广州海珠地区非O1/非O139群霍乱弧菌流行状况调查及生物学特征研究

目的 了解广州海珠地区2001-2009年非O1/非O139群霍乱弧菌的流行、分布状况及生物学特征,为防控该类病原菌引起的急性腹泻病提供科学依据.方法 参照<霍乱防治手册>(第5版)中的检测方法,对本区珠江河水、饮用水、养殖水、各种海(水)产品、市售熟食等外环境标本764份、健康人群标本189份及肠道门诊腹泻患者标本3398份,共计4351份标本进行非O1/非O139群霍乱弧菌的检定.结果 4351份标本共检出非O1/非O139群霍乱弧菌101株,总检出率为2.32%;其中有66株可分型,分属于26个血清型,分型率为65.3%;VBO9、VBO38和VBO76为优势菌.外环境与腹泻患者标本中分离到9株同型别非O1/非O139群霍乱弧菌,提示两者可能存在关联.结论 非O1/非O139群霍乱弧菌在本地区的血清型分布呈现多样性,在人群中有一定的感染率;外环境中该类菌群广泛存在,对人群是一个潜在的危害因素.     

应用胶体金免疫层析法检测水产品中霍乱弧菌的研究

〔目的〕应用胶体金免疫层析法,研究水产品中霍乱弧菌的快速检测方法。〔方法〕通过模拟水产品棉拭子标本比对试验及现场应用比对试验,比较不同检验方法的差异。〔结果〕70份增菌液中霍乱弧菌理论浓度为1.5×10CFU/ml的模拟水产品棉拭子标本分别按检验方法1、检验方法2检验,同时检出阳性51份,检验方法2的检出率(87%)显著高于检验方法1的检出率(74%);30份增菌液中霍乱弧菌理论浓度为1.5×101CFU/ml的“模拟水产品棉拭子标本”按检验方法2进行检验的检测结果均为阳性。对2000份水产品进行检测的现场应用对照实验结果显示:检验方法3的检出率(0.4%)与检验方法1的检出率(0.3%)差异无显著性;胶体金试纸条实验的假阳性率为8%。〔结论〕在胶体金免疫层析法实验中,增菌液增菌前霍乱弧菌含量为1CFU/ml,通过增菌后可被检出;增菌液霍乱弧菌含量为10CFU/ml,增菌12h后均可被检出;水产品棉拭子标本增菌12h检测阴性者可报告阴性,阳性者则参照《霍乱防治手册》[1]进一步分离鉴定。鱼贝类霍乱弧菌污染菌数较少(103CFU/100g以下)[2],无法满足使用胶体金免疫层析法所要求的含菌量(106CFU/ml)[3],可以通过改良增菌培养液成分,以提高待检菌繁殖能力,在短时间内提高增菌液待检菌浓度,使胶体金免疫层析法的灵敏性高于或接近常规的分离鉴定法。     

三峡库区长江水非O1群霍乱弧菌L型的研究

为了解三峡库区长江水非Ｏ１群霍乱弧菌Ｌ型的流行病学状况,１９９４￣１９９８年作者在三峡库区选三地取长江水同时进行非Ｏ１群霍乱弧菌与非Ｏ１群霍乱弧菌Ｌ型的检测。结果发现Ｏ１群霍乱弧菌Ｌ型冬天阳性检出率较高,夏天阳性检出率较低。但各月份总阳性率无明显差异。非Ｏ１群霍乱弧菌Ｌ型返祖回复后证实为非Ｏ１群霍乱弧菌。表明非Ｏ１群霍乱弧菌Ｌ型为非Ｏ１群霍乱弧菌越冬的一种重要形式。在卫生监督中,应加强对非为群霍乱     

2005年-2010年武汉市霍乱弧菌监测分析

目的:对2005年-2010年霍乱弧菌监测中分离的25株疑似菌株进行系统鉴定。方法:分离培养、血清学试验、系统生化鉴定(VITEK32)、PCR检测其分型及毒素基因、药敏试验。结果:25株菌鉴定为霍乱弧菌的有21株,与霍乱弧菌发生交叉凝集4株;病例监测主要为O139霍乱弧菌、外环境监测主要为O1群霍乱弧菌;病例监测菌株携带霍乱弧菌毒素基因(ctxAB)、其他菌株均不携带霍乱弧菌毒素基因(ctxAB)。药敏实验结果对诺氟沙星和环丙沙星敏感,对其他抗生素有不同程度的耐药。结论:武汉市霍乱弧菌在病例监测中主要为O139群霍乱弧菌产毒株、外环境监测中主要为O1群霍乱弧菌非产毒株,不同型别菌株存在不同程度耐药性。     

霍乱流行区江河霍乱弧菌污染的监测和消除研究

目的：全面掌握霍乱流行区江河段霍乱弧菌污染的状况并评价消除污染的措施及其效果。方法：于198年5月4日－7月1日对流行区A、B、C河流按每隔300－500米设立取水点，每2－3天采样1次；1998年7月－2001年12月对13个曾反复阳性点按月进行霍乱弧菌监测。采取对下水道污水消毒，拆除沿江两岸厕所和人群查源等措施以消除污染。结果：1998年4月28日－6月1日在流行区29个取水点检出霍乱弧菌。毒力测定人源株以中等毒力为主（68．8％），而水源株以弱毒为主（66．7％）。1998年7月至2001年12月采水796份，均未检出霍乱弧菌，24份水样L型菌，非可培养状态也为阴性。结论：该流行区江河段遭受一时性霍乱弧菌污染，1998年6月2日以后已彻底消除，未发生水型传播疫情，1998年8月－2001年12月，该区域也未再有霍乱病例发生，表明消除措施效果显著，也证明当地霍乱疫情系输入性。     

Surveillance methodology for Vibrio cholerae in environmental samples

The purpose of this study was to examine the prevalence of Vibrio cholerae in environmental water samples by using a series of biochemical tests. A total of 223 V. cholerae-like bacteria were isolated from TCBS agar after spreading the alkaline peptone water enriched sewer (n = 21) and water (n = 16) samples. All oxidase positive isolates were subjected to confirmation for V. cholerae by seven other biochemical tests and polymerase chain reaction. Only 74.2% isolates were found to be V. cholerae by PCR using primers against an outer membrane protein (ompW) gene, out of which only 2 isolates were positive for cholera toxin (ctxAB) gene. Among the various biochemical tests studied, arginine hydrolysis, arabinose fermentation and string test showed 92 - 100% sensitivity and 42 - 67% specificity. Eight isolates including the toxigenic ones, showed agglutination with V. cholerae O1 antiserum. The present study showed that no biochemical test is 100% specific for V. cholerae. However, a few tests, if performed in a sequence after growing the alkaline peptone water enriched samples onto TCBS media can be used for screening of V. cholerae from the environmental samples. This study also showed that most of the environmental isolates are non-O1/non-O139 and the chances of presence of toxigenic V. cholerae are very rare in the environment.     

非可培养状态霍乱弧菌的间接免疫荧光检测

目的建立检测水中活的非可培养状态霍乱弧菌(O1群、O139群)的间接免疫荧光法。方法用本实验室人工转化的非可培养状态霍乱弧菌的菌液制成抗原片,用混合诊断血清(抗O1群、O139群)作为一抗,异硫氰酸荧光素(FITC)标记猪抗兔IgG作为二抗,进行间接免疫荧光检测。结果该方法可以成功检测出水中非可培养状态霍乱弧菌,其检测下限约为104cells/400 ml。结论该方法具有较好的特异性和灵敏度,可作为自然水体中非可培养状态霍乱弧菌的检测方法,用于霍乱弧菌的生态研究。     

O1群霍乱弧菌胶体金法快速检测

目的 了解胶体金法检测O1群霍乱弧菌的特异性、灵敏度和实用性.方法 取患者粪便标本,先用碱性蛋白胨水35℃增菌6 h,然后用胶体金法进行检测,同时作霍乱弧菌常规培养,培养出细菌后用血清凝集法作为鉴别诊断O1群霍乱弧菌的金标准,用统计学方法对胶体金法诊断试验的评价指标进行计算..结果 700份样本中,常规法培养出O1霍乱弧菌105株,胶体金法试验阳性108株,经χ2检验,差异无统计学意义(χ2=0.24,P>0.05),表明胶体金法检测O1群霍乱弧菌与常规培养法检测结果之间的差异无统计学意义.胶体金法检测O1群霍乱弧菌诊塑试验的灵敏度为93.33%,特异性为98.32%,阳性预测值为90.74%,阴性预测值为97.18%,假阴性率为1.68%,假阳性率为6.67%.结论 胶体金法检测O1群霍乱弧菌特异性高(98.32%),灵敏度好,与常规培养法的一致性也较高,在快速检测O1群霍乱弧菌中有一定应用价值.     

1984-2002年重庆市59株O_1群霍乱临床分离株耐药性及分子分型研究

目的分析重庆市1984-2002年O1群霍乱临床分离株的耐药性和菌株的相关性。方法对59株O1群霍乱临床分离株(小川型20株,稻叶型39株)采用K-B法检测对16种抗菌药物的敏感性,用NotI酶切基因组DNA,经脉冲场凝胶电泳(PFGE)获得电泳图谱,利用BioNumerics软件对图谱进行聚类分析。结果重庆地区O1群霍乱弧菌对复方新诺明耐药、痢特灵和链霉素的耐药严重,耐药率分别为28.81%(17/59)、61.02%(36/59)和30.51%(18/59),人群分离株对丁胺卡那、庆大霉素、妥布霉素、氨苄西林、新霉素和强力霉素敏感,未发现耐药菌株。21株小川型和40株稻叶型O1群霍乱弧菌被分为13种PFGE型18个亚型,从患者分离到的59株O1群霍乱弧菌的相似性值约在88%～100%。结论未发现重庆市O1群霍乱弧菌人群分离株的耐药性有明显改变,小川型和稻叶型的耐药性不同。重庆市O1群霍乱弧菌人群分离株同源性较高,可能为来自同一来源的流行菌株。     

New qnr gene cassettes associated with superintegron repeats in Vibrio cholerae O1

A novel qnr determinant emerged in ciprofloxacin-resistant Vibrio cholerae O1 from the Amazon region of Brazil. This qnrVC1 was in a typical class 1 integron. Its attC showed 89% identity with V. parahaemolyticus superintegron repeats. Analysis showed V. cholerae O1 carrying qnrVC2 associated with a V. cholerae superintegron repeat.     

产色素非O1群霍乱弧菌的发现及其实验研究

１９９４年６月自饮用水源中分离到４株产色素的革兰氏阴性弧菌，运动活泼似流星样．经形态学、培养特性、生化学和血清学方面的系统鉴定，均符合非Ｏ１群霍乱弧菌定义．因该菌在营养琼脂、硷性胆盐琼脂和血琼脂上培育后产生水溶性褐色色素，故被正式命名为产色素非Ｏ１群霍乱弧菌ＶＢＯ８型和ＶＢＯ３８型．     

The prevalence of Vibrio spp. in drinking water and environmental samples in Vellore South India.

The prevalence of Vibrio cholerae in drinking water, lakes and sewage outfalls during July and August 1996 in Vellore, India was determined. Drinking water samples were collected on single occasions from 12 sites in different geographic areas of the town where cholera had been reported. Samples of water, plankton and sediment were collected from fixed sites at three lakes on three occasions separated by at least 3 days during the course of the study. Samples from open sewers were taken from two representative sites in four areas of the town. Bacteria isolated from samples were identified by standard biochemical tests and isolated strains of V. cholerae tested for their ability to agglutinate O1 and O139 antisera. Water samples from lakes were also tested for the presence of V. cholerae O1 and O139 by fluorescent antibody staining. Non-O1, non-O139 strains of V. cholerae were detected in 41% of drinking water samples and 100% of water, sediment and plankton samples from the test lakes. Eighty-seven per cent of open sewers sampled contained viable non-O1, non-O139 V. cholerae. Fluorescent antibody staining gave positive results for V. cholerae O1 and O139 for all water samples from the three lake sites. Strains of Aeromonas spp. were isolated from 58% of drinking water samples and from 66% of sediment, 77% of plankton and 55% of water samples from lakes. All open sewers sampled contained Aeromonas spp. PCR amplification employing specific primers demonstrated that none of the non-agglutinating V. cholerae isolates contained the ctx operon. The non-O1, non-O139 V. cholerae isolates showed different patterns of antibiotic resistance to ampicillin, ciprofloxacin, chloramphenicol, tetracycline and trimethoprim.     

广西130株非O1群霍乱弧菌血清学分型及噬菌体-生物分型研究

目的 将噬菌体－生物分型方法应用到非Ｏ１群霍乱弧菌的分型,以掌握广西地区非Ｏ１群霍乱弧菌血清型别的分布,并对此分型方法进行探讨。方法 收集广西部分地区１３０株非Ｏ１群霍乱弧菌进行血清学分型和噬菌体－生物分型,并用多聚酯链反应（ＰＣＲ）进行霍乱肠毒素检测。结果 在１３０例菌株中有９０株可以分型,占６９．２３％,分布于３４个血清型,腹泻病人菌株可分为１４个血清型,其中以Ｏ２以优势血清型,外环境菌株分布     

Characterization of toxigenic Vibrio cholerae from Haiti, 2010-2011

In October 2010, the US Centers for Disease Control and Prevention received reports of cases of severe watery diarrhea in Haiti. The cause was confirmed to be toxigenic Vibrio cholerae, serogroup O1, serotype Ogawa, biotype El Tor. We characterized 122 isolates from Haiti and compared them with isolates from other countries. Antimicrobial drug susceptibility was tested by disk diffusion and broth microdilution. Analyses included identification of rstR and VC2346 genes, sequencing of ctxAB and tcpA genes, and pulsed-field gel electrophoresis with SfiI and NotI enzymes. All isolates were susceptible to doxycycline and azithromycin. One pulsed-field gel electrophoresis pattern predominated, and ctxB sequence of all isolates matched the B-7 allele. We identified the tcpETCIRS allele, which is also present in Bangladesh strain CIRS 101. These data show that the isolates from Haiti are clonally and genetically similar to isolates originating in Africa and southern Asia and that ctxB-7 and tcpET(CIRS) alleles are undergoing global dissemination.     

霍乱疫情分离株流行菌型及耐药性分析

目的了解霍乱疫情分离株的流行菌型及其耐药性,为霍乱疫情的快速有效控制和临床治疗提供科学依据。方法对2008年6月海南省某地霍乱疫情采集的6株霍乱弧菌分离株进行血清学分型、噬菌体-生物分型、霍乱毒素基因检测、脉冲场凝胶电泳(PFGE)分子分型和药物敏感性试验。结果 6株菌均属埃尔托生物型霍乱弧菌,除1株为O1群小川型霍乱弧菌流行株(1c),其余5株均为O1群稻叶型霍乱弧菌流行株(1c);6株菌均具有霍乱毒素基因;PFGE分型为2个型,但聚类图谱分析相似性为100%;6株菌均对链霉素、复方新诺明、磺胺异噁唑、多粘菌素B耐药;对诺氟沙星、头孢噻肟、头孢噻吩等9种抗生素均敏感;结论该起霍乱疫情的流行菌型为O1群埃尔托霍乱弧菌稻叶型流行株(1c);诺氟沙星、头孢噻肟、头孢噻吩等可作为病例和带菌者治疗的指导用药,链霉素、复方新诺明、磺胺异噁唑、多粘菌素B则不宜使用。     

Consecutive outbreaks of Vibrio cholerae O139 and V. cholerae O1 cholera in a fishing village near Karachi, Pakistan

In July 2002 and June 2003, cholera outbreaks were detected by a diarrhoea surveillance system in a village outside Karachi, Pakistan. Specimens were culture confirmed. The first outbreak was caused by Vibrio cholerae O139 (n = 30) and the second outbreak by V. cholerae O1 (n = 39). Demographic and clinical features of patients were recorded and case-control studies were conducted following each outbreak. Clinical information was obtained for 29 of the 30 patients in the first outbreak, and 2 of the patients in the second outbreak were either out of the area or lost to follow-up, leaving 29 and 37 cases in the analysis for the first and second outbreak, respectively. Eighteen (49%) of the 37 V. cholerae O1 patients were under 2 years of age compared with 6 (21%) of the 29 V. cholerae O139 patients (P = 0.02). Vibrio cholerae O139-infected patients were more likely to be febrile (16/29) than those infected with V. cholerae O1 (2/37; P<0.001). A household contact with cholera was a risk factor in both outbreaks; water source was a risk factor in the first outbreak only. Geographically, cases were clustered during the first outbreak but not during the second. Person-to-person contact and water reservoirs appear to be the main transmission routes for cholera in this setting.     

1株不典型非O1群霍乱弧菌的发现及鉴定

目的 对1株从肝硬化患者血液中分离的病原菌进行鉴定.方法 按霍乱弧菌的鉴定标准和方法进行细菌学表型特征(包括形态学、培养特性、生化特征)、血清学鉴定及16S rDNA序列分析.结果 从患者血液中分离获得1株病原菌HN 9837,该菌株的形态学、培养特性、生化特征、血清学诊断和分子生物学诊断结果基本符合非O1群霍乱弧菌定义;但与普通霍乱弧菌在甘露醇发酵及赖氨酸利用、柠檬酸盐利用试验结果存在差异.结论 海南省首次在肝硬化患者血液中分离出非O1群霍乱弧菌,该菌为1株不发酵甘露醇、赖氨酸且柠檬酸盐利用试验阴性的不典型霍乱弧菌.     

多重荧光定量PCR甄别霍乱弧菌方法的建立

目的 利用多重荧光定量PCR技术,建立一种快速、准确、特异甄别霍乱弧菌的定量方法.方法 分别根据霍乱弧菌霍乱毒素A亚单位基因(ctxA)和糖基转移酶基因(LPSgt)作为检测的靶基因,设计引物和TaqMan-MGB探针,探针的5'端分别用FAM和VIC进行荧光标记,3'端标记MGB.优化PCR扩增体系,对多重实时荧光定量PCR方法的特异性、灵敏度、重复性评价,同时进行一定数量临床样本考核鉴定,与常规方法进行比较.结果 该方法可准确、特异地鉴定霍乱弧菌,同时能够甄别O139群霍乱弧菌.全部的霍乱弧菌用ctxA基因对应引物和探针检测均为阳性,其中只有O139群霍乱弧菌出现LPSgt基因阳性,其他菌株均无阳性结果.检测的灵敏度为2×102 cfu/ml.同时对收集的45例临床样本进行鉴定,结果显示10例为霍乱弧菌,其中O139群有4例,其余均为阴性结果,与常规鉴定方法一致.结论 研究建立的多重荧光定量PCR方法特异、灵敏、快速,可有效鉴定产毒霍乱弧菌及甄别O139群霍乱弧菌,可用于霍乱监测和防制工作中的实验室诊断.     

Association of a disease approximating cholera caused by Vibrio cholerae of serogroups other than O1 and O139.

One hundred and six patients suffering from severe dehydrating diarrhoea were studied of whom 36 patients were positive for Vibrio cholerae. Out of 36, 15 were positive for V. cholerae O1, 10 for V. cholerae O139 and 11 for V. cholerae non-O1 non-O139. O1 and O139 were positive for the 301-bp ctxA amplicon and 471-bp tcpA amplicon indicating that the strains possessed toxigenic capability whereas no non-O1 non-O139 strain possessed ctxA or tcpA genes. Post-admission severity of purging and amount of ORS required were less in the V. cholerae non-O1 non-O139 group (P < 0.05) compared to the V. cholerae O1 and O139 groups. It appears from this study that a cholera-like clinical condition can be caused in the absence of CT as exemplified by strains of non-O1 non-O139.