Avian MHC : Characterization and expression patterns of classical and non-classical MHC-I genes

Sammanfattning: The function of the vertebrate immune system is to enable recognition and elimination of microorganisms that can cause harm (pathogens). A key component in adaptive immunity is the major histocompatibility complex (MHC) that codes for molecules with antigen presentation function. There are two main types of MHC molecules, class-I (MHC-I) and class-II (MHC-II) and the focus of my thesis has been MHC-I. The number of MHC gene copies can vary greatly between populations, species and orders. Passerine birds (order Passeriformes) seem to have particularly high MHC diversity (defined as number of different MHC alleles per individual). The main aim of my thesis has been to understand the high MHC-I diversity in passerines focusing on two aspects: gene expression and presence of classical and non-classical MHC-I genes. Classical MHC molecules present antigens and trigger adaptive immune responses, whereas non-classical genes have an as yet largely unclear function in immunity. Non-classical MHC genes have lower diversity and often have lower expression. Non-classical genes have been found in several different bird orders although never been fully characterized in passerines. I investigated the presence of classical and non-classical genes using a phylogenetic approach by comparing three closely related sparrow species; house sparrow (Passer domesticus), Spanish sparrow (Passer hispaniolensis) and tree sparrow (Passer montanus). All three species had putatively classical and non-classical genes. All putatively non-classical alleles formed a highly supported cluster, independent of species, indicating that the presence of classical and non-classical genes predates the speciation event of these sparrows. Moreover, only one classical gene was found to be highly expressed in house sparrows and tree sparrows (Paper I). The high diversity of MHC-I alleles makes them difficult to genotype, but with high throughput sequencing (HTS) it is feasible. Initially, I used Roche 454 and then Illumina MiSeq amplicon sequencing. These two HTS methods were evaluated using house sparrow MHC-I and they both performed well (Paper II). We characterized MHC-I in a non-passerine bird, the Icelandic black-tailed godwits (Limosa limosa islandica) and showed that there were no putatively non-classical genes in this species despite that such genes were found in two closely related Charadriiformes species (Paper III). I partly characterized MHC-I in siskins (Spinus spinus) of the order Passeriformes and it had putatively non-classical genes; one highly supported cluster contained only low expression alleles that also had low diversity (Paper IV). Moreover, I found that as many as three classical genes could have high expression. Expression of MHC-I was then studied in an infection experiment with a mild and a severe avian malaria strain (Paper V). We showed that classical alleles were continuously expressed to a higher degree than non-classical alleles. Moreover, MHC-I was differently expressed in infected individuals compared to control individuals and there was a tendency for MHC-I to be more highly expressed soon after the acute phase of the severe malaria infection. In my thesis, I have shown that the presence of classical and non-classical MHC-I genes most likely is a common feature in passerine birds which has so far been overlooked. Moreover, the expression of MHC-I in passerine birds seems to differ considerably, not only between species but also between classical MHC-I alleles within individuals of the same species – an avenue for future research.