多重PCR方法检测大肠杆菌O157:H7的初步研究

目的研究一种快速、特异的检测大肠杆菌O157：H7的多重PCR方法。方法以大肠杆菌O157：H7的O157抗原基因（rfbE基因）、鞭毛H7抗原基因（fliC基因）、粘附素基因（eaeA基因）和志贺样毒素Ⅰ、Ⅱ（SLT-Ⅰ、SLT-Ⅱ基因）为扩增对象，设计能导至目的片段大小不同的5对引物，通过优化条件，建立了可用于粪便检测的大肠杆菌5重PCR。结果在同一扩增体系中，检测了5株不同来源的大肠杆菌O157：H7及27株非大肠杆菌O157：H7细菌。结果表明，该方法能从2株大肠杆菌O157：H7中扩增出rfbE、fliC、eaeA、SLT-Ⅰ、SLT-Ⅱ基因，它们的大小分别为582bp、802bp、487bp、368bp和177bp。而另外3株大肠杆菌O157：H7也能扩增出除eaeA、SLT-Ⅱ基因外的其它3个基因片段。表明该方法可用于检测大肠杆菌O157：H7，还可检测细菌是否含有毒素基因。在检测27株非大肠杆菌O157：H7．除O149出现SLT-Ⅱ扩增产物和01：H7出现flic基因扩增产物外，其它非大肠杆菌O157：H7的扩增结果均为阴性，表明该方法有较高的特异性。当粪便样品经增菌培养12h，用该5重PCR体系能检测出最初接种量为5cfu／g粪便的样品。结论本5重PCR方法具有较高的特异性和敏感性，可迅速、有效地将大肠杆菌O157：H7与其它非大肠杆菌O157：H7相区别，同时还能检测O157：H7中的毒力因子。因此，通过进一步研究，本方法有望应用于临床大肠杆菌O157：H7感染的辅助诊断。     

2012年东台市肠出血性大肠埃希菌O157：H7监测分析

目的了解肠m血性大肠埃希菌O157：H7在东台市动物宿主和腹泻患者中带菌及毒力基因携带与耐药情况。方法于流行季节采集动物宿主、腹泻患者粪便及生熟食品和苍蝇标本,mEC肉汤增菌后,川特异性免嫂磁珠集菌、科玛嘉显色平板分离,纯化的菌株经生化鉴定,血清分型,以K—B纸片法进行药敏试验,采用PCR方法检测uida缺闪和stx1、stx2、eaeA、hly等4种毒力基因。结果683份标本共检m0157：H7菌株35株,检出率为5．12％,疑小动物粪便检出率为10．59％（34／321）,检出率最高的为牛粪（26．92％）,其次为山羊粪（7．50％）和猪粪（7．32％）,腹泻患者的检出率为0．39％（1／256）。35株菌株均检meaeA和hly毒力基因,而牛粪中stx2、eaeA和hly等3种毒力基因组合型检出率达12．82％。35株菌株对氨苄西林敏感率为14．3％,对其余17种抗牛素敏感率均〉90％。结论东台市动物宿主和腹泻患者均不同程度携带肠出血性大肠埃希菌O157：H7,且存在流行的潜在危险。     

肠出血性大肠杆菌O157：H7实验室诊断技术进展

肠出血性大肠杆菌O157：H7（E．coli O157：H7）是肠出血性大肠杆菌（EHEC）的一个血清型，主要引起人的食源性疾病，以突发性腹痛、腹泻、血便为主要症状，严重时并发肾功能衰竭，病死率高。E．coli O157：H7广泛分布于自然界，在生猪、猪肉、牛及牛肉及其粪便中分离率极高，对公众健康构成严重威胁。因此，早期发现及时治疗和预防是控制该病的关键。E．coli O157：1-17实验室诊断以细菌培养、分离及生化和血清鉴定为主。近年来出现的新型免疫学方法和聚合酶链反应（PCR）对E．coli O157：H7诊断具有较高的敏感性和特异性。本文就E．coli O157：H7检测技术的研究进展综述如下。     

抗出血性大肠杆菌O157:H7单克隆抗体的制备及鉴定

目的制备抗出血性大肠杆菌O157：H7(E．coli O157：H7)特异性单克隆抗体(MAbs)。方法福尔马林灭活的E．coli O157：H7免疫BALB／c小鼠，利用细胞融合技术建立分泌抗E．coli O157：H7 MAbs的杂交瘤细胞株，对配对较好的6株MAbs用ELISA法测定其免疫球蛋白类及亚类，用ELISA法、凝集法和Western blot鉴定MAbs的特异性。结果6株MAbs免疫球蛋白均为小鼠IgM。这些MAbs均能与27个E．coli O157：H7菌株发生凝集反应。与部分弗劳地杆菌发生凝集反应，与11株鼠伤寒沙门氏菌、7株伤寒杆菌、2株痢疾杆菌、致病性大肠杆菌、产毒性大肠杆菌、侵袭性大肠杆菌、出血性大肠杆菌O26：H11和O111、肠集聚性大肠杆菌、42株非定血清型大肠杆菌、霍乱弧菌O1群和O139群不发生凝集反应；ELISA结果显示6株MAbs与粪链球菌、变形杆菌、粘质沙雷氏菌、肺炎克雷伯杆菌均无交叉反应；ELISA和Western blot结果显示。3株MAbs针对E．coli O157：H7酚相脂多糖。结论6株MAbs具有较高的特异性，有可能用于制备检测E．coli O157：H7的病原检测试剂。     

大肠埃希菌O157:H7脂肪酸组分及DNA特征研究

目的探讨大肠埃希菌O157：H7脂肪酸组分及DNA指纹图谱特征。方法用气相色谱（Gaschromatography，GC）及随机扩增多态性DNA（Randomly amplified polymorphyi cDNA，RAPD）对2株大肠埃希菌O157：H7和其他6株大肠埃希菌全细胞脂肪酸组分及DNA指纹图谱特征进行分析。结果大肠埃希菌O157：H7不仅含有15：0和17：0脂肪酸组分，不含或仅含微量14：02OH和19：0w8c脂肪酸，而且sum4和sum7脂肪酸峰值也明显高于其他菌株，DNA指纹谱也显示了其独有特征。结论大肠埃希菌O157：H7脂肪酸组分及相对含量和DNA指纹图谱特征与O26：H11及其他大肠埃希菌显著不同，与大肠埃希菌O26：H11等在遗传进化关系上存在一定距离。     

应用多重PCR方法检测大肠埃希菌O157：H8的初步研究

目的 研究一种快速、特异的检测大肠埃希菌（以下称大肠杆菌）O157：H7的复合PCR方法。方法 选用针对大肠杆菌O157：H7志贺样毒素Ⅰ、Ⅱ（SLT-Ⅰ、SLT-Ⅱ）和溶血素（Hly)基因的三对引物,在同一扩增体系中进行PCR,检测12株不同来源的O157：H7大肠杆菌及其它致病性大肠杆菌及沙门菌、志贺菌15株。结果 除质粒缺失株（933）外,其余11株O157：H7大肠杆菌均在361bp处有溶血素基因产物出现;而12株O157：H8大肠杆菌扩增后的两条志贺样毒素基因产物存在差异,其中6株在210bp、484bp处出现两条相应产物,3株仅有210bp一条SLT－Ⅰ基因产物,另3株仅有484bp的SLT－Ⅱ基因产物。其它致病性大肠杆菌及沙门菌、志贺菌的扩增结果均为阴性。结论 本复合PCR方法具有较高的特异性,可迅速、有效地将O157：H7大肠杆菌与其它常见致病性大肠杆菌及沙门菌、志贺菌相鉴别,通过进一步研究有望应用于临床大肠杆菌O157：H7感染的辅助诊断。     

肠出血性大肠埃希菌(O157:H7)的基因同源性的分析

目的 对江苏省徐州地区O157：H7的病原学进行分析。方法 采用聚合酶链反应对O157：H7菌株毒力基因谱进行检测，同时用脉冲凝胶电泳(PFGE)和随机扩增多态性DNA(RAPD)方法对O157：H7菌株的同源性分析比较。结果 流行地区分离的O157：H7菌株，100％携带Hly、eaeA基因，95．35％携带SLT2基因，11．63％携带SLT1基因。脉冲凝胶电泳图谱表明流行地区分离的O157：H7菌株与日本分离的O157：H7菌株有明显差异，为不相关菌株；与国内标准菌株882364为近似型(相似，但不相同)。流行地区患者分离菌株与外环境家畜家禽粪便及昆虫肠道分离菌株的脉冲凝胶电泳图谱完全相同。结论 携带O157：H7菌株的家畜家禽可能是导致疫情发生的传染源。脉冲凝胶电泳方法用于O157：H7病原学分析，对流行病学研究有重要意义。随机扩增多态性DNA方法用于O157：H7病原学分析，技术简便、省时。     

肠出血性大肠杆菌O157：H7脂多糖抗原的制备及鉴定

目的提取纯化肠出血性大肠杆菌（EHEC）O157：H7脂多糖（LPS）抗原并对其鉴定,以探讨其应用于检测抗O157抗体的价值。方法应用热酚水法提取EHECO157：H7S型933株LPS抗原,并经超速离心纯化。提取物以蒽酮硫酸法、紫外光谱吸收法、考马斯亮蓝染色法及鲎试剂分别测定多糖、核酸、蛋白含量和尝试剂凝集活性。LPS抗原致敏醛化鸡血球,IHA法检测EHEC157：H7及其它肠道病原菌的抗血清。对凝集阳性的抗血清,经菌体抗原吸收、O157LPS抑制及去脂质多糖抗原抑制试验,确定抗血清与LPS的反应位置。结果纯化后的O157LPS抗原告多精,具鲎试剂凝集活性,未检出核酸,蛋白含量＜0．5％。O157LPSIHA检测O157多克隆和单克隆抗体均呈高效价凝集阳性反应,而大部分其它肠道病原菌抗血清均呈阴性反应,交叉反应仅见于沙门氏菌O30、EIECO144：K？及ESIECⅣ抗血清。EHEC157和沙门氏菌O30抗血清与O157LPS反应位置为多糖部分,而EIECO144：K？及ESIECⅣ抗血清的反应位置为脂质A部分。结论热酚水法提取的EHECO157LPS抗原纯度高、特异性强,可望应用于抗O157抗体的检测。O157LPS抗原的去脂质可进一步提高反应的特异性。     

从同一份市售活鸡中检出E.coli O157:H7和沙门菌的报告

E．coli O157：H7和沙门菌均为致病菌，它们可以通过食物、水源污染和接触传播使人类感染致病或食物中毒，各国食品安全卫生指标对这些致病菌均规定禁止检出。近日，黄石市疾病预防控制中心中心实验室在进行食品污染物食源性致病菌的检测中，在市售的一份活鸡中同时检出了E．coli O157：H7和沙门菌两种致病菌。现将有关情况报告如下：     

在奶牛粪便中检出O157：H7大肠埃希菌的报告

从奶牛粪便中检出1株O157：H7大肠埃希菌，但未发现人与其他家禽、家畜感染及食品污染。     

The isolation rate of Escherichia coli 0157:H7 in Toronto and surrounding communities

Verocytotoxin-producing strains of Escherichia coli, most often serotype 0157:H7, have been associated with both sporadic and epidemic diarrheal disease in Canada. In order to determine the isolation rate of E coli 0157:H7 in outpatients with diarrhea, all stool specimens submitted for culture to Med-Chem Laboratories in Metropolitan Toronto between June 1988 and September 1989 were cultured on MacConkey-Sorbitol agar in addition to standard enteric media. A total of 46 (0.3%) of 16,125 stool specimens yielded E coli 0157:H7 or verotoxin-producing E coli 0157:H(-). These isolates came from 31 patients with diarrhea; only 16 (52%) had a history of hemorrhagic colitis and one patient developed hemolytic uremic syndrome. Although MacConkey-Sorbitol agar was useful as a differential medium for detecting E coli 0157:H7, 14.5% of all specimens yielded nonsorbitol-fermenting isolates. It is not certain whether the routine use of MacConkey-Sorbitol agar is justified when isolation rates of E coli 0157:117 are very low.     

Verocytotoxin-producing Escherichia coli EH 0157:H7 colitis

0157:H7 is a known etiologic agent of hemorrhagic colitis. The clinical and histologic picture of colitis is largely similar to that of ischemic colitis, with areas of submucosal hemorrhage and edema, erosions, and ulcerations. We present a case report and review of the literature. A 52-year-old HIV-positive man, in apparently good immunologic condition, developed severe hemorrhagic colitis characterized by the onset of multiple colonic perforations and an unfavorable outcome. The diagnosis of 0157:H7 colitis should therefore be considered in all patients with indeterminate hematic diarrhea. Further studies are warranted to verify whether HIV infection may play a determinant role in the clinical course of 0157:H7 infection.     

Clinical and laboratory patterns of the haemolytic uraemic syndrome and thrombotic thrombocytopenic purpura in southern Iran

OBJECTIVE: The haemolytic uraemic syndrome (HUS) and thrombotic thrombocytopenic purpura (TTP) are rare disorders characterised by intravascular platelet aggregation and widespread thrombus formation in the microcirculation resulting in tissue ischaemia. A retrospective analysis was carried out in 136 patients with HUS or TTP hospitalized from April 1991 through March 2004 in three tertiary referral hospitals of Shiraz (the largest city in southern Iran) to evaluate the epidemiological aspects, clinical characteristics and laboratory findings of the two diseases. METHODS: One hundred and one cases of HUS (49 females and 52 males) and 35 cases of TTP (21 females and 14 males) were identified. The mean age was 3.5 years for HUS, 30.8 years for TTP. RESULTS: The mean annual incidence rate of HUS decreased approximately 30-fold throughout the observed period, while the incidence rate of TTP increased approximately 6-fold. A seasonal pattern was noted for both TTP and HUS, with the highest incidence during the summer months. Twenty patients with HUS and 16 patients with TTP died, resulting in case fatality rates of 19.8% and 45.7%, respectively. No prognostic factor was identified for TTP, whereas signs of neurological impairment and high leucocyte counts had an adverse effect on the prognosis of HUS patients. CONCLUSIONS: This study shows that, in contrast to other countries, the incidence rate of HUS shows a decreasing trend in southern Iran, probably related to the prevention of gastrointestinal diarrhoeal infections (especially Escherichia coli 0157:H7) and to their improved management. The incidence of TTP is increasing in Iran as in other countries.     

Simple colonoscopy reporting system checking the detection rate of colon polyps

AIM: To present a simple colonoscopy reporting system that can be checked easily the detection rate of colon polyps.METHODS: A simple colonoscopy reporting system Kosin Gastroenterology (KG quality reporting system) was developed. The polyp detection rate (PDR), adenoma detection rate (ADR), serrated polyp detection rate (SDR), and advanced adenoma detection rate (AADR) are easily calculated to use this system.RESULTS: In our gastroenterology center, the PDR, ADR, SDR, and AADR test results from each gastroenterologist were updated, every month. Between June 2014, when the program was started, and December 2014, the overall PDR and ADR in our center were 62.5% and 41.4%, respectively. And the overall SDR and AADR were 7.5% and 12.1%, respectively.CONCLUSION: We envision that KG quality reporting system can be applied to develop a comprehensive system to check colon polyp detection rates in other gastroenterology centers.     

High incidence of haemorrhagic colitis due to Escherichia coli 0157 in one Scottish town: Clinical and epidemiological features

Verotoxin-producing strains of Escherichia coli (VTEC), in particular serotype 0157:H7, are now recognised as the major cause of haemorrhagic colitis and the haemolytic uraemic syndrome (HUS) in the U.K. and in North America, and increasingly so in other countries. Over a 3-year period (1989–1991), 16 cases of E. coli 0157 infection occurred in one town (Peterhead) in north-east Grampian. Four patients required admission to hospital, of whom three developed HUS. The bovine source of VTEC infection has now been clearly established with foodborne, waterborne, person-to-person and zoonotic transmission described. Despite extensive local enquiries, the source(s) of infection of the 16 cases in Peterhead was not established. Much still needs to be learned about the epidemiology, risk factors and long-term clinical sequelae of VTEC infection and HUS. Close collaboration between the medical and veterinary professions is of paramount importance in order to provide better understanding of the prevalence of E. coli 0157 infection in cattle and the route(s) of transmission to humans.     

冠状动脉心肌桥的检出率及临床分析

目的:分析冠状动脉心肌桥的检出率及其临床特点。方法:回顾性分析冠状动脉造影检测出的53例心肌桥病例的临床资料。结果:冠状动脉造影发现心肌桥的检出率为3.29%(53/1610),心肌桥患者并发冠心病的发生率约为26.4%,孤立性心肌桥与心肌桥并发冠心病在传统的冠心病易患因素方面无统计学差异(P0.05),但在收缩期壁冠状动脉狭窄程度与心肌桥长度方面差异有显著统计学意义(P0.01)。结论:经冠状动脉造影,心肌桥的检出率为3.29%,心肌桥患者心肌桥长度与收缩期壁冠状动脉狭窄程度可能是心肌桥并发冠心病的危险因素。     

Symptoms of food-borne diseases and gastroenteritis in Kyushu, Japan

In this study we analyzed the symptoms of gastroenteritis or food-borne disease caused by the 10 most prevalent pathogens: Norovirus, Salmonella, Vibrio parahaemolyticus, Campylobacter jejuni, Clostridium perfringens, Shiga toxin-producing Escherichia coli (STEC), enterotoxigenic E. coli (ETEC), Shigella sonnei/flexneri (Shigella), Staphylococcus aureus, and emetic-type Bacillus cereus. The symptoms diarrhea, vomiting, fever, abdominal pain, and headache, and the incubation period in 646 cases in 10 districts of Kyushu between January 2000 and December 2004 were recorded. The pathogen with the shortest mean incubation period was B. cereus (0.8 h), and was followed by S. aureus (3.3 h), C. perfringens (10.7 h) and V. parahaemolyticus (16.4 h). All the patients infected with B. cereus and S. aureus developed symptoms within 6 hours, and those infected with V. parahaemolyticus and C. perfringens developed symptoms within 24 hours. Bloody diarrhea was associated with STEC and Shigella, but rare with other pathogens. Vomiting was associated with almost all cases of S. aureus and B. cereus infection, and occurred in 71.5% of the Norovirus cases and 56.1% of the V. parahaemolyticus cases. Vomiting was less common in the C. perfringens (22.0%) and the ETEC and STEC (both about 5%). Bloody diarrhea, abdominal pain, and vomiting were statistically significantly more common with STEC 0157 infection than with STEC non-0157 infection. Since the cases analyzed in this study included all degrees of illness, mild to severe, and a wide range of ages, the information obtained will serve as a good reference material for administrative and laboratory work when an outbreak takes place.     

Study on Chlamydia trachomatis antigen detection in first-voided urine sediments

C. trachomatis antigen in first-voided urine sediments was detected by a new EIA kit using a monoclonal antibody, IDEIA CHLAMYDIA (IDEIA, Novo Nordisk), in males with urethritis and females with cervicitis. The result was compared with that by Chlamydiazyme (Abbott). 1. C. trachomatis antigen detection in male urethritis (285 cases) by the IDEIA test: The antigen detection rate was 37.9% (108/285) in urethral smears, and 33.7% (96/285) in first-voided urine sediments of the patients. The positive co-incidence rate between urethral smears and first-voided urine sediments was 82.4% (98/108). Thus, the detection of the antigen seems feasible in first-voided urine sediments. 2. Comparison of C. trachomatis antigen detection by the IDEIA and Chlamydiazyme tests: In 78 male cases with urethritis undergoing both tests, the rates of antigen detection from urethral smears and first-voided urine sediments were studied. The detection rate from urethral smears was 41.0% (32/78) for IDEIA, and 37.2% (29/78) for Chlamydiazyme. In first-voided urine sediments, the rate was 35.9% (28/78) for IDEIA and 24.4% (19/78) for Chlamydiazyme. In both specimens, the detection sensitivity was higher for IDEIA. 3. C. trachomatis antigen detection in chlamydial cervicitis (28 cases) by the IDEIA test: The antigen detection rate was 46.4% (13/28) in urethral smears and 60.7% (17/28) in first-voided urine sediments. The detection rate in first voided urine sediments was higher. Thus, in patients suspected of having chlamydial cervicitis, it seems necessary not only to search the antigen in cervical smears but also to study the first-voided urine sediments.(ABSTRACT TRUNCATED AT 250 WORDS)