重庆市首次分离出巴达维亚群钩端螺旋体

钩端螺旋体病（简称钩体病）是一种人畜共患的动物源性疾病,而鼠类是钩体病的主要带菌动物。我站根据《卫生部三峡库区疾病监测方案》的计划布署。于１９９７年开始每年进行两次监测,在 １９９７年 １０月这次监测中分离出一株巴达维亚群钩体．现将结果报告如下。１　材料与方法１．１　标本来源 在渝北区硌碛镇长江边稻田及野外菜地布鼠夹１００个,捕鼠７只,其中黑线姬鼠５只,褐家鼠２只。１．２　培养基 柯大氏（Ｋｏｕｔｈｏｆ）培养基：重庆市卫生防疫站细菌科提供。１．３　实验方法 按《三峡库区重庆段．鼠类钩端螺旋体病原的检…     

湖南省洪涝灾害高发地区钩端螺旋体病流行因素的研究

目的研究湖南省洪涝灾害高发地区钩端螺旋体病的流行因素。方法选择洞庭湖区6个县研究传染源密度、带茵情况和自然人群、病人与动物抗体水平，按国家有关标准进行实验室和现场工作。结果(1)传染源：灭鼠前、后和钩体病流行后期鼠密度分别为7．02％、2．31％和4．64％，优势鼠种为黑线姬鼠，灭鼠后鼠密度下降了67．09％：村民家庭家畜饲养以圈养形式占99．15％，喂商品混合饲料的占72．33％。(2)病原学：对鼠、猪、犬和病人的标本进行钩体培养，分离出82株钩体，黄疸出血群占70．73％，1株国内新型茵，黑线姬鼠分离率高达11．03％。(3)血清学：检测1263名自然人15个群钩体抗体，一群以上抗体阳性率未接种钩体苗为50．88％，接种一针四价钩体为72．55％，人群抗体有14群之多；检测85例病人双份血清抗体，确诊63例，符合率74．11％，黄疸出血群感染比例最高；流行季节牛、犬和出栏猪抗体分别有14、8和7个群，牛阳性率最高。结论流行前灭鼠效果好，不仅降低了鼠密度，还降低了带茵率；主要传染源是鼠和牛，牛作为传染源的意义大于猪和犬；人群接种一针四价钩体菌苗亦有效；病人感染黄疸出血、澳洲和秋季群为多。     

2007年湖南省钩端螺旋体宿主动物感染状况结果分析

目的 掌握湖南省钩端螺旋体宿主动物主要种类、密度、以及感染率和感染类型等情况,为防治钩体病提供科学依据.方法 按照2007年<湖南钩端螺旋体病监测实施方案>,在全省6个常规监测点和环洞庭湖区5个应急监测点系统开展钩体的宿主动物带菌率与带菌种群调查.以夹夜法在室外捕获鼠类,采用无菌操作采集鼠肾,Korthof方法 进行培养;应用分群血清与新分离的钩端螺旋体作凝集试验确定菌群,应用交叉凝集素吸收试验或分型因子血来确定菌型.结果 常规监测点有效培养459份,感染率为10.24%.湘潭县、沅江市、双蜂县的感染率分别为3.81%、6.67%、44.19%(p＜0.01).黑线姬鼠、东方田鼠、小家鼠、褐家鼠的感染率分别为13.96%、12.50%、5.19%、4.55(p＜0.05).浏阳市的蛙肾中培养出1株秋季型、3株澳洲型钩体.应急监测点有效培养296份,感染率为16.89%,其中沅江市、大通湖区、君山区、岳阳县、汉寿县的感染率分别是10.84%、24.39%、27.27%、41.46%、2.30%(p＜0.01).东方田鼠、褐家鼠、黑线姬鼠、黄毛鼠的感染率分别为28.36%、10.17%、9.26%.5.03%(p＜0.01).菌型主要是黄疸出血群赖型、赛罗群乌尔夫型.结论 湖南省宿主动物密度和带菌事差异较大,以黄疸出血群和赛罗群为主,感染率较往年升高,必须加强综合防制措施,预防爆发和流行.     

2007年湖南省钩端螺旋体宿主动物感染状况结果分析

目的 掌握湖南省钩端螺旋体宿主动物主要种类、密度、以及感染率和感染类型等情况,为防治钩体病提供科学依据.方法 按照2007年<湖南钩端螺旋体病监测实施方案>,在全省6个常规监测点和环洞庭湖区5个应急监测点系统开展钩体的宿主动物带菌率与带菌种群调查.以夹夜法在室外捕获鼠类,采用无菌操作采集鼠肾,Korthof方法 进行培养;应用分群血清与新分离的钩端螺旋体作凝集试验确定菌群,应用交叉凝集素吸收试验或分型因子血来确定菌型.结果 常规监测点有效培养459份,感染率为10.24%.湘潭县、沅江市、双蜂县的感染率分别为3.81%、6.67%、44.19%(p＜0.01).黑线姬鼠、东方田鼠、小家鼠、褐家鼠的感染率分别为13.96%、12.50%、5.19%、4.55(p＜0.05).浏阳市的蛙肾中培养出1株秋季型、3株澳洲型钩体.应急监测点有效培养296份,感染率为16.89%,其中沅江市、大通湖区、君山区、岳阳县、汉寿县的感染率分别是10.84%、24.39%、27.27%、41.46%、2.30%(p＜0.01).东方田鼠、褐家鼠、黑线姬鼠、黄毛鼠的感染率分别为28.36%、10.17%、9.26%.5.03%(p＜0.01).菌型主要是黄疸出血群赖型、赛罗群乌尔夫型.结论 湖南省宿主动物密度和带菌事差异较大,以黄疸出血群和赛罗群为主,感染率较往年升高,必须加强综合防制措施,预防爆发和流行.     

清远市2004年钩端螺旋体病监测分析

目的 对2004年清远市钩端螺旋体病疫情、人群及宿主动物监测情况进行分析,为制定防治措施提供科学依据.方法 采取疑似病人血、鼠类、水禽类和蛙类动物肾组织分离培养病原体,用MAT法检测健康人、疑似病人、鼠类和水禽类动物血清抗体.结果 报告发生钩体病10例,发病率0.25/10万,死亡1例,病死率10.00%.从鼠肾组织中分离鉴定了5株钩体菌,分离阳性率为5.00%,均为爪哇群.流行前、后期健康人血清和流行期疑似病人血清抗体阳性率分别为13.27%、4.00%和15.79%,菌群以黄疸出血群和七日热群为主,其次为拜伦群和爪哇群.鼠血清抗体阳性率为22.00%,黄毛鼠、板齿鼠、褐家鼠和黄胸鼠的血清抗体阳性率在8.57%～50.00%之间,菌群以秋季热群为主,其次为犬热群.水禽（鸭）血清抗体阳性率为4.00%,菌群为拜伦群和致热群.结论 我市人群钩体隐性感染水平和鼠带菌率均较高,开始出现钩体病发病年龄组后移及钩体带菌优势鼠种由野栖鼠种向家栖鼠种交叉转移的动向,从水禽类动物（鸭）血清中检测出钩体带菌抗体在我市尚属首次.继续加强监测,采取综合性防治措施,可有效地控制钩体病的流行.     

三峡库区兴山县钩端螺旋体宿主动物带菌状况及人群免疫水平分析

目的掌握兴山县钩端螺旋体病(钩体病)宿主动物带菌及人群免疫水平,为进一步做好钩体病的预防与控制提供依据。方法按照《全国钩端螺旋体病监测方案》开展鼠、家畜、青蛙等宿主动物带菌调查及健康人群血清学监测。结果平均鼠密度为1.80%。鼠种构成以黄胸鼠、小家鼠和黑线姬鼠为优势种,平均带菌率为3.75%。共检测病人尿液及家畜、青蛙、稻田疫水等7类样本565份,培养阳性菌株4株,阳性率为0.71%,其中稻田疫水2株,病人尿液、蛙肾各1株,菌株经鉴定黄疸出血群1株,秋季热群3株。人群钩体抗体阳性率平均为28.62%,青壮年阳性率较高,以黄疸出血型、七日热型、流感伤寒型、波摩那型为主。在监测中发现1例在农贸市场感染的经济型钩体病。结论兴山县钩体宿主动物带菌率低,血清学显示人群钩体隐性感染率较低,在山区型钩体自然疫源地存在的情况下,其发病率与社会因素和自然因素密切相关。农贸市场、畜牧养殖区及其周边为经济型钩体病疫源地,病例分析已经得到证实,建议今后应将其纳入预防和监测范围。     

清远市2000～2003年钩端螺旋体病监测分析

目的：了解清远市2000～2003年钩端螺旋体病人群及宿主动物带菌情况，为制定防治措施提供科学依据。方法：采取病人血、动物脏器分离培养病原体。用MAT法检测健康人、疑似病人和鼠血清抗体。结果：从病人血和动物脏器中分离鉴定了16株钩端螺旋体，分属4种菌群，分别为犬热群、秋季热群、赛罗群和爪哇群。健康人血清和疑似病人血清阳性率分别为29．22％和3．81％，菌群均以黄疸出血群为主。鼠血清阳性率为29．71％，菌群以爪哇群为主，板齿鼠、黄毛鼠、黄胸鼠和褐家鼠的带菌率在26％～50％之间。结论：我市人群钩端螺旋体隐性感染水平和鼠带菌率均较高，从人群中分离到的赛罗群钩端螺旋体在我省尚属首次。继续加强监测，采取综合性防治措施，可有效地控制钩端螺旋体病的流行。     

重庆市钩端螺旋体病流行因素监测分析

目的分析重庆市钩体病的发病规律和影响因素,为制定针对性的防制措施提供依据,防止钩体病疫源地向三峡库区和长江下游地区扩散。方法选择1个钩体疫区县和2个三峡库区县作为监测点,用显微镜凝集试验(AMT)检测病人、健康人群及耕牛血清抗体。用夹夜法捕鼠,鉴定鼠种,计算鼠密度,分离培养病原体,同时用PCR法进行比较。结果1961~2004年重庆市钩体发病率在0.09/10万~12.16/10万之间,死亡率在0~0.44/10万之间,病死率在0%~10.34%之间波动。彭水县1998年和2000年的洪灾年出现2次局部爆发。监测点鼠密度在1.6%~4.4%之间波动,食虫目小兽(51%)、褐家鼠(25%)和黑线姬鼠(12%)为优势鼠种。鼠带菌率为1.7%(10/583),褐家鼠和食虫目小兽中分离出钩体病原,均为澳洲群澳洲型。病人血清检测阳性率为73.33%(33/45),血清群以澳洲群和黄疸出血群为主;健康人群感染率为45%(598/1330),疫区点血清抗体阳性率(74.7%)显著高于库区点(28.0%)。牛血清检测优势菌群为澳洲群。结论鼠类是重庆市钩体病的主要传染源,主要传播因素为稻田作业,主要流行菌群为澳洲群,库区人群免疫水平较低。三峡库区蓄水后水流变缓,接触疫水的频率增加,可能引起钩体疫情暴发流行。     

曹娥江流域钩端螺旋体病自然疫源地垂直结构模式的调查

本文报告在曹娥江支流发源地至出海口的钩端螺旋体病自然疫源地的垂直调查结果是:1.中山梯田:未发现自然疫源地。2.低山梯田:黑线姬鼠黄疸出血群纳姆型自然疫源地。3.盆地、丘陵、平原水网:黑线姬鼠黄疸出血群沃尔登型自然疫源地。4.滨海平原:黑线姬鼠黄疸出血群沃尔登型,爪哇群爪哇型自然疫源地。5.海涂旱地;罗赛鼠爪哇群爪哇型自然疫源地。     

四川省大竹县钩端螺旋体病两个集中暴发点流行病学特征调查

目的了解大竹县钩端螺旋体病(简称钩体病)两个集中暴发点流行病学特征,为制定防治措施提供科学依据。方法根据疫情报告,选取钩体病两个集中暴发点进行调查,用MAT法检测病人和蛙血清抗体;并分离培养钩体病原体。用夹夜法捕鼠,无菌采集鼠肾、猪肾、牛中段尿和蛙肾分离培养钩体病原体,分析钩体病流行病学特征和感染菌群情况。结果发病户数占总户数的68.13%;发病例数占总人数的28.41%;病人和宿主动物标本中检出阳性菌株51份,经鉴定以七日热群为主,占60.78%,其次分别为黄疸出血群、澳洲群、波摩那群、犬群,分别占29.41%、3.92%、3.92%、1.96%。结论病人和宿主动物感染钩体菌群复杂,应切实加强钩体病的监测,为制定钩体病防治方案提供科学依据。     

钩端螺旋体重组DNA探针对致病性黄疸出血群钩体DNA的杂交识别

Two recombinant DNA probes-PLIpso1 (15kb) and PLIEc34 (4kb), derived from Leptospira serovaricterohaemorrhagiae genomic libraries, were applied for the hybridization and identification of 13 strains of Leptospira in serogroup icterohaemorrhagiae. Difference in hybridization signal in combination with the banding pattern provide a good way for identification of serovars and strains. The recombinant DNA, specific to L. serogroup icterohaemorrhagiae, hybridized with a limited number of DNA fragments which had been digested by several restriction endonucleases. The less complex banding pattern and higher sensitivity facilitate characterization of various serovars and strains in serogroup icterohaemorrhagiae. In general the DNA patterns recognized by both probes have extensive genomic homology in same serovar (but still can distinguish strains in same serovar by some unique bands) and apparent difference in various serovars (especially serovars naam, nanxi and honghe). The results indicated that Southern blotting with recombinant DNA probe might provide tools for identification, characterization and analysis of leptospira.     

Molecular Typing of Leptospira interrogans Strains Isolated from Rattus tanezumi in Guizhou Province, Southwest of China

ObjectiveTo identify and type three leptospires isolated from Rattus tanezumi in Guizhou Province by using three molecular techniques (PFGE, MLVA, and MLST), reveal the molecular characteristic of causative agents of local leptospirosis and evaluate these three molecular methods based on their detection resolution and efficiency.MethodsThree Leptospira strains were isolated from the kidney of Rattus tanezumi and cultured with EMJH medium. PFGE, MLVA, and MLST assays were applied to type the three strains isolated from Rattus tanezumi in Guizhou Province.ResultsPFGE, MLVA, and MLST typing showed that the three leptospiral isolates matched with leptospiral serogroup Icterohaemorrhagiae serovar Lai. The findings of the genotyping methods were consistent. MLVA and MLST defined genotypes, whereas PFGE allowed the recognition of additional subgroups within the genotypes, and the findings of molecular typing were also consistent with those of traditional techniques.ConclusionThree leptospiral isolates from Guizhou Province matched with leptospiral serogroup Icterohaemorrhagiae serovar Lai, and PFGE, MLVA, and MLST, as reliable molecular techniques for identifying and typing of Leptospira interrogans, would contribute to the active surveillance, outbreak investigation and source tracking for leptospirosis in Guizhou Province.     

Homology study of leptospires by molecular hybridization]

Nick translation and random primer labelling method were applied to prepare three genomic DNA probes from Leptospira interrogans strain 017, Leptospira biflexa strain Patoc I and Leptonema illini strain 3055, and then hybridized with DNA of 17 strains leptospires from different genus, species, serogroup and serovar. The results showed no homology between Leptospira and Leptonema, and only a low degree of homology between L. interrogans and L. biflexa but it showed a high degree of homology among L. interrogans. The study also proved the possibility to establish a DNA probe prepared from a single leptospira strain to detect different serovars.     

2005年贵州省流行性感冒监测结果分析

目的: 了解2005年贵州省人群中流行性感冒流行的趋势及病毒型别,及时发现新的变异毒株,为流感防治提供科学依据.方法: 选择哨点医院进行流感的流行病学和病原学监测,采用鸡胚和MDCK细胞培养,进行病毒分离,用血凝抑制试验进行型和亚型鉴定,同时开展全省疫情监测.结果: 2005年检测1 054份标本,分离阳性病毒株56株(其中监测点标本24株,疫情标本32株),分离的病毒型别主要是甲1型42株,甲3型7株,乙型6株,有1株未定型.结论: 2005年贵州省流感流行的高峰在3～4月和10～12月,有甲1、甲3及乙型流感病毒,但以甲1型流感病毒较为活跃.     

聚合酶链反应检测致病性钩端螺旋体微量DNA的研究

Leptospirosis is a severe zoonosis in the world. The methods for detecting leptospira are not sensitive and specific so far. The problems in early diagnosis and epidemiological identification of Leptospirosis remain unsolved. Two recombinant DNA fragments of serogroup Icterohaemorrhagiae, Leptospira were selected by repeated molecular cloning and screening in this study firstly. One of them can hybridize with the DNA of various serogroups of Leptospira interrogans; the other can only hybridize with the DNA of serogroup Icterohaemorrhagiae. After the nucleotides sequence analysis, from these 2 recombinant DNA fragments, 2 pairs of polymerase chain reaction (PCR) oligonucleotide primers were synthesized, named primer B 1, 2 and primer B 3, 4 PCRs were carried out with these 2 primers for detecting the microquantity (0.1 ng) of various serovars, serogroup of leptospires. All the DNA of Leptospira interrogans can be amplified by primer B 1, 2, and only the DNA of serogroup icterohaemorrhagiae, Leptospira reacted specially with primer B 3, 4. The DNA of non-pathogenetic Leptospira and some other microbes, however, had no amplification at all. This study is first reported at home and abroad. The results demonstrate that PCR is a very sensitive and specific technique of DNA amplification, which can be used as a powerful tool in the early diagnosis and epidemiological identification of leptospirosis.     

霍乱弧菌抑制子基因及间隔区(rstR-ig2)多态性研究

目的研究霍乱弧菌CTXφ基因组抑制子基因及间隔区（rstR-ig2）多态性。方法聚合酶链反应（PCR）及限制性片段长度多态性分析（PCR-RFLP）、型特异性引物PCR分型。结果霍乱弧菌（VC）01群古典型（CVC）5株均为class类型。VC01群埃尔托型（EVC）155株共分为4个类型：class、ET、class＋ET、class ＋newET,后2个类型为首次发现。不同地区分离的EVC均以ET类型为主,以class和class＋ET类型为辅。O139群VC82株共分为4个类型：ET、ET＋new O139、ET＋calc、ET＋calc＋newO139,后3个类型为首次发现;福建省分离的菌株以ET4-newO139类型为主,其它地区分离的均以ET类型为主。结论EVC和O139群VC的rstR-ig2基因均以ET类型为主．首次发现5种不同类型rstR-ig2基因共存的新类型。     

问号状钩端螺旋体外膜单克隆抗体凝集反应特性的研究

Three McAb were produced against an outer envelope preparation from Leptospira, interrogans, serovar Lai by fusion of SP2/0 myeloma cells with immune BALB/c mice spleen cells. The fusion rate was 96% and the antibody positive rate was 50%. One of the hybridomas, E4B11C9, reacted with 13 of the 13 serovars of the Icterohaemorrhagiae serogroup in microscopic agglutination test (MAT) but did not react with the 18 representative serovars of L. interrogans and L. biflexa serovar patoc and Leptonema illini. For all non-reactive serovars the MAT titres were greater than 1:25. The McAb, E4B7G5, reacted similarly with all serovars except smithi and tonkini. E4B7D4 reacted also similarly with all serovars except serovars birkini, ndambari, bogvere, smithi and tonkini. Therefore, 3 McAb showed serogroup specificity and partial serogroup specificity by agglutination. The agglutination titres were high and hybridomas were stable, so it might be useful in providing a simple, rapid method for the classification and identification of clinical isolates such as pathogenic L. interrogans in place of the complicated and time-consuming conventional methods.     

Leptospiral carriage by mice and mongooses on the island of Barbados

Leptospirosis is a zoonotic disease, maintained by chronic infection of the kidneys of reservoir animals, usually small mammals. Infection in humans is acquired from direct or indirect exposure to the urine of infected animals. Leptospirosis has a high incidence in tropical regions, and has been studied extensively in several Caribbean countries. We studied the carriage of Leptospira serovars by two small mammals which are potential maintenance hosts of the disease in Barbados. A total of 136 mongooses (Herpestes auropunctatus) and 97 mice (Mus musculus) were caught in live traps. Leptospiral antibodies were detected by microscopic agglutination test (MAT) using antigens representing 12 serogroups, and kidney tissues were inoculated into polysorbate medium for isolation of leptospires. The seroprevalence (at a titre of > or = 100) in mice was 28.2% (24/85, 95% CI 19.0, 39.1) and in mongooses 40.7% (48/118, 95% CI 31.7, 50.1). In mice, antibodies were detected predominantly against serogroups Ballum and Autumnalis, while in mongooses the predominant serogroup was Autumnalis. Leptospires were isolated from 28 mice (28.9%, 95% CI 20.1, 39.0) and from 4 mongooses (2.9%, 95% CI 0.8, 7.4). Mouse isolates were identified as serovars arborea (17) and bim (7). As in other parts of the world, common house mice (Mus musculus) represent a significant reservoir of leptospirosis. Although carriage of the Ballum serovar, arborea, was not unexpected, this represents the first time that an animal reservoir of serovar bim has been identified. This is significant because bim causes about 63% of human leptospirosis in Barbados, and control efforts and education for prevention can now be targeted at a specific reservoir.     

问号钩端螺旋体胶原酶体内和体外的活性研究

目的 通过检测致病性钩端螺旋体(钩体)胶原酶在体内、外的活性,探讨问号钩体黄疸出血群赖型赖株(赖株)假定的胶原酶(LA0872)在钩体病出血中的可能作用.方法 在大肠杆菌中克隆、表达重组赖株la0872,并行相关鉴定及蛋白酶学活性检测.比较不同毒力菌株赖株、赖株减毒株和双曲钩体三宝垄群montevalerio型Monte Valerio株钩体胶原酶的基因序列特异性及转录和酶活水平的差异.测定豚鼠和沙鼠赖株感染模型的血清胶原酶活性水平变化.结果 成功构建的重组质粒经酶切和测序鉴定显示位点连接正确,插入序列与GenBank公布的la0872序列完全一致,并证实其具有胶原酶活性;不同毒力菌株中的钩体胶原酶基因序列一致,转录和酶活水平均无显著性差异;豚鼠和沙鼠感染赖株后,宿主体内血清胶原酶活性水平与未感染赖株动物比较差异无统计学意义(P>0.05).结论 首次表达和鉴定了有活性的钩体胶原酶,但其在钩体病出血中的作用尚待证实和探讨.