Isolation,identification and biological characters of bacteriophages of Proteus mirabilis

Objective To isolate and identify the bacteriophages of Proteus mirabilis,and analyze its biological characters.Methods Bacteriophages specific to P.mirabilis were isolated from the raw sewage by double-agar plaque method using P.mirabilis as host cells,then purified and observed for morphology and size by electron microscopy using negative staining.The bacteriophages and host cells were co-cultured at a ratio of 10 ∶ 1,based on which the one-step growth curve of bacteriophages was plotted,and the bacteriophage DNA was extracted and digested with restriction endonucleases.Results Four bacteriophage strains named as PrM-01,PrM-02,PrM-03 and PrM-04 were isolated,which formed round and transparent plaques with clear borders on double-agar plate,at diameters of 2 ～ 3 mm.The titers of bacteriophages after repeat purification were(1.2 ～ 2.3) × 1012 PFU / ml.Electron microscopy showed that the head of bacteriophages exhibited an icosahedra symmetry,at a diameter of about 40 nm,while the tail was at a length of about 30 nm.The one-step growth curve of PrM-04 revealed incubation period,time of outbreak and amount of lysis of 15 min,25 min and 3.2 × 104 respectively.Restriction analysis showed that the genome of bacteriophages was at a length of about 26 000 bp,with restriction sites of EcoRⅠ,Hind Ⅲ and BamHⅠ.Conclusion Four bacteriophage strains specific to P.mirabilis were successfully isolated,which showed characters of virulent bacteriophages,such as high specificity and strong lytic ability.