bStreptococcus suis serotype 2 is a highly invasive, extracellular pathogen in pigs with the capacity to cause severe infections in humans. This study was initiated by the finding that IgM degradation products are released after opsonization of S. suis. The objective of this work was to identify the bacterial factor responsible for IgM degradation. The results of this study showed that a member of the IdeS family, designated Ide Ssuis (Immunoglobulin M-degrading enzyme of S. suis), is responsible and sufficient for IgM cleavage. Recombinant Ide Ssuis was found to degrade only IgM but neither IgG nor IgA. Interestingly, Western blot analysis revealed that Ide Ssuis is host specific, as it exclusively cleaves porcine IgM but not IgM from six other species, including a closely related member of the Suidae family. As demonstrated by flow cytometry and immunofluorescence microscopy, Ide Ssuis modulates binding of IgM to the bacterial surface. Ide Ssuis is the first prokaryotic IgM-specific protease described, indicating that this enzyme is involved in a so-far-unknown mechanism of host-pathogen interaction at an early stage of the host immune response. Furthermore, cleavage of porcine IgM by Ide Ssuis is the first identified phenotype reflecting functional adaptation of S. suis to pigs as the main host.

a b s t r a c tStreptococcus pyogenes employs an IgG specific endopeptidase, IdeS, to counteract the effector functions of specific IgG. The physiological significant step in disarming specific IgG is the cleavage of one IgG heavy chain. So far, characterizations of IdeS enzymatic activity have employed techniques that failed to differentiate between the first and the second cleavage step. The present data demonstrate that IdeS is active as a monomer and that IdeS activity follows classical Michaelis-Menten kinetics arguing against the previously proposed formation of a functional IdeS dimer. Our results show that IdeS inactivates IgG 100-fold faster than previously reported.

Recently we have discovered an IgG degrading enzyme of the endemic pig pathogen S. suis designated IgdE that is highly specific for porcine IgG. This protease is the founding member of a novel cysteine protease family assigned C113 in the MEROPS peptidase database. Bioinformatical analyses revealed putative members of the IgdE protease family in eight other Streptococcus species. The genes of the putative IgdE family proteases of S. agalactiae, S. porcinus, S. pseudoporcinus and S. equi subsp. zooepidemicus were cloned for production of recombinant protein into expression vectors. Recombinant proteins of all four IgdE family proteases were proteolytically active against IgG of the respective Streptococcus species hosts, but not against IgG from other tested species or other classes of immunoglobulins, thereby linking the substrate specificity to the known host tropism. The novel IgdE family proteases of S. agalactiae, S. pseudoporcinus and S. equi showed IgG subtype specificity, i.e. IgdE from S. agalactiae and S. pseudoporcinus cleaved human IgG1, while IgdE from S. equi was subtype specific for equine IgG7. Porcine IgG subtype specificities of the IgdE family proteases of S. porcinus and S. pseudoporcinus remain to be determined. Cleavage of porcine IgG by IgdE of S. pseudoporcinus is suggested to be an evolutionary remaining activity reflecting ancestry of the human pathogen to the porcine pathogen S. porcinus. The IgG subtype specificity of bacterial proteases indicates the special importance of these IgG subtypes in counteracting infection or colonization and opportunistic streptococci neutralize such antibodies through expression of IgdE family proteases as putative immune evasion factors. We suggest that IgdE family proteases might be valid vaccine targets against streptococci of both human and veterinary medical concerns and could also be of therapeutic as well as biotechnological use.

Streptococcus suis is a major endemic pathogen of pigs causing meningitis, arthritis, and other diseases. Zoonotic S. suis infections are emerging in humans causing similar pathologies as well as severe conditions such as toxic shock-like syndrome. Recently, we discovered an IdeS family protease of S. suis that exclusively cleaves porcine IgM and represents the first virulence factor described, linking S. suis to pigs as their natural host. Here we report the identification and characterization of a novel, unrelated protease of S. suis that exclusively targets porcine IgG. This enzyme, designated IgdE for immunoglobulin G-degrading enzyme of S. suis, is a cysteine protease distinct from previous characterized streptococcal immunoglobulin degrading proteases of the IdeS family and mediates efficient cleavage of the hinge region of porcine IgG with a high degree of specificity. The findings that all S. suis strains investigated possess the IgG proteolytic activity and that piglet serum samples contain specific antibodies against IgdE strongly indicate that the protease is expressed in vivo during infection and represents a novel and putative important bacterial virulence/colonization determinant, and a thus potential therapeutic target.The Gram-positive bacterium Streptococcus suis commonly colonizes the respiratory and genital tracts of domestic pigs and wild boars, but is also a significant porcine pathogen causing pathologies such as arthritis, abscesses, endocarditis, septicemia, and meningitis (1). Streptococcal meningitis is a common problem in intense pig farming and associated with substantial economic losses. S. suis strains can, based on the structure of capsular polysaccharides, be classified into at least 35 serotypes with S. suis serotype 2 as the significant zoonotic agent causing mainly septicemia and meningitis in humans (2-4). Mortality rates from zoonotic infection range between 3 and 18% (3-6). Transmission of S. suis between humans has not been confirmed, but only few data explaining S. suis functional adaptation to pigs as their dominant host are available (7). The routine use of antibiotics in the swine industry has caused increasing concern with reports of widespread resistance of S. suis strains against tetracyclines, and macrolides, but also cephalosporins, fluoroquinolone, and an emerging resistance against ␤-lactam antibiotics (2, 8, 9).Several virulence-associated factors of S. suis have been described, among which the polysaccharide capsule is so far the most prominent one in protecting the pathogen against phagocytosis (10). Numerous surface-bound and secreted proteins of S. suis were identified as homologous to virulence factors from other pathogenic streptococci and shown to exert similar functions (10). Among these are several proteases that appear to be involved in S. suis pathogenicity, but only few have been characterized, e.g. the IL-8 degrading protease SspA, a putative collagenase, and an IgA protease (11). Interestingly, most virulence factors and phagocytosis evasion mechan...

Significance Pneumococcal infections are major contributors to morbidity and mortality worldwide. Introduction of pneumococcal conjugated vaccines (PCVs) into the childhood vaccination program has led to a decrease in invasive pneumococcal disease (IPD) in vaccinated children but concurrently to an increase of nonvaccine-type IPD, also in nonvaccinated age groups such as the elderly. Thus, novel vaccine approaches are urgently needed, especially for the elderly, targeting all pneumococci causing IPD. Here, we show that pneumococcal membrane particles (MPs) evoke a serotype-independent cross-protection against IPD. This protection is dependent on the presence of the two conserved lipoproteins MalX and PrsA. We suggest that MPs can be used for pneumococcal vaccine development.