Most invasive bacterial infections are caused by species that more commonly colonize the human host with minimal symptoms. Although phenotypic or genetic correlates underlying a bacterium's shift to enhanced virulence have been studied, the in vivo selection pressure governing such shifts are poorly understood. The globally disseminated M1T1 clone of group A Streptococcus (GAS) is linked with rare but life-threatening syndromes of necrotizing fasciitis and toxic shock syndrome. Mutations in the GAS control of virulence regulatory sensor kinase (covRS) operon are associated with severe invasive disease, abolishing expression of a broad spectrum cysteine protease (SpeB)2,3 and allowing the recruitment and activation of host plasminogen on the bacterial surface. Here we describe how bacteriophage-encoded GAS DNase (Sda1), which facilitates the pathogen's escape from neutrophil extracellular traps (NETs)5,6, serves as a selective force for covRS mutation. The results provide a paradigm whereby natural selection exerted by the innate immune system generate hypervirulent bacterial variants with increased risk of systemic dissemination. Keywords CMMB Disciplines Life Sciences | Physical Sciences and Mathematics | Social and Behavioral Sciences Publication DetailsThis article was originally published as Walker, MJ et al, DNase Sda1 provides selection pressure for a switch to invasive group A streptococcal infection, Nature Medicine 13, 2007, 981- GAS is estimated to cause ~700 million cases of self-limited throat or skin infection each year worldwide 7 . Invasive GAS disease occurs in approximately 1/1,000 cases, with associated mortality of 25% 7 . Epidemic invasive disease is associated with the emergence of the globally disseminated GAS M1T1 clone 1,8 , which is distinguished from related strains by acquisition of prophages encoding virulence determinants such as superantigen SpeA and DNase Sda1 9,10 . In the M1T1 GAS clone, the transition from local to systemic infection can be linked to mutations in the two-component covRS regulator. The effect of these mutations is a distinct shift in the transcriptional 3 profile of invasive GAS isolates compared to mucosal (throat) isolates 3 . The covRS mutation and changes in gene expression are recapitulated upon subcutaneous challenge of mice and analysis of GAS disseminating to the spleen in comparison with those in the original inocolum 3 . Prominent changes in the transcriptional profile of invasive GAS isolates include a strong up-regulation of the DNase gene sda1, and a marked decrease in expression of the gene encoding the cysteine protease SpeB 3 .Sda1 is a virulence factor that protects GAS against neutrophil killing by degrading the DNA framework of NETs 5,6 . Abolishment of SpeB expression allows accumulation and activation of the broad spectrum host protease plasmin on the GAS bacterial surface 4 . A clinical correlation of GAS invasive disease severity and diminished SpeB expression has been established 2 .To elucidate the selection pressure for the rap...

SummaryMycoplasma hyopneumoniae , the causative agent of porcine enzootic pneumonia, colonizes the respiratory cilia of affected swine causing significant economic losses to swine production worldwide. Heparin is known to inhibit adherence of M. hyopneumoniae to porcine respiratory epithelial cilia. M. hyopneumoniae cells bind heparin but the identity of the heparinbinding proteins is limited. Proteomic analysis of M. hyopneumoniae lysates identified 27 kDa (P27), 110 kDa (P110) and 52 kDa (P52) proteins representing different regions of a 159 kDa (P159) protein derived from mhp494. These cleavage fragments were surface located and present at all growth stages.

Acute rheumatic fever is a serious autoimmune sequela of pharyngitis caused by certain group A streptococci. One mechanism applied by streptococcal strains capable of causing acute rheumatic fever is formation of an autoantigenic complex with human collagen IV. In some geographic regions with a high incidence of acute rheumatic fever pharyngeal carriage of group C and group G streptococci prevails. Examination of such strains revealed the presence of M-like surface proteins that bind human collagen. Using a peptide array and recombinant proteins with targeted amino acid substitutions, we could demonstrate that formation of collagen complexes during streptococcal infections depends on an octapeptide motif, which is present in collagen binding M and M-like proteins of different ␤-hemolytic streptococcal species. Mice immunized with streptococcal proteins that contain the collagen binding octapeptide motif developed high serum titers of anti-collagen antibodies. In sera of rheumatic fever patients such a collagen autoimmune response was accompanied by specific reactivity against the collagen-binding proteins, linking the observed effect to clinical cases. Taken together, the data demonstrate that the identified octapeptide motif through its action on collagen plays a crucial role in the pathogenesis of rheumatic fever. Eradication of streptococci that express proteins with the collagen binding motif appears advisable for controlling rheumatic fever. Acute rheumatic fever (ARF)3 is one of the most serious diseases caused by streptococci and occurs as an autoimmune sequela of untreated or inadequately treated group A streptococcal pharyngitis (1). ARF and the subsequent rheumatic heart disease (RHD) remain significant causes of cardiovascular disease today (2, 3). The most devastating effects are on children and young adults in their most productive years (2-4). According to a recent estimate more than 15 million people have RHD, more than 0.5 million acquire ARF each year, and about 0.25 million deaths annually are directly attributable to ARF or RHD (2). The fact that penicillin has clearly failed to eradicate ARF and that streptococcal vaccines are still years away from being available underlines the need for novel control strategies (5, 6). Identification of the pathogenic mechanisms underlying ARF is a prerequisite for the development of the necessary diagnostic and preventive approaches.Major virulence factors of streptococci that infect humans are the M and M-like proteins. They exert anti-phagocytic effects (7-10) and facilitate streptococcal survival within polymorph nuclear neutrophils (11). Variability in the N-terminal of M proteins generated more than 100 distinct M serotypes.

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MJ, M protein mediated plasminogen binding is essential for the virulence of an invasive Streptococcus pyogenes isolate, The FASEB Journal, 22(8), 2008, 2715-2722 M protein mediated plasminogen binding is essential for the virulence of an invasive Streptococcus pyogenes isolate AbstractThe human protease plasmin plays a crucial role in the capacity of the group A streptococcus (Streptococus pyogenes; GAS) to initiate invasive disease. The GAS strain NS88.2 was isolated from a case of bacteremia from the Northern Territory of Australia, a region with high rates of GAS invasive disease. Mutagenesis of the NS88.2 plasminogen binding M protein Prp was undertaken to examine the contribution of plasminogen binding and cell surface plasmin acquisition to virulence. The isogenic mutant NS88.2prp was engineered whereby four amino acid residues critical for plasminogen binding were converted to alanine codons in the GAS genome sequence. The mutated residues were reverse complemented to the wildtype sequence to construct GAS strain NS88.2prpRC. In comparison to NS88.2 and NS88.2prpRC, the NS88.2prp mutant exhibited significantly reduced ability to bind human plasminogen and accumulate cell surface plasmin activity during growth in human plasma. Utilising a humanised plasminogen mouse model of invasive infection, we demonstrate that the capacity to bind plasminogen and accumulate surface plasmin activity plays an essential role in GAS virulence. ABSTRACTThe human protease plasmin plays a crucial role in the capacity of the group A streptococcus (Streptococus pyogenes; GAS) to initiate invasive disease. The GAS strain NS88.2 was isolated from a case of bacteremia from the Northern Territory of Australia, a region with high rates of GAS invasive disease. Mutagenesis of the NS88.2 plasminogen binding M protein Prp was undertaken to examine the contribution of plasminogen binding and cell surface plasmin acquisition to virulence.The isogenic mutant NS88.2prp was engineered whereby four amino acid residues critical for plasminogen binding were converted to alanine codons in the GAS genome sequence. The mutated residues were reverse complemented to the wildtype sequence to construct GAS strain NS88.2prpRC. In comparison to NS88.2 and NS88.2prpRC, the NS88.2prp mutant exhibited significantly reduced ability to bind human plasminogen and accumulate cell surface plasmin activity during growth in human plasma. Utilising a humanised plasminogen mouse model of invasive infection, we demonstrate that the capacity to bind plasminogen and accumulate surface plasmin activity plays an essential role in GAS virulence.