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Identification of a new iron-regulated virulence gene, ireA, in an extraintestinal pathogenic isolate of Escherichia coli.

ABSTRACT: Our laboratory is studying an extraintestinal pathogenic isolate of Escherichia coli (CP9) as a model pathogen. We have been using human urine, ascites, and blood ex vivo to identify genes with increased expression in these media relative to expression in Luria-Bertani (LB) broth. Such genes may represent new or unrecognized virulence traits. In this study, we report the identification of a new gene, ireA (iron-responsive element). This gene has an open reading frame of 2,049 nucleotides, and its peptide has a molecular mass of 75.3 kDa on sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Its expression is increased a mean of 3.6-fold in human urine, 16.2-fold in human ascites, and 6.6-fold in human blood relative to expression in LB medium, and it is Fe repressible. IreA also exhibits peptide similarities (48 to 56%) to previously identified proteins that function as siderophore receptors, suggesting that IreA is involved in iron acquisition. PCR-based analysis of ireA's phylogenetic distribution detected ireA in none (0%) of 14 fecal isolates that represented probable commensal strains, but in 13 (26%) of 50 random urine and blood clinical isolates (P = 0.05) and in 5 (100%) of 5 representatives of the J96-like, clonal group of which CP9 is a member (P < 0.001). In a mouse urinary tract infection model, the presence of ireA contributed significantly to CP9's ability to colonize the bladder (P < 0.02), evidence that IreA is a urovirulence factor. Taken together, these findings demonstrate that ireA encodes a new virulence factor, which is likely involved in Fe acquisition.

SUBMITTER: Russo TA

PROVIDER: S-EPMC98753 | biostudies-other | 2001 Oct

REPOSITORIES: biostudies-other

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Identification of a new iron-regulated virulence gene, ireA, in an extraintestinal pathogenic isolate of Escherichia coli.

Russo T A TA Carlino U B UB Johnson J R JR

Infection and immunity 20011001 10

Our laboratory is studying an extraintestinal pathogenic isolate of Escherichia coli (CP9) as a model pathogen. We have been using human urine, ascites, and blood ex vivo to identify genes with increased expression in these media relative to expression in Luria-Bertani (LB) broth. Such genes may represent new or unrecognized virulence traits. In this study, we report the identification of a new gene, ireA (iron-responsive element). This gene has an open reading frame of 2,049 nucleotides, and it ...[more]

PMID: 11553562

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Project description:Strains of extraintestinal pathogenic Escherichia coli (ExPEC) can invade and colonize extraintestinal sites and cause a wide range of infections. Genomic analysis of ExPEC has mainly focused on isolates of human and avian origins, with porcine ExPEC isolates yet to be sequenced. To better understand the genomic attributes underlying the pathogenicity of porcine ExPEC, we isolated two E. coli strains PCN033 and PCN061 from pigs, assessed their in vivo virulence, and completed and compared their genomes.Animal experiments demonstrated that strain PCN033, but not PCN061, was pathogenic in a pig model. The chromosome of PCN033 was 384 kb larger than that of PCN061. Among the PCN033-specific sequences, genes encoding adhesins, unique lipopolysaccharide, unique capsular polysaccharide, iron acquisition and transport systems, and metabolism were identified. Additionally, a large plasmid PCN033p3 harboring many typical ExPEC virulence factors was identified in PCN033. Based on the genetic variation between PCN033 and PCN061, corresponding phenotypic differences in flagellum-dependent swarming motility and metabolism were verified. Furthermore, the comparative genomic analyses showed that the PCN033 genome shared many similarities with genomic sequences of human ExPEC strains. Additionally, comparison of PCN033 genome with other nine characteristic E. coli genomes revealed 425 PCN033-special coding sequences. Genes of this subset included those encoding type I restriction-modification (R-M) system, type VI secretion system (T6SS) and membrane-associated proteins.The genetic and phenotypic differences between PCN033 and PCN061 could partially explain their differences in virulence, and also provide insight towards the molecular mechanisms of porcine ExPEC infections. Additionally, the similarities between the genomes of PCN033 and human ExPEC strains suggest that some connections between porcine and human ExPEC strains exist. The first completed genomic sequence for porcine ExPEC and the genomic differences identified by comparative analyses provide a baseline understanding of porcine ExPEC genetics and lay the foundation for their further study.

Project description:BACKGROUND:Classification of pathogenic Escherichia coli (E. coli) has traditionally relied on detecting specific virulence associated genes (VAGs) or combinations thereof. For E. coli isolated from faecal samples, the presence of specific genes associated with different intestinal pathogenic pathovars will determine their classification and further course of action. However, the E. coli genome is not a static entity, and hybrid strains are emerging that cross the pathovar definitions. Hybrid strains may show gene contents previously associated with several distinct pathovars making the correct diagnostic classification difficult. We extended the analysis of routinely submitted faecal isolates to include known virulence associated genes that are usually not examined in faecal isolates to detect the frequency of possible hybrid strains. METHODS:From September 2012 to February 2013, 168 faecal isolates of E. coli routinely submitted to the Norwegian Institute of Public Health (NIPH) from clinical microbiological laboratories throughout Norway were analysed for 33 VAGs using multiplex-PCR, including factors associated with extraintestinal pathogenic E. coli (ExPEC) strains. The strains were further typed by Multiple Locus Variable-Number Tandem-Repeat Analysis (MLVA), and the phylogenetic grouping was determined. One isolate from the study was selected for whole genome sequencing (WGS) with a combination of Oxford Nanopore's MinION and Illumina's MiSeq. RESULTS:The analysis showed a surprisingly high number of strains carrying ExPEC associated VAGs and strains carrying a combination of both intestinal pathogenic E. coli (IPEC) and ExPEC VAGs. In particular, 93.5% (101/108) of isolates classified as belonging to an IPEC pathovar additionally carried ExPEC VAGs. WGS analysis of a selected hybrid strain revealed that it could, with present classification criteria, be classified as belonging to all of the Enteropathogenic Escherichia coli (EPEC), Uropathogenic Escherichia coli (UPEC), Neonatal meningitis Escherichia coli (NMEC) and Avian pathogenic Escherichia coli (APEC) pathovars. CONCLUSION:Hybrid ExPEC/IPEC E. coli strains were found at a very high frequency in faecal samples and were in fact the predominant species present. A sequenced hybrid isolate was confirmed to be a cross-pathovar strain possessing recognised hallmarks of several pathovars, and a genome heavily influenced by horizontal gene transfer.

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Dataset Information

Identification of a new iron-regulated virulence gene, ireA, in an extraintestinal pathogenic isolate of Escherichia coli.

Publications

Identification of a new iron-regulated virulence gene, ireA, in an extraintestinal pathogenic isolate of Escherichia coli.

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