ABSTRACT
Klebsiella pneumoniae strains typically express both smooth lipopolysaccharide (LPS) with O antigen molecules and capsule polysaccharide (K antigen) on the surface. A single mutation in a gene that codes for a UDP galacturonate 4-epimerase (uge) renders a strain with the O-:K- phenotype (lack of capsule and LPS without O antigen molecules and outer core oligosaccharide). The uge gene was present in all the K. pneumoniae strains tested. The K. pneumoniae uge mutants were unable to produce experimental urinary tract infections in rats and were completely avirulent in two different animal models (septicemia and pneumonia). Reintroduction of the single uge wild-type gene in the corresponding mutants completely restored the wild-type phenotype (presence of capsule and smooth LPS) independently of the O or K serotype of the wild type. Furthermore, complemented uge mutants recovered the ability to produce experimental urinary tract infections in rats and virulence in the septicemia and pneumonia animal models.
Subject(s)
Carbohydrate Epimerases/genetics , Genes, Essential , Klebsiella pneumoniae/pathogenicity , Pneumonia, Bacterial/physiopathology , Urinary Tract Infections/physiopathology , Animals , Bacterial Proteins/genetics , Bacterial Proteins/metabolism , Carbohydrate Epimerases/metabolism , Carbohydrate Sequence , Disease Models, Animal , Female , Klebsiella Infections/microbiology , Klebsiella Infections/physiopathology , Klebsiella pneumoniae/genetics , Male , Mice , Mice, Inbred ICR , Molecular Sequence Data , Pneumonia, Bacterial/microbiology , Rats , Rats, Wistar , Sequence Analysis, DNA , Urinary Tract Infections/microbiology , VirulenceABSTRACT
To determine the function of the waaE gene in the biosynthesis of the inner-core LPS of Klebsiella pneumoniae, a waaE non-polar mutant has been constructed. Data obtained from the comparative chemical analysis of LPS samples obtained from the wild-type, the mutant strain and the complemented mutant demonstrated that the waaE gene is involved in substitution of alpha-L-glycero-D-manno-heptopyranose I (L,D-HeppI) at the O-4 position by a beta-D-glucopyranose (beta-D-Glcp) residue. In addition, DNA amplification and nucleotide sequence determination studies revealed that waaE homologues located between the waaA and coaD genes are present in clinical isolates of Enterobacteriaceae containing the structure beta-D-Glcp-(1-->4)-alpha-L,D-HeppI (K. pneumoniae, Proteus mirabilis and Yersinia enterocolitica), as well as in strains of Serratia marcescens and Enterobacter aerogenes of unknown LPS-core structures. Complementation studies using non-polar waaE mutants prove that all the waaE homologues perform the same function. Furthermore, K. pneumoniae, Ser. marcescens and P. mirabilis non-polar waaE mutants showed reduced adhesion and pathogenicity. In addition, the Ser. marcescens and P. murabilis waaE mutants showed reduced swarming motility and ability to form biofilms in vitro. All these characteristics were rescued by reintroduction of the waaE gene independently of its origin. An easy DNA amplification method to detect this gene was established, which also helps in finding the potential presence of this structural feature [beta-D-Glcp-(1-->4)-alpha-L,D-HeppI] in the inner-core LPS of Enterobacteriaceae members with unknown LPS-core structures.
