一株虾源假交替单胞菌(Pseudoalteromonas sp.)的分离鉴定及口服后对对虾体内弧菌的影响

Isolation,identification and effects of a strain of Pseudoalteromonas sp.on Vibrio sp.in shrimp by oral administration

为探讨对虾细菌性病原的微生物防控技术,采用牛津杯打孔法对分离自对虾育苗池的一株编号为2021041402-14的细菌(简称“02HN”)进行了抗菌测试,分析其对欧文氏弧菌(Vibrio owensii)、哈维氏弧菌(V.harveyi)、坎贝氏弧菌(V.campbellii)、溶藻弧菌(V.alginolyticus)、副溶血弧菌(V.parahaemolyticus)和美人鱼发光杆菌(Photobacterium damselae)等6株虾病原菌的抑菌效果及最低抑菌浓度,通过浸泡感染测试了菌株02HN对凡纳滨对虾(Litopenaeus vannamei)的生物毒性,采用16S rRNA、gyrB-rpoD串联基因序列比对法及生理生化方法对该菌进行了鉴定,并将该菌添加到饲料中投喂对虾,评价该菌对对虾体内弧菌数量的影响。结果表明:菌株02HN对欧文氏弧菌、哈维氏弧菌、坎贝氏弧菌、溶藻弧菌、副溶血弧菌和美人鱼发光杆菌均具有抑菌效果,其最低抑菌浓度分别为2.32×10~4、2.32×10~3、2.32×10~6、2.32×10~6、2.32×10~3、2.32×10~6 CFU/mL;采用02HN浓度为1×10~7 CFU/mL以下的菌液浸泡凡纳滨对虾,生物毒性不明显;分子鉴定显示,该菌与杀鱼假交替单胞菌(Pseudoalteromonas piscicida)及金丽假交替单胞菌(P.flavipulchra)最为接近,为一株假交替单胞菌(Pseudoalteromonas sp.);对虾摄食菌含量分别为1×10~5、1×10~7cells/g的饵料后,其存活及生长无显著性变化,但能显著降低对虾肝胰腺中的弧菌数量(P<0.05)。研究表明,菌株02HN在对虾养殖中具有开发应用潜能,本研究结果可为对虾细菌性病原的生物防控提供生物材料及技术思路。

In order to explore the microbial control technology of bacterial pathogens of shrimp,an antibacterial test was performed by Oxford cup method on a strain of bacteria numbered 2021041402-14(referred to as 02HN)isolated from shrimp nursery pond.The bacteriostatic effect and minimum inhibitory concentration of strain 02HN against six shrimp pathogens were analyzed,including Vibrio owensii,V.harveyi,V.campbellii,V.alginolyticus,V.parahaemolyticus and Photobacterium damselae.Meanwhile the strain 02HN was identified by comparison of 16S rRNA gene,gyrB-rpoD tandem gene sequence and physiological biochemical indices.Also the effect of this bacterium on Vibrio density was evaluated in Pacific white leg shrimp(Litopenaeus vannamei)fed diets containing the strain 02HN at concentrations of 0(control group),1×10~5 and 1×10~7 cells/g diet.The results showed that strain 02HN had obvious bacteriostatic effects on V.owensii,V.harveyi,V.campbellii,V.alginolyticus,V.parahaemolyticus and Photobacterium damselae with the minimal inhibitory concentrations at 2.32×10~4,2.32×10~3,2.32×10~6,2.32×10~6,2.32×10~3 and 2.32×10~6 CFU/mL respectively.Challenge test at concentration of 1×10~7 CFU/mL showed that there was no biological toxicity of strain 02HN to the Pacific white leg shrimp.Molecular biological identification results revealed that strain 02HN was a strain of Pseudodotalteromonas sp.closing to P.piscicida and P.flavipulchra in taxonomic status.There was no significant effect of strain 02HN content of 1×10~5 cells/g and 1×10~7 cells/g diet on survival and growth of the shrimp fed the diets containing strain 02HN,but significant decrease in the density of Vibrio in hepatopancreas.It can be seen that strain 02HN has the potential for development and application in shrimp farming.The findings provide biological materials and technical support for the biological control of bacterial pathogens of shrimp.