Numerous studies have implicated bacteria in cardiovascular disease, but there is a paucity of information on the mechanism involved. In this study we show how the common oral bacterium Streptococcus sanguis can directly interact with platelets, resulting in activation and aggregate formation. Platelet aggregation was dependent on glycoprotein IIb/IIIa (GPIIb/ IIIa) and thromboxane. Platelets could also directly bind to S sanguis, but this interaction was not inhibited by GPIIb/IIIa antagonists. Antibodies to GPIb could inhibit both platelet aggregation and platelet adhesion to bacteria. This suggested a direct interaction between GPIb and S sanguis; however, this interaction did not require von Willebrand factor, the normal ligand for GPIb. By use of a range of monoclonal antibodies to GPIb and the enzyme mocharagin, which cleaves GPIb at amino acid 282, the interaction was localized to a region within the N-terminal 1-225 portion of GPIb␣. Furthermore S sanguis failed to induce aggregation of platelets from a patient with BernardSoulier disease, the organism bound to Chinese hamster ovary cells transfected with the GPIb␣ gene but did not bind to mock-transfected cells and biotin-labeled S sanguis cells bound to purified GPIb in ligand blots. It is suggested that the interaction between S sanguis and GPIb is important in the pathogenesis of infective endocarditis and may also play a contributory role in some cases of myocardial infarction. ( IntroductionRecent reports suggest a role for infectious agents in cardiovascular disease. Much of this work is in the form of clinical evidence of infection [1][2][3][4][5][6] or the effect of antibiotics on the incidence of cardiovascular disease. 7,8 Although studies have found evidence of bacteria in atherosclerotic plaques, 1,2,4,5,9-13 their role in the etiology of cardiovascular disease is uncertain. In contrast, the role of bacteria in infective endocarditis is well established and the molecular mechanisms involved may also occur in other forms of cardiovascular disease.Infective endocarditis involves inflammation of the heart valves due to infection and if untreated can lead to valve failure and death. In most cases there is one or more predisposing factors, which results in damage to the endothelium on or adjacent to the valves. This area of damage becomes covered with a platelet-fibrin vegetation and these can become colonized by bacteria that gain access to the blood. The 2 species most commonly involved are Streptococcus sanguis 14 and Staphylococcus aureus. 15 Historically oral streptococci have been referred to as Streptococcus viridans, but this name was never accepted as a recognized taxon because of the biochemical and serologic heterogeneity among isolates. Subsequent detailed biochemical and genetic studies allowed the definition of at least 17 taxa within what was originally called S viridans. However, the term viridans has survived but is now used as viridans group of streptococci to recognize the existence of various taxa and S sanguis is one taxon w...

By comparison of the cell surface proteins derived from the outer membrane and fibrils from 14 Prevotella intermedia and 19 Prevotella nigrescens strains using SDS and analysed by SDS-PAGE, it was possible to distinguish the two species. A polypeptide of approx. 21 kDa distinguished P. intermedia strains, whereas two polypeptides of approx. 18 and 22 kDa could be used to identify P. nigrescens strains. Four other human oral black pigmented bacterial species (Porphyromonas gingivalis, Prevotella denticola, Prevotella loescheii and Prevotella melaninogenica) did not have the 18-, 21- or 22-kDa polypeptides shown by P. intermedia or P. nigrescens. The cell-associated proteolytic activity of eight strains of P. intermedia, 14 strains of P. nigrescens and one strain of P. gingivalis (W50) was assessed using four chromogenic substrates. The hydrolysis of the substrate GPPNA (indicative of dipeptidyl peptidase IV-like activity) and SAAPPNA (elastase-like activity) by P. intermedia strains varied from 32 to 114 units and 0.5 to 12.6 units of activity respectively, where one unit was defined as the amount of protease enzyme catalysing the formation of 1 nmol of p-nitroaniline under experimental conditions. 37.5% (3 of 8) of P. intermedia strains hydrolysed SAAPPNA (chymotrypsin-like enzyme activity) with activities of between 7 and 12 units. The hydrolysis of GPPNA and SAAAPNA by P. nigrescens strains was 32-149 and 3-16 units, respectively. 57% (8 of 14) of P. nigrescens strains hydrolysed SAAPPPNA with activities ranging from 3 to 8 units. None of the P. intermedia or P. nigrescens strains examined were found to have trypsin-like enzyme activity (BAPNA hydrolysis). The GPPNA and SAAAPNA hydrolytic activity associated with the proteases from Porphyromonas gingivalis W50 was at least twice that of P. intermedia and P. nigrescens strains. The similar peptidase activities of P. intermedia and P. nigrescens against chromogenic substrates cannot be used to differentiate the species, but SDS-PAGE of cell surface protein extracts allowed unambiguous speciation between P. intermedia and P. nigrescens. This simple technique of cell surface protein analysis can be performed in most laboratories and offers a convenient way by which to differentiate the two species.