Immunogenetic variation at MHC class I loci in desert bighorn sheep (Ovis canadensis nelsoni) reflects natural and anthropogenic fragmentation

Abstract

Desert bighorn sheep (Ovis canadensis nelsoni) are a charismatic ungulate native to western North America that live in mountainous regions. In the desert of the southwest United States, populations of bighorn sheep can be isolated by both natural and anthropogenic barriers. Isolation limits gene flow, exacerbates loss of genetic diversity to drift, and thus could affect immune responses to infectious agents. We sought to develop a relatively rapid method for determining MHC class I gene diversity in individual sheep. Both DNA and RNA were isolated from leukocytes collected from 154 adult bighorn sheep from across different mountain ranges in Southern California. Previously identified primers from domestic sheep were used to amplify exons 2 and 3 from OMHC I genes while appending adapters for PacBio circular consensus sequencing. We successfully identified over 40 unique MHC class I sequences expressed by bighorn sheep. Using a combination of phylogenic tree analysis and mapping MHC sequences to the domestic sheep genome, we detect at least 4 unique classical MHC class I regions. Comparison of results using either genomic DNA or mRNA (cDNA) as a template for analysis suggest that certain MHC alleles are expressed at different levels within leukocytes, while many potential pseudogenes were identified when genomic DNA was used as a template. Our data suggests using messenger RNA (cDNA) as a template to identify MHC class I diversity is superior to genomic DNA. Finally, we found clear patterns of MHC class I genotypes differentiated by population, which suggests that both natural and anthropogenic-induced population fragmentation can limit the diversity of MHC genes within a given population.