SELECTION AND PHENOTYPIC CHARACTERIZATION OF NONHEMAGGLUTINATING MUTANTS OF PORPHYROMONAS-GINGIVALIS

To further investigated the relationship between fimbriae and the hema gglutinating adhesin HA-Ag2 of Porphyromonas gingivalis, three spontan eous mutants of the type strain ATCC 33277 were selected by a hemadsor ption procedure. They were characterized for hemagglutination, trypsin -like and lectin-binding activities, and hydrophobicity and for the pr esence of fimbriae. The presence of the 42-kDa (the fimbrilin subunit) and the 43- and 49-kDa (the HA-Ag2 components) polypeptides was inves tigated by immunoblotting using polyclonal and monoclonal antibodies d irected to fimbriae and to the hemagglutinating adhesin HA-Ag2. Cells from two of the three mutants (M1 and M2) exhibited no or little hemag glutination activity and very low trypsin-like activity and did not sh ow the 43- and 49-kDa polypeptides. Abnormal fimbriation in M1 was ded uced from the following observations of cells grown for 18 h: absence of the 42-kDa polypeptide and of a 14-kDa polypeptide and no fimbriae visible on electron micrographs. While the cells of mutant M2, irrespe ctive of the age of the culture, were found to lack the 43- and 49-kDa polypeptides and hemagglutination activity, the supernatants of cultu res grown for 72 h had high hemagglutination and trypsin-like activiti es and revealed tile presence of the 42-, 43-, and 49-kDa polypeptides . This suggests that M2 may be missing some molecules which anchor the components to the cell surface. Mutant M3 showed levels of activities similar to those of the parental strain but lacked the 43-kDa polypep tide. Other pleiotropic effects observed for the mutants included loss of dark pigmentation and lower hydrophobicity. The data from this stu dy fuel an emerging consensus whereby fimbriation, hemagglutination, a nd proteolytic activities, as well as other functions in P. gingivalis , are intricate.