The exopolysaccharide Poly-N-Acetyl-Glucosamine (PNAG) coats Klebsiella pneumoniae in vivo
The conserved bacterial polysaccharide Poly-N-Acetyl-Glucosamine (PNAG) is a potential broad-spectrum vaccine candidate. While the immunogenicity of PNAG-based vaccine candidates has been established, characterisation of PNAG production across clinically relevant bacteria remains largely unknown. In particular, PNAG production in the Gram-negative pathogen Klebsiella pneumoniae (KP) is not well understood. Here, we demonstrate that PNAG production is prevalent in clinical KP isolates, where it is secreted as extracellular networks during adherent growth conditions. However, during severe KP pulmonary infection, KP PNAG production undergoes a switch to a cell-associated phenotype, coating the bacterial cell surface. By screening a panel of isogenic KP mutants in prominent cell surface components ({Delta}wcaJ,{Delta} rmpADC,{Delta} rfb,{Delta} ompA and{Delta} ompk36), we identified KP capsular polysaccharide as a key determinant underpinning the phenotype. Deleting genes involved in capsule synthesis ({Delta}wcaJ) and regulation ({Delta}rmpADC) resulted in cell-associated PNAG during adherent growth and infection of alveolar epithelial cells in vitro. Taken together, we describe a novel interaction between KP surface polysaccharides and detect for the first time, cell-associated PNAG in KP during lung infection, highlighting PNAG as an attractive KP vaccine antigen. Author summaryThe Gram-negative pathogen Klebsiella pneumoniae (KP) is a leading cause of hospital-associated lung and bloodstream infections worldwide. As KP exhibits resistance to most frontline antibiotics, there is a growing demand for immune-based strategies to treat KP infections. Poly-N-Acetyl-Glucosamine (PNAG) is a surface sugar produced by most clinically relevant bacteria, including KP. However, relatively little is known about PNAG production in KP. Therefore, we set out to characterise PNAG production in KP during in vitro growth and following lung infection in a pulmonary mouse model. During in vitro growth, KP produces extracellular PNAG networks. In contrast, during an in vivo severe lung infection, PNAG is found cell-associated, coating the bacterial surface. We propose that the visible change in KP PNAG between in vitro and in vivo environments is due to crosstalk with capsule, another polysaccharide on the KP surface. Together, this supports PNAG as an attractive KP antigen.