Genome Based Phylogeny and Comparative Genomic Analysis of Intra-Mammary Pathogenic Escherichia coli

Escherichia coli is an important cause of bovine mastitis and can cause both severe inflammation with a short-term transient infection, as well as less severe, but more chronic inflammation and infection persistence. E. coli is a highly diverse organism that has been classified into a number of different pathotypes or pathovars, and mammary pathogenic E. coli (MPEC) has been proposed as a new such pathotype. The purpose of this study was to use genome sequence data derived from both transient and persistent MPEC isolates (two isolates of each phenotype) to construct a genome-based phylogeny that places MPEC in its phylogenetic context with other E. coli pathovars. A subsidiary goal was to conduct comparative genomic analyses of these MPEC isolates with other E. coli pathovars to provide a preliminary perspective on loci that might be correlated with the MPEC phenotype. Both concatenated and consensus tree phylogenies did not support MPEC monophyly or the monophyly of either transient or persistent phenotypes. Three of the MPEC isolates (ECA-727, ECC-Z, and ECA-O157) originated from within the predominately commensal clade of E. coli, referred to as phylogroup A. The fourth MPEC isolate, of the persistent phenotype (ECC-1470), was sister group to an isolate of ETEC, falling within the E. coli B1 clade. This suggests that the MPEC phenotype has arisen on numerous independent occasions and that this has often, although not invariably, occurred from commensal ancestry. Examination of the genes present in the MPEC strains relative to the commensal strains identified a consistent presence of the type VI secretion system (T6SS) in the MPEC strains, with only occasional representation in commensal strains, suggesting that T6SS may be associated with MPEC pathogenesis and/or as an inter-bacterial competitive attribute and therefore could represent a useful target to explore for the development of MPEC specific inhibitors.     

Genomic Comparative Study of Bovine Mastitis Escherichia coli

Escherichia coli, one of the main causative agents of bovine mastitis, is responsible for significant losses on dairy farms. In order to better understand the pathogenicity of E. coli mastitis, an accurate characterization of E. coli strains isolated from mastitis cases is required. By using phylogenetic analyses and whole genome comparison of 5 currently available mastitis E. coli genome sequences, we searched for genotypic traits specific for mastitis isolates. Our data confirm that there is a bias in the distribution of mastitis isolates in the different phylogenetic groups of the E. coli species, with the majority of strains belonging to phylogenetic groups A and B1. An interesting feature is that clustering of strains based on their accessory genome is very similar to that obtained using the core genome. This finding illustrates the fact that phenotypic properties of strains from different phylogroups are likely to be different. As a consequence, it is possible that different strategies could be used by mastitis isolates of different phylogroups to trigger mastitis. Our results indicate that mastitis E. coli isolates analyzed in this study carry very few of the virulence genes described in other pathogenic E. coli strains. A more detailed analysis of the presence/absence of genes involved in LPS synthesis, iron acquisition and type 6 secretion systems did not uncover specific properties of mastitis isolates. Altogether, these results indicate that mastitis E. coli isolates are rather characterized by a lack of bona fide currently described virulence genes.     

肠外致病性大肠杆菌致病机制及公共卫生学意义

致病性大肠杆菌包括肠致病性大肠杆菌(intestinal pathogenic Escherichia coli, IPEC)和肠外致病性大肠杆菌(extraintestinalpathogenicE.coli,ExPEC),可引起人和动物多种感染性疾病。ExPEC主要在肠道外其他组织脏器定殖并导致感染,包括尿道致病性大肠杆菌(uropathogenicE.coli, UPEC)、新生儿脑膜炎大肠杆菌(newborn meningitis E. coli, NMEC)和禽致病性大肠杆菌(avian pathogenic E. coli, APEC)。人源ExPEC (UPEC和NMEC)主要引起人尿道感染、肾盂肾炎和新生儿脑膜炎,而APEC可导致禽类的大肠杆菌病,造成家禽业的巨大经济损失。另外,乳腺致病性大肠杆菌(mammary pathogenic E. coli, MPEC)和猪源ExPEC可导致奶牛乳房炎、猪的肺炎及急性败血症等病症。研究发现,ExPEC类菌株在基因组结构上很相似,与IPEC本质区别在于致病机制不同,ExPEC具有很多相同的毒力基因和耐药基因,而且动物源ExPEC...     

Lactic Acid Bacteria Isolated from Bovine Mammary Microbiota: Potential Allies against Bovine Mastitis

Bovine mastitis is a costly disease in dairy cattle worldwide. As of yet, the control of bovine mastitis is mostly based on prevention by thorough hygienic procedures during milking. Additional strategies include vaccination and utilization of antibiotics. Despite these measures, mastitis is not fully under control, thus prompting the need for alternative strategies. The goal of this study was to isolate autochthonous lactic acid bacteria (LAB) from bovine mammary microbiota that exhibit beneficial properties that could be used for mastitis prevention and/or treatment. Sampling of the teat canal led to the isolation of 165 isolates, among which a selection of ten non-redundant LAB strains belonging to the genera Lactobacillus and Lactococcus were further characterized with regard to several properties: surface properties (hydrophobicity, autoaggregation); inhibition potential of three main mastitis pathogens, Staphylococcus aureus, Escherichia coli and Streptococcus uberis; colonization capacities of bovine mammary epithelial cells (bMEC); and immunomodulation properties. Three strains, Lactobacillus brevis 1595 and 1597 and Lactobacillus plantarum 1610, showed high colonization capacities and a medium surface hydrophobicity. These strains are good candidates to compete with pathogens for mammary gland colonization. Moreover, nine strains exhibited anti-inflammatory properties, as illustrated by the lower IL-8 secretion by E. coli-stimulated bMEC in the presence of these LAB. Full genome sequencing of five candidate strains allowed to check for undesirable genetic elements such as antibiotic resistance genes and to identify potential bacterial determinants involved in the beneficial properties. This large screening of beneficial properties while checking for undesirable genetic markers allowed the selection of promising candidate LAB strains from bovine mammary microbiota for the prevention and/or treatment of bovine mastitis.     

Free-Ranging Frigates (Fregata magnificens) of the Southeast Coast of Brazil Harbor Extraintestinal Pathogenic Escherichia coli Resistant to Antimicrobials

Seabirds may be responsible for the spread of pathogenic/resistant organisms over great distances, playing a relevant role within the context of the One World, One Health concept. Diarrheagenic E. coli strains, known as STEC (shiga toxin-producing E. coli), and the extraintestinal pathogenic E. coli (ExPEC and the subpathotype APEC), are among the E. coli pathotypes with zoonotic potential associated with the birds. In order to identify health threats carried by frigates and to evaluate the anthropic influence on the southern coast of Brazil, the aim of this work was to characterize E. coli isolated from free-ranging frigates in relation to virulence genotypes, serotypes, phylogenetic groups and antimicrobial resistance. Cloacal and choanal swabs were sampled from 38 Fregata magnificens from two oceanic islands and one rescue center. Forty-three E. coli strains were recovered from 33 out of the 38 birds (86.8%); 88.4% of strains showed some of the virulence genes (VGs) searched, 48.8% contained three or more VGs. None of the strains presented VGs related to EPEC/STEC. Some of the isolates showed virulence genotypes, phylogenetic groups and serotypes of classical human ExPEC or APEC (O2:H7, O1:H6, ONT:H7, O25:H4). Regarding antimicrobial susceptibility, 62.8% showed resistance, and 11.6% (5/43) were multidrug-resistant. The E. coli present in the intestines of the frigates may reflect the environmental human impact on southeast coast of Brazil; they may also represent an unexplored threat for seabird species, especially considering the overlap of pathogenic potential and antimicrobial resistance present in these strains.     

Genome sequence analyses of two isolates from the recent <Emphasis Type="Italic">Escherichia coli</Emphasis> outbreak in Germany reveal the emergence of a new pathotype: Entero-Aggregative-Haemorrhagic <Emphasis Type="Italic">Escherichia coli</Emphasis> (EAHEC)

The genome sequences of two Escherichia coli O104:H4 strains derived from two different patients of the 2011 German E. coli outbreak were determined. The two analyzed strains were designated E. coli GOS1 and GOS2 (German outbreak strain). Both isolates comprise one chromosome of approximately 5.31 Mbp and two putative plasmids. Comparisons of the 5,217 (GOS1) and 5,224 (GOS2) predicted protein-encoding genes with various E. coli strains, and a multilocus sequence typing analysis revealed that the isolates were most similar to the entero-aggregative E. coli (EAEC) strain 55989. In addition, one of the putative plasmids of the outbreak strain is similar to pAA-type plasmids of EAEC strains, which contain aggregative adhesion fimbrial operons. The second putative plasmid harbors genes for extended-spectrum β-lactamases. This type of plasmid is widely distributed in pathogenic E. coli strains. A significant difference of the E. coli GOS1 and GOS2 genomes to those of EAEC strains is the presence of a prophage encoding the Shiga toxin, which is characteristic for enterohemorrhagic E. coli (EHEC) strains. The unique combination of genomic features of the German outbreak strain, containing characteristics from pathotypes EAEC and EHEC, suggested that it represents a new pathotype Entero-Aggregative-Haemorrhagic E scherichia c oli (EAHEC).     

Comparative Genomics and Characterization of Hybrid Shigatoxigenic and Enterotoxigenic Escherichia coli (STEC/ETEC) Strains

Background

Shigatoxigenic Escherichia coli (STEC) and enterotoxigenic E. coli (ETEC) cause serious foodborne infections in humans. These two pathogroups are defined based on the pathogroup-associated virulence genes: stx encoding Shiga toxin (Stx) for STEC and elt encoding heat-labile and/or est encoding heat-stable enterotoxin (ST) for ETEC. The study investigated the genomics of STEC/ETEC hybrid strains to determine their phylogenetic position among E. coli and to define the virulence genes they harbor.

Methods

The whole genomes of three STEC/ETEC strains possessing both stx and est genes were sequenced using PacBio RS sequencer. Two of the strains were isolated from the patients, one with hemolytic uremic syndrome, and one with diarrhea. The third strain was of bovine origin. Core genome analysis of the shared chromosomal genes and comparison with E. coli and Shigella spp. reference genomes was performed to determine the phylogenetic position of the STEC/ETEC strains. In addition, a set of virulence genes and ETEC colonization factors were extracted from the genomes. The production of Stx and ST were studied.

Results

The human STEC/ETEC strains clustered with strains representing ETEC, STEC, enteroaggregative E. coli, and commensal and laboratory-adapted E. coli. However, the bovine STEC/ETEC strain formed a remote cluster with two STECs of bovine origin. All three STEC/ETEC strains harbored several other virulence genes, apart from stx and est, and lacked ETEC colonization factors. Two STEC/ETEC strains produced both toxins and one strain Stx only.

Conclusions

This study shows that pathogroup-associated virulence genes of different E. coli can co-exist in strains originating from different phylogenetic lineages. The possibility of virulence genes to be associated with several E. coli pathogroups should be taken into account in strain typing and in epidemiological surveillance. Development of novel hybrid E. coli strains may cause a new public health risk, which challenges the traditional diagnostics of E. coli infections.     

Identification of Unconventional Intestinal Pathogenic Escherichia coli Isolates Expressing Intermediate Virulence Factor Profiles by Using a Novel Single-Step Multiplex PCR

Intestinal pathogenic Escherichia coli represents a global health problem for mammals, including humans. At present, diarrheagenic E. coli bacteria are grouped into seven major pathotypes that differ in their virulence factor profiles, severity of clinical manifestations, and prognosis. In this study, we developed and evaluated a one-step multiplex PCR (MPCR) for the straightforward differential identification of intestinal pathotypes of E. coli. The specificity of this novel MPCR was validated by using a subset of reference strains and further confirmed by PCR-independent pheno- and genotypic characterization. Moreover, we tested 246 clinical E. coli isolates derived from diarrhea patients from several distinct geographic regions. Interestingly, besides strains belonging to the defined and well-described pathotypes, we identified five unconventional strains expressing intermediate virulence factor profiles. These strains have been further characterized and appear to represent intermediate strains carrying genes and expressing factors associated with enteropathogenic E. coli, Shiga toxin-producing E. coli, enterotoxigenic E. coli, and enteroaggregative E. coli alike. These strains represent further examples of the extraordinary plasticity of the E. coli genome. Moreover, this implies that the important identification of specific pathotypes has to be based on a broad matrix of indicator genes. In addition, the presence of intermediate strains needs to be accounted for.     

Persistent Colonization of Sheep by Escherichia coli O157:H7 and Other E. coli Pathotypes

Shiga toxin-producing Escherichia coli (STEC) is an important cause of food-borne illness in humans. Ruminants appear to be more frequently colonized by STEC than are other animals, but the reason(s) for this is unknown. We compared the frequency, magnitude, duration, and transmissibility of colonization of sheep by E. coli O157:H7 to that by other pathotypes of E. coli. Young adult sheep were simultaneously inoculated with a cocktail consisting of two strains of E. coli O157:H7, two strains of enterotoxigenic E. coli (ETEC), and one strain of enteropathogenic E. coli. Both STEC strains and ETEC 2041 were given at either 10⁷ or 10¹⁰ CFU/strain/animal. The other strains were given only at 10¹⁰ CFU/strain. We found no consistent differences among pathotypes in the frequency, magnitude, and transmissibility of colonization. However, the STEC strains tended to persist to 2 weeks and 2 months postinoculation more frequently than did the other pathotypes. The tendency for persistence of the STEC strains was apparent following an inoculation dose of either 10⁷ or 10¹⁰ CFU. One of the ETEC strains also persisted when inoculated at 10¹⁰ CFU. However, in contrast to the STEC strains, it did not persist when inoculated at 10⁷ CFU. These results support the hypothesis that STEC is better adapted to persist in the alimentary tracts of sheep than are other pathotypes of E. coli.     

Complete Genome Sequence and Comparative Analysis of the Wild-type Commensal Escherichia coli Strain SE11 Isolated from a Healthy Adult

We sequenced and analyzed the genome of a commensal Escherichia coli (E. coli) strain SE11 (O152:H28) recently isolated from feces of a healthy adult and classified into E. coli phylogenetic group B1. SE11 harbored a 4.8 Mb chromosome encoding 4679 protein-coding genes and six plasmids encoding 323 protein-coding genes. None of the SE11 genes had sequence similarity to known genes encoding phage- and plasmid-borne virulence factors found in pathogenic E. coli strains. The comparative genome analysis with the laboratory strain K-12 MG1655 identified 62 poorly conserved genes between these two non-pathogenic strains and 1186 genes absent in MG1655. These genes in SE11 were mostly encoded in large insertion regions on the chromosome or in the plasmids, and were notably abundant in genes of fimbriae and autotransporters, which are cell surface appendages that largely contribute to the adherence ability of bacteria to host cells and bacterial conjugation. These data suggest that SE11 may have evolved to acquire and accumulate the functions advantageous for stable colonization of intestinal cells, and that the adhesion-associated functions are important for the commensality of E. coli in human gut habitat.Key words: Escherichia coli, commensal, human gut, genome sequencing     

Genomic Comparison of Translocating and Non-Translocating Escherichia coli

Translocation of E. coli across the gut epithelium can result in fatal sepsis in post-surgical patients. In vitro and in vivo experiments have identified the existence of a novel pathotype of translocating E. coli (TEC) that employs an unknown mechanism for translocating across epithelial cells to the mesenteric lymph nodes and the blood stream in both humans and animal models. In this study the genomes of four TEC strains isolated from the mesenteric lymph nodes of a fatal case of hospitalised patient (HMLN-1), blood of pigs after experimental shock (PC-1) and after non-lethal haemorrhage in rats (KIC-1 and KIC-2) were sequenced in order to identify the genes associated with their adhesion and/or translocation. To facilitate the comparison, the genomes of a non-adhering, non-translocating E. coli (46–4) and adhering but non-translocating E. coli (73–89) were also sequenced and compared. Whole genome comparison revealed that three (HMLN-1, PC-1 and KIC-2) of the four TEC strains carried a genomic island that encodes a Type 6 Secretion System that may contribute to adhesion of the bacteria to gut epithelial cells. The human TEC strain HMLN-1 also carried the invasion ibeA gene, which was absent in the animal TEC strains and is likely to be associated with host-specific translocation. Phylogenetic analysis revealed that the four TEC strains were distributed amongst three distinct E. coli phylogroups, which was supported by the presence of phylogroup specific fimbriae gene clusters. The genomic comparison has identified potential genes that can be targeted with knock-out experiments to further characterise the mechanisms of E. coli translocation.     

SslE Elicits Functional Antibodies That Impair In Vitro Mucinase Activity and In Vivo Colonization by Both Intestinal and Extraintestinal Escherichia coli Strains

SslE, the Secreted and surface-associated lipoprotein from Escherichia coli, has recently been associated to the M60-like extracellular zinc-metalloprotease sub-family which is implicated in glycan recognition and processing. SslE can be divided into two main variants and we recently proposed it as a potential vaccine candidate. By applying a number of in vitro bioassays and comparing wild type, knockout mutant and complemented strains, we have now demonstrated that SslE specifically contributes to degradation of mucin substrates, typically present in the intestine and bladder. Mutation of the zinc metallopeptidase motif of SslE dramatically impaired E. coli mucinase activity, confirming the specificity of the phenotype observed. Moreover, antibodies raised against variant I SslE, cloned from strain IHE3034 (SslE_IHE3034), are able to inhibit translocation of E. coli strains expressing different variants through a mucin-based matrix, suggesting that SslE induces cross-reactive functional antibodies that affect the metallopeptidase activity. To test this hypothesis, we used well-established animal models and demonstrated that immunization with SslE_IHE3034 significantly reduced gut, kidney and spleen colonization by strains producing variant II SslE and belonging to different pathotypes. Taken together, these data strongly support the importance of SslE in E. coli colonization of mucosal surfaces and reinforce the use of this antigen as a component of a broadly protective vaccine against pathogenic E. coli species.     

Predicting Essential Metabolic Genome Content of Niche-Specific Enterobacterial Human Pathogens during Simulation of Host Environments

Microorganisms have evolved to occupy certain environmental niches, and the metabolic genes essential for growth in these locations are retained in the genomes. Many microorganisms inhabit niches located in the human body, sometimes causing disease, and may retain genes essential for growth in locations such as the bloodstream and urinary tract, or growth during intracellular invasion of the hosts’ macrophage cells. Strains of Escherichia coli (E. coli) and Salmonella spp. are thought to have evolved over 100 million years from a common ancestor, and now cause disease in specific niches within humans. Here we have used a genome scale metabolic model representing the pangenome of E. coli which contains all metabolic reactions encoded by genes from 16 E. coli genomes, and have simulated environmental conditions found in the human bloodstream, urinary tract, and macrophage to determine essential metabolic genes needed for growth in each location. We compared the predicted essential genes for three E. coli strains and one Salmonella strain that cause disease in each host environment, and determined that essential gene retention could be accurately predicted using this approach. This project demonstrated that simulating human body environments such as the bloodstream can successfully lead to accurate computational predictions of essential/important genes.     

Small Heat-Shock Proteins,IbpAB, Protect Non-Pathogenic Escherichia coli from Killing by Macrophage-Derived Reactive Oxygen Species

Many intracellular bacterial pathogens possess virulence factors that prevent detection and killing by macrophages. However, similar virulence factors in non-pathogenic bacteria are less well-characterized and may contribute to the pathogenesis of chronic inflammatory conditions such as Crohn’s disease. We hypothesize that the small heat shock proteins IbpAB, which have previously been shown to reduce oxidative damage to proteins in vitro and be upregulated in luminal non-pathogenic Escherichia strain NC101 during experimental colitis in vivo, protect commensal E. coli from killing by macrophage-derived reactive oxygen species (ROS). Using real-time PCR, we measured ibpAB expression in commensal E. coli NC101 within wild-type (wt) and ROS-deficient (gp91phox^-/-) macrophages and in NC101 treated with the ROS generator paraquat. We also quantified survival of NC101 and isogenic mutants in wt and gp91phox^-/- macrophages using gentamicin protection assays. Similar assays were performed using a pathogenic E. coli strain O157:H7. We show that non-pathogenic E. coli NC101inside macrophages upregulate ibpAB within 2 hrs of phagocytosis in a ROS-dependent manner and that ibpAB protect E. coli from killing by macrophage-derived ROS. Moreover, we demonstrate that ROS-induced ibpAB expression is mediated by the small E. coli regulatory RNA, oxyS. IbpAB are not upregulated in pathogenic E. coli O157:H7 and do not affect its survival within macrophages. Together, these findings indicate that ibpAB may be novel virulence factors for certain non-pathogenic E. coli strains.     

Aryl hydrocarbon receptor activation by Lactobacillus reuteri tryptophan metabolism alleviates Escherichia coli-induced mastitis in mice

The intestinal microbiota has been associated with the occurrence and development of mastitis, which is one of the most serious diseases of lactating women and female animals, but the underlying mechanism has not yet been elucidated. Aryl hydrocarbon receptor (AhR) activation by microbiota tryptophan metabolism-derived ligands is involved in maintaining host homeostasis and resisting diseases. We investigated whether AhR activation by microbiota-metabolic ligands could influence mastitis development in mice. In this study, we found that AhR activation using Ficz ameliorated mastitis symptoms, which were related to limiting NF-κB activation and enhancing barrier function. Impaired AhR activation by disturbing the intestinal microbiota initiated mastitis, and processed Escherichia coli (E. coli)-induced mastitis in mice. Supplementation with dietary tryptophan attenuated the mastitis, but attenuation was inhibited by the intestinal microbiota abrogation, while administering tryptophan metabolites including IAld and indole but not IPA, rescued the tryptophan effects in dysbiotic mice. Supplementation with a Lactobacillus reuteri (L. reuteri) strain with the capacity to produce AhR ligands also improved E. coli-induced mastitis in an AhR-dependent manner. These findings provide evidence for novel therapeutic strategies for treating mastitis, and support the role of metabolites derived from the intestinal microbiota in improving distal disease.     

Comparative Genomic Analysis of Two Novel Sporadic Shiga Toxin-Producing Escherichia coli O104:H4 Strains Isolated 2011 in Germany

A large outbreak of gastrointestinal disease occurred in 2011 in Germany which resulted in almost 4000 patients with acute gastroenteritis or hemorrhagic colitis, 855 cases of a hemolytic uremic syndrome and 53 deaths. The pathogen was an uncommon, multiresistant Escherichia coli strain of serotype O104:H4 which expressed a Shiga toxin characteristic of enterohemorrhagic E. coli and in addition virulence factors common to enteroaggregative E. coli. During post-epidemic surveillance of Shiga toxin-producing E. coli (STEC) all but two of O104:H4 isolates were indistinguishable from the epidemic strain. Here we describe two novel STEC O104:H4 strains isolated in close spatiotemporal proximity to the outbreak which show a virulence gene panel, a Shiga toxin-mediated cytotoxicity towards Vero cells and aggregative adherence to Hep-2 cells comparable to the outbreak strain. They differ however both from the epidemic strain and from each other, by their antibiotic resistance phenotypes and some other features as determined by routine epidemiological subtyping methods. Whole genome sequencing of these two strains, of ten outbreak strain isolates originating from different time points of the outbreak and of one historical sporadic EHEC O104:H4 isolate was performed. Sequence analysis revealed a clear phylogenetic distance between the two variant strains and the outbreak strain finally identifying them as epidemiologically unrelated isolates from sporadic cases. These findings add to the knowledge about this emerging pathogen, illustrating a certain diversity within the bacterial core genome as well as loss and gain of accessory elements. Our results do also support the view that distinct new variants of STEC O104:H4 repeatedly might originate from yet unknown reservoirs, rather than that there would be a continuous diversification of a single epidemic strain established and circulating in Germany after the large outbreak in 2011.     

Occurrence of Virulence and Antimicrobial Resistance Genes in Escherichia coli Isolates from Different Aquatic Ecosystems within the St. Clair River and Detroit River Areas

Although the number of Escherichia coli bacteria in surface waters can differ greatly between locations, relatively little is known about the distribution of E. coli pathotypes in surface waters used as sources for drinking or recreation. DNA microarray technology is a suitable tool for this type of study due to its ability to detect high numbers of virulence and antimicrobial resistance genes simultaneously. Pathotype, phylogenetic group, and antimicrobial resistance gene profiles were determined for 308 E. coli isolates from surface water samples collected from diverse aquatic ecosystems at six different sites in the St. Clair River and Detroit River areas. A higher frequency (48%) of E. coli isolates possessing virulence and antimicrobial resistance genes was observed in an urban site located downstream of wastewater effluent outfalls than in the other examined sites (average of 24%). Most E. coli pathotypes were extraintestinal pathogenic E. coli (ExPEC) pathotypes and belonged to phylogenetic groups B2 and D. The ExPEC pathotypes were found to occur across all aquatic ecosystems investigated, including riverine, estuarine, and offshore lake locations. The results of this environmental study using DNA microarrays highlight the widespread distribution of E. coli pathotypes in aquatic ecosystems and the potential public health threat of E. coli pathotypes originating from municipal wastewater sources.     

Complete genome sequence and comparative genome analysis of enteropathogenic Escherichia coli O127:H6 strain E2348/69

Enteropathogenic Escherichia coli (EPEC) was the first pathovar of E. coli to be implicated in human disease; however, no EPEC strain has been fully sequenced until now. Strain E2348/69 (serotype O127:H6 belonging to E. coli phylogroup B2) has been used worldwide as a prototype strain to study EPEC biology, genetics, and virulence. Studies of E2348/69 led to the discovery of the locus of enterocyte effacement-encoded type III secretion system (T3SS) and its cognate effectors, which play a vital role in attaching and effacing lesion formation on gut epithelial cells. In this study, we determined the complete genomic sequence of E2348/69 and performed genomic comparisons with other important E. coli strains. We identified 424 E2348/69-specific genes, most of which are carried on mobile genetic elements, and a number of genetic traits specifically conserved in phylogroup B2 strains irrespective of their pathotypes, including the absence of the ETT2-related T3SS, which is present in E. coli strains belonging to all other phylogroups. The genome analysis revealed the entire gene repertoire related to E2348/69 virulence. Interestingly, E2348/69 contains only 21 intact T3SS effector genes, all of which are carried on prophages and integrative elements, compared to over 50 effector genes in enterohemorrhagic E. coli O157. As E2348/69 is the most-studied pathogenic E. coli strain, this study provides a genomic context for the vast amount of existing experimental data. The unexpected simplicity of the E2348/69 T3SS provides the first opportunity to fully dissect the entire virulence strategy of attaching and effacing pathogens in the genomic context.     

Comparison of 61 Sequenced Escherichia coli Genomes

Escherichia coli is an important component of the biosphere and is an ideal model for studies of processes involved in bacterial genome evolution. Sixty-one publically available E. coli and Shigella spp. sequenced genomes are compared, using basic methods to produce phylogenetic and proteomics trees, and to identify the pan- and core genomes of this set of sequenced strains. A hierarchical clustering of variable genes allowed clear separation of the strains into clusters, including known pathotypes; clinically relevant serotypes can also be resolved in this way. In contrast, when in silico MLST was performed, many of the various strains appear jumbled and less well resolved. The predicted pan-genome comprises 15,741 gene families, and only 993 (6%) of the families are represented in every genome, comprising the core genome. The variable or ‘accessory’ genes thus make up more than 90% of the pan-genome and about 80% of a typical genome; some of these variable genes tend to be co-localized on genomic islands. The diversity within the species E. coli, and the overlap in gene content between this and related species, suggests a continuum rather than sharp species borders in this group of Enterobacteriaceae.