Helmut Schaschl

MHC genes and oxidative stress in sticklebacks an immuno-ecological approach

Individual variation in the susceptibility to infection may result from the varying ability of hosts to specifically recognize different parasite strains. Alternatively, there could be individual host differences in fitness costs of immune defence. Although, these two explanations are not mutually exclusive, they have so far been treated in separate experimental approaches. To analyse potential relationships, we studied body condition and oxidative stress, which may reflect costs of immunity, in three-spined sticklebacks that had been experimentally exposed to three species of naturally occurring parasite. These sticklebacks differed in a trait, which is crucial to specific parasite defence, i.e. individual genetic diversity at major histocompatibility complex (MHC) class IIB loci. Oxidative stress was quantified as tissue acrolein, a technique that has been applied to questions of immuno-ecology for the first time. We measured gene expression at the MHC and other estimates of immune activation. We found that fish with high levels of MHC expression had poor condition and elevated oxidative stress. These results indicate that MHC-based specific immunity is connected with oxidative stress. They could, thus, also be relevant in the broader context of the evolution of sexually selected signals that are based on carotenoids and are, thus supposed to reflect oxidative stress resistance.

Genetic variation in MHC class II expression and interactions with MHC sequence polymorphism in three-spined sticklebacks

Genes of the major histocompatibility complex (MHC) have been studied for several decades because of their pronounced allelic polymorphism. Structural allelic polymorphism is, however, not the only source of variability subjected to natural selection. Genetic variation may also exist in gene expression patterns. Here, we show that in a natural population of three‐spined sticklebacks (Gasterosteus aculeatus) the expression of MHC class IIB genes was positively correlated with parasite load, which indicates increased immune activation of the MHC when infections are frequent. To experimentally study MHC expression, we used laboratory‐bred sticklebacks that were exposed to three naturally occurring species of parasite. We found strong differences in MHC class IIB expression patterns among fish families, which were consistent over two generations, thus demonstrating a genetic component. The average number of MHC class IIB sequence variants within families was negatively correlated to the MHC expression level suggesting compensatory up‐regulation in fish with a low (i.e. suboptimal) MHC sequence variability. The observed differences among families and the negative correlation with individual sequence diversity imply that MHC expression is evolutionary relevant for the onset and control of the immune response in natural populations.

Selection and recombination drive the evolution of MHC class II DRB diversity in ungulates

Major histocompatibility complex (MHC) antigen-presenting genes are the most variable loci in vertebrate genomes. Host–parasite co-evolution is assumed to maintain the excessive polymorphism in the MHC loci. However, the molecular mechanisms underlying the striking diversity in the MHC remain contentious. The extent to which recombination contributes to the diversity at MHC loci in natural populations is still controversial, and there have been only few comparative studies that make quantitative estimates of recombination rates. In this study, we performed a comparative analysis for 15 different ungulates species to estimate the population recombination rate, and to quantify levels of selection. As expected for all species, we observed signatures of strong positive selection, and identified individual residues experiencing selection that were congruent with those constituting the peptide-binding region of the human DRB gene. However, in addition for each species, we also observed recombination rates that were significantly different from zero on the basis of likelihood-permutation tests, and in other non-quantitative analyses. Patterns of synonymous and non-synonymous sequence diversity were consistent with differing demographic histories between species, but recent simulation studies by other authors suggest inference of selection and recombination is likely to be robust to such deviations from standard models. If high rates of recombination are common in MHC genes of other taxa, re-evaluation of many inference-based phylogenetic analyses of MHC loci, such as estimates of the divergence time of alleles and trans-specific polymorphism, may be required.

Recombination and the origin of sequence diversity in the DRB MHC class II locus in chamois (Rupicapra spp.).

We examined the evolutionary processes contributing to genetic diversity at the major histocompatibility complex (MHC) class II DRB locus in chamois (Rupicapra spp., subfamily Caprinae). We characterised the pattern of intragenic recombination (or homologous gene conversion) and quantified the amount of recombination in the genealogical history of the two chamois species, Pyrenean chamois (Rupicapra pyrenaica) and Alpine chamois (Rupicapra rupicapra). We found evidence for intragenic recombination, and the estimated amount of population recombination suggests that recombination has been a significant process in generating DRB allelic diversity in the genealogical history of the genus Rupicapra. Moreover, positive selection appears to act on the same peptide-binding residues in both analysed chamois species, but not in identical intensity. Recombination coupled with positive selection drives the rapid evolution at the peptide-binding sites in the MHC class II DRB gene. Many chamois MHC class II DRB alleles are thus much younger than previously assumed.

Spatial patterns of mitochondrial and nuclear gene pools in chamois (Rupicapra r. rupicapra) from the Eastern Alps

We have assessed the variability of maternally (mtDNA) and biparentally (allozymes) inherited genes of 443 chamois (Rupicapra r. rupicapra) from 19 regional samples in the Eastern Alps, to estimate the degree and patterns of spatial gene pool differentiation, and their possible causes. Based on a total mtDNA-RFLP approach with 16 hexanucleotide-recognizing restriction endonucleases, we found marked substructuring of the maternal gene pool into four phylogeographic groups. A hierarchical AMOVA revealed that 67.09% of the variance was partitioned among these four mtDNA-phylogroups, whereas only 8.04% were because of partitioning among regional samples within the populations, and 24.86% due to partitioning among individuals within regional samples. We interpreted this spatial pattern of mtDNA variability as a result of immigration of chamois from different Pleistocene refugia surrounding the Alps after the withdrawal of glaciers, rather than from topographic barriers to gene flow, such as Alpine valleys, extended glaciers or woodlands. However, this striking geographical structuring of the maternal genome was not paralleled by allelic variation at 33 allozyme loci, which were used as nuclear DNA markers. Wrights hierarchical F-statistics revealed that only ⩽0.45% of the explained allozymic diversity was because of partitioning among the four mtDNA-phylogroups. We conclude that this discordance of spatial patterns of nuclear and mtDNA gene pools results from a phylogeographic background and sex-specific dispersal, with higher levels of philopatry in females.

Polymorphic MHC loci in an asexual fish, the amazon molly (Poecilia formosa; Poeciliidae).

Genes of the major histocompatibility complex (MHC) encode molecules that control immune recognition and are highly polymorphic in most vertebrates. The remarkable polymorphisms at MHC loci may be maintained by selection from parasites, sexual selection, or both. If asexual species show equal (or higher) levels of polymorphisms at MHC loci as sexual ones, this would mean that sexual selection is not necessary to explain the high levels of diversity at MHC loci. In this study, we surveyed the MHC diversity of the asexual amazon molly (Poecilia formosa) and one of its sexual ancestors, the sailfin molly (P. latipinna), which lives in the same habitat. We found that the asexual molly has polymorphic MHC loci despite its clonal reproduction, yet not as polymorphic as the sexual species. Although the nucleotide diversity was similar between the asexual and sexual species, the sexual species exhibited a greater genotypic diversity compared to the asexual one from the same habitats. Within‐genome diversity was similar for MHC class I loci, but for class IIB, the sexual species had higher diversity compared to the asexual — despite the hybrid origins and higher levels of heterozygosity at microsatellite loci in the asexual species. The level of positive selection appears to be similar between the two species, which suggests that these polymorphisms are maintained by selection. Thus, our findings do not allow us to rule out the sexual selection hypothesis for the evolution of MHC diversity, and although the sexual fish has higher levels of MHC‐diversity compared to the asexual species, this may be due to differences in demography, parasites, or other factors, rather than sexual selection.

Contrasting mode of evolution between the MHC class I genomic region and class II region in the three-spined stickleback (Gasterosteus aculeatus L.; Gasterosteidae: Teleostei)

Major histocompatibility complex (MHC) class I molecules display peptides on cell surfaces for subsequent T-cell recognition and are involved in the immune response against intracellular pathogens. In this study, a BAC library was created from a single three-spined stickleback and screened for clones containing MHC class I genes. In a 163.2-kb genomic sequence segment of a single clone, we identified three MHC class I genes in the same transcriptional orientation. Two class I genes are potentially expressed and functional. In one class I gene, the transmembrane region is missing and could therefore present a pseudogene. Alternatively, it presents a functional gene that encodes a soluble MHC class Ib molecule. Despite genomic similarities to the MHC class II region, which is characterized by interlocus recombination, we did not find any evidence for this kind of recombination in the class I genes. It thus seems that interlocus recombination may play a rather minor role in generating class I diversity in stickleback and that the class I region displays a higher genomic stability (i.e., lower local recombination rate). In addition, two non-MHC genes (Oct-2 beta and Na+,K+-ATPaseα3) have been identified in the analyzed class I region. The Oct-2 beta gene is a transcription factor that is expressed primarily in B lymphocytes, in activated T-cells, and in neuronal cells. The Na+,K+-ATPaseα3 gene is primarily expressed in the brain and heart and mediates catalytic activities. Both genes are located on the same linkage group together with the MHC class I genes in the zebra fish. In humans, however, homologues of Oct-2 beta and ATPaseα3 lie outside the MHC region, which indicates that the concentration of immune genes found in mammalian genomes is a derived state.

On allozyme and cyt-b gene characteristics of Cretan hedgehogs, Erinaceus concolor nesiotes Bate, 1906

To infer the phylogenetic position of Cretan hedgehogs, En’naceus concolor nesiotes, we compared directly allozymic variation at 27 Loei and 383 bp-long sequences of the mitochondrial cytb gene of eleven and three hedgehogs, respectively, collected at diverse Locations in Crete with already published data of western and eastern hedgehogs (F. europaeus and E. concolor). Horizontal starch gel electrophoresis revealed 4 alleles at 2 polymorphic loci (Gpi, Acy) in Cretan hedgehogs. Indices of allozymic diversity were similar to those of regional samples of E. europaeus and E. concolor from Central Europe. A. Wagner dendrogram based on pairwise Rogers distances showed that Cretan hedgehogs clustered clearly with the E. concolor from Central Europe. The cytb sequences of the Cretan hedgehogs revealed 3 closely related haplotypes, that were tightly connected to haplotypes off. concolor from various provenances in the Balkans. In contrast, published haplotypes off. concolor from Asia Minor and Palestine differed distinctly from the Cretan haplotypes. We conclude that Cretan hedgehogs originate from mainlandGreece or other places in the Balkans.