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

Abstract

Klebsiella pneumoniae ( K. pneumoniae ) spp. are important nosocomial and community-acquired opportunistic pathogens, which cause various infections. We observed that K. pneumoniae 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. GT-1 , GT-2 , and wcaJ are all located in the gene cluster of capsular polysaccharide (CPS). Upon deletion, even of single component, of GT-1 , GT-2 , and wcaJ resulted clearly in significant decline of CPS synthesis with concomitant development of GH-K3 resistance and decline of virulence of K. pneumoniae , 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 GT-1 ) and K7(Δ wcaJ ) 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 GT-1 or wcaJ 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 K. pneumoniae in bacterial virulence and generation of rough-type mutation under the pressure of bacteriophage.

Genomic Profiling Reveals Distinct Routes To Complement Resistance in Klebsiella pneumoniae

“…3 ). Note that mutagenesis of genes of the capsule locus can cause secondary cell envelope defects (shown for wza and wzi [ 37 ]), and not all cps locus mutations entirely eliminate capsule production ( 34 , 38 ); therefore, complete consistency of selection across all genes of the cps locus is not expected. With K. pneumoniae B5055, which produces copious amounts of K2 capsule, 11/18 genes of the cps locus were called as hits, accounting for half of the serum survival determinants of this strain.…”

A Frameshift Mutation in wcaJ Associated with Phage Resistance in Klebsiella pneumoniae

“…Colanic acid was previously demonstrated to be associated with virulence in K. pneumoniae, including evidence that colanic acid mutants cannot survive with macrophages. It was also found that wcaJ is essential for virulence in mice [12]. However, some studies do not support previous conclusions regarding the role of colanic acid in biofilm formation or virulence [28].…”

“…WcaJ is known as the initiating enzyme for colanic acid synthesis, transferring the glucose-1-phosphate moiety from undecaprenyl-phosphate glucose (UDP-Glc) onto the carrier lipid undecaprenyl phosphate [11]. Interestingly, Cai et al [12] reported that WcaJ was significantly down-regulated in phage GH-K3 mediated K. pneumoniae K7R R mutants. In their study, the potential mechanism was not likely to have involved genetic mutations, as they proposed that epigenetic modifications such as DNA methylation could be linked to the down-regulation of wcaJ transcription and expression levels.…”

<p>AmpR Increases the Virulence of Carbapenem-Resistant <em>Klebsiella pneumoniae</em> by Regulating the Initial Step of Capsule Synthesis</p>

“…The potential role of WcaJ in the virulence of K. pneumoniae was defined recently. 33 The results of proteome analysis suggest that AmpR enhances the virulence of K. pneumoniae by regulating the initial step of capsule synthesis. As a regulator, AmpR needs to bind to the gene promoter to exert its regulatory role.…”