Capsular Polysaccharide of Erysipelothrix rhusiopathiae, the Causative Agent of Swine Erysipelas, and Its Modification with Phosphorylcholine

Abstract

ABSTRACTThe capsule has been implicated in the virulence of the swine pathogenErysipelothrix rhusiopathiae, a rod-shaped, intracellular Gram-positive bacterium that has a unique phylogenetic position in the phylumFirmicutesand is a close relative ofMollicutes(mycoplasma species). In this study, we analyzed the genetic locus and composition of the capsular polysaccharide (CPS) of the Fujisawa strain ofE. rhusiopathiae. Genome analysis of the Fujisawa strain revealed that the genetic locus for capsular polysaccharide synthesis (cps) is located next to anlicoperon, which is involved in the incorporation and expression of phosphorylcholine (PCho). Reverse transcription-PCR analysis showed thatcpsandlicare transcribed as a single mRNA, indicating that the loci form an operon. Using the cell surface antigen-specific monoclonal antibody (MAb) ER21 as a probe, the capsular materials were isolated from the Fujisawa strain by hot water extraction and treatment with DNase, RNase, pronase, andN-acetylmuramidase SG, followed by anion-exchange and gel filtration chromatography. The materials were then analyzed by high-performance liquid chromatography, mass spectrometry, and nuclear magnetic resonance (NMR) spectroscopy. The CPS ofE. rhusiopathiaeis heterogeneous and consists of the major monosaccharides galacturonic acid, galactose, mannose, glucose, arabinose, xylose, andN-acetylglucosamine and some minor monosaccharides containing ribose, rhamnose, andN-acetylgalactosamine. In addition, the capsule is modified by PCho, which comigrates with the capsular materials, as determined by Western immunoblotting, and colocalizes on the cell surface, as determined by immunogold electron microscopy. Virulence testing of PCho-defective mutants in mice demonstrated that PCho is critical for the virulence of this organism.