【作者】 郭刚；

【导师】 姚华；

【作者基本信息】 新疆医科大学 ， 劳动卫生与环境卫生学， 2016， 博士

【摘要】 目的:2010-2015年期间在新疆北疆地区选择四类不同生境(包括乌鲁木齐、伊宁及阿拉山口在内的城区、精河县南山牧区、准噶尔盆地西缘半荒漠生境和新源县那拉提草原),选择不同方法捕获生境内鼠类和采集体外寄生生物,以优势鼠种和寄生媒介为主分别进行汉坦病毒、莱姆螺旋体、立克次体、利氏曼原虫等病原体检测及开展遗传进化分析,完善北疆地区鼠传疾病研究并为针对相关疾病做到早期管控提供依据。方法:1)采用不同捕获方法,收集北疆地区四类不同生境鼠种及体表寄生生物,开展物种形态学鉴定,并将宿主动物按要求进行解剖取材和保存;2)利用分子生物技术检测褐家鼠汉坦病毒的M基因部分片段,通过做遗传进化分析确定褐家鼠病毒感染及变异情况。同时检测褐家鼠线粒体细胞色素b基因(Cytb),了解新疆输入鼠种的遗传变异情况;3)检测伊犁普通田鼠Cytb基因和HV携带情况,选择初筛阳性样本开展S基因测定并做遗传进化分析;4)开展大沙鼠及寄生媒介蜱、蚤携带莱姆螺旋体、利什曼原虫及立克次体等病原分子生物学检测,掌握病原携带流行病学情况;5)开展长尾黄鼠立克次体、莱姆螺旋体等病原携带检测,并对检测到立克次体运用多位点序列分析(MLSA)技术,研究病原种属地位。结果:1)从北疆区域不同生境共捕获鼠类优势种1135只(其中褐家鼠458只,大沙鼠226只,长尾黄鼠253只,普通田鼠198只),蚤5123匹,螨699只,蜱3635只;2)从乌鲁木齐市区褐家鼠脏器组织中检测到汉城病毒(SEOV)核酸阳性样本,阳性率为15%,呈高携带状态。M片段核酸序列与疆外及国外相比,未表现出遗传多样性及地理聚集特征;3)新疆地区褐家鼠Cytb全基因检测,并与疆外及国外褐家鼠Gen Bank上传序列开展比对分析,未发现明显独立进化的现象;4)通过开展Cytb基因检测确定伊犁新源县那拉提草原采集普通田鼠为普通田鼠北疆亚种(Microtus arvali obscurus),并首次在中国境内从普通田鼠北疆亚种检出图拉型汉坦病毒,阳性率为16%,通过对S基因做遗传进化分析发现其与哈萨克斯坦东部检出毒株更为近源;5)从准噶尔盆地西缘阿拉山口口岸周边捕获大沙鼠和体外寄生亚洲璃眼蜱检出狭义莱姆螺旋体(B.burgdorferi sensu strict)阳性核酸片段,阳性率分别为24%,3%;检出嗜吞噬细胞无形体(Anaplasma phagocytophilum)和查菲埃立克体(Ehrlichia chaffeensis)阳性核酸片段,阳性率分别为8%,31%和5%,34%;从大沙鼠和体外寄生臀突客蚤检出都兰利氏曼原虫(Leishmania turanica)阳性核酸片段,阳性率分比为6%和2%;6)从长尾黄鼠和体外寄生草原革蜱检出立克次体核酸阳性片段,阳性率分别为34%和53%,经多序列位点分析确定为立克次体属斑点热群劳氏立克次体(Rickettsiae raoultii)。结论:1)虽然目前新疆无本地肾综合征出血热(HFRS)病例,但乌鲁木齐部分区域褐家鼠SEOV出现高携带率,提示在乌鲁木齐褐家鼠已经作为保存宿主参与SEOV在当地的自然循环,乌鲁木齐存在导致HFRS发病的病原、宿主和易感人群基础。今后需要开展以乌鲁木齐为重点的北疆城区人群和宿主动物汉坦病毒感染的长期监测和研究工作,并采取措施减少人群暴露机会;2)本研究中除在乌鲁木齐和伊犁新源县那拉提草原分别捕获褐家鼠和普通田鼠检测到汉坦病毒核酸以外,其余所有鼠种均为阴性,提示其他鼠种所在生境可能不存在汉坦病毒传播,但也不排除该宿主动物对汉坦病毒不易感的可能;3)从通过分子鉴定确认的普通田鼠北疆亚种组织样本检测到图拉病毒核酸,为我国境内新发现的新型汉坦病毒,丰富了我国境内汉坦病毒的种属分型,为全面做好我国汉坦病毒感染防控提供了依据;4)从准噶尔盆地西缘鼠疫宿主-大沙鼠及体外寄生生物检出莱姆、无形体、埃立克体及利氏曼病原核酸片段阳性样本,表明准噶尔盆地大沙鼠栖息地半荒漠生境应当是多种人兽共患病和病媒生物性疾病病原混合共存的自然疫源地,应当加强相关宿主、媒介和病原监测工作。另外从臀突客蚤检出都兰利氏曼原虫,提示除白蛉作为主要媒介传播皮肤利什曼病以外,蚤类也有可能作为伴随媒介参与人群感染,应继续开展相关研究;5)首次运用分子生物检测技术和MLSA方法从精河天山山地草原生境鼠疫宿主-长尾黄鼠及寄生草原革蜱中检出劳氏立克次体,提示在该生境内可能会造成当地人群立克次体感染,在该区域开展包括鼠疫、立克次体等病原在内的疾病监测研究具有重要的公共卫生学意义。更多还原

【Abstract】 Objective: To determine the presence, distribution, hereridity and evolution of the relative pathogens of hantavirus, Borrelia burgdorferi, Rickettsiae, Leishmania spp., etc from the rodents and ectoparasites, dominant species especially in the four types of ecotopes in northen region of Xinjiang Uygur Autonomous Region(XUAR), northwest of China, which consisted of semi-desert area(surrounding the port of Alashankou, west of Junggar basin), urban areas(Alashankou, Urumqi and Yining city), subalpine meadow(Nalat Grassland), mountain meadows(Nanshan mountain around Jinghe County) through collecting the rodents and ectoparasites by the different capture methods during the years 2010 to 2014. The theory basis is offered for the further study and early control of relative diseases in northern region of Xinjiang. Methods: 1) The samples of rodents and ectoparasites were collected and reserved accordance with the requirements after finishing the identification by morphology; 2) M gene segments of hantavirus were detected for identifying the Seoul virus infection and gene variation in Norway rat from diferent urban city of Northern Xinjiang, and Cytb genes of Norway rat were sequenced for understanding the evolution progress of Norway rat, as an immigrant species from outside Xinjiang in China; 3) Cytob gene and hantavirus nucleic acids were detected in Microtus arvalis from Nalat Grassland and the relations of hereridity and evolution about the positive samples were analysed ed for virus taxonomy. Meanwhile, Cytb genes of Microtus arvalis were sequenced and compared for affirming the taxnonomy; 4) The nucleic acids of B. Burgdorferi, Rickettsiae, Leishmania spp. in great gerbils and parasites(ticks and fleas) were detected for understanding the pathogenic infection status; 5) MLSA was done on the Rickettsiae positve sample from long-tailed ground squirrels and the parasites in Nanshan mountain meadows around Jinghe city for identifying the taxonomy and nucleic acid sequence of Borrelia burgdorferi, Orientia tsutsugamushi were also screened by PCR. Results: 1) The number of dominant mice speices that were sampled in different ecotopes is in total 1135 including great gerbils(Rhombomys opimus) 226, long-tailed ground squirrels(Citellus undulates) 253, Norway rats(Rattus norvegicus) 458 and Common vole(Microtus arvalis) 198, and parasites including fleas 5123, ticks 3635, mites 699; 2) The nucleic acids of SEOV were detected from the tissue samples of Norway rats with a high positive rate(15%) which were captured in the city of Urumqi; 3) Compared Cytb gene sequences of Norway rats from this study with those of outside Xinjiang in China and foreign countries in Gen Bank. There was no evidence that independent evolution happened in Norway rats from Xinjiang; 4) The rodent samples from Nalat grassland in Xinyuan County were identified as Microtus arvalis obscurus via Cytb gene sequencing and it was the first time that Tula virus(TULV, genus hantavirus) was found in Microtus arvalis obscurus in China; the positive rate is 16%. It showed that it was more related to the strains isolated in eastern Kazkhstan after PCR was done for S gene, which compared with the nucleotide sequences of TULV deposited in Gen Bank; 5) The nucleic acid segments of B. burgdorferi sensu strict were detected from great gerbils(24%) and its parasites-Hyalomma asiaticum(3%) around Alashankou in westen Junggar basin. The nucleic acid segments of AP(Anaplasma phagocytophilum) and EC(Ehrlichia chaffeensis) were also found and the positive rates were 8% and 31% for AP, 5% and 34% for EC. The nucleic acid segments of Leishmania turanica were detected from great gerbils and its parasites- Xenopsylla minax. The positive rates were 6% and 12%, respectively; 6) The nucleic acid segments of Rickettsiae(R.raoultii) were detected from long-tailed ground squirrels and the parasites- Dermacentor nuttalli. The positive rates were 34% and 53%, respectively. Conclusion: 1) Although local cases that involve hantavirus infections which results in HFRS have not been reported, our results indicated a positive rate(15 %) of the Norway rats carrying the SEOV, suggesting a high risk for the occurrence of HFRS in Urumqi City. The hantavirus circulates in Urumqi City where the Norway rat may act as a natural reservoir for the virus, indicative by its high hantavirus-positive rate. Our negative results indicate the possibility that these species are not susceptible for hantavirus infections or lack exposure to hantavirus infection. Based on our studies, long-term surveillance and examination of hantaviruses in human and rodent populations located in XUAR, and especially in Urumqi City, is necessary to identify novel hantavirus strains and ultimately reduce the rate of exposure; 2) Except for the hantavirus-positive resluts in the sampled Norway rats and Microtus arvalis obscurus from Urumqi and Nalat grassland respectively, the results of the other samples are all negative, which indicates the possibility that these species are not susceptible for hantavirus infections or lack exposure to hantavirus infection; 3) The nucleic acids of TULV detected from the Microtus arvalis obscure is a new type of hantavirus founded in China firstly and significant to the control and prevent of hantavirus; 4) The nucleic acid segments of B. b.s, AP, EC and L. Turanica were detected from the great gerbils and parasites in western Junggar basin; it indicates the semi-desert habitat should be a natural epidemic foci where multiple kinds of zoonosis pathogens coexist. More attention and control measures should be put into action. In addition, apart from the main vectors-sandfly, flea also could be a vector of infections occasionally, which leads to Cutaneous leishmaniasis among human as L. turanica was detected in Xenopsyllaminax and the related study should go further; 5) The nucleic acid segments of R.raoultii were detected from long-tailed ground squirrels and Dermacentor nuttalli. It suggested that there was an exposure risk to spotted fever pathogens in Nanshan moutain grassland and it deserved to make a long term monitoring on Rickettsiae.更多还原