Moon Kira, Ryu Sangdon, Song Seung Hui, Chun Se Won, Lee Nakyeong, Lee Aslan Hwanhwi
Division of Environmental Materials, Honam National Institute of Biological Resources, Mokpo 58762, Republic of Korea.
J Microbiol. 2025 May;63(5):e2502009. doi: 10.71150/jm.2502009. Epub 2025 May 27.
The widespread use of antibiotics in aquaculture has led to the emergence of multidrug-resistant pathogens and environmental concerns, highlighting the need for sustainable, eco-friendly alternatives. In this study, we isolated and characterized three novel bacteriophages from aquaculture effluents in Korean shrimp farms that target the key Vibrio pathogens, Vibrio harveyi, and Vibrio parahaemolyticus. Bacteriophages were isolated through environmental enrichment and serial purification using double-layer agar assays. Transmission electron microscopy revealed that the phages infecting V. harveyi, designated as vB_VhaS-MS01 and vB_VhaS-MS03, exhibited typical Siphoviridae morphology with long contractile tails and icosahedral heads, whereas the phage isolated from V. parahaemolyticus (vB_VpaP-MS02) displayed Podoviridae characteristics with an icosahedral head and short tail. Whole-genome sequencing produced complete, circularized genomes of 81,710 bp for vB_VhaS-MS01, 81,874 bp for vB_VhaS-MS03, and 76,865 bp for vB_VpaP-MS02, each showing a modular genome organization typical of Caudoviricetes. Genomic and phylogenetic analyses based on the terminase large subunit gene revealed that although vB_VhaS-MS01 and vB_VhaS-MS03 were closely related, vB_VpaP-MS02 exhibited a distinct genomic architecture that reflects its unique morphology and host specificity. Collectively, these comparative analyses demonstrated that all three phages possess genetic sequences markedly different from those of previously reported bacteriophages, thereby establishing their novelty. One-step growth and multiplicity of infection (MOI) experiments demonstrated significant differences in replication kinetics, such as burst size and lytic efficiency, among the phages, with vB_VhaS-MS03 maintaining the most effective bacterial control, even at an MOI of 0.01. Additionally, host range assays showed that vB_VhaS-MS03 possessed a broader spectrum of activity, supporting its potential use as a stand-alone agent or key component of phage cocktails. These findings highlight the potential of region-specific phage therapy as a targeted and sustainable alternative to antibiotics for controlling Vibrio infections in aquaculture.
水产养殖中抗生素的广泛使用导致了多重耐药病原体的出现以及环境问题，这凸显了对可持续、生态友好型替代方案的需求。在本研究中，我们从韩国虾养殖场的养殖废水里分离并鉴定了三种新型噬菌体，它们靶向关键弧菌病原体哈维氏弧菌和副溶血性弧菌。通过环境富集和双层琼脂试验进行连续纯化来分离噬菌体。透射电子显微镜显示，感染哈维氏弧菌的噬菌体，命名为vB_VhaS-MS01和vB_VhaS-MS03，呈现典型的长尾噬菌体科形态，具有长收缩尾和二十面体头部，而从副溶血性弧菌分离出的噬菌体（vB_VpaP-MS02）表现出短尾噬菌体科特征，有二十面体头部和短尾。全基因组测序产生了vB_VhaS-MS01的81,710 bp完整环状基因组、vB_VhaS-MS03的81,874 bp完整环状基因组以及vB_VpaP-MS02的76,865 bp完整环状基因组，每个基因组都显示出尾病毒目典型的模块化基因组结构。基于末端酶大亚基基因的基因组和系统发育分析表明，尽管vB_VhaS-MS01和vB_VhaS-MS03密切相关，但vB_VpaP-MS02呈现出独特的基因组结构，反映了其独特的形态和宿主特异性。总体而言，这些比较分析表明，所有这三种噬菌体都拥有与先前报道的噬菌体明显不同的基因序列，从而确定了它们的新颖性。一步生长和感染复数（MOI）实验表明，噬菌体之间在复制动力学方面存在显著差异，如爆发大小和裂解效率，其中vB_VhaS-MS03即使在MOI为0.01时也能保持最有效的细菌控制。此外，宿主范围试验表明，vB_VhaS-MS03具有更广泛的活性谱，支持其作为单独制剂或噬菌体鸡尾酒关键成分的潜在用途。这些发现突出了区域特异性噬菌体疗法作为控制水产养殖中弧菌感染的靶向性和可持续性抗生素替代方案的潜力。
