Intranasal immunization with C5a peptidase prevents nasopharyngeal colonization of mice by the group A Streptococcus

Abstract

Early inflammatory events are initiated by phased production of C5a and interleukin-8 in tissue. Most serotypes of group A streptococci express a surface-bound peptidase (SCPA) which specifically cleaves mouse and human C5a chemotaxins. This study investigates the impact of SCPA on colonization of the nasopharyngeal mucosa of mice and evaluates its potential to induce protective immunity. Two strains, serotypes M6 and M49, which contain insertion and deletion mutations in the SCPA gene (scpA) and represent the two major subdivisions of group A streptococci, were characterized and compared in a mouse intranasal infection model. In this model, SCPA mutants were more rapidly cleared from the nasopharynges of inoculated mice compared with wild-type strains. A 2,908-bp fragment of scpA49 gene, obtained by PCR, was ligated to the expression vector pGEX-4T-1 and expressed in Escherichia coli. The affinity-purified ⌬SCPA49 protein proved to be highly immunogenic in mice and rabbits. Although the purified ⌬SCPA49 immunogen lacked enzymatic activity, it induced high titers of rabbit antibodies which were able to neutralize peptidase activity associated with M1, M6, M12, and M49 streptococci in vitro. This result confirmed that antipeptidase antibodies lack serotype specificity. Intranasal immunization of mice with the deleted form of the SCPA49 protein stimulated significant levels of specific salivary secretory immunoglobulin A (IgA) and serum IgG antibodies and reduced the potential of wild-type M1, M2, M6, M11, and M49 streptococci to colonize. These experiments suggest a new approach to vaccine development for prevention of streptococcal pharyngitis.

Signaling by a Conserved Quorum Sensing Pathway Contributes to Growth Ex Vivo and Oropharyngeal Colonization of Human Pathogen Group A Streptococcus

“…We next tested the hypothesis that the SIP signaling pathway contributes to GAS colonization of the mouse oropharynx. Although GAS is a humanonly pathogen, the murine model of oropharyngeal GAS colonization has been extensively used to investigate GAS survival at the oropharynx (35)(36)(37)(38)(39). Mice were infected intranasally with 10 8 CFU of each GAS strain.…”

Group A streptococcal pharyngitis: Immune responses involved in bacterial clearance and GAS-associated immunopathologies

“…SCPA cleaves the complement component C5a, a potent polymorphonuclear leukocyte chemotactic factor, and intranasal vaccination against SCPA prevented GAS nasopharyngeal colonization in mice. 35 SIC inhibits complement-mediated formation of the MAC by blocking the membrane-insertion site on C5b-7, 126 and inactivation of SIC impaired M1 GAS colonization of the mouse oropharynx. 121 GAS, like all Gram-positive bacteria, is also resistant to MAC-dependent lysis.…”

“…SCPA cleaves murine and human polymorphonuclear leukocyte chemotactic factor C5a, and intranasal vaccination against SCPA generated serum IgG/IgA responses and prevented GAS nasopharyngeal colonization of mice. 35 In a murine air-sac model, SCPA mutant GAS were cleared more efficiently than wild-type GAS and induced the infiltration of significantly higher numbers of polymorphonuclear leukocytes, 36 suggesting that subversion of leukocyte recruitment is critical for GAS to establish an infection. Immunization with SpyCEP (or ScpC), which cleaves chemokines that attract phagocytes, including neutrophils, protected mice against nasopharyngeal infection.…”

Virulence factors of the group A streptococci and genes that regulate their expression

“…Husmann et al suggested that the contribution of C5a peptidase might be strain dependent as they were unable to find a difference between mutant and wild type strains in their ability to colonize the throat or to cause pneumonia in mice (89). Evidence that C5a peptidase plays a role in virulence includes the increased clearance from subdermal infection sites and nasopharyngeal mucosa in scpA mutants and a strong antibody response to the protein in adults but not in uninfected children (88,90,91). This later response may, in part, explain why children are more susceptible to streptococcal infections than adults.…”