蝙蝠体表吸血昆虫携带病毒及其与蝙蝠和其他人畜动物感染关系研究

Many ectoparasites (bat insects) known as spider flies, bat flies etc. feed on blood of bats. Several pathogenic agents such as gammaproteobacteria, bartonella and malaria have been isolated from these insects, which indicate that bat insects maybe act as the “external reservoir” for pathogenic agents carried and transmitted by bats. However, up to now, no viruses were isolated from these bat insects in the world. Since 2012, we isolated several strains of viruses from bat insects. Preliminary studies showed that some of them are members of Flavivirus, Orthobunyavirus and Reoviridae, some of them leave unknown. This is the first to isolate viruses from blood-feeding ectoparasites on bats in the world. This project plan to perform following study: 1) Biological and molecular biological characteristic of these isolates, and clearing their taxonomic classification; 2) Further investigate the virus resources carried by bat insects and their genetic evolution using virus culture and deep sequencing techniques; 3) Serological study for the infection of the viruses carried by bat insects with bats, livestock and human beings. The results will elucidate the relationship between viruses carried by bat insects and viruses carried or transmitted by bats. In addition, the investigation of infection of viruses carried by bat insects in animals and human beings will be important for public health.

蝙蝠体表吸血昆虫（下称蝙蝠昆虫）如蛛蝇、蝠蝇等以吸食蝙蝠血为生，已经从蝙蝠昆虫分离到多种细菌、巴尔通体和疟疾等病原微生物，提示蝙蝠昆虫可能是蝙蝠携带和传播病原体的“外储存库”，但至今国内外未见从蝙蝠昆虫分离病毒报道。2012年以来本课题组从蝙蝠体表吸血昆虫标本分离到多株病毒分离物，初步鉴定结果显示病毒分离物中含黄病毒、布尼亚病毒和呼肠孤病毒及多株目前尚未鉴定的病毒，提示病毒也是蝙蝠昆虫携带的重要病原体。本申请拟开展1）对已经分离到病毒分离物开展生物学表型和分子生物学特征研究，明确其分类地位等；2）使用病毒分离培养和大规模基因组序列测定技术进一步挖掘蝙蝠昆虫携带病毒资源并开展病毒分子遗传进化分析；3）开展血清流行病学研究以明确蝙蝠及人畜动物对蝙蝠昆虫携带病毒感染状况。研究结果对于阐明蝙蝠体表吸血昆虫携带病毒及其与蝙蝠携带与传播病毒的关系，及其与人畜动物感染关系等具有重要理论意义与公共卫生意义

蝙蝠体表吸血昆虫如蛛蝇、蝠蝇等以吸食蝙蝠血为生，已经从吸血昆虫中发现多种细菌、巴尔通体和疟疾等病原微生物，提示吸血昆虫可能是蝙蝠携带和传播病原体的“外储存库”。本项目在云南省12个采集点采集到不同种类的蝙蝠及其体表吸血昆虫，对采集的吸血昆虫在形态学分类的基础上，利用DNA条形码技术对吸血昆虫利用节肢动物线粒体细胞色素氧化酶Ⅰ进行PCR扩增后，分为蛛蝇、蝠蝇、革螨、蜱和跳蚤5个种类。将5类昆虫分为9个pools提取核酸后送华大基因进行RNASeq二代测序，发现病毒reads数占总reads数的4.07%。从蝙蝠血清和脾脏中分离到2株病毒，蛛蝇中分离到8株病毒，这是国际上首次从蝙蝠体表蛛蝇中分离到病毒。从3个采集点的蛛蝇中分离到6株肯科伊病毒（Kaeng Khoi virus，KKV），提示KKV病毒不仅具有传播媒介的多样性、地域分布的广泛性同时还具有物种多样性，各分离株在M基因核苷酸和氨基酸存在较大变异。用间接免疫荧光实验对人血清进行IgG抗体检测，结果有9份IgG为阳性，证实蝙蝠体表吸血昆虫中分离的KKV与人类疾病之间可能存在关系。从水鼠耳蝠血清中分离到1株横须贺病毒（Yokose virus），与原始分离株之间在病毒分子生物学和病毒抗原性方面存在较大差异，横须贺病毒日本分离株（Oita36）与黄热病毒之间存在中和抗体的交叉反应，但是横须贺病毒云南分离株（XYBX1332）与乙脑病毒间存在中和抗体反应但是与黄热病毒间不存在中和抗体反应。从蛛蝇和蝙蝠的脾脏中分离到3株呼肠孤病毒（Orthoreovirus），提示蛛蝇有可能是该病毒的传播媒介。还从蝙蝠体表吸血昆虫中检测到巴尔通体和沃尔巴克氏体。这些研究结果均提示蝙蝠及其体表吸血昆虫携带多种重要病原体，与人畜动物感染关系等具有重要理论意义与公共卫生意义。目前研究结果发表SCI论文3篇，英文论文1篇，中文论文8篇。经费使用过程按照预算和经费管理制度执行。本项目已按考核指标完成任务，按要求可顺利结题。