Attaching And Effacing E. Coli Research Articles

Phylogenetic Analysis of Attaching and Effacing E. coli Strains Isolated From Pet Birds in Iran

Background: Enteropathogenic E. coli (EPEC) and Shiga toxin-producing E. coli (STEC) are categorized as attaching and effacing E. coli (AEEC) due to their eae gene. One of the essential causes of diarrhea in humans is AEEC, which affects birds, too, thereby being considered a zoonotic pathogen. Objectives: Our study aimed to determine AEEC and evaluate its antibiotic resistance and phylogroups. Methods: A total of 200 fecal samples were collected from pet birds referred to the Veterinary Medicine Hospital, University of Tehran. PCR methods were used to detect AEEC using uspA, eae, bfpA, stx1, and stx2 gene-specific primers. The antimicrobial susceptibility of the recovered isolates was determined by the agar disk diffusion and MIC methods. Their phylogroups were analyzed based on Clermont phylotyping methods. Results: Of 200 samples, we isolated 26 (13%) E. coli strains, 9 harbor eae genes. None of the ease-positive samples possessed the bfpA gene, but 4 had stx2, and 5 had stx1 and stx2 genes. Phylogenetic analysis identified the phylogenetic groups of all AEEC isolated strains but 2 (duck and cockatiel). Detected phylogroups include four B2 and three D. Based on our results, 7 out of 9 AEEC isolated strains showed multi-drug resistance. Conclusion: The discovery of common phylogroups of AEEC in pet birds (a common companion animal in Iran with intimate contact with their owners, especially children) and humans, as well as their resistance to a wide range of antibiotics used in human medicine, verifies AEEC as a serious public health threat.

Occurrence of Mobile Colistin Resistance Genes mcr-1–mcr-10 including Novel mcr Gene Variants in Different Pathotypes of Porcine Escherichia coli Isolates Collected in Germany from 2000 to 2021

In the European Union, gastrointestinal disease in pigs is the main indication for the use of colistin, but large-scale epidemiologic data concerning the frequency of mobile colistin resistance (mcr) genes in pig-associated pathotypes of Escherichia coli (E. coli) are lacking. Multiplex polymerase chain reactions were used to detect virulence-associated genes (VAGs) and mcr-1–mcr-10 genes in 10,573 porcine E. coli isolates collected in Germany from July 2000 to December 2021. Whole genome sequencing was performed on 220 representative mcr-positive E. coli strains. The total frequency of mcr genes was 10.2%, the most frequent being mcr-1 (8.4%) and mcr-4 (1.6%). All other mcr genes were rarely identified (mcr-2, mcr-3, mcr-5) or absent (mcr-6 to mcr-10). The highest frequencies of mcr genes were found in enterotoxigenic and shiga toxin-encoding E. coli (ETEC/STEC hybrid) and in edema disease E. coli (EDEC) strains (21.9% and 17.7%, respectively). We report three novel mcr variants, mcr-1.36, mcr-4.8, and mcr-5.5. In 39 attaching and effacing E. coli (AEEC) isolates analyzed in our study, the eae subtype β1 was the most prevalent (71.8%). Constant surveillance for the presence of mcr genes in various sectors should consider the different frequency of mcr-positive isolates in pathogenic E. coli.

The Prevalence of Shiga Toxin-Producing Escherichia coli and Enteropathogenic Escherichia coli Isolated from Raw Chicken Meat Samples.

Background Shiga toxin-producing Escherichia coli (STEC) is known as a crucial zoonotic food-borne pathogen. A total of 257 raw chicken meat samples were collected from different markets in Hamadan, west of Iran, from January 2016 to May 2017. Materials and Methods The samples were cultured in selective and differential culture media, and the virulence genes of E. coli isolates were analyzed by PCR assay. The antibiotic resistance patterns of E. coli isolates were determined by the disk diffusion method. The genetic relatedness of the E. coli O157 isolates was analyzed by ERIC-PCR. Results In total, 93 (36% ± 3.12) of the isolates were identified as E. coli in this study. Based on serological and microbiological tests, 36 (38.7% ± 9.9), 7 (7.5% ± 5.35), and 12 (12.9% ± 6.81) of the E. coli isolates were characterized as STEC, enteropathogenic E. coli (EPEC), and attaching and effacing E. coli (AEEC) strains, respectively. A high level of resistance to nalidixic acid (91.4% ± 5.7), tetracycline (89.2% ± 6.31), ampicillin (82.8% ± 7.67), and trimotoprime-sulfametoxazole (71% ± 9.22) was detected among the E. coli isolates. The analysis of the ERIC-PCR results showed five different ERIC types among the E. coli O157 isolates. Conclusions Based on our findings, control and check-up of poultry meats should be considered as a crucial issue for public health.

The Mobilome; A Major Contributor to Escherichia coli stx2-Positive O26:H11 Strains Intra-Serotype Diversity.

Shiga toxin-producing Escherichia coli of serotype O26:H11/H- constitute a diverse group of strains and several clones with distinct genetic characteristics have been identified and characterized. Whole genome sequencing was performed using Illumina and PacBio technologies on eight stx2-positive O26:H11 strains circulating in France. Comparative analyses of the whole genome of the stx2-positive O26:H11 strains indicate that several clones of EHEC O26:H11 are co-circulating in France. Phylogenetic analysis of the French strains together with stx2-positive and stx-negative E. coli O26:H11 genomes obtained from Genbank indicates the existence of four clonal complexes (SNP-CCs) separated in two distinct lineages, one of which comprises the “new French clone” (SNP-CC1) that appears genetically closely related to stx-negative attaching and effacing E. coli (AEEC) strains. Interestingly, the whole genome SNP (wgSNP) phylogeny is summarized in the cas gene phylogeny, and a simple qPCR assay targeting the CRISPR array specific to SNP-CC1 (SP_O26-E) can distinguish between the two main lineages. The PacBio sequencing allowed a detailed analysis of the mobile genetic elements (MGEs) of the strains. Numerous MGEs were identified in each strain, including a large number of prophages and up to four large plasmids, representing overall 8.7–19.8% of the total genome size. Analysis of the prophage pool of the strains shows a considerable diversity with a complex history of recombination. Each clonal complex (SNP-CC) is characterized by a unique set of plasmids and phages, including stx-prophages, suggesting evolution through separate acquisition events. Overall, the MGEs appear to play a major role in O26:H11 intra-serotype clonal diversification.

Targeted Amplicon Sequencing for Single-Nucleotide-Polymorphism Genotyping of Attaching and Effacing Escherichia coli O26:H11 Cattle Strains via a High-Throughput Library Preparation Technique.

Enterohemorrhagic Escherichia coli (EHEC) O26:H11, a serotype within Shiga toxin-producing E. coli (STEC) that causes severe human disease, has been considered to have evolved from attaching and effacing E. coli (AEEC) O26:H11 through the acquisition of a Shiga toxin-encoding gene. Targeted amplicon sequencing using next-generation sequencing technology of 48 phylogenetically informative single-nucleotide polymorphisms (SNPs) and three SNPs differentiating Shiga toxin-positive (stx-positive) strains from Shiga toxin-negative (stx-negative) strains were used to infer the phylogenetic relationships of 178 E. coli O26:H11 strains (6 stx-positive strains and 172 stx-negative AEEC strains) from cattle feces to 7 publically available genomes of human clinical strains. The AEEC cattle strains displayed synonymous SNP genotypes with stx2-positive sequence type 29 (ST29) human O26:H11 strains, while stx1 ST21 human and cattle strains clustered separately, demonstrating the close phylogenetic relatedness of these Shiga toxin-negative AEEC cattle strains and human clinical strains. With the exception of seven stx-negative strains, five of which contained espK, three stx-related SNPs differentiated the STEC strains from non-STEC strains, supporting the hypothesis that these AEEC cattle strains could serve as a potential reservoir for new or existing pathogenic human strains. Our results support the idea that targeted amplicon sequencing for SNP genotyping expedites strain identification and genetic characterization of E. coli O26:H11, which is important for food safety and public health.

Genetic Diversity and Pathogenic Potential of Attaching and Effacing Escherichia coli O26:H11 Strains Recovered from Bovine Feces in the United States.

Escherichia coli O26 has been identified as the most common non-O157 Shiga toxin-producing E. coli (STEC) serogroup to cause human illnesses in the United States and has been implicated in outbreaks around the world. E. coli has high genomic plasticity, which facilitates the loss or acquisition of virulence genes. Attaching and effacing E. coli (AEEC) O26 strains have frequently been isolated from bovine feces, and there is a need to better characterize the relatedness of these strains to defined molecular pathotypes and to describe the extent of their genetic diversity. High-throughput real-time PCR was used to screen 178 E. coli O26 isolates from a single U.S. cattle feedlot, collected from May to July 2011, for the presence or absence of 25 O26 serogroup-specific and virulence-associated markers. The selected markers were capable of distinguishing these strains into molecularly defined groups (yielding 18 unique marker combinations). Analysis of the clustered regularly interspaced short palindromic repeat 1 (CRISPR1) and CRISPR2a loci further discriminated isolates into 24 CRISPR types. The combination of molecular markers and CRISPR typing provided 20.8% diversity. The recent CRISPR PCR target SP_O26-E, which was previously identified only in stx2-positive O26:H11 human clinical strains, was identified in 96.4% (161/167 [95% confidence interval, 99.2 to 93.6%]) of the stx-negative AEEC O26:H11 bovine fecal strains. This supports that these stx-negative strains may have previously contained a prophage carrying stx or could acquire this prophage, thus possibly giving them the potential to become pathogenic to humans. These results show that investigation of specific genetic markers may further elucidate our understanding of the genetic diversity of AEEC O26 strains in bovine feces.

Shiga toxin-producing Escherichia coli isolated from chicken meat in Iran: Serogroups, virulence factors, and antimicrobial resistance properties

The aim of the current study was to determine the virulence factors, serogroups, and antibiotic resistance properties of Shiga toxin-producing Escherichia coli isolated from chicken meat samples. A total of 422 chicken meat samples were collected from 5 townships of Iran. Specimens were immediately transferred to the laboratory in a cooler with an ice pack. Samples were cultured, and the positive culture samples were analyzed by PCR assays. Finally, the antimicrobial susceptibility test was performed using the disk diffusion method in Mueller–Hinton agar. According to the results, out of 422 samples, 146 (34.59%) were confirmed to be E. coli positive and among E. coli-positive samples, 51 (34.93%) and 31 (21.23%) were from attaching and effacing E. coli (AEEC) and enterohemorrhagic E. coli (EHEC) subgroups, respectively. All of the EHEC-positive samples had all stx1, eaeA, and ehly virulence genes, whereas only 5 (9.80%) of AEEC subgroup had all stx1, stx2, and eaeA genes. As the data revealed, O157 was the most prevalent and O111 was the least prevalent strains in the Shiga toxin-producing E. coli (STEC) population. Among STEC strains, sulI and blaSHV had the highest and lowest incidence rate, respectively. There was a high resistance to tetracycline (76.82%), followed by chloramphenicol (73.17%) and nitrofurantoin (63.41%), but there was low resistance to cephalotine (7.31%) antibiotics in isolated strains. Results shows that the PCR technique has a high performance for detection of serogroups, virulence genes, and antibiotic resistance genes in STEC strains. This study is the first prevalence report of detection of virulence genes, serogroups, and antibiotic resistance properties of STEC strains isolated from chicken meat samples in Iran. Based on the results, chicken meat is one of the main sources of STEC strains and its virulence factors in Iran, so an accurate meat inspection would reduce disease outbreaks.

PCR MULTIPLEX PARA DETERMINAÇÃO DE MARCADORES DE VIRULÊNCIA EM Escherichia coli ISOLADAS DE BOVINOS COM DIARRÉIA EM MINAS GERAIS

Diarrhea is one of the most common diseases in calves and leads to substantial economic loss due to treatment costs, increased mortality and morbidity rates, and growth constraints. Enterotoxigenic Escherichia coli (ETEC), attaching and effacing E. coli (AEEC) and Shiga toxin-producing E. coli (STEC) have been identified as causes of diarrhea and dysentery in calves. This study investigates virulence factors of E. coli strains isolated from calves by multiplex PCR. The animals were up to 60 days old and were raised in Minas Gerais State, Brazil. Samples were obtained from semi-intensive dairy farms in the center-west and Alto Paranaiba regions of Minas Gerais. Feces were collected and processed routinely. The DNA templates of E. coli strains were subjected to multiplex PCR for the detection of the genes encoding the virulence factors Stx1, Stx2, Int, F41, K99, and STa. Out of 156 isolates, 51 (32,7%) were positive, whereas 105 (67,3%) did not present any of the virulence factors investigated. Results show that E. coli is an important cause of diarrhea in calves in Minas Gerais and that multiplex PCR test is a useful tool for the identification and characterization of pathogenic E. coli strains.

Attaching and Effacing Escherichia coli Downregulate DNA Mismatch Repair Protein In Vitro and Are Associated with Colorectal Adenocarcinomas in Humans

BackgroundMucosa-associated Escherichia coli are frequently found in the colonic mucosa of patients with colorectal adenocarcinoma, but rarely in healthy controls. Chronic mucosal E. coli infection has therefore been linked to colonic tumourigenesis. E. coli strains carrying eae (encoding the bacterial adhesion protein intimin) attach intimately to the intestinal mucosa and are classed as attaching and effacing E. coli (AEEC). Enteropathogenic Escherichia coli (EPEC) are the most common form of AEEC identified in man. EPEC utilise a type III secretion system to translocate effector proteins into host cells and infection induces wide-ranging effects on the host cell proteome. We hypothesised that EPEC infection could influence molecular pathways involved in colorectal tumourigenesis.Methodology/Principal FindingsWhen co-cultured with human colorectal cell lines, EPEC dramatically downregulated the expression of key DNA mismatch repair proteins MSH2 and MLH1 in an attachment specific manner. Cytochrome c staining and TUNEL analysis confirmed that this effect was not a consequence of apoptosis/necrosis. Ex vivo human colonic mucosa was co-cultured with EPEC and probed by immunofluorescence to locate adherent bacteria. EPEC entered 10% of colonic crypts and adhered to crypt epithelial cells, often in the proliferative compartment. Adenocarcinoma and normal colonic mucosa from colorectal cancer patients (n = 20) was probed by immunofluorescence and PCR for AEEC. Mucosa-associated E. coli were found on 10/20 (50%) adenocarcinomas and 3/20 (15%) normal mucosa samples (P<0.05). AEEC were detected on 5/20 (25%) adenocarcinomas, but not normal mucosa samples (P<0.05).Significance/ConclusionsThe ability of EPEC to downregulate DNA mismatch repair proteins represents a novel gene-environment interaction that could increase the susceptibility of colonic epithelial cells to mutations and therefore promote colonic tumourigenesis. The potential role of AEEC in colorectal tumourigenesis warrants further investigation.

Occurrence of non-O157 Shiga toxin-producing Escherichia coli in dogs with diarrhea

Shiga toxigenic Escherichia coli (STEC) and Attaching and effacing E. coli (AEEC) have been associated with diarrhea illness in dogs. From January to December 2006, 92 E. coli isolates from 25 diarrheic dogs were analyzed, by screening for the presence of Shiga toxin-producing (stx 1 and stx 2) and intimin (eae) genes. Twelve isolates were detected by PCR to harbor the Shiga toxin genes (7 the stx 1 (7.6%); 5 the stx 2 (5.4%); and none both of them). Nine (9.8%) of the E. coli isolates studied were eae positive non Shiga toxin-producing. Thirteen (62.0%) isolates, carrying stx or eae gene, also showed a hemolysin production. The strains with virulence genes were also examined for resistance to 12 antimicrobial agents. Resistances to cephalothin (85.7%), streptomycin (81.0%), amoxicillin (71.4%) and gentamicin (71.4%) were predominantly observed.

Prevalence of enterohaemorrhagic Escherichia coli from serotype O157 and other attaching and effacing Escherichia coli on bovine carcasses in Algeria

Bovine meat is the principal source of human contamination of attaching and effacing Escherichia coli, including enterohaemorrhagic E. coli O157. The aim was to study the prevalence of these strains on bovine carcasses in Algeria. Two-hundred and thirty carcasses were swabbed and analysed by classical microbiological methods for total E. coli counts and for the presence of pathogenic E. coli. The E. coli counts were high, with a 75th percentile of 444.75 CFUs cm(-2). For pathogenic E. coli, more than 7% of the tested carcasses were positive for E. coli O157. Eighteen E. coli O157 strains were isolated and typed by multiplex PCR. The main isolated pathotype (78%) was eae+ stx2+ ehxA+. In addition to E. coli O157, other attaching and effacing E. coli (AEEC) were also detected from carcasses by colony hybridization after pre-enrichment and plating on sorbitol MacConkey agar using eae, stx1 and stx2 probes. Thirty carcasses (13%) on the 230 analysed harboured at least one colony positive for one of the tested probes. These positive carcasses were different from those positive for E. coli O157. Sixty-six colonies (2.9%) positive by colony hybridization were isolated. The majority (60.6%) of the positive strains harboured an enteropathogenic E. coli-like pathotype (eae+ stx-). Only three enterohaemorrhagic E. coli (EHEC)-like (eae+ stx1+) colonies were isolated from the same carcass. These strains did not belong to classical EHEC serotypes. In this study, the global hygiene of the slaughterhouse was low, as indicated by the high level of E. coli count. The prevalence of both E. coli O157 and other AEEC was also high, representing a real hazard for consumers. This is the first study of this type in Algeria, which indicates that the general hygiene of the slaughterhouse must be improved.

Prevalence of Pathogenic Escherichia coli in a Swine Breeding Environment in Can Tho Province, Vietnam

To determine the presence of enterotoxigenic Escherichia coli (E. coli) (ETEC), Shiga toxin-producing E. coli (STEC) and attaching and effacing E. coli (AEEC) were collected in swine and in a water system in 17 swine farms in the Can Tho province of the Mekong delta. A total of 258 samples which were collected from swine feces (n=169), from feed (n=39) and from water (n=50) in the irrigation system were examined by PCR to determine whether they harbored the stx, eae, and enterotoxin genes. STEC was detected in 10 of 169 (6%) fecal samples, and AEEC in 5 fecal samples (3%). Both STEC and AEEC were detected in one fecal sample. AEEC was mainly detected in fecal samples from pigs that were less than 4 months old. STEC or AEEC was detected in 18 samples from 5 out of 17 (29%) farms. On the other hand, STEC and AEEC were also detected in the water and feed samples. ETEC was not detected in diarrhea samples.

Gastric Attaching and Effacing Escherichia coli Lesions in a Puppy with Naturally Occurring Enteric Colibacillosis and Concurrent Canine Distemper Virus Infection

A puppy suffering from chronic diarrhea was humanely killed at 90 days of age. Numerous Gram-negative bacilli were found adhering to the surface of as well as within epithelial cells from the stomach to the colon. Canine distemper virus inclusions were in the epithelial cytoplasm of the esophageal, gastric, and intestinal mucosa. Typical attaching and effacing ultrastructural lesions were in the stomach, and some bacilli were in the cytoplasm of the epithelial cells. Escherichia coli, isolated from the contents of the small intestine, belonged to serotype 0118: NM and were negative for plasmid-encoded EPEC adherence factor (EAF) and positive for the E. coli attaching effacing (eae) gene. Immunohistologically, bacilli attached to the epithelium from the stomach to the colon were positive for antisera against E. coli 0118. E. coli 0118: NM inoculated into human tissue culture cells (HEp-2 cells) were attached to the surface of the cells and within the cytoplasm. This is the first report of attaching and effacing E. coli (AEEC) infection in the stomach of the dog.

Dual infection with attaching and effacing Escherichia coli and enterotoxigenic Escherichia coli in post-weaning pigs

Post-weaning diarrhoea in pigs occurred on two farms in Hokkaido, Japan, in 1994. Four piglets aged 35 or 45 days were examined after death. At necropsy, ecchymotic haemorrhages were seen on the mucosal surface of the caecum and colon. Histopathologically, numerous Gram-negative bacilli adhered to the brush border of the small intestines, but the brush border itself was intact. Typical attaching and effacing (AE) lesions were seen in the caecum and colon. Immunohistochemically, the bacilli which adhered to the brush border gave positive results with antisera against serogroup O149 of Escherichia coli; the bacilli which caused the AE lesions, however, belonged to serogroup O45. It was concluded that the disease resulted from dual infection with attaching and effacing E. coli (AEEC) and enterotoxigenic E. coli (ETEC).

Biotype, serotype, and pathogenicity of attaching and effacing enteropathogenic Escherichia coli strains isolated from diarrheic commercial rabbits.

A total of 568 strains of Escherichia coli isolated from healthy and diarrheic rabbits were separated into 11 different biotypes according to the fermentation patterns of four carbohydrates. Strains belonging to biotypes 1 to 3, 6, and 8 induced lesions characteristic for attaching and effacing E. coli (AEEC). They attached to the intestinal epithelium of the terminal small intestine and the large intestine of 5-week-old rabbits after experimental infection and caused effacement of the microvillous brush border. However, pathogenicity for weaned rabbits, as judged by diarrhea score, anorexia, and reduced weight gain, varied according to the biotypes of the strains. Strains belonging to biotypes 1 and 6 produced only discrete clinical signs, strains belonging to biotypes 2 and 3+ (motile) induced diarrhea and growth depression, whereas strains belonging to biotypes 3- (immotile) and 8 caused severe clinical signs and high mortality. This confirms evidence from the field. Biotypes 3- and 8, accounting for 35.5 and 7.1% of AEEC strains in weaned diarrheic rabbits, respectively, were not detected in weaned healthy rabbits, while biotype 2 was the predominant strain in weaned healthy rabbits (62.3%). Finally, serotyping showed a close relationship between biotype and serotype of the AEEC examined. Most strains of biotypes 1+ and 2+ tested were O109:K-:H2 and O132:K-:H2, respectively, whereas all strains tested of biotype 3- were O15:K-:H- and those of biotype 8 were O103:K-:H2. These data indicate that specific clones of AEEC might be involved in juvenile rabbit enteritis. It was concluded that determination of biotypes allows the screening of highly pathogenic AEEC in weaned rabbits (biotypes 3- and 8).