Identification of a fimbriae-associated haemagglutinin from Prevotella intermedia.

Prevotella intermedia, a putative periodontopathic microorganism, possesses various types of fimbriae (surface appendages). Some of these surface structures mediate the adherence of the organism to several mammalian erythrocytes, resulting in the agglutination of the erythrocytes. Prevotella intermedia fimbriae were solubilized and separated by preparative SDS gel electrophoresis followed by preparative isoelectric focusing to determine which fimbrial component(s) were responsible for the haemagglutinating activity exhibited by these bacteria. Heat treatment of isolated fractions which exhibited haemagglutinating activity revealed the presence of two types of haemagglutinating activity which were either heat sensitive or resistant. Analysis of isolated fractions, which exhibited haemagglutinating activity that were heat labile, by Tricine-sodium dodecyl sulfate-polyacrylamide gel electrophoresis revealed the presence of a 3.8 kD peptide. The purified peptide had a pI of 4.8-5.0. Examination of fractions containing this peptide by electron microscopy showed the presence of fimbriae. Immunogold labelling showed that mouse antibodies raised against the 3.8 kD peptide bound specifically and extensively to P. intermedia fimbriae. It appears that this peptide is a fimbriae-associated haemagglutinin and may represent a major constituent of the fimbriae. Further, fractions exhibiting haemagglutinating activity that were heat resistant, which were recovered at a pH of 3.5 in preparative isoelectric focusing of fimbrial samples, did not possess any detectable major protein bands as shown by analytical gel electrophoresis. However, silver stained gels for the detection of lipopolysaccharide (LPS) revealed the presence of LPS-like components in these fractions. In addition, LPS isolated from whole cells showed a similar electrophoretic pattern and exhibited the haemagglutinating activity that was heat resistant. The results of this study strongly suggest that P. intermedia possesses the machinery to agglutinate erythrocytes, which may be a contributing factor in its colonization in vivo.