目的:分离获得含半透明仔虾病(translucent post-larvae disease,TPD)毒力因子的副溶血弧菌及裂解该弧菌的噬菌体,并评估噬菌体用于防治水产养殖行业弧菌感染的可能性.方法:利用选择平板法和分子生物学技术分离鉴定弧菌菌株,然后通过双层平板法分离获得噬菌体,并对其进行表征和全基因组测序分析.结果:分离到一株含TPD毒力因子的副溶血弧菌VP392和可裂解该菌株的噬菌体BP471,该噬菌体感染宿主菌最佳感染复数(multiplicity of infection,MOI)为1,潜伏期为20 min,裂解量为181 PFU/cell;在pH 3～11、温度55℃以下、盐浓度50 g/L以下均具有良好的稳定性,可明显抑制宿主菌生长且具有较广的宿主谱;全基因组测序表明其为双链DNA病毒,无毒力因子和抗生素抗性基因.结论:成功分离到一株可裂解含TPD毒力因子的副溶血弧菌噬菌体,其稳定性好、裂解谱广、安全性好,有望作为安全、环保型生物制剂应用到TPD防治中.
Isolation and Characterization of a Highly Eficient Lytic Phage against Highly Pathogenic Vibrio parahaemolyticus
Objective:To isolate phages with good lytic ability to Vibrio parahaemolyticus with translucent post-larvae disease(TPD)virulence factors and evaluate their potential application in the prevention or treatment of Vibrio infections in aquaculture industry.Methods:Vibrio strains were isolated using selective plates and screened by molecular identification.A phage was separated by double-layer plate method,and evaluated through a series of characterization experiments and whole genome sequencing analysis.Results:Vibrio parahaemolyticus strain VP392 with a translucent virulence factor and a lytic phage BP471 capable of lysing VP392 were isolated.The optimal multiplicity of infection(MOI)of phage BP471 was 1,the latent period was 20 min,and the burst size was 181 PFU/cell;phage BP471 retained good stability at temperatures below 55℃,salt concentration of 50 g/L,and pH 3-11.Phage BP471,characterized as a double-stranded DNA virus devoid of virulence factors and antibiotic resistance genes by whole-genome sequencing analysis,was able to inhibit the growth of its host bacterium in vitro.Conclusion:A lytic phage with good activity against Vibrio parahaemolyticus containing TPD virulence factors was successfully isolated.Phage BP41 has good stability,safety,and broad host spectrum,and may be a safe and environmentally friendly biological agent for the treatment of TPD in aquaculture.
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