It seems that Orwell was right: all pigs are equal, but some are more equal than others.
The question of how diverse different breeds of domesticated animals may be has begun to be addressed with the completion of many "genome projects" directed towards household animals like dogs and cats. Pigs are animals that are extremely important economically both historically and today, and that come in many different breeds throughout the world. This species (Sus scrofu) arose from the common ancestor with wild boars about 1.2 million year ago, and then diverged into European pigs and Chinese pigs. This was not a speciation event but a population one (similar in its way with the divergence between human races), which in pigs occurred sometime in the Pleistocene and possibly related to the last large glaciation event in the Northern Hemisphere (about 130,000 years ago). These events were complete long before pigs were domesticated by humans, in both China and the Middle East, about 10,000 years ago. China has almost one-third of the breeds of pigs in the world (~100 breeds), with there being a recognized difference in phenotype between pigs in the north and south of the country, inter alia, differences in thermoregulatory mechanisms that have adapted these pigs to their different environments.
Now a group led by researchers from Jiangxi Agricultural College* has sequenced genomes of 69 pigs (Sus scrofa) from different parts of China, specifically 15 geographically distinct regions, as reported in Nature Genetics ("Adaptation and possible ancient interspecies introgression in pigs identified by whole-genome sequencing," Nature Genetics (2015) (Published online 26 January 2015)). These pigs were members of eleven "geographically diverse" breed plus three populations of wild boar, which included the breeds Bamaxiang, Wuzhishan, and Luchuan and southern Chinese wild boars (comprising the southern population); and Erhualian, Hetao, Laiwu, Min, Tibetan (Gansu), Tibetan (Sichuan) Tibetan (Tibet), and Tibetan (Yunnan) breeds and the northern Chinese wild boar (comprising the northern population; although the Tibetan pigs are at similar latitudes as the southern populations their elevation is closer in climate to the northern pigs).
The types of studies reported by the Jiangxi group have been used (on pigs and other species) to identify "genomic signature of selection" indicative of domestication (wherein both natural selection and artificial (human) selection has been at work). This work extends that to individuals in distinct populations within a species to identify such signatures related to geographic condition-related adaptations.
While there were a variety of interesting results to this study, one broad conclusion these authors made is that pigs have adapted to a hot and wet vs. a cold and dry climate that is unique to China in view of its geographic range.
The statistics of the genomic DNA sequencing from these 69 pigs is as follows:
• Greater than 25-fold coverage of the pig genomes, 95% genome coverage;
• About 41 million (40,820,483) SNPs were detected, with 21 million of these that are not in the reference pig genomic database;
• 26.6 million of these SNPs were intergenic;
• 188,664 SNPs were exonic;
• About 500,000 SNPs were intronic (the authors interpret the non-exonic SNPs as indicating that regulatory variants were involved in the observed genetic adaptations);
• An average of 13.3 million SNPs were detected per individual, with Chinese wild boar showing the highest average (14.5 million); wild boars also had the highest number of population-specific SNPs (767,770);
• An average of 794 "loss-of-function" mutations in each pig breed but only 3 breed-specific LOF variants, including "a nonsense mutation in the PKD1L3 gene in Laiwu pigs, a splice-site mutation in the IFLTD1 gene in Luchuan pigs and a splice-site mutation in the SKIL gene in Min pigs";
• 5,663,829 indels (insertion/deletion events) and 44,170 structural variations;
• 2,000,000 copies of tRNA-derived short interspersed repeat (SINEs) distributed in the chromosomes in such a way that suggested these elements were "an important source of genetic diversity in pigs."
Pig genes associated with geographically relevant diversity (i.e., differences between "northern" breeds and "southern" breeds) included genes involved in blood coagulation and circulation (TBC1D8, KIF2B, VPS13A, and GNA14); vasculature development (FZD10), neural tube closure (SUFU), cellular chloride ion homeostasis (ABCC2), and hair development (DCAF17).
Locus-specific branch-length analyses revealed 774 "sweep" regions implicating 219 genes with population-related reductions in heterozygosity and recombination. These regions comprised on average 51kb on pig autosomes and one large ~14 Mb sweep in the X chromosome. On the autosomes, the strongest sweep signal from chromosome 15 spanning 300 kb, comprising the locus for the DCAF17 gene. This region was found to be "nearly devoid of genetic variability" and the hair development gene DCAF17 showed "clear allelic imbalance" between the two pig populations. This was consistent with the authors' hypothesis that the pig population genomes had adapted to the climate in which the different pig breeds were found in China, and with the observed phenotypes of sparse hair in southern pigs and dense, insulating hair in northern pigs.
These studies also implicated 219 genes in these sweep regions, and the authors reported finding missense mutations in 10 of these 219 genes. One particular gene, the VPS13A gene that encodes chorein, was found to have two nonsynonymous substitutions; the significance of this finding is that chorein was recently characterized as a key regulator of the secretion and aggregation of blood platelet and showed "marked" allelic differences between the two populations, with the authors "hypothesiz[ing] that the VPS13A missense mutations might contribute to reducing the risk of thrombosis by modulating platelet counts and blood viscosity in southern Chinese pigs in hot environments."
Gene Ontology (GO) analysis identified a significant over-representation in these 219 genes of genes involved in biological processes that contribute to the maintenance of thermostatic status during heat or cold stress. These processes were related to hair cell differentiation (ATOH1, JAG1 and RAC1) and hair follicle maturation (BARX2 and TBC1D); forebrain neuron differentiation (DLX1, DLX2, RAC1, ROBO1 and SALL1); kidney development (BMP4, BMP7, MYC, SALL1, SPRY1 and KLHL3); energy metabolism and blood circulation, including artery development (BMP4, CITED2 and JAG1) and embryonic heart tube development (CITED2, INVS, RYR2, SUFU and TBC1D8), among others.
Also of interest was the large sweep region (~14 Mb) detected on the X chromosome identified by LSBL analyses as having suppressed heterozygosity/low recombination rate and complete linkage disequilibrium. This region was found to contain a total of 84,373 LSBL outlier SNPs over the 14 Mb span, with 74,515 SNPs with "extreme differences in allele frequencies" between the northern and southern pig populations. Interestingly, all northern pigs had the northern pig haplotype, including ones that were otherwise phenotypically diverse. This sweep region was embedded in a larger region of the X chromosome (48 Mb) that was found to have a low recombination rate. There was no evidence detected of structural variation (such as one or more inversions that would make meiotic pairing unlikely or impossible and thus suppress recombination frequency) in this region. The region comprises the X chromosome centromere and reduced GC content, both of which had been associated with reduced recombination in other species. Unique to the pigs in this study was a greater extent of repeat sequences in this region, specifically enriched for 6 kb poly(T) sequences. Haplotype analysis showed "three major groups of haplotypes in the 48-Mb region: one in European pigs (domestic breeds and wild boars) and northern Chinese wild boars; a highly differentiated one in southern Chinese pigs (domestic breeds and wild boars) and a third haplogroup in northern Chinese domestic pigs, which appeared to be a recombinant between the other two haplogroups."
The authors concluded that this sweep was caused by natural selection, in part because there was a strong reduction in heterozygosity (10-15 fold reduction in nucleotide variability, compared with two-fold reductions between different Sus species) in this region in both northern and southern pigs, despite the fact that the two groups were themselves highly divergent. The age of this sweep was determined by these authors to be ~0.13 million years, around the time of the last glaciation event, perhaps reflecting population bottlenecks at that time.
Another unique finding in this study was evidence for ancient interspecies introgression being involved in the establishment of the sweep, with divergence between the two populations happening ~8.5 million years ago (alternatively the authors hypothesized that two different haplotypes arose in two subpopulations of a species ancestral to S. scrofa). As explained in the report:
"To explore the introgression hypothesis, we further analyzed whole-genome sequence data for one Phacochoerus species (African warthog; P. africanus) and four Sus species, including bearded pig (Sus barbatus), Celebes warty pig (Sus celebensis), Java warty pig (Sus verrucosus) and Visayan warty pig (Sus cebifrons)," finding the northern haplotype to be distinct from all other Sus haplotypes, and that further analysis suggested that "the northern haplotype was likely introgressed from a (possibly) extinct Sus species and may have spread through the northern Chinese population as well as the European pig population, before the split with wild boars ~1.2 million years ago."
Alternatively this haplotype:
[M]ight have been caused by gene flow between wild boars of northern China and Europe during the Pleistocene era after their divergence, a hypothesis supported by an excess of derived similarity between European and northern Chinese wild boars on the autosomes as reported previously.
The authors assert that this is the first report showing genetic adaptation to local climate conditions, and that "the adaptive haplotype in the northern Chinese populations was likely introduced from another divergent Sus species, providing the first evidence, to our knowledge, that inter-species introgression has driven adaptation in a mammal." They also acknowledged their good fortune by noting that "[i]f the introgressed segment had not fallen in a region with such a strong reduction in recombination rate, we would likely never have detected the signal of introgression as introgression fragments in other systems degenerate quickly, owing to recombination."
* The researchers were from Key Laboratory for Animal Biotechnology of Jiangxi Province and the Ministry of Agriculture of China, Jiangxi Agricultural University, Nanchang, China; GI-Tech, BGI-Shenzhen, Shenzhen, China; Department of Integrative Biology, University of California, Berkeley, Berkeley, California, USA.
