Grindelia (Asteraceae). We examined 439 individuals in 10 different populations. The plants grouped broadly into a coastal and an inland set of populations. The coastal group contained plants from salt marshes and coastal bluffs, as well as a population growing in a serpentine grassland close to the coast, while the inland group contained grassland plants. No evidence for hybridization was found at the single location where adjacent populations of the two groups were sampled. In addition to differentiation along ecotypic lines, there was also a strong signal of local differentiation, with the plants grouping strongly by population. The strength of local differentiation is consistent with the extensive morphological variation observed across populations and the history of taxonomic confusion in the group. The Pacific Clade of Grindelia and other young Californian plant groups warrant additional analysis of evolutionary divergence along the steep coast-to-inland climatic gradient, which has been associated with local adaptation and ecotype formation since the classic studies of Clausen, Keck, and Hiesey.Studies of ecotypic differentiation in the California Floristic Province have contributed greatly to plant evolutionary biology since the pioneering work of Clausen, Keck, and Hiesey. The extent of gene flow and genetic differentiation across interfertile ecotypes that span major habitats in the California Floristic Province is understudied, however, and is important for understanding the prospects for local adaptation to evolve or persist in the face of potential gene flow across populations in different ecological settings. We used microsatellite data to examine local differentiation in one of these lineages, the Pacific Coast polyploid complex of the plant genus  Ecotypes or ecological races have been widely studied in order to understand the processes of adaptation and diversification Timema cristinae Vickery walking sticks associated with different chaparral shrubs (Ceanothus spinosus Nutt. and Adenostoma fasciculatum Hook. & Arn.) Gasterosteus aculeatus L. (three-spined stickleback) G. aculeatusPungitius pungitius L. (nine-spined stickleback) Ecotypes have been widely investigated in both plants and animals, with some particularly well-studied examples in animals including Eucalyptus globulus Labill. Senecio lautus G.Forst. ex Willd. complex Mimulus guttatus DC. has been extensively studied One of the best-studied patterns of ecotypic differentiation in plants is that between coastal and inland ecotypes Clarkia Pursh Collinsia Nutt. Downingia Torr. Layia Hook. & Arn. ex DC. Achillea L. Mimulus L., The California Floristic Province (CA-FP) has long been recognized as an area where many groups of plants and animals have undergone recent diversification, in part reflected by the large number of closely related, but often ecologically divergent, species that are endemic to the area Grindelia Willd. (Asteraceae: Astereae). Throughout its range in North America and South America, members of the genus occupy a wide variety of open, predominantly xeric habitats, such as grasslands, deserts, and early successional sites, as well as coastal bluffs and salt marshes. One clade of Grindelia is restricted to far western North America, from Baja California, Mexico, to British Columbia, Canada and has its center of morphological and ecological diversity in the CA-FP. This Pacific Clade was well-supported in molecular analyses of nuclear ribosomal DNA sequences, although resolution within the clade was poor due to lack of sequence divergence Grindelia by a chromosomal rearrangement One group that shows considerable ecological and morphological diversity within the CA-FP is the New World genus Grindelia, and plants from two different habitats often have striking morphological differences, it is generally very difficult to draw clear boundaries between putative taxa when they are examined across their entire ranges. This problem is reflected in the variety of taxonomic hypotheses that have been proposed for the group, ranging from treatment of the plants as a group of 16 related species Given the lack of sequence divergence among members of the Pacific Clade, the group as a whole appears to be quite young The phylogenetic difficulties are compounded by variation in ploidy. The Pacific Clade includes both diploids and tetraploids Grindelia using microsatellite data. Ten populations, with 29 to 50 individuals per population, were sampled , California State Parks , the East Bay Regional Park District , the Sacramento National Wildlife Refuge Complex , and the San Francisco Bay National Wildlife Refuge Complex . All necessary permits were obtained for the described study, which complied with all relevant regulations.All sampled populations of Pacific Between 29 and 50 individuals from each of ten populations were sampled, for a total of 439 individuals . All popGrindelia at a given site. All collection localities were separated from one another by large areas of unsuitable habitat, except for the two localities at Point Pinole (populations 220 and 221), where morphologically different populations were found within ca. 100 m of each other in grassland and saltmarsh habitats. This distance is likely within the flight distance of their bee pollinators The term population is used to refer to a group of plants growing in the same habitat in a given area. In non-wetland habitats, the boundaries of the populations were quite distinct, and samples were obtained from throughout the entire area where the plants occurred at a collection locality. In wetland habitats, the plants occurred in much more extensive stands, and we were only able to sample plants from part of the area occupied by Moore, Silviera, and Anderson 551 (JEPS), collected in the Sacramento National Wildlife Refuge, near the site of collection of population 156. Primer sequences for the variable loci are described in Sequences of microsatellite-containing loci were obtained using the protocol of DNA was extracted from fresh or silica-dried material using the Qiagen Plant Mini Kit . Samples were ground directly in AP1 extraction buffer using a mortar and pestle or ground dry using glass beads in a Mini-Bead-Beater-16 .Taq polymerase (New England Biolabs), 0.4 µM each primer, 0.6 mM dNTPs, 0.5 µg BSA, and 3 µl DNA that was diluted 1∶10 from the concentration of the originally extracted DNA. Loci 045 and 113 were amplified using AccuPower PCR PreMix  in 20 µl reactions using 0.375 µM concentration of each primer and 3 µl of DNA at 1∶10 dilution. The touchdown PCR program of Most loci were amplified with component-based PCR in 25 µl reactions with 1× ThermoPol reaction buffer , 1.5 units of Samples were run on ABI 3730×l capillary sequencing machines  at the U.C. Berkeley DNA Sequencing Facility using the GeneScan 500 ROX size standard (Applied Biosystems). Samples were scored using Peak Scanner v.1.0 software (Applied Biosystems). A subset of individuals (ca. 5%) was run twice to ensure that amplification and scoring of alleles were consistent across runs.All individuals produced between 1 and 4 score-able bands at each locus (or between 1 and 2 score-able bands in the diploid population 218), with the exception of individuals 9 and 23 from population 223 , for which locus 024 did not amplify; individuals of 4, 7, and 14 from population 222 , which produced 5 bands at locus 026; and individual 10 from the diploid population 218 (grassland), which produced 4 bands at locus 024.Microsatellite data were scored by recording the presence or absence of the alleles (phenotypic scoring), instead of by attempting to determine how many copies of each allele were present in a given individual (genotypic scoring). We chose to perform phenotypic instead of genotypic scoring because at least some individuals at each of the different loci had stutter bands or had peaks that were overloaded, so the full area of the peaks could not be accurately determined. The scored microsatellite data have been deposited in Dryad (doi:10.5061/dryad.rv524).Dice’s similarity coefficient structureK = 2, with the two intermediate populations not included. After the discriminant analysis in which the plants from seven of the ten populations were grouped according to the structure groups, the discriminant classification function was used to classify the plants from the remaining three populations into one of the existing groups. A finding that both discriminant analyses are significant would indicate not only that the populations are significantly different, but also that there is a hierarchical structure to these differences, where the larger groups are also significantly different from each other.Discriminant analyses were performed using SPSS version 20.0.0.1  to determine whether populations and groups differed significantly in terms of the principal coordinates that had corresponding eigenvalues greater than zero. Discriminant analyses were performed with the plants grouped two different ways: into populations (ten groups) and into the two groups from the FST that is calculated using allelic phenotypes. ρ is independent of both the amount of double reduction and the degree of inbreeding. It can therefore be used to compare diploids and tetraploids SPAGeDi v. 1.3a StructureStructure cannot calculate the likelihood values of the individual runs accurately when there is genotypic uncertainty K) was allowed to vary from 2–15; 20 replicate runs were performed for each value of K. Each replicate was run for 100,000 generations preceded by a burn-in period of 30,000 generations. Admixture was allowed, and allele frequencies were independent in the different populations. As the analysis sometimes had trouble converging at higher values of K, the two populations with the lowest log likelihood values were removed prior to comparing the likelihood values at the different values of K. We also considered the delta K method of Evanno et al. The first set of analyses, in which it was assumed that the genotypes were known unambiguously, was performed with data sets in which 4-allele genotypes were created by replacing the unknown alleles with one of the known alleles with equal probability. For example, if an individual had three different alleles at a given locus, the fourth allele had a one-third probability of being a repeat of any of the other three. Diploids were coded with two or four copies of each allele, depending on whether they were heterozygous or homozygous at a given locus. Four different data sets, with different random resolutions of the tetraploid genotypes, were constructed. For these analyses, it was assumed that there were no recessive alleles and no ambiguity (RECESSIVEALLELES = 0). The number of groups (K) was set to 2 or to 10, which were the optimal numbers from the previous analysis. Each replicate was run for 100,000 generations preceded by a burn-in period of 10,000 generations. Admixture was allowed, and allele frequencies were independent in the different populations.The second set of analyses, in which genotypic uncertainty was taken into account, was performed with one of the data sets from the first set of analyses. Replicate data sets were not necessary, as individuals with different numbers of the same alleles would be treated identically structure, the two individuals lacking locus 024 were coded as having missing data at that locus (ca. 0.08% missing data), while only the four shortest alleles were used for the three individuals with five alleles at locus 026. Each of the five alleles present in those three individuals was also found in other plants, so it was not clear which allele was the extra one and the choice of not using the longest allele was arbitrary.For the analyses with SPAGeDi and structure analysis with K = 2 (with only the seven populations that could be unambiguously classified included).A hierarchical analysis of molecular variance AMOVA,  was perfAcross the six loci, 134 alleles in total were found, with the number of alleles per locus ranging from 7 to 31 overall and 3 to 19 within a population . For eacThe first two principal coordinates explained 9.69% and 6.04% of the variance in the data, respectively. While these two dimensions explained less than 20% of the variance in the data, they provide the best two-dimensional representation and are shown to help visualize the data. In the plot of those two coordinates ; in addition, 98.6% of individuals  were classified in the correct population  gave the same results.In the ulations . The twostructure was set to two, the distribution of individuals across groups corresponded closely to the division of individuals along the first principal coordinates axis . In this analysis, 99.4% of individuals  were correctly classified . When the plants were divided into coastal and inland populations , 9.9% of the variance was between coastal and inland groups, 13.5% was among populations within groups, and 76.7% was within populations  was also relatively high between all pairs of populations. The relative genetic isolation of the different populations supports Strother and Wetter’s Grindelia (“facies”) correspond to local or regional differentiation. Local differentiation was also shown by the morphological differences between populations of Grindelia collected in grasslands of California’s Central Valley, which persisted when the plants were grown in a common garden Evidence for individual populations being the strongest groups was found in the Grindelia is due to few opportunities for gene exchange between populations or to selection against migrants . The idea that lack of opportunity for gene exchange could be a contributing factor is supported by the current isolation of populations of Grindelia in this part of its range. Even within areas of apparently suitable upland habitat, Grindelia is found in small patches  that are generally isolated from other patches by 1 km or more . The major exceptions to this patchy distribution are the marsh plants, which occur in linear populations extending along the banks of sloughs and the shores of San Francisco Bay. Even these large populations are presently separated from each other by areas of unsuitable habitat . However, there are also cases (including the two Point Pinole populations included in this study), where two morphologically different populations occur in slightly to entirely different habitats in close proximity to each other. In these cases, the plants are able to maintain their morphological differences , despite the opportunity to interbreed.It is unclear whether this local differentiation in In addition to grouping the plants into populations, there was evidence in the microsatellite data that the populations themselves could be grouped according to distribution and ecology, with a coastal group, an inland group, and three populations that were somewhat intermediate between the two groups. The coastal group consisted of populations growing in the salt marshes surrounding San Francisco Bay (3 populations), coastal bluffs (1), and serpentine grasslands (1). This last population was collected from Mt. Tamalpais, ca. 3 km from the coast, in an area heavily influenced by fog. The inland group consisted of two populations from non-serpentine grasslands. Although we refer to these groups as coastal and inland, increased sampling may indicate that the groups are not divided strictly along coastal and inland lines or that additional coastal or inland groups may be found.structure analysis and were split between coastal and inland groups in the discriminant analysis. Population 156 was the only population from outside of the San Francisco Bay Area, coming from approximately 175 km north, near the Sacramento River. It grew in vernal pools, areas that are filled with water during the winter and spring and completely dry in the summer and fall, when these plants flower. Thus, it is possible that genes from the coastal ecotype would be selectively advantageous at its inland locality.Three populations  were intermediate between the coastal and inland groups in the Population 217 was from a grassland that was heavily influenced by fog, although it was at some distance from the Pacific Ocean in the East Bay hills. Unlike the other grassland populations we sampled, which flowered starting in July, this population flowered from May to June.G. paludosa Greene or G.×paludosa), and subsequently hypothesized on morphological grounds to be hybrids between inland plants that grew in the Central Valley grasslands and Inner Coast Ranges (G. camporum Greene) and the coastal plants that grew in the salt marshes surrounding San Francisco Bay (G. stricta DC. var. angustifolia (A.Gray) M.A.Lane/G. humilis Hook. & Arn.) The other intermediate population, 154, grew in a brackish marsh in the Suisun Delta, where the Sacramento and San Joaquin rivers come together before they flow into San Francisco Bay. The plants from the Suisun Delta were described as a distinct taxon in the 1890s , and do not show the wide variation of morphology expected in recent, unstabilized hybrids. Their genetic intermediacy could be the signal of a hybrid origin or some lower level of gene exchange in the past.Grindelia, has been found in many other plant groups along the Pacific coast of North America. Classic biosystematic studies found coastal and inland species or coastal and inland ecotypes of species with wide geographic ranges based on morphological and ecological differences  DC. that was congruent with a geographical division between populations from the Outer Coast Ranges and Inner Coast Ranges.A division into coastal and inland entities, as shown here for CA-FP Grindelia, including common garden investigations, were not oriented around ecotype discovery G. arenicola Steyerm., G. blakei Steyerm., G. hirsutula Hook. & Arn. (in part), G. humilis, G. maritima (Greene) Steyerm., G. rubricaulis DC., and G. stricta, while inland plants were classified in G. camporum and G. hirsutula (in part) Although biosystematic studies of CA-FP Layia gaillardioides were also apparent from DNA sequence data Achillea millefolium complex Eucalyptus globulusSenecio lautusDowningia yina Applegate, molecular studies found three cryptic species along a coast-to-inland transect, although these were not suspected based on previous morphological work: D. willamettensis M. Peck occurs west of the Cascade Ranges , D. pulcherrima M. Peck occurs to the east (inland), and D. yina s.s. is localized in the Cascade Range For some plant groups, molecular data have reinforced or extended the ecological findings of past biosystematic studies, but for others they have not. While coastal and inland lineages of Mimulus guttatus into coastal and inland ecotypes has been extensively studied, both molecularly and experimentally M. guttatus differ adaptively, with the coastal plants being more salt tolerant GrindeliaThe differentiation of M. guttatus ecotypes, the coastal and inland ecotypes in CA-FP Grindelia have overlapping flowering periods, and they occasionally do occur close enough together for cross-pollination to occur, as at our site on Point Pinole. More study is needed to determine whether the lack of gene flow is due to one or multiple factor(s), such as hybrids simply not being formed  or to selection against hybrids at the seedling stage due to distinct habitat adaptations of the parents. In any case, the populations of the two plants at the site where they co-occur are just as morphologically uniform as they are in sites where the ecotypes occur alone .In contrast to the Grindelia is unclear. They could potentially represent two separate tetraploidization events. They could also have arisen from greater gene exchange within the coastal group and within the inland group than between the groups (either due to geographic circumstances or to barriers to gene exchange between groups). In the one instance where we found two populations of different habitats occurring adjacent to each other without exchanging genes, the populations were from the two different ecotypic groups, but it is unclear if similar phenomena could be observed between populations of different habitats within the coastal or inland groups.The origin of the coastal and inland groups in CA-FP Grindelia, or at least all of the tetraploids, are classified as one species, the oldest available name is G. hirsutula. If, instead, the inland and coastal groups are treated as distinct species, as could be warranted if further evidence of genetic isolation is found, the taxonomic situation is more complicated, in part because plants that have been classified as G. hirsutula are found in both groups, and the type specimen of G. hirsutula does not correspond exactly with any of the plants we have sampled thus far. If the type of G. hirsutula is found to belong to the coastal group, then the oldest available name for the inland group would likely be G. camporum. Instead, if the type of G. hirsutula is found to belong to the inland group, the oldest available name for the coastal group would likely be G. stricta.Given our limited sampling and the relatively large number of intermediate populations, it would be premature to draw any firm taxonomic conclusions. If all CA-FP Grindelia is not alone in posing such difficulties, especially among young perennial lineages that retain interfertility across populations and that span ecological gradients. Such groups represent an important frontier for the application of genomic and other methods in order to understand, communicate, and protect some of the most interesting examples of recent evolutionary change.Early differentiation of lineages in evolutionary hotspots, such as the CA-FP, poses serious challenges for resolving and classifying biodiversity. Table S1Pairwise matrix of ρ distances between populations.(CSV)Click here for additional data file.


Ostrinia includes two allopatric maize pests across Eurasia, namely the European corn borer  and the Asian corn borer . A third species, the Adzuki bean borer , occurs in sympatry with both the ECB and the ACB. The ABB mostly feeds on native dicots, which probably correspond to the ancestral host plant type for the genus Ostrinia. This situation offers the opportunity to characterize the two presumably independent adaptations or preadaptations to maize that occurred in the ECB and ACB. In the present study, we aimed at deciphering the genetic architecture of these two adaptations to maize, a monocot host plant recently introduced into Eurasia. To this end, we performed a genome scan analysis based on 684 AFLP markers in 12 populations of ECB, ACB and ABB. We detected 2 outlier AFLP loci when comparing French populations of the ECB and ABB, and 9 outliers when comparing Chinese populations of the ACB and ABB. These outliers were different in both countries, and we found no evidence of linkage disequilibrium between any two of them. These results suggest that adaptation or preadaptation to maize relies on a different genetic architecture in the ECB and ACB. However, this conclusion must be considered in light of the constraints inherent to genome scan approaches and of the intricate evolution of adaptation and reproductive isolation in the Ostrinia spp. complex.The genus  An intriguing yet unresolved question in evolutionary biology is the degree to which evolution is canalized by intrinsic developmental properties MC1R gene involved in color variation in lizards, birds, felids, mice and black bears “Rerunning the tape of life” being… somewhat difficult, an alternative approach is to study naturally occurring cases of repeated adaptations, i.e. adaptations that occurred independently as a response to similar ecological changes. For a long time, evolutionary biologists considered that a trait evolving in closely related lineages  is likely to rely on the same genetic architecture, whereas a trait evolving in already distant lineages (‘convergent evolution’) is more likely to involve different genetic architectures. However, recent reviews and discussions Nymphalini butterflies, different species feed on different plant families, and some specific associations have evolved independently several times Trirhabda, in which four independent host-plant shifts from Asteraceae to Anthemideae have been documented The evolution of food preference within the large and diverse group of phytophagous insects is an outstanding source of case studies to analyze repeated adaptations. Phytophagous insects evolve in close interaction with their host-plants, and some authors consider the use of a particular host-plant type as a homoplasic character. For example, in Ostrinia . Two Ostrinia species have been particularly studied over the last decades because they are major pests of maize (Zea mays L.): Ostrinia nubilalis Hübner sensuOstrinia furnacalis Guénée, the Asian corn borer (ACB). These two species are mostly allopatric at Eurasian scale, the ECB occurring in Europe and the ACB in Asia Ostrinia scapulalis Walker sensuArtemisia vulgaris L.: Asteraceae), hop (Humulus lupulus L.: Cannabaceae) and hemp (Cannabis sativa L.: Cannabaceae), which are all dicots and native to Eurasia, unlike maize.Here, we conducted a comparative analysis of repeated adaptation in phytophagous moths of the genus At least in France, the ECB and ABB do not only differ in host plant use. A large number of behavioral and ecological differences have indeed been documented between these species, most of which can be related either to differences in host plant use and/or to reproductive isolation: oviposition preferences for different host-plants ca. 8,700 years ago in the highlands of Mexico. It then spread across the Americas during the next millennia, and was introduced very recently (ca. 500 years ago) into Europe Maize has been domesticated Ostrinia spp. genome is not sequenced yet, we used non-targeted amplified fragment length polymorphisms (AFLPs). From a total of 684 marker loci, we identified those markers that displayed a pattern of adaptive divergence between pairs of ECB and ABB populations in France, as well as between pairs of ACB and ABB populations in China.Since the No specific permits were required for the described field studies.Ostrinia spp. larvae were collected from three locations in France  and three locations in China  was increased to 30 for preselective PCRs. The selective PCRs consisted in a 5-min denaturation at 95°C, followed by 13 cycles of a 30-s denaturation at 94°C, a 1-min hybridization starting at 65°C with a 0.7°C decrement per cycle, and a 2-min elongation at 72°C. These 13 cycles were followed by 27 cycles similar to those of preselective PCRs, and by a final 10-min extension at 72°C. To generate AFLP fragments, we used the same set of 13 primer pairs as in The genomic DNA of each individual was isolated from a piece of the larval body, using either the Qiagen DNEasy Blood and Tissue extraction kit or a standard phenol-chloroform procedure AFLP products were electrophoresed on an ABI 3130XL (Applied Biosystems) sequencer, together with the Internal Lane Standard 600 ladder (Applied Biosystems). Raw data were analyzed with GENEMAPPER© software version 4.0. Samples were first scored for the presence or absence of any given AFLP band (between 80 and 450 bp) on the basis of the automatic procedure in GENEMAPPER©, with intensity threshold set to 20. Then, all samples were visually corrected for, e.g., overlapping bands, unusual peak shape, or dubious peaks. AFLP loci were chosen and scored on the whole data set without considering the origin of the samples (host-plant and country), in order to avoid any ascertainment bias in the definition of the final set of AFLP markers.aflp-surv 1.0 aflp-surv, assuming Hardy-Weinberg equilibrium within each population and for each locus. Nei's He) was computed from these allele frequency estimates, and we used Lynch & Milligan's method aflp-surv to measure genetic differentiation between populations (FST).If any two independent AFLP fragments (corresponding to distinct DNA sequences) have exactly the same length, they may be analyzed as a single locus. Since a typical signature for such homoplasy is a negative correlation between fragment size and frequency, we evaluated the level of homoplasy in our dataset by means of the Pearson's correlation test implemented in the software structureK, the number of putative clusters, from 1 to 20, and performed 20 independent runs for each value of K. We used the admixture model, with the default initial value for α, the hyperparameter of the symmetric Dirichlet distribution for the admixture proportions, and assumed that allele frequencies were correlated across populations, using default priors for the kF parameters. Each Markov chain was run for 500,000 steps, after 50,000 steps of burn-in. We characterized the uppermost hierarchical level of clustering using Evanno et al.'s statistic ΔKK. We further checked the occurrence of multimodal solutions for a given value of K using the greedy algorithm implemented in the software clumppClustering analyses based on multilocus genotypes were performed with the Bayesian method implemented in the software package ayeScanOstrinia species. BayeScan is based on the Multinomial-Dirichlet model for allele frequencies in an island model of population structure. At each locus, the variance of allele frequency between each subpopulation and the common pool of migrants is given by a subpopulation-specific FST parameter. In BayeScan, as in Beaumont & Balding's FST is decomposed into a locus-specific component (iα) shared by all populations, and a population-specific component (jβ) shared by all loci. Positive or negative values of iα are taken as evidence for selection. BayeScan is based on a reversible-jump Markov chain Monte Carlo (MCMC) algorithm, which provides estimates of the posterior probabilities of two alternative models: one purely neutral (iα = 0) and one including selection (iα ≠ 0). For each locus, Q-values are computed from the posterior probability that the locus is under selection, allowing for the control of false discovery rate (FDR).We used the software package BayeScan version 2.1  for all possible pairs of populations, within a country, that involved samples collected from distinct host-plant types (inter-host comparisons) as well as for all possible pairs of populations, within a country, that involved the same host-plant type (intra-host comparisons). Following Nosil et al.iα estimates for all outlier loci, and for each inter-host comparison, as a proxy for the nature and strength of selection: positive iα values indeed suggest divergent selection while negative values suggest balancing selection.We ran BgdaWe performed a hierarchical analysis of molecular variance using the program DAB between any given pair of loci A and B within each population. The pairwise gametic correlation coefficient between pairs of loci was estimated following Dasmahapatra et al. pA and pB are the frequencies of the dominant alleles at locus A and locus B, respectively, estimated using Zhivotovsky's We tested for a possible clustering of outlier loci, as different AFLP markers may be located in the same genomic region. To this end, we calculated the level of linkage disequilibrium (LD): (i) within each French population, between pairs of outlier loci detected in French populations; (ii) within each Chinese population, between pairs of outlier loci detected in Chinese populations; (iii) within each population (Chinese or French), between all pairs of loci consisting in one outlier locus detected in French populations and one outlier locus detected in Chinese populations. The latter analyses were aimed at identifying a potential association between the genomic regions under selection in the ACB and ECB. Since AFLPs are dominant markers, we used Hill's et al. P-value for each locus pair within each population was then computed as the probability of getting values as large or larger than the one observed.We developed an original method to test for the significance of gametic correlation coefficients for dominant markers. Hereafter, we refer to the dominant allele (which corresponds to the amplification of one AFLP fragment) by “+” and to the recessive allele (which corresponds to the non-amplification of the AFLP fragment) by “−”. With dominant markers, randomizing the “band absence” and “band presence” phenotypes across individuals at two loci is not an appropriate way of characterizing the distribution of LD under the null hypothesis of linkage equilibrium, since the “band presence” phenotype may correspond either to the “+/+” or to the “+/−” genotype. An appropriate test must therefore be based on the randomization of the underlying genotype frequencies. To this end, we used Zhivotovsky's P-values were combined for each of the following categories: populations collected (i) in France on dicots, (ii) in France on maize, (iii) in China on dicots and (iv) in China on maize. To this end, we used the Z-transform method, as advocated by Whitlock survcompP-values were then corrected for the FDR due to multiple testing oF+ Finally, LD Ostrinia spp. populations (6 pairs of populations). The number of AFLP loci per pair of AFLP primers ranged from 35 to 82, with an average of 53. We did not detect any significant correlation between AFLP fragment size and frequency .We analyzed 684 AFLP loci in 271 individuals sampled from 12 He) per population ranged from 0.231 to 0.260 with an average of 0.245 and was homogeneous across populations .A total of 558 markers out of 684 were polymorphic using a minimum allele frequency of 0.05 across the whole dataset . Among them, 92% were shared between French and Chinese samples. Only 1% of the loci were private to pooled French samples and 7% to pooled Chinese samples . Furthermore, most loci were also shared between host-plants within countries, with only 6%, 7%, 5% and 1.5% of the loci being private to French samples on maize, French samples on dicots, Chinese samples on maize and Chinese samples on dicots, respectively. Nei's FST estimates provided in FST values were observed between populations collected on maize in France (FST ranging from 0.009 to 0.026), while the intra-host differentiation was a little higher on dicots in France, on dicots in China and on maize in China (FST ranging from 0.060 to 0.119) and to a lesser extent in France  for K = 7 putative clusters, and the highest value of Evanno et al.'s K) for K = 2. For K = 2, one cluster . These putative migrants were nevertheless kept assigned to the populations they were sampled from, in all subsequent analyses, including BayeScan; (ii) some individuals were assigned to a cluster that was not representative of any of the populations in our data set  were positive in all within-country inter-host comparisons where they were significantly detected as outliers  outliers were 0.678 (resp. 0.631), and 0.102 (resp. 0.098) for the presumably neutral markers. Both in France and China, the largest component of genetic variance was between host-plant types for outlier loci, and between individuals within populations for presumably neutral loci , neither between the 2 outliers detected in France, nor between the 36 possible pairs of outliers detected in China. Likewise, we did not find any significant LD between the 18 possible pairs consisting of one outlier detected in France and one detected in China.We did not find any significant LD  and the ACB (in Asia)  see . Whetheret al.et al.'s review referred to quantitative genetics studies, recent molecular studies on host shifts in Drosophila and aphids carried on this characterization by identifying two major classes of gene functions underlying host plant adaptation. The first class refer to chemoreceptor genes such as odorant, gustatory receptor genes and genes encoding salivary proteins in the pea aphid, Acyrthosiphon pisumDrosophila sechelliaD. mojavensisD. sechelliaOstrinia, the association between AFLP outliers detected in our study and candidate genes remains to be established. Yet, chemoreceptor genes would appear as relevant candidates since a recent flight tunnel study showed that ECB adults discriminate volatile organic compounds emitted by host plants We detected a small proportion of the genome to be involved in the adaptive divergence between the dicot-feeding and the maize-feeding taxa both in France and in China. This result is in line with those compiled by Matsubayashi We found that different and unlinked outliers differentiated ECB from ABB in France, and ACB from ABB in China. Detecting different genomic regions under selection in the ACB and ECB has two major implications. First, it strengthens the hypothesis of two independent adaptation events to maize. Indeed, if there had been only one adaptation event to a maize-like plant followed by a reversion to the ancestral host plant(s) in the ABB , others tend to show that genetic signatures of selection can spread over long genomic regions  trait We have implicitly assumed so far that adaptation to maize or to an intermediate “maize-like” host plant was the main force driving differentiation between species in each pair. We also implicitly assumed that this adaptation has occurred earlier in the ACB than in the ECB, as suggested (but not demonstrated) by the fact that the ACB and ABB are more differentiated than the ECB and ABB. However, more complex scenarios are possible. For example, in one or both species pairs, divergence may have been initially triggered – and possibly driven for some time – by other factors than host-plant use. If so, adaptation to a new host could have occurred much later. We must therefore keep in mind that, as pointed out by Bierne a priori reason to expect the former to be identical in both species pairs. Hence, shared outliers in both species pairs are better candidates for adaptation to the host-plant than to reproductive isolation – unless reproductive isolation and adaptation to host plant are ensured by the same or tightly linked loci/genomic regions (see above). As genome scans are not exhaustive though, and since none of the outliers we detected was shared between the two pairs of species, we cannot exclude that most, if not all, outlier loci are involved in reproductive isolation.If genome scans were exhaustive, we should therefore detect loci involved in reproductive isolation together with loci involved in host-plant adaptation. However, there is no et al.Ostrinia spp. may also help deciphering the putative historical scenarios Click here for additional data file.Figure S2structure analyses for K = 2 to K = 7. Each individual is represented by a vertical line, divided into up to K coloured segments representing the individual's estimated fraction of membership of each of the K clusters. Vertical black lines separate samples from different localities and host-plants, as labelled at the bottom of the figure. Host-plant and country are indicated at the top. For each value of K, the less supported clustering solutions, in terms of number of runs out of 20 (number given to the right of the plot) are represented, whereas the most supported solution is given in Estimated clustering from (EPS)Click here for additional data file.Table S1Number of outlier markers detected using methods alternative to BayeScan.(DOC)Click here for additional data file.


Bactrocera dorsalis, was initially recorded in Taiwan Island in 1912, and has dispersed to many areas in the Pacific-Asia region over the last century. The area of origin of the species may be confidently placed in South-East China. However, routes of range expansion to new areas and underlying population processes remain partially unclear, despite having been the subject of several studies. To explore the invasion history of this species, a partition of the cox1 gene of mitochondrial DNA was used to investigate genetic diversity, haplotype phylogeny and demographic history of 35 populations, covering China and South-East Asia and including marginal populations from Pakistan and Hawaii. Based on neighbor-joining tree analysis and the distribution of haplotypes, two main invasion routes are inferred: one from South-East China to Central China, another from South-East China to South-East Asia, with both routes probably coinciding in Central China. Populations in Taiwan Island and Hainan Island might have originated in South-East China. The marginal populations in Pakistan and Hawaii might have undergone founding events or genetic bottlenecks. Possible strategies for the control of this species are proposed based on the invasion history and reconstructed expansion routes.The oriental fruit fly,  Biological invasions constitute a growing threat to human economic activities and health, agriculture, and natural environments. Although many species of plants, animals and other organisms have been introduced around the world by human activities An understanding of the history of invasion processes, i.e. a description of geographical pathways of invading populations, provides useful information about the origin and genetic composition of such populations and can be highly advantageous in attempts to quarantine, control or eradicate an invading species. Investigating the genetic architecture of invading populations offers an opportunity to infer a species' invasion history using molecular techniques Although the reliability of some traits of mitochondrial DNA (mtDNA), such as neutrality and constant mutation rate, has been criticized Bactrocera dorsalis, a phytophagous species of the Tephritidae family, was first recorded in Taiwan Island, China, in 1912 The oriental fruit fly In this study, we aim to a) expand our previous work on oriental fruit fly expansion to cover all areas of the species' worldwide distribution, b) reconstruct the routes of the species' expansion from its origin in South-East China to areas of recent colonization in South-East Asia and to marginal populations in Pakistan and Hawaii, and c) discuss the application of these results to the planning of appropriate control measures.B. dorsalis adults collected from 14 locations in China and one in Pakistan in 2008–2010  DNA was extracted using DNeasy Blood and Tissue Kits (QIAGEN) on each individual fly. A 505 base-pair partial segment of the mtDNA Sequences were aligned using ClustalX 2.0 The Kimura two-parameter model in MEGA 5.0 To infer asymmetric immigration rates between different regions, seven regions were defined based on geographic location and previous studies M = m/μ, where m is immigration rate and μ is mutation rate per site per generation), and mutation-scaled effective population sizes , by applying a Bayesian search strategy. Four independent MIGRATE runs of 20,000,000 generations with different random start seeds were performed to examine the consistency of the results, with the first 10,000 generations discarded as “burn-in”. Analysis of molecular variation (AMOVA) and computation of fixation indices (FST) between pairwise regions were implemented in ARLEQUIN.MIGRATE 3.27 0θ), population size after expansion (1θ), population expansion time (τ), Tajima's D, Fu's FS, and sum of squared deviations (SSD) between observed and expected mismatches. All parameters were calculated using ARLEQUIN and tested against the expected values of a recent population expansion with 1000 bootstrap replicates.The demographic history of each region and of all populations pooled together was examined using mismatch distributions, population size before expansion  was to be found within populations, while genetic differentiation among groups (6.41%) and among populations within each group (7.36%) was limited. All fixation indices tested as highly significant (P<0.01) .ST values ranged from −0.0018 (Taiwan Island and Hainan Island) to 0.4948 (South Asia and Hawaii), and increased with geographic distance. FST values between Southeast China, Taiwan Island, Hainan Island, Central China and Southeast Asia were low. Relatively high and highly significant, FST values were found between South Asia or Hawaii and the other five regions (Pairwise F regions .Θ×M) entering these two regions were very limited  intervals between immigration rates into and out of each region. Due to the low effective population size, 0.00372 and 0.00141 in South Asia and Hawaii respectively, gene flows  located in the center and other rare haplotypes connected to them through several mutation steps. The haplotypes belonging to the Hawaii region were clearly separated from others in the network, with the exception of H208. H208 connected to H19 through one mutation step, as did the main haplotype of H210 with H154 . Other hFS values were found among pooled populations, as well as in the separate populations from Southeast China, Central China, Hainan Island and Southeast Asia, suggesting that the B. dorsalis populations of those distribution regions did not conform to the theory of neutral evolution (Significantly negative Tajima's D and Fu's volution .SSD = 0.0032) and the Central China region (PSSD = 0.0169).The unimodal mismatch distribution  of theseτ) in the seven regions ranged from 2.506–5.828. Ratios between the effective population after expansion (1θ) (3.680–9999.000) and effective population size before expansion (0θ) (0.001–0.039) implied large population growth.Population expansion time  suggest a gradual invasion process along this route, coupled with rapid population expansion One invasion route may run from South-East China to Central China, as indicated by asymmetric gene flows from South-East China to inland China FS values suggest that due to population expansion, selective neutrality is not currently present in this species in South-East Asia Another invasion route may run from South-East China to South East Asia. The fact that the PX population  located on the border of China and Vietnam shared haplotypes with South-East China and other South-East Asia populations  indicateThe two independent invasion routes from South-East China may be coinciding in Central China. Populations in this area share haplotypes with both the South-East China and South-East Asia regions and are distributed in both clades of the NJ tree. Genetic admixture supplied by different independent routes in the form of multiple introductions, which have also been found in our previous study B. dorsalis in Hawaii is equivocal. No shared haplotypes were found between Hawaii and other regions. MJ network analysis indicates that haplotype H208, unique to Hawaii, is derived by mutation from H19, mainly found in Taiwan Island, South-East China, Hainan Island and South-East Asia. However the other haplotypes found in this region are mutations of the unique haplotype H154 attributed to Wanzhou, a recently invaded area of Central China. This discrepancy may be due to insufficient haplotype sampling size. Based on historical records and the results of PCR-RFLP analysis of a mitochondrial control region 0θ = 0.01) may be the main cause of this genetic homogeneity; constant pest-control pressure associated with strong quarantine regulations imposed on the fruit trade may be another reason. Very limited gene flows detected between Hawaii and other regions indicate that the Pacific Ocean is a barrier to migrational exchange. No shared haplotypes  may have been directly from South-East China, from the nearby region of Myanmar. This interpretation is based on haplotype H6 being shared with the FZ and NN populations, H215, H216 and H217 being mutated from H3 (shared by Taiwan Island and South-East China), H22  and H193 (unique haplotype of Myanmar). The high genetic diversity of HP  is likelBecause of the considerable differences in ecological conditions between the endemic and non-native regions, natural selection and adaption may be determinants for the success of an invasion before or during the settlement phase. Sufficient genetic variation is essential for evolutionary adaption in response to environmental change and can facilitate the adaption to new environments The oriental fruit fly has to be considered a highly invasive species, as it is able to disperse efficiently and establish adventive populations in various tropical and sub-tropical climate zones. It may however be possible to develop some management strategies based on available invasion history information. For populations that are genetically well-connected and show high gene flow , any intervention that is geographically limited to one region is likely to fail, as neighboring populations would readily re-colonize the region. Area-wide interventions aimed at reducing population numbers and economic damage would be a more feasible choice in this case, which would be also appropriate for HON population with relatively low genetic diversity but high population density Table S1cox1 sequences obtained from GenBank.Accession numbers of the (DOC)Click here for additional data file.Table S2Haplotypes constitution of each population.(DOC)Click here for additional data file.


Bruguiera gymnorrhiza, using ten nuclear gene regions. Genetic variation of individual populations covering its distribution range was low, but as the entire species it was comparable to other plant species. Genetic differentiation among the investigated populations was high. They could be divided into two genetic clusters: the West and East clusters of the Malay Peninsula. Our results indicated that these two genetic clusters derived from their ancestral population whose effective size of which was much larger compared to the two extant clusters. The point estimate of speciation time between B. gymnorrhiza and Bruguiera sexangula was two times older than that of divergence time between the two clusters. Migration from the West cluster to the East cluster was much higher than the opposite direction but both estimated migration rates were low. The past Sundaland and/or the present Malay Peninsula are likely to prevent gene flow between the West and East clusters and function as a geographical or land barrier.Inference of genetic structure and demographic history is fundamental issue in evolutionary biology. We examined the levels and patterns of genetic variation of a widespread mangrove species in the Indo-West Pacific region,  Mangrove species constitute a unique ecosystem in coastal zones of tropical and subtropical regions. The distribution of mangrove species is classified into two major regions, Indo-West Pacific (IWP) and Atlantic-East Pacific (AEP)  markers and chloroplast DNA (cpDNA) sequences  Voris  and thusBruguiera gymnorrhiza populations covering its natural distribution range. B. gymnorrhiza (L.) Lamk. (Rhizophoraceae) is a typical mangrove species and one of the predominant mangrove species in the IWP region. It has the broadest distribution in the genus: from eastern coast of Africa in the west through Asia, northward to the Ryukyu Islands of southern Japan and southward in tropical Australia (Tomlinson B. gymnorrhiza has a unique viviparous propagule (Tomlinson B. gymnorrhiza is one of the bird-pollinated large-flowered species of Bruguiera (Tomlinson Bruguiera sexangula (Lour.) Poir., was also examined.Our main objective is to clarify genetic structure and demographic history of typical mangrove species on a global scale. For this purpose, we examined genetic variation of ten nuclear gene regions as well as one cpDNA region using samples from ten We address the following specific questions: (1) What is the extent of nucleotide sequence variation? (2) What is the extent of genetic differentiation between populations? (3) Do the Sundaland and/or the Malay Peninsula play a role of a barrier to prevent gene flow between the Pacific and Indian Oceans? (4) When did the two species diverge?B. gymnorrhiza were sampled from different individuals at the following ten locations  method , VVHP (BP944085), PO (BP939063), GM (BP939421), SF (BP939928), EXP2 (BP942725), EPCRF (BP946081), and UNK (BP945250), where the corresponding B. gymnorrhiza ESTs  of Hamilton , were alPCR amplification was carried out under the following protocols: an initial denaturation at 95°C for 3 min followed by 30 cycles of denaturation at 95°C for 30 sec, annealing at 50°C, 55°C, or 60°C for 30 sec, polymerization at 72°C for 1.5 min, and a final extension at 72°C for 7 min. The details of the sequence and annealing temperature of PCR primers are listed in Two haplotype sequences were determined for each individual. After the direct sequencing when we found no or only one heterozygous site in a sequence of a single individual, we inferred sequences of two haplotypes for each individual. On the other hand, when two or more heterozygous sites or indels were found in a sequence of a single individual, purified PCR product was cloned into the pGEM T-easy vector . Then, we sequenced each clone using T7 and Reverse primers designed at the promoter sites of the vector (Table S1). Four to sixteen clones were sequenced for each individual. Sequences of two haplotypes of a heterozygous individual were determined when we obtained at least two exactly identical sequences for each haplotype.EXP2 gene of one individual sampled from the AB population from the east coast of Australia, we treated it as missing data in the following analysis. A continuous alignment gap was treated as a single indel. All sequences obtained in this study were deposited in the DNA Data Bank of Japan (DDBJ). Accession numbers are AB813916-AB817042.Sequences of each gene were aligned using CLUSTAL X 2.1 , and Ka (number of nucleotide substitutions per replacement site between species), were estimated using the DnaSP ver. 5.10.00  among populations, (ii) among individuals/within populations, and (iii) within individuals, were examined for nuclear genes by the locus-by-locus analysis, and two levels, (i) among populations and (ii) within populations, were examined for cpDNA. Each haplotype in a gene was regarded as an allele. Overall values of F-statistics for all the ten nuclear genes were obtained by summing variance components over the genes. In the AMOVA analyses, genotypic data of nuclear genes, where gametic phase of genotype is unknown, and haplotypic data of the cpDNA region were used. A haplotype network of the cpDNA region was constructed using the TCS ver. 1.21  approach  to confirm the convergence of Markov chain Monte Carlo (MCMC) chains. In each run, 500,000 MCMC iterations were performed after a burn-in period of 1,000,000 iterations. To estimate the number of clusters, we applied the method proposed by Evanno et al.  model using the IMa2 program (Hey and Nielsen T = tu), population sizes (θ = 4Nu), and migration rates (2Nm) between genetic clusters (or species) were estimated, where u is the mutation rate per locus per generation, t is the population splitting (or speciation) time, N is the effective population size, and m is the migration rate. The infinite site model  to 0.02506 (EPCRF) for the nuclear genes (Table NAC and PO) were monomorphic. In the total population, the average πs over the ten nuclear genes was 0.00800. The πs value of the cpDNA region was 0.00008. Nucleotide diversity of the nuclear genes was ∼100 times higher than that of the cpDNA region, indicating that the nuclear genes are more useful markers than the cpDNA region.The level of nucleotide variation was different among the genes. For example, nucleotide diversity Nei  per sileπs value over the ten nuclear genes ranged from 0 (G and K populations from the southwest and northeast India) to 0.00424 (V population from the east of the Asia Continent) (Table S2).The level of nucleotide variation also varied among the ten populations. For example, the average B. gymnorrhiza and B. sexangula is summarized in Table Ks) ranged from 0.00437 to 0.04925 for the nuclear genes and 0.00334 for the cpDNA region. The average Ks value over the ten nuclear genes was 0.02162. Divergence at silent sites of nuclear genes was ∼10 times higher than that of the cpDNA region. The number of nucleotide substitution per replacement site between the species (Ka) ranged from 0.00000 to 0.00485 for the nuclear genes. The highest value of Ka/Ks was 0.32071 for PAL1, indicating that these nuclear genes are not pseudogenes.Nucleotide divergence between D values were not significant except for the VVHP gene of MO population (the west of Madagascar) and the mang-1 gene of the V population (the eastern part of the Asian Continent). No significant results were obtained in Fay and Fu's test (Table S3). The result of the multilocus HKA test was weakly significant (P = 0.048). In MK test no significant result was found (data not shown). These results suggest that gene regions examined could be treated as neutral markers in B. gymnorrhiza.Results of neutrality tests are summarized in F-statistics among the ten populations estimated by the AMOVA approach were shown in Table NAC and PO) had no variation, and therefore they did not have any information for population structure. The estimated FST values were highly significant both for nuclear genes and cpDNA region, indicating that there is genetic differentiation among the ten populations. In nuclear genes most population pairwise FST values were highly significant except for population pairs, AB  – MK (northwestern Borneo), AB – ML , AC  – ML, BMJ (eastern Java) – MO (western Madagascar), V (eastern part of Asian Continent) – K (northeastern India), MK – ML, and G (southwestern India) – SL (southeastern Madagascar) (Table S4), indicating that most population pairs are genetically differentiated. Although the pairwise FST values in nuclear genes were not significant for population pairs, AB  – MK (northwestern Borneo), AB – ML , and AC  – ML, a relationship of cpDNA haplotypes . This result indicates the speciation event between B. gymnorrhiza and B. sexangula is roughly two times older than the divergence of the two clusters in B. gymnorrhiza.To infer the population history of the two genetic clusters identified by the STRUCTURE analysis, population parameters were estimated under the IM model using IMa2 program (Hey and Nielsen θ for the East cluster (θE) was 0.17  and θ for the West cluster (θW) was 0.08 . θ for the ancestral population was 2.05 . This result means that the effective population size of East cluster is larger than that of West cluster, and the ancestral population is much larger than either of the clusters. Larger size in ancestral populations than the extant populations was also reported in the other mangrove species  than from the East to West , although in both cases the migration was rare (2Nm < 1). No migration was estimated between B. gymnorrhiza and B. sexangula after the speciation event (data not shown).The estimate of B. gymnorrhiza, leaf materials were collected from 10 localities covering its distribution range and DNA sequences were determined. General pattern of nucleotide variation of B. gymnorrhiza was as follows: (1) low polymorphism in each local population but (2) heterogeneous variation between the local populations (Table S2). As the entire species, however, the level of polymorphism was comparable to other plant species  genes, each population is likely to have little potential for genetic adaptability. This suggests that extinction risk of each population is high. Alternatively, it may be the nature of this species, for example, the species may repeat the following extinction-recolonization cycle; (1) a population starts with a small number of individuals, which adapted (or survived) to a given environment, (2) the population increase in size , and (3) when the environment changes accompanying with climatic and/or geographical changes, the population disappears.B. gymnorrhiza and B. sexangula was ∼2%. It was similar or smaller that nucleotide divergence between Sonneratia species  was comparable to those of Sonneratia alba (0.00432) and S. caseolaris (0.01003)  and R. mucronata (0.00003) . Similar trend was observed in the cpDNA region. In this study, in the trnS-trnG spacer, three and one segregating sites were found in B. gymnorrhiza and B. sexangula, respectively, and one fixed substitution was observed between the two species. Zhou et al. . As is the case of other mangrove species in the South East Asia , the estimated population splitting parameters need to be divided by the mutation rate per locus per year. As it is difficult to estimate mutation rate of io Table . If we amang-1 and a unique haplotype in the SF, EXP2, and GM (data not shown). Except for comparisons with the V (eastern part of the Asian Continent) population, pairwise FST values were much higher in comparisons with the BMJ (eastern Java) population (Table S4: BMJ – MK (northwestern Borneo): 0.69, BMJ (the east of the Java) – ML : 0.68, MK (northwestern Borneo) – ML : 0.10). In the Southeast Asia the BMJ (eastern Java) population could be an ancient population compared with others.During the LGM, the sea level dropped ∼120 m compared with the present geographic area, and the Sundaland was formed in the Southeast Asia. At that time three localities examined in this study  were not coast, while one of them  was considered to be the edge of the Sundaland. Extant populations located on the previous Sundaland should be established after the LGM. Consistent with this, the BMJ (eastern Java) population showed higher polymorphism than the ML  and MK (northwestern Borneo) populations (Table S2). In addition, the BMJ (eastern Java) population harbors five of thirteen haplotypes in the


Oncorhynchus mykiss. Samples included 805 individuals from 11 anadromous and resident populations from the northwestern United States and British Columbia, and represented two major lineages including paired populations of each life history within single drainages of each lineage. Overall patterns of variation affirmed clear distinctions between lineages and in most instances, isolation by distance within them. Evidence for divergent selection at eight candidate loci included significant landscape correlations, particularly with temperature. High diversity of two nonsynonymous mutations within the peptide-binding region of the major histocompatibility complex (MHC) class II (DAB) gene provided signatures of balancing selection. Weak signals for potential selection between sympatric resident and anadromous populations were revealed from genome scans and allele frequency comparisons. Our results suggest an important adaptive role for immune-related functions and present a large genomic resource for future studiesRecent advances in molecular interrogation techniques now allow unprecedented genomic inference about the role of adaptive genetic divergence in wild populations. We used high-throughput genotyping to screen a genome-wide panel of 276 single nucleotide polymorphisms (SNPs) for the economically and culturally important salmonid  Inference of the structure and relatedness of natural populations exploded in the 1960s and 1970s with the development of molecular genetics and a deepening of our understanding of the genetic basis driving evolutionary change e.g., . Our abiOncorhynchus mykiss  (The purpose of this study was to screen a new genome-wide SNP resource in . nerka)  and browo trutta). Using an = 24–95) of O. mykiss from nine rivers throughout the Pacific Northwest of North America , were discarded in order to assume neutrality and independence among a set of 246 remaining markers . PCR amplification and genotyping was performed in 96.96 Dynamic Arrays using the Fluidigm IFC thermal cyclers and BioMark instruments following the protocols of results, . Genome results, .FST estimates among the 15 collections were generated in Arlequin 3.5  (FST = −0.002–0.011) compared to all spatial comparisons (FST = 0.013–0.375). Assuming a conservative α value of 0.001, all temporal comparisons remained nonsignificant while spatial comparisons were all significant. Temporal replicates were pooled to optimize sample sizes for a total of 11 populations in subsequent analyses.Pairwise quin 3.5  using 10k = 105) . These tP values were corrected using the sequential Bonferroni method (k = 3069) (P < 0.05) with at least six performed tests among 11 populations, (3) for all locus pairs showing significant LD, SNPs potentially involved in multiple pairs were discarded.Conformance to HWE was tested independently for each locus in each of the 11 populations using the MC algorithm implemented in Genepop 4.0 . P value = 3069) . Linkage = 3069) . Thus, w = 3069) . Due to FST values were estimated for each locus in Genepop 4.0 (EH) and observed (OH) heterozygosity were calculated for each locus and population using GenAlEx 6.4 (RA), a measure of the number of alleles corrected for minimum sample size, was calculated for all populations using FSTAT v2.9.4  to generate a multi dimensional scaling (MDS) plot in ViSta 5.6.3  in relation to observed heterozygosity. Observed values by locus were then projected onto this distribution, and loci lying above or below the simulated 99% confidence threshold for neutral variation were considered as candidates for divergent or balancing selection, respectively. We applied the hierarchical test by grouping populations into two groups representing the coastal and inland lineages (P < 0.01) detected for both FST  and FCT , we plotted major allele frequencies over all populations. We also performed a genome scan over all anadromous populations using BayeScan 1.0  using 10 simulated demes and 100,000 simulations in Arlequin 3.5. Again, we plotted major allele frequencies over all populations for all outliers above the 99% confidence levels. BayeScan 1.0 was not considered here, since it is expected to perform poorly with few samples .We further tested for associations between landscape variables and allelic distributions for each SNP to reinforce evidence of natural selection acting on outlier loci  e.g., . Underlya priori focus to six newly developed markers residing within the classical MHC class I  and the classical MHC class II  genes  with a simulated value  expected at mutation drift equilibrium for a gene with a similar number of alleles  genes . We obsec) genes . To testc) genes  using thc) genes  to test  alleles , and wheP < 0.05) in more than half of the performed tests leading to the exclusion of four loci, of which some were involved in multiple pairs, to avoid potential pseudoreplication by including markers in LD (n = 23–95) and 266 SNPs , 0.09 to 0.24 (OH), and 1.45 to 1.80 (RA) with highly reduced levels in the SKAGRES population  at the FST level (FCT level (P < 0.01) found with Arlequin as well as three new outliers  were also detected in local genome scans for selection between within river populations exhibiting alternate life histories. We observe one outlier in the allopatric Skagit River and six in the sympatric Twisp River comparisons  global candidates for divergent selection , had smaller than expected F values pointing toward balancing selection (One SNP within the MHC class I gene (Omy_UBA2a) was discarded due to significant deviation from HWE . Remainielection .O. mykiss from this region using allozymes and mtDNA  with consequently strong genetic drift. A similar scenario has been shown for another physically isolated population of resident O. mykiss . Despite potential gene flow between the sympatric anadromous and resident Twisp River populations see also , our resFCT outliers for distinguishing between the two lineages. Population history and dynamics of populations spawning within or in close proximity to the transition zone has been difficult to infer in previous studies applying fewer and assumingly neutral markers , in particular, may play important roles in shaping adaptive genetic variation in O. mykiss. An effect of temperature would be in accordance with three regional (FST) outliers following a latitudinal trend within the coastal lineage  and a nucleoside diphosphate kinase (Omy_ndk-152) gene . A recenlineages . A recenlineages . Indeed,lineages . Due to lineages . Neverth lineage . A recenFST = 0.30, P < 0.001). However, the observation of six outlier loci  at the 99% confidence level between the Twisp populations, together with another marker showing signatures of selection in both locations, suggest ongoing selection between anadromous and resident life-history types. This overall result is consistent with recent studies identifying signatures of divergent selection between different migratory variants in O. mykiss  and found similar evidence for balancing selection. Sequence-based analyses are in general expected to be more powerful for detecting balancing selection compared to individual marker based outlier tests (The two known nonsynonymous mutations in the peptide-binding region of a MHC class II gene (Omy_DAB_431 and Omy_DABb) generally showed high levels of diversity in most populations with MAF ranging between 0.26 and 0.48 . Althoug. mykiss . Furtherer tests .O. mykiss populations from the Klickitat basin in the Columbia tributary, Washington. This result is in agreement with our observations at this locus showing a signature of divergent selection between the resident and anadromous populations in the Twisp River  and a MHC class II gene . Here, the latter is represented by three SNPs hitherto unscreened in wild populations. These candidate genes will inevitably prove valuable in future studies of Genome scans including functional genetic variation have proven very promising for identifying (and understanding) adaptively important genes and traits in nonmodel organisms (e.g., Nielsen et al.


Catla catla Hamilton 1822) in the Halda, Jamuna and Padma rivers and one hatchery population in Bangladesh. Five decamer random primers were used to amplify RAPD markers from 30 fish from each population. Thirty of the 55 scorable bands were polymorphic, indicating some degree of genetic variation in all the populations. The proportion of polymorphic loci and gene diversity values reflected a relatively higher level of genetic variation in the Halda population. Sixteen of the 30 polymorphic loci showed a significant  departure from homogeneity and the FST values in the different populations indicated some degree of genetic differentiation in the population pairs. Estimated genetic distances between populations were directly correlated with geographical distances. The unweighted pair group method with averages (UPGMA) dendrogram showed two clusters, the Halda population forming one cluster and the other populations the second cluster. Genetic variation of C. catla is a useful trait for developing a good management strategy for maintaining genetic quality of the species. Genetic variation is a key component for improving a stock through selective breeding programs. Randomly amplified polymorphic DNA (RAPD) markers were used to assess genetic variation in three wild population of the catla carp ( Catla catla Hamilton 1822: Cypriniformes, Cyprinidae) is an Indian major carp and one of the major aquaculture species of Bangladesh, India, Myanmar (Burma) and Pakistan , randomly amplified polymorphic DNA (RAPD), restriction fragment length polymorphisms (RFLP) and allied methodologies are useful for piscine gene pool conservation and enhancement of aquaculture production of fish species  from each pond (representing one distinct population) for RAPD marker analysis. Caudal fin tissue samples were clipped from each fish and preserved in 95% ethanol.® Genesys5 UV-spectrophotometer . We used sub-samples of three fish from each population to screen 27 random sequence decamer primers , 10 of which were selected on the basis of the intensity or resolution of bands without smearing, repeatability of markers and consistency within individual fish. Of the 10 pre-selected primers, seven, exhibiting the highest quality banding patterns, were used for analysis of all the samples.Genomic DNA was extracted from fin tissue following the standard method of ++, and Taq DNA polymerase concentrations and to determine the optimum annealing temperature essentially following the method described by Initial PCR amplifications were ran to test the effect of template DNA, dNTPs, MgH), gene flow (Nm) and homogeneity test at different loci using the POPGENE software version 1.31 . The Tools for Population Genetic Analyses (TFPGA) program version 1.3  was applied for the estimation of population differentiation by the Reynolds coancestry coefficient (FST or θ) (DCH) (D) (Si) and between populations (Sij) were calculated according to the formula given by All distinct fragments were given identification numbers according to their size as measured by the DNAfrag software version 3.03  and scorθ) (DCH) , Nei's gDCH) (D)  and ReynDCH) (D)  based onDCH) (D)  implemenOf the 27 decamer primer sets initially tested, seven primers  yielded more numerous bands with high intensity and minimal smearing, although two (OPAB05 and OPU20) produced monomorphic bands and were excluded from the analysis. The five remaining primers generated 55 distinct bands, 30 of which were polymorphic. The frequency of the most common allele was 0.95 or less.P) and Nei gene diversity (H)  in the Halda river population, followed by the Padma  and Jamuna river  populations. However, the lowest percentage of polymorphic loci (p = 41.82%) and less gene diversity (H = 0.174) was found in the hatchery population.The overall proportion of polymorphic loci (sity (H)  across aSi = 87.86%) than between population similarity (mean Sij = 87.07%). The Si value was found to be highest in the Jamuna population (89.15%) followed by the hatchery (88.15%), the Padma (87.44%) and the Halda (86.72%) populations. On the other hand, the Sij value for the Jamuna-Padma population pair was the highest (88.74%) compared with all the other between population comparisons. The highest gene flow (Nm) was between the Padma and hatchery populations (Nm = 18.16), while the lowest was for the Halda-hatchery population pair (Nm = 6.34). Among the wild populations, the highest Nm value (17.17) was between the Padma and Jamuna river populations. No significant population differentiation (FST) was found in the studied populations at the 95% confidence interval (CI)  . Of the DCH = 0.043 to 0.145, D = 0.052 to 0.136 and θ = 0.013 to 0.040  and reported 24 RAPD polymorphic loci and some degree of genetic variation. In our present study, to obtain a more precise knowledge of the genetic variation in C. catla, we included one hatchery population together with three river populations and screened a total of 120 fish and used five different random primers . Our study revealed 30 polymorphic loci, indicating a high level of polymorphism in the four C. catla populations studied and supporting the suitability of RAPD markers for effectively discriminating different populations. Moreover, our present study compared the genetic variation between hatchery and river populations, which might assist in formulating good management practices for increased aquaculture production.Catla catla, Cirrhinus mrigala, Labeo rohita and Labeo calbasu) an average of 45% of loci were polymorphic  in the wild populations, especially the Halda and Padma populations, compared to the hatchery population. The Halda is a fresh water tidal river which is one of the principal spawning grounds of Indian major carps (Ne) that possibly resulted in higher genetic variation. Lower levels of genetic variation in the hatchery population might be due to, among other factors, inbreeding, bottlenecking or genetic drift. To keep the maintenance costs to a minimum farmers maintain a smaller number of broods in most hatcheries and thus induce breeding between related fish, which means that the genetic variation of hatchery reared fish may be reduced over a period of captive management in the hatcheries. Similar findings to ours were reported in a population genetic analysis of L. rohita (We found that 54.55% of the loci in our study were polymorphic as compared to the 75% reported by ymorphic . Our stuor carps . Similar. rohita .FST) in each population pair, with different FST values and significant deviations from homogeneity at a number of loci in the different populations indicating some degree of population differentiation in the different population pairs  produced UPGMA dendrograms with similar clustering patterns. Mantel tests detected correlations between genetic and geographical distances for some population pairs, with large genetic distances being found between distantly located populations. The Halda, an isolated river in south-eastern Bangladesh, is far from the other three populations, resulting in larger genetic distances between the Halda and any one of the other three populations. Contrastingly, the close relationship between the Padma and Jamuna population pair, as shown by the small genetic distance between this pair, may have been due to the connection between these two rivers  and simple sequence repeat (SSR) markers to better characterize populations of C. catla and other fish.In spite of still being used in assessing population structure, RAPD markers have many shortcomings, such as difficulty in demonstrating Mendelian inheritance of the bands (which are not true genetic loci), the inability to distinguish between homozygotes and heterozygotes, the presence of paralogous PCR products and the low reproducibility all limit the application of RAPD marker in some cases. To produce more precise data on the genetic structure of


The study of genetic information can reveal a reconstruction of human population’s history. We sequenced the entire mtDNA control region  of 605 individuals from seven Mesoamerican indigenous groups and one Aridoamerican from the Greater Southwest previously defined, all of them in present Mexico. Samples were collected directly from the indigenous populations, the application of an individual survey made it possible to remove related or with other origins samples. Diversity indices and demographic estimates were calculated. Also AMOVAs were calculated according to different criteria. An MDS plot, based on FST distances, was also built. We carried out the construction of individual networks for the four Amerindian haplogroups detected. Finally, barrier software was applied to detect genetic boundaries among populations. The results suggest: a common origin of the indigenous groups; a small degree of European admixture; and inter-ethnic gene flow. The process of Mesoamerica’s human settlement took place quickly influenced by the region’s orography, which development of genetic and cultural differences facilitated. We find the existence of genetic structure is related to the region’s geography, rather than to cultural parameters, such as language. The human population gradually became fragmented, though they remained relatively isolated, and differentiated due to small population sizes and different survival strategies. Genetic differences were detected between Aridoamerica and Mesoamerica, which can be subdivided into “East”, “Center”, “West” and “Southeast”. The fragmentation process occurred mainly during the Mesoamerican Pre-Classic period, with the Otomí being one of the oldest groups. With an increased number of populations studied adding previously published data, there is no change in the conclusions, although significant genetic heterogeneity can be detected in Pima and Huichol groups. This result may be explained because populations historically assigned as belonging to the same group were, in fact, different indigenous populations. The American continent was peopled across the Bering Strait from Asia after the LastGlacial Maximum (LGM) about 15,000 years before the present (YBP) During the process of human expansion, fragmentation and diversification of the aboriginal populations increased. In fact, there are currently between 400 and 1,500 indigenous groups in America Different molecular genetic studies of Native American populations are based on the mitochondrial genome (mtDNA) and five haplogroups: A2, B2, C1, D1 and X2 The characterization of a large number of Native Mexican populations should provide insights into the American model of colonization and expansion and the possible factors that have given rise to the current genetic and cultural diversity of the region.In this paper 605 mtDNA control region sequences from eight indigenous Mesoamerican and Aridoamerican groups were analyzed. Note that Aridoamerica corresponds to the area proposed by Kroeber in 1939 The 605 samples  came from 8 indigenous groups . Four maTo determine whether the frequency distribution of haplogroups in Mesoamerica follows a geographical pattern, correlations between these and the variables latitude and longitude were calculated. In Mesoamerica, and hgC1 hgB2 show the Same trend, Increasing with latitude  and decreasing with longitude . While you have to reverse hgA2 trend, decreasing with latitude  and with Increasing longitude , presents no correlation hgD1 With The Above-Mentioned parameters.Genetic diversity indices are shown in No significant values of Tajima’s D were found in the study groups. All eight populations therefore fit the neutral theory model. This result contrasts with the value obtained for all populations taken together , in which there is an excess of rare haplotypes, indicating a rapid growth of the indigenous population.K and θS indices  and geographical distances were calculated. Correlation between these matrices was checked using the Mantel test Two matrices representing genetic distances . In this group, and unlike other geographical associations, it was decided that western slope populations in Sierra Madre Occidental, such as both Otomi groups and the Nahuas de la Huasteca, should be included in the Mexican central highland group, while those that were located on the eastern side, such as the Tepehuas, were classified as coastal. A detailed analysis of this geographic association shows that it overlaps with cultural regions AMOVAs were also performed to detect population structure according to geographical, cultural, and linguistic criteria . Based oThere are a number of distinct regions within Mesoamerica that are defined by a convergence of geographic and cultural attributes, being more conceptual than culturally meaningful. The demarcation of their limits is not rigid. The areas include Maya, Central Mexico, West Mexico, the Gulf Coast Lowlands, Oaxaca, the Southern Pacific Lowlands, and Southeast Mesoamerica (including northern Honduras).ST distances represented in a MDS plot (stress = 0.097) (F= 0.097)  showed tThe four median-joining networks  and S4 rThe A2 median-joining network  has two In order to obtain a broader view of the diversity and history of Mesoamerica, we compared data for twelve indigenous populations from the scientific literature Note that in this paper the terminology used by the National Commission for the Development of Indigenous Peoples (CNI Mexico) has been followed. For this reason groups O’odham Akimel O’odham and Tohoma have been called Pimas_k and Papago. The diversity and demographics indices of these populations are presented in The geographic discontinuities in the haplogroup frequency were computed, detecting genetic boundaries that delimit geographical regions . The mosThe aim of this study is provide information about the origin, phylogenetic relationships, and history of Mesoamerican indigenous groups. Samples representing the different cultural and geographical areas of Mesoamerica and Aridoamerica were collected by the investigators directly from indigenous communities. This strategy permitted the exclusion of relatives and samples of dubious origin, which guaranteed they were representative of the selected indigenous groups. The analysis of 605 complete mtDNA control region sequences for seven Mesoamerican and one Aridoamerican indigenous groups makes it possible to reconstruct the female biological past of these populations.The four Pan-American haplogroups arrived on the continent at the same time The distribution of Amerindian haplogroups does not fit the general model proposed by some authors: hgA2 north-south gradient, hgC1 and hgD1 south-north gradients, and a lack of distribution for hgB2 Genetic diversity is an estimation of a population’s genetic variation and the result of its demographic history. In this sense, the lowest values were detected in Pima, Tepehua and Mayo populations, although each of them does so for different reasons. The first two populations were tipically hunter-gatherer communities until recently K and θS, while populations with a lower diversity index usually present positive values for D and moderate values for θK and θS.The demographic model based on genetic diversity is corroborated by the results of the θ and Tajima’s D indices. Groups with a high diversity index show negative values for D, suggesting an excess of rare alleles in the population and high values of θMesoamerican groups have a very similar demographic history characterized by ancestral demographic uniformity, and have grown over time. Northern populations differ from this model, however, since some are based on smaller sizes than the Mesoamerican populations, though they have grown in recent times.AMOVA results verify that Mesoamerican populations are genetically structured according to geographical parameters. This structure, allows us to draw a distinction between a northern region and Mesoamerica, which in turn can be subdivided into “East”, “Center”, “West” and “Southeast”. It is noteworthy that this geographical description matches the cultural area classification. The fact that these two areas overlap would seem to indicate that a combination of geographical distribution and relative isolation was at the origin of the blossoming of these indigenous cultures. The language parameter was found not to influence the distribution of genetic markers The distribution of populations in the MDS plot  providesThe four networks feature a star pattern, indicative of populations that have experienced processes of explosive growth. In some of the four central nodes of the four haplogroups, eight populations are represented, which confirms single, unique origin for all of theml. A global analysis of the network information enables us to propose a general diversification model in Mesoamerican indigenous populations. The node defined by variant C16295T in B2 network, represented by Nahua, Otomí Valle and Tepehua, in addition to the absence of haplotypes belonging to Otomí Valle suggests that this node may be a product of admixture in the Sierra Madre Oriental. This information is also confirmed by the high number of derived haplotypes shared by the Sierra Madre populations. The G16274A variation in D1 network (4145.85 YBP) defines an exclusively Otomí node, the first ethnic group differentiated from the common gene pool. The presence of a unique Pima and Mayo haplotype dated to 2984.49 YBP (T16295C polymorphism of C1) which does not include the Nahua people and the polymorphism A16182C (4064.55 YBP) in B2 marks the possible time range for the separation of the precursors of the Aztec culture, that is to say the Nahua, and the northern groups. The two central nodes in A2 are composed of the same groups with one exception: the Maya population found in the principal node but not in derivatives. This gives scope to the suggestion that when the C16111T variant appeared, separating both nodes (dating to 3960.72 YBP), the Mayan group was already differentiated from the rest of the population pool. The network information is not detailed enough to provide information on the history of the Huichol and Tepehua groups. In fact, virtually no haplotypes of these populations appear outside the central node, which would indicate they are of a relatively recent origin. Only a few B2 variants typical in Tepehua date the origin of this population to the Middle Classic period 3071 YBP.The barrier software result is consistent with the descriptive information about haplogroup distribution. The clearest genetic boundaries were detected in the Pima population, although this frontier extends northwards, separating the Zuni from other northern populations. Another deep limit is detected in the southern region, isolating the Maya from the remaining Mesoamerican populations. The next frontier in intensity divides the Mesoamerican region into two strands, the Atlantic and Pacific regions. Finally, note that lowest-intensity genetic boundaries were detected in Oaxaca populations relative to other central highland populations located in the Center of the Mexican territory.The case of the Pima indigenous group is interesting since some results (MDS and barriers) show them to be genetically distinct. There are several possible explanations for these differences. One is that they may be the result of a sampling problem. A more plausible option, however, is that the Pima group is more heterogeneous than had previously been considered. This heterogeneity could be due to admixing with other groups in a post-colonization period, as has been detected in other groups in the New Continent Although it was not the objective of this study, the data point to a late colonization of the continent by Asian populations, and a shared, single origin for all of them. The first groups that came to the Mexican highlands colonized the region in a process of expansion and rapid growth in accordance with a model of settlement, expansion and differentiation. This process of fragmentation or tribalisation occurred mainly on the basis of geographical parameters but in the process of expansion did not necessarily follow a north-south axis Geographical isolation was not absolute, however, since gene flow has been detected in pre-Hispanic periods The differences of demographic histories between Aridoamerica and Mesoamerica are the result of very different survival strategies; in the north a hunter-gatherer strategy and in the center and southeast a sedentary agriculture, with livestock, trade and sometimes complex societies.The Otomí population was the first of these populations to separate from the original gene pool and supports the hypothesis that the Otomí culture is one of the oldest in Mesoamerica Gene flow between population and admixture with European people was detected in the region of the Sierra Madre Oriental. This result shows that orographically complex regions can be permeable to gene flow as long as certain precise circumstances exist, in this case a natural passageway between two regions. Anyway it should be remembered that the mtDNA only provides information about femenine history and that in America, male and female migrations were different, as shown by studies based on Y chromosome markers We have detected a high intra-population heterogeneity in Pima and Huichol groups. This result shows that the genetic diversity of the indigenous population in pre-Hispanic times was greater than currently. It is also possible that many indigenous groups were considered and classified as the same group when they were in fact different groups sharing the same geographical area. On the other hand, the heterogeneity of the Nahuas supports the idea that this group is composed of genetically distinct cultural groups absorbed by the Aztecs.Apart from the well-known limitations of mtDNA in reconstructing the history of our species, we consider it an appropriate tool with which to address the reconstruction of the history of mankind. The study of the mtDNA control region and detailed and thorough sampling are powerful tools for investigating our past.In order to gain an accurate estimate of genetic variability, a careful selection of the indigenous communities was undertaken . Each paParticipants completed a survey to collect data relating to their origins and predecessors. Only those people who spoke an indigenous language were included in the study, with all four grandparents belonging to a particular group and coming from the selected region.The D-loop or control region was sequenced for all 605 individuals using the Applied Biosystems BigDye Terminator kit (v1.1), in two or three fragments depending on the samples between positions (16024 and 576 following CRS).Amplified fragments were analyzed on Applied Biosystems genetic analyzers, ABI PRISM 3100 Genetic Analyzer and ABI PRISM 3130. All sequences are available from Various statistical analyses focused on the 600 control region mtDNA sequences of the eight indigenous groups were performed. The results were then compared with those available in the scientific literature http://www.mitosearch.org); PhyloThree A first descriptive approach involved the allocation of haplogroups K and θSK expresses better the recent female population effective size ST distances ST distances, and using XL-STAT 3.2 software, an MDS  was plotted in order to establish approximate relationships among populations. The Monmonier’s maximum difference algorithm Subsequently, the sequences were aligned and edited when necessary with the Bioedit 7.0.5 sequence alignment editor Figure S1Median-joning network of the mtDNA sequences belonging to the A2 haplogroup in the indigenous populatons. (numbers represent the position of mutation defining a haplotype).(TIF)Click here for additional data file.Figure S2Median-joning network of the mtDNA sequences belonging to the B2 haplogroup in the indigenous populatons. (numbers represent the position of mutation defining a haplotype).(TIFF)Click here for additional data file.Figure S3Median-joning network of the mtDNA sequences belonging to the C1 haplogroup in the indigenous populatons. (numbers represent the position of mutation defining a haplotype).(TIF)Click here for additional data file.Figure S4Median-joning network of the mtDNA sequences belonging to the D1 haplogroup in the indigenous populatons. (numbers represent the position of mutation defining a haplotype).(TIF)Click here for additional data file.Figure S5ST distances of twenty populations from Mesoamerica and Aridoamerica (Stress = 0.145).Multidimensional scaling (MDS) based on F(TIF)Click here for additional data file.Table S1Sample geographic and demographic information. The percentage of indigenous population has been calculated in reference to the municipality (INEGI 2000).(DOCX)Click here for additional data file.Table S2Number of samples (N) and haplogroups found in the eight populations.(DOCX)Click here for additional data file.Table S3Diversity indices for mtDNA control region (16024-576) sequences and lineages in twelve indigenous populations. N, sample size; K, number of different sequences; S, number of polymorphic sites; Ĥ, sequence diversity; π, nucleotide diversity; θ, mean number of pairwise differences between sequences; D, Tajima test of selective neutrality (s.d. standard deviation).(DOCX)Click here for additional data file.Table S4Theta estimators for the twelve populations .(DOCX)Click here for additional data file.Table S5AMOVAs based on different classification criteria. We have used the mtDNA control region of twenty populations from Mesoamerica and Aridoamerica. In bold the eight populations studied in this paper. *Significance level 0.05.(DOC)Click here for additional data file.Table S6ST calculated for twenty populations from Mesoamerica and Aridoamerica.F In bold the eight populations studied.(DOC)Click here for additional data file.Table S7Number of haplotypes shared between the twenty populations. Lower hemimatrix shows the number of haplotypes shared in the control region, upper hemimatrix shows the number of haplotypes shared in HV-I region. In bold the eight populations studied in this paper.(DOC)Click here for additional data file.Table S8Pearson correlation coefficients (r) and their significance (p) between geographic distance from Bering Strait to the communities and genetic diversity indices. K, number of different sequences; S, number of polymorphic sites; Ĥ, sequence diversity; π, nucleotide diversity; θ, mean number of pairwise differences between sequences.(DOC)Click here for additional data file.Table S9Control region haplotypes distribution in the eight populations studied.(XLSX)Click here for additional data file.