一株具有抑菌活性的芽孢杆菌的分离鉴定及生物特性研究

Isolating, identification and biological characteristics of a Brevibacillus laterosporus strain with antibacterial activity

试验旨在分离出具有抑菌作用和抗逆性优良的饲用芽孢杆菌。从健康仔猪粪便和周围土壤中分离到18株芽孢杆菌,用大肠埃希氏菌和金黄色葡萄球菌作为指示菌,通过抑菌试验,筛选出一株对2种病原菌抑菌作用最好的菌株。利用生化试验和16S rDNA基因序列分析的方法鉴定菌株,研究其对高温、胆盐、人工胃肠液的耐受能力及其致病菌的抑菌能力,并对该菌株发酵液中抑菌物质进行研究。结果表明,该菌株经鉴定为侧孢短芽孢杆菌(Brevibacillus laterosporus),命名为LP-9。该菌株对高温和胆盐有较好的耐受力,85℃处理10 min后,芽孢存活率达到96%,能耐受0.3%的胆盐;对多种抗生素具有良好的耐受性;经过人工胃液和肠液处理后,活菌数达95%以上。LP-9上清液经高温和蛋白酶K处理后,对两种指示菌的抑菌作用明显下降,说明细菌上清液中含蛋白类的抑菌物质。结果表明,LP-9对高温、胆盐和人工胃液和肠液有极好的耐受性,在生产和加工的过程中能保持较好的活性,并产生蛋白类抑菌活性物质,在饲料应用方面有极好的应用前景。

The study was to isolate to forage Bacillus with excellent bacteriostatic effect and stress resistance.Eighteen strains of Bacillus were isolated from feces piglets and the surrounding soil.E.coli K88 and Staphylococcus aureus were used as the indicator bacteria,and a strain was found to exhibite the best antibacterial effect against the two pathogenic bacteria by bacteriostatic experiment.The strains were identified by biochemical identification and 16 S rDNA gene sequence analysis,and the tolerance to high temperature,bile salts,artificial gastric juice and artificial intestinal juice and the antagonism to pathogenic bacteria were explored.Next,the stability of bacteriostat in fermentation broth were further studied.The results showed that a strain named LP-9 was isolated and identified as Brevibacillus laterosporus,which had good tolerance to high temperature,various antibiotics and 0.3%of bile salts.After treatment at 85℃for 10 min,the survival rate of spores reached 96%.After artificial gastric juice and intestinal juice treatment,the number of viable bacteria reached more than 95%.The supernatant fluid of LP-9 with high temperature and proteinase K treatment,the bacteriostasis against two indicator bacteria were obviously decreased.The data indicated that the supernatant fluid contains antibacterial components with protein.The above results suggested that B.laterosporus PL-9 has excellent resistence to high temperature,bile salts and artificial gastric juice and artificial intestinal juice.In the process of production and processing,PL-9 can maintain great activity and produce bacteriostatic substances,which has excellent application prospects in feed applications.