冷藏石斑鱼特定腐败菌菌相高通量测序分析及蛋白降解菌筛选鉴定  

Screening and Identification of Specific Spoilage Bacteria in Chilled Grouper Based on High-throughput Sequencing and Protein Degradation Ability

作　　者：黎铸毅 刘美娇 陈秋翰 杨学博 王敏杰 钟赛意 洪鹏志 朱春华 刘寿春[1,2] LI Zhuyi;LIU Meijiao;CHEN Qiuhan;YANG Xuebo;WANG Minjie;ZHONG Saiyi;HONG Pengzhi;ZHU Chunhua;LIU Shouchun(College of Food Science and Technology,Guangdong Ocean University/Guangdong Provincial Key Laboratory of Aquatic Product Processing and Safety/Guangdong Provincial Engineering Technology Research Center of Seafood/Guangdong Provincial Engineering Technology Research Center of Prefabricated Seafood Processing and Quality Control,Zhanjiang 524088,China;Southern Marine Science and Engineering Guangdong Laboratory(Zhanjiang),Zhanjiang 524004,China;College of Fisheries,Guangdong Ocean University,Zhanjiang 524088,China)

机构地区：[1]广东海洋大学食品科技学院/广东省水产品加工与安全重点实验室/广东省海洋食品工程技术研究中心/广东省水产预制食品加工与品质控制工程技术研究中心,广东湛江524088 [2]南方海洋科学与工程广东省实验室(湛江),广东湛江524004 [3]广东海洋大学水产学院,广东湛江524088

出　　处：《广东海洋大学学报》2024年第4期105-112,共8页Journal of Guangdong Ocean University

基　　金：广东海洋大学科研启动项目(060302042311);广东省高等教育创新团队项目(2021KCXTD021);茂名市罗非鱼优势特色产业集群项目(22282109-1);广东省现代农业产业技术体系创新团队建设项目(2023KJ150)。

摘　　要：【目的】研究珍珠龙胆石斑鱼(Epinephelus fuscoguttatus♀×Epinephelus lanceolatus♂)冷藏过程优势腐败菌的菌相变化,筛选鉴定具有强蛋白分解能力的特定腐败菌。【方法】通过质量指数法(QIM)评价、菌落计数、挥发性盐基氮(TVB-N)含量等指标判断鱼肉腐败进程;对新鲜(冷藏0 d)、次新鲜(冷藏4 d)和腐败(冷藏8 d)样品进行细菌16S V3-V4区域高通量测序,通过物种组成、Alpha和Beta多样性分析、物种韦恩图分析其菌群的结构、多样性和差异;通过传统培养法分离腐败样品菌株,酪蛋白琼脂筛选蛋白分解菌,对筛选出的菌进行革兰氏染色和16S rDNA测序鉴定。【结果】冷藏石斑鱼质量指数法的感官拒绝点为12 d,菌落总数(TVC)超标临界期为4 d,TVB-N超标临界期约为10 d;随冷藏时间延长,鱼肉表面菌群多样性下降,新鲜鱼肉、冷藏4 d、冷藏8 d鱼肉菌群结构在属水平上具有明显差异;新鲜鱼肉中菌群主要是不动杆菌属(Acinetobacter)、嗜冷杆菌属(Psychrobacter)、金黄杆菌属(Chryseobacterium),相对丰度分别为20.44%、15.93%、9.24%;冷藏4 d和8 d鱼肉菌群主要物种是希瓦氏菌属(Shewanella)、气单胞菌属(Aeromonas)、嗜冷菌属(Psychrobacter)、不动杆菌属(Acinetobacter),其相对丰度分别为32.27%、34.89%,30.28%、16.02%,12.90%、16.65%以及13.99%、5.67%;冷藏8 d时鱼肉中具有良好蛋白分解能力的菌株为气单胞菌属和希瓦氏菌属,其组成比例接近高通量测序结果。【结论】气单胞菌属和希瓦氏菌属是珍珠龙胆石斑鱼冷藏腐败过程的优势菌并具有较强的蛋白降解能力,本研究可为后续探究冷藏石斑鱼优势腐败菌的致腐能力和开发靶向抑菌保鲜技术提供基础。【Objective】 To study the microbial phase changes of dominant spoilage bacteria in pearl gentian grouper(Epinephelus fuscoguttatus♀×Epinephelus lanceolatus♂) during chilled storage,and to screen and identify specific spoilage bacteria with strong protein decomposition ability.【Method】The spoilage process of grouper meat was judged by quality index method(QIM),colony count and TVB-N content.Fresh(0 d chilled),sub-fresh(4 d chilled) and spoilaged(8 d chilled) samples were subjected to high-throughput sequencing of bacterial 16S V3-V4 region,and the structure,diversity and differences of the flora were analyzed by species composition,Alpha and Beta diversity analysis,and species Venn diagram.The putrefied sample strains were isolated by traditional culture method,and the protein-decomposing bacteria were screened by casein AGAR and identified by Gram staining and 16S rDNA sequencing.【Result】 The sensory rejection point of chilled grouper QIM came on 12th,the total number of colonies(TVC) exceeded the limit critical period on the 4th day,and the TVB-N exceeded the limit critical period on about 10th day.With the extension of refrigeration time,the diversity of surface microbiota decreased.There were significant differences in the composition of microbiota among fresh grouper,4 d and 8 d chilledstorage grouper at the genus level.The main flora of bacteria in fresh grouper were Acinetobacter,Psychrobacter,and Chrysobacter(relative abundance were 20.44%,15.93% and 9.24% respectively).Shewanella,Aeromonas,Psychrobacter and Acinetobacter,with relative abundance of 32.27% with 34.89%,30.28% with 16.02%,12.90% with 16.65% and 13.99% with 5.67% were the main bacteria in chilled fish meat stored for 4 days and 8 days,respectively.Aeromonas and Shewella had good protein-degrading ability in fish chilled stored for 8 days,and their composition ratio was close to the results of high-throughput sequencing.【Conclusion】 Aeromonas and Shewanella were the dominant bacteria in the chilled grouper,with strong protei

关 键 词：石斑鱼 冷藏 腐败 高通量测序 蛋白降解菌 

分 类 号：S983[农业科学—捕捞与储运]