抗奶牛乳房炎主要病原菌细菌素的筛选及其生物学特性分析

Screening and Biological Characteristics Analysis of Bacteriocins against Main Pathogenic Bacteria of Bovine Mastitis

为了筛选出对奶牛乳房炎主要病原菌具有抑菌活性的细菌素,并对其生物学特性进行分析。本试验采用涂布和平板划线方法从吉林市传统辣白菜中分离纯化乳酸菌,并以金黄色葡萄球菌为指示菌通过琼脂扩散法进行抑菌试验,同时结合排酸和排过氧化氢试验初步筛选出拮抗菌株。通过菌株形态学特征和16S rRNA序列分析鉴定拮抗菌株。测定菌株生长和抑菌动力学曲线确定最佳发酵时间。采用蛋白酶敏感性、热稳定性和酸碱稳定性试验测定抗菌活性物质的理化特性,并测定其抑菌谱。结果显示,本试验分离出1株可产细菌素的拮抗菌株MX1,经鉴定其为植物乳杆菌,最佳发酵时间为22 h;MX1菌株分泌的细菌素具有良好的热稳定性和酸稳定性,经高温处理2 h或处于pH为3.0和4.0等酸性较强的环境下仍能保持较强的抗菌活性;MX1菌株分泌的细菌素对奶牛乳房炎主要致病菌金黄色葡萄球菌、大肠杆菌和肺炎克雷伯菌具有良好的抗菌活性。结果表明,本试验筛选获得的细菌素有望作为替抗产品用于细菌性奶牛乳房炎的防治。

This study aimed to screen bacteriocins with antibacterial activity against the main pathogenic bacteria of bovine mastitis and analyze their biological characteristics.Lactic acid bacteria were isolated and purified from traditional Korean kimchi from Jilin City using streak plate and spread plate methods.The inhibitory activity was tested using the agar diffusion method with Staphylococcus aureus as the indicator bacterium,along with acid and hydrogen peroxide exclusion tests to preliminarily screen antagonistic strains.The antagonistic strains were identified by morphological characteristics and 16S rRNA sequence analysis.The growth and antibacterial kinetics curves of the strains were determined to establish the optimal fermentation time.The physicochemical properties of the antibacterial substances were assessed by testing their sensitivity to protease,thermal stability,and pH stability,and their antibacterial spectrum was determined.The results showed that one strain,designated MX1,capable of producing bacteriocin was isolated and identified as Lactobacillus plantarum.The optimal fermentation time was 22 hours.The bacteriocin secreted by strain MX1 exhibited good thermal and acid stability,retaining strong antibacterial activity after 2 hours of high-temperature treatment or in highly acidic environments with pH of 3.0 and 4.0.The bacteriocin produced by strain MX1 demonstrated effective antibacterial activity against the primary pathogenic bacteria of bovine mastitis,including Staphylococcus aureus,Escherichia coli,and Klebsiella pneumoniae.These results suggest that the bacteriocin obtained in this study has potential as an alternative antibacterial product for the prevention and treatment of bacterial bovine mastitis.