Genetic origins and diversity of bushpigs from Madagascar (Potamochoerus larvatus, family Suidae)

Abstract

The island of Madagascar, situated off the southeast coast of Africa, shows the first evidence of human presence ~ 10,000 years ago; however, other archaeological data indicates a settlement of the modern peoples of the island distinctly more recent, perhaps > 1500 years ago. Bushpigs of the genus Potamochoerus (family Suidae), are today widely distributed in Madagascar and presumed to have been introduced from Africa at some stage by human immigrants to the island. However, disparities about their origins in Madagascar have been presented in the literature, including the possibility of endemic subspecies, and few empirical data are available. Furthermore, the separation of bushpigs in Madagascar from their mainland relatives may have favoured the evolution of a different repertoire of immune genes first due to a founder effect and then as a response to distinct pathogens compared to their ancestors. Molecular analysis confirmed the species status of the bushpig in Madagascar as P. larvatus, likely introduced from the central region of southern Africa, with no genetic evidence for the recognition of eastern and western subspecies as suggested from previous cranial morphology examination. Investigation of the immunologically important SLA-DQB1 peptide-binding region showed a different immune repertoire of bushpigs in Madagascar compared to those on the African mainland, with seventeen exon-2 haplotypes unique to bushpigs in Madagascar (2/28 haplotypes shared). This suggests that the MHC diversity of the Madagascar populations may have enabled Malagasy bushpigs to adapt to new environments.

Figure 1

Map of native bushpig distribution as illustrated in the IUCN. Map is a modified version by mapchart.net.

Figure 2

Phylogenetic tree indicating the position of Malagasy bushpigs as Potamochoerus larvatus. A Maximum Likelihood tree was generated, based on concatenated mtDNA (CR and Cytb) and nuDNA (GPIP and MC1R) sequences, using the RAxML-NG v0.6.0 web-server (https://raxml-ng.vital-it.ch/#/). Statistical support was assessed using a bootstrap cut-off of 0.03 (Bootstrap support indicated above the branches). Numbers in brackets indicate the number of sequences from this study within each node.

Figure 3

Median joining network of Malagasy bushpigs based on concatenated mtDNA (CR and cytb). Coloured circles represent the haplotypes identified by DnaSP v6 with each colour represented by a location as indicated by the figure key, and the size of the circle is proportional to the frequency of each haplotype (with ‘mt’ as a prefix to indicate mitochondrial). Numbers in parentheses along the branches represents the mutational steps between each haplotype.

Figure 4

Malagasy bushpig mtDNA (CR and cytb) sequence by sampling location. Each colour indicates the different haplotypes (with ‘mt’ as a prefix to indicate mitochondrial) with the number of individuals in each population shown in brackets. The number of different haplotypes within each location is highlighted by the black box adjacent to each circle. Figure includes an edited map generated by mapchart.net and haplotypes produced in PopART (https://popart.otago.ac.nz/index.shtml).

Figure 5

Median joining network of Malagasy bushpigs based on MHC loci SLA-DQB1 exon-2 sequence. Coloured circles represent the haplotypes identified by DnaSP v6 with each colour represented by a location as indicated by the figure key, and the size of the circle is proportional to the frequency of each haplotype. Numbers in parentheses along the branches represents the mutational steps between each haplotype. Haplotypes with the same translated amino acid sequence are outlined.

Figure 6

Malagasy bushpig MHC class II SLA-DQB1 sequences by sampling location. Each colour indicates the different haplotypes with the number of individuals in each population shown in brackets. The number of different haplotypes within each location is highlighted by the black box adjacent to each circle. Figure includes an edited map generated by mapchart.net and haplotypes produced in PopART (https://popart.otago.ac.nz/index.shtml).

Figure 7

Phylogenetic analysis of the SLA-DQB1 haplotypes including all haplotypes (and samples). Haplotype numbers are aligned to their position on the maximum likelihood tree with their frequencies (%). The presence of each haplotype in the specific localities is indicated by the shaded box. Group numbers indicate identical antigen-binding sites of the relevant haplotypes.

Figure 8

Amino-acid sequence and selection of SLA-DQB1 exon-2 haplotypes. Analysis was completed using DataMonkey HyPhy server. Dots represent identical residues to DQB1-1 at the positions shown. Grey shaded sites indicate the antigen-binding sites and yellow indicate the conserved cysteines. Positions significant for positive (+) or negative (−) selection for at least two methods (MEME/FEL/SLAC/REL) are indicated for each population/location category. Dotted lines indicate haplotypes found in bushpigs and haplotypes in bold font are found in mainland bushpigs.