传统生理生化鉴定技术结合PCR法分析复合保鲜剂对冷藏带鱼贮藏期间菌相变化的影响

Effect of complex biological fresh-keeping agents for the main bacteria composition on cutlassfish Trichiurus haumela under the cold storage by PCR and physiology-biochemistry technology

将微生物传统生理生化鉴定技术与PCR法相结合,研究了复合保鲜剂对冷藏带鱼贮藏期间菌相变化的影响。通过对冷藏对照组与保鲜剂处理组样品贮藏期间的主要微生物进行分离纯化、16SrDNA序列分析与生理生化鉴定,并作系统发育树分析,最终鉴定出13株具有典型特征的纯菌株。对照组样品在货架期终点时,其主要微生物的种类与所占比例依次为:腐败希瓦氏菌(34.7%)、荧光假单胞菌(14.5%)、松鼠葡萄球菌(10.2%)、嗜水气单胞菌(8.2%)、弧菌(6.1%)、恶臭假单胞菌(6.1%)、绿色气球菌(4.1%)、金黄色葡萄球菌(4.1%)、蜡样芽孢杆菌(4.0%)、铜绿假单胞菌(2.0%)、嗜冷杆菌(2.0%)、成团肠杆菌(2.0%)、约氏不动杆菌(2.0%)。其中,腐败希瓦氏菌为特定优势腐败微生物。假单胞菌属在带鱼冷藏过程中所占比例大小依次为荧光假单胞菌>恶臭假单胞菌>铜绿假单胞菌。经复合保鲜剂处理后,能够使冷藏带鱼的二级鲜度货架期延长9d,并使其贮藏期间的细菌菌相组成比例发生变化,细菌种类减少到9种。主要优势菌的比例明显减少,表明复合保鲜剂对带鱼体内腐败菌具有不同程度的抑制作用。

Microbial physiology-biochemistry technology in traditional method combined with PCR technology were used to study the effect of Complex Biological Fresh- keeping Agents （CBFA） on Trichiurus haumela.After the separation and purification of pre-dominate spoilage bacteria on Trichiurus haumela used by CBFA under the cold storage, ]6S rDNA sequence analysis, morphological and biochemical tests were used to identify the composition of spoilage microfiora, and phylogenetic tree were analyzed.The result showed that there were mainly 13 typical bacterial strains of the controlled, and their proportions at the end of shelf life in control groups were： Shewanella putrefaciens 34.7%, Pseudomonas fluorescens ]4.5%, Staphylococcus sciuri ]0.2%, Aeromonas hydrophila 8.2%, Vibrio rumoiensis 6. ] %, Pseudomonas putida d.1% , Aerococcus viridans 4. ] %, Staphyloccocus aureus 4.1%, Bacillus cereus 4.0%, Pseudomonas aeruginosa 2.0%, Psychrobacter sp. 2.0% Enterobacter agglomerans 2.0% and Acinetobacterjohnsonii2.0% respectively.Among them, Shewanella putrefaciens（34.7%）was the Specific Spoilage Organism（SSO）, the proportion of Pseudomonas sp. in descending order was Pseudomonas fluorescens 〉 Pseudomonas putida 〉 Pseudomonas aeruginosa. It was found that the dominant spoilage bacteria of Trichiurus haumela was extended 9d by CBFA, the spoilage microflora was also changed, and the species of bacteria decreased to 9 strains.CBFA showed different inhibitory activity on different spoilage bacteria.