海洋湿地环境中拮抗水产品腐败菌的乳酸菌筛选与鉴定

从渤海锦州海岸湿地植物根系中筛选对腐败希瓦氏菌、铜绿假单胞菌和副溶血弧菌具有较强抑制作用的乳酸菌,为研发一种海产品防腐保鲜生物制剂奠定基础。应用牛津杯打孔法从碱蓬草、芦苇根部筛选拮抗性乳酸菌菌株,应用生长曲线分析菌株拮抗作用变化,通过pH、蛋白酶、温度、NaCl浓度等因素,分析乳酸菌无细胞上清液(CFS)的拮抗性能。从碱蓬草根部获得1株拮抗能力较强的乳酸菌株JP-12,初步判定其抑菌活性物质存在于CFS中,生长曲线结果显示其抑菌活性在培养28 h时达到峰值。CFS经不同蛋白酶处理后,抑菌活性均有所下降。对胃蛋白酶最为敏感,活性丧失率为24.21%;CFS经40~121 ℃等不同温度处理30 min,其抑菌活性基本不变;在pH3.0~4.5范围内抑菌活性保持稳定;3.0%~3.5% NaCl条件下其抑菌活性较强,抑菌直径达到18.46、18.15 mm。经生理生化实验和16S rRNA菌株JP-12被鉴定为植物乳杆菌(Lactobacillus plantarum)。本文为挖掘拮抗性乳酸菌的资源及在水产品防腐保鲜中的应用提供一条新的途径。

Screening and Identification of Lactic Acid Bacteria with Antimicrobial Activity to Aquatic Products Spoilage Bacteria from Marine Wetland

Lactic acid bacteria(LAB)strains with strong antagonistic activity to Shewanella putrefaciens,Pseudomonas aeruginosa,Vibrio parahaemolyticus were isolated from roots of plants in Jinzhou Bohai coast wetland to develop a biological preparation for the preservation of seafood. LAB with antibacterial activity was isolated from Suaeda glauca roots using the Oxford cup method. The formation process of antibacterial substance was analyzed by growth curve. The antimicrobial activity of cell-free supernatant(CFS)produced by LAB was analyzed from the stability of pH,protease,temperature and NaCl concentration. Strain JP-12 with strong inhibitory activity was obtained from roots of Suaeda glauca,and its antagonistic activity mainly existed in CFS. The growth curve analysis showed that the maximum activity of strain JP-12 was reached after 28 h of incubation. Antibacterial activity of CFS decreased treated by different proteases. And the most sensitive to pepsin,the loss rate of activity was 24.21%. The antagonistic activity of CFS was remained after treatment with 40~121 ℃ for 30 min and kept stable within pH3.0~4.5.CFS possessed strong antibacterial activity at the concentration of NaCl in 3.0% and 3.5%,and its diameter of inhibition zone reached 18.46,18.15 mm,respectively. The strain JP-12 was identified as Lactobacillus plantarum by physiological and biochemical characteristics and 16S rRNA gene sequencing. This study provided a new approach to excavate resource of antagonistic LAB and its application in aquatic products preservation.