Dataset Information

Identification of commensal Escherichia coli genes involved in biofilm resistance to pathogen colonization.

ABSTRACT: Protection provided by host bacterial microbiota against microbial pathogens is a well known but ill-understood property referred to as the barrier effect, or colonization resistance. Despite recent genome-wide analyses of host microbiota and increasing therapeutic interest, molecular analysis of colonization resistance is hampered by the complexity of direct in vivo experiments. Here we developed an in vitro-to-in vivo approach to identification of genes involved in resistance of commensal bacteria to exogenous pathogens. We analyzed genetic responses induced in commensal Escherichia coli upon entry of a diarrheagenic enteroaggregative E. coli or an unrelated Klebsiella pneumoniae pathogen into a biofilm community. We showed that pathogens trigger specific responses in commensal bacteria and we identified genes involved in limiting colonization of incoming pathogens within commensal biofilm. We tested the in vivo relevance of our findings by comparing the extent of intestinal colonization by enteroaggregative E. coli and K. pneumoniae pathogens in mice pre-colonized with E. coli wild type commensal strain, or mutants corresponding to identified colonization resistance genes. We demonstrated that the absence of yiaF and bssS (yceP) differentially alters pathogen colonization in the mouse gut. This study therefore identifies previously uncharacterized colonization resistance genes and provides new approaches to unravelling molecular aspects of commensal/pathogen competitive interactions.

SUBMITTER: Da Re S

PROVIDER: S-EPMC3646849 | biostudies-literature | 2013

REPOSITORIES: biostudies-literature

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Identification of commensal Escherichia coli genes involved in biofilm resistance to pathogen colonization.

Da Re Sandra S Valle Jaione J Charbonnel Nicolas N Beloin Christophe C Latour-Lambert Patricia P Faure Philippe P Turlin Evelyne E Le Bouguénec Chantal C Renauld-Mongénie Geneviève G Forestier Christiane C Ghigo Jean-Marc JM

PloS one 20130507 5

Protection provided by host bacterial microbiota against microbial pathogens is a well known but ill-understood property referred to as the barrier effect, or colonization resistance. Despite recent genome-wide analyses of host microbiota and increasing therapeutic interest, molecular analysis of colonization resistance is hampered by the complexity of direct in vivo experiments. Here we developed an in vitro-to-in vivo approach to identification of genes involved in resistance of commensal bact ...[more]

PMID: 23667443

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Project description:Leafy green vegetables, such as lettuce, have been increasingly implicated in outbreaks of foodborne illnesses due to contamination by Escherichia coli O157:H7. While E. coli can survive in soils, colonize plants, and survive on produce, very little is known about the interaction of E. coli with the roots of growing lettuce plants. In these studies a combination of microarray analyses and microbial genetics were used to gain a comprehensive understanding of bacterial genes involved in the colonization and growth of E. coli K12 on lettuce roots using a hydroponic assay system. Here we report that after three days of interaction with lettuce roots, 193 and 131 genes were significantly up-regulated and down-regulated at least 1.5 fold, respectively. Forty-five out of the 193 up-regulated genes (23%) were involved in protein synthesis and were highly induced. Genes involved in stress response, attachment and biofilm formation were up-regulated in E. coli when they interacted with lettuce roots under conditions of hydroponic growth. In particular crl, a gene regulating the cryptic csgA gene for curli production, was significantly up regulated. The crl, csgA and fliN mutants had a reduced capacity to attach to roots as determined by bacterial counts and by confocal laser scanning microscopy. Our microarray data showed that E. coli K12 increased the synthesis of proteins indicated that a dramatic change was induced in the physiology of the microorganism. This study indicates that E. coli K12 can efficiently colonize lettuce roots by using attachment and biofilm modulation genes and can readily adapt to the rhizosphere of lettuce plants. Further studies are needed to better characterize this interaction in pathogenic strains of this species. Escherichia coli MG1655 strains were grown in the lettuce rhizosphere for three days. Transcriptional profiling of E. coli was compared between cells grown with and without rhizosphere . Three biological replicates of each treatment were prepared, and six microarray slides were used.

Project description:Honey has been widely used against bacterial infection for centuries. Previous studies suggested that honeys in high concentrations inhibited bacterial growth due to the presence of anti-microbial compounds, such as methylglyoxal, hydrogen peroxide, and peptides. In this study, we found that three honeys (acacia, clover, and polyfloral) in a low concentration as below as 0.5% (v/v) significantly suppress virulence and biofilm formation in enterohemorrhagic E. coli O157:H7 affecting the growth of planktonic cells while these honeys do not harm commensal E. coli K-12 biofilm formation. Transcriptome analyses show that honeys (0.5%) markedly repress quorum sensing genes (e.g., AI-2 import and indole biosynthesis), virulence genes (e.g., LEE genes), and curli genes (csgBAC). We found that glucose and fructose in honeys are key compounds to reduce the biofilm formation of E. coli O157:H7 via suppressing curli production, but not that of E. coli K-12. Additionally, we observed the temperature-dependent response of honeys and glucose on commensal E. coli K-12 biofilm formation; honey and glucose increase E. coli K-12 biofilm formation at 37Â°C, while they decrease E. coli K-12 biofilm formation at 26Â°C. These results suggest that honey can be a practical tool for reducing virulence and colonization of the pathogenic E. coli O157:H7, while honeys do not harm commensal E. coli community in the human. For the microarray experiments, E. coli O157:H7 EDL933 was inoculated in 250 ml of LB in 1000 ml flasks with overnight cultures that were diluted at 1:100. Cells were shaken with 10g of glass wool at 100 rpm and 37Â°C for 7 hrs. Cells were immediately chilled with dry ice and 95% ethanol (to prevent RNA degradation) for 30 sec before centrifugation in 50 ml centrifuge tubes at 13,000 g for 2 min; cell pellets were frozen immediately with dry ice and stored -80Â°C. RNA was isolated using Qiagen RNeasy mini Kit (Valencia, CA, USA). To eliminate DNA contamination, Qiagen RNase-free DNase I was used to digest DNA. RNA quality was assessed by Agilent 2100 bioanalyser using the RNA 6000 Nano Chip (Agilent Technologies, Amstelveen, The Netherlands), and quantity was determined by ND-1000 Spectrophotometer (NanoDrop Technologies, Inc., DE, USA).

Dataset Information

Identification of commensal Escherichia coli genes involved in biofilm resistance to pathogen colonization.

Publications

Identification of commensal Escherichia coli genes involved in biofilm resistance to pathogen colonization.

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