The observation that highly encapsulated mutant CovS strains are attenuated in keratinocyte attachment suggested that the capsule might prevent the interaction of bacterial surface molecules with

specific receptors on keratinocytes by blocking the function of different adhesins through physical shielding. A similar finding was made previously by Darmstadt and co-workers, who reported that hyaluronic acid capsule impedes the interaction of bacterial adhesins with the keratinocyte receptor [30]. The adherence buy CHIR-99021 of a mutant lacking hyaluronic acid capsule (has mutants) was increased 13-fold [30]. Furthermore, Schrager and others pointed out that acapsular GAS exhibit enhanced adherence to human keratinocytes [28]. Therefore, we assume that CovS inactivation in different serotype GAS strains led to reduction in the adherence ability of the mutant strains in comparison with the corresponding wild type strains, which might be explained by the overexpressed capsule in the CovS defective mutants. However, the CovS influence on keratinocyte adherence among the tested GAS serotypes is apparently a uniform feature. Figure 4 Adherence to HaCaT cells. The adherence of CovS mutant strains is presented as a percentage of the data determined for the corresponding parental strains. The data represent the mean values of three independently performed experiments. *, the significance level (p < 0.05)

Torin 1 price for differences between wild type and isogenic mutant strains was determined by two-tailed paired

Student’s t test. Contribution of CovS to survival of GAS in whole blood GAS are known to be very well equipped for survival in whole human blood by expression of a diverse armamentarium of virulence factors that interfere with primary host defense mechanisms in the blood, in particular the complement system and phagocytosis [17]. Increased capsule expression leads to mucoid strains that are very often more virulent compared to unencapsulated strains [31] and have an increased resistance to phagocytic killing [1]. Thus, exponential-phase wild type and CovS mutant strains were tested for survival in whole human blood. As shown in Fig. 5, mutation of CovS in GAS serotypes M2, M6 and else M18 leads to a significantly reduced ability of the strains to survive and multiply in blood. This finding was unexpected since the increase in capsule amounts should allow for a better survival and multiplication. However, many other GAS surface-associated and secreted virulence factors have been described to act as defense against phagocytic killing [4, 32] and some of them might be more dominant in their protective effect compared to capsule. At least for M18 it was shown that capsule may be responsible for phagocytosis resistance in serum, whereas survival in blood to a larger extend relied on M protein expression [33]. Lack of CovS protein expression had no effect on blood survival of the GAS M49 serotype (Fig.