日本血吸虫胞蚴的体外培养和操作初探  

Preliminary investigation on in vitro-cultivation and manipulation of Schistosoma japonicum sporocysts

作　　者：江艳[1,2] 龚仁敏[1] 杨艺[2] 李小红[2] 刘毅[2] 朱传刚[2] 陆珂[2] 郑浩[2] 

机构地区：[1]安徽师范大学生命科学学院,芜湖241000 [2]中国农业科学院上海兽医研究所 动物寄生虫病重点实验室,上海200241

出　　处：《国际医学寄生虫病杂志》2011年第3期158-162,共5页International JOurnal of Medical Parasitic Diseases

基　　金：基金项目：国家公益性行业（农业）科研专项基金（200903036）

摘　　要：目的探究一种体外培养日本血吸虫胞蚴的新方法，并通过体外注射，感染阴性钉螺，培养阳性钉螺。方法将日本血吸虫虫卵孵化的毛蚴与作为滋养层的草地夜蛾（Spodoptera frugiperda）卵巢细胞（庐细胞）共培养，观察毛蚴的生长情况，并拍摄生长照片。用显微注射针将3～5只母胞蚴注射到钉螺体内，计算钉螺的存活率及尾蚴的阳性率，用钉螺逸出的尾蚴人工感染昆明鼠，检测尾蚴的活力，用组织化学染色法和PCR方法鉴定尾蚴的相关基因特性。结果毛蚴培养至第12小时观察到纤毛板已脱落，培养2～3d的虫体呈不断的伸缩运动，此后母胞蚴维持该状态并不转化成子胞蚴。3次实验的钉螺存活率依次为24．72％、28．23％和57．89％，活螺阳性率依次为4．55％、8．57％和14．29％。阳性钉螺释放的尾蚴具有感染性，且具有与日本血吸虫相同的基因特性。结论体外培养毛蚴通过显微注射可以人工感染钉螺并逸出有感染活力的尾蚴，为日本血吸虫幼虫的体外操作提供一种新的方法。Objective To explore a new method of in vitro cultivation of Schistosoma japonicum sporocysts, infection of negative snails through in vitro-injection, and cultivation of positive snails. Methods Iso- lated miracidia were directly cultivated in sf9 cells-conditioned medium, the growth of miracidia was observed and the growth picture was taken. 3-5 mother sporocysts were inoculated into the snails body using a microscopic needle. The survival rate of the snails and the positive rate of cercariae were calculated. Kunming mice were infected with the cercariae and the activity of cercariae was detected. The related gene characteristics of cercariae was identified by histochemical examination and PCR method. Results The pictures of miracidia were recorded after co-cultivation for 1 h, 12 h, 24 h and 3 d. The shedding of the cilia of miracidia occurred within 12 h, parasites exhibited continuous stretching movements after co-cultivated for 2-3 d. Since then the mother sporoeysts kept the status, did not develop to daughter sporocysts. The survival rates of snails in 3 experiments were 24.72% , 28.23 % and 57.89% respectively, while the positive rates of cereariae were 4.55% , 8.57% and 14.29% respectively. Animal experiments showed that the cercariae released from artificial cultivation of positive snails were infectious and had same gene characteristics of Schistosoma japonicum. Conclusion Sf9 cells could be feeder cells for in vitro-cultivation of miracidia and positive snails could be obtained by in vitro micro-injection of cercariae thus to provide a new approach for in vitro operation of Schistosoma japonicum larvae.

关 键 词：日本血吸虫 Spodopterafrugiperda卵巢细胞 钉螺 显微注射 

分 类 号：R383.24[医药卫生—医学寄生虫学]