Biological Characteristics and Comparative Genomics Analysis of a Phage of Multi-drug Resistant Escherichia coli from Brown Bear
[Objective]The aim of this study was to explore the biological and genomic characteristics of a lytic phage of multi-drug resistant Escherichia coli,so as to provide reference for the scientific prevention and control of multi-drug resistant strains.[Method]The phage was isolated and purified from farm sewage by double-plate method with Escherichia coli M719-6WT as host bacteria from Brown bear.The morphology of phage was observed by transmission electron microscopy.The host spectrum of phage was determined by plaque method and its biological characteristics were investigated.Phage bioinformatics analysis was conducted based on phage whole genome sequencing data,and bacteriostatic effects of phage were evaluated by in vitro and in vivo bacteriostatic tests.[Result]Through double-plate method,a phage specific to Escherichia coli was isolated and purified,named pEC-M719-6WT.2,with clear and transparent plaques.Transmission electron microscopy and whole-genome analysis revealed that phage pEC-M719-6WT.2 belonged to the subfamily Tevenvirinae of the Myoviridae family.Host range determination showed pEC-M719-6WT.2 could lyse 11 strains of Escherichia coli.The optimum multiplicity of infection(MOI)was 0.1,the optimum temperature was 25 ℃,the optimum pH was 7.0,the incubation period was less than 10 min,the lysis period was 70 min,and the cleavage volume was about 190 PFU/cell.Genomic analysis showed that phage pEC-M719-6WT.2 had a genome size of 170.441 kb and no known genes related to drug resistance,lysogen or virulence were found.The results of receptor binding tests showed that phage pEC-M719-6WT.2 could bind to lipopolysaccharide receptors on the surface of host bacteria.In vitro bacteriostatic results showed,when MOI was 100,phage pEC-M719-6WT.2 combined with 8 μg/mL tetracycline could prolong the bacteriostasis time to 24 h.The results of in vivo protection test showed that after 2.8×105 CFU/mL M719-6WT was infected with Galleria mellonella larvae,the survival rate of the larvae was increased to 100％and 70％within 24 and 48 h after injection of 2×107 PFU/mL phage pEC-M719-6WT.2.[Conclusion]A phage against multi-drug resistant Escherichia coli was isolated.The phage had good environmental stability and antibacterial activity,clear bioinformatics characteristics,and good antibacterial effect in vivo and in vitro.It could produce synergistic effect when combined with antibiotics,and had potential application value in the treatment of clinical drug-resistant Escherichia coli infection.
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