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Fis is essential for capsule production in Pasteurella multocida and regulates expression of other important virulence factors.

ABSTRACT: P. multocida is the causative agent of a wide range of diseases of animals, including fowl cholera in poultry and wild birds. Fowl cholera isolates of P. multocida generally express a capsular polysaccharide composed of hyaluronic acid. There have been reports of spontaneous capsule loss in P. multocida, but the mechanism by which this occurs has not been determined. In this study, we identified three independent strains that had spontaneously lost the ability to produce capsular polysaccharide. Quantitative RT-PCR showed that these strains had significantly reduced transcription of the capsule biosynthetic genes, but DNA sequence analysis identified no mutations within the capsule biosynthetic locus. However, whole-genome sequencing of paired capsulated and acapsular strains identified a single point mutation within the fis gene in the acapsular strain. Sequencing of fis from two independently derived spontaneous acapsular strains showed that each contained a mutation within fis. Complementation of these strains with an intact copy of fis, predicted to encode a transcriptional regulator, returned capsule expression to all strains. Therefore, expression of a functional Fis protein is essential for capsule expression in P. multocida. DNA microarray analysis of one of the spontaneous fis mutants identified approximately 30 genes as down-regulated in the mutant, including pfhB_2, which encodes a filamentous hemagglutinin, a known P. multocida virulence factor, and plpE, which encodes the cross protective surface antigen PlpE. Therefore these experiments define for the first time a mechanism for spontaneous capsule loss in P. multocida and identify Fis as a critical regulator of capsule expression. Furthermore, Fis is involved in the regulation of a range of other P. multocida genes including important virulence factors.

SUBMITTER: Steen JA

PROVIDER: S-EPMC2816674 | biostudies-literature | 2010 Feb

REPOSITORIES: biostudies-literature

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Fis is essential for capsule production in Pasteurella multocida and regulates expression of other important virulence factors.

Steen Jason A JA Steen Jennifer A JA Harrison Paul P Seemann Torsten T Wilkie Ian I Harper Marina M Adler Ben B Boyce John D JD

PLoS pathogens 20100205 2

P. multocida is the causative agent of a wide range of diseases of animals, including fowl cholera in poultry and wild birds. Fowl cholera isolates of P. multocida generally express a capsular polysaccharide composed of hyaluronic acid. There have been reports of spontaneous capsule loss in P. multocida, but the mechanism by which this occurs has not been determined. In this study, we identified three independent strains that had spontaneously lost the ability to produce capsular polysaccharide. ...[more]

PMID: 20140235

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Project description:Pasteurella multocida is a Gram-negative capsulated bacterium responsible for a range of diseases that cause severe morbidity and mortality in livestock animals. The hyaluronic acid (HA) capsule produced by P. multocida serogroup A strains is a critical virulence factor. In this study, we utilized transposon-directed insertion site sequencing (TraDIS) to identify genes essential for in vitro growth of P. multocida and combined TraDIS with discontinuous density gradients (TraDISort) to identify genes required for HA capsule production and regulation in this pathogen. Analysis of mutants with a high cell density phenotype, indicative of the loss of extracellular capsule, led to the identification of 69 genes important for capsule production. These genes included all previously characterized genes in the capsule biosynthesis locus and fis and hfq, which encode known positive regulators of P. multocida capsule. Many of the other capsule-associated genes identified in this study were involved in regulation or activation of the stringent response, including spoT and relA, which encode proteins that regulate the concentration of guanosine alarmones. Disruption of the autoregulatory domains in the C-terminal half of SpoT using insertional mutagenesis resulted in reduced expression of capsule biosynthesis genes and an acapsular phenotype. Overall, these findings have greatly increased the understanding of hyaluronic acid capsule production and regulation in P. multocida. IMPORTANCE The bacterial pathogen P. multocida can cause serious disease in production animals, including fowl cholera in poultry, hemorrhagic septicemia in cattle and buffalo, atrophic rhinitis in pigs, and respiratory diseases in a range of livestock. P. multocida produces a capsule that is essential for systemic disease, but the complete mechanisms underlying synthesis and regulation of capsule production are not fully elucidated. A whole-genome analysis using TraDIS was undertaken to identify genes essential for growth in rich media and to obtain a comprehensive characterization of capsule production. Many of the capsule-associated genes identified in this study were involved in the stringent response to stress, a novel finding for this important animal pathogen.

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Fis is essential for capsule production in Pasteurella multocida and regulates expression of other important virulence factors.

Publications

Fis is essential for capsule production in Pasteurella multocida and regulates expression of other important virulence factors.

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