Spatial Epidemiology of Escherichia coli O157:H7 in Dairy Cattle in Relation to Night Roosts Of Sturnus vulgaris (European Starling) in Ohio,USA (2007–2009)

The goal of our study was to use spatial scan statics to determine whether the night roosts of European starlings (Sturnus vulgaris) act as point sources for the dissemination of Escherichia coli O157:H7 among dairy farms. From 2007 to 2009, we collected bovine faecal samples (n = 9000) and starling gastrointestinal contents (n = 430) from 150 dairy farms in northeastern Ohio, USA. Isolates of E. coli O157:H7 recovered from these samples were subtyped using multilocus variable‐number tandem repeat analysis (MLVA). Generated MLVA types were used to construct a dendrogram based on a categorical multistate coefficient and unweighted pair‐group method with arithmetic mean (UPGMA). Using a focused spatial scan statistic, we identified statistically significant spatial clusters among dairy farms surrounding starling night roosts, with an increased prevalence of E. coli O157:H7‐positive bovine faecal pats, increased diversity of distinguishable MLVA types and a greater number of isolates with MLVA types from bovine‐starling clades versus bovine‐only clades. Thus, our findings are compatible with the hypothesis that starlings have a role in the dissemination of E. coli O157:H7 among dairy farms, and further research into starling management is warranted.     

Risk factors associated with faecal shedding of verocytotoxin-producing Escherichia coli O157 in eight known-infected Danish dairy herds

A risk-factor study was performed in eight dairy herds found to excrete verocytotoxin-producing Escherichia coli (VTEC) O157 in a former prevalence study. Associations between excretion of VTEC O157 and management factors such as housing and feeding were analysed in a generalised linear mixed model. The animals were stratified in three age groups and sampled four times during 1 year. The risk of excreting VTEC O157 was higher among weaned calves than non-weaned calves. Among the calves aged 1–4 months, the risk was reduced if the calf had suckled colostrum from the mother or if the calf had stayed >2 days with the mother after calving. Calves aged 5–24 months that had been moved within the last 2 weeks had a higher risk, but risk was reduced if fed barley silage. Cows fed grain or molasses had a higher risk of excreting VTEC O157.     

1株大肠杆菌O157鸭源噬菌体的分离、纯化及其特性鉴定

将安徽某鸭场的400份鸭粪样品离心,微孔滤器过滤后提取上清液,将上清液与均匀涂布于LB固体培养基上的大肠杆菌O157ATCC43889滴定,6 h后初步鉴定噬菌体。将疑似存在噬菌体的上清液采用双层琼脂平板法进一步鉴定,结果分离鉴定了1株噬菌体,命名为EC43889-30。3次纯化后得到直径约为2 mm的噬菌斑,富集得到纯培养物。电镜观察,噬菌体粒子无尾,为直径50 nm左右的正六边形,形态与复层病毒科(Tectiviridae)的成员相似。将纯培养物稀释至107 pfu/mL,每隔10 min(至50 min)分别测其在50℃,60℃和70℃时噬菌体存活数,热稳定曲线显示,50℃时存活率均在70%左右,60℃时存活率明显下降,70℃时完全失活。宿主谱检测结果显示,其宿主范围较窄,对测试的42株鸭源大肠杆菌无裂解作用。     

Factors associated with the presence of Escherichia coli O157 in feedlot-cattle water and feed in the Midwestern USA

Our objective was to generate hypotheses about associations between management, climate, and the presence of Escherichia coli O157 in feedlot–cattle water tanks and in feedlot–cattle feed. Water samples from 710 tanks on 73 feedlots, and feed-samples from a subset of 504 pens on 54 feedlots, in four US states were tested for E. coli O157. Management and climate factors were ascertained by survey and observation. Escherichia coli O157 were isolated from 13% of the water tanks and at least one water tank was positive on 60% of the feedlots. The factors significantly associated with E. coli O157 in water were greater percentage of cattle shedding E. coli O157 in faeces within the same pen, higher concentration of total E. coli in the water, lack of the clarity of the water, the use of fly traps, the reported frequency of rodent sightings in the pen or alley area, and the weather at the time of sampling. Escherichia coli O157 were isolated from 14.9% of the feed samples obtained from the feedbunks. Factors positively associated with E. coli O157 in feed were higher heat index at the time of sampling, the presence of cottonseed meal in the ration, and the feedlot location (state). Coliform counts in feed, presence of E. coli O157 in water tanks and faecal prevalence of E. coli O157 were not associated with the presence of E. coli O157 in feed.     

双抗夹心ELISA检测肠出血性大肠杆菌O157方法的建立

以前期建立并纯化的肠出血性大肠杆菌O157：H7单克隆抗体3D6,建立一种适宜食品样品检测的双抗夹心ELISA检测方法。该方法只对O157菌株有特异性反应,对非O157型肠出血性大肠杆菌及其他菌株无交叉反应。敏感性试验结果表明,对大肠杆菌O157纯培养菌液检出限为1.2×105 CFU/mL。选择性增菌培养后,对牛肉、猪肉和鸡肉与模拟样品中的大肠杆菌O157的检出限为0.4～4CFU/g。     

Risk factors for hide contamination of Scottish cattle at slaughter with Escherichia coli O157

In the slaughter processing of cattle, contaminated hides have been identified as one of the major sources of Escherichia coli O157 carcase contamination. Logistic regression analysis was applied to data collected in a large scale study in Scotland involving 222 cattle forming 34 groups sent for slaughter from 30 farms to 10 slaughterhouses. Aspects of individual animal characteristics, farm management practices and slaughterhouse features were examined to identify potential risk factors for hide contamination at harvest. Two models were developed, the first in which slaughterhouse was modelled as a fixed effect, and a second model where slaughterhouse and farm groups were modelled as random effects. In the first model, there was a significantly increased risk of a carcase testing positive for E. coli O157 on the hide if either the hide of the carcase immediately before or after it on the line was contaminated (OR 3.6; 95% CI: 1.4–9.9). If both adjacent carcases had contaminated hides, the odds ratio for the study carcase having a contaminated hide rose to 11.5 (95% CI: 4.4–32.5). If animals were held in lairage, receiving hay as feed appeared to have a protective effect on hide contamination. Transportation to the slaughterhouse by haulier, as opposed to transport by the farmer, was associated with a 5.4 increase in the odds of E. coli O157 contamination. The use of a crush in the lairage, often employed when reading ear tags, was also found to significantly increase the odds of hide contamination with E. coli O157. In the second model, the inclusion of slaughterhouse and farm group as random effects resulted in two of the previously identified factors being associated with hide contamination. If at least one of the adjacent carcases on the line had a contaminated hide, the associated odds ratio was 6.6 (95% CI: 2.8–15.9), which rose to 22.7 (95% CI: 9.3–55.5) if both adjacent hides were contaminated. Receiving hay in lairage was found to be important to the model, although not significant in itself (OR 0.005; 95% CI: 1.2e⁻⁶–20.7). These results suggest that modifiable risk factors for hide contamination exist. However, in order best to reduce the prevalence of hide contamination at slaughter, individual slaughterhouse risk assessment and intervention strategies are appropriate.     

A preliminary study of Salmonella, verocytotoxigenic Escherichia coli/Escherichia coli O157 and Campylobacter on four mixed farms

The aims of this study were to investigate the incidence of Salmonella, verocytotoxigenic Escherichia coli (VTEC)/Escherichia coli O157 and Campylobacter on four mixed farms and to characterize the isolates in terms of a range of virulence factors. Eighty-nine composite (five different samples from the same animal species combined) faecal [cattle (24), pigs (14), sheep (4), poultry (4), horses (7), deer (4), dogs (9), rodents (2) and wild birds (20)] samples, 16 composite soil samples plus 35 individual water samples were screened using culture-based, immunomagnetic separation and molecular methods. Salmonella was detected in bovine faeces, cattle and poultry house water. Salmonella serotypes/phage types included Dublin, Kiel and Typhimurium DT193, and most isolates were spvC, invA and rck positive. The pefA and rck genes were found exclusively in the non-Typhimurium strains, while Salmonella Dublin and Salmonella Kiel strains carried Salmonella genomic island I marker(s). VTEC/E. coli O157 were found in deer and dog faeces only. The E. coli O157 isolate was an enteroinvasive E. coli, while the VTEC isolate was untypable but carried the vt1, eaeA, hlyA, tir and eptD genes. This article reports the first confirmed carriage of E. coli O157 in Irish deer. Campylobacter species were not detected over the course of this study. It was concluded that [1] Salmonella, VTEC and Campylobacter have low (<5%) prevalence or are absent on the farms in this study; [2] water was an important source of bacterial pathogens; [3] both dogs and deer may act as a source of pathogenic E. coli and [4] key virulence and resistance determinants are widespread in farm Salmonella strains. This study highlights the need to control water as a source of pathogens and suggests that the domestic pets and deer should be considered in any farm risk assessment.     

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.     

Associations between management, climate, and Escherichia coli O157 in the faeces of feedlot cattle in the Midwestern USA

Our objective was to generate hypotheses for potential on-farm control strategies for Escherichia coli O157 by identifying associations between management practices and climate, and the presence of E. coli O157 in feedlot cattle. Faeces were obtained from 10,622 cattle in 711 pens on 73 feedlots between May and August 2001. Management and climate information was obtained by questionnaire and observation at the time of sampling. The prevalence of E. coli O157 was 10.2% at the sample level, 52.0% at the pen-level, and 95.9% at the feedlot-level. The factors associated with the presence of E. coli O157 in cattle faeces were the frequency of observing cats in the pens or alleys (most common when observed daily), the presence of E. coli O157 in the water tanks (positive association), the historical use of injectable mass medication (positive association), the use of antibiotics in the ration or water (negative association), the wetness of the pen, number of cattle in the pen (negative association), wind velocity (positive association), and height of the feed bunk (positive association).     

Study on Seasonal Impact to the Distribution of Bovine E.coli O157: H7 in Xinjiang

To study the impact of season to the distribution of bovine E.coli O157:H7,samples of anus swabs (399),feces (68),water (29) and feed (43) were collected in the spring, summer,autumn and winter from A,B and C farms of Xinjiang. After enrichment by EC broth, SMAC and MUG selective culture were then performed. Finally,PCR was used for identification and virulence gene detection of isolated strains. A total of 5 E.coli O157:H7 strains were isolated from 539 samples from three farms (0.93%,5/539), 2 of them were from spring (1.44%,2/139),1 from autumn(0.56%,1/180),2 from winter (1.38%,2/145) and no strains were isolated from summer samples. One strain were isolated from anus swab samples in farm B (0.69%,1/145) and one were isolated from anus swabs (0.66%,1/152) and three strains were isolated from feed samples in farm C (20.00%,3/15),and no target strains were isolated from water samples. The distribution of bovine E. coli O157:H7 had obvious seasonal characteristics.One E.coli O157:H7 strain of farm B was isolated from autumn and four of farm C were from isolated spring (2 strain) and winter (2 strain),and the isolation rate of E. coli O157:H7 in spring and winter were higher than that in summer and autumn. In conclusion,under the special climate characteristics and feeding mode in Xinjiang,to prevent and control the spreading of E. coli O157:H7 of cattle,we must pay great attention on hygiene management of pens at cold season, specially avoiding the feed contaminated by feces.     

An investigation of factors associated with the prevalence of verocytotoxin producing Escherichia coli O157 shedding in Scottish beef cattle

The prevalence of verocytotoxin-producing Escherichia coli (VTEC) O157 in 12-30-month-old beef finishing cattle in Scotland was determined using 1g faeces samples enriched in buffered peptone water, followed by immunomagnetic separation (IMS) and isolation on sorbitol MacConkey agar with cefixime and tellurite supplement (CT-SMAC). A validated questionnaire was used to collect information that could be associated with the samples. Generalised Linear Models and Generalised Linear Mixed Models were used to identify factors associated with shedding both between and within groups. A total of 14,856 samples were collected from 952 farms, of which 1231 were positive for VTEC O157. Prevalence levels were calculated with 95% confidence intervals as follows: 7.9% (6.5%, 9.6%) of animals sampled were estimated to be shedding VTEC O157, while 22.8% (19.6%, 26.3%) of farms were estimated as having at least one animal shedding in the group sampled. The median percentage of animals shedding in positive groups was 25% (20%, 32%). An increased probability of a group containing a shedding animal was associated with larger numbers of finishing cattle, the presence of pigs on the farm, or the farm being classed as a dairy unit stocking beef animals. Farms that spread slurry on grazing land were more likely to have shedding animals, while those that spread manure were at lower risk. Groups with older animals were less likely to be identified as positive. There was no significant regional difference in group shedding probabilities, but the proportion of positive groups dropped over two successive years of the study. Higher mean levels of shedding in positive groups were associated with animals being housed rather than at pasture, and this effect was stronger in groups which had recently had a change in housing or diet. Farms with animals at pasture had lower mean prevalence where water was supplied from a natural source, as had farms with higher numbers of finishing cattle. There remained unexplained variability in mean prevalence levels on positive farms in different areas of Scotland.     

猪源大肠杆菌O157:H7体外氟苯尼考耐药诱导及对抗菌药物敏感性变化研究

为初步研究猪源大肠杆菌O157:H7 (E.coli O157:H7)对氟苯尼考耐药性的产生和消除机制,本研究采用亚抑菌浓度体外耐药诱导的方法将两株猪源大肠杆菌O157:H7诱导成氟苯尼考高度耐药菌株,采用无氟苯尼考压力下连续传代培养的方法将获得的氟苯尼考耐药菌株的氟苯尼考耐药性消除,检测耐药诱导菌和耐药消除菌对抗菌药物的敏感性,并检测菌株质粒携带的耐药基因。结果显示,经氟苯尼考耐药诱导,猪源大肠杆菌O157:H7对氟苯尼考、阿莫西林、头孢唑啉、头孢拉定和头孢噻吩由敏感变为耐药,对头孢噻肟的敏感性由敏感变为中介,对氧氟沙星、环丙沙星和阿奇霉素由中介变为耐药;而经耐药消除后,菌株恢复对上述药物的敏感性;在菌株的质粒中检测到氟苯尼考耐药基因、喹诺酮类耐药基因和β-内酰胺酶基因,与耐药表型相符。结果表明,在氟苯尼考压力的长期存在下,猪源大肠杆菌O157:H7对氟苯尼考产生耐药,且对青霉素类、头孢类和喹诺酮类药物产生交叉耐药,在去除氟苯尼考压力下连续培养,可消除菌株的部分耐药性。     

大肠杆菌O157:H7 rfbE基因和fliC基因的克隆与序列分析

根据已发表的E．coli O157：H7 EDL933株的rfbE基因和fliC基因，设计了两对引物，分别对两个O157：H7，菌株的rfbE基因和一个菌株的fliC基因进行PCR，并将PCR产物克隆于pMD18-T栽体质粒。测序结果表明，获得到了与理论相符的582bp rfbE基因片段和802bp fliC基因片段。一株菌的rfbE基因存在无意义突变（两个）。而另一株菌fliC基因有一个有意义突变（aac→gac）     

动物源E.coli O157∶H7 stx基因的检测与系统进化分析

为了解2009－2010年间在河南、甘肃地区分离鉴定的5株大肠埃希菌O157(E.coli O157)携带stx的情况及不同分离株间stx分子进化与变迁的情况,本研究利用PCR方法对分离株进行了stx基因检测,并完成了序列测定与系统演化分析。结果表明,5株不同动物源的分离株均含有stx1及stx2基因。序列分析结果显示5株分离株间stx1、stx2的核苷酸及氨基酸同源性均较高;stx1基因均与参考株中的山羊源和食品源E.coli O157菌株的同源性较高,进化树中遗传距离最近;分离株的stx2基因与多株牛源及少数人源参考株也具有较高的同源性,进化树中虽然5株分离菌均在一个大主干分支中,但分离株27与其他各分离株及参照株遗传距离最远,独自处于一次级分支中;分离株L37与W、12与50分别分布于牛源、人源E.coli O157小次级分支中;由此可推测,分离株所携带的stx1很有可能是经食品源或羊源E.coli O157传递而来;分离株L37与W、分离株12与50的stx2可能是由牛源、人源E .coli O157菌株传递而来,分离株27的stx2来源不清楚。研究结果表明,5株E.coli O157分离株均含有stx1、stx2基因,但两个基因的起源存在差异。     

Detection of Escherichia coli O157 and Escherichia coli O157:H7 by the immunomagnetic separation technique and stx1 and stx2 genes by multiplex PCR in slaughtered cattle in Samsun Province, Turkey

This study was conducted to investigate the presence of Escherichia (E.) coli O157 and E. coli O157:H7 and stx1 and stx2 genes on cattle carcasses and in rectal samples collected from Samsun Province of Turkey. A total of 200 samples collected from cattle carcasses and the rectal contents of 100 slaughtered cattle from two commercial abattoirs were tested using the immunomagnetic separation technique and multiplex PCR methods. E. coli O157 and E. coli O157:H7 were detected in 52 of the 200 samples (26%) tested. Of the positive samples, 49 were E. coli O157 and three were E. coli O157:H7. The E. coli O157 strain was isolated from 24 carcasses and 25 rectal samples, while E. coli O157:H7 was isolated from two carcasses and one rectal sample. Of the 49 samples positive for E. coli O157, 32 were from the rectal and carcass samples of the same animal, while two E. coli O157:H7 isolates were obtained from rectal swabs and carcasses of the same animal. The stx1 and stx2 genes were both detected in 35 E. coli O157 isolates and one E. coli O157:H7 isolate, but the stx2 gene was only detected alone in two E. coli O157 isolates. Overall, 16 carcasses tested positive for E. coli O157 and one carcass tested positive for E. coli O157:H7 based on both carcass and rectal samples. Overall, the results of this study indicate that cattle carcasses pose a potential risk to human health due to contamination by E. coli O157 and E. coli O157:H7 in the feces.     

Effectiveness of simulated interventions in reducing the estimated prevalence of E. coli O157:H7 in lactating cows in dairy herds

A transmission model developed to investigate the dynamics of Escherichia coli O157:H7 bacteria in a typical Dutch dairy herd was used to assess the effectiveness of vaccination, diet modification, probiotics (colicin) and hygienic measures as to water troughs and bedding, when they are applied single or in combination, in reducing the prevalence of infected animals. The aim was to rank interventions based on their effectiveness in reducing the baseline prevalence of infected animals in the lactating group. The baseline prevalence of the lactating group and the within-herd prevalence were estimated by the model to be 5.02% and 13.96% respectively. The results show that all four interventions, if applied to all four animal groups or only to young stock, are the most effective and will reduce the baseline prevalence by 84% to 99%. In general, combinations of hygiene (applied in all groups) and one other intervention had the highest effectiveness in reducing prevalence in the lactating group. Vaccination and diet modification show a slightly higher effectiveness than colicin and hygiene.     

奶牛乳和血中大肠杆菌和轮状病毒抗体检测比较

用轮状病毒和大肠杆菌给妊娠后期奶牛免疫,使之产生抗这两种病原的抗体,定期采血和乳,分别用试管凝集反应和反向间接血凝抑制试验检测大肠杆菌和轮状病毒抗体,并比较血和乳中抗体相互关系,试验结果说明：奶牛在未免疫情况下,只在初乳中才能检出抗体;在人工免疫情况下,在初乳和近２个月的常乳中均可检测到抗体,但乳中抗体比血中抗体低２个滴度左右。     

亨得拉病毒核蛋白基因(N)在大肠杆菌的高效表达与反应原性鉴定

从引进的含亨得拉病毒（Hendra virus，HeV）基因的克隆质粒中扩增得到该病毒核蛋白基因（N），将其克隆到pET-28a（＋）原核表达栽体，转化大肠杆菌后成功地表达了N蛋白，SDS-PAGE电泳分析结果表明目的基因在JM109（DE3）大肠杆菌菌株中得到了良好的表达，目的蛋白量占菌体总蛋白的14．6％。Western-blotting检测表明，表达的蛋白可以被兔抗亨得拉全病毒阳性血清识别，表明该蛋白具有良好的反应性。该研究结果可进一步应用于诊断试剂和单克隆抗体的制备以及流行病学调查，以防范该病在我国的流行。     

甜叶菊绿原酸增强大肠杆菌感染蛋雏鸡免疫力研究

旨在评价甜叶菊绿原酸增强人工腹气囊感染大肠杆菌O78蛋雏鸡免疫力的作用效果,为功能性抗生素替代品研发提供基础参数支持.本试验随机将1日龄、体重无显著差异的健康海兰蛋鸡360只分为6组:空白对照组(C)、大肠杆菌O78处理组(EC0)、1.0g·L-1杜仲素+大肠杆菌O78处理组(ED1)、1.0g·L-1甜叶菊绿原酸+...