# H-NS Regulates the Virulence of Klebsiella pneumoniae by Affecting Capsular Polysaccharide Chain Synthesis and Anchoring

## TL;DR

This study shows that H-NS, a bacterial protein, controls virulence in Klebsiella pneumoniae by regulating capsule production and anchoring.

## Contribution

The novel finding is that H-NS regulates virulence by affecting capsule synthesis and anchoring in K. pneumoniae.

## Key findings

- Deleting hns in K. pneumoniae leads to a hypermucoid phenotype with increased polysaccharide chains but reduced capsule anchoring.
- H-NS represses capsule synthesis genes and downregulates wabG, which is crucial for capsule anchoring.
- Loss of H-NS results in unstable capsule attachment and reduced bacterial virulence in a mouse model.

## Abstract

H-NS (histone-like nucleoid-structuring protein) is a global regulator affecting diverse bacterial processes. This study aimed to elucidate the regulatory role of H-NS in the virulence of Klebsiella pneumoniae (K. pneumoniae), particularly in relation to capsule synthesis and anchoring. A clinically isolated ST11-KL64 strain of K. pneumoniae FK6741 with low virulence was used. The role of H-NS was evaluated using colony morphology, the string test, viscosity measurement, capsule quantification, transmission electron microscopy, growth curve, biofilm assay, a mouse infection model, transcriptomic analysis, and RT-qPCR. Deletion of hns converted FK6741 into a hypermucoid phenotype in the positive string test; capsule quantification and transmission electron microscopy (TEM) showed increased polysaccharide chains but a reduced and tightly bound capsule. The mutant was initially found to grow slowly but formed stronger biofilms. In vivo, it displayed reduced virulence but induced stronger inflammation. Molecular assays revealed upregulation of capsule synthesis genes (galF, wzi, wcaJ, and wzc) and downregulation of wabG, which is involved in capsule anchoring. H-NS represses capsule synthesis genes, limiting capsule formation in K. pneumoniae. In contrast, loss of H-NS downregulates wabG, a key gene involved in GalA-mediated capsule anchoring, resulting in unstable surface attachment and loss of capsular polysaccharides. Consequently, these unanchored polysaccharides fail to confer effective protection, resulting in reduced bacterial virulence.