Comparative genomics of two super-shedder isolates of Escherichia coli O157:H7

Shiga toxin-producing Escherichia coli O157:H7 (O157) are zoonotic foodborne pathogens and of major public health concern that cause considerable intestinal and extra-intestinal illnesses in humans. O157 colonize the recto-anal junction (RAJ) of asymptomatic cattle who shed the bacterium into the environment through fecal matter. A small subset of cattle, termed super-shedders (SS), excrete O157 at a rate (104 CFU/g of feces) that is several orders of magnitude greater than other colonized cattle and play a major role in the prevalence and transmission of O157. To better understand microbial factors contributing to super-shedding we have recently sequenced two SS isolates, SS17 (GenBank accession no. CP008805) and SS52 (GenBank accession no. CP010304) and shown that SS isolates display a distinctive strongly adherent phenotype on bovine rectal squamous epithelial cells. Here we present a detailed comparative genomics analysis of SS17 and SS52 with other previously characterized O157 strains (EC4115, EDL933, Sakai, TW14359). The results highlight specific polymorphisms and genomic features shared amongst SS strains, and reveal several SNPs that are shared amongst SS isolates, including in genes involved in motility, adherence, and metabolism. Finally, our analyses reveal distinctive patterns of distribution of phage-associated genes amongst the two SS and other isolates. Together, the results of our comparative genomics studies suggest that while SS17 and SS52 share genomic features with other lineage I/II isolates, they likely have distinct recent evolutionary histories. Future comparative and functional genomic studies are needed to decipher the precise molecular basis for super shedding in O157

Accelerating the Deactivation of \u3cem\u3eSalmonella enterica\u3c/em\u3e Serovar Newport and \u3cem\u3eEscherichia coli\u3c/em\u3e O157:H7 in Dairy Manure by Modifying pH or Temperature

To assess methods for control of disease-causing bacteria in animal manures prior to field application, we manipulated the temperature or adjusted pH of dairy manure to high (3.5 to 5) or low (10 to 12) values with aluminum sulfate or hydrated lime, and inoculated the manure with Salmonella enterica serovar Newport or Escherichia coli O157:H7, then incubated the manure at ambient temperature. At pH 4.2, S. Newport was eliminated within 6 days; however at pH \u3e4.2 S. Newport was suppressed only temporarily and recovered to concentrations near the unamended controls. pH required to eliminate E. coli O157:H7 was 4.5. Both pathogens were killed by pH 11.0. The pathogens were eliminated within 2 weeks when inoculated manure was incubated at 37°C, whereas at 22°C and 4°C, the organisms persisted for much longer periods. S. Newport survived for over 300 days at 4°C, which has implications for manure spreading in colder seasons

Associations Between Multidrug Resistance, Plasmid Content, and Virulence Potential Among Extraintestinal Pathogenic and Commensal Escherichia coli from Humans and Poultry

The emergence of plasmid-mediated multidrug resistance (MDR) among enteric bacteria presents a serious challenge to the treatment of bacterial infections in humans and animals. Recent studies suggest that avianEscherichia coli commonly possess the ability to resist multiple antimicrobial agents, and might serve as reservoirs of MDR for human extraintestinal pathogenic Escherichia coli (ExPEC) and commensal E. coli populations. We determined antimicrobial susceptibility profiles for 2202 human and avian E. coli isolates, then sought for associations among resistance profile, plasmid content, virulence factor profile, and phylogenetic group. Avian-source isolates harbored greater proportions of MDR than their human counterparts, and avian ExPEC had higher proportions of MDR than did avian commensal E. coli. MDR was significantly associated with possession of the IncA/C, IncP1-α, IncF, and IncI1 plasmid types. Overall, inferred virulence potential did not correlate with drug susceptibility phenotype. However, certain virulence genes were positively associated with MDR, including ireA,ibeA, fyuA, cvaC, iss, iutA, iha, and afa. According to the total dataset, isolates segregated significantly according to host species and clinical status, thus suggesting that avian and human ExPEC and commensal E. coli represent four distinct populations with limited overlap. These findings suggest that in extraintestinal E. coli, MDR is most commonly associated with plasmids, and that these plasmids are frequently found among avian-source E. coli from poultry production systems

Antimicrobial Resistance of Escherichia coli O26, O103, O111, O128, and O145 from Animals and Humans

Susceptibilities to fourteen antimicrobial agents important in clinical medicine and agriculture were determined for 752 Escherichia coli isolates of serotypes O26, O103, O111, O128, and O145. Strains of these serotypes may cause urinary tract and enteric infections in humans and have been implicated in infections with Shiga toxin–producing E. coli (STEC). Approximately 50% of the 137 isolates from humans were resistant to ampicillin, sulfamethoxazole, cephalothin, tetracycline, or streptomycin, and approximately 25% were resistant to chloramphenicol, trimethoprim-sulfamethoxazole, or amoxicillin-clavulanic acid. Approximately 50% of the 534 isolates from food animals were resistant to sulfamethoxazole, tetracycline, or streptomycin. Of 195 isolates with STEC-related virulence genes, approximately 40% were resistant to sulfamethoxazole, tetracycline, or streptomycin. Findings from this study suggest antimicrobial resistance is widespread among E. coli O26, O103, O111, O128, and O145 inhabiting humans and food animals

Global Distribution of O Serotypes and Antibiotic Resistance in Extraintestinal Pathogenic Escherichia coli Collected From the Blood of Patients With Bacteremia Across Multiple Surveillance Studies

Background: Extraintestinal pathogenic Escherichia coli (ExPEC) is the leading cause of bacteremia worldwide, with older populations having increased risk of invasive bacterial disease. Increasing resistance to first-line antibiotics and emergence of multidrug-resistant (MDR) strains represent major treatment challenges. ExPEC O serotypes are key targets for potential multivalent conjugate vaccine development. Therefore, we evaluated the O serotype distribution and antibiotic resistance profiles of ExPEC strains causing bloodstream infections across 4 regions. Methods: Blood culture isolates from patients aged ≥60 years collected during 5 retrospective E. coli surveillance studies in Europe, North America, Asia-Pacific, and South America (2011-2017) were analyzed. Isolates were O serotyped by agglutination; O genotyping was performed for nontypeable isolates. Antimicrobial susceptibility testing was also conducted. Results: Among 3217 ExPEC blood culture isolates, the most ubiquitous O serotype was O25 (n = 737 [22.9%]), followed by O2, O6, O1, O75, O15, O8, O16, O4, O18, O77 group, O153, O9, O101/O162, O86, and O13 (prevalence of ≥1%). The prevalence of these O serotypes was generally consistent across regions, apart from South America; together, these 16 O serotypes represented 77.6% of all ExPEC bacteremia isolates analyzed. The overall MDR frequency was 10.7%, with limited variation between regions. Within the MDR subset (n = 345), O25 showed a dominant prevalence of 63.2% (n = 218). Conclusions: Predominant O serotypes among ExPEC bacteremia isolates are widespread across different regions. O25 was the most prevalent O serotype overall and particularly dominant among MDR isolates. These findings may inform the design of multivalent conjugate vaccines that can target the predominant O serotypes associated with invasive ExPEC disease in older adults

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Not AvailableFourier transform–infrared spectroscopy, in conjunction with artificial neural networks, has been used for identification and classification of selected foodborne pathogens. Five bacterial species (Enterococcus faecium, Salmonella Enteritidis, Bacillus cereus, Yersinia enterocolitica, Shigella boydii) and five Escherichia coli strains (O103, O55, O121, O30, O26) suspended in phosphate-buffered saline were enumerated to provide seven different concentrations ranging from 109 to 103 CFU/ml. The trained artificial neural networks were then validated with an independent subset of samples and compared with the traditional plate count method. It was found that the concentration-based classification of the species was 100% correct and the strain-based classification was 90 to 100% accurate.Not Availabl

Comparison of antimicrobial resistant genes in chicken gut microbiome grown on organic and conventional diet

Antibiotics are widely used in chicken production for therapeutic purposes, disease prevention and growth promotion, and this may select for drug resistant microorganisms known to spread to humans through consumption of contaminated food. Raising chickens on an organic feed regimen, without the use of antibiotics, is increasingly popular with the consumers. In order to determine the effects of diet regimen on antibiotic resistant genes in the gut microbiome, we analyzed the phylotypes and identified the antimicrobial resistant genes in chicken, grown under conventional and organic dietary regimens. Phylotypes were analyzed from DNA extracted from fecal samples from chickens grown under these dietary conditions. While gut microbiota of chicken raised in both conventional and organic diet exhibited the presence of DNA from members of Proteobacteria and Bacteroidetes, organic diet favored the growth of members of Fusobacteria. Antimicrobial resistance genes were identified from metagenomic libraries following cloning and sequencing of DNA fragments from fecal samples and selecting for the resistant clones (n=340) on media containing different concentrations of eight antibiotics. The antimicrobial resistant genes exhibited diversity in their host distribution among the microbial population and expressed more in samples from chicken grown on a conventional diet at higher concentrations of certain antimicrobials than samples from chicken grown on organic diet. Further studies will elucidate if this phenomena is widespread and whether the antimicrobial resistance is indeed modulated by diet. This may potentially assist in defining strategies for intervention to reduce the prevalence and dissemination of antibiotic resistance genes in the production environment

Emerging Approaches for Typing, Detection, Characterization, and Traceback of Escherichia coli

Pathogenic Escherichia coli strains cause a large number of diseases in humans, including diarrhea, hemorrhagic colitis, hemolytic uremic syndrome, urinary tract infections, and neonatal meningitis, while in animals they cause diseases such as calf scours and mastitis in cattle, post-weaning diarrhea and edema disease in pigs, and peritonitis and airsacculitis in chickens. The different E. coli pathotypes are characterized by the presence of specific sets of virulence-related genes. Therefore, it is not surprising that pathogenic E. coli constitutes a genetically heterogeneous family of bacteria, and they are continuing to evolve. Rapid and accurate molecular methods are critically needed to detect and trace pathogenic E. coli in food and animals. They are also needed for epidemiological investigations to enhance food safety, as well as animal and human health and to minimize the size and geographical extent of outbreaks. The serotype of E. coli strains has traditionally been determined using antisera raised against the >180 different O- (somatic) and 53 H- (flagellar) antigens. However, there are many problems associated with serotyping, including: it is labor-intensive and time consuming; cross reactivity of the antisera with different serogroups occurs; antisera are available only in specialized laboratories; and many strains are non-typeable. Molecular serotyping targeting O-group-specific genes within the E. coli O-antigen gene clusters and genes that are involved in encoding for the different flagellar types offers an improved approach for determining the E. coli O- and H-groups. Furthermore, molecular serotyping can be coupled with determination of specific sets of virulence genes carried by the strain offering the possibility to determine O-group, pathotype, and the pathogenic potential simultaneously. Sequencing of the O-antigen gene clusters of all of the known O-groups of E. coli is now complete, and the sequences have been deposited in the GenBank database. The sequence information has revealed that some E. coli serogroups have identical sequences while others have point mutations or insertion sequences and type as different serogroups in serological reactions. There are also a number of other ambiguities in serotyping that need to be resolved. Furthermore, new E. coli O-groups are being identified. Therefore, there is an essential need to resolve these issues and to revise the E. coli serotype nomenclature based on these findings. There are emerging technologies that can potentially be applied for molecular serotyping and detection and characterization of E. coli. On a related topic, the genome sequence of thousands of E. coli strains have been deposited in GenBank, and this information is revealing unique markers such as CRISPR (clustered regularly interspaced short palindromic repeats) and virulence gene markers that could be used to identify E. coli pathotypes. Whole genome sequencing now provides the opportunity to study the role of horizontal gene transfer in the evolution and emergence of pathogenic E. coli strains. Whole genome sequencing approaches are being investigated for genotyping and outbreak investigation for regulatory and public health needs; however, there is a need for establishing bioinformatics pipelines able to handle large amounts of data as we move toward the use of genetic approaches for non-culture-based detection and characterization of E. coli and for outbreak investigations

Emerging Approaches for Typing, Detection, Characterization, and Traceback of Escherichia coli, 2nd Edition

Pathogenic Escherichia coli strains cause a large number of diseases in humans, including diarrhea, hemorrhagic colitis, hemolytic uremic syndrome, urinary tract infections, and neonatal meningitis, while in animals they cause diseases such as calf scours and mastitis in cattle, post-weaning diarrhea and edema disease in pigs, and peritonitis and airsacculitis in chickens. The different E. coli pathotypes are characterized by the presence of specific sets of virulence-related genes. Therefore, it is not surprising that pathogenic E. coli constitutes a genetically heterogeneous family of bacteria, and they are continuing to evolve. Rapid and accurate molecular methods are critically needed to detect and trace pathogenic E. coli in food and animals. They are also needed for epidemiological investigations to enhance food safety, as well as animal and human health and to minimize the size and geographical extent of outbreaks. The serotype of E. coli strains has traditionally been determined using antisera raised against the >180 different O- (somatic) and 53 H- (flagellar) antigens. However, there are many problems associated with serotyping, including: it is labor-intensive and time consuming; cross reactivity of the antisera with different serogroups occurs; antisera are available only in specialized laboratories; and many strains are non-typeable. Molecular serotyping targeting O-group-specific genes within the E. coli O-antigen gene clusters and genes that are involved in encoding for the different flagellar types offers an improved approach for determining the E. coliO- and H-groups. Furthermore, molecular serotyping can be coupled with determination of specific sets of virulence genes carried by the strain offering the possibility to determine O-group, pathotype, and the pathogenic potential simultaneously. Sequencing of the O-antigen gene clusters of all of the known O-groups of E. coli is now complete, and the sequences have been deposited in the GenBank database. The sequence information has revealed that some E. coli serogroups have identical sequences while others have point mutations or insertion sequences and type as different serogroups in serological reactions. There are also a number of other ambiguities in serotyping that need to be resolved. Furthermore, new E. coli O-groups are being identified. Therefore, there is an essential need to resolve these issues and to revise the E. coli serotype nomenclature based on these findings. There are emerging technologies that can potentially be applied for molecular serotyping and detection and characterization of E. coli. On a related topic, the genome sequence of thousands of E. coli strains have been deposited in GenBank, and this information is revealing unique markers such as CRISPR (clustered regularly interspaced short palindromic repeats) and virulence gene markers that could be used to identify E. coli pathotypes. Whole genome sequencing now provides the opportunity to study the role of horizontal gene transfer in the evolution and emergence of pathogenic E. coli strains. Whole genome sequencing approaches are being investigated for genotyping and outbreak investigation for regulatory and public health needs; however, there is a need for establishing bioinformatics pipelines able to handle large amounts of data as we move toward the use of genetic approaches for non-culture-based detection and characterization of E. coli and for outbreak investigations

Comparative genomics of two super-shedder isolates of Escherichia coli O157:H7

Shiga toxin-producing Escherichia coli O157:H7 (O157) are zoonotic foodborne pathogens and of major public health concern that cause considerable intestinal and extra-intestinal illnesses in humans. O157 colonize the recto-anal junction (RAJ) of asymptomatic cattle who shed the bacterium into the environment through fecal matter. A small subset of cattle, termed super-shedders (SS), excrete O157 at a rate (104 CFU/g of feces) that is several orders of magnitude greater than other colonized cattle and play a major role in the prevalence and transmission of O157. To better understand microbial factors contributing to super-shedding we have recently sequenced two SS isolates, SS17 (GenBank accession no. CP008805) and SS52 (GenBank accession no. CP010304) and shown that SS isolates display a distinctive strongly adherent phenotype on bovine rectal squamous epithelial cells. Here we present a detailed comparative genomics analysis of SS17 and SS52 with other previously characterized O157 strains (EC4115, EDL933, Sakai, TW14359). The results highlight specific polymorphisms and genomic features shared amongst SS strains, and reveal several SNPs that are shared amongst SS isolates, including in genes involved in motility, adherence, and metabolism. Finally, our analyses reveal distinctive patterns of distribution of phage-associated genes amongst the two SS and other isolates. Together, the results of our comparative genomics studies suggest that while SS17 and SS52 share genomic features with other lineage I/II isolates, they likely have distinct recent evolutionary histories. Future comparative and functional genomic studies are needed to decipher the precise molecular basis for super shedding in O157