A natural inactivating mutation in the CovS component of the CovRS regulatory operon in a pattern D streptococcal pyogenes strain influences virulence-associated genes

A skin-tropic invasive group A Streptococcus pyogenes (GAS) strain, AP53, contains a natural inactivating mutation in the covS gene (covS^M) of the two-component responder (CovR)/sensor (CovS) gene regulatory system. The effects of this mutation on specific GAS virulence determinants have been assessed, with emphasis on expression of the extracellular protease, streptococcal pyrogenic exotoxin B (SpeB), capsular hyaluronic acid, and proteins that allow host plasmin assembly on the bacterial surface, viz. a high affinity plasminogen (Pg)/plasmin receptor, Pg-binding group A streptococcal M protein (PAM), and the human Pg activator streptokinase. To further illuminate mechanisms of the functioning of CovRS in the virulence of AP53, two AP53 isogenic strains were generated, one in which the natural covS^M gene was mutated to WT-covS (AP53/covS^WT) and a strain that contained an inactivated covR gene (AP53/ΔcovR). Two additional strains that do not contain PAM, viz. WT-NS931 and NS931/covS^M, were also employed. SpeB was not measurably expressed in strains containing covR^WT/covS ^M, whereas in strains with natural or engineered covR ^WT/covS^WT, SpeB expression was highly up-regulated. Alternatively, capsule synthesis via the hasABC operon was enhanced in strain AP53/covS^M, whereas streptokinase expression was only slightly affected by the covS inactivation. PAM expression was not substantially influenced by the covS mutation, suggesting that covRS had minimal effects on the mga regulon that controls PAM expression. These results demonstrate that a covS inactivation results in virulence gene alterations and also suggest that the CovR phosphorylation needed for gene up- or down-regulation can occur by alternative pathways to CovS kinase.