动物乳杆菌的分离鉴定及其抑菌蛋白的特性分析

Screening and Identification of Lactobacillus Animalis Strain and Its Inhibitory Protein Characteristics

摘要:

通过滤纸片法从健康肉猪猪大肠、小肠中分离得到212株抗生物质产生菌, 以杯碟法复筛, 得到1株对溶壁微球菌(Micrococcus lysodeikticus)、金黄色葡萄球菌(Staphylococcus aureus)、枯草芽孢杆菌(Bacillus subtilis)、蜡状芽孢杆菌(Bacillus cereus)等革兰氏阳性菌和大肠杆菌(Escherichia coli)、鼠伤寒沙门氏菌(Salmonella typhimurium)等革兰氏阴性菌以及部分真菌如禾谷镰刀霉(Fusarium graminearum)均有强烈抑制作用的乳酸菌。经形态学、生理生化特征及16S rDNA序列同源性分析等手段鉴定该菌株为动物乳杆菌(Lactobacillus animalis)。排除酸和过氧化氢的干扰后, 该菌株的发酵上清液对指示菌仍有明显抑菌活性; 用蛋白酶处理该菌株的发酵上清液后, 抑菌活性丧失; 发酵液粗提物具有较好的热稳定性(经121°C处理20 min仍有较强抑菌活性)以及较宽的抑菌活性pH值范围(3.5~5.5), 因此初步认为该菌株产生一类具有广谱抑菌活性的类细菌素物质。

关键词:猪肠道, 动物乳杆菌, 类细菌素, 鉴定, 抑菌特性

Abstract:

A bacteriocin-like substance producing strain T12 was screened from 212 lactic acid bacteria isolated from intestine of healthy pigs by paper disk test and was identified as Lactobacillus. The supernatant of T12 exhibited strong inhibitory activity against the gram-positive bacteria, including Micrococcus lysodeikticus, Staphylococcus aureus, Bacillus subtilis and Bacillus cereus. A strong activity was also observed against the gram–negative bacteria and some eumycete, such as Escherichia coli, Salmonella typhimurium and Fusarium graminearum. Through detection of its appearance, physiological and biochemical characteristics and 16S rDNA gene sequence homology analysis, it was identified as Lactobacillus animalis. Eliminated the organic acids and hydrogen peroxide the fermented production exhibited strong inhibitory activity against indicat bacterias still; the inhibitory activity was totally lost after treatment with protease; The bacteriocin-like substance was heat-stable even at the autoclaving temperature (121°C for 20 min) and was active over a wide pH range (3.5~5.5). The protein nature and restricted spectrum of the substance points to its being bacteriocin-like substance.

Key words:Pig intestinal tract, Lactobacillus animalis, Bacteriocin-like substance, Identification, Characterization