Characterization of Major Histocompatibility Complex (MHC) DRB Exon 2 and DRA Exon 3 Fragments in a Primary Terrestrial Rabies Vector (Procyon lotor)

The major histocompatibility complex (MHC) presents a unique system to explore links between genetic diversity and pathogens, as diversity within MHC is maintained in part by pathogen driven selection. While the majority of wildlife MHC studies have investigated species that are of conservation concern, here we characterize MHC variation in a common and broadly distributed species, the North American raccoon (Procyon lotor). Raccoons host an array of broadly distributed wildlife diseases (e.g., canine distemper, parvovirus and raccoon rabies virus) and present important human health risks as they persist in high densities and in close proximity to humans and livestock. To further explore how genetic variation influences the spread and maintenance of disease in raccoons we characterized a fragment of MHC class II DRA exon 3 (250bp) and DRB exon 2 (228 bp). MHC DRA was found to be functionally monomorphic in the 32 individuals screened; whereas DRB exon 2 revealed 66 unique alleles among the 246 individuals screened. Between two and four alleles were observed in each individual suggesting we were amplifying a duplicated DRB locus. Nucleotide differences between DRB alleles ranged from 1 to 36 bp (0.4–15.8% divergence) and translated into 1 to 21 (1.3–27.6% divergence) amino acid differences. We detected a significant excess of nonsynonymous substitutions at the peptide binding region (P = 0.005), indicating that DRB exon 2 in raccoons has been influenced by positive selection. These data will form the basis of continued analyses into the spatial and temporal relationship of the raccoon rabies virus and the immunogenetic response in its primary host

Average nucleotide and amino acid distances among raccoon MHC DRB exon 2 alleles.

<p>Standard errors (in parentheses) were obtained through 1000 bootstrap replicates. Distances were corrected for multiple substitutions using K2P model for nucleotide distances and Poisson distribution for amino acid differences. Putative peptide binding region (PBR) sites were those determined by Brown et al. <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0012066#pone.0012066-Brown1" target="_blank">[38]</a> and Stern et al. <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0012066#pone.0012066-Stern1" target="_blank">[39]</a> Distances are given as a percentage per site.</p

Innate immune responses in raccoons after raccoon rabies virus infection

Zoonotic wildlife diseases pose significant health risks not only to their primary vectors but also to humans and domestic animals. Rabies is a lethal encephalitis caused by rabies virus (RV). This RNA virus can infect a range of terrestrial mammals but each viral variant persists in a particular reservoir host. Active management of these host vectors is needed to minimize the negative impacts of this disease, and an understanding of the immune response to RV infection aids strategies for host vaccination. Current knowledge of immune responses to RV infection comes primarily from rodent models in which an innate immune response triggers activation of several genes and signalling pathways. It is unclear, however, how well rodent models represent the immune response of natural hosts. This study investigates the innate immune response of a primary host, the raccoon, to a peripheral challenge using the raccoon rabies virus (RRV). The extent and temporal course of this response during RRV infection was analysed using genes predicted to be upregulated during infection (IFNs; IFN regulatory factors; IL-6; Toll like receptor-3; TNF receptor). We found that RRV activated components of the innate immune system, with changes in levels of transcripts correlated with presence of viral RNA. Our results suggest that natural reservoirs of rabies may not mimic the immune response triggered in rodent models, highlighting the need for further studies of infection in primary hosts.</jats:p

Bayesian phylogenetic relationship of raccoon MHC DRB exon 2.

<p>This tree was constructed using the best fit model from the JModel test <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0012066#pone.0012066-Guindon1" target="_blank">[42]</a>, <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0012066#pone.0012066-Posada1" target="_blank">[43]</a>. In addition to the 66 raccoon alleles, three MHC alleles belonging to other mammals were included as outgroups: <i>Zalophus californianus</i> (GenBank Accession AY491456), <i>Ailuropoda melanoleuca</i> (GenBank Accession EF125965), and <i>Mustela lutreola</i> (GenBank Accession EU263550).</p

Number of individuals and geographic locations of MHC DRB exon 2 alleles.

<p>Number of individuals and geographic locations of MHC DRB exon 2 alleles.</p