Genetic relationship among wild,landraces and cultivars of hazelnut (Corylus avellana) from Portugal revealed through ISSR and AFLP markers

The genus Corylus, a member of the birch family Betulaceae, includes several species that are widely distributed throughout temperate regions of the Northern Hemisphere. This study assesses the genetic diversity in 26 international cultivars and 32 accessions of Corylus avellana L. from Portugal: 13 wild genotypes and 19 landraces. The genetic relationships among the 58 hazelnuts (Corylus avellana L.) were analyzed using inter simple sequence repeat (ISSR) and amplified fragment length polymorphism (AFLP) markers. Eighteen ISSR primers and seven AFLP primer pairs generated a total of 570 unambiguous and repeatable bands, respectively, from which 541 (95.03 %) were polymorphic for both markers. Genetic similarity index values ranged from 0.239 for wild types and cultivars to 0.143 for landraces and wild types. The genetic relationships were presented as a Neighbor-Joining method dendrogram and a two-dimensional principal coordinate analysis (PCoA) plot. The Neighbor-Joining dendrogram showed three main clusters, and the PCoA analysis has shown to be congruent with the hierarchical analysis. Bayesian analysis clustered all individuals into three groups showing a good separation among wild genotypes, landraces and cultivars. The genetic diversity found on wild genotypes and Portuguese landraces may provide relevant information for the diversity conservation and it will be useful in breeding programs and to identify local selections for preservation.     

Genetic diversity of Carica papaya as revealed by AFLP markers.

Genetic relationships among Carica papaya cultivars, breeding lines, unimproved germplasm, and related species were established using amplified fragment length polymorphism (AFLP) markers. Seventy-one papaya accessions and related species were analyzed with nine EcoRI-MseI primer combinations. A total of 186 informative AFLP markers was generated and analyzed. Cluster analysis suggested limited genetic variation in papaya, with an average genetic similarity among 63 papaya accessions of 0.880. Genetic diversity among cultivars derived from the same or similar gene pools was smaller, such as Hawaiian Solo hermaphrodite cultivars and Australian dioecious cultivars with genetic similarity at 0.921 and 0.912, respectively. The results indicated that self-pollinated hermaphrodite cultivars were as variable as open-pollinated dioecious cultivars. Genetic diversity between C. papaya and six other Carica species was also evaluated. Carica papaya shared the least genetic similarity with these species, with an average genetic similarity of 0.432; the average genetic similarity among the six other species was 0.729. The results from AFLP markers provided detailed estimates of the genetic variation within and among papaya cultivars, and supported the notion that C. papaya diverged from the rest of Carica species early in the evolution of this genus.     

Assessment of genetic diversity in Azadirachta indica using AFLP markers

 Genetic diversity was estimated in 37 neem accessions from different eco-geographic regions of India and four exotic lines from Thailand using AFLP markers. Seven AFLP selective primer combinations generated a total of 422 amplification products. The average number of scorable fragments was 60 per experiment, and a high degree (69.8%) of polymorphism was obtained per assay with values ranging from 58% to 83.8%. Several rare and accession-specific bands were identified which could be effectively used to distinguish the different genotypes. Genetic relationships within the accessions were evaluated by generating a similarity matrix based on the Jaccard index. The phenetic dendrogram generated by UPGMA as well as principal correspondence analysis separated the 37 Indian genotypes from the four Thai lines. The cluster analysis indicated that neem germplasm within India constitutes a broad genetic base with the values of genetic similarity coefficient ranging from 0.74 to 0.93. Also, the Indian genotypes were more dispersed on the principal correspondence plot, indicating a wide genetic base. The four lines from Thailand, on the other hand, formed a narrow genetic base with similarity coefficients ranging from 0.88 to 0.92. The lowest genetic similarity coefficient value (0.47) was observed between an Indian and an exotic genotype. The level of genetic variation detected within the neem accessions with AFLP analysis suggests that it is an efficient marker technology for delineating genetic relationships amongst genotypes and estimating genetic diversity, thereby enabling the formulation of appropriate strategies for conservation and tree improvement programs. Received: 20 October 1998 / Accepted: 28 November 1998     

The origin of cultivated <Emphasis Type="Italic">Coffea arabica</Emphasis> L. varieties revealed by AFLP and SSR markers

Molecular markers were used to assess polymorphism between and within the genetic bases of coffee (i.e. Typica and Bourbon) spread from Yemen since the early 18th century that have given rise to most arabica cultivars grown world-wide. Eleven Coffea arabica accessions derived from the disseminated bases were evaluated by amplified fragment length polymorphism (AFLP) using 37 primer combinations and simple-sequence repeats (SSRs) produced by six microsatellites. Four cultivars growing in Yemen and 11 subspontaneous accessions collected in the primary centre of diversity of the species were included in the study in order to define their relationship with the accessions derived from the genetic bases of cultivars. One hundred and seven AFLP markers were used to calculate genetic distances and construct a dendrogram. The accessions derived from the disseminated bases were grouped separately, according to their genetic origin, and were distinguished from the subspontaneous accessions. The Yemen cultivars were classified with the Typica-derived accessions. Except for one AFLP marker, all AFLP and SSR markers present in the cultivated accessions were also detected in the subspontaneous accessions. Polymorphism among the subspontaneous accessions was much higher than among the cultivated accessions. It was very low within the genetic bases, confirming the historical documentation on their dissemination. The results enabled a discussion of the genetic diversity reductions that successively occurred during the dissemination of C. arabica from its primary centre of diversity.     

Analysis of diversity and relationships among mango cultivars using Start Codon Targeted (SCoT) markers

Genetic variation and relationships among 47 mango germplasm and 3 relative species from Guangxi province in China, were analyzed using Start Codon Targeted (SCoT) markers. Using 33 selected SCoT primers 273 bands were generated with an average of 8.27 bands per primer among the 50 accessions, of which 208 (76.19%) were polymorphic. Genetic relationships estimated using the SM similarity coefficient generated values between different pairs of accessions that varied from 0.531 to 0.923 with an average of 0.782. These coefficients were utilized to construct a dendrogram using the UPGMA. All 50 accessions were basically classified into six clusters and correspond well with their recorded pedigrees. The results will provide much more useful information for the management of germplasm and will also be useful to improve the current breeding strategies. The results also demonstrate that the SCoT marker system is useful for identification and genetic diversity analysis of mango cultivars.     

Assessment of genetic variation and differentiation of hop genotypes by microsatellite and AFLP markers.

Microsatellites have many desirable marker properties and have been increasingly used in crop plants in genetic diversity studies. Here we report on the characterisation of microsatellite markers and on their use for the determination of genetic identities and the assessment of genetic variability among accessions from a germplasm collection of hop. Thirty-two polymorphic alleles were found in the 55 diploid genotypes, with an average number of eight alleles (3.4 effective alleles) for four microsatellite loci. Calculated polymorphic information content values classified three loci as informative markers and two loci as suitable for mapping. The average observed heterozygosity was 0.7 and the common probability of identical genotypes was 3.271 x 10(-4). An additional locus, amplified by one primer pair, was confirmed by segregation analysis of two crosses. The locus discovered was heterozygous, with a null allele in the segregating population. The same range of alleles was detected in nine triploid and five tetraploid hop genotypes. Cultivar heterozygosity varied among all 69 accessions, with only one cultivar being homozygous at four loci. Microsatellite allele polymorphisms distinguished 81% of all genotypes; the same allelic profile was found mainly in clonally selected cultivars. Cultivar-specific alleles were found in some genotypes, as well as a specific distribution of alleles in geographically distinct hop germplasms. The genetic relationship among 41 hop accessions was compared on the basis of microsatellite and AFLP polymorphisms. Genetic similarity dendrograms showed low correlation between the two marker systems. The microsatellite dendrogram grouped genetically related accessions reasonably well, while the AFLP dendrogram showed good clustering of closely related accessions and, additionally, separated two geographically distinct hop germplasms. The results of microsatellite and AFLP analysis are discussed from the point of view of the applicability of the two marker systems for different aspects of germplasm evaluation.     

AFLP analysis of genetic diversity within and among Coffea arabica cultivars

Genetic diversity of Coffea arabica cultivars was estimated using amplified fragment length polymorphism (AFLP) markers. Sixty one Coffea accessions composed of six arabica cultivars, including Typica, Bourbon, Catimor, Catuai, Caturra and Mokka Hybrid, plus two diploid Coffea species, were analyzed with six EcoRI- MseI primer combinations. A total of 274 informative AFLP markers were generated and scored as binary data. These data were analyzed using cluster methods in the software package NTSYSpc. The differences among cultivars at the DNA level were small, with an average genetic similarity of 0.933. Most accessions within a cultivar formed a cluster, although deviant samples occurred in five of the six cultivars examined due to residual heterozygosity from ancestral materials. Among the six cultivars fingerprinted, the highest level of genetic diversity was found within the cultivar Catimor, with an average genetic similarity of 0.880. The lowest level was found within Caturra accessions, with an average genetic similarity of 0.993. Diversity between C. arabica and two other Coffea species, Coffea canephora and Coffea liberica, was also estimated with average genetic similarities of 0.540 and 0.413, respectively, suggesting that C. canephora is more closely related to C. arabica than is C. liberica. The genetic variation among arabica cultivars was similar to the variation within cultivars, and no cultivar-specific DNA marker was detected. Although arabica cultivars appear to have a narrow genetic base, our results show that sufficient polymorphism can be found among some arabica cultivars with a genetic similarity as low as 0.767 for genetic/QTL mapping and breeding. The assessment of genetic diversity among arabica cultivars provided the necessary information to estimate the potential for using marker-assisted breeding for coffee improvement.     

野生鸭茅种质遗传多样性的AFLP分析

本文应用扩增片段长度多态性（AFLP）技术,从64个引物对中筛选出9个,分别对37份鸭茅种质进行扩增,共得到400条清晰条带。其中多态性带336条,平均多态位点水平为84.0%,表现出较高的多态性。利用NTSYS-pc软件计算种质间的遗传距离,变化范围为0.0692～0.4214。用UPGMA方法进行聚类分析,在遗传相似系数为0.81水平上37份鸭茅种质可分为8个类群。用EIGEN方法进行主坐标分析,建立种质间相互关系的二维图。各种质在二维图中的分布与UPGMA聚类基本吻合。从分子水平分析,鸭茅遗传变异与染色体倍性和地理分布密切相关。     

Molecular Analysis of Chickpea (Cicer arietinum L) Cultivars Using AFLP and STMS Markers

Fifteen AFLP and eighteen STMS primer pairs were employed to reveal genetic diversity and relationship in twenty-one cultivars of chickpea (Cicer arietinum L). Fifteen AFLP primer pairs generated 1804 amplicons, out of which 1732 amplicons (96%) were polymorphic and 600 amplicons (&#x223C;33%) were genotype specific. Eighteen polymorphic STMS primer pairs generated 64 amplicons with an average of 3.55 amplicons per primer pair. Polymorphic information content (PIC) varied from 0.52 to 1.0 for STMS markers. The genetic similarity between cultivars varied from 0.30 to 0.85 for AFLP and 0.22 to 0.83 for STMS markers. Dendrogram constructed after combining both AFLP and STMS markers data with Bootstrap analysis, grouped all the cultivars into four clusters. Association of varietal type and flower colour was observed as cultivars E 100Ymu and Nabin (Both Desi type and pink flower) clustered together in the dendrogram.     

Molecular genetic diversity and association mapping of nut and kernel traits in Slovenian hazelnut (<Emphasis Type="Italic">Corylus avellana</Emphasis>) germplasm

European hazelnut (Corylus avellana L.), cultivated in several areas of the world including Europe, Anatolia, and the USA, is an economically important nut crop due to its high mineral, oleic acid, amino acid, and phenolic compound content and pleasant flavor. This study examined molecular genetic diversity and population structure of 54 wild accessions and 48 cultivars from the Slovenian national hazelnut collection using amplified fragment length polymorphism (AFLP) and simple sequence repeat (SSR) markers. Eleven AFLP primer combinations and 49 SSR markers yielded 532 and 504 polymorphic fragments, respectively. As expected for a wind-pollinated, self-incompatible species, levels of genetic diversity were high with cultivars and wild accessions having mean dissimilarity values of 0.50 and 0.60, respectively. In general, cultivars and wild accessions clustered separately in dendrogram, principal coordinate, and population structure analyses with regional clustering of the wild material. The accessions were also characterized for ten nut and seven kernel traits and some wild accessions were shown to have breeding potential. Morphological principal component analysis showed distinct clustering of cultivars and wild accessions. An association mapping panel composed of 64 hazelnut cultivars and wild accessions had considerable variation for the nut and kernel quality traits. Morphological and molecular data were associated to identify markers controlling the traits. In all, 49 SSR markers were significantly associated with nut and kernel traits [P < 0.0001 and LD value (r ²) = 0.15–0.50]. This work is the first use of association mapping in hazelnut and has identified molecular markers associated with important quality parameters in this important nut crop.     

Assessment of genetic diversity among and within Achillea species using amplified fragment length polymorphism (AFLP)

Amplified fragment length polymorphism (AFLP) markers were used to assess the genetic diversity of 57 Achillea accessions belonging to five species, A. millefolium, A. filipendulina, A. tenuifolia, A. santolina and A. biebersteinii. Nine AFLP primer combinations were used, which produced 301 polymorphic bands. In most species, a high level of genetic variation was detected among the genotypes. The Jaccard's similarity indices (J), based on AFLP profiles, were subjected to UPGMA cluster analysis. Application of Mantel's test for cophenetic correlation to the cluster analysis indicated the high fitness of the accessions to a group (r = 0.918). The dendrogram generated revealed five major groups corresponding to five species. The principle coordinate analysis (PCoA) data confirmed the results of the clustering. Among the species, A. teunifolia and A. santolina showed the greatest and the least genetic diversity, respectively. A. filipendulina accessions were acquired primarily from the same ecological regions of western Iran. Accessions belonging to A. biebersteinii originated from the Isfahan province and were separated from other species at the root of the dendrogram. The results of the clustering method, based on AFLP markers, corresponded closely with the geographical origins of the genotypes. The results of the present study could contribute to a better understanding and management of conservation and exploitation of the Achillea germplasm.     

感病与抗病圭亚那柱花草遗传多样性的AFLP分析

圭亚那柱花草(Stylosanthes guianensts Swartz)原产中南美洲及非洲,是一种重要的热带豆科牧草,已在我国华南热带、南亚热带地区种植并利用.由胶孢炭疽菌(Colletotrichum gloeosporioides(Penz.)Sacc.)引起的炭疽病是柱花草的主要病害.采用扩增片段长度多态性(AFLP)技术分析了42个圭亚那柱花草品系的遗传多样性,同时对其抗病性进行了接种鉴定.从96个选择性引物对中筛选出较好的4个,分别对42个圭亚耶柱花草品系进行扩增,共获得出225条带,其中多态性带215条,平均多态性水平为95.5%,表现出高度的多态性.采用NTSYS-pc软件计算了品系间的遗传相似系数,其变化范围为0.31～0.95.根据非加权成对平均数法(uPGMA)进行聚类分析,建立了42个品系的聚类树系图,以所有品系的平均遗传相似系数0.48为阈值,共分为5类.主成分分析表明:第一主成分和第二主成分对全部品系间遗传变异的贡献率分别为56.04%和6.40%,并建立了品系间相互关系的二维图,各品系在二维图中的分布与UPGMA分类相吻合.抗病性鉴定结果表明:各品系对两种典型的病原菌的抗性有差异,其中抗病品系对两种病原菌的抗病相关系数达到0.904,表明抗病品系对两种病原菌有共同抗性.此外,抗病品系在UPGMA聚类中呈随机分布.这些结果表明,AFLP技术是分析圭亚那柱花草遗传多样性的有效方法.     

Assessment of genetic variation withinBrassica campestris cultivars using amplified fragment length polymorphism and random amplification of polymorphic DNA markers

Genetic relationships were evaluated among nine cultivars ofBrassica campestris by employing random amplification of polymorphic DNA (RAPD) and amplified fragment length polymorphism (AFLP) markers. RAPDs generated a total of 125 bands using 13 decamer primers (an average of 9.6 bands per assay) of which nearly 80% were polymorphic. The per cent polymorphism ranged from 60–100%. AFLP, on the other hand generated a total of 319 markers, an average of 64 bands per assay. Of these, 213 were polymorphic in nature (66.8%). AFLP methodology detected polymorphism more efficiently than RAPD approach due to a greater number of loci assayed per reaction. Cultivar-specific bands were identified, for some cultivars using RAPD, and for most cultivars with AFLP. Genetic similarity matrix, based on Jaccard’s index detected coefficients ranging from 0.42 to 0.73 for RAPD, and from 0.48 to 0.925 for AFLPs indicating a wide genetic base. Cluster analyses using data generated by both RAPD and AFLP markers, clearly separated the yellow seeded, self-compatible cultivars from the brown seeded, self-incompatible cultivars although AFLP markers were able to group the cultivars more accurately. The higher genetic variation detected by AFLP in comparison to RAPD was also reflected in the topography of the phenetic dendrograms obtained. These results have been discussed in light of other studies and the relative efficiency of the marker systems for germplasm evaluation.     

羊草种质基因组DNA的AFLP多态性研究

羊草是禾本科牧草之王 ,在当前我国西部生态建设和草原畜牧业发展中发挥着重要作用。用AFLP方法对2 7份我国不同地区分布的羊草 (Leymuschinensis (Trin .)Tzvel)材料进行了基因组DNA多态性分析 ,8对AFLP引物组合在 2 7个不同羊草基因型中共扩增出 5 37条带 ,产生出的DNA片段大小分布在 75bp - 5 30bp之间。其中单态性带 89条 ,占 16 .6 % ,多态性带 32 9条 ,占 6 1.3%。平均每对引物组合扩增的DNA带数为 6 6 .13,总的多态性比率为 78.84%。AFLP多态信息含量PIC值分布于 0 .0 - 0 .5之间 ,平均PIC值为 0 .2 16 ,出现的PIC最大值 (0 .5 )约占AFLP标记的 8.5 % ,说明羊草基因组DNA的多态性比较丰富。以 5 37个AFLP标记为原始数据 ,根据Nei和Li的方法对 2 7份羊草材料进行遗传变异和聚类分析的结果表明 :羊草种内有高频率的遗传变异发生 ,且与地理分布和生态环境密切相关 ;2 7份羊草不同基因型被划分为四大类群 ,不同类群相互间的遗传距离相对较大 ,在树状图中表现为较远的亲缘关系。对羊草种内遗传变异发生的原因和品种的形成进行了初步讨论。     

Genetic similarity among cultivars of Phyllostachys pubescens

Phyllostachys pubescens is the most important economic bamboo species in China, which grows widely in the South of China. There are more than ten cultivars in this species but their genetic relationship still remains unknown. We used both amplified fragment length polymorphism (AFLP) and inter-simple sequence repeat (ISSR) techniques to determine genetic similarity among ten cultivars of P. pubescens and two related species. Eight hundred and twenty seven bands, in which 495 are polymorphic, were detected using 15 pairs of AFLP primers whereas total 231 bands, in which 154 bands are polymorphic, were scored using 16 ISSR primers. Statistic analysis showed that the genetic similarity matrices obtained from these two sets of molecular markers had a significant correlation (R = 0.959, P = 0.013). The dendrogram generated with AFLP and ISSR markers could clearly genetically identify ten cultivars of P. pubescens that had high similarity with genetic distances ranging from 0.023 to 0.108, and could be divided into three groups based on their genetic variation and similarity. Our results suggest that these molecular markers are useful to genetically classify cultivars or varieties of a species, particularly a bamboo species.     

Diversity and Species Relationship in Pearl Millet (Pennisetum typhoides) and Related Species

Thirteen accessions of pearl millet (Pennisetum typhoides (L) Leeke) collected from different states of India and eight wild species of the genus Pennisetum across the world were analyzed for genetic diversity using AFLP markers. A combined analysis of eight primer combinations showed 35% polymorphism among P. typhoides accessions while analysis with five primer combinations showed 99% polymorphism among the wild species. The dendrogram constructed for the P. typhoides accessions based on the UPGMA method revealed two major clusters with samples from Gujarat forming a separate cluster from the rest of the samples. Principal component analysis of the same data set revealed similar results with the first principal component accounting for 65% of the total variation. The percentage of rare and common alleles contributing to the diversity in the sample was analyzed using the Shannon Weiner diversity index. The SW index revealed that the samples from Gujarat contributed significantly to the overall diversity among the accessions. Among accessions of each geographical region, considerable variation was revealed by SW index with samples from Tamil Nadu being most polymorphic. The genetic diversity in the accessions could be utilized for future breeding work. The dendrogram constructed for the wild species revealed the extent of genetic diversity among them. Analysis with one primer combination showed P. typhoides being closer to P. mollissimum than to the other analyzed species.     

Comparative study of different molecular markers for classifying and establishing genetic relationships in Coffea canephora

The genetic variability characterization of the accessions of the germplasm collection, using molecular markers, is being applied as a complementary strategy to the traditional approaches to redefine the plant genetic resources. In this study, we compared the informativeness and efficiency of the molecular markers RAPD, AFLP and SSR in the analysis of 94 accessions of Coffea canephora germplasm held by the breeding program of the Brazilian Agricultural Research Corporation (Embrapa), Rondônia State, Brazil. For this, we considered the marker’s discriminatory power and level of polymorphism detected and also the genetic relationships and clustering (dendrogram) analysis. The RAPD marker yielded low-quality data and problems in the discrimination of some accessions, being less recommended for genetic studies of C. canephora. The SSRs had a higher level of information content and yielded high-quality data, while AFLP was the most efficient marker system because of the simultaneous detection of abundant polymorphism markers per few reactions. Our results indicate that AFLP and SSR, allies to the intrinsic characteristics of each technique, are the most suitable molecular markers for genetic studies of C. canephora. However, the choice of AFLP or SSR in the species characterization should be made in agreement with some characteristics that are discussed in this work.     

全国7省本地早橘基因组间相关性的AFLP分析

采用AFLP分子标记对全国７省的12份本地早橘资源进行遗传多样性分析。使用了28对选择性引物组合,共扩增出882个遗传位点,其中192条（21.8％）为多态性位点;以黄岩本地早橘为对照,各地的本地早橘的多态性位点数不多（3.5%～10.54%）,且来源于浙江省外的本地早橘样品的AFLP多态性位点数要多于浙江省内的,说明区域生态差异对遗传多样性有着一定的影响。对12个样品进行聚类分析结果表明,样品间的遗传距离最大为0.229,说明差异不大,引种后的变异率不高。这些结果有助于揭示其遗传变异与生态条件的关系,对柑橘品种的遗传改良提供参考意见。      

Evaluating genetic relationships between tropical maize inbred lines by means of AFLP profiling

Diversity among tropical maize inbred lines that compose breeding programs, is not well known. The lack of this information has made the arrangement of heterotic groups to be used for breeding purposes difficult. Methods of molecular analysis have been used as efficient alternatives for evaluating genetic diversity, aiming at heterotic group arrangement and acquisition of new hybrids. In this study, AFLP (amplified fragment length polymorphism) was used to investigate the genetic relationships among 96 tropical maize inbred lines from two different origins. The polymorphism level among the genotypes and the possibility of their allocation in heterotic groups were evaluated. Besides, correlations among genetic diversity and flowering time were analyzed. Nine primer combinations were used to obtain AFLP markers, producing 638 bands, 569 of which were polymorphic. Genetic similarities (GS), determined by Jaccard's similarity coefficient, varied from 0.345 to 0.891, with an average of 0.543. The dendrogram based on the GS and on the UPGMA cluster method did not separate the inbred lines in well-defined groups. Aiming at separating the lines into more accurate groups, Tocher's optimization procedure was carried out, 17 groups being identified. Association between flowering time and germplasm pools was detected. AFLP showed itself to be a robust assay, revealing a great power of detection of genetic variability in the tropical germplasm, and also demonstrated to be very useful for guiding breeding programs.     

Genetic diversity and population structure of Chrysichthys nigrodigitatus from Niger Delta based on AFLP analysis

The silver catfish, Chrysichthys nigrodigitatus is an important food fish in the Niger Delta. To understand the genetic variations and population structure in Nigeria’s Niger Delta, eighty-eight individuals collected from 7 locations were analyzed based on amplified fragment length polymorphism (AFLP) analysis. A total of 243 bands were identified using 4 AFLP primer combinations. The average gene diversity was 0.1708 ± 0.1547 and Shannon’s information index was 0.2792 ± 0.2224. The pairwise F_st values ranged from 0.133 to 0.796. The results of AMOVA analysis indicated that 19.03% of the genetic variation was contained among three groups. The gene flow estimates (Nm) demonstrated that different degrees of gene flow existed among populations ranging from 0.064 to 1.630. Two clades were recognized on the UPGMA tree. The results supported that freshwater or brackishwater habitat, limited long-distance dispersal of the adult and juveniles and physical barriers may be responsible for the current genetic structure.