Exploring the genetic features and signatures of selection in South China indigenous pigs

Guangdong Provincial Key Lab of Agro-Animal Genomics and Molecular Breeding/National Engineering Research Center for Breeding Swine Industry/College of Animal Science,South China Agricultural University,Guangzhou 510642,P.R.China

Abstract To explore the genetic features and signatures of selection in indigenous pigs from South China and Duroc pigs,259 pigs from six populations were genotyped using single-nucleotide polymorphism (SNP) BeadChips.Principal component analysis (PCA),effective population size (Ne),linkage disequilibrium (LD),and signatures of selection were explored and investigated among the six pig populations.The results showed the Ne of five South China indigenous pig populations has been decreasing rapidly since 100 generations ago.The LD between pairwise SNP distance at 100 kb ranged from 0.16 to 0.20 for the five indigenous pig populations,while it was 0.32 for the Duroc population.However,the LD of all six pig populations showed the opposite order at long distances (>5 Mb).Furthermore,15 potential signatures of selection associated with meat quality and age at puberty were exclusively detected in South China indigenous pigs,while eight potential signatures of selection associated with growth traits were detected in Duroc pigs.Our work provides valuable insights for the utilization and conservation of South China indigenous pigs.

Keywords:pigs,population structure,effective population size,SNP

1.Introduction

Animal domestication was a milestone of human evolution that led to the emergence of farming during the Neolithic period (Vigne 2011).During domestication,animals adapt to the environment and undergo substantial morphological,behavioral,and physiological changes (Price 1999).Today,fast-developing genomic technologies and tools enable scientists to resolve the domestication of major livestock species (Troyet al.2001;Rubinet al.2010;Groenenet al.2012).

Evidence indicates that pigs (Sus scrofa) were domesticated from wild boars in Asia and Europe 8 000–10 000 years ago (Giuffraet al.2000;Cucchiet al.2011;Groenenet al.2012).From the viewpoint of culture in contemporary society,the meat quality of most Chinese indigenous pigs has received more attention,while the meat quantity is intensively selected in influential commercial breeds (Wanget al.2017).The different breeding goals lead to different selection directions,which may have contributed to the observed differences between Chinese indigenous and commercial pig breeds.

Chinese indigenous pigs are known for high fertility,excellent stress resistance,and superior pork quality.People in South China like to eat roast suckling pigs,and as a result,South China indigenous pig breeds are reputed for their excellent pork quality and early maturity after terms of selection.Modern commercial breeds,which undergo a period of artificial selection,are known for their fast growth rate,high dressing percentage,and high lean meat percentage,compared with Chinese indigenous pig breeds.For instance,Duroc pigs are known for high feed conversion and growth rate (Briggs and Briggs 1969).

Liangguangxiaohua (LGXH),Hainan,and Wuzhishan pigs originated in South China,while Duroc pigs are Europeantype domestic pigs and developed mainly in the USA (Giuffraet al.2000).Luchuan and Guangdongxiaoerhua (XE) pigs are subpopulations of LGXH pigs (NCAGRC 2011),and Tunchang and Ding’an pigs are subpopulations of Hainan pigs(NCAGRC 2011).The different selection directions of South China indigenous pigs and Duroc pigs may have resulted in different appearances and characteristics.As we had previously detected some potential signatures of selection in Hainan pig genomes compared with Baoshan and Saba pigs (Diaoet al.2019a),a total of 347 potential signatures of selection associated with meat quality were detected in a comparison of indigenous pigs from Guangdong Province and Guangxi Zhuang Autonomous Region,western commercial pig breeds,and European and Asian wild pig populations(Diaoet al.2019b).Another study based on mitochondrial DNA showed the Berkshire and Yorkshire pigs had significant Asian matrilineal footprints from South China indigenous pigs(Wanget al.2019).

The geographic isolation between Hainan indigenous pigs and LGXH pigs over the long term may have resulted in genetic isolation.However,the different genetic characteristics among Hainan indigenous pigs and LGXH pigs remain unclear.Therefore,we hypothesized that the genetic features and signatures of selection in Hainan indigenous pigs and LGXH pigs could be uncovered through single-nucleotide polymorphism (SNP) BeadChips.Hence,the aim of this study was to explore the genetic features and signatures of selection in Hainan indigenous pigs and LGXH pigs.Our results will be beneficial to the conservation and utilization of these South China indigenous pigs.

2.Materials and methods

2.1.Sample collection

Three indigenous pig breeds,LGXH,Wuzhishan,and Hainan pigs,and one western breed,Duroc pigs,were used in this study.Duroc pigs were used as the control group to explore the genetic features and signatures of selection in South China indigenous pigs.A total of five populations from South China were used for sample collection:XE pigs (a subpopulation of LGXH pigs),Luchuan pigs (a subpopulation of LGXH pigs),Ding’an pigs (a subpopulation of Hainan pigs),Tunchang pigs (a subpopulation of Hainan pigs),and Wuzhishan pigs.To obtain reliable samples,Luchuan pigs,Tunchang pigs and Wuzhishan pigs were collected from the national reservation farms of China (Yulin,Guangxi Zhuang Autonomous Region;Tunchang,Hainan Province;and Haikou,Hainan Province),while LGXH pigs and Duroc pigs were collected from breeding herds in farms of two companies (Zhanjiang,Guangdong Province and Fuqing,Fujian Province),and Ding’an pigs were collected from two pig farms located in Ding’an,Hainan Province.Since the population sizes and structures were varied and limited for indigenous pigs,the individuals for sample collection were selected with the goal of maximizing the genetic representativeness within each population according to available records.

Ear tissues of the pigs were collected for genotyping in consideration of animal welfare.More details of the sampling parameters are shown in Table 1.

2.2.Genotyping and data quality control

The genomic DNA from ear tissue was extracted for all individuals using the E.Z.N.A.®Tissue DNA Kit (D3396-02,Omega Bio-tek,Norcross,GA,USA) with a concentration>50 ng μL–1.All Chinese indigenous pigs were genotyped with the GeneSeek-Neogen PorcineSNP80 BeadChip,which contains 68 516 SNPs.The Duroc pigs were genotyped with the GeneSeek Genomic Profiler (GGP)Porcine BeadChip,which contains 50 697 SNPs.

Quality control of the genotypic data was conducted with the following criteria:(1) retaining the mutual SNP locus between the two SNP BeadChips;(2) retaining the SNPs located on autosomes;(3) filtering out individuals with a call rate below 0.90;(4) removing the SNPs with a call rate less than 0.90;and (5) removing the SNPs with a minor allele frequency (MAF) below 0.01.The data quality control was conductedviaPLINK Software (Changet al.2015).

2.3.Phylogenetic tree

A phylogenetic tree was constructed in this study to filter the outer individuals of a given pig population.PLINK Software(Changet al.2015) was used to calculate pairwise identity by state (IBS) among individuals within/between populations.Dstis described as the average proportion of alleles shared among all individuals,and the genetic distance between pairs of individuals was denoted as 1–Dst.Then,MEGASoftware (version 7.0) (Kumaret al.2016) was used to construct the phylogenetic tree based on genetic distances.

Table 1 Sample sizes and sites of the pig populations

2.4.Population structure analysis

Principal component analysis (PCA) and genetic structure analysis were carried out to further investigate the relationships between the individuals and populations in this study.To mitigate the effects of linkage disequilibrium(LD) and reduce the running time,SNPs with pairwise LD(r2>0.5) were filtered out in this step through PLINK Software(Changet al.2015) with the command:--indep-pairwise 50 5 0.5.The remaining SNPs were implemented for PCA and genetic structure analysis.

PCA was carried out with GCTA Software (version 1.91.1)(Yanget al.2011),while STRUCTURE Software (Falushet al.2003) was implemented to infer the most probable number of ancestral populations (K) of individuals,with K varying from 2 to 6.R Software (version 3.5.2) (R Core Team 2017) was used to visualize the results of PCA and genetic structure analysis.

2.5.Linkage disequilibrium and effective population size analysis

LD,defined as genotype correlation coefficient (r2),was calculated within each population usingSNePSoftware(Barbatoet al.2015).Ther2was calculated with the following formula:,whereXiandYiare the genotypes at first and second locus of individuali,andandare the mean genotype frequencies for the first and second locus,respectively.All SNPs were divided into 100 groups according to the physical distances between pairwise SNPs within 20 Mb.As a result,pairwise SNPs with a physical distance from 0 to 0.20 Mb were the first group,pairwise SNPs with a distance from 0.20 to 0.40 Mb were the second group,and so on.Then,the averages of physical distance andr2values in each group were calculated.A nonlinear model was used to fit the relationship betweenr2and physical distance:,where,LDrepresentsr2in the previous procedure;xrepresents the physical distance of SNPs;a,b,canddare the parameters of this model;andeis the random residual.

The effective population size(Ne) of each population was based on the LD,with the following calculating formula:(Sved 1971),whereNe(t)represents the ancestralNeoftgenerations ago,kequals 4,mrepresents the linkage map in Morgans estimated by the equation(Sved and Feldman 1973),distis the physical distance of pairwise SNPs,r2is estimated by the LD decline model,andαequals 1.R Software (version 3.5.2)(R Core Team 2017) was used to visualize the results of LD andNein each population.

2.6.Signatures of selection detection

Based on the genetic structures revealed by the previous steps and their geographical distribution,the six populations fall into a 3-group contrasting model.The first group contains Hainan indigenous pigs,including Ding’an,Tunchang,and Wuzhishan pigs;the second group contains LGXH pigs,including XE and Luchuan pigs;and the last group is Duroc pigs.

Cross population extended haplotype homozygosityCross population extended haplotype homozygosity (XP-EHH)was used to detect signatures of selection in this study.The XP-EHHscoreswas calculated with the following formula:,whereIAis the value integrated with respect to genetic distance for population A,andIBis defined analogously for population B (Sabetiet al.2007).A positive value suggests selection in population A;similarly,a negative value suggests selection in population B.XP-EHH Software(Pickrellet al.2009) was used to calculate XP-EHHscores.The top 1% of SNPs were treated as significant SNPs detected by XP-EHH (Liuet al.2018),hence the 0.50% right-tail/left-tail of XP-EHHscoreswere treated as the significant SNPs detected in population A/B.Before calculating XP-EHH scores,fastPHASE Software (Scheet and Stephens 2006)was used to infer haplotypes with the parameters:-KL10-KU30 -Ki5.

Fixation indexThe Fixation index (FST) statistic (Weir and Cockerham 1984) was calculated with the genepop R package (Rousset 2008),using the following formula:,whereMSPrepresents the observed mean square errors for loci between populations,MSGdenotes the observed mean square errors for loci within populations,andncis the average sample size across samples,which also incorporates and corrects for the variance in sample sizes over the population.The top 1%of SNPs were treated as significant SNPs detected byFST(Liuet al.2018).

2.7.Bioinformatics annotation

The significant SNPs detected simultaneously by the two methods extending 200 kb upstream and downstream were treated as potential selection regions in this study,as in Liuet al.(2018).Genome annotations were based onSus scrofagenome 10.2 (http://may2017.archive.ensembl.org).Genes overlapping with potential selection regions were treated as candidate genes.The Animal Quantitative Trait Loci (QTL) Database (Huet al.2018) was used to annotate the potential functions of the detected selection regions.QTLs overlapping with the potential selection regions were treated as functional annotations for potential selection regions.

2.8.Enrichment analysis

To further explore the biological functions of candidate genes overlapping with potential selection regions,the Kyoto Encyclopedia of Genes and Genomes (KEGG)Pathway Database (Kanehisa and Goto 1999) and Gene Ontology (GO) Database (Ashburneret al.2000) were used for enrichment analysis based on candidate genesviathe Database for Annotation,Visualization,and Integrated Discovery (DAVID) version 6.8 (Huanget al.2009;Weiet al.2009).TheP-value for the KEGG pathways and GO terms less than 0.05 was treated as significant.

3.Results

3.1.Data quality control

A total of 35 494 SNPs were identified as the mutual SNPs located on autosomes between two SNP BeadChips.One individual was filtered out because of an individual call rate below 0.90.In addition,803 SNPs were removed because call rates less than 0.90,and 1 719 SNPs were filtered out due to MAF values below 0.01.Finally,258 individuals and 32 972 SNPs passed the data quality control criteria.

3.2.Phylogenetic tree

A phylogenetic tree was constructed based on 32 972 SNPs(Fig.1).All individuals were clustered together by their breeds,which indicated the individuals reflected genetic representativeness within the breeds.The phylogenetic tree was divided into two clusters,South China indigenous pig populations and Duroc pig.In the South China indigenous cluster,Luchuan and XE pigs clustered closely,and Ding’an and Tunchang pigs clustered closely,while Wuzhishan pigs clustered alone;moreover,the cluster of Wuzhishan pigs was located between Hainan and LGXH pigs.More details of genetic distances between individuals are shown in Appendix A.The results of the phylogenetic tree were identical to the population constituents of individuals.

3.3.Population structure analysis

A subset of 32 972 SNPs (20 217 LD-pruned SNPs) were selected to conduct PCA and genetic structure analysis.The results of PCA are shown in Fig.2.The top three principal components contributed 13.85,6.27 and 3.05%of the genetic variance,respectively.Principal component(pc) 1 divided individuals into two groups,South China indigenous pig populations separated from Duroc pigs.The discrimination of Hainan indigenous pigs from LGXH pigs was clear on pc2.The pc3 separated Wuzhishan pigs from other South China indigenous pigs.The top three pcs divided all individuals into four groups,which was consistent with the phylogenetic tree.

The results of genetic structure analysis are shown in Fig.3,and the likelihood of each K-value is shown in Appendix B.When K=2,South China indigenous pig populations were separated from Duroc pigs.When K=3,five South China indigenous pig populations were divided into two groups:LGXH pigs and Hainan indigenous pigs.The results of the genetic structure analysis of the six pig populations showed that XE and Tunchang pigs had a fraction of the same lineages with Duroc pigs many years ago when K=4 and 5.When K=6,Duroc pigs were divided into two parts,while Luchuan pigs and XE pigs were particularly difficult to distinguish.As a result,the division of all individuals in this study into five clusters was appropriate.Moreover,Ding’an and Wuzhishan pigs experienced introgression from other ambiguous pig breeds.The genetic structure of Ding’an pigs was similar to that of Tunchang pigs.

Fig.1 Phylogenetic tree of the six pig populations.DRC,Duroc pigs;LC,Luchuan pigs;XE,Guangdongxiaoerhua pigs;WZS,Wuzhishan pigs;DA,Ding’an pigs;TC,Tunchang pigs.

3.4.Linkage disequilibrium and effective population size analysis

The LD of each pairwise SNPs was calculated for the six populations (Fig.4-A).The model-fitting degrees of all populations reached 0.97 (Appendix C).The degree of LD decreased with increased SNP marker distance.In five South China indigenous pig populations,the extent of LD between pairwise SNP distance at 100 kb for XE pigs was 0.16,while it was 0.20 for Ding’an pigs.Tunchang and Wuzhishan pigs showed a similar declining tendency when the SNP distance was below 5 Mb.In comparison,LD of Duroc pigs declined rapidly from 0.32 to 0.03.

Based on the LD results of the six pig populations,historicalNewas estimated from three to 100 generations ago (Fig.4-B).DecreasedNewas observed for all six populations from 100 generations ago to the present.About 40 generations ago,theNeof each population followed the descending order of XE pigs,Luchuan pigs,Wuzhishan pigs,Tunchang pigs,Ding’an pigs,and Duroc pigs.More details ofNeare shown in Appendix D.Taken together,theNeof the five South China indigenous pig populations ranged from 125.27 to 245.53 at 40 generations ago,while it was 96.95 for Duroc pigs.But theNeof the six pig populations showed the opposite order in recent decades (Appendix D).

3.5.Signatures of selection detection

Hainan indigenous pigs/LGXH pigsThe five South China indigenous pig populations were used to detect signatures of selection.Hainan indigenous pigs (Ding’an,Tunchang,and Wuzhishan) were used as an observation group,and LGXH pigs (XE and Luchuan) as a reference group.

Fig.2 Principal component analysis (PCA) results of the six pig populations.DRC,Duroc pigs;XE,Guangdongxiaoerhua pigs;TC,Tunchang pigs;LC,Luchuan pigs;WZS,Wuzhishan pigs;DA,Ding’an pigs;pc,principal component.

Fig.3 Genetic structure results of the six pig populations.DRC,Duroc pigs;LC,Luchuan pigs;XE,Guangdongxiaoerhua pigs;DA,Ding’an pigs;TC,Tunchang pigs;WZS,Wuzhishan pigs.

The results of signatures of selection detection are shown in Fig.5.XP-EHH scores ranged from–1.5617 to 0.8135 with an average value of–0.2803 (Appendix E).A total of 165 significant SNPs (XP-EHH scores>0.3957) were detected by XP-EHH in Hainan indigenous pigs (Fig.5-A),while 165 significant SNPs (XP-EHH scores<–1.1627)were detected in Duroc pigs.FSTranged from–0.0174 to 0.6736 with an average value of 0.0847 (Appendix E).In total,322 significant SNPs were detected byFST(Fig.5-B).Two significant SNPs,located onSus scrofachromosome(SSC) 13 and SSC18,were simultaneously detected by the two methods in Hainan indigenous pigs,while 13 significant SNPs were simultaneously detected by the two methods in LGXH pigs (Table 2;Fig.5).

Fig.4 Population genetic properties of the six pig populations.A,linkage disequilibrium results.B,effective population size estimation.DRC,Duroc pigs;LC,Luchuan pigs;XE,Guangdongxiaoerhua pigs;DA,Ding’an pigs;TC,Tunchang pigs;WZS,Wuzhishan pigs.

Fig.5 Genome-wide distribution of signatures of selection in Hainan indigenous pigs/Liangguangxiaohua pigs.A,cross population extended haplotype homozygosity (XP-EHH) scores across all autosomes.B,FSTacross all autosomes.Diamonds represent significant SNPs detected simultaneously by the two methods.Red line shows the threshold value.

The two potential selection regions detected in Hainan indigenous pigs enriched three candidate genes and 33 QTLs (Table 2).The QTLs overlapping with the two potential selection regions were associated withActinobacillus pleuropneumoniaesusceptibility,age at puberty,and so on(Table 2;Appendix F).

Meanwhile,a total of 35 candidate genes were enriched in 13 potential selection regions in LGXH pigs.Moreover,GO:0006355～regulation of transcription,DNA-templated(candidate genes:HINT1,MYB,HMGA1,YBX1;P-value:0.0342),and GO:0006284～base-excision repair (candidate genes:HMGA1,SMUG1;P-value:0.0455) were enriched in this study.In total,423 QTLs overlapping with 13 potential selection regions were associated with backfat (average backfat thickness,backfat at last rib,backfat at tenth rib,and so on),meat color (meat color a*,meat color b*,meat color L*,and so on),etc.(Table 2;Appendix F).

Hainan indigenous pigs/Duroc pigsHainan indigenous pigs and Duroc pigs were used to detect signatures of selection,with Hainan indigenous pigs (Ding’an,Tunchang,and Wuzhishan) used as an observation group and Duroc pigs used as a reference group.The results of signatures of selection detection in Hainan indigenous pigs and Duroc pigs are shown in Appendix G.

XP-EHHscores ranged from–3.1389 to 1.1419 with an average value of–0.6795 (Appendix H).A total of 165 significant SNPs(XP-EHH scores >0.7112) were detected by XP-EHH in Hainan indigenous pigs (Appendix G),while 165 significant SNPs (XP-EHH scores<–2.2621) were detected in Duroc pigs.FSTranged from–0.0137 to 0.8337 with an average value of 0.1612 (Appendix H).A total of 330 significant SNPs were detected byFST(Appendix G).In summary,a total of six significant SNPs were detected simultaneously by the two methods in Duroc pigs,located in SSC1,SSC2,SSC4,and SSC8 (Table 3;Appendix G).No significant SNPs were detected simultaneously by the two methods in Hainan indigenous pigs.

A total of 13 candidate genes were enriched in six potential selection regions (Table 3).No KEGG pathway or GO terms were enriched with these candidate genes.In total,277 QTLs were overlapping with six potential selection regions (Table 3;Appendix I).The QTLs overlapping with potential selection regions were associated with average daily gain,carcass weight (3,17,20,30 and 46 weeks),and so on.It is worth noting that QTLs associated with average daily gain were overlapping with six potential selection regions,with 37 report counts.Moreover,three potential selection regions located on SSC8 in this study were overlapping with days to 100 kg QTL,with 42 report counts.

LGXH pigs/Duroc pigsLGXH and Duroc pigs were used to detect signatures of selection.LGXH pigs(XE and Luchuan) were used as an observation group,and Duroc pigs were used as a reference group.The results of signatures of selection detection in LGXH and Duroc pigs are shown in Appendix J.

XP-EHHscores ranged from–2.9330 to 1.2656 with an average value of–0.4881 (Appendix K).A total of 165 significant SNPs(XP-EHH scores>0.9052) were detected by XP-EHH in LGXH pigs(Appendix J),while 165 significant SNPs (XP-EHH scores<–2.0387)were detected in Duroc pigs.FSTranged from–0.0141 to 0.9779 with an average value of 0.1757 (Appendix K).A total of 322 significant SNPs were detected byFST(Appendix J).Importantly,two significant SNPs were detected simultaneously by the two methods in Duroc pigs,both located on SSC14 (Table 4;Appendix J).Similarly,no significant SNPs were detected simultaneously by the two methods in LGXH pigs.

Only one candidate gene was enriched in two potential selection regions (Table 4).A total of 105 QTLs were overlapping with two potential selection regions (Table 4;Appendix L).The QTLs overlapping with potential selection regions were associated with aspartate aminotransferase activity,fat androstenone level,adipocyte diameter,and so on.

4.Discussion

This study explored the genetic features and signatures of selection between Hainan indigenous pigs and LGXH pigs.The results showed the five South China indigenous pig populations were divided into two clusters due to the geographic isolation between Hainan indigenous pigs and LGXH pigs,and theNein recent decades was decreasing rapidly for all South China indigenous pig populations.Moreover,two and 13 potential signatures of selection were detected in Hainan indigenous pig populations and LGXH pigs,respectively,while eight were detected in Duroc pig genomes.We indeed explored the genetic features and signatures of selection in Hainan indigenous pigs and LGXH pigs through SNP BeadChips.Moreover,our study provides insights into the conservation and utilization of these South China indigenous pigs.

We found that South China indigenous pig populations and Duroc pigs originated from different pigs,which was in line with previous study (Larsonet al.2005).The phylogenetic tree divided the individuals into two clusters:Duroc pigs and South China indigenous pig populations (Fig.1).Among the South China indigenous pigs,individuals were clustered by population.Ding’an,Tunchang,and Wuzhishan pigs clustered closely,and XE and Luchuan pigs clustered closely,illustrating that different genetic variations may exist in pig populations as a result of the geographical isolation of Guangdong and Hainan provinces.In addition,as shown in Fig.1,Ding’an and Tunchang pigs were mixing,and this occurrence may be caused by the origin of the pigs.Both pig populations were the subpopulations of Hainan pigs,and the sample sites of the two pig populations were adjoining which may result in gene flow in recent generations of the two pig populations,and PCA showed similar results to the phylogenetic tree.Wuzhishan pigs clustered between XE,Luchuan pigs and Hainan pigs (Fig.2),and Wuzhishan pigs were close to Hainan pig populations.Our results reflected the origin of Wuzhishan pigs (NCAGRC 2011).Moreover,the genetic structure results showed that when K=2,all individuals were divided into two groups;the Chinese indigenous pig populations except for Luchuan pigs only shared a small proportion of lineage with Duroc pigs.When K=3,pigs from Hainan province share a major proportion of lineage with other ambiguous pig populations;thus,it was supposed that pigs from Hainan province might have hybridized with other pigs,but this supposition requires more verification.When K=5,we also found that four South China indigenous pig populations except for Luchuan pigs were involved in the lineage with Duroc pigs,and this result might mean that these four pig populations had been hybridized with Duroc pigs.Ding’an and Tunchang pigs showed similar genetic structures in this study.This result was consistent with cultivar classification of Chinese indigenous pigs,in which Ding’an and Tunchang pigs are subpopulations of Hainan pigs.The genetic structure results of Luchuan and Wuzhishan pigs in this study were similar to the results of Xuet al.(2019),indicating the reliability of the results in this study.

The level of utilization of pig populations may affect theirNeand LD.The LD between pairwise SNP distance at 100 kb ranged from 0.16 to 0.20 for the five indigenous pig populations,while it was 0.32 for the Duroc populations.But the LD of the six pig populations showed the opposite order at long distances (>5 Mb).However,other studies based on other Chinese indigenous pig breeds and Duroc pigs showed that the LD degree of Duroc pigs was higher than those of Jiangquhai pigs,Litang pigs,Rongchang pigs and others (Wanget al.2018;Xuet al.2019).TheNeof the five South China indigenous pig populations in this study ranged from 125.27 to 245.53 at 40 generations ago,while it was 96.95 for Duroc pigs.But theNeof the six pig populations showed the opposite order in recent decades.It is possible that Duroc pigs have undergone intensive selection in recent years,but not the indigenous pig populations.Commercial pig breeds that were largely brought into and fed in China based on consumer demand had high carcass lean meat percentages a few years ago,which resulted in theNeof the five South China indigenous pig populations declining rapidly in recent decades.As a result,individuals of those five south China indigenous pig populations might inbreed,which may be the reason why their LD degree with long SNP distances was high.Collectively,these results implied that the five indigenous pig populations were subjected to less selection pressure than Duroc pigs.

TheNeof the other six Chinese indigenous breeds in the Taihu Lake region declined rapidly recent decades (Zhaoet al.2019),which showed a similar trend as the five South China indigenous pig populations in this study.Together,these results indicate the urgent need for the conservation of Chinese indigenous pigs.

The different eating habits of local populations might influence the selection direction of pigs.People in Guangdong and Hainan provinces may have different eating habits,resulting in differences in the selection directions among the five South China indigenous pig populations.Two potential signatures of selection were detected in Hainan indigenous pigs (Ding’an,Tunchang,and Wuzhishan),while 13 potential signatures of selection were detected in LGXH pigs.The two potential signatures were overlapping with QTLs associated with age at puberty and other traits.Hainan indigenous pigs are known to have precocious puberty compared with other Chinese indigenous pigs:Hainan boars reach sexual maturity at three months old and sows reach sexual maturity at three or four months old,and Wuzhishan pigs reach sexual maturity and are in estrus at 61 days old (NCAGRC 2011).The candidate geneIGFBP1,enriched in Hainan indigenous pigs,was reported to contribute to insulin resistance and impaired metabolic control during puberty of adolescents with insulin-dependent diabetes mellitus (Riihimaaet al.2002).This may imply thatIGFBP1is related to the sexual maturity of Hainan indigenous pigs.

QTLs overlapping with 13 potential selection regions detected in LGXH pigs were associated with meat quality traits,such as:backfat (average backfat thickness,backfat at last rib,backfat at tenth rib,and so on),meat color (meat color a*,meat color b*,meat color L*,and so on),as previously described.These results are in accordance with the good meat quality of LGXH pigs,as previously reported (Zhanget al.2012).In addition,two potential selection regions (SSC7 34.67–36.07 Mb and SSC7 36.00–36.40 Mb) overlapping with significant SNPs associated with abdominal fat were detected by Zhanget al.(2016),which may be related to the percentage of abdominal fat in LGXH pigs.

The potential selection regions detected in Duroc pigs were associated with growth rate.A total of six signatures of selection were detected in the genome of Duroc pigs compared with indigenous pigs from Hainan province(Hainan and Wuzhishan pigs).These potential selection regions were overlapping with QTLs associated with average daily gain and carcass weight (3,17,20,30 and 46 weeks) as previously described.These results are in accordance with the characteristics of Duroc pigs,which show good growth and feed conversion efficiency (Briggs and Briggs 1969).

In this study,no potential selection regions were detected in South China indigenous pigs,with Duroc pigs used as the reference group.There are three possible reasons for this.(1) The destiny of SNP markers.Two SNP BeadChips were used to genotype all individuals in this study;in addition,the gender compositions of the populations were different,so that the mutual SNPs of the two SNP BeadChips and SNPs located on autosomes were used to do quality control.In total,32 972 SNPs passed quality control,and the mean marker distance reached 74.33 kb.(2) The detection power of the two methods.The detection power of XP-EHH is higher than that ofFST,as Maet al.(2015) reported.However,the signatures of selection detected byFSTshowed better agreement with the results of genome-wide association study (GWAS) (Kimet al.2015).Therefore,GWAS could be used to enhance the accuracy of the signatures of selection detection.(3)Different selection pressures the pigs had undergone.Duroc pigs have undergone more intensive selection than South China indigenous pigs.As a result of the similar selection pressures which South China indigenous pigs have undergone,potential signatures of selection were detected in Hainan indigenous pigs when LGXH pigs were used as the reference group.

5.Conclusion

The rapid decrease of effective population size in recent decades indicates the urgent need for the conservaiton of South China indigenous pigs.Population structure analysis and signatures of selection uncover the potential differential selection directions of different South China indigenous pig populations.This study not only provides new insights into the origins,historical population sizes,and selection of South China indigenous pigs,but it also provides useful information for future use regarding these indigenous pigs.

Acknowledgements

This research was supported by the National Key R&D Program of China (2018YFD0501200),the earmarked fund for China Agriculture Research System (CARS-35)and the Key R&D Program of Guangdong Province,China(2018B020203003).

Declaration of competing interest

The authors declare that they have no conflict of interest.

Ethical approval

All experimental procedures were approved by the Animal Care Committee of South China Agricultural University,China (SCAU#2013-10).

Appendicesassociated with this paper are available on http://www.ChinaAgriSci.com/V2/En/appendix.htm