Submitted to: Journal of Bacteriology
Publication Type: Peer reviewed journal
Publication Acceptance Date: 7/17/2009
Publication Date: 10/1/2009
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. Characterization and Comparative Analysis of the Genes Encoding Haemophilus parasuis Outer Membrane Proteins P2 and P5. Journal of Bacteriology. 191(19):5988-6002. Interpretive Summary: Haemophilus parasuis causes Glasser’s disease and systemic ailments in swine. These problems are of significant concern and economic importance to pork producers, but little is known about how H. parasuis causes disease. A related human pathogen, Haemophilus influenzae, has been better studied and its disease mechanisms are much better understood. On the basis of the demonstrated importance of the H. influenzae proteins P2 and P5 in human infection and immune response, we attempted to identify genes for the corresponding proteins in H. parasuis from a collection of 35 strains differing widely in virulence and representing all known serotypes. Gene sequences for both P2 and P5 were discovered in all strains. Comparison of the predicted protein sequences revealed extensive variability leading to 25 different types of P2 and 17 different types of P5. Additionally, we determined that the genes for both proteins are expressed by all strains examined. These results suggest that P2 and P5 proteins of H. parasuis are likely to be important in the bacterial/host interaction and may play roles both in causing disease and in immune protection. This knowledge will be of benefit in efforts to develop more effective treatments, diagnostic tools and preventative measures.
Technical Abstract: Haemophilus parasuis is a swine pathogen of significant industry concern, but little is known about how this organism causes disease. A related human pathogen, Haemophilus influenza, 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. The goals of this study were to identify, analyze and compare the genetic relatedness of orthologs of the genes encoding these two proteins in a diverse group of 35 strains of H. parasuis. Genes encoding P2 and P5 proteins were present in all H. parasuis strains evaluated. The predicted amino acid sequences for both exhibited considerable heterogeneity, particularly in regions corresponding to predicted extracellular loops. Twenty-five variants of P2 and 17 variants of P5 were identified. The P2 protein of 7 strains was predicted to contain a highly conserved extra loop as compared to the remaining strains and to H. influenzae P2. Antigenic site predictions coincide with predicted extracellular loop regions of both P2 and P5 . Neighbor-joining trees constructed using P2 and P5 sequences predict divergent evolutionary histories distinct from those predicted by a multilocus sequence typing phylogeny based on partial sequencing of 7 housekeeping genes. Real-time reverse transcription PCR indicates that both genes are expressed in all strains. Inter- and intra-species variation in surface-exposed loop regions suggests that these proteins may play important roles in virulence, host specificity, and evasion of host immune response.