【摘要】 我国草鱼(Ctenopharyngodon idella)养殖过程中面临多种细菌病的威胁，传统的细菌病防控依赖抗生素、消毒剂等化学药物，这些药物的不合理使用会造成细菌性病原的耐药性增强并威胁水产品质量安全。本研究旨在筛选具有防控鱼类细菌病潜力的水生动物源益生菌。采集健康草鱼肠道以及体表黏液，匀浆后利用MRS培养基分离纯化乳酸菌。通过测定分离菌株的拮抗活性、溶血活性，从肠道中筛选出候选菌株GCC10，并分析菌株的形态特征和分类学地位，测定其胞外酶活性、药物敏感性、胆汁耐受能力、肠道定殖能力和生物安全性。全基因组测序分析GCC10菌株的毒力基因和耐药基因存在情况以及潜在的益生功能。结果显示，分离菌株GCC10为革兰氏阳性乳酸乳球菌(Lactococcus lactis)，无溶血活性，具有胞外蛋白酶活性、耐受胆汁能力和肠道定殖能力。致病性试验结果显示GCC10菌株对草鱼表现出良好的安全性。药敏试验结果显示GCC10菌株对恩诺沙星、氟苯尼考、多西环素、磺胺间甲氧嘧啶等多种抗菌药物敏感，对庆大霉素表现为耐药。拮抗抑菌结果显示该菌对草鱼常见致病菌如嗜冷黄杆菌(Flavobacterium psychrophilum)、美人鱼发光杆菌(Photobacterium damselae)、嗜水气单胞菌(Aeromonas hydrophila)、温和气单胞菌(Aeromonas sobria)和维氏气单胞菌(Aeromonas veronii)等有较好的抑菌作用。全基因组测序分析表明GCC10菌株未携带主要毒力基因，但是存在与应对冷热应激、耐胆酸盐、耐酸、免疫调节、蛋白水解和黏附因子等基因以及产乳球菌素相关的基因簇。本研究筛选出1株生物安全性良好、拮抗抑菌活性强且能够在肠道定殖的乳酸乳球菌GCC10，为防控鱼类细菌病提供了一种优良的种质资源，在水产养殖中作为减抗或替抗制剂具有巨大的开发应用潜力。更多还原

【Abstract】 Grass carp(Ctenopharyngodon idella) aquaculture in China faces serious challenges from various bacterial diseases. Conventional prevention and control strategies mainly rely on antibiotics and chemical disinfectants, the excessive use of which has led to increasing bacterial resistance and potential threats to the quality and safety of aquatic products. This study aimed to screen aquatic animal-derived probiotics with potential for the prevention and control of fish bacterial diseases. Intestinal and mucus samples were collected from healthy grass carp, homogenized, and used for the isolation and purification of lactic acid bacteria using MRS medium. Strain GCC10 was selected based on its antagonistic activity and hemolytic activity from intestinal. The morphological characteristics and taxonomic classification of strain GCC10 were determined,and its extracellular enzyme activity, antibiotic susceptibility, bile salt tolerance, intestinal colonization capasity, and biosafety were evaluated. Whole-genome sequencing was performed to evaluate the genomic safety and potential probiotic functions of strain GCC10. Strain GCC10 was identified as a Gram-positive Lactococcus lactis with no hemolytic activity. It exhibited extracellular protease activity, strong bile salt tolerance, and the ability to colonize the intestinal tract. The antibiotic susceptibility test showed that strain GCC10 was sensitive to several antimicrobial agents, including enrofloxacin, florfenicol, doxycycline,sulfamonomethoxine, but resistant to gentamicin. Antagonistic assays indicated that GCC10 displayed strong inhibitory activity against several common fish pathogens, such as Flavobacterium psychrophilum,Photobacterium damselae, Aeromonas hydrophila, Aeromonas sobria, and Aeromonas veronii. Whole-genome analysis revealed that GCC10 lacked major virulence genes. However, the genome contained genes related to heat and cold stress responses, bile salt and acid tolerance, immune modulation, proteolysis, and adhesion, as well as gene clusters involved in the production of lactococcal bacteriocins. This study screened out a Lactococcus lactis strain GCC10, which possessed good biosafety, strong antagonistic antibacterial activity, and effective intestinal colonization capability. This study provides a promising biological resource for controlling bacterial diseases in fish and has great potential for development and application as an antibiotic-reducing or antibiotic-replacing agent in aquaculture.更多还原