H7 In Cattle Research Articles

Reduction of Escherichia coli O157:H7 in Finishing Cattle Fed Corn Genetically Modified to Produce Increased Concentrations of Alpha Amylase in the Corn Kernel.

Cattle are recognized as the principal reservoir for Escherichia coli O157:H7 and preharvest food safety efforts often focus on decreasing shedding of this pathogen in cattle feces. Enogen® corn (EC; Syngenta Seeds, LLC) is genetically modified to produce enhanced concentrations of α-amylase in the corn kernel endosperm. Research has demonstrated improvements in feed efficiency for cattle fed EC and research has not yet explored whether improved digestion impacts foodborne pathogen populations in cattle. Therefore, this study explored effects of finishing diets containing EC on Escherichia coli O157:H7 prevalence in cattle. A 2 × 2 factorial experiment was conducted with steers (n = 960) fed diets consisting of 2 types of silage (EC or Control) and grain (EC or Control), fed daily ad libitum. Steers were grouped into 12 blocks by incoming body weight, blocks were randomly assigned to one of four pens, and pens were randomly assigned to one diet. Cattle were sampled using rectoanal mucosal swabs in cohorts of 298-337 cattle per day, for a total of 3 sampling days (15-16 days apart). Escherichia coli O157:H7 prevalence rates ranged from not detected (0/75) to 10.0% (8/80) depending on sampling day. Tests for the silage × corn interaction, and the main effects of silage and corn, were not significant (p > 0.05); however, EC reduced the odds of Escherichia coli O157:H7 prevalence by 43% compared to the control corn diet (p = 0.07). Diets containing EC tended to decrease Escherichia coli O157:H7 prevalence in feedlot cattle; however, this reduction was not significant. Before a conclusion can be drawn about impact of EC on Escherichia coli O157:H7 in cattle, further research is necessary to (1) determine if this tendency is due to increased alpha amylase activity and (2) elucidate impact on Escherichia coli O157:H7 prevalence and concentration, as well as a possible mechanism of action.

Commensal Escherichia coli Strains of Bovine Origin Competitively Mitigated Escherichia coli O157:H7 in a Gnotobiotic Murine Intestinal Colonization Model with or without Physiological Stress.

Cattle are a primary reservoir of enterohemorrhagic Escherichia coli (EHEC) O157:H7. Currently, there are no effective methods of eliminating this important zoonotic pathogen from cattle, and colonization resistance in relation to EHEC O157:H7 in cattle is poorly understood. We developed a gnotobiotic EHEC O157:H7 murine model to examine aspects of the cattle pathogen-microbiota interaction, and to investigate competitive suppression of EHEC O157:H7 by 18 phylogenetically distinct commensal E. coli strains of bovine origin. As stress has been suggested to influence enteric colonization by EHEC O157:H7 in cattle, corticosterone administration (±) to incite a physiological stress response was included as an experimental variable. Colonization of the intestinal tract (IT) of mice by the bovine EHEC O157:H7 strain, FRIK-2001, mimicked characteristics of bovine IT colonization. In this regard, FRIK-2001 successfully colonized the IT and temporally incited minimal impacts on the host relative to other EHEC O157:H7 strains, including on the renal metabolome. The presence of the commensal E. coli strains decreased EHEC O157:H7 densities in the cecum, proximal colon, and distal colon. Moreover, histopathologic changes and inflammation markers were reduced in the distal colon of mice inoculated with commensal E. coli strains (both propagated separately and communally). Although stress induction affected the behavior of mice, it did not influence EHEC O157:H7 densities or disease. These findings support the use of a gnotobiotic murine model of enteric bovine EHEC O157:H7 colonization to better understand pathogen-host-microbiota interactions toward the development of effective on-farm mitigations for EHEC O157:H7 in cattle, including the identification of bacteria capable of competitively colonizing the IT.

Prevalence, virulence genes, and antimicrobial profiles of Escherichia coli O157:H7 isolated from healthy cattle in Tunisia.

Shiga toxin-producing Escherichia coli (STEC) O157:H7 is associated with intestinal infection in humans and is considered an important cause of food-borne diseases. The aim of the study was to assess the incidence of E. coli O157:H7 in fecal samples of healthy cattle collected in slaughterhouses (n = 160) and from five farms (n = 100). E. coli isolates were detected on MacConkey agar. A total of 236 E. coli isolates were recovered from fecal samples of healthy cattle. We used sorbitol MacConkey medium to detect non-sorbitol fermenting colonies. These bacteria were examined for the presence of O157:H7 antigen by latex agglutination. The isolation of E. coli O157:H7 has been confirmed with PCR amplification of rfbEO157 and fliCH7 specific genes for serogroup O157 and with multiplex PCR of stx1, stx2, eaeA, and ehxA. All isolates were examined for their susceptibility to 21 antibiotics by the disc diffusion method. Of the 236 E. coli isolates, 4.2% (10/236) were positive for STEC O157:H7. Shiga toxin gene (stx2) and ehxA were present in 70% of isolates, stx1 and eae were confirmed in 60% of the isolates. Other virulence factors screened (fimH, sfa/focDE, cdt3, traT, iutA, and hlyA) were present among the 10 isolates. All E. coli O157:H7 isolates were sensitive to amoxicillin/clavulanic acid, cefotaxime, cefepime, aztreonam, colistin, and sulfamethoxazole/trimethoprim. All isolates belong to the phylo-group E. This is the first study of the incidence of E. coli O157:H7 in cattle in Tunisia. Our finding proves the existence of STEC O157:H7 in healthy animals producing food for human consumption which could be a source of food-borne disease.

Enteric Escherichia coli O157:H7 in Cattle, and the Use of Mice as a Model to Elucidate Key Aspects of the Host-Pathogen-Microbiota Interaction: A Review.

Enterohemorrhagic Escherichia coli (EHEC) serotype O157:H7 is responsible for foodborne disease outbreaks, typically associated with the consumption of undercooked foods contaminated with cattle manure containing the bacterium. At present, effective mitigations do not exist. Many of the factors regulating enteric colonization by E. coli O157:H7 in cattle, and how cattle respond to the bacterium are unknown. In this regard, intestinal colonization locations, shedding patterns, interactions with the enteric microbiota, and host immune responses to infection are current knowledge gaps. As disturbances to host homeostasis are believed to play an important role in the enteric survival of the bacterium, it is important to consider the potential importance of stress during cattle production. Husbandry logistics, cost, and the high genetic, physiological, and microbial heterogeneity in cattle has greatly hampered the ability of researchers to elucidate key aspects of the host-pathogen-microbiota interaction. Although mice have not been extensively used as a cattle model, the utilization of murine models has the potential to identify mechanisms to facilitate hypothesis formulation and efficacy testing in cattle. Murine models have been effectively used to mechanistically examine colonization of the intestine, host responses to infection, and to interactively ascertain how host physiological status (e.g., due to physiological stress) and the enteric microbiota influences colonization and disease. In addition to reviewing the relevant literature on intestinal colonization and pathogenesis, including existing knowledge gaps, the review provides information on how murine models can be used to elucidate mechanisms toward the development of rationale-based mitigations for E. coli O157:H7 in cattle.

Escherichia coli 0157:H7 virulence factors and the ruminant reservoir.

This review updates recent findings about Escherichia coli O157:H7 virulence factors and its bovine reservoir. This Shiga toxin (Stx)-producing E. coli belongs to the Enterohemorrhagic E. coli (EHEC) pathotype causing hemorrhagic colitis. Its low infectious dose makes it an efficient, severe, foodborne pathogen. Although EHEC remains in the intestine, Stx can translocate systemically and is cytotoxic to microvascular endothelial cells, especially in the kidney and brain. Disease can progress to life-threatening hemolytic uremic syndrome (HUS) with hemolytic anemia, acute kidney failure, and thrombocytopenia. Young children, the immunocompromised, and the elderly are at the highest risk for HUS. Healthy ruminants are the major reservoir of EHEC and cattle are the primary source of human exposure. Advances in understanding E. coli O157:H7 pathogenesis include molecular mechanisms of virulence, bacterial adherence, type three secretion effectors, intestinal microbiome, inflammation, and reservoir maintenance. Many aspects of E. coli O157:H7 disease remain unclear and include the role of the human and bovine intestinal microbiomes in infection. Therapeutic strategies involve controlling inflammatory responses and/or intestinal barrier function. Finally, elimination/reduction of E. coli O157:H7 in cattle using CRISPR-engineered conjugative bacterial plasmids and/or on-farm management likely hold solutions to reduce infections and increase food safety/security.

Repeated Oral Vaccination of Cattle with Shiga Toxin-Negative Escherichia coli O157:H7 Reduces Carriage of Wild-Type E. coli O157:H7 after Challenge.

Subcutaneous vaccination of cattle for enterohemorrhagic Escherichia coli O157:H7 reduces the magnitude and duration of fecal shedding, but the often-required, repeated cattle restraint can increase costs, deterring adoption by producers. In contrast, live oral vaccines may be repeatedly administered in feed, without animal restraint. We investigated whether oral immunization with live stx-negative LEE+E. coli O157:H7 reduced rectoanal junction (RAJ) colonization by wild-type (WT) E. coli O157:H7 strains after challenge. Two groups of cattle were orally dosed twice weekly for 6 weeks with 3 × 109 CFU of a pool of three stx-negative LEE+E. coli O157:H7 strains (vaccine group) or three stx-negative LEE- non-O157:H7 E. coli strains (control group). Three weeks following the final oral dose, animals in both groups were orally challenged with a cocktail of four stx+ LEE+E. coli O157:H7 WT strains. Subsequently, WT strains at the RAJ were enumerated weekly for 4 weeks. Serum antibodies against type III secretion protein (TTSP), the translocated intimin receptor (Tir), and EspA were determined by enzyme-linked immunosorbent assay (ELISA) at day 0 (preimmunization), day 61 (postimmunization, prechallenge), and day 89 (postchallenge). Vaccine group cattle had lower numbers of WT strains at the RAJ than control group cattle on postchallenge days 3 and 7 (P ≤ 0.05). Also, vaccine group cattle shed WT strains for a shorter duration than control group cattle. All cattle seroconverted to TTSP, Tir, and EspA, either following immunization (vaccine group) or following challenge (control group). Increased antibody titers against Tir and TTSP postimmunization were associated with decreased numbers of WT E. coli O157:H7 organisms at the RAJ.IMPORTANCE The bacterium E. coli O157:H7 causes foodborne disease in humans that can lead to bloody diarrhea, kidney failure, vascular damage, and death. Healthy cattle are the main source of this human pathogen. Reducing E. coli O157:H7 in cattle will reduce human disease. Using a randomized comparison, a bovine vaccine to reduce carriage of the human pathogen was tested. A detoxified E. coli O157:H7 strain, missing genes that cause disease, was fed to cattle as an oral vaccine to reduce carriage of pathogenic E. coli O157:H7. After vaccination, the cattle were challenged with disease-causing E. coli O157:H7. The vaccinated cattle had decreased E. coli O157:H7 during the first 7 days postchallenge and shed the bacteria for a shorter duration than the nonvaccinated control cattle. The results support optimization of the approach to cattle vaccination that would reduce human disease.

Prevalence and Concentration of Escherichia coli O157:H7 in Cattle, Products, and the Environment in the United States of America: A Meta-Analysis Study

Shiga toxin-producing Escherichia coli O157:H7 are bacterial pathogens that cause foodborne infections in humans. The objectives of this study were to find the pooled prevalence and concentration of Escherichia coli O157:H7 in cattle, hides, carcass, and the environment in the United States of America using meta-analysis. The PRISMA and MOOSE research protocols were employed in the methodology. Weighted effect size was calculated using MetaXL software. A total of 1737 publications were screened, out of which 53 were selected for the final analysis. The pooled prevalence in feedlot cattle was 10.96% (95% CI: 4.2-18.8%). In dairy cattle a pooled prevalence of 1.5% (95% CI: 0.11-3.5%) was observed. The prevalence between feedlot and dairy cattle was significantly different (p<0.05). The herd prevalence in combined feedlot and dairy cattle was 31.7% (95% CI: 10.2-55.5%). Hide and carcass samples’ pooled prevalences were 54.7% (95% CI: 41.7-67.5%) and 21.3% (95% CI: 9.7-34.2%), respectively. Prevalence of environmental samples was 8.1% for produce (95% CI: 0-29.6%), 4.6% for watershed and sediment samples (95% CI: 0-12.2%), and 2.4% for water taken from troughs (95% CI: 0.39-5.1%). Significant difference was observed in individual, herd, and environment prevalence between regions (χ2 =903.14, p=0.0000; χ2 =11.06, p=0.0039; χ2 =13.59, p=0.0004, respectively). E. coli O157:H7 concentrations were highest in feces (900- 300,000 cfu/g), followed by hides (5-9,800 cfu/100 square cm), and carcass (1-189 cfu/100 square cm). At least one supershedder exists in a herd. The findings in this study showed that Escherichia coli O157:H7 serotype is widespread in feedlots, herds, hides, and carcass in the United States of America necessitating appropriate measures to prevent human illnesses. Improving management programs in cattle herds, reduction of environmental contamination, and hygienic slaughter practices are targets of intervention.

Scalable Bayesian Inference for Coupled Hidden Markov and Semi-Markov Models

Bayesian inference for coupled hidden Markov models frequently relies on data augmentation techniques for imputation of the hidden state processes. Considerable progress has been made on developing such techniques, mainly using Markov chain Monte Carlo (MCMC) methods. However, as the dimensionality and complexity of the hidden processes increase some of these methods become inefficient, either because they produce MCMC chains with high autocorrelation or because they become computationally intractable. Motivated by this fact we developed a novel MCMC algorithm, which is a modification of the forward filtering backward sampling algorithm, that achieves a good balance between computation and mixing properties, and thus can be used to analyze models with large numbers of hidden chains. Even though our approach is developed under the assumption of a Markovian model, we show how this assumption can be relaxed leading to minor modifications in the algorithm. Our approach is particularly well suited to epidemic models, where the hidden Markov chains represent the infection status of an individual through time. The performance of our method is assessed on simulated data on epidemic models for the spread of Escherichia coli O157:H7 in cattle. Supplementary materials for this article are available online.

Long-term survival of Escherichia coli O157:H7 and Salmonella Typhimurium in cowpats on pasture.

To investigate the survival of Escherichia coli O157:H7 and Salmonella Typhimurium in cowpats on pasture in a temperate Nordic climate. The study consists of two parts, the first part using artificially created cowpats inoculated with E.coli O157:H7 and S. Typhimurium and the second part using cowpats from empty pastures on which cattle herds positive for E.coli O157:H7 had grazed 6month previously. Artificial cowpats were created, placed in an outdoor field station in June, August and October, and sampled over 1year. Escherichia coli O157:H7 and S. Typhimurium were analysed by standard culture methods. The results showed viable E.coli O157:H7 and S. Typhimurium in the sampled cowpats throughout the 365-day sample period for the June trial, 250days for the August trial and 40-70days for the October trial. In addition, 200 natural cowpats were sampled from eight pastures that had previously held E.coli O157:H7 positive cattle herds. Five positive E.coli O157:H7 isolates were obtained, all with the same multi-locus variable number tandem repeat analysis pattern as had been found on the pasture the previous grazing season. Escherichia coli O157:H7 and S. Typhimurium can survive in cowpats up to a year and persist throughout a winter season. Therefore, there is a possibility that cowpats can act as a reservoir and be a source of re-infection of Salmonella or E.coli O157:H7 in cattle between grazing seasons. The obtained results can provide valuable information for managing the risk posed by zoonotic pathogens originating from farm environments.

Occurrence of Escherichia coli and Escherichia Coli O157:H7 in Cattle, Farm Environment, Milk and Beef

Occurrence of Escherichia coli and Escherichia Coli O157:H7 in Cattle, Farm Environment, Milk and Beef

Molecular detection and quantification of viable probiotic strains in animal feedstuffs using the commercial direct fed microbial Lactobacillus animalis NP51 as a model

Lactobacillus animalis NP51 is a direct-fed microbial strain (DFM) extensively used as a pre-harvest food safety mitigation in feedlot cattle due to its antagonistic effects against human foodborne pathogens such as Salmonella and Escherichia coli O157:H7. NP51 not only promotes overall gut health but interferes with the ability of these pathogens to colonize the gastrointestinal tract of cattle. As a result, NP51 reduces fecal shedding of Salmonella and E. coli O157:H7 in cattle presented for harvest and the load of these pathogens that enter the human food chain. Cattle are administered a high dose (1 × 109 CFU/head/day) of NP51 to reduce fecal shedding of foodborne pathogens. Ensiled animal feedstuffs naturally contain a high load of lactic acid bacteria (LAB) and it is not possible to detect and quantify the level of a specific LAB strain (e.g., NP51) in this matrix using traditional microbiological culture. The purpose of this study was to develop a molecular method to detect and quantify viable populations of a specific LAB strain (e.g., NP51) in cattle feedstuffs. The NP51 whole genome sequence was aligned with closely related LAB clustering within the same well-supported clade in a LAB phylogeny derived from 30 conserved amino acid encoding sequence to identify orthologs. A sequence encoding recombinational DNA repair protein RecT was found to be unique to NP51 and used to design primers and a probe for molecular detection and quantification of NP51. The primers and probe were confirmed to be specific to NP51 in vitro. Total RNA was extracted from silage samples, including samples naturally inoculated in the field and control samples that were artificially spiked with a range of NP51 concentrations in the laboratory. Reverse-transcriptase quantitative real-time (RT-qRTi) PCR was used to quantify cDNA copies in samples and cycle threshold (Ct) values were compared to a standard curve to estimate NP51 concentrations. Our results indicate this novel molecular method is suitable to confirm the presence and estimate the concentration of a specific LAB strain in animal feedstuffs containing high background levels of LAB.

Corn-Based Distillers' Grains in Diets for Feedlot Cattle Are Associated with the Burden of Escherichia coli O157 in Feces.

Inclusion of distillers' grains (DGs) has been associated with increased prevalence of Escherichia coli O157:H7 in cattle housed in research settings. Our objective was to quantify the relationship between inclusion of DGs in commercial feedlot rations and the burden of E. coli O157. A convenience sample of 10 feedlots was enrolled based on DG use in finishing diets; 1 cohort included 5 feedlots in which DGs were greater than 15% of the dietary dry matter and the other cohort consisted of 5 feedlots at a concentration less than 8%. Sampling occurred at each feedlot on four occasions at ∼6-week intervals. At each feedlot visit, 4 pens of cattle within 3 weeks of slaughter were selected and 24 freshly voided fecal pats were sampled. Ten-gram samples were enriched in 90 mL of modified tryptic soy broth with novobiocin (20 mg/L) for 14 h at 42°C. Enrichments were subjected to immunomagnetic separation, plating onto chromogenic agar with novobiocin (5 mg/L) and potassium tellurite (2.5 mg/L), incubation for 18 h at 37°C, and latex agglutination of morphologically typical colonies. E. coli O157 was recovered from 16.7% of 3840 samples. Adjusted prevalence was 14.3% after controlling for within-feedlot and within-pen clustering. Prevalence during each sampling period was 19.9% (round 1), 21.0% (round 2), 14.1% (round 3), and 11.7% (round 4). Prevalence varied between cohorts, but this difference varied over time (p = 0.06). Among those with greater than 15% of the diet as DGs, prevalence was greater than those with less than 8% inclusion for all rounds of sampling (p < 0.01). Averaged across time, prevalence was 23.9% and 9.4% for those with greater than 15% and those with less than 8% of DGs, respectively. While observational, these data provide real-world support of reports of increased E. coli O157:H7 burden associated with DG use in cattle diets.

The prevalence of Escherichia coli O157:H7 fecal shedding in feedlot pens is affected by the water-to-cattle ratio: A randomized controlled trial.

Escherichia coli O157:H7 fecal shedding in feedlot cattle is common and is a public health concern due to the risk of foodborne transmission that can result in severe, or even fatal, disease in people. Despite a large body of research, few practical and cost-effective farm-level interventions have been identified. In this study, a randomized controlled trial was conducted to assess the effect of reducing the level of water in automatically refilling water-troughs on fecal shedding of E. coli O157:H7 in feedlot cattle. Pens in a feedlot in the Texas Panhandle were randomly allocated as control (total number: 17) or intervention (total number: 18) pens. Fecal samples (2,759 in total) were collected both at baseline and three weeks after the intervention, and tested for the presence of E. coli O157:H7 using immunomagnetic bead separation and selective culture. There was a strong statistical association between sampling date and the likelihood of a fecal sample testing positive for E. coli O157:H7. Pen was also a strong predictor of fecal prevalence. Despite accounting for this high level of clustering, a statistically significant association between reduced water levels in the trough and increased prevalence of E. coli O157:H7 in the feces was observed (Odds Ratio = 1.6; 95% Confidence Interval: 1.2–2.0; Likelihood Ratio Test: p = 0.02). This is the first time that such an association has been reported, and suggests that increasing water-trough levels may be effective in reducing shedding of E. coli O157:H7 in cattle feces, although further work would be needed to test this hypothesis. Controlling E. coli O157:H7 fecal shedding at the pre-harvest level may lead to a reduced burden of human foodborne illness attributed to this pathogen in beef.

Interactions of the Hindgut Mucosa-Associated Microbiome with Its Host Regulate Shedding of Escherichia coli O157:H7 by Cattle.

Cattle are the primary carrier of Escherichia coli O157:H7, a foodborne human pathogen, and those shedding >104 CFU/gram of feces of E. coli O157:H7 are defined as supershedders (SS). This study investigated the rectoanal junction (RAJ) mucosa-associated microbiota and its relationship with host gene expression in SS and in cattle from which E. coli O157:H7 was not detected (nonshedders [NS]), aiming to elucidate the mechanisms involved in supershedding. In total, 14 phyla, 66 families, and 101 genera of RAJ mucosa-associated bacteria were identified and Firmicutes (61.5 ± 7.5%), Bacteroidetes (27.9 ± 6.4%), and Proteobacteria (5.5 ± 2.1%) were the predominant phyla. Differential abundance analysis of operational taxonomic units (OTUs) identified 2 OTUs unique to SS which were members of Bacteroides and Clostridium and 7 OTUs unique to NS which were members of Coprococcus, Prevotella, Clostridium, and Paludibacter Differential abundance analysis of predicted microbial functions (using PICRUSt [phylogenetic investigation of communities by reconstruction of unobserved states]) revealed that 3 pathways had higher abundance (log2 fold change, 0.10 to 0.23) whereas 12 pathways had lower abundance (log2 fold change, -0.36 to -0.20) in SS. In addition, we identified significant correlations between expression of 19 differentially expressed genes and the relative abundance of predicted microbial functions, including nucleic acid polymerization and carbohydrate and amino acid metabolism. Our findings suggest that differences in RAJ microbiota at both the compositional and functional levels may be associated with E. coli O157:H7 supershedding and that certain microbial groups and microbial functions may influence RAJ physiology of SS by affecting host gene expression.IMPORTANCE Cattle with fecal E. coli O157:H7 at >104 CFU per gram of feces have been defined as the supershedders, and they are responsible for the most of the E. coli O157:H7 spread into farm environment. Currently, no method is available for beef producers to eliminate shedding of E. coli O157:H7 in cattle, and the lack of information about the mechanisms of supershedding greatly impedes the development of effective methods. This study investigated the role of the rectoanal junction (RAJ) mucosa-associated microbiome in E. coli O157:H7 shedding, and our results indicated that the compositions and functions of RAJ microbiota differed between supershedders and nonshedders. The identified relationship between the differentially abundant microbes and 19 previously identified differentially expressed genes suggests the role of host-microbial interactions involved in E. coli O157:H7 supershedding. Our findings provide a fundamental understanding of the supershedding phenomenon which is essential for the development of strategies, such as the use of directly fed microbials, to reduce E. coli O157:H7 shedding in cattle.

Risk assessment modelling of fecal shedding caused by extended-spectrum cephalosporin-resistant Escherichia coli transmitted through waste milk fed to dairy pre-weaned calves

Waste milk feeding is a common practice in dairy operations. Regardless of the benefits of this practice to the dairy farmers, concerns from the potential dissemination of antimicrobial-resistant bacteria through the gut and subsequent shedding by calves into the environment are increasing. In this study, we employed Monte Carlo simulation to assess the risk of shedding extended-spectrum cephalosporin-resistant Escherichia coli (ESC-R E. coli) caused by waste milk feeding in pre-weaned calves using an exponential dose-response model fit to data for E. coli O157:H7 in cattle. Data from pertinent studies were included in our model to predict the risk of shedding. The median (5th and 95th percentiles) for the daily risk of shedding ESC-R E. coli by calves fed only contaminated waste milk was predicted to be 2.9 × 10-3 (2.1 × 10-3, 3.7 × 10-3), representing a median daily risk of 29 out of 10,000 calves shedding ESC-R E. coli due to exclusive feeding of waste milk containing ESC-R E. coli. This median value was reduced by 94% when accounting for the proportion of waste milk that does not contain ESC-R E. coli. The overall risk of shedding ESC-R E. coli through the pre-weaning period for farms that feed waste milk to calves was 5.7 × 10-3 (2.4 × 10-3, 1.1 × 10-2), representing 57 out of 10,000 calves. When accounting for the proportion of farms that do not feed waste milk, the pre-weaning period risk was reduced by 23%. By varying the prevalence of ESC-R E. coli in waste milk using values of 3, 1.5, and 1%, the daily risk of shedding decreased by factors of 50, 65, and 82%, respectively, which supports the reduction of contamination or discontinuation of feeding waste milk containing ESC-R E. coli as major mitigation measures to reduce the risk of shedding caused by ingestion of resistant bacteria. It is anticipated that the effects of antimicrobial residues in waste milk, which was not considered herein due to lack of data, would further increase risks. Although waste milk feeding to calves may be economically beneficial to the dairy farmers, there exists the risk of dissemination of ESC-resistant bacteria into the environment.

Persistence of Escherichia coli O157:H7 and Total Escherichia coli in Feces and Feedlot Surface Manure from Cattle Fed Diets with and without Corn or Sorghum Wet Distillers Grains with Solubles

Feeding corn wet distillers grains with solubles (WDGS) to cattle can increase the load of Escherichia coli O157:H7 in feces and on hides, but the mechanisms are not fully understood. The objective of these experiments was to examine a role for the persistence of E. coli O157:H7 in the feces and feedlot pen surfaces of cattle fed WDGS. In the first study, feces from steers fed 0, 20, 40, or 60% corn WDGS were inoculated with E. coli O157:H7. The E. coli O157:H7 numbers in feces from cattle fed 0% corn WDGS rapidly decreased (P < 0.05), from 6.28 to 2.48 log CFU/g of feces by day 14. In contrast, the E. coli O157:H7 numbers in feces from cattle fed 20, 40, and 60% corn WDGS were 4.21, 5.59, and 6.13 log CFU/g of feces, respectively, on day 14. A second study evaluated the survival of E. coli O157:H7 in feces from cattle fed 0 and 40% corn WDGS. Feces were collected before and 28 days after the dietary corn was switched from high-moisture corn to dry-rolled corn. Within dietary corn source, the pathogen persisted at higher concentrations (P < 0.05) in 40% corn WDGS feces at day 7 than in 0% WDGS. For 40% corn WDGS feces, E. coli O157:H7 persisted at higher concentrations (P < 0.05) at day 7 in feces from cattle fed high-moisture corn (5.36 log CFU/g) than from those fed dry-rolled corn (4.27 log CFU/g). The percentage of WDGS had no effect on the E. coli O157:H7 counts in feces from cattle fed steam-flaked corn-based diets containing 0, 15, and 30% sorghum WDGS. Greater persistence of E. coli O157:H7 on the pen surfaces of animals fed corn WDGS was not demonstrated, although these pens had a higher prevalence of the pathogen in the feedlot surface manure after the cattle were removed. Both or either the greater persistence and higher numbers of E. coli O157:H7 in the environment of cattle fed WDGS may play a part in the increased prevalence of E. coli O157:H7 in cattle by increasing the transmission risk.

Phenotypic identification of Escherichia coli O157:H7 isolates from cattle at Suleja Abattoir, Nigeria

Escherichia coli O157:H7 is a well-known pathogen of man and animals and a very low infection dose is needed to propagate the infection and clinical disease. In this study, a total of 515 rectal swab samples were collected from cattle and subjected to conventional biochemical tests. Presumptive identification on Eosine Methylene Blue (EMB) yielded an overall prevalence of 83.1%. Cefixime, Tallurite, Sorbitol MacConkey Agar (CT-SMAC) test yielded 18(4.2) isolates while Indole test, Methyl Red, Voges Proskauer and Citrate utilization test (IMVIC) biochemical test showed prevalence rate of 11(61.1%) and MicrogenTM test performed on the 18 Isolates yielded a prevalence rate of 33.3%. A total of 6 of the isolates were subjected to latex agglutination test, in which 2(0.4%) were confirmed to be E. coli O157. The results of the somatic flagella antigen test performed on the 2 isolates revealed that 1(50%) belonged to E. coli O157:H7. Thus this study is therefore the first research work to confirm the presence of E. coli O157:H7 in cattle presented for slaughter in Suleja abattoir in Nigeria. Humans can contract the infection through exposure, handling and consumption of beef or animal products. Control measures are therefore necessary especially during processing and evisceration of beef, to ensure safety of cattle offal presented to the public for human consumption. Key words: Escherichia coli O157:H7, cattle, Microgen kit, flagella staining, phenotypic identification, cattle, latex agglutination test.

Frequency and risk-factors analysis of Escherichia coli O157:H7 in Bali-cattle

Cattle are known as the main reservoir of zoonotic agents verocytotoxin-producing Escherichia coli. These bacteria are usually isolated from calves with diarrhea and/or mucus and blood. Tolerance of these agents to the environmental conditions will strengthen of their transmission among livestock. A total of 238 cattle fecal samples from four sub-districts in Badung, Bali were used in this study. Epidemiological data observed include cattle age, sex, cattle rearing system, the source of drinking water, weather, altitude, and type of cage floor, the cleanliness of cage floor, the slope of cage floor, and the level of cattle cleanliness. The study was initiated by culturing of samples onto eosin methylene blue agar, then Gram stained, and tested for indole, methyl-red, voges proskauer, and citrate, Potential E.coli isolates were then cultured onto sorbitol MacConkey agar, and further tested using O157 latex agglutination test and H7 antisera. Molecular identification was performed by analysis of the 16S rRNA gene, and epidemiological data was analyzed using STATA 12.0 software. The results showed, the prevalence of E. coli O157:H7 in cattle at Badung regency was 6.30% (15/238) covering four sub districts i.e. Petang, Abiansemal, Mengwi, and Kuta which their prevalence was 8.62%(5/58), 10%(6/60), 3.33%(2/60), and 3.33(2/60)%, respectively. The analysis of 16S rRNA gene confirmed of isolates as an E. coli O157:H7 strain with 99% similarities. Furthermore, the risk factors analysis showed that the slope of the cage floor has a highly significant effect (P<0.05) to the distribution of infection. Consequently, implementing this factor must be concerned in order to decrease of infection.

Effects of lactoferrin treatment on Escherichia coli O157:H7 rectal colonization in cattle

The terminal rectal mucosa has been identified as the predominant colonization site of Escherichia coli O157:H7 in cattle, thus a possible intervention approach should directly target this colonization site. To determine the effect of lactoferrin on E. coli O157:H7 mucosal colonization at the rectum, five 6-month-old Holstein-Friesian calves were experimentally infected with E. coli O157:H7 and received daily rectal treatment with bovine lactoferrin. Three calves that did not receive the lactoferrin served as control group. The treatment decreased faecal shedding of E. coli O157:H7 and completely eliminated the infection in all animals (n=5) after 19 days administration. The rectal mucosa of all animals (n=5) was cleared from E. coli O157:H7 within 13 days of lactoferrin treatment. To evaluate the local immune responses, three calves treated previously with lactoferrin and three calves of the control group were re-infected when E. coli O157:H7 excretion was no longer detected. The rectal administration of lactoferrin resulted in an EspA- and EspB-specific IgA responses at the rectal mucosa. These mucosal antibodies were not detected in the animals which did not receive the lactoferrin powder. Interestingly, no serum IgA antibodies could be found in animals of the group that received the lactoferrin. These findings emphasize the ability of bovine lactoferrin to clear E. coli O157:H7 colonization in cattle, where lactoferrin may influence the local immune processes against E. coli O157:H7 infection. Thus, bovine lactoferrin treatment could be used in the field to eliminate high-level faecal excretion of E. coli O157:H7 in cattle.

Differing populations of endemic bacteriophages in cattle shedding high and low numbers of Escherichia coli O157:H7 bacteria in feces.

The objectives of this study were to identify endemic bacteriophages (phages) in the feedlot environment and determine relationships of these phages to Escherichia coli O157:H7 from cattle shedding high and low numbers of naturally occurring E. coli O157:H7. Angus crossbred steers were purchased from a southern Alberta (Canada) feedlot where cattle excreting ≥ 10(4) CFU · g(-1) of E. coli O157:H7 in feces at a single time point were identified as supershedders (SS; n = 6), and cattle excreting <10(4) CFU · g(-1) of feces were identified as low shedders (LS; n = 5). Fecal pats or fecal grabs were collected daily from individual cattle for 5 weeks. E. coli O157:H7 in feces was detected by immunomagnetic separation and enumerated by direct plating, and phages were isolated using short- and overnight-enrichment methods. The total prevalence of E. coli O157:H7 isolated from feces was 14.4% and did not differ between LS and SS (P = 0.972). The total prevalence of phages was higher in the LS group (20.9%) than in the SS group (8.3%; P = 0.01). Based on genome size estimated by pulsed-field gel electrophoresis and morphology determined by transmission electron microscopy, T4- and O1-like phages of Myoviridae and T1-like phage of Siphoviridae were isolated. Compared to T1- and O1-like phages, T4-like phages exhibited a broad host range and strong lytic capability when targeting E. coli O157:H7. Moreover, the T4-like phages were more frequently isolated from feces of LS than SS, suggesting that endemic phages may impact the shedding dynamics of E. coli O157:H7 in cattle.