Septicemia due to Vibrio cholerae serogroup non-O1/non-O139 strain in a cirrhotic patient

We describe a case of non-O1/ non-O139?Vibrio cholerae septicemia in a 65-year-old male patient with alcoholic liver cirrhosis. He was admitted due to septic shock from non-O1/ non-O139 V. cholerae. An intravenous empiric antibiotic, ceftriaxone sodium hydrate, was administered together with amikacin sulfate, gamma globulin and dopamine. He was discharged feeling well 17?days after admission. Poor host defense mechanisms as seen in cirrhotic patients seem to be a determinant for systemic infection of non-O1/ non-O139 V. cholerae. Such patients should be warned and educated against eating raw or undercooked seafood to avoid the occurrence of non-O1/non-O139 V. cholerae septicemia.     

云南省非O1/O139群霍乱弧菌分子特征分析

目的非O1/O139群霍乱弧菌能够引起人类急性腹泻性疾病,但与O1群和O139群相比,人们往往容易忽视其危害性。因此,我们对分离于云南省的31株非O1/O139群霍乱弧菌开展了分子特征分析。方法采用纸片法(K-B)开展抗生素敏感试验;多聚酶链反应(PCR)扩增检测毒力基因;同时,我们对菌株进行了脉冲场凝胶电泳(PFGE)和多位点序列分析(MLST)分子分型研究。结果药敏实验结果显示,67.74%的菌株对利福平耐药,对萘啶酸和复方新诺明的耐药率为29.03%,所有的菌株对庆大霉素和环丙沙星敏感;PCR结果显示,所有菌株的ompW基因为阳性,87.10%的菌株hly基因为阳性,25.81%的菌株rstREl tor为阳性,16.13%的菌株rstRClassical和tcpAEl tor为阳性,而CT、rfbO1和rfbO139基因全为阴性;PFGE结果显示,31株非O1/O139群霍乱弧菌呈现出高度的离散趋势,几乎没有相同带型的菌株出现;在MLST结果分析中,发现了1个gyrB新的等位基因,mdh基因发现了4个新的等位基因,metE基因发现了6个新的等位基因,pntA中发现了2个新的等位基因,purM中发现了3个新等位基因,pyrC中发现了4个新等位基因。经过排列组合后,共发现了17个新的霍乱弧菌ST型别(ST273-ST289)。结论非O1/O139群霍乱弧菌呈现出高度的多样性特征,同时,云南省的非O1/O139群霍乱弧菌具有一定的地域特点,丰富了现有的霍乱弧菌分子分型数据库。     

兔抗霍乱弧菌O139血清群rMSHA多克隆抗体的制备和鉴定

目的制备兔抗霍乱弧菌O139血清群rMSHA多克隆抗体。方法用经Ni-NTA亲和层析法纯化的重组蛋白(rMSHA)免疫日本大耳兔,收集免疫兔血清。Western Blot检测重组蛋白的免疫原性;ELISA测定该抗体的效价。结果兔抗rMSHA抗体能与重组蛋白发生特异性反应;间接ELISA法测定该抗体效价为1∶25 600。结论成功地制备了兔抗rM-SHA抗体,该抗体能够识别原核表达的重组蛋白rMSHA,为研制霍乱弧菌疫苗及相关诊断试剂盒奠定了基础。     

The Vibrio cholerae O139 serogroup antigen includes an O-antigen capsule and lipopolysaccharide virulence determinants.

Vibrio cholerae serogroup O139 emerged on the Indian subcontinent in October 1992 to become the first non-O1 V. cholerae serogroup documented to cause epidemic cholera. Although related to V. cholerae El Tor O1 strains, O139 strains have unique surface structures that include a capsular surface layer and lipopolysaccharide (LPS). Immunoblot analysis of either whole-cell lysates or LPS preparations revealed three electrophoretic forms of the O139 antigen: two slowly migrating forms and one rapidly migrating form that appeared identical to O139 LPS. All three forms of the antigen shared an epitope defined by an O139-specific monoclonal antibody. A serum-sensitive nonencapsulated mutant was isolated that lacks only the slow migrating forms. The slow migrating forms did not stain with silver whereas the rapidly migrating form did, suggesting that the former might constitute highly polymerized O-antigen side-chain molecules that were not covalently bound to core polysaccharide and lipid A (an "O-antigen capsule"). A single transposon insertion resulted in the loss of immunoreactivity of both the LPS and the O-antigen capsule, implying that there are genes common to the biosynthesis of both these macromolecules. The O139 LPS and O-antigen capsule were both important for colonization of the small intestine of the newborn mouse and for serum resistance, demonstrating that both of these forms of the O139 serogroup antigen are virulence factors.     

Distribution of virulence markers in clinical and environmental Vibrio cholerae non-O1/non-O139 strains isolated in Brazil from 1991 to 2000

One hundred seventy nine Vibrio cholerae non-O1/non-O139 strains from clinical and different environmental sources isolated in Brazil from 1991 to 2000 were serogrouped and screened for the presence of four different virulence factors. The Random Amplification of Polymorphic DNA (RAPD) technique was used to evaluate the genetic relatedness among strains. Fifty-four different serogroups were identified and V. cholerae O26 was the most common (7.8%). PCR analysis for three genes (ctxA, zot, ace) located of the CTX genetic element and one gene (tcpA) located on the VPI pathogenicity island showed that 27 strains harbored one or more of these genes. Eight (4.5%) strains possessed the complete set of CTX element genes and all but one of these belonged to the O26 serogroup suggesting that V. cholerae O26 has the potential to be an epidemic strain. The RAPD profiles revealed a wide variability among strains and no genetic correlation was observed.     

Increasing drug resistance in Vibrio cholerae O1 and non-O1 strains isolated from diarrheal cases in Japan

Drug resistance trends were investigated for 271 Vibrio cholerae O1 (V.c O1) and 401 V. cholerae non-O1 (V.c non-O1) strains isolated from mainly imported diarrheal cases during 1981-2001 in Japan. The results of drug resistance test using 8 drugs (CP, TC, SM, KM, ABPC, ST, NA, and NFLX) showed that 34.7% of the V. c O1 strains and 15.7% of V.c non-O1 strains were multi-drug or mono-drug resistant. The incidence of drug resistant strains has increased since 1991, and it has been remarkable in V.c O1 strains that increased from 1.2% in 1981-1985 to 70.8% in 1996-2001. The drug resistance patterns of the resistant strains classified into 6 types in V.c O1 and 21 types in V.c non-O1. The prevalent patterns recognized were SM (75.5%), CP.TC.SM.ST (10.6%) and CP.SM.ST (8.5%) in V.c O1, and SM (25.4%) and ABPC (25.4%) in V.c non-O1. Ten V.c O1 strains (3.7%) and 10 V.c non-O1 strains (2.5%) were multi-drug resistant including TC. Among those, 13 strains were isolated from travelers who returned to Japan from Thailand. One V.c O1 strain (0.4%) and 6 V.c non-O1 strains (1.5%) were NA high-resistant and fluoroquinolones low-sensitive. Among those, 4 strains were isolated from travelers who returned to Japan from India.     

肝硬化患者非O1非O139型霍乱弧菌败血症

目的分析非O1非O139型霍乱弧菌感染的肝硬化败血症患者的临床和实验室特点。方法回顾调查病人的临床基本情况、原发病、实验室结果、治疗和预后;BacT/Alert全自动血培养微生物检测仪培养病人全血标本,用VITEK仪器鉴定细菌。药敏试验根据CLSI(2005)规定,K-B药敏纸片测定法进行。结果3例患者均为肝硬化病人;感染发生在夏季;经左氧氟沙星、加替沙星或联用头孢曲松治疗后痊愈。结论非O1非O139型霍乱弧菌败血症推荐使用头孢三代或氟喹诺酮类抗生素治疗。     

Genomic characterization of non-O1, non-O139 Vibrio cholerae reveals genes for a type III secretion system

Non-O1, non-O139 Vibrio cholerae can cause gastroenteritis and extraintestinal infections, but, unlike O1 and O139 strains of V. cholerae, little is known about the virulence gene content of non-O1, non-O139 strains and their phylogenetic relationship to other pathogenic V. cholerae. Comparative genomic microarray analysis of four pathogenic non-O1, non-O139 strains indicates that these strains are quite divergent from O1 and O139 strains. Genomic sequence analysis of a non-O1, non-O139 strain (AM-19226) that appeared particularly pathogenic in experimental animals suggests that this strain carries a type III secretion system (TTSS) that is related to the TTSS2 gene cluster found in a pandemic clone of Vibrio parahaemolyticus. The genes for this V. cholerae TTSS system appear to be present in many clinical and environmental non-O1, non-O139 strains, including at least one clone that is globally distributed. We hypothesize that the TTSS present in some pathogenic strains of non-O1, non-O139 V. cholerae may be involved in the virulence and environmental fitness of these strains.     

Are the environmental niches of Vibrio cholerae O139 different from those of Vibrio cholerae O1 El Tor?

BACKGROUND: Vibrio cholerae are known to be normal inhabitants of surface water. However, the environmental niches of the different strains of cholera are not well known, and therefore, populations at risk for cholera outbreaks cannot be clearly identified. METHODS: This study identifies environmental risk factors for cholera caused by V. cholerae O1 El Tor and O139 and environmental niches of the two strains present in Matlab, a cholera endemic area of Bangladesh. The study year was 1993, the year that the O139 strain first appeared in the study area. Patients who had either strain of cholera identified in a laboratory were included in the study. A geographic information system was used to map the household locations of the patients, to describe the human sanitary environment and population density, and to address potential anthropogenic and environmental risk factors of the disease. Spatial point pattern and exploratory spatial data analysis techniques were used to define the environmental niches of the two cholera strains. RESULTS: The study suggests the niches of O1 El Tor and O139 strains of V. cholerae appear to be similar, based on common environmental risk factors. CONCLUSIONS: The results of this study support a theory that O1 El Tor could possibly be replaced by the newer O139 strain in the future.     

Emergence and evolution of Vibrio cholerae O139

The emergence of Vibrio cholerae O139 Bengal during 1992-1993 was associated with large epidemics of cholera in India and Bangladesh and, initially, with a total displacement of the existing V. cholerae O1 strains. However, the O1 strains reemerged in 1994 and initiated a series of disappearance and reemergence of either of the two serogroups that was associated with temporal genetic and phenotypic changes sustained by the strains. Since the initial emergence of the O139 vibrios, new variants of the pathogen derived from multiple progenitors have been isolated and characterized. The clinical and epidemiological characteristics of these strains have been studied. Rapid genetic reassortment in O139 strains appears to be a response to the changing epidemiology of V. cholerae O1 and also a strategy for persistence in competition with strains of the O1 serogroup. The emergence of V. cholerae O139 has provided a unique opportunity to witness genetic changes in V. cholerae that may be associated with displacement of an existing serogroup by a newly emerging one and, thus, provide new insights into the epidemiology of cholera. The genetic changes and natural selection involving both environmental and host factors are likely to influence profoundly the genetics, epidemiology, and evolution of toxigenic V. cholerae, not only in the Ganges Delta region of India and Bangladesh, but also in other areas of endemic and epidemic cholera.     

Isolation of Vibrio cholerae O139 from the drinking water supply during an epidemic of cholera

In mid 1994, the public water supply was investigated in a medium-sized town in south India during an epidemic of cholera due to Vibrio cholerae O139. Vibrio cholerae O139 was isolated from the public water supply including one of the wells supplying the town, the central overhead tank, and domestic taps connected to the public supply. Following chlorination, the organism was no longer isolated from the water supply and the epidemic subsided. This demonstration of V. cholerae O139 in the drinking water supply of a town underlines the need for adequate treatment of the water supply.     

Genetic rearrangements in the rfb regions of Vibrio cholerae O1 and O139.

The recent emergence of a pathogenic new non-O1 serotype (O139) of Vibrio cholerae has led to numerous studies in an attempt to identify the origins of this new strain. Our studies indicate that O139 strains have clear differences in the surface polysaccharides when compared with O1 strains: the lipopolysaccharide can be described as semi-rough. Southern hybridization with the O1 rfb region demonstrates that O139 strains no longer contain any of the rfb genes required for the synthesis of the O1 O-antigen or its modification and also lack at least 6 kb of additional contiguous DNA. However, O139 strains have retained rfaD and have a single open reading frame closely related to three small open reading frames of the O1 rfb region. This region is closely related to the H-repeat of Escherichia coli and to the transposases of a number of insertion sequence elements and has all the features of an insertion sequence element that has been designated VcIS1. Transposon insertion mutants defective in O139 O-antigen (and capsule) biosynthesis map to the same fragment as VcIS1. Preliminary sequence data of complementing clones indicate that this DNA encodes a galactosyl-transferase and other enzymes for the utilization of galactose in polysaccharide biosynthesis. We propose a mechanism by which both the Ogawa serotype of O1 strains and the O139 serotype strains may have evolved.     

福建省非典型O1群埃尔托型霍乱弧菌的研究

目的 探索非典型O1群埃尔托型霍乱弧菌在福建省1962-2005年霍乱菌株中的存在及其意义。方法 选择1962-2005年代表性霍乱菌株69株(稻叶21、小川48),运用PCR技术进行ctxB、tcpA、rstR、hlyA 各毒力因子的古典型(Cl)和埃尔托型(El)基因的扩增;同时对部分菌株的ctxB 和rstR 基因的序列进行测定和比对;分析福建省O1群埃尔托型霍乱弧菌(EVC)中非典型菌株出现的时间和存在意义。结果 1962年福建省分离的O1群EVC菌株中携带有ctxB-Cl、tcpA-Cl、hlyA-Cl基因;rstR-Cl基因仅在1994-2000年的霍乱菌株中检出。序列比对结果发现ctxB 基因出现新的204位碱基突变位点(T→G);同时报道两种新的ctxB 基因型别(10和11),且发现存在于同一个菌株中;ctxB基因型11的115位碱基为C,表现为古典型ctxB 位点特征,203位碱基为T,表现为埃尔托型ctxB 位点特征。结论 福建省在1962年的O1群EVC菌株中就出现杂合有ctxB-Cl、tcpA-Cl、hlyA-Cl基因的非典型EVC菌株,是迄今为止报道出现非典型EVC最早的年代;1994年以后出现整合有rstR-Cl基因的非典型EVC菌株。另外,在福建省O1群EVC中存在国内外未曾报道的新的ctxB 碱基突变位点和ctxB 基因型别;同时在同一菌株中存在不同的ctxB 基因型别杂合形式;ctxB 序列也存在不同生物型别的特异位点组合形式。福建省O1群EVC表现为独特的地方性分子特征。     

杭州市健康人群粪便中检出O1和O139群霍乱弧菌无毒力株

目的　对杭州市饮食、服务行业人员粪便标本进行霍乱弧菌监测。方法　 2 0 0 0年 5月～ 2 0 0 1年 7月采集 2 6 2 5 2名饮食、服务行业从业人员健康体检者粪便标本 ,分别接种于碱性蛋白胨水和 4号琼脂中。通过培养特性、菌落形态、革兰染色、动力和生化试验等检查 ,对所分离的疑似霍乱弧菌菌株进行初步鉴定。采用霍乱弧菌O1群及O139群单克隆抗体的玻片凝集试验、噬菌体分型、霍乱弧菌ctxA和tcpA基因的PCR进一步鉴定各疑似霍乱菌株。 结果　上述标本中检出O1血清群和O139血清群霍乱弧菌各 2株。 2株O1血清群霍乱弧菌的噬菌体分型分别为 19k和 5k ,为埃尔托生物型非流行菌株 ;2株O139血清群霍乱弧菌的噬菌体分型分别为 2 0k和 31k ;但ctxA和tcpA基因PCR检测结果均为阴性。结论　尽管所检出的 4株霍乱弧菌均为无毒力的非流行菌株 ,但因携带者从事职业的特殊性 ,仍应引起高度重视。     

霍乱弧菌O1群和O139群nhaA基因的克隆和变异性研究

目的 克隆霍乱弧菌O1群和O139群nhaA基因，并分析其变异性。方法收集我国1988—2000年散发霍乱弧菌O1群和O139群40株，用聚合酶链反应(PCR)扩增nhaA基因，克隆至pcDNA3载体，通过序列测定分析其同源性和变异性。结果 分别从霍乱弧菌Ol群和O139群扩增出约1．2kb的nhaA基因片段，基因序列分析表明，我国霍乱散发O1群和O139群的nhaA基因与Genbank中霍乱弧菌O1群野毒株参考序列同源性分别为99％和96％。编码氨基酸的突变率分别为2％和11％，nhaA基因重要的结构和功能决定簇(Aspl33、Aspl63、Aspl64、His225、Leu73和Gly338)未发生变异；第203、221位的氨基酸，O1群和O139群发生相同的变异。结论 nhaA基因编码氨基酸的变异可能是霍乱弧菌适应外环境变化的结果。     

Induction of protective immunity by synthetic Vibrio cholerae hexasaccharide derived from V. cholerae O1 Ogawa lipopolysaccharide bound to a protein carrier

Synthetic antigens that mimic the terminal hexasaccharide epitope of the O-specific polysaccharide of Vibrio cholerae O1, serotype Ogawa, were conjugated to bovine serum albumin (BSA). Conjugates with carbohydrate-to-carrier molar ratios of 15.5:1, 9.2:1, and 4.6:1 were tested for immunogenicity and efficacy in mice. The role of preimmunity to BSA and the use of adjuvant in the generation of the serologic response to the O-specific polysaccharide and protection against virulent V. cholerae was examined. Preimmunity to BSA did not affect the anti-Ogawa titers but seemed to enhance the protective capacity of antiserum. All 3 conjugates were immunogenic, but adjuvant was effective at inducing higher and earlier antibody responses. In tertiary serum samples, a correlation was found between vibriocidal activity and protection. The protective capacity of antiserum was evident in serum from mice immunized with all conjugates, but it was highest in the groups that received the conjugate with the lowest level of substitution. Further studies are required to increase understanding of the reason for differential protection.     

Helicobacter pylori and epidemic Vibrio cholerae O1 infection in Peru

In a cross-sectional study of the 1991 Peruvian cholera epidemic, Vibrio cholerae O1 infection was associated with Helicobacter pylori infection, particularly in young children. These data support the hypothesis that hypochlorhydria induced by H. pylori is important in the pathogenesis of diarrhoeal disease.