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Reprogramming of Yersinia from virulent to persistent mode revealed by complex in vivo RNA-seq analysis.

Avican K, Fahlgren A, Huss M, Heroven AK, Beckstette M, Dersch P, Fällman M - PLoS Pathog. (2015)

Bottom Line: We found that the Crp/CsrA/RovA regulatory cascades influence the pattern of bacterial gene expression during persistence.Furthermore, arcA, fnr, frdA, and wrbA play critical roles in persistence.Our findings suggest a model for the life cycle of this enteropathogen with reprogramming from a virulent to an adapted phenotype capable of persisting and spreading by fecal shedding.

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular Biology, Umeå Centre for Microbial Research (UCMR), Umeå University, Umeå, Sweden; Department of Molecular Biology, Laboratory for Molecular Infection Medicine Sweden (MIMS), Umea University, Umeå, Sweden.

ABSTRACT
We recently found that Yersinia pseudotuberculosis can be used as a model of persistent bacterial infections. We performed in vivo RNA-seq of bacteria in small cecal tissue biopsies at early and persistent stages of infection to determine strategies associated with persistence. Comprehensive analysis of mixed RNA populations from infected tissues revealed that Y. pseudotuberculosis undergoes transcriptional reprogramming with drastic down-regulation of T3SS virulence genes during persistence when the pathogen resides within the cecum. At the persistent stage, the expression pattern in many respects resembles the pattern seen in vitro at 26oC, with for example, up-regulation of flagellar genes and invA. These findings are expected to have impact on future rationales to identify suitable bacterial targets for new antibiotics. Other genes that are up-regulated during persistence are genes involved in anaerobiosis, chemotaxis, and protection against oxidative and acidic stress, which indicates the influence of different environmental cues. We found that the Crp/CsrA/RovA regulatory cascades influence the pattern of bacterial gene expression during persistence. Furthermore, arcA, fnr, frdA, and wrbA play critical roles in persistence. Our findings suggest a model for the life cycle of this enteropathogen with reprogramming from a virulent to an adapted phenotype capable of persisting and spreading by fecal shedding.

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Y. pseudotuberculosis undergoes transcriptional reprogramming for adaption to persistence.(A) Comparison of genes up-regulated in Y. pseudotuberculosis in vitro at 26°C and 37°C compared to in vivo during early (2 dpi) and persistent (42 dpi) stages of infection. Similarities are shown with the number of genes up-regulated in both groups. (B) Functional annotation of Y. pseudotuberculosis genes up-regulated during early and persistent infection (KEGG pathway mapping tool). (C) Comparison of the in vivo gene expression profiles and the expression profiles of bacteria grown under anaerobic conditions in vitro. The analysis included genes up-regulated (>1.8-fold) during anaerobic or aerobic growth in both the exponential and stationary growth phase compared to genes up-regulated during early and persistent infection. Similarities are shown with the number of genes up-regulated in both groups.
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ppat.1004600.g005: Y. pseudotuberculosis undergoes transcriptional reprogramming for adaption to persistence.(A) Comparison of genes up-regulated in Y. pseudotuberculosis in vitro at 26°C and 37°C compared to in vivo during early (2 dpi) and persistent (42 dpi) stages of infection. Similarities are shown with the number of genes up-regulated in both groups. (B) Functional annotation of Y. pseudotuberculosis genes up-regulated during early and persistent infection (KEGG pathway mapping tool). (C) Comparison of the in vivo gene expression profiles and the expression profiles of bacteria grown under anaerobic conditions in vitro. The analysis included genes up-regulated (>1.8-fold) during anaerobic or aerobic growth in both the exponential and stationary growth phase compared to genes up-regulated during early and persistent infection. Similarities are shown with the number of genes up-regulated in both groups.

Mentions: A total of 1288 genes were found to be differentially expressed (log2 fold change ≥0.7) in vivo. RPKMO values detected by RNA-seq are shown in a histogram and heat map in Fig. 3A to highlight the differences in individual ORFs during early and persistent infection (see also S3 Table). Surprisingly, the T3SS components encoded on the virulence plasmid that were highly expressed during the early stage of infection were distinctly down-regulated during persistent infection. Another conspicuous finding was the up-regulation of flagella and chemotaxis genes. T3SS is known to be induced at 37°C, but flagella are down-regulated at this temperature; in vitro, flagella are expressed only at 26°C (confirmed by qPCR in the same samples used in RNA-seq; Fig. 4A–B). In analogy, the T3SS master regulator lcrF was up-regulated during early infection and down-regulated during persistence (S3 Table), and the flagellar regulator flhCD was down-regulated during early infection and up-regulated during persistence (Fig. 4B). In addition, up-regulation of the gene encoding the adhesion protein invA, which is co-regulated with flagella [28], and its positive regulator rovA [29] suggested that other genes that are only expressed at 26°C in vitro could be up-regulated during persistence. Accordingly, a comparison of the expression patterns of in vivo and in vitro-derived samples showed that, during the early phase of infection, bacteria have an expression pattern similar to that seen in vitro at 37°C, whereas the expression pattern of persistent bacteria was much more similar to that of bacteria grown in vitro at 26°C (Fig. 5A and S4 Table). These results clearly indicate that, though increased temperature triggers T3SS and associated virulence genes during initial infection, other environmental cues are responsible for the observed transcriptional reprogramming of Y. pseudotuberculosis during prolonged infection.


Reprogramming of Yersinia from virulent to persistent mode revealed by complex in vivo RNA-seq analysis.

Avican K, Fahlgren A, Huss M, Heroven AK, Beckstette M, Dersch P, Fällman M - PLoS Pathog. (2015)

Y. pseudotuberculosis undergoes transcriptional reprogramming for adaption to persistence.(A) Comparison of genes up-regulated in Y. pseudotuberculosis in vitro at 26°C and 37°C compared to in vivo during early (2 dpi) and persistent (42 dpi) stages of infection. Similarities are shown with the number of genes up-regulated in both groups. (B) Functional annotation of Y. pseudotuberculosis genes up-regulated during early and persistent infection (KEGG pathway mapping tool). (C) Comparison of the in vivo gene expression profiles and the expression profiles of bacteria grown under anaerobic conditions in vitro. The analysis included genes up-regulated (>1.8-fold) during anaerobic or aerobic growth in both the exponential and stationary growth phase compared to genes up-regulated during early and persistent infection. Similarities are shown with the number of genes up-regulated in both groups.
© Copyright Policy
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC4295882&req=5

ppat.1004600.g005: Y. pseudotuberculosis undergoes transcriptional reprogramming for adaption to persistence.(A) Comparison of genes up-regulated in Y. pseudotuberculosis in vitro at 26°C and 37°C compared to in vivo during early (2 dpi) and persistent (42 dpi) stages of infection. Similarities are shown with the number of genes up-regulated in both groups. (B) Functional annotation of Y. pseudotuberculosis genes up-regulated during early and persistent infection (KEGG pathway mapping tool). (C) Comparison of the in vivo gene expression profiles and the expression profiles of bacteria grown under anaerobic conditions in vitro. The analysis included genes up-regulated (>1.8-fold) during anaerobic or aerobic growth in both the exponential and stationary growth phase compared to genes up-regulated during early and persistent infection. Similarities are shown with the number of genes up-regulated in both groups.
Mentions: A total of 1288 genes were found to be differentially expressed (log2 fold change ≥0.7) in vivo. RPKMO values detected by RNA-seq are shown in a histogram and heat map in Fig. 3A to highlight the differences in individual ORFs during early and persistent infection (see also S3 Table). Surprisingly, the T3SS components encoded on the virulence plasmid that were highly expressed during the early stage of infection were distinctly down-regulated during persistent infection. Another conspicuous finding was the up-regulation of flagella and chemotaxis genes. T3SS is known to be induced at 37°C, but flagella are down-regulated at this temperature; in vitro, flagella are expressed only at 26°C (confirmed by qPCR in the same samples used in RNA-seq; Fig. 4A–B). In analogy, the T3SS master regulator lcrF was up-regulated during early infection and down-regulated during persistence (S3 Table), and the flagellar regulator flhCD was down-regulated during early infection and up-regulated during persistence (Fig. 4B). In addition, up-regulation of the gene encoding the adhesion protein invA, which is co-regulated with flagella [28], and its positive regulator rovA [29] suggested that other genes that are only expressed at 26°C in vitro could be up-regulated during persistence. Accordingly, a comparison of the expression patterns of in vivo and in vitro-derived samples showed that, during the early phase of infection, bacteria have an expression pattern similar to that seen in vitro at 37°C, whereas the expression pattern of persistent bacteria was much more similar to that of bacteria grown in vitro at 26°C (Fig. 5A and S4 Table). These results clearly indicate that, though increased temperature triggers T3SS and associated virulence genes during initial infection, other environmental cues are responsible for the observed transcriptional reprogramming of Y. pseudotuberculosis during prolonged infection.

Bottom Line: We found that the Crp/CsrA/RovA regulatory cascades influence the pattern of bacterial gene expression during persistence.Furthermore, arcA, fnr, frdA, and wrbA play critical roles in persistence.Our findings suggest a model for the life cycle of this enteropathogen with reprogramming from a virulent to an adapted phenotype capable of persisting and spreading by fecal shedding.

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular Biology, Umeå Centre for Microbial Research (UCMR), Umeå University, Umeå, Sweden; Department of Molecular Biology, Laboratory for Molecular Infection Medicine Sweden (MIMS), Umea University, Umeå, Sweden.

ABSTRACT
We recently found that Yersinia pseudotuberculosis can be used as a model of persistent bacterial infections. We performed in vivo RNA-seq of bacteria in small cecal tissue biopsies at early and persistent stages of infection to determine strategies associated with persistence. Comprehensive analysis of mixed RNA populations from infected tissues revealed that Y. pseudotuberculosis undergoes transcriptional reprogramming with drastic down-regulation of T3SS virulence genes during persistence when the pathogen resides within the cecum. At the persistent stage, the expression pattern in many respects resembles the pattern seen in vitro at 26oC, with for example, up-regulation of flagellar genes and invA. These findings are expected to have impact on future rationales to identify suitable bacterial targets for new antibiotics. Other genes that are up-regulated during persistence are genes involved in anaerobiosis, chemotaxis, and protection against oxidative and acidic stress, which indicates the influence of different environmental cues. We found that the Crp/CsrA/RovA regulatory cascades influence the pattern of bacterial gene expression during persistence. Furthermore, arcA, fnr, frdA, and wrbA play critical roles in persistence. Our findings suggest a model for the life cycle of this enteropathogen with reprogramming from a virulent to an adapted phenotype capable of persisting and spreading by fecal shedding.

Show MeSH
Related in: MedlinePlus