| Escherichia coli O157: H7 is an important foodborne pathogen that causes severe bloody diarrhea,hemorrhagic colitis,and hemolytic uremic syndrome.The contamination of E.coli O157 has become an important factor affecting the global public safety of food.Every year,there have been outbreaks of diarrhea and even death in the world due to eating vegetable and meat contaminated with E.coli O157.However,traditional food sterilizing agents have not been able to meet the increasingly serious food safety problems.Bacteriophage has gradually attracted people's attention due to its specific sterilization method.In this study,two lytic phages,named FEC14 and FEC19 against Escherichia coli O157: H7,were isolated from hospital sewage.Both phages belong to the family Myoviridae with highly lytic,broad-spectrum,and highly stable characteristics,but they were significantly different from morphology,Transmission electron microscopy(TEM)analysis showed that phage FEC19 has an isometric head 58±3 nm in diameter and contractile tails whose terminal spikes presents pin shape,while phage FEC14 has an isometric head 80 ±5nm in diameter and contractile tails whose terminal spikes presents star shape.The host range test revealed that phage FEC14 showed specific inhibition against all of 14 strains of E.coli O157,while phage FEC19 only specifically inhibits 11 strains of E.coli.The host range of phage FEC14 is broader than that of phage FEC19.The optimal MOI of FEC14 and FEC19 were 0.001 and 0.1,respectively.FEC14 had latent period of 15 min,growing period of 65 min.FEC19 had latent period of 10 min,growing period of 80 min.Phage FEC14 is more sensitive to temperature than FEC19.At 70 ?,phage FEC14 was gradually inactivated within 40 min,while phage FEC19 was completely inactivated at 80 ? for 20 min.Phage FEC14 was stable over a board range of pH from 3 to 12.while phage FEC19(pH 4-11)was less stable than phage FEC14 in acid-base environment.The genome of FEC19 was digested by EcoR??Hind??Xba?,but The genome of FEC14 was resistant to ten types of restriction endonucleases used in laboratory,which may suggest the presence of modified nucleotides.Phage FEC14 and FEC19 were sequenced and uploaded to Genbank,and their Accession Numbers were MG383452 and MH816966,respectively.Because phage FEC14 contains more gene information than phage FEC19,only the phage FEC14 genome was selected for further analysis.Sequence analysis showed that the genome of phage FEC14 was 158639 bp in length with a GC content of 44.6%,and the genome contained 210 ORFs and 4 tRNAs of which 73 are predicted to have known functions,and the existence of virulence genes and drug resistance genes has not yet been found in the database.Evolutionary analysis revealed that phage FEC14 belongs to the latest division of Myoviridae family,Tevenvirinae subfamily,Vilvirus genus.It is concluded that the phage has different characteristics distinguishing from other Myoviridae phages: morphology,genome size and organization,gene synteny,use of non-canonical nucleotide,etc.Finally,a preliminary study on the application of phage to evaluate the bactericidal effect of phage on artificially contaminated beef showed that the application of FEC14,FEC19 phage separately or cocktail method on the infected beef can effectively reduce the number of host bacteria.The control effect of phage FEC19 is stronger than that of phage FEC14.The control effect at 4? is stronger than that at room temperature.The single application of phage is equivalent to the control effect of cocktail.However,considering the tolerance of host bacteria,it is best to use the cocktail method to prevent and control the host bacteria.In summary,the lytic bacteriophage FEC14 and FEC19 have great potential in controlling the application of Escherichia coli O157 contamination in food,and can be developed into products for food safety in the future. |
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