Genetic Inactivation of an Extracellular Cysteine Protease (SpeB) Expressed byStreptococcus pyogenesDecreases Resistance to Phagocytosis and Dissemination to Organs

Abstract

Streptococcal pyrogenic exotoxin B (SpeB), a conserved cysteine protease expressed by virtually all Streptococcus pyogenes strains, has recently been shown to be an important virulence factor (S. Lukomski, S. Sreevatsan, C. Amberg, W. Reichardt, M. Woischnik, A. Podbielski, and J. M. Musser, J. Clin. Invest. 99:2574–2580, 1997). Genetic inactivation of SpeB significantly decreased the lethality of a serotype M49 strain for mice and abolished the lethality of a serotype M3 strain after intraperitoneal (i.p.) injection. In the present study, a wild-type M3 isolate and an M3 speB mutant derivative were used to investigate the mechanism responsible for altered virulence. Following i.p. injection, the mutant and wild-type strains induced virtually identical cellular inflammatory responses, characterized largely by an influx of polymorphonuclear leukocytes (PMNs). In addition, the mutant and wild-type strains rapidly entered the blood and were recovered from all organs examined. However, significantly fewer (P < 0.05) CFUs of the isogenic mutant derivative than of the wild-type parent strain were recovered from blood and organs. PMNs effectively cleared the M3 speB mutant from the peritoneum by 22 h, thereby sparing the host. In contrast, the wild-type M3 strain continued to replicate intraperitoneally and had the ability to kill phagocytes. This process allowed the wild-type strain to continuously disseminate, resulting in host death. Our results indicate that genetic inactivation of the cysteine protease decreased the resistance of the mutant to phagocytosis and impaired its subsequent dissemination to organs. These results provide insight into the detrimental effect of SpeB inactivation on virulence.

Adhesion determinants of the Streptococcus species

“…The SpeB is an extracellular protease; however, it has been shown to be surface exposed with binding activity to Lm and other glycoproteins (Hytonen et al, 2001). In addition, different activities were ascribed to SpeB: it was demonstrated to directly bind to integrins, and, as a secreted molecule to degrade host ECM proteins, thus contributing to bacterial dissemination, colonization and invasion (Kapur et al, 1993;Lukomski et al, 1998). BLAST sequence analysis revealed that SpeB is specific for S. pyogenes, whereas Lmb/Lbp homologues, with an extremely high degree of conservation, are present in several other Streptococcus species, including S. pneumoniae, S. gordonii and S. sanguinis (Fig.…”

Is streptolysin S of group A streptococci a virulence factor?

“…Revertants of the mutants in which Tn916 had been excised regained the hemolytic phenotype and the virulence profile of the wild-type strain. A cysteine proteinaseproducing streptococcal strain, which presumably also produced SLS, participated in the pathogenesis of invasive skin infections in mice (50) and augmented, in a synergistic manner, lung injury induced by several products of group A streptococci, including SLO and cellwall components (51). On the other hand, a genetic inactivation of an extracellular cysteine proteinase decreased resistance to phagocytosis and dissemination to organs (52).…”

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

“…In the case of the streptococcal protease a number of different roles in the disease process have been investigated with effects on producer strain proteins, the host immune system, and degradation of host extracellular matrix material (5, 6, 49,169,174). Insertional inactivation of the gene decreased mouse lethality (175)(176)(177), decreased resistance to phagocytosis (178), enhanced internalization by human cells using a fibronectin-dependent process (107,179), eliminated fibrinogen cleavage (180), and in a strain specific manner affected expression of the hyaluronic acid capsule (181)(182)(183). Other results, again using an insertional mutant, suggested that SpeB has no effect on virulence (182,184).…”