食品科学

• 生物工程 • 上一篇    下一篇

拮抗阪崎肠杆菌乳酸菌的筛选鉴定及抑菌特性

杜静芳,缪璐欢,马欢欢,吕欣然,李 莹,白凤翎*,李 春,励建荣   

  1. 渤海大学食品科学与工程学院,辽宁省食品安全重点实验室,生鲜农产品贮藏加工及安全控制技术国家地方联合工程研究中心,辽宁 锦州 121013
  • 出版日期:2016-06-15 发布日期:2016-06-27

Screening and Identification of Lactic Acid Bacteria Antagonistic against Enterobacter sakazakii

DU Jingfang, MIAO Luhuan, MA Huanhuan, LÜ Xinran, LI Ying, BAI Fengling*, LI Chun, LI Jianrong   

  1. National & Local Joint Engineering Research Center of Storage, Processing and Safety Control Technology for Fresh Agricultural
    and Aquatic Products, Food Safety Key Laboratory of Liaoning Province, College of Food Science and Technology, Bohai University,
    Jinzhou 121013, China
  • Online:2016-06-15 Published:2016-06-27

摘要:

阪崎肠杆菌(Enterobacter sakazakii)是引起新生儿败血症、脑膜炎和坏死性小肠结肠炎的条件致病菌,感染后死亡率高达80%。实验采用双层琼脂扩散法从淡水鱼肠道(19 株)和泡菜(7 株)中筛选出1 株来源于鲤鱼肠道的对Enterobacter sakazakii(106 CFU/mL)具有较强拮抗作用的乳酸菌菌株LY-4。结果表明,抑菌活性物质存在于乳酸菌无细胞上清液(cell free supermatant,CFS)中,抑菌最佳培养时间为28 h。CFS经胃蛋白酶处理可完全丧失抑菌活性,中性蛋白酶和木瓜蛋白酶处理后抑菌活性降至76.44%和54.38%,对α-淀粉酶不敏感;在pH 3.0~4.5范围内保持抗菌活性稳定;经40~80 ℃范围内处理2 h后其拮抗活性基本不变,100 ℃和121 ℃处理2 h后活性降至79.57%和75.71%。初步判定菌株LY-4产生的抑菌物质为非糖基化类细菌素,并筛选出初步纯化抑菌物质的最佳萃取剂为乙酸乙酯,粗提物的最小抑菌浓度为6.0 mg/mL。经生理生化和16S rDNA鉴定为植物乳杆菌(Lactobacillus plantarum)。

关键词: 乳酸菌, 阪崎肠杆菌, 鲤鱼肠道, 筛选与鉴定, 抑菌作用

Abstract:

Enterobacter sakazakii is an opportunistic pathogen that can cause neonatal septicemia and cephalomeningitis and
necrotizing enterocolitis with mortality rate of 80%. Lactic acid bacteria with inhibitory activity against E. sakazakii were
isolated from the intestine of freshwater fish and pickles by double-layer agar diffusion method. The lactic acid bacterial
strain isolated from the intestine of common carp, named as LY-4, showed the highest inhibitory activity, which was mainly
derived from its cell-free culture supernatant (CFS). The CFS showed the strongest antagonistic activity when the strain was
cultured for 28 h. The antagonistic activity was lost entirely after treatment with pepsin, and degraded sharply to 76.44% and
54.38% by treatment with neutral protease and papain, respectively, but it was not sensitive to α-amylase, and was stable
at pH 3.0–4.5. The antagonistic activity remained basically unchanged after thermal treatment at temperatures between 40
and 80 ℃ for 2 h, but declined to 79.57% and 75.71% of its original value after incubation at 100 ℃ and 121 ℃ for 2 h,
respectively. The inhibitory substance was determined as a non-glycosylated bacteriocin, which was found to be the most
extractable with ethyl acetate. The minimum inhibitory concentration (MIC) of this bacteriocin was 6.0 mg/mL. The strain
LY-4 was identified as Lactobacillus plantarum by physiological and biochemical characteristics and 16S rDNA sequence.

Key words: lactic acid bacteria, Enterobacter sakazakii, common carp intestine, screening and identification, antagonism

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