Three Capsular Polysaccharide Synthesis-Related Glucosyltransferases, GT-1, GT-2 and WcaJ, Are Associated With Virulence and Phage Sensitivity of Klebsiella pneumoniae

Abstract

<i>Klebsiella pneumoniae</i> (<i>K. pneumoniae</i>) spp. are important nosocomial and community-acquired opportunistic pathogens, which cause various infections. We observed that <i>K. pneumoniae</i> strain K7 abruptly mutates to rough-type phage-resistant phenotype upon treatment with phage GH-K3. In the present study, the rough-type phage-resistant mutant named K7R^R showed much lower virulence than K7. Liquid chromatography-tandem mass spectrometry (LC-MS-MS) analysis indicated that WcaJ and two undefined glycosyltransferases (GTs)- named GT-1, GT-2- were found to be down-regulated drastically in K7R^R as compared to K7 strain. <i>GT-1</i>, <i>GT-2</i>, and <i>wcaJ</i> are all located in the gene cluster of capsular polysaccharide (CPS). Upon deletion, even of single component, of <i>GT-1</i>, <i>GT-2</i>, and <i>wcaJ</i> resulted clearly in significant decline of CPS synthesis with concomitant development of GH-K3 resistance and decline of virulence of <i>K. pneumoniae</i>, indicating that all these three GTs are more likely involved in maintenance of phage sensitivity and bacterial virulence. Additionally, K7R^R and GT-deficient strains were found sensitive to endocytosis of macrophages. Mitogen-activated protein kinase (MAPK) signaling pathway of macrophages was significantly activated by K7R^R and GT-deficient strains comparing with that of K7. Interestingly, in the presence of macromolecular CPS residues (>250 KD), K7(Δ<i>GT-1</i>) and K7(Δ<i>wcaJ</i>) could still be bounded by GH-K3, though with a modest adsorption efficiency, and showed minor virulence, suggesting that the CPS residues accumulated upon deletion of <i>GT-1</i> or <i>wcaJ</i> did retain phage binding sites as well maintain mild virulence. In brief, our study defines, for the first time, the potential roles of GT-1, GT-2, and WcaJ in <i>K. pneumoniae</i> in bacterial virulence and generation of rough-type mutation under the pressure of bacteriophage.