食品科学 ›› 2019, Vol. 40 ›› Issue (8): 79-86.doi: 10.7506/spkx1002-6630-20180518-258

• 生物工程 • 上一篇    下一篇

1株T1样噬菌体vB_EcoS_IME18的分离鉴定及其受体分析

李 萍,林 洪,童贻刚,王静雪   

  1. 1.中国海洋大学食品科学与工程学院,山东 青岛 266003;2.军事科学院军事医学研究所,微生物流行病研究所病原微生物安全国家重点实验室,北京 100071
  • 出版日期:2019-04-25 发布日期:2019-05-05
  • 基金资助:
    “十三五”国家重点研发计划重点专项(2017YFC1600703);国家重点研发计划项目(AWS16J020);山东省重点研发计划项目(2017GNC1318)

Isolation and Identification of a T1-Like Bacteriophage vB_EcoS_IME18 and Analysis of Its Receptor

LI Ping, LIN Hong, TONG Yigang, WANG Jingxue   

  1. 1. College of Food Science and Engineering, Ocean University of China, Qingdao 266003, China; 2. State Key Laboratory of Pathogenic Microbiology Safety, Institute of Microbiological Epidemiology, Institute of Military Medicine, Academy of Military Sciences, Beijing 100071, China
  • Online:2019-04-25 Published:2019-05-05

摘要: 以大肠杆菌BL21为指示菌分离得到1 株噬菌体vB_EcoS_IME18(IME18),通过电镜观察噬菌体形态,并测定其最佳感染复数、一步生长曲线、裂解谱和体外杀菌效果;使用Illumina MiSeq测序仪完成噬菌体全基因组测序,利用比较基因组学分析噬菌体基因组特征;通过筛选噬菌体抗性株分析其耐受原因,利用受体基因沉默突变实验、噬菌体斑点实验和噬菌体吸附实验验证噬菌体受体。结果表明,噬菌体IME18具有二十面体头部和非收缩性尾部,属于长尾噬菌体科,Tunavirinae亚科,T1类噬菌体;噬菌体IME18的最佳感染复数为0.01,潜伏期为5 min,裂解期50 min,爆发量高达223 PFU/cell;在体外,噬菌体IME18能在90 min内有效杀死处于对数期的BL21;噬菌体IME18基因组全长50 354 bp,G+C含量为45.6%;蛋白FhuA、TonB在噬菌体IME18的吸附和侵染过程中发挥重要作用。

关键词: 大肠杆菌, 噬菌体, 鉴定, 噬菌体受体

Abstract: A strain of phage, designated vB_EcoS_IME18 (IME18), was isolated using Escherichia coli BL21 as the indicator bacterium. The morphology of the phage was observed by electron microscopy and the multiplicity of infection, one-step growth curve, lysis spectrum and in vitro bactericidal effect were determined; the genomic sequencing was performed by Illumina MiSeq sequencer and genomic features were analyzed by comparative genomics; the cause of phage resistance was analyzed by screening for phage resistant strains and confirmed by receptor gene silencing mutation, phage spotting, and phage adsorption assay. The results showed that phage IME18 had an icosahedral head and a non-contractile tail, belonging to the T1-like in the Tunavirinae subfamily in the family of Siphoviridae. The multiplicity of infection of phage IME18 was 0.01, latent period 5 min, burst period 50 min and burst size 223 PFU/cell; in vitro, phage IME18 could effectively kill BL21 in the log phase in 90 min. The total length of the genome was 50 354 bp with a G + C content of 45.6%. In addition, the proteins FhuA and TonB were proved to play a key role in the adsorption and infestation of phage IME18.

Key words: Escherichia coli, bacteriophage, identification, phage receptor

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