Figure 1.
MHC DRB allele diversity in Alpine ibex and chromosomal localization of genetic markers.
(A) Frequency distribution of the two MHC DRB exon 2 alleles in Alpine ibex populations across the Swiss Alps. The marker OLADRB1 was used to assess the frequency of Caib-DRB*2. All populations with n≥12 are shown except for Weisshorn (n = 9; marked by an asterisk). See Table S1 for complete allele frequency data. (B) Genetic diversity of MHC DRB alleles in different Caprinae species. The proportions of sites that differ between each pair of sequences are shown based on 227 bp sequence length. (C) Chromosomal locations of MHC genes and genetic markers were mapped to the goat chromosome 23 using the homologous cattle (Bos taurus) chromosome 23 as a reference (for further details see [40]). The MHC DRB exon 2 is shown in orange. Marker distances between the microsatellites OLADRB1 and OLADRB2 are based on cattle chromosome 23 and sheep chromosome 20. (D) Observed linkage disequilibrium (connected lines) between sequence and microsatellite alleles. Caib-DRB*2 was completely associated to allele 184 of microsatellite OLADRB1 (previously shown for a subset of 98 individuals in Alasaad et al. [28]) and allele 277 of OLADRB2 (except 2 individuals out of 707).
Figure 2.
Sequence alignments of coding and non-coding regions of the MHC DRB gene in Alpine ibex and domestic goat.
The DRB gene (11 kb in domestic sheep) is located on to the goat reference genome scaffold2167 at position 79'094 bp. A total of 2253 bp of the MHC DRB gene was sequenced including partial sequences of introns 1–4 and the complete intron 5 as well as a 3′ UTR sequence and complete sequences of exons 3, 5 and 6. Exon 6 is not shown as no polymorphism was found. Seven Alpine ibex homozygous for Caib-DRB*1 (red), seven Alpine ibex homozygous for Caib-DRB*2 (orange) and five domestic goat individuals selected from a screening of diverse breeds (grey) were sequenced. All seven Alpine ibex homozygous for Caib-DRB*2 were homozygous for the same haplotype at all four loci. This haplotype was highly distinct from the haplotype carried by all seven Alpine ibex homozygous for Caib-DRB*1. The domestic goat VBN.4 shared a nearly identical haplotype with individuals homozygous for Caib-DRB*2 at all four sequenced loci (99.8% sequence identity across 2253 bp). The domestic goat VBN.4 was homozygous for the DRB exon 2 Cahi-DRB*16, which is identical to Caib-DRB*2 (not shown). The phylogenetic trees are based on neighbor-joining. See Supporting Text S1-S4 for full sequence alignments.
Figure 3.
Alpine ibex and goat genotypes in 750DRB.
RAD sequencing SNP genotypes in a 750(see Figure S3). A representative set of Alpine ibex individuals (A) homozygous for Caib-DRB*1 (n = 10) and (B and C) carrying Caib-DRB*2 (n = 10) are shown in the top panels. (D) The corresponding RAD sequencing SNP genotypes for 9 domestic goat individuals. SNPs with a minor allele frequency of less than 0.1 within Alpine ibex were excluded. Alpine ibex carrying Caib-DRB*2 are nearly exclusively heterozygous at the RAD sequencing SNP loci and share alleles both with Alpine ibex homozygous for Caib-DRB*1 and with domestic goats.
Figure 4.
Linkage disequilibria (LD) and haplotype clustering in the region surrounding the MHC of Alpine ibex.
(A) LD heatmap (r2) of chromosome 23 for the Alpine ibex population Cape au Moine. The color gradient scale represents the range of r2 values. Red is used for the highest estimates of linkage disequilibrium. The chromosomal position of SNP16397 (diagnostic for Caib-DRB*2) is indicated by a blue asterisk. The red circle shows a SNP of a low minor allele frequency (0.06) that may explain the low LD. (B) NeigborNet networks were constructed from SNP haplotypes in three regions of chromosome 23. Caib-DRB*1 and Caib-DRB*2 haplotypes are colored in green and red, respectively. Networks are shown separately for three chromosomal sections of similar length comprising 7, 21 and 3 SNPs, respectively.
Figure 5.
Linkage disequilibrium (LD) heatmaps of chromosome 23 for Alpine ibex populations and domestic goat breeds.
(A) The Alpine ibex populations Albris, Rheinwald and Cape au Moine (shown on Figure 4A) have a high frequency of allele Caib-DRB*2 (Table S5). The Weisshorn population has a low frequency of Caib-DRB*2 (Table S5). Populations with a high frequency of Caib-DRB*2 showed larger blocks of strong LD than the Weisshorn population. The asterisks show the position of SNP16397, which was diagnostic for Caib-DRB*2. See Figure S5 for LD heatmaps of all chromosomes of population Albris. (B) In all six domestic goat breeds, LD blocks were generally small.
Figure 6.
Decay of linkage disequilibria (LD) on chromosome 23.
(A) The decay of pairwise linkage disequilibria (r2) among each pair of SNPs is shown against the corresponding physical distances. The three Alpine ibex populations Albris, Cape au Moine and Rheinwald showed high Caib-DRB*2 frequencies and a slow decay in LD over distance. Caib-DRB*2 was rare in the Weisshorn population and the LD decay is steeper. Rho (ρ) values provide estimates of the LD decay with higher values indicating steeper decays and shorter LD blocks. For details on Caib-DRB*2 allele frequencies see Table S5. (B) LD decay in six domestic goat breeds. The LD decay is much steeper in domestic goats than in Alpine ibex populations.
Figure 7.
Extended haplotype homozygosity (EHH) plots and bifurcation diagrams.
(A) The EHH plot (left panel) of the Alpine ibex population Cape au Moine shows the length of conserved haplotypes on both sides of the SNP diagnostic for Caib-DRB*2 (SNP16397). EHH = 1 indicates that all haplotypes containing the SNP allele (either SNP16397/A or SNP16397/G) are identical up to this position. EHH for SNP16397/G (diagnostic for Caib-DRB*2) is shown in red and the EHH for SNP16397/A (diagnostic for Caib-DRB*1) is shown in green. The bifurcation diagrams (center and right panel) show the branching of haplotypes on both sides of SNP16397. Branches at nodes suggest historical recombination events and the splitting of the haplotype at the node position. The bifurcation diagram for SNP16397/G (diagnostic for Caib-DRB*2) shows much longer haplotypes and fewer branchings at nodes than the bifurcation diagram for SNP16397/A (diagnostic for Caib-DRB*1). (B) The EHH plot and bifurcation diagram for the domestic goat breed Gemsfarbige Gebirgsziege show much shorter haplotypes and more extensive branching at nodes for both SNP alleles. See Figures S6 and S7 for additional Alpine ibex populations and domestic goat breeds.