China Animal Husbandry and Veterinary Medicine ›› 2023, Vol. 50 ›› Issue (4): 1684-1694.doi: 10.16431/j.cnki.1671-7236.2023.04.039

• Basic Veterinary Medicine • Previous Articles     Next Articles

Isolation,Identification and Biological Characteristics Analysis of a Bacteriophage EP_H11 from Escherichia coli

HUANG Qiaochen1, LIAO Binru1, YU Chunyan1, MO Yupeng1,2, HUANG Weiqian1,3, LI Lei1, HAN Kaiou1, WANG Leping1, LI Xun1, WANG Xiaoye1   

  1. 1. College of Animal Science and Technology, Guangxi University, Nanning 530004, China;
    2. Guilin Liyuan Group, Guilin 541012, China;
    3. Guangdong Rongda Biological Co., Ltd., Qingyuan 515000, China
  • Published:2023-04-06

Abstract: 【Objective】 It was planned to isolate a strain of phages with good bacteriostatic effect and wide adaptability for Escherichia coli,which provided a basis for the development of new Escherichia coli control methods.【Method】 Taking Escherichia coli H11 as the host bacteria,a strain of Escherichia coli phage was isolated and purified from sewage samples by double-layer agar plate method.The morphology of the phage was observed under electron microscope,and the titer,host spectrum,optimal multiplicity of infection(MOI),one-step growth curve,pH stability and temperature sensitivity of the phage were determined.The whole phage genome was analyzed and sequenced,and the inhibitory effect of the phage on H11 in liquid medium was explored.【Result】 A lytic Escherichia coli phage was successfully isolated and named EP_H11(accession number:OP688485).The results of double-layer agar culture method showed that the phage EP_H11 could form plaques with uniform size,transparent bright spots in the middle and halo around,and the phage cleavage titer could reach 109-1010 PFU/mL.Electron microscopy showed that the phage EP_H11 had a regular polyhedral head with a diameter of about 85 nm,a tail with a diameter of about 23 nm and a length of about 109 nm.Using Escherichia coli H11 as host bacteria,the optimal MOI of phage EP_H11 was 0.1.The one-step growth curve showed that the incubation period was 40 min and the outbreak period was 80 min.The amount of cleavage was about 200 PFU/cell.The phage could survive for at least 2 h in the environment of pH 4.0-10.0,and could remain stable for 90 min at 10-60 ℃.Combined with transmission electron microscopy and whole genome analysis,the phage EP_H11 belonged to the Caudovirales,Siphoviridae family,Myoviridae genus.The total length of the phage genome was 111 373 bp and the GC content was 44.49%.According to the results of phage linear comparison,Escherichia coli phage EP_H11 and phage Escherichia phage vB_EcoM_IME392 had the highest homology.Phylogenetic tree analysis showed that the phage EP_H11 and Escherichia phage vB_EcoM_IME392 (Myoviridae genus) were located in the same branch,belonging to the order of Caudovirales and the Siphoviridae family.BLAST sequence analysis showed that there were 153 CDSs in genome of EP_H11,28 of which were predicted to encode functional proteins.In vitro bacteriostasis results showed that within 8 h,EP_H11 showed good bacteriostatic effect when MOI was 10. 【Conclusion】 In this study,a bacteriophage with strong lytic ability,wide adaptability and good bacteriostatic effect in vitro was isolated.The bacteriophage had the potential to be further developed as an antimicrobial agent for Escherichia coli.

Key words: Escherichia coli; phage; whole genome analysis

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