Streptococcal β - hemolysins : genetics and role in disease pathogenesis

38, 6th ASM Conference on Streptococcal Genetics, Ashville, NC, USA]. GBS β-h/c contributes to virulence in animal studies. When administered to neonatal rats via the pulmonary route, NH GBS mutants possess a 1000-fold greater LD50 than the wild-type parent strains, confirming a role in the pathogenesis of pneumonia and systemic spread [52]. In an adult mouse model of GBS arthritis, hemolysin expression is associated with higher mortality, increased bacterial loads, greater degrees of joint injury, and release of the proinflammatory cytokines IL-6 and IL-1α systemically and intra-articularly [53]. Challenge of rabbits with isogenic GBS mutants showed that hemolysin production was associated with significantly higher mortality and evidence of liver necrosis with hepatocyte apoptosis [54]. It appears that GBS hemolysin is a pluripotent virulence factor that contributes to disease pathogenesis by cytotoxicity and inflammatory activation. Several virulence properties of GBS β-h/c can be blocked by DPPC, the major component of pulmonary surfactant. DPPC significantly reduces β-h/c-associated cellular toxicity [46,55], cytokine activation [51,56] and macrophage apoptosis [57]. Lack of DPPC inhibition of β-h/c toxicity might contribute to the increased incidence and severity of GBS pneumonia and sepsis in premature, surfactant-deficient neonates [58]. Experiments in the ventilated premature rabbit model of GBS pneumonia and retrospective analysis of clinical data from human neonates with early-onset GBS infection corroborate the beneficial effects of surfactant therapy against GBS-induced lung injury [59,60]. Conclusion The β-hemolysins of two major pathogenic streptococci, GAS and GBS, are both broad-spectrum cytotoxins that have long been postulated to play a role in the tissue injury and systemic spread associated with severe human infections. Although these toxins are responsible for a similar colony phenotype on blood-agar and share other phenotypic properties, the discovery of the genetic loci for GAS and GBS β-hemolysin production revealed their molecular basis is fundamentally distinct. Table 1 summarizes some of the known features of these two streptococcal β-hemolysins. Human leukocytes, epithelial cells and endothelial cells respond variably to the streptococcal β-hemolysins, with release of proinflammatory and/or chemotactic defense factors such as IL-8 (GBS) or cathelicidin antimicrobial peptides (GAS; V. Nizet and R.L. Gallo, unpublished). The ability to detect proportionately and respond to sub-cytolytic levels of these exotoxins is probably adaptive for the host in the vast majority of encounters – serving to orientate the innate immune response and contain the organism on the mucosal epithelial surface. However, in special circumstances such as the overwhelming in utero exposure of a premature, surfactant-deficient TRENDS in Microbiology Vol.10 No.12 December 2002 http://tim.trends.com 578 Review Table 1. Comparison of key features of GAS and GBS β-hemolysins Feature Streptolysin S (GAS) β-hemolysin/cytolysin (GBS) Spectrum and mode of action Broad, pore-forming Broad, pore-forming Genes required for β-hemolytic Entire nine-gene sag operon (except sagE?) A single gene (cylE) phenotype Genes sufficient to confer β-hemolytic Entire nine-gene sag operon (to Lactococcus A single gene (cylE) phenotype lactis) (to Escherichia coli) Predicted size 2.9 kDa 78 kDa Homologies Bacteriocin None Precise molecular structure Unknown Unknown Stabilizers RNA-core, lipoteichoic acid, albumin, non-ionic Starch, albumin, non-ionic detergents detergents Inhibitors Trypan blue, phospholipids (e.g. DPPC) Phospholipids (e.g. DPPC), proteases (e.g. subtilin) Linked phenotypes SpeB, M-protein ? Pigment production Found in other species? Yes (GCS, GGS, Streptococcus iniae) Unknown Naturally immunogenic? No No Experimentally immunogenic? Yes Unknown Proinflammatory effects Neutrophil activation Cytokines, nitric oxide Virulence factor (models)? Yes (necrotizing cellulitis) Yes (pneumonia, septicemia) Abbreviations: DPPC, dipalmitoyl phosphatidylcholine; GAS, group A Streptococcus; GBS, group B Streptococcus; GGS, group G Streptococcus; SpeB, streptococcal pyogenic exotoxin B. • Will the availability of new genetic information and targeted hemolysin-negative mutants allow purification and characterization of the mature β-hemolysin toxin proteins? • Does streptolysin S possess bacteriocin activity against other human microflora? • Through what signal transduction pathways (e.g. Toll-like receptors) does the GBS β-hemolysin/cytolysin activate cytokines and other innate immune response factors? • Will active immunization with unprocessed or inactivated streptococcal β-hemolysin toxins afford protection against subsequent challenge with the infectious organism? • Do strain differences in regulation of β-hemolysin production in vivo contribute to the epidemiology of invasive streptococcal infection? Questions for future research neonate to invasive GBS, over-activation of these same response pathways could contribute to massive release of cytokines and the manifestations of septic shock. More puzzling perhaps is the well-documented ability of GAS to cause life-threatening invasive infections such as necrotizing fasciitis and toxic shock syndrome in previously healthy children and young adults. Do these individuals have subtle deficits in their innate immune recognition of GAS SLS production, allowing deeper penetration and multiplication of the organism? Alternatively, do certain GAS strains ‘phase-shift’ in vivo towards constitutive high-level production of SLS with attendant injury to host tissues? Elucidating the molecular genetic basis of the streptococcal β-hemolysins promises to open a new era of detailed investigation of these unique and potent toxins. The availability of precise isogenic mutants and cloned hemolysin determinants should facilitate purification of these elusive proteins and allow careful analysis of their contribution to disease pathogenesis. Curiously, humans do not appear to generate neutralizing antibodies to either toxin during the course of natural infection. One can speculate that this non-immunogenicity derives from the potent cytotoxicity of these compounds against macrophages and other antigen-presenting cells at the initial steps of the humoral immune response pathways. The recent observations that non-toxic synthetic peptides of the SLS precursor maintain enough sequence features of the mature toxin to elicit neutralizing antibodies [17,27] suggest that subunits or inactivated toxoids of each hemolysin could have significant immunogenicity. Vaccine or chemotherapeutic strategies (e.g. phospholipid inhibitors) designed to neutralize β-hemolysin activity could be beneficial in managing human GAS and GBS infection. TRENDS in Microbiology Vol.10 No.12 December 2002 http://tim.trends.com 579 Review