广谱抑菌乳酸菌的筛选及其细菌素相关基因分析

Screening for a Strain of Lactic Acid Bacteria with Broad-Spectrum Antimicrobial Activity and Analysis of Its Bacteriocin-Related Genes

采用琼脂扩散法,从分离自新疆传统酸奶的8株乳酸菌中筛选出1株抑菌效果较好并具有广谱抑菌性的菌株B6,通过革兰氏染色、生理生化及16S rDNA序列比对,鉴定该菌为植物乳杆菌。植物乳杆菌B6发酵液经排除酸、过氧化氢实验后,仍有抑菌效果,且对蛋白酶敏感,判定该菌产细菌素。通过设计10对编码合成细菌素相关基因的特异性引物,以菌株B6的DNA为模板,进行聚合酶链式反应快速鉴定。结果表明,菌株B6含有编码IIb类细菌素结构基因plnE/F/J/K,plnJ与植物乳杆菌C11相比,序列仅在前导肽区出现一处氨基酸突变,相似度为99%,plnE/F/K序列与植物乳杆菌C11的这3个基因序列完全相同。因此鉴定植物乳杆菌B6产生的是IIb类细菌素。

Using agar diffusion method,one strain,named B6,with potent and broad-spectrum antimicrobial activity was selected from the 8 strains of lactic acid bacteria(LAB)isolated from traditional yogurt in Xinjiang,China.It was identified as Lactobacillus plantarum by morphological,physiological and biochemical characteristics and 16S rDNA sequence alignment.The cell-free fermented broth of B6 retained its antibacterial activity after sequential removal of acids and hydrogen peroxide,which indicated that L.plantarum B6 could produce bacteriocin.Polymerase chain reaction(PCR)analysis was carried out to identify the bacteriocin genes of B6 by using 10 pairs of primers designed to be specific for the genes related to the biosynthesis of bacteriocin with the total DNA of strain B6 as the template.The results showed that strain B6 had the structural genes plnE/F/J/K for class IIb bacteriocin.The plnJ gene shared 99%similarity with that of L.plantarum C11 with only one amino acid mutation in the leader peptide region,and the plnE/F/K genes shared 100%similarity with those of L.plantarum C11,which indicated that L.plantarum B6 produced class IIb bacteriocin.