Megan E. Jacob

Effects of Dried Distillers' Grain on Fecal Prevalence and Growth of Escherichia coli O157 in Batch Culture Fermentations from Cattle†

ABSTRACT Distillers’ grains (DG), a by-product of ethanol production, are fed to cattle. Associations between Escherichia coli O157 prevalence and feeding of DG were investigated in feedlot cattle (n = 379) given one of three diets: steam-flaked corn (SFC) and 15% corn silage with 0 or 25% dried distillers’ grains (DDG) or SFC with 5% corn silage and 25% DDG. Ten fecal samples were collected from each pen weekly for 12 weeks to isolate E. coli O157. Cattle fed 25% DDG with 5 or 15% silage had a higher (P = 0.01) prevalence of E. coli O157 than cattle fed a diet without DDG. Batch culture ruminal or fecal microbial fermentations were conducted to evaluate the effect of DDG on E. coli O157 growth. The first study utilized microbial inocula from steers fed SFC or dry-rolled corn with 0 or 25% DDG and included their diet as the substrate. Ruminal microbial fermentations from steers fed DDG had higher E. coli O157 contents than ruminal microbial fermentations from steers fed no DDG (P < 0.05) when no substrate was included. Fecal fermentations showed no DDG effect on E. coli O157 growth. In the second study with DDG as a substrate, ruminal fermentations with 0.5 g DDG had higher (P < 0.01) E. coli O157 concentrations at 24 h than ruminal fermentations with 0, 1, or 2 g DDG. In fecal fermentations, 2 g DDG resulted in a higher concentration (P < 0.05) at 24 h than 0, 0.5, or 1 g DDG. The results indicate that there is a positive association between DDG and E. coli O157 in cattle, and the findings should have important ramifications for food safety.

Dietary interactions and interventions affecting Escherichia coli O157 colonization and shedding in cattle.

Escherichia coli O157 is an important foodborne pathogen affecting human health and the beef cattle industry. Contamination of carcasses at slaughter is correlated to the prevalence of E. coli O157 in cattle feces. Many associations have been made between dietary factors and E. coli O157 prevalence in cattle feces. Preharvest interventions, such as diet management, could reduce the fecal prevalence and diminish the impact of this adulterant. Dietary influences, including grain type and processing method, forage quality, and distillers grains have all been associated with E. coli O157 prevalence. In addition, several plant compounds, including phenolic acids and essential oils, have been proposed as in-feed intervention strategies. The specific mechanisms responsible for increased or decreased E. coli O157 shedding or survival are not known but are often attributed to changes in hindgut ecology induced by diet types. Some interventions may have a direct bacterial effect. Frequently, results of studies are conflicting or not repeatable, which speaks to the complexity of the hindgut ecosystem, variation in animal feed utilization, and variation within feed products. Still, understanding specific mechanisms, driven by diet influences, responsible for E. coli O157 shedding will aid in the development and implementation of better and practical preharvest intervention strategies.

Animal- and Truckload-Level Associations between Escherichia coli O157:H7 in Feces and on Hides at Harvest and Contamination of Preevisceration Beef Carcasses†

Cattle feces and hides contribute to carcass contamination with Escherichia coli O157:H7, ultimately impacting beef safety. Primary objectives of our cross-sectional study were to evaluate associations among fecal, hide, and preevisceration carcass prevalence of E. coli O157:H7 and to assess factors affecting carcass contamination. Fecal, hide, and preevisceration carcass samples were collected from up to 32 cattle on each of 45 truckloads presented to a midwestern U.S. abattoir. Enrichment and selective culture were used to assess fecal, hide, and carcass prevalence, and direct plating was used to identify cattle shedding high levels of E. coli O157:H7 in feces. Fecal, hide, and carcass prevalence of E. coli O157:H7 within truckload were significantly correlated (P < 0.05) with each other. Enriched fecal sample prevalence was 13.8%, and high shedder prevalence was 3.3%; 38.5% of hides and 10.5% of carcasses were positive for E. coli O157:H7. We used logistic regression to assess animal- and truckload-level variables affecting the probability of carcasses testing positive for E. coli O157:H7. All truckload-level predictors significantly affected the probability of an E. coli O157:H7-positive carcass, including presence of a high shedder within the truckload (odds ratio [OR] = 4.0; confidence interval [CI], 1.6 to 10.1), high (>25%) within-truckload fecal prevalence (OR = 19.3; CI, 4.7 to 79.0), and high (>50%) within-truckload hide prevalence (OR = 7.7; CI, 3.1 to 19.6). The only significant animal-level predictor was having a positive hide (OR = 1.6; CI, 1.0 to 2.6). Our results suggest that preharvest interventions for reducing E. coli O157:H7 contamination of carcasses should focus on truckload (cohort)-level and hide mitigation strategies.

Antimicrobial susceptibility of foodborne pathogens in organic or natural production systems: an overview.

Organic and natural food production systems are increasing in popularity, at least partially because consumers perceive that these niche markets provide healthier and safer food products. One major difference between these niche markets and conventional production systems is the use of antimicrobials. Because antimicrobial agents exert selective pressures for antimicrobial resistance, relating antimicrobial susceptibility of foodborne bacteria to niche market production systems is of interest. Other differences between production systems might also influence the susceptibility of foodborne pathogens. The objective of this review is to compare the impact of food animal production systems on the antimicrobial susceptibility of common foodborne bacterial pathogens. Studies comparing the susceptibility of such pathogens were diverse in terms of geographic location, procedures, species of bacteria, and antimicrobials evaluated; thus, it was difficult to draw conclusions. The literature is highly variable in terms of production type and practices and susceptibility associations, although few studies have compared truly organic and conventional practices. When statistical associations were found between production type and minimum inhibitory concentrations or percentage of isolates resistant for a particular pathogen, the isolates from conventionally reared animals/products were more commonly resistant than the comparison group (organic, antibiotic free, etc.). Therefore, further studies are needed to better assess public health consequences of antimicrobial resistance and food animal production systems, specifically organic or natural versus conventional.

Evaluation of feeding dried distiller's grains with solubles and dry-rolled corn on the fecal prevalence of Escherichia coli O157:H7 and Salmonella spp. in cattle.

Escherichia coli O157:H7 and Salmonella are foodborne pathogens that reside in the gut of cattle and are shed in the feces. Previous work indicated a positive association between feeding cattle distillers grains (DG) and an increase in E. coli O157:H7 prevalence. Feeding processed grains also has been shown to affect fecal prevalence of E. coli O157:H7. The objective of this study was to evaluate the effect of feeding DG and dry-rolled corn (DRC), alone or in combination, on fecal prevalence of E. coli O157:H7 and Salmonella in finishing cattle. Cattle were allotted to pens (n = 28), and fed dietary treatments (n = 150 days) structured in a 2 x 2 factorial arrangement; the factors were 0% or 25% dried corn DG with solubles (DDGS) and 0% or 25% DRC in steam-flaked corn-based high-grain diets. Fecal samples were collected from each pen floor before initiating dietary treatments and at least once every 2 weeks after final diets began. Overall prevalence of E. coli O157:H7 in fecal samples was 5.1%. There were no significant effects of DDGS, DRC, or sampling time on E. coli O157:H7 prevalence (p > 0.20). Overall prevalence of Salmonella in pen floor fecal samples was 23.7%, and sampling week affected prevalence (p < 0.01), ranging from < 1% (week 1) to 77.5% (week 17). Salmonella prevalence was not affected by cattle diet, and no work had previously reported an association between either DG or DRC and Salmonella prevalence. Lack of an association between E. coli O157:H7 prevalence and feeding DG or DRC is contrary to previous observations. Further research is needed to understand inconsistencies between studies of E. coli O157:H7 prevalence and potential associations with DG and grain-processing methods.

Effects of feeding elevated concentrations of copper and zinc on the antimicrobial susceptibilities of fecal bacteria in feedlot cattle.

Cattle are fed elevated concentrations of copper and zinc for growth promotion. The potential mechanisms of growth promotional effects of these elements are attributed to their antimicrobial activities, similar to that of antibiotics, in that gut microbial flora are altered to reduce fermentation loss of nutrients and to suppress gut pathogens. Copper and zinc fed at elevated concentrations may select for bacteria that are resistant not only to heavy metals but also to antibiotics. Our objectives were to determine the effects of feeding elevated copper and zinc on the antimicrobial susceptibilities of fecal bacteria in feedlot cattle. Twenty heifers, fed corn-based high-grain diets, were randomly assigned to treatments in a 2 x 2 factorial arrangement with 1X or 10X National Research Council recommended copper and/or zinc. Feces, collected on days 0, 14, and 32, were cultured for commensal bacteria (Escherichia coli and Enterococcus) to determine their susceptibilities to copper, zinc, and antibiotics. Fecal DNA was extracted to detect tcrB gene and quantify erm(B) and tet(M) genes. In E. coli and Enterococcus sp., minimal differences in minimum inhibitory concentrations (MICs) of copper, zinc, and antibiotics were noticed. The mean copper MIC for E. coli increased (p < 0.05) between days 0 and 32 and days 14 and 32. The only treatment effect detected was increased zinc MIC of E. coli isolates (p < 0.01). The tcrB gene was not detected in feces or in enterococcal isolates. Proportions of erm(B) and tet(M) were unaffected by copper or zinc supplementation. However, the proportion of tet(M) increased (p < 0.05) between days 0 and 14. Feeding elevated copper and/or zinc to feedlot cattle had marginal effects on antimicrobial susceptibilities of fecal E. coli and enterococci.

Niche Marketing Production Practices for Beef Cattle in the United States and Prevalence of Foodborne Pathogens

Niche-marketed food products are rapidly gaining market share in todays society. Consumers are willing to pay premium prices for food perceived to be safer, healthier, more nutritious, and better tasting than conventional food. This review outlines typical production practices for niche-market beef production systems in the United States and compares prevalence estimates of foodborne pathogens in animals and produce from conventional and niche-market production systems. The two main niches for food animal production are organic and natural productions. Organic and natural beef productions are becoming increasingly popular and there is high consumer demand. Two major differences between conventional beef production systems and niche-market production systems (natural and organic) are in the use of antimicrobials and growth-promoting hormones. The impacts of these production systems on foodborne pathogens in beef cattle are variable and often data are nonexistent. Studies directly comparing conventional and niche-market production systems for dairy, swine, poultry, and produce have observed that the prevalence of foodborne pathogens was seldom statistically different between production systems, but when differences were observed, prevalence was typically greater for the niche-market production systems than the conventional production system. The published literature suggests that the perception of niche-marketed food products being safer and healthier for consumers with regard to foodborne pathogens may not be justified.

Occurrence of tcrB, a Transferable Copper Resistance Gene, in Fecal Enterococci of Swine

High concentration of copper, fed as copper sulfate, is often used to increase growth rates in swine. Bacteria exposed to copper may acquire resistance, and in Enterococcus faecium and Enterococcus faecalis, a plasmid-borne transferable copper resistance (tcrB) gene that confers copper resistance has been reported. Our objectives were to determine the occurrence of tcrB in fecal enterococci from weaned piglets fed diets with a normal supplemental level (16.5 ppm) or an elevated supplemental level (125 ppm) of copper and to determine the association of tcrB with copper, erythromycin, and vancomycin resistance. A total of 323 enterococcal isolates were examined and 15 (4.6%) isolates (14 E. faecium and 1 E. faecalis) were positive for tcrB. Fifteen tcrB-positive and 15 randomly chosen tcrB-negative isolates from piglets fed the normal supplemental level of copper were tested for erm(B), tet(M), vanA, and vanB genes and susceptibilities to copper, erythromycin, tetracyclines, and vancomycin. All tcrB-positive and -negative isolates contained erm(B) and tet(M), but not vanA and vanB. The mean minimum inhibitory concentration of copper for tcrB-positive (21.1 mM) was higher (p < 0.001) compared with tcrB-negative isolates (6.1 mM). All isolates were resistant to erythromycin and tetracyclines and susceptible to vancomycin. The transferability of the tcrB gene from tcrB-positive strains to tcrB-negative strains was demonstrated by conjugation. The potential link between tcrB and antibiotic resistance genes and the propensity of enterococci to transfer tcrB to other strains raises the possibility that copper supplementation may exert selection pressure for antibiotic-resistant enterococci. This study is the first report on the occurrence of the tcrB gene in enterococci isolated from swine in the United States.

Evaluation of a Multiplex Real-Time Polymerase Chain Reaction for the Quantification of Escherichia coli O157 in Cattle Feces

Cattle are asymptomatic reservoirs for Escherichia coli O157, a major foodborne pathogen. The organism generally colonizes the hindgut of cattle and is shed in the feces at low concentrations. The objective of this research was to evaluate a multiplex, real-time polymerase chain reaction (mqPCR) assay for quantification of E. coli O157 in cattle feces using stx1, stx2, and rfbE gene targets. Primer efficiency and analytical sensitivity of the assay were evaluated with a single or pooled (five strain) culture of E. coli O157. In pure culture, the minimum detection limit of the assay was 1.4×10(3) CFU/mL and 3.6×10(3) CFU/mL for the single and five-strain mixture of E. coli O157, respectively. Diagnostic sensitivity was analyzed using DNA extracted from cattle feces spiked with E. coli O157. In feces spiked with the pooled mixture of five E. coli O157 strains, the minimum detection limit was 3.6×10(4) CFU/g. We also evaluated the assay with feces from cattle experimentally inoculated with E. coli O157 by comparing the results to a culture-based method. For the majority of samples tested, the concentration of E. coli O157 detected by the real-time and culture methods was within one log difference. However, the assay could only be evaluated for cattle shedding high concentrations of E. coli O157. In conclusion, the mqPCR quantifying E. coli O157 in cattle feces using stx1, stx2, and rfbE gene targets may have use in detecting and quantifying super shedders, but is not applicable for quantification in animals shedding low concentrations (10(2) to 10(3) CFU/g feces).

Vaginal Microbiota of Spayed Dogs with or without Recurrent Urinary Tract Infections

Background Limited information is available regarding the vaginal microbiota of normal spayed dogs and spayed dogs with recurrent UTIs. Vaginal lactic acid‐producing bacteria (LAB) have been associated with decreased frequency of recurrent urinary tract infection in women and may have a protective role within the urinary tract of female dogs. Hypothesis/Objectives Spayed dogs with historical recurrent UTI will have decreased prevalence of LAB and increased prevalence of uropathogenic bacterial populations in the vaginal microbiota when compared with the vaginal microbiota of healthy, spayed dogs. Animals Twenty‐one client‐owned adult spayed female dogs with historical recurrent UTI and 23 healthy, spayed female dogs without a history of recurrent UTI. Methods Dogs were placed into a recurrent UTI group or control group in this prospective study. Bacterial populations were isolated and characterized from vaginal swabs obtained from each dog. Results The most common bacterial isolates obtained from the vaginal tract of all dogs were Escherichia coli (11/44) and S. pseudintermedius (13/44). E. coli was isolated from the vaginal tract of 8 of 21 (38%) dogs in the rUTI group and 3 of 23 (13%) dogs in the control group (P = .08). LAB were isolated from 7 of the 44 dogs. Two of these 7 dogs were in the rUTI group and 5 of the 7 dogs were in the control group. Conclusions and Clinical Importance The vaginal microbiota of spayed female dogs with recurrent UTI was similar to the control population of normal, spayed female dogs.