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Co-evolution with lytic phage selects for the mucoid phenotype of Pseudomonas fluorescens SBW25.

ABSTRACT: The effects of co-evolution with lytic phage on bacterial virulence-related traits are largely unknown. In this study we investigate the incidence of the mucoid phenotype of the bacterium Pseudomonas fluorescens SBW25 in response to co-evolution with the lytic phage phi2 (?2). The mucoid phenotype of Pseudomonas spp. is due to overproduction of alginate and is a considerable virulence factor contributing to the intractability of infections most notably in cystic fibrosis (CF) lung, but also in pathogenic infections of plants. Our data show that this phenotype can evolve as an adaptive response to phage predation and is favoured under specific abiotic conditions, in particular a homogenous spatial structure and a high rate of nutrient replacement. The mucoid phenotype remains partially sensitive to phage infection, which facilitates 'apparent competition' with phage-sensitive competitors, partially offsetting the costs of alginate production. Although P. fluorescens SBW25 is not a pathogen, several key characteristics typical of Pseudomonas aeruginosa clinical isolates from CF lung were noted, including loss of motility on mucoid conversion and a high rate of spontaneous reversion to the wild-type phenotype. Although the genetic mechanisms of this phenotype remain unknown, they do not include mutations at many of the commonly reported loci implicated in mucoid conversion, including mucA and algU. These data not only further our understanding of the potential role phage have in the ecology and evolution of bacteria virulence in both natural and clinical settings, but also highlight the need to consider both biotic and abiotic variables if bacteriophages are to be used therapeutically.

SUBMITTER: Scanlan PD

PROVIDER: S-EPMC3358020 | biostudies-literature | 2012 Jun

REPOSITORIES: biostudies-literature

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Co-evolution with lytic phage selects for the mucoid phenotype of Pseudomonas fluorescens SBW25.

Scanlan Pauline Deirdre PD Buckling Angus A

The ISME journal 20111222 6

The effects of co-evolution with lytic phage on bacterial virulence-related traits are largely unknown. In this study we investigate the incidence of the mucoid phenotype of the bacterium Pseudomonas fluorescens SBW25 in response to co-evolution with the lytic phage phi2 (φ2). The mucoid phenotype of Pseudomonas spp. is due to overproduction of alginate and is a considerable virulence factor contributing to the intractability of infections most notably in cystic fibrosis (CF) lung, but also in p ...[more]

PMID: 22189495

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Project description:HutC is known as a transcriptional repressor specific for histidine utilization (hut) genes in Gram-negative bacteria, including Pseudomonas fluorescens SBW25. However, its precise mode of protein-DNA interactions hasn't been examined with purified HutC proteins. Here, we performed electrophoretic mobility shift assay (EMSA) and DNase I footprinting using His6-tagged HutC and biotin-labeled probe of the hut promoter (PhutU). Results revealed a complex pattern of HutC oligomerization, and the specific protein-DNA interaction is disrupted by urocanate, a histidine derivative, in a concentration-dependent manner. Next, we searched for putative HutC-binding sites in the SBW25 genome. This led to the identification of 143 candidate targets with a P value less than 10-4 HutC interaction with eight selected candidate sites was subsequently confirmed by EMSA analysis, including the type IV pilus assembly protein PilZ, phospholipase C (PlcC) for phosphatidylcholine hydrolyzation, and key regulators of cellular nitrogen metabolism (NtrBC and GlnE). Finally, an isogenic hutC deletion mutant was subjected to transcriptome sequencing (RNA-seq) analysis and phenotypic characterization. When bacteria were grown on succinate and histidine, hutC deletion caused upregulation of 794 genes and downregulation of 525 genes at a P value of <0.05 with a fold change cutoff of 2.0. The hutC mutant displayed an enhanced spreading motility and pyoverdine production in laboratory media, in addition to the previously reported growth defect on the surfaces of plants. Together, our data indicate that HutC plays global regulatory roles beyond histidine catabolism through low-affinity binding with operator sites located outside the hut locus.IMPORTANCE HutC in Pseudomonas is a representative member of the GntR/HutC family of transcriptional regulators, which possess a N-terminal winged helix-turn-helix (wHTH) DNA-binding domain and a C-terminal substrate-binding domain. HutC is generally known to repress expression of histidine utilization (hut) genes through binding to the PhutU promoter with urocanate (the first intermediate of the histidine degradation pathway) as the direct inducer. Here, we first describe the detailed molecular interactions between HutC and its PhutU target site in a plant growth-promoting bacterium, P. fluorescens SBW25, and further show that HutC possesses specific DNA-binding activities with many targets in the SBW25 genome. Subsequent RNA-seq analysis and phenotypic assays revealed an unexpected global regulatory role of HutC for successful bacterial colonization in planta.

Dataset Information

Co-evolution with lytic phage selects for the mucoid phenotype of Pseudomonas fluorescens SBW25.

Publications

Co-evolution with lytic phage selects for the mucoid phenotype of Pseudomonas fluorescens SBW25.

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