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.     

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.     

Influence of processed grains on fecal pH, starch concentration, and shedding of Escherichia coli O157 in feedlot cattle

Manipulation of cattle diets has been proposed as a possible preharvest control measure for Escherichia coli O157. Altering hindgut fermentation through diet changes may be a means to reduce fecal shedding of E. coli O157. In Exp. 1, the objective was to determine whether fecal shedding of E. coli O157 was related to fecal starch concentration. Beginning on d 20, and every week thereafter until d 61, steers in 54 pens (6 to 7 steers per pen) were sampled (n = 122) by fecal collection and rectoanal mucosal swabs (RAMS) for E. coli O157 and fecal starch concentration determinations. Escherichia coli O157 prevalence was 3.3% in fecal samples, 4.1% as measured by RAMS, and 4.9% by fecal or RAMS samples. Steers positive for E. coli O157 contained 21% more (P < 0.05) fecal starch than steers that were negative for E. coli O157. In Exp. 2, we attempted to alter the concentration of starch escaping rumen fermentation by feeding finishing diets based on steam-flaked corn (SFC) and dry-rolled corn (DRC) to 30 heifers prescreened for being culture positive for fecal E. coli O157. Beginning on d 13, heifers were sampled (feces and RAMS) weekly to monitor fecal pH and starch concentration, and prevalence of E. coli O157. Prevalence of E. coli O157 remained above 30% for the first 13 d, but declined (P < 0.05) over the entire 7-wk period. Based on RAMS, the prevalence of E. coli O157 tended to be greater (P = 0.08) for heifers fed SFC than for those fed the DRC diet. After d 20, heifers fed DRC had greater (P < 0.05) fecal starch and lower (P < 0.05) fecal pH than heifers fed SFC. Fecal pH was negatively correlated (r = - 0.34; P < 0.05; n = 143) with fecal starch concentration. Fecal starch concentration and pH were not different (P > 0.05) for heifers that were positive or negative for E. coli O157. Our data suggest that fecal shedding of E. coli O157 was not related to fecal pH or starch concentration in cattle fed grain-based diets.     

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.     

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.     

Feeding supplemental dried distiller's grains increases faecal shedding of Escherichia coli O157 in experimentally inoculated calves

Escherichia coli O157 is an important foodborne pathogen and asymptomatic cattle serve as major reservoirs for human infection. We have shown a positive association between feeding distiller's grains and E. coli O157 prevalence in feedlot cattle. The objective of this study was to determine the effect of feeding dried distiller's grain (DDG) on faecal shedding of E. coli O157 in calves experimentally inoculated with E. coli O157. Holstein calves (five per treatment group), fed steam-flaked corn-based high-grain diets supplemented with 0% (control) or 25% DDG, were orally inoculated with a five-strain mixture (6 x 10(9) CFU/calf) of nalidixic acid-resistant (NalR) E. coli O157. Faecal samples were taken three times per week for 6 weeks to determine the prevalence and concentration of Nal E. coli O157. At the end of the study (day 43), calves were euthanized and necropsied. Ruminal, caecum, colon, and rectal contents, and rectoanal mucosal swab (RAMS) samples were collected at necropsy to determine NalR E. coli O157 concentration. There was a trend for an interaction between treatment and faecal sampling day. The concentration of NalR E. coli O157 in the faeces was significantly higher in faecal samples from calves fed DDG compared with control calves on days 35, 37, 39 and 42. At necropsy, the concentration of NalR E. coli O157 was higher in the caecum (P = 0.01), colon (P = 0.03) and rectum (P = 0.01) from calves fed DDG compared with control animals. The number of sites at necropsy positive for NalR E. coli O157 was higher in calves fed DDG compared with calves in the control treatment (P < 0.001). Our results indicate that E. coli O157 gut persistence and faecal prevalence increased in calves fed DDG, which potentially have important implications for food safety.     

Fecal Escherichia coli O157:H7 shedding patterns of orally inoculated calves.

To assess the duration of fecal shedding upon initial infection, the duration of shedding after subsequent re-infection and the effects of dietary restriction and antibiotic treatment on shedding recrudescence, four, one-week-old calves were orally inoculated on three separate occasions with 5x10(8) cfu of Escherichia coli O157:H7 strain 86-24 Nal-R. Fecal shedding was followed by serial culture three times weekly. Following the first inoculation, the calves shed E. coli O157:H7 in their feces for a mean of 30 days, with a range of 20 to 43 days. Following the second and third inoculations, the calves shed E. coli O157:H7 in their feces for 3-8 days. In each of the three inoculations, feed was withheld from the calves for 24 h after they had become fecal culture negative. Two calves resumed shedding, one for 1 day and the other for 4 days, after food was withheld after the third inoculation, but not in the first two inoculations. In the third inoculation, one calf resumed shedding for one day after treatment with oxytetracycline. No E. coli O157:H7 strain 86-24 Nal-R was found in the calves at necropsy. These calves did not exhibit persistent low-level shedding, and did not appear to be persistently colonized with E. coli O157:H7.     

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.     

Intermittent Escherichia coli O157:H7 colonisation at the terminal rectum mucosa of conventionally-reared lambs

In cattle, the lymphoid rich regions of the rectal-anal mucosa at the terminal rectum are the preferred site for Escherichia coli O157:H7 colonisation. All cattle infected by rectal swab administration demonstrate long-term E. coli O157:H7 colonisation, whereas orally challenged cattle do not demonstrate long-term E. coli O157:H7 colonisation in all animals. Oral, but not rectal challenge of sheep with E. coli O157:H7 has been reported, but an exact site for colonisation in sheep is unknown. To determine if E. coli O157:H7 can effectively colonise the ovine terminal rectum, in vitro organ culture (IVOC) was initiated. Albeit sparsely, large, densely packed E. coli O157:H7 micro-colonies were observed on the mucosa of ovine and control bovine terminal rectum explants. After necropsy of orally inoculated lambs, bacterial enumeration of the proximal and distal gastrointestinal tract did suggest a preference for E. coli O157:H7 colonisation at the ovine terminal rectum, albeit for both lymphoid rich and non-lymphoid sites. As reported for cattle, rectal inoculation studies were then conducted to determine if all lambs would demonstrate persistent colonisation at the terminal rectum. After necropsy of E. coli O157:H7 rectally inoculated lambs, most animals were not colonised at gastrointestinal sites proximal to the rectum, however, large densely packed micro-colonies of E. coli O157:H7 were observed on the ovine terminal rectum mucosa. Nevertheless, at the end point of the study (day 14), only one lamb had E. coli O157:H7 micro-colonies associated with the terminal rectum mucosa. A comparison of E. coli O157:H7 shedding yielded a similar pattern of persistence between rectally and orally inoculated lambs. The inability of E. coli O157:H7 to effectively colonise the terminal rectum mucosa of all rectally inoculated sheep in the long term, suggests that E. coli O157:H7 may colonise this site, but less effectively than reported previously for cattle.     

Oral infection with a Shiga toxin-negative Escherichia coli O157:H7 strain elicits humoral and cellular responses but does not protect sheep from colonisation with the homologous strain

We have previously shown that rectally inoculated sheep excrete Escherichia coli O157:H7 during weeks to months without developing a clear antibody response. However, antibodies against this bacterium were observed in naturally infected sheep, which most likely became orally infected. To understand this difference, sheep were orally inoculated with the same Shiga toxin-negative E. coli O157:H7 strain that was used for the rectal inoculation. A primary oral inoculation resulted in shedding of E. coli O157:H7 in the faeces and detection of antibody responses against intimin, EspA and EspB. The antibody titres waned as shedding decreased. A secondary inoculation resulted in longer shedding, even though a booster antibody response occurred. Cellular responses followed a similar pattern as the antibody levels, albeit with a lower secondary response. The presence of antigen-specific antibody-secreting cells indicates involvement of both a systemic response in the spleen and a local immune response in the terminal rectum. These results suggest that E. coli O157:H7 has to pass the small intestine to evoke antibody responses.     

Dry-rolled or steam-flaked grain-based diets and fecal shedding of Escherichia coli O157 in feedlot cattle

Hindgut is a major colonization site for Escherichia coli O157 in cattle. In this study, diets were formulated to effect changes in hindgut fermentation to test our hypothesis that changes in the hindgut ecosystem could have an impact on fecal shedding of E. coli O157. Feedlot heifers (n = 347) were prescreened for the prevalence of E. coli O157 by fecal and rectoanal mucosal swab cultures. A subset of 40 heifers identified as being positive for fecal shedding of E. coli O157 was selected, housed in individual pens, and randomly allocated to 4 dietary treatments. Treatments were arranged as a 2 x 2 factorial, with factor 1 consisting of grain type (sorghum or wheat) and factor 2 being method of grain processing (steam-flaking or dry-rolling). Four transition diets, each fed for 4 d, were used to adapt the animals to final diets that contained 93% concentrate and 7% roughage. The grain fraction consisted of dry-rolled sorghum, steam-flaked sorghum, a mixture of dry-rolled wheat and steam-flaked corn, or a mixture of steam-flaked wheat and steam-flaked corn. Wheat diets contained 52% wheat and 31% steam-flaked corn (DM basis). Fecal and rectoanal mucosal swab samples were obtained 3 times a week to isolate (enrichment, immunomagenetic separation, and plating on selective medium) and identify (sorbitol negative, indole production, and agglutination test) E. coli O157. The data were analyzed as repeated measures of binomial response (positive or negative) on each sampling day. Method of processing (dry-rolled vs. steam-flaked), sampling day, and the grain type x day interaction were significant (P < 0.05), but not the method of processing x grain type interaction. The average prevalence of E. coli O157 from d 9 was greater (P < 0.001) in cattle fed steam-flaked grains (65%) compared with those fed dry-rolled grains (30%). Average prevalence in cattle fed sorghum (51%) or wheat (43%) were similar (P > 0.10) on most sampling days. Results from this study indicate that feeding dry-rolled grains compared with steam-flaked grains reduced fecal shedding of E. coli O157. Possibly, dry-rolling allowed more substrate to reach the hindgut where it was fermented, thus making the hindgut inhospitable to the survival of E. coli O157. Dietary intervention to influence hindgut fermentation offers a simple and practical mitigation strategy to reduce the prevalence of E. coli O157 in feedlot cattle.     

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.     

Study of porcine postweaning diarrhea involving K88(-) hemolytic Escherichia coli

The onset of fecal shedding, serogroup involvement, and association of hemolytic Escherichia coli with the postweaning diarrhea syndrome were studied in swine. The only E coli O antigen, detected by slide agglutination with the antisera used, was O157; K antigens were not detected. The ligated intestinal loop test (LILT) was used for enterotoxigenicity testing. All O157 serogroup isolates (n = 9) were hemolytic, and 89% (8 of 9) were LILT positive. Of all hemolytic isolates tested, 59% (10 of 17) were LILT positive. Twenty-seven nonhemolytic isolates were tested for enterotoxigenicity; of these, 45% (12) were LILT positive. The onset of postweaning diarrhea coincided with the modal onset of hemolytic E coli shedding at postweaning day 7. At postweaning day 7, hemolytic E coli shedding was concurrently associated with diarrhea (P less than 0.0005), whereas later during the postweaning period, this was not the situation.     

Effects of short-term early gestational exposure to endophyte-infected tall fescue diets on plasma 3,4-dihydroxyphenyl acetic acid and fetal development in mares

Consumption of wild-type (toxic) endophyte-infected tall fescue (E+) by horses during late gestation is known to adversely affect pregnancy outcome; however, little is known of the potential disruptive consequences of E+ consumption by mares during the critical phases of placentation and fetal development in early pregnancy. The objective of this study was to evaluate the detrimental effects of feeding E+ to mares during early gestation. Mares (n = 12) paired by stage of gestation (d 65 to 100) were assigned to diets (six per diet) consisting of endophyte-free (E-) or E+ tall fescue seed (50% E- or E+ tall fescue seed, 45% sweet feed, and 10% molasses fed at 1.0% of BW/d). Mares also had ad libitum access to E+ or E- annual ryegrass hay, and were fed diets for 10 d. Following removal from the tall fescue diet on d 11, mares were placed on common bermudagrass pasture and monitored until d 21. Morning and evening rectal temperatures were recorded and daily blood samples were collected for progesterone and prolactin (PRL) analyses, whereas samples for 3,4-dihydroxyphenyl acetic acid (a catecholamine metabolite) analysis were collected on alternate days. For clinical chemistry analysis, blood samples were collected on d 0, 5, 10 and 21. Daily urine samples were collected for ergot alkaloid analysis, and ultrasonography was performed for presence of echogenic material in fetal fluids. Rectal temperatures (E+ 37.76+/-0.03; E- 37.84+/-0.03 degrees C) and serum PRL concentrations (E+ 14.06< or =0.76; E- 12.11+/-0.76 ng/mL) did not differ (P = 0.96) between treatments. Measuring the change in basal serum concentration from d 0 over time, progesterone concentrations did not differ (-0.64 +/-1.49 and -0.55+/-1.47 ng/mL for E+ and E- mares, respectively). There was no negative pregnancy outcome, and ultrasonography indicated no increase in echogenic material in fetal fluids. Plasma 3,4-dihydroxyphenyl acetic acid concentrations decreased (P < 0.05) in E+ compared with E- mares (2.1+/-0.14 and 4.4+/0.43 ng/mL, respectively). Urinary ergot alkaloid concentration was greater (P < 0.01) in mares consuming E+ compared with E- (532.12+/- 52.51 and 13.36+/-2.67 ng/mg of creatinine, respectively). Although no fetal loss was observed during the current study, elevated concentrations of urinary ergot alkaloid were consistent with depressed endogenous catecholamine activity, suggestive of an endocrine disruptive effect of hypothalamic origin.     

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.     

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.     

Acid resistance of Escherichia coli O157:H7 from the gastrointestinal tract of cattle fed hay or grain

There has been strong debate as to whether feeding cattle hay prior to slaughter will reduce the number and/or virulence of Escherichia coli O157:H7 in the bovine gastrointestinal tract (GIT). This study addressed this issue by comparing numbers, persistence, and acid resistance of generic coliforms and E. coli O157:H7 from various gastrointestinal tract sites of cattle fed grain or hay. Mature Angus steers, doubly cannulated into the rumen and duodenum were inoculated with E. coli O157:H7. Aliquots of digesta from the rumen, duodenum, and rectum were cultured directly or acid shocked (pH 2.0) and then cultured to determine acid resistance. The culture technique used was as sensitive as standard immunomagnetic bead separation protocols. E. coli O157:H7 from hay-fed or grain-fed cattle were similarly acid resistant in all GIT locations. In contrast, generic coliforms from the rumen and rectum of hay-fed animals were more sensitive to an acid shock than coliforms from those GIT locations in grain-fed animals. E. coli O157:H7 colonized the most distal region of the GIT and was not consistently cultured from the rumen or the duodenum. Numbers in the upper GIT did not predict numbers or persistence of E. coli O157:H7 in rectal samples. Grain-feeding or hay-feeding did not affect survival of E. coli O157:H7 in the rumen, nor its passage through the abomasum (pH 2.0) to the duodenum. These data show that generic coliforms behave differently in the bovine host than E. coli O157:H7 and that E. coli O157:H7 acid resistance was independent of animal diet.     

The characterisation of E. coli O157:H7 isolates from cattle faeces and feedlot environment using PFGE

The objectives of this study were to investigate the diversity of Escherichia coli O157:H7 isolates obtained over a 3-month period from a cattle feedlot in order to assess the relationship between environmental and faecal isolates and to determine the pattern of transmission of E. coli O157:H7 between groups of cattle. Faecal samples were obtained from cattle housed in four adjacent feedlot pens at monthly intervals, with environmental pen samples collected simultaneously. All E. coli O157:H7 isolates obtained were examined by pulsed field gel electrophoresis (PFGE), polymerase chain reaction (PCR) to detect eaeA, ehxA, stx1 and stx2 genes and antibiotic sensitivity profiling. Ten isolates were subjected to acid shock to imitate conditions in the acidic cattle abomasum and assess the effect on PFGE profiles. E. coli O157:H7 was isolated from 69 faecal samples and 26 environmental samples. All isolates (n=95) carried the genes for eaeA, ehxA and stx2 and were sensitive to all antibiotics tested. The PFGE profiles of all isolates differed by no more than two bands and clustered within 80% similarity following dendrogram analysis. Acid shock had no effect on the subsequent PFGE patterns. A total of 8.7% (6/69) of cattle were shedding E. coli O157:H7 in the first month with faecal shedding increasing to 52% (36/69) by the third month of the study. A single isolate of E. coli O157:H7 may be passed rapidly through cattle pens, with the environment acting as a significant reservoir for transmission. PFGE is a useful tool for tracking the direct and indirect transmission of E. coli O157:H7 isolates on the farm.     

A longitudinal study of Shiga-toxigenic Escherichia coli (STEC) prevalence in three Australian diary herds

Over a 12 month period, 588 cattle faecal samples and 147 farm environmental samples from three dairy farms in southeast Queensland were examined for the presence of Shiga-toxigenic Escherichia coli (STEC). Samples were screened for Shiga toxin gene (stx) using PCR. Samples positive for stx were filtered onto hydrophobic grid membrane filters and STEC identified and isolated using colony hybridisation with a stx-specific DNA probe. Serotyping was performed to identify serogroups commonly associated with human infection or enterohaemorrhagic Escherichia coli (EHEC). Shiga-toxigenic Escherichia coli were isolated from 16.7% of cattle faecal samples and 4.1% of environmental samples. Of cattle STEC isolates, 10.2% serotyped as E. coli O26:H11 and 11.2% serotyped as E. coli O157:H7, and the E. coli O26:H11 and E. coli O157:H7 prevalences in the cattle samples were 1.7 and 1.9%, respectively. Prevalences for STEC and EHEC in dairy cattle faeces were similar to those derived in surveys within the northern and southern hemispheres. Calves at weaning were identified as the cattle group most likely to be shedding STEC, E. coli O26 or E. coli O157. In concurrence with previous studies, it appears that cattle, and in particular 1-14-week-old weanling calves, are the primary reservoir for STEC and EHEC on the dairy farm.     

Persistence of Escherichia coli O157:H7 in experimentally infected swine

These experiments determined the ability of Escherichia coli O157:H7 to colonize and persist in pigs simultaneously inoculated with other pathogenic E. coli strains. Three-months-old pigs were inoculated with a mixture of five E. coli strains. The mixture included two Shiga toxigenic E. coli (STEC) O157:H7 strains, two enterotoxigenic E. coli (ETEC) strains and one enteropathogenic E. coli (EPEC) strain. A high dose mixture with all five strains at 10(10)CFU/animal (CFU: colony forming units) and a low dose mixture with the STEC strains at 10(7)CFU and the EPEC and ETEC strains remaining at 10(10)CFU were used. The STEC strains persisted in the alimentary tracts of some pigs at 2 months post-inoculation, following inoculation with both the high and low dose mixtures. When all strains were given at 10(10)CFU (high dose) the STEC strains persisted in greater numbers and in more pigs than did the other E. coli strains. The results demonstrated that persistent colonization (> or =2 months) by E. coli O157:H7 can occur in pigs. These findings were similar to those reported from sheep inoculated with the same mixture of E. coli strains. The results are consistent with reports suggesting that pigs have the potential to be reservoir hosts for STEC O157:H7.