Abstract:

Vibrio parahaemolyticus disease brings huge losses to the aquaculture industry. With the banning of antibiotics， people have started to explore novel therapeutic methods regarding the disease in aquatic animals， such as phage therapy. In this paper， 15 samples of seafood culture water and sewerage effluent were collected from Chengyang wholesale aquatic products market in Qingdao， and 12 strains of Vibrio parahaemolyticus phage were obtained by isolation and purification using the spot-drop method with Vibrio parahaemolyticus as the host bacterium. The biological characteristics of CHY5-M1M， such as the optimal infection complex， one-step growth curve， stability of temperature and pH and sensitivity to UV light， were studied using the double-layer plate method. The results showed that， the potency of CHY5-M1M phage was 8.65×10¹³ CFU·mL^-1； the potency was highest at 100 infection replicates； the latency period of one-step growth curve was 20 min， the lysis time was 100 min， the plateau was reached after 140 min； the potency of phage was highest at 40 ℃， the activity started to decrease above 60 ℃； the pH adaptation range of phage was 5~11. The phage concentration decreased significantly with the increase of UV irradiation time； the genome length was 43 193 bp， 44 genes， no virulence factor gene and antibiotic resistance gene. The results of this study indicated that phage CHY5-M1M has good application prospects in the development of novel anti-vibrio drugs for the prevention and treatment of severe vibriosis in marine animals， and could be an alternative antibacterial product that can be further investigated.

Key words: Vibrio parahaemolyticus, phage, biological properties, genome-wide analysis

摘要：

副溶血弧菌引起的疾病给水产养殖行业带来巨大损失，而随着抗生素的禁用，人们开始探索治疗水产动物病菌的新型方法，如噬菌体治疗法。从青岛市城阳水产品批发市场采集了15份海产品养殖水及下水道污水样品，以副溶血弧菌作为宿主菌，采用点滴法分离纯化获得12株副溶血弧菌噬菌体，通过双层平板法研究副溶血弧菌噬菌体CHY5-M1M的最佳感染复数、一步生长曲线、温度和pH的稳定性以及对紫外线的敏感度等生物学特性。结果显示，CHY5-M1M噬菌体效价为8.65×10¹³ CFU·mL^-1，且在100感染复数时效价最高；一步生长曲线的潜伏期为20 min，裂解时长100 min，140 min后达到平稳期；噬菌体在温度为40 ℃时效价最高，高于60 ℃时活性开始下降；噬菌体pH适应范围为5~11；噬菌体浓度随紫外照射时间增加明显下降；噬菌体基因组共43 193 bp，包含44个基因，无毒力因子基因和抗生素耐药基因。研究结果表明，噬菌体CHY5-M1M在开发新型抗弧菌药物、预防治疗严重海水动物弧菌病等方面具有良好的应用前景，有望作为未来的抗生素替代产品。

关键词: 副溶血弧菌, 噬菌体, 生物学特性, 全基因组分析