Decoration of pasteurella multocida lipopolysaccharide with phosphocholine is important for virulence

Phosphocholine (PCho) is an important substituent of surface structures expressed by a number of bacterial pathogens. Its role in virulence has been investigated in several species, in which it has been shown to play a role in bacterial adhesion to mucosal surfaces, resistance to antimicrobial peptides or sensitivity to complement-mediated killing. The lipopolysaccharide (LPS) structure of the Pasteurella multocida strain Pm70, whose genome sequence is known, has recently been determined and does not contain PCho. However, the LPS structures from the closely related, virulent P. multocida strains VP161 and X-73 were shown to contain PCho on their terminal galactose sugar residues. To determine if PCho was involved in virulence in P. multocida, we used subtractive hybridisation of the VP161 genome against the Pm70 genome to identify a four-gene locus (designated pcgDABC) which we show is required for addition of the PCho residues to LPS. The proteins predicted to be encoded by pcgABC showed identity to proteins involved in choline uptake, phosphorylation and nucleotide sugar activation of PCho. We constructed a mutant in the P. multocida VP161 pcgC gene and demonstrated that this strain produces LPS that lacks PCho on the terminal galactose residues. This pcgC mutant displayed reduced in vivo growth in a chicken infection model and was more sensitive to the chicken antimicrobial peptide fowlicidin-1 than wild-type P. multocida.