Isolation of a bacteriophage specific for a new capsular type of Klebsiella pneumoniae and characterization of its polysaccharide depolymerase

PLoS One. 2013 Aug 2;8(8):e70092. doi: 10.1371/journal.pone.0070092. Print 2013.

Abstract

Background: Klebsiella pneumoniae is one of the major pathogens causing hospital-acquired multidrug-resistant infections. The capsular polysaccharide (CPS) is an important virulence factor of K. pneumoniae. With 78 capsular types discovered thus far, an association between capsular type and the pathogenicity of K. pneumoniae has been observed.

Methodology/principal findings: To investigate an initially non-typeable K. pneumoniae UTI isolate NTUH-K1790N, the cps gene region was sequenced. By NTUH-K1790N cps-PCR genotyping, serotyping and determination using a newly isolated capsular type-specific bacteriophage, we found that NTUH-K1790N and three other isolates Ca0507, Ca0421 and C1975 possessed a new capsular type, which we named KN2. Analysis of a KN2 CPS(-) mutant confirmed the role of capsule as the target recognized by the antiserum and the phage. A newly described lytic phage specific for KN2 K. pneumoniae, named 0507-KN2-1, was isolated and characterized using transmission electron microscopy. Whole-genome sequencing of 0507-KN2-1 revealed a 159 991 bp double-stranded DNA genome with a G+C content of 46.7% and at least 154 open reading frames. Based on its morphological and genomic characteristics, 0507-KN2-1 was classified as a member of the Myoviridae phage family. Further analysis of this phage revealed a 3738-bp gene encoding a putative polysaccharide depolymerase. A recombinant form of this protein was produced and assayed to confirm its enzymatic activity and specificity to KN2 capsular polysaccharides. KN2 K. pneumoniae strains exhibited greater sensitivity to this depolymerase than these did to the cognate phage, as determined by spot analysis.

Conclusions/significance: Here we report that a group of clinical strains possess a novel Klebsiella capsular type. We identified a KN2-specific phage and its polysaccharide depolymerase, which could be used for efficient capsular typing. The lytic phage and depolymerase also have potential as alternative therapeutic agents to antibiotics for treating K. pneumoniae infections, especially against antibiotic-resistant strains.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Bacterial Capsules / genetics
  • Bacterial Capsules / metabolism*
  • Bacteriophages / genetics
  • Bacteriophages / isolation & purification*
  • Bacteriophages / pathogenicity
  • DNA, Bacterial / genetics
  • Genotype
  • Glycoside Hydrolases / genetics
  • Glycoside Hydrolases / metabolism*
  • Humans
  • Immunoblotting
  • Klebsiella Infections / genetics
  • Klebsiella Infections / metabolism
  • Klebsiella Infections / microbiology*
  • Klebsiella pneumoniae / genetics
  • Klebsiella pneumoniae / isolation & purification*
  • Klebsiella pneumoniae / pathogenicity
  • Polymerase Chain Reaction
  • Virulence Factors / genetics
  • Virulence Factors / metabolism*

Substances

  • DNA, Bacterial
  • Virulence Factors
  • Glycoside Hydrolases
  • capsular-polysaccharide galactohydrolase

Grant support

This study was supported by grants from the National Science Council, National Taiwan University, and National Taiwan University Hospital in Taiwan. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.