斑马鱼“裂头病”病原的分离与鉴定及常见疾病诊断

Pathogen Isolation and Identification of The Zebrafish“Hole in The Head”Disease and Diagnosis of Common Diseases

目的斑马鱼是生命科学研究中应用最广的实验动物之一,其健康状况关乎实验数据的质量和结果的可信度。2018年以来,中国斑马鱼鱼房内出现危害性极高的新型鱼病“裂头病”,其病原未知,防治困难。确定“裂头病”的病原,并提出鱼房管理健康建议,可帮助研究者防止鱼病带来的损失。方法对“裂头病”病鱼脑组织进行病原分离,对分离的病原理化特征、16S rDNA同源性进行分析,采用回归感染试验确认病原菌。分析121例鱼病数据,总结中国斑马鱼鱼房各类常见疾病种类及发病因素。结果确定“裂头病”病原为鮰爱德华氏菌。2017至2021年国内斑马鱼鱼房数据显示,50.4%的健康问题由非生物因素直接或者间接导致;致死率最高的问题是气体中毒。在已知病原中细菌性病原最为常见,占比83.46%。结论“裂头病”病原为鮰爱德华氏菌。近一半鱼房健康问题是由非生物因素导致。致病率最高的为细菌性病原。因此,实验室日常工作中,加强设备维护,稳定优良水质,保证卫生管理措施,是防止鱼病发生的主要手段。

Objective Zebrafish is one of the most widely used model animals for scientific research in the world.The health and quality of zebrafish are the basis for accurate data and reliable results.With the rapid development of zebrafish related research in China,the number of fish diseases in research facilities has been increasing in recent years.However,the research and analysis on these diseases have been lacking.A novel lethal disease,“hole in the head”disease has spread in several zebrafish facilities since 2018,whose pathogen was unrecognized and hard to be controlled.Pathogen identification is urgently needed,and the analysis and advice of health administration are required for the fast-growing field.Methods We necropsied the brain tissue from the“hole in the head”disease zebrafish.After the brain tissue was homogenized,the pathogen was isolated and cultured on brain heart infusion agar(BHIA)plates.The isolated pathogen was analyzed for biochemical characteristics and 16S rDNA homology.The Koch’s Postulate was fulfilled to confirm the pathogen.In addition,we analyzed the zebrafish disease data collected by China Zebrafish Resource Center(CZRC)from 2017 to 2021and summarized the characteristics of various common diseases.Results The isolated and identified pathogen of the“hole in the head”disease has the same physicochemical properties with Edwardsiella ictaluri,which also fulfilled the Koch’s Postulate.The 16s DNA sequence of this pathogen was highly homologous to Edwardsiella ictalurid.From 2017 to 2021 CZRC has collected 121 disease and fish death cases in Chinese zebrafish facilities.We found that 50.4%fish health problems were directly or indirectly caused by equipment failure and abiotic factors,including 12.39%fish death caused only by improper equipment maintenance and water quality problems.The most lethal problem was gas poisoning.Among all the known pathogens,bacterial pathogens were the most common,accounting for 83.46%,fungal pathogens accounted for 2.47%,and parasitic pathogens accounted for 1.65%.Among all the known bacterial diseases,the opportunistic pathogens of the genus Aeromonas were the most common,followed by Edwardsiella,accounting for 34.65%and 23.76%respectively.Conclusion Edwardsiella ictaluri was isolated and identified as the pathogen of“hole in the head”disease.The highest incidence of zebrafish disease is bacterial infection,and nearly half of zebrafish health problems are caused by abiotic factors.Well-maintained equipment and proper water quality are the most important prerequisites to ensure the health of zebrafish,managements of daily hygiene are the main means to prevent fish diseases,and the prevention of bacterial diseases is the top priority of zebrafish farming.