Submitted to: American Society for Microbiology General Meeting
Publication Type: Abstract Only
Publication Acceptance Date: 2/17/2009
Publication Date: 5/17/2009
Citation: Mullins, M.A., Register, K.B., Bayles, D.O., Loving, C.L., Nicholson, T.L., Brockmeier, S.L., Dyer, D.W., Phillips, G.J. 2009. Identification and comparative analysis of genes encoding outer membrane proteins P2 and P5 in Haemophilus parsuis [abstract]. 109th American Society for Microbiology General Meeting. Poster No. Z-034.
Technical Abstract: Haemophilus parasuis is a serious swine pathogen but little is known about how it causes disease. A related human pathogen, Haemophilus influenzae, has been better studied and many of its virulence factors have been identified. Two of these, outer membrane proteins P2 and P5, have been shown to have important virulence properties, including binding to human mucin and CEACAM1, and immune evasion. The goals of this study were to identify, compare, and analyze orthologs of genes encoding these two proteins in a diverse group of H. parasuis. Thirty-five strains of H. parasuis were selected for study. P2- and P5-specific PCR primers were designed using the draft genome of strain 29755. Amplicons covering the coding region for both genes were sequenced and assembled. DNA sequences were translated into predicted protein sequences, aligned, and analyzed using predictive software. Neighbor-joining and maximum parsimony trees were constructed. A multilocus sequence typing tree, using regions of seven housekeeping genes from the same 35 strains, was made for comparison. Gene expression in all strains was measured using real-time PCR. Genes encoding P2 and P5 proteins were present in all 35 H. parasuis strains. Sequences for both exhibited considerable heterogeneity, particularly in regions corresponding to predicted extracellular loops. Twenty-six variants of P2 and 17 variants of P5 were identified. For seven strains, the P2 protein was predicted to contain an extra loop compared to other strains and to H. influenzae P2. Antigenic site predictions coincided with predicted outer loop regions of both proteins. Trees constructed using P2 and P5 sequences predicted different evolutionary relationships from each other and from the MLST tree. Real-time PCR indicated that both genes were expressed in all strains. Conclusions. Orthologs of the H. influenzae genes for outer membrane proteins P2 and P5 are widely present in H. parasuis. Inter- and intra-species variation in surface-exposed loop regions suggest that these proteins may play important roles in virulence, host specificity, and evasion of host immune response.