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Interspecies communication among commensal and pathogenic streptococci.

Cook LC, LaSarre B, Federle MJ - MBio (2013)

Bottom Line: Using isogenic mutant strains and transcriptional reporters, we confirmed that RovS/SHP1520 comprise a QS circuit in GBS.Environmental cues and interspecies signaling between members of the human flora likely play an important role in the transition to a disease state.Here we provide evidence of a bidirectional QS system between GAS, GBS, and Streptococcus dysgalactiae subsp. equisimilis, demonstrating the possibility of evolved communication systems between human pathogens.

View Article: PubMed Central - PubMed

Affiliation: Center for Pharmaceutical Biotechnology, Department of Medicinal Chemistry and Pharmacognosy, College of Pharmacy, University of Illinois at Chicago, Chicago, Illinois, USA.

ABSTRACT

Unlabelled: Quorum sensing (QS) regulates diverse and coordinated behaviors in bacteria, including the production of virulence factors, biofilm formation, sporulation, and competence development. It is now established that some streptococci utilize Rgg-type proteins in concert with short hydrophobic peptides (SHPs) to mediate QS, and sequence analysis reveals that several streptococcal species contain highly homologous Rgg/SHP pairs. In group A streptococcus (GAS), two SHPs (SHP2 and SHP3 [SHP2/3]) were previously identified to be important in GAS biofilm formation. SHP2/3 are detected by two antagonistic regulators, Rgg2 and Rgg3, which control expression of the shp genes. In group B streptococcus (GBS), RovS is a known virulence gene regulator and ortholog of Rgg2, whereas no apparent Rgg3 homolog exists. Adjacent to rovS is a gene (shp1520) encoding a peptide nearly identical to SHP2. Using isogenic mutant strains and transcriptional reporters, we confirmed that RovS/SHP1520 comprise a QS circuit in GBS. More important, we performed experiments demonstrating that production and secretion of SHP1520 by GBS can modulate Rgg2/3-regulated gene expression in GAS in trans; likewise, SHP2/3 production by GAS can stimulate RovS-mediated gene regulation in GBS. An isolate of Streptococcus dysgalactiae subsp. equisimilis also produced a secreted factor capable of simulating the QS circuits of both GAS and GBS, and sequencing confirms the presence of an orthologous Rgg2/SHP2 pair in this species as well. To our knowledge, this is the first documented case of bidirectional signaling between streptococcal species in coculture and suggests a role for orthologous Rgg/SHP systems in interspecies communication between important human pathogens.

Importance: Pathogenic streptococci, such as group A (GAS) and group B (GBS) streptococcus, are able to persist in the human body without causing disease but become pathogenic under certain conditions that are not fully characterized. Environmental cues and interspecies signaling between members of the human flora likely play an important role in the transition to a disease state. Since quorum-sensing (QS) peptides have been consistently shown to regulate virulence factor production in pathogenic species, the ability of bacteria to signal via these peptides may prove to be an important link between the carrier and pathogenic states. Here we provide evidence of a bidirectional QS system between GAS, GBS, and Streptococcus dysgalactiae subsp. equisimilis, demonstrating the possibility of evolved communication systems between human pathogens.

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GAS responds to signals produced by other species, and the GBS signal is dependent on RovS and SHP1520. A shp- GAS strain containing a Pshp2 –lux reporter, BNL177, was grown to log phase and diluted in spent culture supernatants from various strains. Luminescence expression indicates induction of the SHP system in GAS. (A) SHP2 expression is activated by spent culture supernatants from WT GBS (A909) and S. dysgalactiae subsp. equisimilis (GGS-LT1), as well as a Δrgg3 GAS strain (JCC131) that produces high levels of SHP peptide. Fresh medium and WT GAS (NZ131) supernatant do not induce the reporter. (B) Production of the secreted inducer in GBS supernatant is dependent on RovS, SHP1520, and the putative RovS binding site. Spent supernatants from GBS strains with mutations in rovS (BLC100), shp1520 (BLC101), both rovS and shp1520 (BLC102), or the putative RovS binding site (Pshp1520mut; BLC103) are unable to induce luminescence expression in BNL177, unlike the WT A909 parent strain. Data are shown as a function of relative light units (CPS/OD600) versus OD600. Each graph is representative of at least 3 biological replicates.
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fig2: GAS responds to signals produced by other species, and the GBS signal is dependent on RovS and SHP1520. A shp- GAS strain containing a Pshp2 –lux reporter, BNL177, was grown to log phase and diluted in spent culture supernatants from various strains. Luminescence expression indicates induction of the SHP system in GAS. (A) SHP2 expression is activated by spent culture supernatants from WT GBS (A909) and S. dysgalactiae subsp. equisimilis (GGS-LT1), as well as a Δrgg3 GAS strain (JCC131) that produces high levels of SHP peptide. Fresh medium and WT GAS (NZ131) supernatant do not induce the reporter. (B) Production of the secreted inducer in GBS supernatant is dependent on RovS, SHP1520, and the putative RovS binding site. Spent supernatants from GBS strains with mutations in rovS (BLC100), shp1520 (BLC101), both rovS and shp1520 (BLC102), or the putative RovS binding site (Pshp1520mut; BLC103) are unable to induce luminescence expression in BNL177, unlike the WT A909 parent strain. Data are shown as a function of relative light units (CPS/OD600) versus OD600. Each graph is representative of at least 3 biological replicates.

Mentions: To examine if GBS strain A909 or S. dysgalactiae subsp. equisimilis strain GGS-LT1 produced a secreted factor capable of stimulating the Rgg2/3 system of GAS, cell-free spent culture supernatants were tested for their ability to induce the reporter strain BNL177. As negative and positive controls, respectively, supernatants from wild-type (WT) GAS and an isogenic Δrgg3 mutant were also included, since the QS system is known to be off in WT GAS but constitutively on in the Δrgg3 mutant (26). As expected, no reporter induction was detected when BNL177 was grown in fresh medium or in spent supernatant from WT GAS, whereas spent supernatant from the GAS Δrgg3 mutant induced reporter expression (Fig. 2A). More importantly, spent supernatants from both GBS and S. dysgalactiae subsp. equisimilis induced robust reporter expression to levels even greater than that of the positive control (Fig. 2A). These results were the first evidence that a secreted factor produced by GBS A909 and S. dysgalactiae subsp. equisimilis GGS-LT1 was indeed capable of inducing shp gene expression in GAS.


Interspecies communication among commensal and pathogenic streptococci.

Cook LC, LaSarre B, Federle MJ - MBio (2013)

GAS responds to signals produced by other species, and the GBS signal is dependent on RovS and SHP1520. A shp- GAS strain containing a Pshp2 –lux reporter, BNL177, was grown to log phase and diluted in spent culture supernatants from various strains. Luminescence expression indicates induction of the SHP system in GAS. (A) SHP2 expression is activated by spent culture supernatants from WT GBS (A909) and S. dysgalactiae subsp. equisimilis (GGS-LT1), as well as a Δrgg3 GAS strain (JCC131) that produces high levels of SHP peptide. Fresh medium and WT GAS (NZ131) supernatant do not induce the reporter. (B) Production of the secreted inducer in GBS supernatant is dependent on RovS, SHP1520, and the putative RovS binding site. Spent supernatants from GBS strains with mutations in rovS (BLC100), shp1520 (BLC101), both rovS and shp1520 (BLC102), or the putative RovS binding site (Pshp1520mut; BLC103) are unable to induce luminescence expression in BNL177, unlike the WT A909 parent strain. Data are shown as a function of relative light units (CPS/OD600) versus OD600. Each graph is representative of at least 3 biological replicates.
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Related In: Results  -  Collection

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fig2: GAS responds to signals produced by other species, and the GBS signal is dependent on RovS and SHP1520. A shp- GAS strain containing a Pshp2 –lux reporter, BNL177, was grown to log phase and diluted in spent culture supernatants from various strains. Luminescence expression indicates induction of the SHP system in GAS. (A) SHP2 expression is activated by spent culture supernatants from WT GBS (A909) and S. dysgalactiae subsp. equisimilis (GGS-LT1), as well as a Δrgg3 GAS strain (JCC131) that produces high levels of SHP peptide. Fresh medium and WT GAS (NZ131) supernatant do not induce the reporter. (B) Production of the secreted inducer in GBS supernatant is dependent on RovS, SHP1520, and the putative RovS binding site. Spent supernatants from GBS strains with mutations in rovS (BLC100), shp1520 (BLC101), both rovS and shp1520 (BLC102), or the putative RovS binding site (Pshp1520mut; BLC103) are unable to induce luminescence expression in BNL177, unlike the WT A909 parent strain. Data are shown as a function of relative light units (CPS/OD600) versus OD600. Each graph is representative of at least 3 biological replicates.
Mentions: To examine if GBS strain A909 or S. dysgalactiae subsp. equisimilis strain GGS-LT1 produced a secreted factor capable of stimulating the Rgg2/3 system of GAS, cell-free spent culture supernatants were tested for their ability to induce the reporter strain BNL177. As negative and positive controls, respectively, supernatants from wild-type (WT) GAS and an isogenic Δrgg3 mutant were also included, since the QS system is known to be off in WT GAS but constitutively on in the Δrgg3 mutant (26). As expected, no reporter induction was detected when BNL177 was grown in fresh medium or in spent supernatant from WT GAS, whereas spent supernatant from the GAS Δrgg3 mutant induced reporter expression (Fig. 2A). More importantly, spent supernatants from both GBS and S. dysgalactiae subsp. equisimilis induced robust reporter expression to levels even greater than that of the positive control (Fig. 2A). These results were the first evidence that a secreted factor produced by GBS A909 and S. dysgalactiae subsp. equisimilis GGS-LT1 was indeed capable of inducing shp gene expression in GAS.

Bottom Line: Using isogenic mutant strains and transcriptional reporters, we confirmed that RovS/SHP1520 comprise a QS circuit in GBS.Environmental cues and interspecies signaling between members of the human flora likely play an important role in the transition to a disease state.Here we provide evidence of a bidirectional QS system between GAS, GBS, and Streptococcus dysgalactiae subsp. equisimilis, demonstrating the possibility of evolved communication systems between human pathogens.

View Article: PubMed Central - PubMed

Affiliation: Center for Pharmaceutical Biotechnology, Department of Medicinal Chemistry and Pharmacognosy, College of Pharmacy, University of Illinois at Chicago, Chicago, Illinois, USA.

ABSTRACT

Unlabelled: Quorum sensing (QS) regulates diverse and coordinated behaviors in bacteria, including the production of virulence factors, biofilm formation, sporulation, and competence development. It is now established that some streptococci utilize Rgg-type proteins in concert with short hydrophobic peptides (SHPs) to mediate QS, and sequence analysis reveals that several streptococcal species contain highly homologous Rgg/SHP pairs. In group A streptococcus (GAS), two SHPs (SHP2 and SHP3 [SHP2/3]) were previously identified to be important in GAS biofilm formation. SHP2/3 are detected by two antagonistic regulators, Rgg2 and Rgg3, which control expression of the shp genes. In group B streptococcus (GBS), RovS is a known virulence gene regulator and ortholog of Rgg2, whereas no apparent Rgg3 homolog exists. Adjacent to rovS is a gene (shp1520) encoding a peptide nearly identical to SHP2. Using isogenic mutant strains and transcriptional reporters, we confirmed that RovS/SHP1520 comprise a QS circuit in GBS. More important, we performed experiments demonstrating that production and secretion of SHP1520 by GBS can modulate Rgg2/3-regulated gene expression in GAS in trans; likewise, SHP2/3 production by GAS can stimulate RovS-mediated gene regulation in GBS. An isolate of Streptococcus dysgalactiae subsp. equisimilis also produced a secreted factor capable of simulating the QS circuits of both GAS and GBS, and sequencing confirms the presence of an orthologous Rgg2/SHP2 pair in this species as well. To our knowledge, this is the first documented case of bidirectional signaling between streptococcal species in coculture and suggests a role for orthologous Rgg/SHP systems in interspecies communication between important human pathogens.

Importance: Pathogenic streptococci, such as group A (GAS) and group B (GBS) streptococcus, are able to persist in the human body without causing disease but become pathogenic under certain conditions that are not fully characterized. Environmental cues and interspecies signaling between members of the human flora likely play an important role in the transition to a disease state. Since quorum-sensing (QS) peptides have been consistently shown to regulate virulence factor production in pathogenic species, the ability of bacteria to signal via these peptides may prove to be an important link between the carrier and pathogenic states. Here we provide evidence of a bidirectional QS system between GAS, GBS, and Streptococcus dysgalactiae subsp. equisimilis, demonstrating the possibility of evolved communication systems between human pathogens.

Show MeSH
Related in: MedlinePlus