Significance of fecal volatile fatty acids in shedding of Escherichia coli O157 from calves: experimental infection and preliminary use of a probiotic product

Cattle have been recognized as a principal reservoir of Escherichia coli O157:H7. This organism appears to be confined to the gastrointestinal tract and is shed in feces. A probiotic product containing lactic acid-producing Streptococcus bovis LCB6 and Lactobacillus gallinarum LCB 12 isolated from adult cattle was developed, and a preliminary experiment was conducted to evaluate its effect on the elimination of E. coli O157 from experimentally infected calves. Eight 4-month-old Holstein calves were orally challenged with E. coli O157 and the probiotic product was administered against four calves continued fecal shedding of E. coli O157 by the 7th day after infection. Fecal shedding of E. coli O157 was completely inhibited and re-shedding was not detected in any of the animals. Remarkable increase of VFAs, especially that of acetic acid in feces after the administration of probiotic bacteria correlated with the diminution of E. coli O157. Four calves that had spontaneously ceased fecal shedding of E. coli O157 by the 7th day exhibited a high concentration of VFAs in feces before and after experimental infection. Although our results are preliminary and obtained from calves under limited conditions, the possible application of probiotic product to reduce fecal shedding of E. coli O157 from cattle is suggested.     

Effect of antibiotics in milk replacer on fecal shedding of Escherichia coli O157:H7 in calves

The objective of this study was to compare the concentration and duration of fecal shedding of Escherichia coli O157:H7 between calves fed milk replacer with or without antibiotic (oxytetracycline and neomycin) supplementation. Eighteen 1-wk-old Holstein calves were orally inoculated with a strain of E. coli O157:H7 (3.6 x 10(8) cfu/calf) made resistant to nalidixic acid (NA). Rectal samples were obtained three times weekly for 8 wk following oral inoculation. Fecal shedding of NA-resistant E. coli O157:H7 was quantified by direct plating or detected by selective enrichment procedure. Eight weeks after inoculation, calves were killed, necropsied, and tissues (tonsils, retropharyngeal and mesenteric lymph nodes, and Peyer's patches) and gut contents (rumen, omasum, abomasum, ileum, cecum, colon, and rectum) were sampled to quantify or detect NA-resistant E. coli O157:H7. The percentage of calves shedding NA-resistant E. coli O157:H7 in the feces in the antibiotic-fed group was higher (P < 0.001) early in the study period (d 6 and 10) compared with the control group fed no antibiotics. There was no difference between treatment and control groups in the concentration of E. coli O157 in feces that were positive at quantifiable concentrations. A comparison of the duration of fecal shedding between treated and untreated calves showed no significant difference between groups. At necropsy, E. coli O157:H7 was recovered from the rumen and omasum of one calf in the control group and from retropharyngeal lymph node and Peyer's patch of two calves in the antibiotic group. Supplementation of milk replacer with antibiotics may increase the probability of E. coli O157:H7 shedding in dairy calves, but the effect seems to be of low magnitude and short duration.     

Experimental infection of calves with Escherichia coli O157:H7

Three 3-month-old Japanese Black calves were experimentally infected with Escherichia coli O157:H7 to define the magnitude (CFU/g) and duration of fecal shedding of the organism. In two of the three calves, fecal shedding of E. coli O157:H7 ceased in 5 and 9 weeks. The remaining calf continued shedding E. coli O157:H7 for more than 31 weeks, and the magnitude of the shedding ranged from 10(1) to 10(4) CFU/g of feces. The possibility is suggested that a percentage of animals naturally infected with E. coli O157:H7 on farms may become long-term shedders, transmitting the organism to other animals in the herd and to the proximate environment.     

Isolation of Escherichia coli O157:H7 from the gall bladder of inoculated and naturally-infected cattle

To determine if Escherichia coli O157:H7 is capable of residing in the gall bladder of cattle, inoculation studies were conducted with O157:H7 strain 86-24 in weaned Holstein calves. Strain 86-24 was isolated from the gall bladders of five calves 36 days after inoculation. Two other calves contained the inoculation strain in the distal colon but the organism was absent in their gall bladders. A second trial in which the calves were euthanized 15 days after inoculation found strain 86-24 in six of seven inoculated calves but only in colon and/or rumen samples. In a third trial that inoculated eight calves with a four-strain cocktail of O157:H7 strains, the gall bladders from all eight animals were positive 9 days after inoculation. The colon and rumen samples from these calves were also positive. E. coli O157:H7 isolates recovered from bile samples and subtyped by pulsed field gel electrophoresis found that three of the four inoculation strains were present in one or more of the calves. Thus, residence in the gall bladder is not restricted to a single strain. Additional evidence of the ability to localize in the gall bladder of cattle was provided by testing the bile from 150 gall bladders (five collection dates, 30 samples each) obtained at an abbatoir and the isolation of E. coli O157:H7 from four samples (2.7%). This study establishes that E. coli O157:H7 can reside transiently or permanently at a low level in the gall bladder of cattle.     

Isotype-specific antibody responses against Escherichia coli O157:H7 locus of enterocyte effacement proteins in adult beef cattle following experimental infection

Escherichia coli O157:H7 is an important food-borne pathogen and cause of hemorrhagic colitis and hemolytic uremic syndrome in humans. Cattle are an important reservoir of E. coli O157:H7, in which the organism colonizes the intestinal tract and is shed in the feces. Vaccination of cattle has significant potential as a pre-harvest intervention strategy for E. coli O157:H7; however, basic information about the bovine immune responses to important bacterial colonization factors resulting from infection has not been reported. The serum and fecal IgG and IgA antibody responses of adult cattle to E. coli O157:H7 intimin, translocated intimin receptor (Tir), E. coli-secreted proteins (Esp)A, EspB and O157 lipopolysaccharide (LPS) in response to infection were determined. All animals were seropositive for all five antigens prior to inoculation, with antibody titers to EspB and O157 LPS significantly higher (P<0.05) than those to Tir, intimin and EspA. After inoculation, the cattle became colonized and developed significant increases in their serum antibody titers to intimin, Tir, EspB, EspA and O157 LPS (P<0.05); however, by 42 days post-inoculation the titers to all except EspB were on the decline. In contrast, pre- and post-inoculation fecal IgG and IgA antibodies to these same antigens were not detected (<1:5). These results indicate that cattle respond serologically to E. coli O157:H7 type III secreted proteins, intimin and O157 LPS during the course of infection and the response is correlated with the extent of fecal shedding.     

Effect of pooling bovine fecal samples on the sensitivity of detection of E. coli O157:H7

To assess the effect of pooling fecal samples on the sensitivity of detection of E. coli O157:H7, 12 calves, inoculated orally with 10(8)cfu per calf of nalidixic acid resistant E. coli O157:H7, were used to provide positive fecal samples. After inoculation, calves were sampled twice weekly. Negative fecal samples were from calves at a local dairy. Samples from inoculated calves were incubated without pooling or were mixed with known negative fecal samples in a 1:4 ratio or a 2:3 ratio (positive:negative) for detection of E. coli O157:H7. Samples were enriched 6h in Gram negative broth with vancomycin, cefixime, and cefsoludin, underwent immunomagnetic separation with Dynabeads, and were plated onto sorbitol MacConkey agar with cefixime, and tellurite (SMACct). Morphologically typical colonies were plated onto blood agar, incubated overnight at 37 degrees C and an indole test was performed on each colony. Indole positives colonies were plated on SMAC agar with 20 microg/ml nalidixic acid (SMACnal). Colonies that grew on SMACnal were confirmed by O157 agglutination. Sensitivity of detection in non-pooled samples was 77%. Samples pooled 1:4 and 2:3 with negative samples were 55 and 52% sensitive, respectively. Pooling decreased sensitivity of detection for E. coli O157:H7 in bovine fecal samples (P<0.01). A deterministic binomial probability model was developed to assess the probability of detecting pens of cattle shedding E. coli O157 using a pooling protocol or individual samples. Pooling decreased sensitivity of detection at low pen prevalence compared to individual samples but was similar at high prevalence.     

Escherichia coli O157:H7 in the gallbladders of experimentally infected calves.

Fifteen weaned calves (age 89-141 days) were treated with dexamethasone (0.25 mg/kg, IV) for 3 days before, the day of, and the day after inoculation with 10 colony-forming units of either Escherichia coli O157:H7 (strain 86-24, which produces Shiga toxin 2 and intimin; n = 13) or nonpathogenic E. coli (strain 123, which does not produce Shiga toxin or intimin; n = 2). All calves were necropsied 4 days after inoculation. Histologic lesions of attaching and effacing bacteria were observed in the large intestine (12/13) and in the gallbladder mucosa (5/13) of calves inoculated with E. coli 86-24. Cholecystitis was present in 12 of 13 calves that received E. coli 86-24. Inoculum bacteria were recovered from the distal colons or feces (13/13) and gallbladders (3/4) of calves inoculated with 86-24.     

Prevalence of Escherichia coli O157:H7 in white-tailed deer sharing rangeland with cattle.

OBJECTIVE: To determine the prevalence of fecal shedding of Escherichia coli O157:H7 in white-tailed deer (Odocoileus virginianus) with access to cattle pastures. DESIGN: Survey study. SAMPLE POPULATION: 212 fecal samples from free ranging white-tailed deer. PROCEDURE: Fresh feces were collected on multiple pastures from 2 farms in north central Kansas between September 1997 and April 1998. Escherichia coli O157:H7 was identified by bacterial culture and DNA-based methods. RESULTS: Escherichia coli O157:H7 was identified in 2.4% (5/212) of white-tailed deer fecal samples. CONCLUSIONS AND CLINICAL RELEVANCE: There is considerable interest in the beef industry in on-farm control of E coli O157:H7 to reduce the risk of this pathogen entering the human food chain. Results of our study suggest that the design of programs for E coli O157:H7 control in domestic livestock on pasture will need to account for fecal shedding in free-ranging deer. In addition, the results have implications for hunters, people consuming venison, and deer-farming enterprises.     

Escherichia coli O157:H7 and other Shiga toxin-producing E. coli in white veal calves

The aims of the study were to determine the prevalence of enterohemorrhagic Escherichia coli O157:H7 (EHEC O157) and other Shiga toxin-producing E. coli (STEC) in feces of white veal calves in an operation in Ontario, to evaluate exposure of the calves to EHEC O157, and to investigate the milk replacer diet and antimicrobial resistance as factors that might influence the prevalence of EHEC O157. Feces from three cohorts of 20-21 calves were collected weekly for 20 weeks and processed for isolation of EHEC O157:H7 and detection of STEC by an ELISA. Exposure to EHEC O157 was also investigated by measuring IgG and IgM antibodies to the O157 lipopolysaccharide (O157 Ab) in sera by ELISA. The prevalences of EHEC O157 were 0.17% of 1151 fecal samples and 3.2% of 62 calves, and for STEC were 68% of 1005 fecal samples and 100% of 62 calves. Seroconversion to active IgG and IgM O157 Ab responses in some calves was not associated with isolation of EHEC O157. The milk replacer contained low levels of antibodies to EHEC antigens and without antimicrobial drugs, it did not inhibit the growth of EHEC O157 in vitro. Two E. coli O157:H7 that were isolated were totally drug sensitive whereas 60 commensal E. coli isolates that were examined were highly resistant. Antibodies in milk replacer that might be protective in vivo, and susceptibility to antimicrobial agents in the milk replacer may contribute to the low prevalence of EHEC O157 in white veal calves.     

Escherichia coli O157:H7 vaccine field trial in 9 feedlots in Alberta and Saskatchewan

A feedlot trial was conducted to assess the efficacy of an Escherichia coli O157:H7 vaccine in reducing fecal shedding of E. coli O157:H7 in 218 pens of feedlot cattle in 9 feedlots in Alberta and Saskatchewan. Pens of cattle were vaccinated once at arrival processing and again at reimplanting with either the E. coli O157:H7 vaccine or a placebo. The E. coli O157:H7 vaccine included 50 microg of type III secreted proteins. Fecal samples were collected from 30 fresh manure patties within each feedlot pen at arrival processing, revaccination at reimplanting, and within 2 wk of slaughter. The mean pen prevalence of E. coli O157:H7 in feces was 5.0%; ranging in pens from 0% to 90%, and varying significantly (P < 0.001) among feedlots. There was no significant association (P > 0.20) between vaccination and pen prevalence of fecal E. coli O157:H7 following initial vaccination, at reimplanting, or prior to slaughter.     

Intranasal immunisation with Stx2B-Tir-Stx1B-Zot protein leads to decreased shedding in goats after challenge with Escherichia coli O157:H7

Ruminants are an important reservoir of Escherichia coli O157:H7. To reduce E coli O157:H7 excretion by these animals could play a key role in prevention and control of human infections. In the present study, the authors used 12 three-month-old goats to evaluate the efficacy of intranasal administration of the Stx2B-Tir-Stx1B-Zot protein. These goats were inoculated on days 0 and 21 and infected with 10(10) colony-forming units (cfu) of E coli O157:H7 by oral inoculation on day 36. Faecal shedding was monitored daily for two weeks. All of six goats immunised with recombinant protein elicited significant Stx2b-Tir-Stx1b-Zot-specific serum IgG antibodies, and three of them also showed production of antigen-specific IgA in faeces. The immunised goats showed much less shedding of E coli O157:H7 after challenge. These results demonstrate the potential for the use of Stx2B-Tir-Stx1B-Zot protein in mucosal vaccine formulations to prevent colonisation and shedding of E coli O157:H7 in goats.     

Results of a longitudinal study of the prevalence of Escherichia coli O157:H7 on cow-calf farms

OBJECTIVE: To describe the frequency and distribution of Escherichia coli O157:H7 in the feces and environment of cow-calf herds housed on pasture. SAMPLE POPULATION: Fecal and water samples for 10 cow-calf farms in Kansas. PROCEDURE: Fecal and water samples were obtained monthly throughout a 1-year period (3,152 fecal samples from 2,058 cattle; 199 water samples). Escherichia coli O157:H7 in fecal and water samples was determined, using microbial culture. RESULTS: Escherichia coli O157:H7 was detected in 40 of 3,152 (1.3%) fecal samples, and 40 of 2,058 (1.9%) cattle had > or = 1 sample with E coli. Fecal shedding by specific cattle was transient; none of the cattle had E coli in more than 1 sample. Significant differences were not detected in overall prevalence among farms. However, significant differences were detected in prevalence among sample collection dates. Escherichia coli O157:H7 was detected in 3 of 199 (1.5%) water samples. CONCLUSIONS AND CLINICAL RELEVANCE: Implementing control strategies for E coli O157:H7 at all levels of the cattle industry will decrease the risk of this organism entering the human food chain. Devising effective on-farm strategies to control E coli O157:H7 in cow-calf herds will require an understanding of the epidemiologic characteristics of this pathogen.     

Effect of feeding the ionophores monensin and laidlomycin propionate and the antimicrobial bambermycin to sheep experimentally infected with E. coli O157:H7 and Salmonella typhimurium

Escherichia coli O157:H7 and Salmonella are widely recognized as important agents of foodborne disease with worldwide distribution. The use of ionophores in feeding growing ruminants is widespread in the United States and has attracted recent interest due to the apparent temporal relationship between initial ionophore use and the increase in human E. coli O157:H7 cases. Two experiments were conducted to evaluate the effects of short-term feeding of ionophores on fecal shedding, intestinal concentrations, and antimicrobial susceptibility of E. coli O157:H7 and S. typhimurium in growing lambs. Sixteen lambs were used in each experiment, four lambs per treatment group: monensin, laidlomycin propionate, bambermycin, and a control treatment. Lambs were fed a grain and hay (50:50) diet with their respective ionophore for 12 d before experimental inoculation with E. coli O157:H7 or S. typhimurium. Animals were maintained on their respective diets an additional 12 d, and fecal shedding of inoculated pathogens was monitored daily. Lambs were killed and tissues and contents were sampled from the rumen, cecum, and rectum. No differences (P > 0.05) in fecal shedding of Salmonella or E. coli O157:H7 were observed due to treatment. Occurrence of Salmonella or E. coli in luminal contents and tissue samples from the rumen, cecum, and rectum did not differ (P > 0.05) among treatments. Feeding monensin decreased (P < 0.05) the incidence of scours in sheep infected with Salmonella compared with the other treatments. No differences in antimicrobial susceptibility were found in any of Salmonella or E. coli O157:H7 isolates. Results from these studies indicate that short-term ionophore feeding had very limited effects on E. coli and Salmonella shedding or on antimicrobial susceptibility in experimentally infected lambs.     

Dietary monensin level, supplemental urea, and ractopamine on fecal shedding of Escherichia coli O157:H7 in feedlot cattle

Inclusion of distillers grains (DG) in cattle diets has been shown to increase fecal shedding of Escherichia coli O157:H7. It is hypothesized that altered gut fermentation by DG may be responsible for the positive association. Therefore, feed additives affecting ruminal or hindgut fermentation of DG also may affect fecal shedding of E. coli O157:H7. The objectives of the study were to evaluate effects of monensin (33 or 44 mg/kg of DM), supplemental urea (0, 0.35, or 0.70% of DM), and ractopamine (0 or 200 mg/steer daily administered during the last 42 d of finishing) in a steam-flaked corn grain-based diet containing 30% wet sorghum DG on fecal shedding of E. coli O157:H7. Seven hundred twenty crossbred beef steers, housed in 48 pens (15 steers/pen), were assigned to dietary treatments in a randomized complete block design with a 2 × 3 × 2 factorial treatment arrangement. Fresh pen floor fecal samples (10 per/pen) were collected every 2 wk for 14 wk (July through November) and cultured for E. coli O157:H7. Isolation of E. coli O157:H7 was by selective enrichment of fecal samples in an enrichment broth, immunomagnetic separation, followed by plating onto a selective medium. Samples that yielded sorbitol-negative colonies, which were positive for indole production, O157 antigen agglutination, and contained rfbE, fliC, and stx2 were considered positive for E. coli O157:H7. Fecal prevalence data were analyzed as repeated measures using negative binomial regression to examine effects and interactions of sampling day, urea, monensin, and ractopamine. Mean fecal prevalence of E. coli O157:H7 was 7.6% and ranged from 1.6 to 23.6%. Cattle fed monensin at 44 mg/kg of feed had less (P = 0.05) fecal E. coli O157:H7 prevalence than cattle fed 33 mg/kg (4.3 vs. 6.8%). Although the reason for the reduction is not known, it is likely because of changes in the microbial ecosystem induced by the greater amount of monensin in the hindgut. Supplemental urea at 0.35 or 0.70% had no effect (P = 0.87) on fecal shedding of E. coli O157:H7. Fecal prevalence of E. coli O157:H7 were 5.3, 5.7, and 5.9% for groups fed 0, 0.35, and 0.7% urea, respectively. The inclusion of ractopamine at 0 or 200 mg/(animal?d) had no effect (P = 0.89) on fecal prevalence of E. coli O157:H7 (4.4 vs. 4.0%). Additional research is needed to confirm the reduction in fecal shedding of E. coli O157:H7 in cattle fed monensin at 44 mg/kg of feed compared with cattle fed 33 mg/kg of feed.     

Effects of wheat or corn distillers dried grains with solubles on feedlot performance, fecal shedding, and persistence of Escherichia coli O157:H7

Distillers dried grains with solubles (DDGS) are a coproduct of the ethanol industry and are often used as a replacement for grain in livestock production. Feeding corn DDGS to cattle has been linked to increased fecal shedding of Escherichia coli O157:H7, although in Canada, DDGS are often produced from wheat. This study assessed the effects of including 22.5% wheat or corn DDGS (DM basis) into barley-based diets on performance, carcass characteristics, animal health, and fecal E. coli O157:H7 shedding of commercial feedlot cattle. Cattle (n = 6,817) were randomly allocated to 10 pens per treatment group: WDDGS (diets including 22.5% wheat DDGS), CDDGS (diets including 22.5% corn DDGS), or CTRL (barley substituted for DDGS). Freshly voided fecal pats (n = 588) were collected and pooled monthly for fecal pH measurement and screened for naturally occurring E. coli O157:H7 by immunomagnetic separation (IMS) and direct plating (DP). Hide swabs (n = 367) were collected from randomly selected cattle from each pen before slaughter. Pen-floor fecal samples (n = 18) were collected from treatment groups at entry to the feedlot (<14 d on the finishing diet) and after adapting to the finishing diet for ≥14 d, inoculated (10(9) cfu of a 5 strain naldixic acid-resistant E. coli O157:H7 mixture), incubated (20°C) and evaluated weekly (IMS and DP) to assess fecal E. coli O157:H7 persistence. The WDDGS group had 3.0% poorer ADG (P = 0.007), 5.3% poorer G:F (P < 0.001), and a decreased proportion of Canada Quality Grade AAA carcasses (P = 0.022) compared with CTRL cattle. The CDDGS group had a similar ADG (P = 0.06), a decreased proportion of Canada Yield Grade (YG) 1 (P < 0.001), and greater proportions of Canada YG 2 (P = 0.003) and YG 3 (P < 0.001) carcasses compared with the CTRL group. There were no differences among groups in any of the animal health parameters assessed. Inclusion of DDGS in cattle finishing diets had no effect on fecal shedding (P = 0.650) or persistence (P = 0.953) of E. coli O157:H7. However, feces from cattle on starter diets <14 d had longer persistence of E. coli O157:H7 (week) than cattle on finishing diets ≥14 d (P < 0.003). Inclusion of DDGS in feedlot diets depends on commodity pricing relative to that of barley and for WDDGS must also include the risk of feedlot performance and carcass grading disadvantages. Feeding cattle barley based-diets with 22.5% corn or wheat DDGS did not affect fecal shedding of E. coli O157:H7.     

Interaction with avian cells and colonisation of specific pathogen free chicks by Shiga-toxin negative Escherichia coli O157:H7 (NCTC 12900)

The prevalence of Escherichia coli O157:H7 infection in birds is low but several deliberate inoculation studies show that poultry are readily and persistently infected by this organism indicating a possible threat to public health. The mechanisms of colonisation of poultry are not understood and the aim is to establish models to study the interaction of E. coli O157:H7, at the cellular and whole animal levels. A non-toxigenic E. coli O157:H7 (NCTC 12900) was used in adherence assays with an avian epithelial cell line (Div-1) and used to inoculate 1-day-old SPF chicks. In vitro, NCTC 12900 induced micro-colonies associated with cytoskeletal arrangements and pedestal formation with intimate bacterial attachment. In the 1-day-old SPF chick, a dose of 1 x 10(5) cfu resulted in rapid and extensive colonisation of the gastrointestinal tract and transient colonisation of the liver and spleen. The number of E. coli O157:H7 organisms attained approximately 10(8) cfu/ml caecal homogenate 24h after inoculation and approximately 10(7) cfu/ml caecal homogenate was still present at day 92. Faecal shedding persisted for 169 days, ceasing 9 days after the birds came into lay and 6% of eggs were contaminated on the eggshell. Histological analysis of tissue samples from birds dosed with 1x10(7) cfu gave evidence for E. coli O157:H7 NCTC 12900 induced micro-colonies on the caecal mucosa, although evidence for attaching effacing lesions was equivocal. These models may be suitable to study those factors of E. coli O157:H7 that mediate persistent colonisation in avian species.     

Transmission of Escherichia coli O157:H7 to cattle by house flies

The main reservoir of Escherichia coli O157:H7 is the digestive tract of cattle; however, the ecology of this food-borne pathogen is poorly understood. House flies (Musca domestica L.) might play a role in dissemination of this pathogen in the cattle environment. In our study, eight calves were individually exposed to house flies that were orally inoculated with a mixture of four strains of nalidixic acid-resistant E. coli O157:H7 (Nal(R)EcO157) for 48h. Another eight calves were individually exposed to uninoculated flies and served as the control. Fresh cattle feces (rectal sampling) and drinking water were periodically sampled and screened for Nal(R)EcO157 up to 19 days after the exposure. At the end of the experiment, all calves were euthanized and the lumen contents of rumen, cecum, colon, and rectum as well as swab samples of gall-bladder mucosa and the recto-anal mucosa were screened for Nal(R)EcO157. On day 1 after the exposure, fecal samples of all eight calves and drinking-water samples of five of eight calves exposed to inoculated flies tested positive for Nal(R)EcO157. The concentration of Nal(R)EcO157 in feces ranged over time from detectable only by enrichment (<10(2)) to up to 1.1 x 10(6)CFU/g. Feces of all calves remained positive for Nal(R)EcO157 up to 11 days after the exposure and 62% were positive until the end of experiment. Contamination of drinking water was more variable and all samples were negative on day 19. At necropsy, the highest prevalence of Nal(R)EcO157 was in the recto-anal mucosa region, followed by rectal and colonic contents.     

Influence of dietary endophyte (Neotyphodium coenophialum)-infected tall fescue (Festuca arundinacea) seed on fecal shedding of antibiotic resistance-selected Escherichia coli O157:H7 in ewes

The objectives were to determine the effects of short-term feeding of a toxic endophyte (Neotyphodium coenophialum)-infected tall fescue seed (Festuca arundinacea, cultivar 'Kentucky 31') on fecal shedding and intestinal concentrations of Escherichia coli O157:H7 and the concentrations of prolactin, cortisol, and NEFA in experimentally inoculated ewes. Twelve ewes (mean BW = 46 +/- 2 kg) were fed a diet containing either high endophyte-infected (HI-E) or low endophyte-infected (LO-E) tall fescue seed for 7 d. Each diet consisted of 50% (as-fed basis) tall fescue seed. Ewes were experimentally inoculated with antibiotic resistance-selected E. coli O157:H7 on d 1 of the feeding treatment, and fecal shedding of inoculated pathogens was monitored daily on d 2 to 6. On d 7, ewes were weighed and euthanized, and tissues and contents were sampled from the ileum, cecum, and rectum for quantitative enumeration of E. coli O157:H7. Urine was collected at euthanization to determine total ergot alkaloid concentrations. Ewes fed HI-E had lower (P < 0.001) DMI than did ewes fed LO-E (0.8 and 1.6 +/- 0.1 kg/d of DMI for HI-E and LO-E ewes, respectively); consequently, there was a tendency (P = 0.06) for HI-E ewes to lose 0.3 +/- 0.4 kg of BW/d and LO-E ewes to gain 0.2 +/- 0.4 kg of BW/d during the 7 d. Urinary ergot alkaloids were increased (P < 0.001) in ewes fed HI-E (47.8 +/- 9.4 ng/mg of creatinine) compared with those fed LO-E (6.2 +/- 9.4 ng/mg of creatinine). Prolactin tended (P = 0.06) to be decreased in ewes fed HI-E (7.2 +/- 7.0 ng/mL) compared with those fed LO-E (27.7 +/- 7.0 ng/mL). Fecal shedding of E. coli O157:H7 tended (P = 0.06) to be increased in HI-E ewes [5.4 cfu (log10)/g of feces] compared with LO-E ewes [4.5 cfu (log10)/g of feces]. The population of E. coli O157:H7 in luminal contents from the ileum, cecum, and rectum did not differ (P > 0.36) between treatments. Treatment did not influence (P = 0.30) the occurrence of E. coli O157:H7 in cecal or rectal tissues; however, ileal tissues from HI-E ewes tended (P = 0.12) to have an increased incidence of E. coli O157:H7. Concentrations of NEFA tended (P = 0.12) to be greater in HI-E ewes than in LO-E ewes, whereas cortisol was similar (P = 0.49) for HI-E and LO-E ewes. We conclude that short-term feeding of HI-E tall fescue seed may alter the concentrations of prolactin and NEFA, and may increase fecal shedding of E. coli O157:H7 in experimentally inoculated ewes.     

Prevalence of Shiga-toxigenic Escherichia coli O157:H7 in adult dairy cattle

OBJECTIVE: To describe shiga-toxigenic Escherichia coil O157:H7 (STEC O157:H7) fecal shedding prevalence, seasonal fecal shedding patterns, and site-specific prevalence from the oral cavity, skin, and feces of dairy cattle. DESIGN: Cross-sectional study. ANIMALS: Adult dairy cattle from 13 herds in Louisiana. PROCEDURE: Samples were cultured for STEC O157 by use of sensitive and specific techniques, including selective broth enrichment, immunomagnetic separation, monoclonal antibody-based O:H enzyme immunoassay serotyping, and polymerase chain reaction virulence gene characterization. Point estimates and 95% confidence intervals were calculated for fecal shedding prevalence as well as site-specific prevalence from the oral cavity, skin, and feces. Logistic regression was used to assess seasonal variation and differences at various stages of lactation with respect to fecal shedding of STEC O157 in cattle sampled longitudinally. RESULTS: Summer prevalence in herds in = 13) was 38.5%, with a cow-level prevalence of 6.5%. Among positive herds, prevalence ranged from 3% to 34.6%. Samples from 3 of 5 herds sampled quarterly over 1 year yielded positive results for STEC O157. In herds with STEC O157, an increase in cow-level prevalence was detected during spring (13.3%) and summer (10.5%), compared with values for fall and winter. Site-specific prevalences of STEC O157:H7 from oral cavity, skin, and fecal samples were 0%, 0.7%, and 25.2%, respectively. CONCLUSIONS AND CLINICAL RELEVANCE: Our data indicated that STEC O157:H7 was commonly isolated from dairy cows in Louisiana, seasonally shed, and isolated from the skin surface but not the oral cavity of cows.     

A systemic vaccine based on Escherichia coli O157:H7 bacterial ghosts (BGs) reduces the excretion of E. coli O157:H7 in calves

Cattle are the main reservoir of enterohemorrhagic Escherichia coli O157:H7, a bacterium that, in humans, causes hemorrhagic colitis and hemolytic uremic syndrome (HUS), a life-threatening disease, especially in children and older people. Therefore, the development of vaccines preventing colonization of cattle by E. coli O157:H7 could be a main tool for an HUS control program. In the present study, we evaluated bacterial ghosts (BGs) of E. coli O157:H7 as an experimental vaccine against this pathogen. BGs are empty envelopes of Gram-negative bacteria, which retain the morphological surface make-up of their living counterparts and are produced by controlled expression of the cloned protein E, which causes loss of all the cytoplasm content. In this work, E. coli O157:H7 BGs were used for subcutaneous immunization of calves. The vaccinated animals elicited significant levels of BG-specific IgG but not IgA antibodies in serum. Low levels of IgA and IgG antibodies against BGs were detected in saliva from vaccinated animals. Following oral challenge with E. coli O157:H7, a significant reduction in both the duration and total bacterial shedding was observed in vaccinated calves compared to the nonimmunized group. We demonstrated that systemic vaccination with E. coli O157 BGs provides protection in a bovine experimental model. Further research is needed to reach a higher mucosal immune response leading to an optimal vaccine.