Biological characteristics and complete genomic analysis of a novel virulent bacteriophage that infects Klebsiella pneumoniae capsular type K63

Abstract:

[Background] Bacteriophages have a specific bactericidal ability to influence the evolution of bacteria and ecology. In recent years, due to the global emergence of multidrug-resistant bacterial strains, phage therapy has received renewed interest. [Objective] Biological characteristics and complete genome sequence of a novel virulent bacteriophage vB_KpnP_IME308 that infects Klebsiella pneumoniae capsular type K63. [Methods] A novel lytic phage was isolated from sewage of Chinese general Hospital of the PLA using a clinical Klebsiella pneumoniae strain as an indicator. The double-layer plate method was conducted the titer, optimal multiplicity of infection (MOI), one-step growth curve, thermostability and pH stability. Phage morphology was observed by transmission electron microscopy after purification. Phage genome was sequenced using the Illumina Miseq sequencing platform. Complete genome sequence was used for genome annotation, comparative genomics and evolutionary analyses. [Results] A novel phage, designated vB_KpnP_IME308, isolated from hospital sewage using a clinical Klebsiella pneumoniae strain as a host. The optimal MOI of phage vB_KpnP_IME308 is 0.001. One step growth curve showed that the latent period and the burst period of vB_KpnP_IME308 were 20 min and 80 min, respectively. The burst size was about 330 PFU/cell. The genome of vB_KpnP_IME308 is 43 091 base pairs (bp) with (G+C)mol% content of 59.3% and (A+T)mol% content of 46.1%. Electron microscopic observation showed that the phage belongs to the family Podoviridae. The BLASTn alignment showed that the genome of the phage had limited similarity with the currently known phages. The evolutionary relationship between phage vB_KpnP_IME308 and other Podoviridae phages was analyzed by major capsid protein and large terminase protein of phage vB_KpnP_IME308, which suggest that phage vB_KpnP_IME308 was a member of the genus Drulisvirus of subfamily Autographivirinae. [Conclusion] A lytic phage, vB_KpnP_IME308, was isolated successfully from hospital sewage, characterization and genome analysis of a novel bacteriophage vB_KpnP_IME308, which shows its potency to be developed as a novel alternative for multi-drug resistant Klebsiella pneumoniae infects control and treatment.

Key words:Klebsiella pneumonia, Bacteriophage, Biological characteristics, Genomic analysis