Characterization of a PB1-like phage PHW2 infecting Pseudomonas aeruginosa

Abstract:

[Background] Pseudomonas aeruginosa is one of the main opportunistic pathogen that has an important role in nosocomial, acute, and chronic infections. The multidrug resistant (MDR) P. aeruginosa remains a common cause of severe nosocomial infection. Phages are viruses that infect and can kill bacteria, and phage therapy is an alternative treatment to eradicate MDR P. aeruginosa infection. [Objective] To isolate virulent phages that infect carbapenem-resistant P. aeruginosa and characterize its biological and genomic properties of phages for future development as phage therapy. [Methods] Environmental water samples were collected, and phages were isolated using double agar overlay plaque assay. The morphology, one-step growth curve, optimal multiplicity of infection and other biological characteristics of phages were studied. Phage genome was sequenced using the Illumina MiSeq sequencing platform. Newbler 3.0, GeneMarkS, BlASTp, and Mauve 2.4.0 were used for genome annotation, comparative genomics and evolutionary analyses. [Results] A new phage, PHW2, which is a member of Myoviridae, was isolated from pond water. It is able to lyse seven carbapenem-resistant P. aeruginosa clinical isolates. The optimal multiplicity of infection (MOI) is 0.1. In addition, one step growth curve showed that the latent period and rise period of PHW2 was 100 min and 360 min, respectively. The burst size was about 25 PFU/cell. The phage PHW2 was stable in the temperature of 25-50℃ and pH 6.0-8.0. The activity of PHW2 decreased significantly after UV irradiation for 7 min. Moreover, it was shown that the activity of PHW2 is unaffected after treatment with 5% chloroform for 100 min. The genome of PHW2 is 65 984 bp, with GC content of 55.69% and comprises of 92 ORFs. tRNAscan-SE analysis indicated that phage PHW2 does not possess tRNAs. The genome of PHW2 had high similarity with other PB1-like phages. Phage PHW2 inhibited the biofilm formation of P. aeruginosa PA001 within 24 h. [Conclusion] We isolated PHW2, a new PB1-like phage that shows potent lytic effect against several carbapenem-resistant P. aeruginosa clinical isolates. Biological characteristics and the in vitro biofilm test indicated that the phage has the potential to be used as a biological agent to control nosocomial multidrug resistant P. aeruginosa infections.

Key words:Pseudomonas aeruginosa;PB1-like phage;biological characteristics;genomic analysis