Isolation and characterization of a novel phage targeting Klebsiella pneumoniae K2 capsular type

Abstract

Phage therapy is a promising therapeutic weapon in the fight against multidrug-resistant Klebsiella pneumoniae infections. A Klebsiella phage, KpTRp1, was isolated from wastewater sample collected in Tunisia. It exhibited specific lytic activity against a reference strain of K. pneumoniae serotype K2. Phage isolation and enrichment were performed using the double-layer agar method. It was characterized by transmission electron microscopy, physiological investigation and genomic sequence analysis. Transmission electron micrographs of KpTRp1 showed a myovirus morphology. The optimal multiplicity of infection by KpTRp1 was 0.0001 with a latent period lasted for 30 min. Its rise phase showed a burst period of almost 150 min and released a small burst size of about six plaque-forming units per infected cell. KpTRp1 was stable over a broad pH and temperature range. The phage genome sequencing revealed a linear double-stranded DNA genome of 48,003 base pairs with a G + C content of 48.79%, defining a new species of Sircambvirus genus of Jameshumphriesvirinae subfamily. Among the 68 open reading frames identified, about half were hypothetical proteins with unknown functions, the others being involved in DNA replication and metabolism, phage structure, host recognition and lysis. Remarkably, lytic activity of KpTRp1 against K. pneumoniae serotype K2 was confirmed by genomic analysis of life cycle, making it suitable candidate for phage therapy, especially since no antibiotic resistance, or lysogenic genes were detected in the phage genome.

Keywords: Klebsiella pneumoniae; Lysis; Comparative genomics; Host range; Phage isolation; Stability.