基于ITS1-5.8S rRNA-ITS2序列的猪源结肠小袋纤毛虫种群特征分析

【目的】为了揭示结肠小袋纤毛虫病在环境中的分子传播机制,研究了猪源结肠小袋纤毛虫的种群特征。【方法】用粪便涂片镜检和改进型DMEM培养基从病猪结肠内容物分离结肠小袋纤毛虫滋养体,然后用显微观察、吖啶橙荧光染色法和基于ITS1-5.8S rRNA-ITS2序列的分子标记技术分析了豫西地区猪群中结肠小袋纤毛虫的种群特征。【结果】结果显示,从来自豫西地区8个县市的病猪中共分离了15株小袋纤毛虫,5.8S rRNA序列相似性高达99.4%以上,同属于结肠小袋纤毛虫,根据ITS1/2序列分析结果,MJ-2和SX-1株属于结肠小袋纤毛虫基因型A,其余13株均属于结肠小袋纤毛虫基因型B。MJ-2和SX-1株滋养体形态特征明显区别其他13株,绝大多数呈球形,运动缓慢,粪便中和体外培养的虫体密度较低;而其他13株的滋养体均呈多形性,运动快速活跃,虫体密度较大。吖啶橙荧光染色显示,2种基因型滋养体的核结构没明显差别。【结论】首次报道中国猪源结肠小袋纤毛虫存在2个基因型,其中基因型B为优势种群,为防控人和动物结肠小袋纤毛虫病提供重要参考。

Population characterization of Balantidium coli from pigs by using ITS1-5.8S rRNA-ITS2 sequence

Abstract: Objective] To determine the species and genotypes of Balantidium isolated from pigs in Henan province, China. Methods] Scatoscopy and the modified DMEM media were used to isolate trophozoites of Balantidium from pig feces. The ITS1-5.8S rRNA-ITS2-based molecular marker method, Acridine orange staining (AO) and microscopic observation were used to determine the population characteristics among different isolates of B. coli from various pigs farms. Results] We isolated 15 isolates from the pigs at diagnosis in the Animal Hospital of Henan University of Science and Technology from the pig farms of the 8 counties or cities of the west of Henan province in total, and all of them belong to the same species B. coli. MJ-2 and SX-1 isolates were genotype A of B. coli, and the remaining 13 isolates were genotype B. Trophozoites of MJ-2 and SX-1 were bigger, moved more slowly and lower density in feces in vitro than other 13 isolates, while structures of their nuclei were not different. Conclusion] Both genotype A and B of B. coli are present in the pig farms of the west of Henan province, China, and genotype B is the determinant population in pigs farms. These findings could provide an important implication for the effective control of balantidiosis of human and other hosts.