Investigation of LuxS-mediated quorum sensing in Klebsiella pneumoniae

Abstract

<b>Introduction.</b> Autoinducer-2 (AI-2) quorum sensing is a bacterial communication system that responds to cell density. The system requires <i>luxS</i> activity to produce AI-2, which can regulate gene expression and processes such as biofilm formation.<b>Aim.</b> To investigate the role of <i>luxS</i> in biofilm formation and gene expression in the nosocomial pathogen <i>Klebsiella pneumoniae</i>.<b>Methodology.</b> A <i>ΔluxS</i> gene deletion was made in <i>K. pneumoniae</i> KP563, an extensively drug-resistant isolate. AI-2 production was assessed in wild-type and <i>ΔluxS</i> strains grown in media supplemented with different carbohydrates. Potential roles of <i>luxS</i> in biofilm formation were investigated using a microtiter plate biofilm assay and scanning electron microscopy. Quantitative RT-PCR evaluated the expression of lipopolysaccharide (<i>wzm</i> and <i>wbbM</i>), polysaccharide (<i>pgaA</i>), and type 3 fimbriae (<i>mrkA</i>) synthesis genes in wild-type and <i>ΔluxS</i> mutant biofilm extracts.<b>Results.</b> AI-2 production was dependent on the presence of <i>luxS</i>. AI-2 accumulation was highest during early stationary phase in media supplemented with glucose, sucrose or glycerol. Changes in biofilm architecture were observed in the <i>ΔluxS</i> mutant, with less surface coverage and reduced macrocolony formation; however, no differences in biofilm formation between the wild-type and <i>ΔluxS</i> mutant using a microtiter plate assay were observed. In <i>ΔluxS</i> mutant biofilm extracts, the expression of <i>wzm</i> was down-regulated, and the expression of <i>pgaA</i>, which encodes a porin for poly-β-1,6-N-acetyl-d-glucosamine (PNAG) polysaccharide secretion, was upregulated.<b>Conclusion.</b> Relationships among AI-2-mediated quorum sensing, biofilm formation and gene expression of outer-membrane components were identified in <i>K. pneumoniae</i>. These inter-connected processes could be important for bacterial group behaviour and persistence.