溶藻弧菌烈性噬菌体分离鉴定及其抑菌作用研究

Isolation and identification of Vibrio alginolyticus bacteriophage and anti-bacteria activity

溶藻弧菌(Vibrio alginolyticus, VA)是水产品中普遍存在的人兽共患病原菌。为研制水产品中VA的生物抑菌剂,以溶藻弧菌VAf 211为宿主菌,应用双层平板法从扬州农贸市场的宰鱼污水中分离纯化获得裂解性噬菌体,研究其微观形态、最佳感染复数、一步生长曲线、裂解谱等生物学特征,并在鱼汁模型中测定噬菌体对VA的抑菌效应以及对VA生物被膜形成的影响。结果分离到1株溶藻弧菌烈性噬菌体,将其命名为VAPf211,透射电镜观察显示其形态归属于肌尾噬菌体科,最佳感染复数为0.01,潜伏期约为10 min,裂解期为50 min,平均裂解量为34.80 PFU·细胞^-1。温度和pH稳定性试验结果显示VAPf211在温度40~60℃、pH值4.0~9.0范围内活性稳定。抑菌试验结果显示噬菌体VAPf211能显著抑制宿主菌在鱼汁中的增长,并能抑制VA生物被膜的形成。这一研究提示VAPf211具有良好的应用前景,为水产食品中溶藻弧菌的减除提供了新型生物减菌剂。

Vibrio alginolyticus(VA), a ubiquitous pathogenic bacteria in marine and marine organism, can infect human beings and aquatic products. In order to isolate and apply the phage to control V.alginolyticus VAf211 was used as the potential host strain for phage isolation from local aquatic product trading markets in Yangzhou, Jiangsu, China by double layer plate method. Morphological characters was observed by transmission electro-microscopy. Optimal multiplicity of infection(MOI), one-step growth curve, lysis spectrum detection, thermal stability, sensitivity to pH, Lytic activity of phage VAPf211 in fish matrices were detected. Microtiter-plate assay was used to test biofilm prevention against the host strain. The results showed that a broad-spectrum lytic phage VAPf211 was isolated. It’s belongs to the family Myoviridae. The optimal MOI was 0.01. One-step growth curve showed that the incubation period was about 10 min, the lysis period was 50 min, and the average amount of lysis was 34.80 PFU·cell^-1. Phage VAPf211 was stable within the temperature of 40-60 ℃ and within the pH of 4.0-9.0. The bacteriostatic test results showed that phage VAPf211 can significantly inhibit the growth of host bacteria in fish juice and the formation of VA biofilm. VAPf211 has a good application prospect and provides a new biological microbicide for the reduction of V.alginolyticus in aquatic foods.