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Evolution of Salmonella enterica virulence via point mutations in the fimbrial adhesin.

Kisiela DI, Chattopadhyay S, Libby SJ, Karlinsey JE, Fang FC, Tchesnokova V, Kramer JJ, Beskhlebnaya V, Samadpour M, Grzymajlo K, Ugorski M, Lankau EW, Mackie RI, Clegg S, Sokurenko EV - PLoS Pathog. (2012)

Bottom Line: We have found that while the low-binding shear-dependent phenotype of the adhesin is preserved in broad host-range (usually systemically non-invasive) Salmonella, the majority of host-adapted serovars express FimH variants with one of two alternative phenotypes: a significantly increased binding to mannose (as in S.The functional diversification of FimH in host-adapted Salmonella results from recently acquired structural mutations.In conclusion, our study demonstrates that point mutations are the target of positive selection and, in addition to horizontal gene transfer and genome degradation events, can contribute to the differential pathoadaptive evolution of Salmonella.

View Article: PubMed Central - PubMed

Affiliation: Department of Microbiology, University of Washington, Seattle, Washington, United States of America.

ABSTRACT
Whereas the majority of pathogenic Salmonella serovars are capable of infecting many different animal species, typically producing a self-limited gastroenteritis, serovars with narrow host-specificity exhibit increased virulence and their infections frequently result in fatal systemic diseases. In our study, a genetic and functional analysis of the mannose-specific type 1 fimbrial adhesin FimH from a variety of serovars of Salmonella enterica revealed that specific mutant variants of FimH are common in host-adapted (systemically invasive) serovars. We have found that while the low-binding shear-dependent phenotype of the adhesin is preserved in broad host-range (usually systemically non-invasive) Salmonella, the majority of host-adapted serovars express FimH variants with one of two alternative phenotypes: a significantly increased binding to mannose (as in S. Typhi, S. Paratyphi C, S. Dublin and some isolates of S. Choleraesuis), or complete loss of the mannose-binding activity (as in S. Paratyphi B, S. Choleraesuis and S. Gallinarum). The functional diversification of FimH in host-adapted Salmonella results from recently acquired structural mutations. Many of the mutations are of a convergent nature indicative of strong positive selection. The high-binding phenotype of FimH that leads to increased bacterial adhesiveness to and invasiveness of epithelial cells and macrophages usually precedes acquisition of the non-binding phenotype. Collectively these observations suggest that activation or inactivation of mannose-specific adhesive properties in different systemically invasive serovars of Salmonella reflects their dynamic trajectories of adaptation to a life style in specific hosts. In conclusion, our study demonstrates that point mutations are the target of positive selection and, in addition to horizontal gene transfer and genome degradation events, can contribute to the differential pathoadaptive evolution of Salmonella.

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Binding phenotypes of natural S. enterica subspecies I FimH variants.Static adhesion of S. Typhimurium LBH4 transformed with plasmids encoding different variants of FimH or plasmids carrying fimH deletion (fimHΔ) to Man-BSA (red bars) and anti-FimHSE antibody (grey bars). The binding of 3H- labeled bacteria was determined as described in Material and Methods. Data are the means ± SD of triplicates from one representative experiment of three experiments that were performed. The strain tags of systemically invasive serovars are in red and the non-invasive ones in black. Bacterial binding was >95% inhibitable in the presence of 50 mM methyl-D-mannopyranoside (not shown). * The non-binding FimH variants of S. Gallinarum were not tested in this study.
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ppat-1002733-g004: Binding phenotypes of natural S. enterica subspecies I FimH variants.Static adhesion of S. Typhimurium LBH4 transformed with plasmids encoding different variants of FimH or plasmids carrying fimH deletion (fimHΔ) to Man-BSA (red bars) and anti-FimHSE antibody (grey bars). The binding of 3H- labeled bacteria was determined as described in Material and Methods. Data are the means ± SD of triplicates from one representative experiment of three experiments that were performed. The strain tags of systemically invasive serovars are in red and the non-invasive ones in black. Bacterial binding was >95% inhibitable in the presence of 50 mM methyl-D-mannopyranoside (not shown). * The non-binding FimH variants of S. Gallinarum were not tested in this study.

Mentions: We next compared functional properties of different structural variants of FimH by examining the level of bacterial binding to Mannose-BSA (Man-BSA) in an isogenic system. Man-BSA, used as model substrate, contains single (mono-) mannose residues, Man1, covalently coupled to BSA. As controls for assessing the level of binding, we used a fimH-knockout variant (fimHΔ) as well as three structural variants of S. Typhimurium FimH characterized previously: a relatively low-binding variant of S. Typhimurium SL1344 (Thm1) and two high-binding variants from strains AJB3 (Thm3) and 5010 (Thm4) [38], [53], [55] (Figure 4). The majority of FimH variants exhibited relatively low but specific (>95% mannose-inhibitable) binding to Man-BSA (Figure 4 and data not shown). However, with the exception of FimH from S. Paratyphi A/Sendai, all of the low binding FimH variants came from systemically non-invasive serovars of S. enterica. In contrast, all high-binding FimH variants were from systemically invasive serovars such as Typhi (strains Typ1–Typ4), Paratyphi C (PaC1), and Choleraesuis (strains Chl5–Chl6). Also, FimH from another systemically invasive serovar Dublin (Dub1–Dub7) bound Man-BSA significantly stronger than low-adhesive FimH variants, though the binding was weaker in comparison to the other high-binding FimH variants. The binding differences were not due to a difference in the expression level of FimH, as bacteria expressing different variants of FimH bound relatively well to polyclonal anti-FimH antibody (Figure 4). Interestingly, FimH variants expressed by the remainder of the invasive strains did not exhibit detectable mannose-specific binding to Man-BSA, and also failed to bind Man5 oligosaccharides carried by ribonuclease B (RNase B, Figure S1), to which all functionally-active FimH variants bind with much higher affinity than Man1 ligands [55]. However, they still retained their interaction with anti-FimH antibodies (Figure 4). Such an ‘inactive’ FimH phenotype may be similar to that shown previously for FimH variants of S. Gallinarum biovars Pullorum (Figure 4 and Figure S1) and Gallinarum ([54], not tested in this study).


Evolution of Salmonella enterica virulence via point mutations in the fimbrial adhesin.

Kisiela DI, Chattopadhyay S, Libby SJ, Karlinsey JE, Fang FC, Tchesnokova V, Kramer JJ, Beskhlebnaya V, Samadpour M, Grzymajlo K, Ugorski M, Lankau EW, Mackie RI, Clegg S, Sokurenko EV - PLoS Pathog. (2012)

Binding phenotypes of natural S. enterica subspecies I FimH variants.Static adhesion of S. Typhimurium LBH4 transformed with plasmids encoding different variants of FimH or plasmids carrying fimH deletion (fimHΔ) to Man-BSA (red bars) and anti-FimHSE antibody (grey bars). The binding of 3H- labeled bacteria was determined as described in Material and Methods. Data are the means ± SD of triplicates from one representative experiment of three experiments that were performed. The strain tags of systemically invasive serovars are in red and the non-invasive ones in black. Bacterial binding was >95% inhibitable in the presence of 50 mM methyl-D-mannopyranoside (not shown). * The non-binding FimH variants of S. Gallinarum were not tested in this study.
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Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3369946&req=5

ppat-1002733-g004: Binding phenotypes of natural S. enterica subspecies I FimH variants.Static adhesion of S. Typhimurium LBH4 transformed with plasmids encoding different variants of FimH or plasmids carrying fimH deletion (fimHΔ) to Man-BSA (red bars) and anti-FimHSE antibody (grey bars). The binding of 3H- labeled bacteria was determined as described in Material and Methods. Data are the means ± SD of triplicates from one representative experiment of three experiments that were performed. The strain tags of systemically invasive serovars are in red and the non-invasive ones in black. Bacterial binding was >95% inhibitable in the presence of 50 mM methyl-D-mannopyranoside (not shown). * The non-binding FimH variants of S. Gallinarum were not tested in this study.
Mentions: We next compared functional properties of different structural variants of FimH by examining the level of bacterial binding to Mannose-BSA (Man-BSA) in an isogenic system. Man-BSA, used as model substrate, contains single (mono-) mannose residues, Man1, covalently coupled to BSA. As controls for assessing the level of binding, we used a fimH-knockout variant (fimHΔ) as well as three structural variants of S. Typhimurium FimH characterized previously: a relatively low-binding variant of S. Typhimurium SL1344 (Thm1) and two high-binding variants from strains AJB3 (Thm3) and 5010 (Thm4) [38], [53], [55] (Figure 4). The majority of FimH variants exhibited relatively low but specific (>95% mannose-inhibitable) binding to Man-BSA (Figure 4 and data not shown). However, with the exception of FimH from S. Paratyphi A/Sendai, all of the low binding FimH variants came from systemically non-invasive serovars of S. enterica. In contrast, all high-binding FimH variants were from systemically invasive serovars such as Typhi (strains Typ1–Typ4), Paratyphi C (PaC1), and Choleraesuis (strains Chl5–Chl6). Also, FimH from another systemically invasive serovar Dublin (Dub1–Dub7) bound Man-BSA significantly stronger than low-adhesive FimH variants, though the binding was weaker in comparison to the other high-binding FimH variants. The binding differences were not due to a difference in the expression level of FimH, as bacteria expressing different variants of FimH bound relatively well to polyclonal anti-FimH antibody (Figure 4). Interestingly, FimH variants expressed by the remainder of the invasive strains did not exhibit detectable mannose-specific binding to Man-BSA, and also failed to bind Man5 oligosaccharides carried by ribonuclease B (RNase B, Figure S1), to which all functionally-active FimH variants bind with much higher affinity than Man1 ligands [55]. However, they still retained their interaction with anti-FimH antibodies (Figure 4). Such an ‘inactive’ FimH phenotype may be similar to that shown previously for FimH variants of S. Gallinarum biovars Pullorum (Figure 4 and Figure S1) and Gallinarum ([54], not tested in this study).

Bottom Line: We have found that while the low-binding shear-dependent phenotype of the adhesin is preserved in broad host-range (usually systemically non-invasive) Salmonella, the majority of host-adapted serovars express FimH variants with one of two alternative phenotypes: a significantly increased binding to mannose (as in S.The functional diversification of FimH in host-adapted Salmonella results from recently acquired structural mutations.In conclusion, our study demonstrates that point mutations are the target of positive selection and, in addition to horizontal gene transfer and genome degradation events, can contribute to the differential pathoadaptive evolution of Salmonella.

View Article: PubMed Central - PubMed

Affiliation: Department of Microbiology, University of Washington, Seattle, Washington, United States of America.

ABSTRACT
Whereas the majority of pathogenic Salmonella serovars are capable of infecting many different animal species, typically producing a self-limited gastroenteritis, serovars with narrow host-specificity exhibit increased virulence and their infections frequently result in fatal systemic diseases. In our study, a genetic and functional analysis of the mannose-specific type 1 fimbrial adhesin FimH from a variety of serovars of Salmonella enterica revealed that specific mutant variants of FimH are common in host-adapted (systemically invasive) serovars. We have found that while the low-binding shear-dependent phenotype of the adhesin is preserved in broad host-range (usually systemically non-invasive) Salmonella, the majority of host-adapted serovars express FimH variants with one of two alternative phenotypes: a significantly increased binding to mannose (as in S. Typhi, S. Paratyphi C, S. Dublin and some isolates of S. Choleraesuis), or complete loss of the mannose-binding activity (as in S. Paratyphi B, S. Choleraesuis and S. Gallinarum). The functional diversification of FimH in host-adapted Salmonella results from recently acquired structural mutations. Many of the mutations are of a convergent nature indicative of strong positive selection. The high-binding phenotype of FimH that leads to increased bacterial adhesiveness to and invasiveness of epithelial cells and macrophages usually precedes acquisition of the non-binding phenotype. Collectively these observations suggest that activation or inactivation of mannose-specific adhesive properties in different systemically invasive serovars of Salmonella reflects their dynamic trajectories of adaptation to a life style in specific hosts. In conclusion, our study demonstrates that point mutations are the target of positive selection and, in addition to horizontal gene transfer and genome degradation events, can contribute to the differential pathoadaptive evolution of Salmonella.

Show MeSH
Related in: MedlinePlus