all strains, were trypsin and sonication sensitive and stained with lead citrate but not with uranyl acetate or ruthenium red. More gold particles were bound by the indirect technique. For grid-mounted bacteria, the gold was mostly bound in clusters at the periphery of the cells. When gold probes were reacted in suspension with bacteria before mounting onto grids, a more even distribution of the gold was seen, but the bacteria were aggregated. Heating the bacteria eliminated beta 2m-gold binding but had no effect on the morphology of the fibrils. Thick sections of embedded bacteria prereacted with beta 2m-conjugated gold probes were analyzed by stereo imaging. A wispy, uranyl acetate-stained fuzzy layer, distinct from the fibrils seen by shadowcasting and extending up to one cell diameter from the cell wall, contained the gold probes. These findings introduce a concept that binding sites for some salivary ligands on mutans streptococci may be clustered on very delicate, nonfibrillar structures extending much further from the cell wall than previously appreciated. As for beta 2m, which composes part of the human histocompatibility antigens, part of the bacterial surface would be coated at a distance from its body with a protein not necessarily recognized as foreign by the host.

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