Article

Escherichia coli O157:H7 strains that persist in feedlot cattle are genetically related and demonstrate an enhanced ability to adhere to intestinal epithelial cells.

Center for Meat Safety & Quality, Department of Animal Sciences, Colorado State University, 7C Animal Sciences, Ft. Collins, CO 80523-1171, USA.
Applied and Environmental Microbiology (Impact Factor: 3.95). 08/2009; 75(18):5927-37. DOI: 10.1128/AEM.00972-09
Source: PubMed

ABSTRACT A longitudinal study was conducted to investigate the nature of Escherichia coli O157:H7 colonization of feedlot cattle over the final 100 to 110 days of finishing. Rectal fecal grab samples were collected from an initial sample population of 788 steers every 20 to 22 days and microbiologically analyzed to detect E. coli O157:H7. The identities of presumptive colonies were confirmed using a multiplex PCR assay that screened for gene fragments unique to E. coli O157:H7 (rfbE and fliC(h7)) and other key virulence genes (eae, stx(1), and stx(2)). Animals were classified as having persistent shedding (PS), transient shedding (TS), or nonshedding (NS) status if they consecutively shed the same E. coli O157:H7 genotype (based on the multiplex PCR profile), exhibited variable E. coli O157 shedding, or never shed morphologically typical E. coli O157, respectively. Overall, 1.0% and 1.4% of steers were classified as PS and NS animals, respectively. Characterization of 132 E. coli O157:H7 isolates from PS and TS animals by pulsed-field gel electrophoresis (PFGE) typing yielded 32 unique PFGE types. One predominant PFGE type accounted for 53% of all isolates characterized and persisted in cattle throughout the study. Isolates belonging to this predominant and persistent PFGE type demonstrated an enhanced (P < 0.0001) ability to adhere to Caco-2 human intestinal epithelial cells compared to isolates belonging to less common PFGE types but exhibited equal virulence expression. Interestingly, the attachment efficacy decreased as the genetic divergence from the predominant and persistent subtype increased. Our data support the hypothesis that certain E. coli O157:H7 strains persist in feedlot cattle, which may be partially explained by an enhanced ability to colonize the intestinal epithelium.

Full-text

Available from: Keith E Belk, Mar 31, 2015
0 Bookmarks
 · 
88 Views
  • [Show abstract] [Hide abstract]
    ABSTRACT: The need to quantify the potential human health risk posed by the bovine reservoir of Escherichia coli O157 has led to a wealth of prevalence studies and improvements in detection methods over the last two decades. Rectoanal mucosal swabs have been used for the detection of E. coli O157 fecal shedding, colonized animals, and those predisposed to super shedding. We conducted a longitudinal study to compare the detection of E. coli O157 from feces and rectoanal mucosal swabs (RAMS) from a cohort of dairy heifers. We collected 820 samples that were tested by immunomagnetic separation of both feces and RAMS. Of these, 132 were detected as positive for E. coli O157 from both samples, 66 were detected as positive from RAMS only, and 117 were detected as positive from feces only. The difference in results between the two sample types was statistically significant (P < 0.001). The relative sensitivities of detection by immunomagnetic separation were 53% (confidence interval, 46.6 to 59.3) from RAMS and 67% (confidence interval, 59.6 to 73.1) from fecal samples. No association between long-term shedding (P = 0.685) or super shedding (P = 0.526) and detection by RAMS only was observed.
    Journal of food protection 06/2014; 77(6):972-6. DOI:10.4315/0362-028X.JFP-13-500 · 1.80 Impact Factor
  • [Show abstract] [Hide abstract]
    ABSTRACT: IntroductionSalmonella, Shigella and Escherichia coliO157: H7 are major foodborne pathogens that cause gastrointestinal diseases worldwide. Apart from food contamination, fecal pollution has been consistently associated with the transmission of these pathogens, and their rapid detection in food and stools is of significance for food safety. However, a variety of factors associated with these complex samples can decrease the sensitivity and specificity of molecular-based methods for detection of these pathogens.Objectives The aim of this study was to develop a DNA-based method for the simultaneous detection of E. coliO157: H7, Salmonella and Shigella in stool and food samples.Methods In this study, a novel magnetic capture–multiplex polymerase chain reaction (PCR) assay was developed and its potential to detect the target pathogens in stool and food samples (including chicken, cucumbers and cooked rice) was tested.ResultsThe results showed that the magnetic particles (MPs) used in the study had a high capacity for bacterial adsorption. The pretreatment protocol, which included the pathogen concentration by MPs, was developed and the sensitivity of the assay was approximately 10° colony-forming unit (CFU) g−1 in food and 1–10 CFUs per stool sample, following an enrichment step. The assay could be completed within 12 h, and was comparable in performance with conventional culture methods, which require several days to complete.Conclusion The assay combines MP-based magnetic capture with multiplex PCR, and offers an efficient, rapid, sensitive and inexpensive alternative for the routine detection of foodborne pathogenic bacteria.
    Quality Assurance and Safety of Crops & Foods 12/2011; 3(4). DOI:10.1111/j.1757-837X.2011.00113.x · 0.73 Impact Factor
  • Source
    [Show abstract] [Hide abstract]
    ABSTRACT: Shiga toxin-producing Escherichia coli O157:H7 (O157) are significant foodborne pathogens and pose a serious threat to public health worldwide. The major reservoirs of O157 are asymptomatic cattle which harbor the organism in the terminal recto-anal junction (RAJ). Some colonized animals, referred to as "super-shedders" (SS), are known to shed O157 in exceptionally large numbers (>104 CFU/g of feces). Recent studies suggest that SS cattle play a major role in the prevalence and transmission of O157, but little is known about the molecular mechanisms associated with super-shedding. Whole genome sequence analysis of an SS O157 strain (SS17) revealed a genome of 5,523,849 bp chromosome with 5,430 open reading frames and two plasmids, pO157 and pSS17, of 94,645 bp and 37,446 bp, respectively. Comparative analyses showed that SS17 is clustered with spinach-associated O157 outbreak strains, and belongs to the lineage I/II, clade 8, D group, and genotype 1, a subgroup of O157 with predicted hyper-virulence. A large number of non-synonymous SNPs and other polymorphisms were identified in SS17 as compared with other O157 strains (EC4115, EDL933, Sakai, TW14359), including in key adherence- and virulence-related loci. Phenotypic analyses revealed a distinctive and strongly adherent aggregative phenotype of SS17 on bovine RAJ stratified squamous epithelial (RSE) cells that was conserved amongst other SS isolates. Molecular genetic and functional analyses of defined mutants of SS17 suggested that the strongly adherent aggregative phenotype amongst SS isolates is LEE-independent, and likely results from a novel mechanism. Taken together, our study provides a rational framework for investigating the molecular mechanisms associated with SS, and strong evidence that SS O157 isolates have distinctive features and use a LEE-independent mechanism for hyper-adherence to bovine rectal epithelial cells.
    PLoS ONE 11/2014; 10(2). DOI:10.1371/journal.pone.0116743 · 3.53 Impact Factor