Functional Analysis of the Quorum-Sensing Streptococcal Invasion Locus (sil)

Abstract

Group A streptococcus (GAS) causes a wide variety of human diseases, and at the same time, GAS can also circulate without producing symptoms, similar to its close commensal relative, group G streptococcus (GGS). We previously identified, by transposon-tagged mutagenesis, the streptococcal invasion locus (sil). sil is a quorum-sensing regulated locus which is activated by the autoinducer peptide SilCR through the two-component system SilA-SilB. Here we characterize the DNA promoter region necessary for SilA-mediated activation. This site is composed of two direct repeats of 10 bp, separated by a spacer of 11 bp. Fusion of this site to gfp allowed us to systematically introduce single-base substitutions in the repeats region and to assess the relative contribution of various positions to promoter strength. We then developed an algorithm giving different weights to these positions, and performed a chromosome-wide bioinformatics search which was validated by transcriptome analysis. We identified 13 genes, mostly bacteriocin related, that are directly under the control of SilA. Having developed the ability to quantify SilCR signaling via GFP accumulation prompted us to search for GAS and GGS strains that sense and produce SilCR. While the majority of GAS strains lost sil, all GGS strains examined still possess the locus and ∼63% are able to respond to exogenously added SilCR. By triggering the autoinduction circle using a minute concentration of synthetic SilCR, we identified GAS and GGS strains that are capable of sensing and naturally producing SilCR, and showed that SilCR can be sensed across these streptococci species. These findings suggest that sil may be involved in colonization and establishment of commensal host-bacterial relationships.

Molecular Epidemiology of sil Locus in Clinical Streptococcus pyogenes Strains

“…This contrasts with the situation in JS95, the strain in which the sil locus was originally described (13,19).…”

“…To identify a possible mutation in the silCR translational start codon as previously described for the JS95 (emm14) strain, we sequenced silCR in the 206 sil locus-positive strains (13,19). Among these strains, no mutation in the silCR start codon was observed; a functional translational start codon was present in silCR of all strains (Fig.…”

“…1 and Table 2) were used to amplify a 860-bp fragment encompassing silC and silCR and a silD gene, respectively. PCR products were sequenced and compared to the published sequence of the JS95 strain in order to identify mutations in this fragment, including the previously described silCR start codon mutation (13,19).…”

“…For this reason, it was deemed important to study its in-volvement in the virulence of other GAS isolates. Previous studies showed that most of the GAS invasive strains either lost sil completely or retained a mutated sil locus (19). A study conducted in Japan on noninvasive strains showed a prevalence of 12% for the sil locus (20).…”

“…However, this result may be GAS strain dependent (18). In the highly invasive strain JS95, the sil locus is inactive due to a point mutation in the start codon of silCR (ATA instead of ATG) (13,19). Thereby, the state of the sil locus seems to account for the virulence of strains such as GAS JS95, which is important as no locus has been strictly associated with GAS virulence.…”