Streptolysin O (SLO) and streptolysin S (SLS) are potent cytolytic toxins produced by almost all clinical isolates of group A streptococci (GAS). Allele-replacement mutagenesis was used to construct nonpolar (in-frame) deletion mutations in the slo and sagB genes of the serotype M5 GAS strain Manfredo, producing isogenic single and double SLO-and SLS-defective mutants. In contrast to recent reports on SLS-defective insertion mutants ( Streptococcus pyogenes (group A streptococci [GAS]) is a highly versatile pathogen that causes a wide variety of important human diseases (55). These range in severity from selflimiting infections of the pharynx and skin to rapidly fulminating streptococcal toxic shock syndrome or devastating tissue infections, such as rapidly spreading myositis or necrotizing fasciitis (5, 55). GAS infections can also lead to serious poststreptococcal sequelae, such as acute glomerulonephritis or acute rheumatic fever (55). This versatility reflects, in part, differences in the abilities of strains to produce a wide range of known or suspected virulence factors. However, some virulence factors are highly conserved and expressed by almost all GAS strains. These include the potent cytolytic toxins streptolysin O (SLO) and streptolysin S (SLS) (2,35,36,51).SLO is a 540-amino-acid secreted protein toxin that binds to cholesterol in eukaryotic cell membranes, where it oligomerizes to produce large transmembrane pores leading to cell lysis (40). Sublytic concentrations of SLO have a wide variety of more subtle effects on target cells in vitro (8,9,29,46,50). In laboratory animals, intravenous (i.v.) administration of small quantities of purified SLO (Ͻ0.2 g in mice) causes death within 2 to 3 min, with cardiotoxicity being the prominent pathophysiological effect (1). Sublethal doses have been reported to have various effects on other tissues, including neurological effects, increased capillary permeability, and dermal necrosis in rabbits (1). Despite these highly potent in vitro and in vivo activities, the first direct studies on the role of SLO during an infection recently revealed that SLO contributed to a surprisingly limited extent to GAS virulence in a murine dermonecrosis-invasive infection model (24). No differences were observed in the abilities of a serotype M3 GAS strain and an isogenic slo mutant derivative to cause weight loss 24 h postinfection or induce necrotic lesions in this model, although SLO did contribute to later stages of infection, producing a significant, albeit small, difference in survival rates (24).SLS also has very potent cytolytic activity against a wide range of cells in vitro and produces a variety of more subtle effects at sublytic concentrations (1,16,17,37). Early studies showed that SLS is a nonimmunogenic peptide that loses activity upon separation from various "carrier" molecules (17, 58). However, its precise molecular nature remained an enigma until recent analysis of an SLS-defective transposon Tn916 insertion mutant identified an operon of nine SLSassociated ge...
