中国“三江并流”自然遗产地兽类蜱媒疾病分子流行病学的调查研究
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摘要
目的调查和了解我国“三江并流”自然遗产地小型兽类、家犬自然感染伯氏疏螺旋体、嗜吞噬细胞无形体、埃立克体、斑点热群立克次体4种蜱媒疾病的情况及其病原基因特征,应用于监测分析山区兽类自然感染新发蜱媒疾病的情况。方法以“三江并流”自然遗产地的高黎贡山、云岭、中甸雪山、怒山等山脉(海拔1000-4500米之间)林区为调查的空间范围,小型兽类和家犬为研究对象,用鼠夹和鼠笼捕捉小型兽类分类鉴定登记编号后,现场分别取脾脏器组织；另取山村家犬全血。所获材料都以低温冻存和运输,带回实验室应用聚合酶链反应(PCR)方法对伯氏疏螺旋体、嗜吞噬细胞无形体、埃立克体、斑点热群立克次体4种蜱媒病原基因片段进行扩增并测.序,并将所测序列与GenBank中注册的基因序列进行相似性比较。结果共采集和检测小型兽类标本5目18属40种共688只,其中,发现3目10属共17种小兽分别与下述3种蜱媒疾病有关：1)莱姆病(8属10种小兽自然感染伯氏疏螺旋体,感染率为2.47%)；2)人粒细胞无形体病(6属11种小兽自然感染率嗜吞噬细胞无形体,感染率为4.65%)；3)埃立克体病(3属4种小兽自然感染埃立克体,感染率为0.73%)；经x2检验三种病原体的感染率有显著差异。检测中未发现小兽感染斑点热群立克次体。此外,共检测63只家犬血液,其中发现9只感染伯氏疏螺旋体,阳性率14.29%,基因序列分析结果显示,当地犬感染的伯氏疏螺旋体为Borrelia garinii基因型。此次调查未发现家犬中感染嗜吞噬细胞无形体、埃立克体、斑点热群立克次体。结论首次证实和发现我国西南横断山区3目10属17种小型曾兽类中自然感染伯氏疏螺旋体、嗜吞噬细胞无形体和埃立克体；证实当地犬自然感染伯氏疏螺旋体,基因型为Borrelia garinii。当地可能存在上述蜱媒病的自然疫源地和可能发生一些相关蜱媒传染病,应引起公共卫生部门的重视。
Objective The study was to investigate the natural infection and genetic characteristics of Borrelia burgdorferi, Anaplasma phagocytophilum, Ehrlichia sp and spotted fever group rickettsiae in wild small mammals and domestic dogs from Three Parallel Rivers Landscape,China.The results were used to monitor natural infection of mammals in mountain emerging tick-borne disease situation. Methods Wild small mammals and domestic dogs were caputured from Gaoligong mountain, Yunling mountain,Zhongdian snow mountain,Nushan mountain in the Natural Protected Territory, "Three Parallel River Region"(altitude:1000-4500m). Box traps and the cages were used to capture wild small mammals. After identification of species, the spleen was removed from each wild small mammal and stored at -196℃until tested. Domestic dog's blood samples were collected using vacuum blood extraction.Samples were transported to laboratory by low temperature freezing. All samples were tested by polymerase chain reactions (PCRs) for tick-borne agents(Borrelia burgdorferi sensu lato, Anaplasma phagocytophilum, Ehrlichia sp and spotted fever group rickettsiae). Positive samples were sequenced and compared with corresponding sequences deposited in Genbank. Results A total number of 688 small mammals, which belongs to 5 orders 18 genera 40 species were collected. Among them,17 species(3 orders 10 genera) of small mammals related with three kinds of tick-borne diseases. 1)Lyme Disease (10 species in 8 genera of small mammals naturally infected with Borrelia burgdorferi, infection rate was 2.47%); 2)Human granulocytic anaplasmosis (11 species in 6 genera of small mammals naturally infected Anaplasma phagocytophilum, infection rate was 4.65%); 3)Ehrlichiosis (4 species in 3 genera of small mammals naturally infected with Ehrlichia sp, infection rate was 0.73%); The positive rate of Borrelia burgdorferi,Anaplasma phagocytophilum, Ehrlichia sp in small mammals was significant difference by theχ2 test. Spotted fever group rickettsia was not found in small mammals in this study. In addition,63 domestic dog's blood were tested.9 naturally infected with Borrelia burgdorferi. The positive rate was 14.29%.The sequence analysis showed that the B. burgdorferi s.1. in dogs all belonged to B. garinii genotypes. Anaplasma phagocytophilum, Ehrlichia sp and spotted fever group rickettsiae were not found in dogs.
Conclusion Borrelia burgdorferi,Anaplasma phagocytophilum, Ehrlichia sp was firstly identified in 17 species(3 orders 10 genera) from Three Parallel Rivers Landscape,China. B. garinii was firstly identified in dogs in this areas.There were more than one tick-borne pathogen prevalence in Three Parallel Rivers Landscape,which should be paid attention by local public health authorities.
Objective The study was to investigate the natural infection and genetic characteristics of Borrelia burgdorferi, Anaplasma phagocytophilum, Ehrlichia sp and spotted fever group rickettsiae in wild small mammals and domestic dogs from Three Parallel Rivers Landscape,China.The results were used to monitor natural infection of mammals in mountain emerging tick-borne disease situation. Methods Wild small mammals and domestic dogs were caputured from Gaoligong mountain, Yunling mountain,Zhongdian snow mountain,Nushan mountain in the Natural Protected Territory, "Three Parallel River Region"(altitude:1000-4500m). Box traps and the cages were used to capture wild small mammals. After identification of species, the spleen was removed from each wild small mammal and stored at -196℃until tested. Domestic dog's blood samples were collected using vacuum blood extraction.Samples were transported to laboratory by low temperature freezing. All samples were tested by polymerase chain reactions (PCRs) for tick-borne agents(Borrelia burgdorferi sensu lato, Anaplasma phagocytophilum, Ehrlichia sp and spotted fever group rickettsiae). Positive samples were sequenced and compared with corresponding sequences deposited in Genbank. Results A total number of 688 small mammals, which belongs to 5 orders 18 genera 40 species were collected. Among them,17 species(3 orders 10 genera) of small mammals related with three kinds of tick-borne diseases. 1)Lyme Disease (10 species in 8 genera of small mammals naturally infected with Borrelia burgdorferi, infection rate was 2.47%); 2)Human granulocytic anaplasmosis (11 species in 6 genera of small mammals naturally infected Anaplasma phagocytophilum, infection rate was 4.65%); 3)Ehrlichiosis (4 species in 3 genera of small mammals naturally infected with Ehrlichia sp, infection rate was 0.73%); The positive rate of Borrelia burgdorferi,Anaplasma phagocytophilum, Ehrlichia sp in small mammals was significant difference by theχ2 test. Spotted fever group rickettsia was not found in small mammals in this study. In addition,63 domestic dog's blood were tested.9 naturally infected with Borrelia burgdorferi. The positive rate was 14.29%.The sequence analysis showed that the B. burgdorferi s.1. in dogs all belonged to B. garinii genotypes. Anaplasma phagocytophilum, Ehrlichia sp and spotted fever group rickettsiae were not found in dogs.
Conclusion Borrelia burgdorferi,Anaplasma phagocytophilum, Ehrlichia sp was firstly identified in 17 species(3 orders 10 genera) from Three Parallel Rivers Landscape,China. B. garinii was firstly identified in dogs in this areas.There were more than one tick-borne pathogen prevalence in Three Parallel Rivers Landscape,which should be paid attention by local public health authorities.
引文
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[29]解放军军事医学科学院微生物流行病研究所.对我国斑点热、莱姆病、人埃立克体病等蜱媒传染病的发现与研究.科技成果,2006,2,15
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[2]权寿瑛,龚正达,张丽云,等.云南横断山区小型兽类多样性的空间分布格局.生命科学,2009,3(7)：26-39.
[3]权寿瑛,岳仁苹,张丽云,等.云南省横断山区小型兽类的组成及空间分布.中国媒介生物学及控制杂志,2010,21(1)：16-22.
[4]龚正达,自登云,冯锡光,等.云南西部横断出区蜱类的组成及其分布.医学动物防制,2001,17(1)：31-33.
[5]栗冬梅,俞东征,刘起勇,龚正达.云南省不同环境鼠形动物巴尔通体感染情况的研究.中华流行病学杂志,1994,25(11)：934
[6]侯中柳,黄文丽,等.首次从云南啮齿和食虫动物中分离到森林脑炎病毒.中国病毒学,1992,7(4)：397-403
[7]陶三菊,张海林,杨冬荣,等.云南省澜沧江下游地区虫媒病毒的调查研究.中华实验和临床病毒学杂志,2003,17(4)：322-326.
[8]张海林,陶三菊,杨冬荣,等.云南省分离到辛德毕斯、巴泰、可提病毒.中国人兽共患病杂志,2005,21(7)：548-557
[9]张海林.云南省虫媒病毒研究进展.中国媒介生物学及控制杂志,2004,15(5)：410-411.
[10]高东旗,曹务春,赵秋敏,等.我国北方蜱中人粒细胞埃立克体16SrRNA斟因的检测.寄生虫学与医学昆虫学报,2000,7(2)：103-108.
[11]Cao WC, Zhao QM, Zhang PH, et al. Prevalence of Anplasma phagocytophila and Borrelia burgdorferi in Ixodes persulcatus ticks from northeastern China. Am J Trop Med Hyg,2003,68(5): 547-550.
[12]詹林,何静,萨仁高娃,等.东北林区鼠类粒细胞埃立克体感染的调查.中华流行病学杂志,2007,28(2)：157-159.
[13]Zhang LJ, Liu Y, Ni DX, et al. Nosocomial Transmission of Human Grannulocytic Anaplasmosis in China. JAMA,2008,300(19):2263-2270.
[14]Zhan L, Chu CY, Zuo SQ, et al. Anaplasma phagocytophilum and Borrelia burgdorferi in rabbits from southeastern China.Vet Parasitol,2009,162(4):354-356.
[15]高燕,何静,王剑波,等.我国东北林区蜱中伯氏疏螺旋体的分离及分型鉴定.中华微生物学和免疫学杂志,2006,26(5)：404-408.
[16]Cao WC., Gao YM., Zhang PH., et al. Identification of Ehrlichia chaffeensis by nested PCR in ticks from Southern China. J. Clin microbiology,2000,38(7):2778-2780.
[17]Wen BH., Cao WC., Pan H., Ehrlichiae and ehrlichial diseases in china. Annals of the New York Academy of Sciences,2002,40(9):3286-3290.
[18]张泮河,曹务春,张习坦,等.我国斑点热立克次体新种检测及其rOmpA基因序列分析.中国公共生,2000,16(6)：815-818
[19]刘国平,吴益民,杨国平,等.我国东北地区蜱类与黑龙江斑点热.沈阳部队医药,2008.21(2)：99-103.
[20]张哲夫,万康林,张金声,等.我国莱姆病的流行病学和病原学研究.中华流行病学杂志,1997,18(1)：8-11
[21]万康林,张哲夫,窦桂兰,等.我国20个省市自治区动物莱姆病初步调查.中国媒介生物学及控制杂志,1998,9(5)：366-371.
[22]Bakken JS, Dumle JS,Chen SM, et al. Human granulocytic ehrlichiosis in the upper Midwest United States. A new species emerging? JAMA,1994,272(3):212-218.
[23]曹务春,张习坦.人埃立克体病的发现与研究进展.中国人兽共患病杂志,1997,13(4)：57-60.
[24]Cao WC, Zhan L, He J, et al. Natural Anaplasma phagocytophilum infection of ticks and rodents from a forest area of Jilin Province, China. Am J Trop Med Hyg,2006,75(4):664-668.
[25]Cao WC, Zhao QM, Zhang PH, et al. Granulocytic Ehrlichiae in Ixodes persulcatus ticks from an area in China where Lyme disease is endemic. J Clin Microbiol,2000,38(11):4208-4210.
[26]Telford SR, Dawson JE, Katavlos P, et al. Perpetuation of the agent of human granulocytic ehrlichiosis in a deer tick-rodent cycle. Proc Natl Acad Sci USA,1996,93(12):6209-6214.
[27]Zhan L, Cao WC, Vlas SD, et al. A newly discovered Anaplasma phagocytophilum variant in rodents from southeastern China. Vector Borne Zoonotic Dis,2008,8(3):369-380.
[28]Wen B., R. Jian, Y Zhang, et al. Simultaneous detection of Anaplasma marginale and a new Ehrlichia species closely related to Ehrlichia chaffeensis by sequence analyses of 16S ribosomal DNA in Boophilus microplus ticks from Tibet. J. Clin. Microbiol.2002,40(9): 3286-3290.
[29]解放军军事医学科学院微生物流行病研究所.对我国斑点热、莱姆病、人埃立克体病等蜱媒传染病的发现与研究.科技成果,2006,2,15
[30]马海滨,张青,杨文映,等.云南西部林区莱姆病血清流行病学调查报告.中国媒介生物学及控制杂志,1992,3(特2)：70-72.
[31]李雪梅,张德荣,陈存,等.云南玉溪地区首次发现斑点热.中华流行病学杂志,2001,22(1)：76.
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