Diversity Of TCBS Group Bacteria In Shrimp Aquaculture System And Biological Characteristics Of Aeromonas Bacteriophages Isolated From A Shrimp Farm

The aquaculture animal diseases caused by explosive proliferation of pathogenicbacteria bring huge economic losses every year. Traditional therapy of bacteriadiseases in aquaculture was using antibiotics, but the frequent uses of antibiotics mayresult in emergence of drug-resistant pathogens as well as food safety problems interms of antibiotic residues in aquatic products. Bacteriophages were suggested to bea potential alternative of antibiotics due to some unqiue characteristics, such as highhost specificity, self-replication, and low environmental impact. The full name ofTCBS agar is thiosulfate-citrate-bile-sucrose agar, this kind of selective medium iswidely used to isolated aquaculture pathogenic Vibrios, other pathogenic bacteria alsocan grow in the medium. This study aims to1) investigate diversity of TCBS groupbacteria in shrimp aquacultute environemts,2) isolate lytic phages specific topotential pathogens in shrimp aquaculture systems,3) and investigate the invasioncharacteristics of phages to host bacteria. The results were summarized as follows.1. A total of73TCBS group bacteria were isolated from7aquaculture watersamples and5shrimp samples in3shrimp farms of Zhangpu, Fujian Province. TheseTCBS group bacteria were isolated from two different types of aquacultureenvironments: natural freshwater and artificial seawater.73strains were identified as13genera,26species based on16S rRNA sequence analysis. All Vibrio strains wereisolated from artificial seawater samples, accounting for45.2%of total isolatedstrains. The remaining strains showed high diversity and were mainly isolated formfreshwater aquaculture samples, as well as artificial seawater and shrimp samples.These results suggested that different water environments have great influences onTCBS group bacterial diversity and species distribution.2. A total of3potential pathogenic bacteria and their lytic phages were isolatedfrom the hepatopancreas of diseased shrimps.16S rRNA sequence analysis showedthat these three bacterial strains were affiliated with Aeromonas hydrophila (higher than99.85%of sequence similarity).3phages named as pIS-4, pIS-6, and pIS-8,showed strong lytic ability. TEM photographs showed that one phage belongs to theMyoviridae family, and the other two phages belong to the Siphoviridae family.3phages showed distinct biological characteristics when using stain IS-4as the targethost. One step growth curves showed that the latent period of pIS-4was20min, burstsize was85; the latent period of pIS-6was30min, burst size was109; the latentperiod of pIS-8was30min, burst size was155. The three phages were thermalrelative stable and the optimal temperature was25℃. The survival rates of threephages were higher than60%under25-60℃. All phage were stable at pH6-9. Theoptimal pH was8.0,7.0,7.0for pIS-4, pSI-6, and pSI-8, respectively.Conclusion: a total of73TCBS group strains were isolated from3shrimp farmsof Zhangpu, Fujian Province. All Vibrio strains were isolated from maricultureenvironment, while the strains from fresh water aquaculture environment showedrelative high diversity.3lytic phages specific to Aeromonas hydrophila were isolatedfrom the hepatopancreas of diseased shrimps. They are different species with distinctmorphological and biological characteristics. These three phages showed strong lyticability to Aeromonas hydrophila, indicating that they have great potential onprevention and treatment of pathogenic Aeromonas aquaculture disease.