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1.
动物及动物产品中大肠埃希氏菌O157带菌情况的调查 总被引：2，自引：0，他引：2
大肠埃希氏菌O157（Escherichia coli O157）是一种出血性致病性大肠杆菌,能引起人类出血性腹泻和溶血性尿毒综合征,其感染已成为世界性公共卫生问题。1982年美国首次报道了大肠埃希氏菌O157：H7引起的出血性肠炎暴发,此后世界各地相继有由该菌所致疫情的报道,尤其在1996年5～8月日本发生了迄今为止最大的一起O157：H7大肠埃希氏菌的流行,累积患者近万人。 相似文献
2.
贺晓龙 《青海畜牧兽医杂志》2006,36(3):34-35
WHO不久前指出在过去的20年里,世界上出现了约30种新的传染病,1982年美国首次报告的由大肠埃希菌O157:H7引起的出血性肠炎就是其中之一。大肠肝菌O157:H7属于肠出血性大肠埃希氏菌(EnterohemorrhagicE.coliEHEC),尽管大肠杆菌O157:H7高致病力的机制尚未明了,但已确定紧密素(int 相似文献
3.
为建立肠出血性大肠埃希菌O157:H7的多重PCR检测方法,针对O157菌特异性eaeA基因、fliC基因、志贺毒素基因(stx1和stx2)及rfbE基因设计5对引物,构建多重PCR反应体系,优化反应的引物浓度和温度检测其特异性和敏感性,并对牛、猪、鸡及犬等不同动物来源的样品进行了大肠埃希菌O157检测。结果显示,该方法具有良好的特异性和敏感性,敏感性达到5×10~3CFU。从检测阳性样本中均分离到目的菌。应用建立的方法在确定样品中是否含有大肠埃希菌O157的同时,还能对菌株毒力进行初步判定。成功建立了肠出血性大肠埃希菌O157:H7多重PCR检测方法,为该病原菌感染的预防和监测提供了简便、快速的技术手段,具有良好的应用前景。 相似文献
4.
《动物医学进展》2021,42(6)
为了解2018年-2019年从广东省内生猪屠宰场分离的19株大肠埃希氏菌O157:H7耐药情况,采用微量肉汤稀释法测定阿莫西林等16种抗菌药物对分离菌株的最小抑菌浓度(MIC),并用普通PCR法检测blaNDM-1等10种耐药基因。结果表明,19株大肠埃希氏菌O157:H7分离菌株对不同抗生素存在耐药差异,其中对阿莫西林、氨苄西林、青霉素、氯霉素和恩诺沙星的耐药率均为100%,对头孢哌酮的耐药率最低为5.26%,其余10种抗菌药物的耐药率位于31.58%～84.21%之间;mcr-1、tetM、floR和rmtB 4种耐药基因未检出,blaNDM-1等6种耐药基因检出率范围为5.26%～89.47%。生猪屠宰场大肠埃希氏菌O157:H7耐药且多重耐药严重,分离株携带丰富的耐药基因。 相似文献
5.
《动物医学进展》2021,42(2)
为了解肠出血性大肠埃希氏菌O157:H7、沙门氏菌和空肠弯曲菌等3种食源性致病菌在生猪屠宰场的污染情况。采集广东惠州和清远两地共10家生猪屠宰场498头猪肉样品,分别经改良EC新生霉素增菌肉汤、SC(亚硒酸盐胱氨酸增菌液)和Bolton肉汤增菌,用大肠埃希氏菌O157:H7、沙门氏菌和空肠弯曲菌的特异性引物进行PCR扩增及测序检测,阳性菌液再分别涂抹于山梨醇麦康凯平板、DHL(胆硫乳琼脂)和Skirrow血琼脂,挑取疑似菌落再次PCR验证,计算检出率。结果显示,检出大肠埃希氏菌O157:H7阳性样本2份,检出率0.4%,检出沙门氏菌阳性样本70份,检出率14.06%,空肠弯曲菌未检出。结果表明,惠州和清远两地屠宰场均存在食源性致病菌污染情况,其中沙门氏菌污染较为严重,惠州地区检出率高于清远地区。提示屠宰场应加强屠宰过程中环境和加工用具的消毒,以保证屠宰环节的食品安全。 相似文献
6.
上海市动物及其产品中大肠埃希菌O157:H7带菌情况的调查 总被引：1，自引：0，他引：1
为了解上海市动物及动物产品中大肠埃希菌O 157∶H 7带菌情况及其毒力采集上海市猪、牛等动物粪便以及市场和超市猪肉、牛肉、牛奶、虾仁等样本568 份,应用免疫胶体金技术、分离培养、形态特征观察生化特性鉴定、血清学试验、多重引物PCR进行大肠埃希菌O 157∶H 7的分离、鉴定及毒力基因分析。结果表明,上海市动物及动物产品中存在大肠埃希菌O 157∶H 7,检出率为2.05%。其中牛粪、猪粪中检出率较高,分别为9.76%和8.57%。 相似文献
7.
为了解O157∶H7型大肠埃希菌中成簇的规律间隔短回文重复序列（clustered regularly interspaced short palindromic repeats,CRISPR）分布和结构特征及cas基因分布情况,本试验通过GenBank数据库和CRISPRdb database获得92株O157∶H7型大肠埃希菌全基因组序列、CRISPR位点位置、CRISPR的侧翼序列及cas基因簇的序列范围,利用多序列比对、启动子预测和RNA二级结构预测等方法分析细菌中CRISPR系统的特点。结果显示,O157∶H7型大肠埃希菌的基因组存在3个CRISPR位点（CRISPR1、CRISPR2和CRISPR3）,每个CRISPR位点上的序列一致;CRISPR1和CRISPR2的重复序列可形成茎环状结构,环上的碱基易发生变化;CRISPR2中存在一段长451 bp的序列（命名为序列X）,该序列X将CRISPR2分为2个部分CRISPR2a和CRISPR2b,其碱基A和T比例为74%,在91株O157∶H7型大肠埃希菌中均存在该序列,在该序列中可预测出至少有1个启动子和9个转录因子结合位点;侧翼序列中的疑似前导序列位于CRISPR2下游,序列长340 bp,其碱基A和T比例为69%,在92株O157∶H7型细菌中均存在该序列,其可预测出至少有1个启动子和3个转录因子结合位点;在20株O157∶H7型大肠埃希菌的全基因组序列中,有15株具有完整的cas基因簇,有5株缺乏cas3基因。本试验结果表明,O157∶H7型大肠埃希菌的CRISPR系统结构稳定,序列具有较高保守性,cas基因簇也相对保守。CRISPR2的结构与其他类型大肠埃希菌有较大差别。本研究发现的序列X在O157∶H7型大肠埃希菌分布广泛且序列保守,可作为鉴定O157∶H7型大肠埃希菌的潜在分子靶标。 相似文献
8.
《中国畜牧兽医》2020,(2)
为了解O157∶H7型大肠埃希菌中成簇的规律间隔短回文重复序列(clustered regularly interspaced short palindromic repeats,CRISPR)分布和结构特征及cas基因分布情况,本试验通过GenBank数据库和CRISPRdb database获得92株O157∶H7型大肠埃希菌全基因组序列、CRISPR位点位置、CRISPR的侧翼序列及cas基因簇的序列范围,利用多序列比对、启动子预测和RNA二级结构预测等方法分析细菌中CRISPR系统的特点。结果显示,O157∶H7型大肠埃希菌的基因组存在3个CRISPR位点(CRISPR1、CRISPR2和CRISPR3),每个CRISPR位点上的序列一致;CRISPR1和CRISPR2的重复序列可形成茎环状结构,环上的碱基易发生变化;CRISPR2中存在一段长451 bp的序列(命名为序列X),该序列X将CRISPR2分为2个部分CRISPR2a和CRISPR2b,其碱基A和T比例为74%,在91株O157∶H7型大肠埃希菌中均存在该序列,在该序列中可预测出至少有1个启动子和9个转录因子结合位点;侧翼序列中的疑似前导序列位于CRISPR2下游,序列长340 bp,其碱基A和T比例为69%,在92株O157∶H7型细菌中均存在该序列,其可预测出至少有1个启动子和3个转录因子结合位点;在20株O157∶H7型大肠埃希菌的全基因组序列中,有15株具有完整的cas基因簇,有5株缺乏cas3基因。本试验结果表明,O157∶H7型大肠埃希菌的CRISPR系统结构稳定,序列具有较高保守性,cas基因簇也相对保守。CRISPR2的结构与其他类型大肠埃希菌有较大差别。本研究发现的序列X在O157∶H7型大肠埃希菌分布广泛且序列保守,可作为鉴定O157∶H7型大肠埃希菌的潜在分子靶标。 相似文献
9.
徐尚荣 《青海畜牧兽医杂志》2008,38(4)
O157:H7大肠埃希氏菌是一种新发现病原微生物,感染大肠杆菌O157:H7主要引起婴幼儿腹泻,出血性结肠炎(haemorrhagic colitis,HC)、溶血性尿毒综合症(Haemolytic Uraemic Syndrome,HUS)、血栓性血小板减少性紫癜(thrombocytopenlc purpura,rrP)及死亡等[1-4]. 相似文献
10.
《动物医学进展》2015,(9)
为了建立一种同时检测O157、O26、O45、O103、O111、O121和O145等7种血清型的出血性大肠埃希菌的高通量检测方法,通过筛选比对大肠埃希菌O157血清型的fbE基因,O26、O103和O111血清型的WZX基因,O45、O121和O145血清型的WZY基因,设计了针对各个血清型的特异性引物和探针。通过在所有的特异性引物5′端加入超级引物的策略,达到了通过一次PCR反应,同时多重扩增7个血清型的7个目标片段的效果。将加尾多重扩增与液相芯片高通量检测相结合,建立了同时检测7种血清型的出血性大肠埃希菌的液相芯片检测方法,并对方法检测体系及反应条件进行了优化。所建立的7种出血性大肠埃希菌液相芯片检测方法灵敏,其灵敏度与荧光PCR的灵敏度相差100倍。所建立的方法特异,单个血清型的探针与相应的PCR扩增的阳性产物之间均有特异性的杂交,LQRR值介于17~63之间。7种血清型的混合探针与各个血清型菌株的PCR与扩增的阳性产物之间均有特异性的杂交,LQRR值介于11~23之间,与其他血清型出血性大肠埃希菌和非出血性大肠埃希菌均无交叉反应。试验结果表明,所建立的液相芯片检测大肠埃希菌O157、O26、O45、O103、O111、O121和O145等7个血清型的检测方法具有快速、特异性强、灵敏度高等特点,可用于主要出血性大肠埃希菌的快速鉴别检测。 相似文献
11.
应用测试片快速检测食品中的大肠杆菌O157：H7 总被引：1，自引：1，他引：0
目的应用大肠杆菌O157:H7测试片快速检测食品中的大肠杆菌O157:H7。方法对大肠杆菌O157:H7测试片的各项指标及影响因素进行测试,并将其应用于食品检测。结果大肠杆菌O157:H7测试片的检测灵敏度高,其对纯菌的检测低限可达3cfu/mL;特异性较强,与鼠伤寒沙门氏菌、志贺氏菌等21种非目的菌无交叉反应;快速,24h内可报告阴性检测结果。应用该测试片检测各种食品206份,检测结果与SN标准的符合率达到98.5%。结论应用测试片检测食品中的大肠杆菌O157:H7具有快速、方便、经济、无需昂贵设备等优点。该测试片可适应于食品中大肠杆菌O157:H7的快速初筛。 相似文献
12.
目的建立一种能同时检测沙门氏菌和大肠杆菌O157:H7的双重荧光PCR方法,应用于动物源性食品的快速检验。方法根据沙门氏菌invA基因和大肠杆菌O157:H7 RFBE基因的保守序列,设计引物和探针,通过优化反应条件,建立可同时检测沙门氏菌和大肠杆菌O157:H7的双重荧光PCR方法,应用于动物源性食品的检验,并与miniVIDAS快速初筛方法和SN标准方法进行比较。结果本研究建立的双重荧光PCR方法可同时快速检测沙门氏菌和大肠杆菌O157:H7,对纯菌的检测灵敏度均低于10CFU/双重荧光PCR反应体系。应用本方法检测36株标准/参考菌株,结果只有9株目的菌标准/参考菌株出现特异性扩增,其余27株非目的菌均呈阴性反应。定量检测重复性试验结果,批内和批间的变异系数均小于2%。应用本方法检测人工染菌样品,结果与miniVIDAS和SN方法检测结果一致,但检测时间比miniVIDAS快了3倍,比SN标准快了10多倍。结论本研究建立的双重荧光PCR方法具有快速、灵敏、特异、重复性好的优点,可在8小时内完成样品沙门氏菌和大肠杆菌O157:H7的检验。 相似文献
13.
根据大肠杆菌Ｏ１５７∶Ｈ７的编码ｅａｅ蛋白的ｅａｅＡ基因和大肠杆菌编码Ｈ７抗原的ｆｌｉＣ基因的核甘酸序列,合成了２对寡核苷酸引物,建立了一个检测大肠杆菌Ｏ１５７∶Ｈ７的ＰＣＲ方法。对１１株已知大肠杆菌Ｏ１５７∶Ｈ７（ＮＭ;无运动性）株和其他不同属的４２株已知肠道致病菌的检测结果表明,该方法只从大肠杆菌Ｏ１５７∶Ｈ７（ＮＭ）株的ＤＮＡ中产生预期的扩增产物,而从其他菌株的ＤＮＡ中未扩增出任何ＤＮＡ产物。该方法从基因水平直接确定大肠杆菌的血清型,特异性强,克服了以往血清学方法有非特异性反应的缺陷,为检测和鉴定大肠杆菌Ｏ１５７∶Ｈ７（ＮＭ）提供了一个新方法。 相似文献
14.
根据GenBank公布的大肠杆菌O157∶H7的Flic(H7)基因序列进行同源性比较分析,选择保守序列设计一对特异性扩增引物,通过优化反应条件,建立一个用于大肠杆菌O157∶H7快速定量检测的实时定量PCR方法。该方法的最低检测极限是103CFU/mL,敏感性比常规PCR提高10倍。方法重复性好、特异性强,重复性检测的变异系数均小于2%;只能检测大肠杆菌O157∶H7,对非大肠杆菌O157∶H7血清型细菌、猪链球菌2型、副猪嗜血杆菌无反应。利用此方法对模拟样本进行定量检测,其结果与平板细菌计数基本一致,表明此方法可作为大肠杆菌O157∶H7快速诊断和疫情监测的一种快速、准确、简便的检测工具。 相似文献
15.
A multiplex real-time PCR (R-PCR) assay was designed and evaluated on the ABI 7700 sequence detection system (TaqMan) to detect enterohemorrhagic Escherichia coli (EHEC) O157:H7 in pure cultures, feces, and tissues. Three sets of primers and fluorogenic probes were used for amplification and real-time detection of a 106-bp region of the eae gene encoding EHEC O157:H7-specific intimin, and 150-bp and 200-bp segments of genes stx1 and stx2 encoding Shiga toxins 1 and 2, respectively. Analysis of 67 bacterial strains demonstrated that the R-PCR assay successfully distinguished EHEC O157:H7 serotype from non-O157 serotypes and provided accurate profiling of genes encoding intimin and Shiga toxins. Bacterial strains lacking these genes were not detected with this assay. The detection range of the R-PCR assay for the three genes was linear over DNA concentrations corresponding from 10(3) to 10(8)CFU/ml of EHEC O157:H7. The R-PCR allowed construction of standard curves that facilitated quantification of EHEC O157:H7 in feces and intestinal tissues. Detection sensitivity of the R-PCR assay ranged from 10(4) to 10(8)CFU/g of feces or tissues without enrichment. Enrichment of feces in a non-selective broth for 4 and 16h resulted in the detection of levels (from 10(0) to 10(3)CFU/g of feces) considered sufficient for infection in humans. The R-PCR assay for eae(O157:H7), stx1, and stx2 proved to be a rapid test for detection of EHEC O157:H7 in complex biological matrices and could also potentially be used for quantification of EHEC O157:H7 in foods or fecal samples. 相似文献
16.
D R Hyatt J C Galland J R Gillespie 《Journal of veterinary diagnostic investigation》2001,13(1):71-73
The performance of a commercially available enzyme immunoassay (EIA) for determining the presence of Shiga toxin I and II in human diarrheal stool samples was evaluated for use as a presumptive test for the presence of Escherichia coli O157:H7 in nondiarrheal bovine fecal samples collected from 10 Kansas cow-calf ranches. The prevalence of E. coli O157:H7 in 2,297 samples, as determined by selective bacterial culture, was 1.6%. The sample prevalence of non-E. coli O157:H7 Shiga toxin-producing bacteria, as detected by the Shiga toxin EIA, was 5.8%. Only 2 of 136 samples that tested positive with the Shiga toxin EIA were positive for E. coli O157:H7 by culture. Compared with bacterial culture, the sensitivity of the Shiga toxin EIA was 5.5% and the specificity was 94.1%. Agreement between the 2 tests, as measured by the kappa statistic, was poor (kappa = -0.002). Although the Shiga toxin EIA was not a good presumptive test for the determination of E. coli O157:H7 in bovine fecal samples because of its low sensitivity (5.5%), it might be a useful test for the detection of Shiga toxin producing non-E. coli O157:H7 organisms in bovine feces. 相似文献
17.
Sargeant JM Gillespie JR Oberst RD Phebus RK Hyatt DR Bohra LK Galland JC 《American journal of veterinary research》2000,61(11):1375-1379
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. 相似文献
18.
J M Sargeant D J Hafer J R Gillespie R D Oberst S J Flood 《Journal of the American Veterinary Medical Association》1999,215(6):792-794
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. 相似文献
19.
Richard D Oberst Michael P Hays Lalit K Bohra Randall K Phebus Jan M Sargeant 《Journal of veterinary diagnostic investigation》2003,15(6):543-552
Currently, methods for recovering and identifying Escherichia coli O157:H7 from cattle feces are inconsistent and hindered by their inability to specifically and rapidly detect small numbers of organisms from this complex and highly variable matrix. A standard approach for isolating and characterizing E. coli O157:H7 from cattle feces was compared with a polymerase chain reaction (PCR)-based 5' nuclease assay specific for E. coli O157:H7 that included a secondary enrichment step. The PCR-based method proved a better indicator of the presence of the organism than the culture procedure. Retests indicated that the inclusion of a secondary enrichment step and the subsequent analysis by the 5' nuclease assay were reproducible and specific. Escherichia coli O157:H7 could be detected in fecal samples that were otherwise negative after a primary enrichment step, immunomagnetic separation, and plating onto sorbitol MacConkey agar plates containing cefixime and tellurite (CT-SMAC). In samples that were initially identified as culture positive but PCR negative, retesting of the culture isolates on CT-SMAC indicated that the sorbitol fermentation interpretations could frequently not be repeated in retests, whereas retesting using the 5' nuclease assay on the original samples demonstrated a high level of agreement with the initial PCR conclusions. These results indicate the necessity of confirmatory evaluation of isolates culturally recovered by standard cultural methods that involve the interpretation of CT-SMAC. The high level of disagreement between initial culture results and retests, and the high level of agreement between initial PCR results and retests, indicates the advantages of a gene-based detection system for identifying E. coli O157:H7 in cattle feces. Screening large numbers of fecal samples for E. coli O157:H7 would appear to be feasible by integrating the use of enrichment media in serial rounds of incubation with a PCR-based fluorogenic detection procedure in high throughput detection systems that had automated liquid-handling capabilities. 相似文献
20.
肠出血性大肠杆菌O157∶H7是一种重要的人畜共患传染病病原菌。为建立一种特异、灵敏的O157∶H7新型检测技术,以O157抗原基因(rfbE基因)为模板设计特异性引物,利用叠氮溴化乙锭(ethidium monoazide bromide,EMA)处理菌液,沸水浴法制备细菌裂解液,优化PCR条件,建立一种快速、有效的O157∶H7活菌EMA-PCR检测方法。结果显示:EMA-PCR可从rfbE基因阳性菌株CVCC248和两株临床分离菌株cd0912、cd0803中扩增出大小为495 bp的特异性条带,检测灵敏度可达12 CFU/mL。经EMA处理,从含有1%～100%O157∶H7 CVCC248活菌混合悬液制备的DNA中均可扩增出目的片段。因此,成功建立了肠出血性大肠杆菌O157∶H7的EMA-PCR检测方法;该方法可避免因分析的样品中含有死细菌而造成的假阳性检测结果,检测的准确性和真实性较传统PCR大大提高。O157∶H7 EMA-PCR技术的建立为O157∶H7的临床诊断提供了新的方法,具有重要的实际应用价值和良好的应用前景。 相似文献