Determination of the relationship between group A streptococcal genome content, M type, and toxic shock syndrome by a mixed genome microarray

Abstract

Group A streptococci (GAS), or Streptococcus pyogenes, are associated with a remarkable variety of diseases, ranging from superficial infections to life-threatening diseases such as toxic-shock-like syndrome (TSS). GAS strains belonging to M types M1 and M3 are associated with TSS. This study aims to obtain insight into the gene profiles underlying different M types and disease manifestations. Genomic differences between 76 clinically well characterized GAS strains collected in The Netherlands were examined using a mixed-genome microarray. Inter-M-type genomic differences clearly outweighed intra-M-type genome variation. Phages were major contributors to observed genome diversification. We identified four novel genes, including two genes encoding fibronectin-binding-like proteins, which are highly specific to a subset of M types and thus may contribute to M-type-associated disease manifestations. All M12 strains were characterized by the unique absence of the citrate lyase complex and reduced growth under hypoxic, nutrient-deprived conditions. Furthermore, six virulence factors, including genes encoding a complement-inhibiting protein (sic), an exotoxin (speA), iron(III) binding factor, collagen binding factor (cpa), and fibrinogen binding factor (prt2-like), were unique to M1 and/or M3 strains. These virulence factors may contribute to the potential of these strains to cause TSS. Finally, in contrast to M-type-specific virulence profiles, we did not identify a common virulence profile among strains associated with TSS irrespective of their M type.

FIG. 1.

2-D hierarchical clustering of 76 S. pyogenes strains. The dendrogram on the x axis shows the clustering of the strains. The dendrogram on the y axis shows the clustering of the 366 differentiating biomarkers. Red represents the presence, and green the absence, of a biomarker. Solid bars on the right, genes unique to one or two M types; open bars, genes that are uniquely absent in one or two M types (see Table 2). Twenty-five representative strains, indicated by arrows at the top, were subjected to PFGE (see Fig. S2 in the supplemental material).

FIG. 2.

Growth comparison between M12 strains (n = 14) and strains belonging to other M types (M1, M3, M28, M4, M6, M11, and M89; three random isolates per M type) under anaerobic, nutrient-rich versus anaerobic, nutrient-deprived conditions. Error bars, standard deviations. All experiments were conducted in duplicate. All OD measurements were normalized by subtracting the initial OD. The lag phase was quantified as the time needed for an increment of 0.2 in the OD. The maximum growth rate was defined as the maximum increase in OD between two measurements (every 20 min). The mean relative increase in lag time or decrease in maximum growth rate for the M12 strains under nutrient-deprived conditions was compared to that for other M types (by an unpaired t test).