冷水鱼固着类纤毛虫PCR检测方法探讨及感染因素分析

Discussion of PCR detection method and investigation of infection factors of sedentarius ciliates in cold-water fish

针对冷水鱼养殖中固着类纤毛虫检测难和污染源不明确、种类鉴定难的问题,对冷水鱼养殖环境中固着类纤毛虫的主要寄生种类及感染因素进行调查分析,在不同季节于河北省内采集冷水鱼和水环境样本。首先,采用形态学方法对鱼体及水环境中的固着类纤毛虫进行初步鉴定;然后,建立PCR方法进行分子水平鉴定,并进行系统发育学分析;最后,对冷水鱼感染寄生虫的原因进行分析。形态学鉴定结果表明,冷水鱼寄生的固着类纤毛虫主要有累枝虫(Epistylis sp.)和钟虫(Vorticella sp.)2类;引物设计、敏感性和特异性试验结果显示,建立的寄生虫PCR检测方法有效,测序比对和系统发育分析结果进一步验证了其形态学结果;运用已建立的PCR方法检测出固着类纤毛虫在水环境中高达94.38%,鱼体广泛寄生,感染率达到了100%;且无季节差异,水体中的虫体可能是造成鱼长期感染的原因之一。研究数据可为冷水鱼病害的科学预测、预防和治疗提供依据,同时为冷水鱼养殖产业的可持续发展提供疫病防治参考。

Aiming at the problem of difficult detection of sedentarius ciliates,unclear pollution sources and difficult identification of species in the cold-water fish farming,the major parasitic species of the parasites and the infective factors were investigated and analyzed.Samples of cold-water fish and water were collected in different seasons in Hebei Province.Firstly,the sedentarius ciliates on the fish body and in the water environment were initially identified by using morphology.Secondly,a PCR method was established for molecular level identification and phylogenetic analysis.Finally,the reasons for cold-water fish infecting the parasite were analyzed.Morphological identification reveals that the mainly parasitic species of sedentarius ciliates were Epistylis sp.and Vorticella sp.;A PCR method to test the parasites is successfully established.The results of sequences alignment and phylogenetic analysis verify the correctness of morphological test.The established PCR method is used to detect the prevalence of sessile ciliates in the water environment,which is as high as 94.38%,and the infection rate of the fish is up to 100%with no seasonal difference.The parasites in water may be one of the reasons for the long-term infection of the fish.The findings provide a scientific basis for predicting,preventing and treating cold-water fish diseases,as well as serving as a reference for disease prevention and control in the sustainable development of the cold-water fish farming industry.