Grapholita ( = Cydia) molesta is a key fruit pest globally. Despite its economic importance, little is known about its population genetics in its putative native range that includes China. We used five polymorphic microsatellite loci and two mitochondrial gene sequences to characterize the population genetic diversity and genetic structure of G. molesta from nine sublocations in three regions of a major fruit growing area of China. Larval samples were collected throughout the season from peach, and in late season, after host switch by the moth to pome fruit, also from apple and pear. We found high numbers of microsatellite alleles and mitochondrial DNA haplotypes in all regions, together with a high number of private alleles and of haplotypes at all sublocations, providing strong evidence that the sampled area belongs to the origin of this species. Samples collected from peach at all sublocations were geographically structured, and a significant albeit weak pattern of isolation-by-distance was found among populations, likely reflecting the low flight capacity of this moth. Interestingly, populations sampled from apple and pear in the late season showed a structure differing from that of populations sampled from peach throughout the season, indicating a selective host switch of a certain part of the population only. The recently detected various olfactory genotypes in G. molesta may underly this selective host switch. These genetic data yield, for the first time, an understanding of population dynamics of G. molesta in its native range, and of a selective host switch from peach to pome fruit, which may have a broad applicability to other global fruit production areas for designing suitable pest management strategies.The oriental fruit moth  In insect field populations, genetic diversity and genetic structure can be affected by various factors, such as range of origin as opposed to invaded regions, geographical distance and host plant species Grapholita ( = Cydia) molesta Busck (Lepidoptera: Tortricidae), a key pest of rosaceae fruit trees wordwide An insect species of global importance is the oriental fruit moth, G. molesta is considered limited based on estimation of its flight range G. molesta individuals have an ecological advantage when they remain and oviposit within the orchard where they emerge, unless environmental change triggers displacement of some individuals G. molesta have been reported twice from invaded countries. In South Africa, G. molesta populations collected from closely situated orchards (<1 km) could be distinguished G. molesta on different continents, and documented that populations of this species were geographically structured on a continental scale Dispersal of G. molesta in its putative range of origin and its genetic structure in a key fruit growing area of China, as well as the consequence of seasonal dynamics with host switch from primary to secondary hosts. We analyzed samples collected from peach throughout the season and from pome fruit after host switch in the late season, using a combination of microsatellite data and mitochondrial gene sequences. We hypothesized that populations will show (a) a number of typical characteristics reflecting a native species, (b) a genetic structure between different regions and possibly also between sublocations within these regions, reflecting the limited flight capacity of this moth species, and (c) little genetic structure across the season irrespective of the host infested.Population genetic studies can reveal the micro-evolution and ecological adaption strategies of an insect pest in agroecosystems No specific permissions were required for the described field studies for this wide spread agriculture pest. We confirm that the locations were not privately owned or protected in any way. The field studies did not involve endangered or protected species.nd to 3rd instar larvae were collected per population . Extraction was performed according to the bench protocol for animal tissues. DNA was eluted in TE buffer and stored at −20°C.Gm01, Gm03, Gm04, Gm05 and Gm07) had to be rejected either because the loci were monomorphic , or because of the presence of stutter peaks and/or imperfect repeats (Gm03 and Gm07) in genotyping results of many samples. Five loci  that were highly polymorphic were selected for further analysis. The forward primer of each pair was labeled at the 5′ end with FAM fluorescent dye 2, 0.2 µM of each dNTP , 0.5 µl of each forward primer (2 µM), 2 µl of each reverse primer (2 µM), 2 µl M13 primer (2 µM), 1.0 U Taq polymerase , and 2 µl genomic DNA (10–30 ng/µl). Amplification comprised a 10 minutes initial denaturation at 95°C, followed by 35 cycles of 30 seconds at 95°C, 45 seconds annealing at a primer-specific temperature, and 45 seconds at 72°C, then 10 cycles of 30 seconds at 95°C, 45 seconds at 53°C and 45 seconds at 72°C, and a final step 10 minutes at 72°C. To examine the length and genotype of the amplified PCR products, an ABI3730XL automated DNA sequencer  and GENESCAN version 4.0  were used.Ten previously reported microsatellite markers 5′-CAACATTTATTTTGATTTTTTGG -3′) and TL2-N-3014 (5′-TCCAATGCACTAATCTGCCATATTA -3′) 5′-ATGGCAGATTATATGTAATGG-3′) and TK-N-3785 (5′-GTTTAAGAGACCAGTACTTG-3′) 2, 0.2 µM of each dNTP , 3.0 µl of each oligonucleotide primer (2 µM), 2 µl genomic DNA (10–30 ng/µl). The thermal profile used a 10 minutes initial denaturation at 95°C, followed by 35 cycles of 30 seconds at 95°C, 45 seconds annealing at a primer-specific temperature (48°C for COI and 57°C for COII), 45 seconds at 72°C, and a final step of 15 minutes at 72°C. After verification via gel electrophoresis, the PCR templates were purified and then sequenced in both directions using the same primer pairs on an ABI3730XL automated DNA sequencer .The 740 bp mitochondrial cytochrome oxidase subunit I (COI) was amplified with the primer pairs C1-J-2183 (oH) and expected heterozygosity (eH), inbreeding coefficient (ISF), the genotypic linkage disequilibrium, deviations from Hardy-Weinberg equilibrium, and the Hardy-Weinberg equilibrium P values. ADZE  version 1.0 We used MICRO-CHECKER version 2.2.3 K) was determined by calculating the probability of the data given a certain prior value of K. Markov chain Monte Carlo (MCMC) was carried out using 20 replicate runs of 1 000 000 iterations following a burn-in period of 50 000 iterations for each K-value from 1 to 10. The most likely value of K was estimated according to Evanno et al. (2005) Population structure from microsatellite data was analyzed using STRUCTURE version 2.3.3 STF), and significance levels were assessed  using 10,000 permutations of diploid multilocus genotypes between samples t test at the significant level 0.05 was used to test whether differentiation between pome fruit populations was less than differences between pome fruit and peach populations; pairwise STF values between pome fruit populations were compared to pairwise STF values difference between pome fruit and peach populations in the test.We used ARLEQUIN version 3.5.1.2 For AMOVA, samples were grouped according to three models: (A) ‘comparison of variance among peach populations from different regions’. In this model, 13 populations from peach were divided into three groups by geographic regions: (i) TaiAn ; (ii) WeiFang ; (iii) YanTai (7YFS-Ph and 8YXS-Ph). (B) ‘comparison of variance among pome fruit populations from different regions’. In this model, genetic differentiation of four pome fruit populations from different regions were separated into two groups according to geographic regions: (i) TaiAn (8TCL-Ap); (ii) YanTai . (C) ‘comparison of variance among populations from peach and pome fruits. In this model, all 17 populations were divided into two groups by the types of host plants: (i) peach ; (ii) pome fruit .STF/(1- STF) of microsatellite data and the geographic distance (lnKm) were compared using the Mantel test with10,000 permutations In order to test for isolation by distance (IBD), the matrices of genetic distance Nh), nucleotide diversity (π) and haplotype diversity (Hd) were calculated using the program DnaSP version 5.0 The obtained mitochondrial DNA sequences were aligned using CLUSTAL_X version1.8 To study the genetic relationships among haplotypes, we used the program NETWORK 4.6.1 P<0.05) was found of all possible combinations after Bonferroni correction, which suggests that the five loci represented independent information across all the samples. The frequency of null alleles ranged from 0.13 to 0.28, which are similar values to previously reported ones from this tortricid pest We genotyped 399 individuals at five microsatellite loci. Population statistics for the 399 analyzed individuals of 17 investigated populations are given in In all the 399 individuals analyzed, we obtained 385 COI (740 bp) and 396 COII (700 bp) gene sequences, and observed 34 haplotypes for the COI gene (GenBank accession numbers are KF552033 to KF552066) and 38 haplotypes for the COII gene (GenBank accession numbers are KF551995 to KF552032).G. molesta populations sampled in China  and 28 (4%) variable characters, respectively, and no indels were observed in either of the two gene regions. As is common for an insect mitochondrial gene, the base composition of each of the two genes was biased toward As and Ts (76.4% of COI and 72.7% of COII). The COI gene covered 28 variable sites, 18 of which were parsimony informative, whilst 13 of the 21 polymorphic sites in COII gene were parsimony informative.Five of the observed 34 Hd) ranged from 0.63 to 0.95 for COII, and from 0.34 to 0.87 for COI (Nh) of COI over all populations was 5.06, ranging from 3 to 10, and for COII, the mean number of haplotypes was 8.12, ranging from 5 to 14. The average nucleotide diversity (π) of COI was 0.14%, ranging from 0.11% to 0.24%, for COII the mean nucleotide diversities (π) was 0.26%, ranging from 0.13% to 0.38% . Cluster 2 comprised all four populations sampled from peach at two sublocations in the WeiFang region . Cluster 3 was mainly made up of two populations sampled from peach in the YanTai region . Cluster 4 primarily contained individuals from the populations sampled in the late season from apple and pear in several regions .The most likely value of To assess whether the four clusters obtained by Bayesian clustering analysis using Structure were genetically distinct, we further analysed the relationship among populations using NJ tree STF analysis was calculated for each pair of populations over five microsatellite loci with values ranging from 0.021 to 0.255. The significance of pairwise STF was determined with a Markov chain analysis. The results showed significant differentiation between all pairs of populations with the exception of one pair (6TYT-Ph and 7TYT-Ph) (t test results showed that the differentiation among pome fruit populations (mean pairwise STF = 0.022±0.0002) was less than the differentiation between pome fruit and peach populations (mean pairwise STF = 0.050±0.002) with a P value of 0.010 .Pairwise 7TYT-Ph) . The twor value of 0.363 (P = 0.005) based on all 13 populations from peach, indicating that there was a significant isolation-by-distance effect present among different geographic populations from this host. In contrast, when the four populations from the pome fruit hosts were added, this correlation between genetic distances and geographic distances was no longer significant .The Mantel test provided an A total of 85 haplotypes were obtained with the combination of COI and COII genes. The network of the haplotypes are shown in G. molesta COI and COII haplotypes based on p-distance is shown in NJ tree of combined AMOVA results for the three models are shown in G. molesta populations from three regions of a main fruit growing area in China, using microsatellite and mitochondrial DNA markers. High genetic diversity was documented in these moth populations. All populations from peach were geographically structured. Whereas genetic structure of populations from peach was similar throughout the growing season within each region, populations collected from apple and pear in the late season surprisingly differed in genetic structure from populations collected from peach.We investigated the genetic diversity and genetic structure of 17 G. molesta. The mean number of alleles per locus (AN) averaged 10.4 over all populations, which was about twice as high as reported for G. molesta populations that invaded Italy (average 5.8) G. molesta, which has the flight ability to move between non-contiguous orchards but not over long distances G. molesta's origin.This study provided an extended sample analysis from the putative native region of G. molestaCydia pomonellaG. molesta populations within regions, as is also the case for cooperatives in Italy Populations from peach were geographically structured in the three sampled regions of China, as documented by the well defined clusters formed by populations from each region based on microsatellite markers. There was weak but significant correlation between geographical distance and genetic distance among these populations. Our microsatellite data provided stronger information on geographical population structure than mitochondrial data did in the NJ tree of the combination of COI and COII haplotypes. This difference may be due to the diploid nature of microsatellites as opposed the haploid nature of mitochondrial markers, as was concluded in a review on population genetics information obtained from these markers P = 0.216) was in a heavily infested abandoned orchard, which seemed to have provided a particularly stable environment for the moth. Otherwise, environmental conditions in single orchards sufficiently changed between sampling times to allow for detection of significant genetic variance. Such environmental changes include the considerable changes in emitted volatile blends from host trees with progressing season, which elicit altered behavioral responses of the oriental fruit moth Genetic variance among all populations sampled existed . The onlG. molesta moths, and this effect is even stronger when ambient temperature is still high G. molesta, there are different olfactory genotypes, and this natural genetic diversity facilitates host switch Populations sampled in the late season from the pome fruit apple and pear differed in genetic structure from populations sampled from the primary host peach, and they form a single clade exclusively comprising populations from pome fruit. Three factors might have contributed to this remarkable finding: the increased flight performance of the moth in the late season The oriental fruit moth is not only a key pest of rosaceae fruit trees in its native range in Asia, requiring major pest management input G. molesta is high in its region of origin. Significant genetic structure exists among populations from peach in the different regions. The weak flight capacity of the moth underlies the pattern of genetic structure observed. Contrary to our original expectation, host switch of the moth results in different genetic structure of populations on the secondary host plants, to which the recently detected different olfactory genotypes of the moth likely contribute. Design and implementation of sustainable pest management strategies could therefore be adjusted to the population genetics and dispersal of this pest insect.Genetic diversity of


Sus scrofa) genomic diversity within and between sites and compared them with human and other livestock. The current Porcine 60K single nucleotide polymorphism (SNP) panel has an average SNP distance in a range of 30 - 40 kb. Most of genetic variation was distributed within populations, and only a small proportion of them existed between populations. The average heterozygosity was lower in pig than in human and other livestock. Genetic inbreeding coefficient (ISF), population differentiation (STF), and Nei’s genetic distance between populations were much larger in pig than in human and other livestock. Higher average genetic distance existed between European and Asian populations than between European or between Asian populations. Asian breeds harboured much larger variability and higher average heterozygosity than European breeds. The samples of wild boar that have been analyzed displayed more extensive genetic variation than domestic breeds. The average linkage disequilibrium (LD) in improved pig breeds extended to 1 - 3 cM, much larger than that in human (~ 30 kb) and cattle (~ 100 kb), but smaller than that in sheep (~ 10 cM). European breeds showed greater LD that decayed more slowly than Asian breeds. We briefly discuss some processes for maintaining genomic diversity in pig, including migration, introgression, selection, and drift. We conclude that, due to the long time of domestication, the pig possesses lower heterozygosity, higher ISF, and larger LD compared with human and cattle. This implies that a smaller effective population size and less informative markers are needed in pig for genome wide association studies.We have reviewed the current pig ( Even so a large number of local breeds are maintained especially in China. It is commonly held that current European and Asian pig breeds are domesticated from different ancestors that might have different genome wide diversity and extent of linkage disequilibrium (LD). Worldwide, domestic pigs can be divided into two main clades: Asian- and European-types, which diverged from each other around 58,000 years ago. The European wild boars were domesticated at least in the 4th millennium BC and then rapidly spread throughout Europe , 2. AsiaInitial genetic studies in the pig mainly focused on investigating the genetic diversity of a breed at relatively small numbers of individual sites, which is essential for sustainable management of genetic resources for future utility. With the identification of the first useful DNA marker HAL1843) in the ryanodine receptor gene for marker assisted selection 3 in the , 6 or byThe development of high throughput genomic techniques provides us with an opportunity to assess genome wide diversity and its structure. Since the earlier draft sequence published for human 2001) and cattle (2009), an increasing number of studies have aimed at studying genomic diversity and LD in human and cattle -16. For  and cattet al. [The focus of this review is on pig, however, we will compare the pig with results from human as well as other livestock species. We begin by discussing pig genomic diversity within sites and then addressed the genomic diversity among sites described by LD. In each case, we briefly compare pig genomic diversity with human and other livestock. The mechanisms for maintaining genomic diversity in pig are briefly discussed, including the factors of migration, introgression, selection and drift. A comprehensive review on population genomic diversity from the perspective of evolutionary theories is detailed in this volume by Hu et al. . 2.2.1Pig genomic diversity within populations is quite variable. Observed heterozygosity ranged from 0.35 to 0.60, with an average ~ 0.5 across 17 autosomal chromosomes in 11 European pig breeds . These vA large number of SNPs are currently available for most species, such as human , cattle These SNPs were used to construct a panel using the Illumina platform, 64,232 SNPs were decoded and selected [2.2.1 [Since pig breeds from Asian and European populations were domesticated from different ancestors, they are expected to have different genetic diversities. One suggestion is that Asian wild boars had higher genetic diversity compared to European wild boars . The nuc1 , 25-26. Nei’s genetic distances between populations in Table 2 [Nei’s genetic distance was 0.531, with a range from 0.194 to 1.188. In European populations, the average genetic distance was 0.495, ranging from 0.163 to 1.122. Higher average genetic distance was observed between European and Asian clades (1.434), which was nearly three times larger than those within populations. The smallest value was even up to 1.119 between Duroc and Tibet pigs [et al. [The genetic distance among populations is very varied reflecting the differences in domestication model and history in the two regions. We have summarized the  Table 2 , 25, 26.STF from the literature [2. The average STF value (0.257) between Asian and European populations was much higher than those within populations. When compared within the two different regions, the overall genetic differentiation for Asian populations was ~ 0.227, ranging from 0.182 to 0.294, much higher than that for European (0.134). The STF value between domestic and wild boars for Asian (0.29) was also higher than that for European (0.194) [STF was between Asian and European breeds , and the smallest was observed within European breeds  [Most genetic variation occurred within populations rather than between breeds from Asia and Europe. A summary of terature , 25, 26 ropean 0.4 [19, 25 (0.194) . A recenrkshire) . All Asian and European pigs were closely related in terms of their maternal lineages, but they were different from each other. In wild boars, the average percentage sequence divergences calculated by mtDNA were 0.2370 within Chinese population, 0.3718 within European, and 0.5603 between Chinese and European. In domestic populations, the pairwise nucleotide sequence divergence was much lower (0.0056) between Chinese and European breeds . Asian d2.3.3. It shows that heterozygosity in pig was smaller than that in human and other livestock, but more flexible. Genetic variation within populations and genetic distance were much larger in pig than in human and other livestock. A comparison of pig genomic diversity with human and other livestock is summarized in Table STF) was lower in human than in pig . In human, STF was 0.05 - 0.13 for autosomal SNPs [Alu insertions polymorphism [et al. [STF).The average expected heterozygosity in human ranged from 0.7 to 0.9 , which wmal SNPs -43, ~ 0.morphism . Biswas  [et al.  indicate3). Recent studies [ISF) ranged from 0.03 to 0.32. The observed and expected heterozygosities were at ranges of 0.42 - 0.67and 0.61 - 0.73, respectively. Among different goat breeds, Nei’s genetic distance differed from 0.067 to 0.830. The genetic diversity between breeds (STF) ranged from 0.012 to 0.200. The overall proportion of genetic differentiation among breeds was around 0.14, such as 0.02 in Guadarrama goat [The genetic diversity was more extensive in goat than in pig . The observed and expected heterozygosities in cattle were at the ranges of 0.47 - 0.74 and 0.45 - 0.78, respectively. While the genetic diversities both within (ISF) and between breeds (STF) seem to be a slightly lower in cattle than in pig. Nei’s genetic distance was much larger in pig (0.163 - 1.794) than in cattle (0.015 - 0.382). Compared to pig, cattle genetic diversity has been more widely studied. Genetic diversity in Chinese breeds was higher than that in European breeds. This is similar to that found for the pig. In Chinese cattle breeds, the mean heterozygosity was at a range of 0.69 - 0.76, and Nei`s genetic distance ranged from 0.025 to 0.352 [ISF) and between (STF) populations were slightly lower. Nei’s distance was at a range of 0.084 - 0.382 [The average heterozygosity in cattle is slightly higher than that in pig  than for SSC15 (0.15 - 0.19) [D´ was higher on SSC7 than on SSC4, and LD decreased faster with the marker distance on SSC4 than on SSC7 [chromosomes 10 and 12 had the lowest average LD, chromosomes 1, 13, and 14 had the highest value [2r). When considering longer SNP pair distance (3 Mb), differences in LD between different chromosomes were even larger up to 0.2  [2r. The significant LD variation among different regions was mainly due to the strong artificial selection on economic traits.Within pig populations, a wide range of LD was observed on different chromosomes. With 29 and 5 microsatellite markers located on SSC15 and SSC2 in different lines, respectively, LD in terms of  - 0.19) . By inve on SSC7 . Using tst value . Using t2 value) . It indiD´ and 2r are widely used to evaluate inter-site associations in livestock. D´ strongly depends on allele frequency, and it decreases as the MAF increases. 2r is more influenced by linkage distance. By using ~ 4,500 autosomal SNPs in more than 6,000 pigs from six commercial lines [D´ was observed in case of MAF as 0.067. The 2r values indicated that the largest LD was in the most tightly linked group. As the linkage distance increased from 0.1 to 40 cM, the average 2 rreduced from 0.371 to 0.008. The observed average 2r began to rapidly decrease when the linkage distance approached to 3 cM. Amaral et al. [2r being 0.3 as a threshold. Marker density and sampling size are the main factors that affect the accuracy of LD evaluation, which further influences the design of whole genome wide association studies. Nsengimana et al. [D´ value was feasible in commercial pig populations for genome-wise association. Du et al. [et al. [al lines , the higl et al.  used higa et al.  suggesteu et al.  recommenu et al.  up to 40u et al.  with com [et al.  predicte3.2.2r = 0.3). However, for Chinese breeds with a similar sample size, the SNP spacing was 0.005 cM, and 500,000 SNPs per individual would be required [The genome wide inter-site associations in terms of LD differ among different populations. Modern breeding programs increased the extent of LD and caused significant differences between European and Chinese pig breeds. LD decayed more rapidly in Chinese breeds than in European breeds, indicating that the extent of LD was smaller in Chinese breeds than in European breeds. In European breeds, LD extended up to 2 cM, and large haploblocks could be up to 400 kb; whereas in Chinese breeds, the extent of LD was smaller (0.05 cM) and generally did not exceed 10 kb , 58. Chirequired . This larequired . 2r > 0.3 and 2 r> 0.2 were 49% and 57%, respectively; while the corresponding percentages for Yorkshire were 52% and 60%. At the average useful 2r > 0.2, the SNPs density in Landrace and Yorkshire extended to 1.0 and 1.5 Mb, respectively, which resulted in a smaller effective population size (eN) for Yorkshire (55) than for Landrace (80). Similar eN were also obtained in American Landrace (74) and Berkshire (77), but higher in Hampshire (109) and Duroc (113) [D´ was also highly significant in Yorkshire, Large White and Landrace populations. The lowest D´ was in Large White breeds, and the highest was in Duroc breeds [Contrary to Chinese breeds, European breeds showed significant differences in LD . By usinoc (113) . This, tc breeds . 3.3.4. It shows that, at the same 2 rthreshold, LD was significantly larger in pig than in human, but much smaller than in sheep. Compared with cattle, LD was much larger in pig at threshold of 2 r= 0.2 but similar to cattle at threshold of 2 r= 0.1. A comparison of LD among pig, human, and other livestock is summarized in Table 3.3.1.et al. [Due to a smaller effective population size and the stronger selection that has occurred in livestock, LD was significantly greater in pig (1 - 3 cM) than in human. The average LD in human was up to ~ 30 kb with a high variability, depending on the different populations and marker density used. Computer simulations and empirical data suggested that human LD extended only 3 - 5 kb for common disease SNPs, meaning that approximately 500,000 SNPs at least was needed for whole-genome studies , 61. Latet al.  analyzedeN in different populations (see above for examples in pigs). For the SNP-pair interval less than 100 kb across all of the genome, thee N estimate was ~3100 for western European, Japanese and Chinese, and ~7500 for the Yoruba population [eN and fewer informative markers required for genome wide studies, at least for the European pig populations.The LD information is very useful for estimating pulation . This re3.3.2.2r > 0.2 [et al. [2r was 0.14 at marker distance of 100 kb. It indicated that at least 50,000 SNP markers were required for whole genomic selection. McKay et al. [2r was 0.15 - 0.2 at a physical distance of 100 kb SNP loci apart, indicating that 30,000 - 50,000 markers were needed to conduct whole genome association studies. When microsatellite markers were used with four cattle breeds to estimate LD [2r value across the populations was 0.16 at a range of 0.11 - 0.22. For the distance greater than 50 cM, the average 2r declined to 0.07. Compared with cattle, pig LD was much stronger, especially for the useful 2r > 0.2.Generally, cattle have larger LD than human, but smaller LD than pig. LD in cattle was at a range of 40 - 100 kb at the threshold of r2 > 0.2 , 15, 67. [et al.  comparedy et al.  reportedimate LD , at a ma2r and 8.2 Mb for D´. It indicated that at least 75,000 SNPs and a sample size of 75 or 400 would be required for whole genome association study. Qanbari et al. [2r) was observed for SNPs less than 100 kb apart. For American Holstein cattle, 2r was much larger (0.59) at the same marker distance (100 kb) [2r > 0.8) in genomic regions with marker distance less than 50 kb. LD for SNP pair intervals 100 kb apart (2r = 0.14) was similar to that in Holstein-Friesian cattle. Compared with pig, LD among populations varied less in cattle. This is in agreement with the common notion that a larger proportion of genetic variation exists within than between populations in most organisms. Extensive LD in cattle varied a little among different populations. In Holstein-Friesian cattle, by analyzing 1,566,890 syntenic and 365,400 non-syntenic SNP pairs that cover all autosomes , the sigi et al.  used Ill(100 kb) . Kim & K(100 kb)  also rev3.3.3.D`) was observed for syntenic marker-pairs with the interval less than 60 cM. By using 490 markers in Soay sheep [D´ > 0.2 as for marker-pair distance < 10 cM. Meadows et al. [D´ value between non-syntenic marker pairs ranged from 0.266 (Poll Dorset) to 0.322 (Merino × Border Leicester). 2 rdecayed faster within Merino and Merino × Border Leicester. LD varied a lot among populations in sheep, similar to the results in pig.Compared with pig, LD in sheep seems much more extensive and flexible. High LD extending to 10 - 60 cM was observed in two domestic sheep breeds evaluated by microsatellite markers . Large fay sheep , 22% mars et al.  used 28 4.4.1.th centuries. When European wild boars were domesticated, they rapidly became the predominant lineage within European domestic swine [e N, subsequent migration can lead to a high level of polymorphism. While maternal introgression from European domestic pig has no or very little impact on Chinese breeds. That is why some European haplotypes were detected in some local breeds, but no European mtDNA haplotypes were detected [A significant differentiation in genetic diversity and LD exists between Asian and European pig populations. One reasonable explanation is that their domestications originated from different ancestors. However, they shared some important haplotypes with each other, which might come from extensive migration between populations and incomplete lineage sorting. Historic and demographic events revealed that Near Eastern pigs were definitely introduced into Europe during the 18 – 19ic swine . So the ic swine . There wic swine , 34, 35.4.2.Efficient artificial selection has a dramatic influence on livestock genome diversity and linkage disequilibrium. This effect depends on the direction, intensity, duration and consistency of selection over time. For a long time, in pigs selection focused on growth and fatness (representing demand for cheap lean meat). The availability of dense marker panels provides the opportunity to consider LD between regions and to search for signatures of selection as a means to identify causative mutations or other useful markers as has been done in several species. For example, two of the first QTL to be identified in the pig were on chromosomes 4 and 7. These include QTL for growth and fatness traits. However, despite significant efforts the genetic variation underlying these QTL has not been elucidated. The extent of LD for SSC7 was significantly larger compared to SSC4 . FurtherModern breeding practices and best linear unbiased prediction (BLUP) selection starting in the last century led to a rapid increase of genetic gain and large LD in pig breeds. This can cause inbreeding that increases LD but reduces genetic diversity. Taking Meishan for example, in order to increase the population size and reproductive performance, an inbred line of Meishan was developed in the late 1980s in a selection scheme. This may explain the lower genetic variability observed within Meishan  comparedSelection reduces genetic diversity on the next generation but enhances LD. Selection on one locus or multiple loci, will increase LD between the neighbouring loci and the selected locus. As a result of intensive artificial selection, livestock have a larger LD than human populations. In addition, selection can cause LD between unlinked loci that contribute to phenotypes undergoing selection . This co4.3.eN (bottleneck effects) and directional artificial selection. The reduction ine N can lead to lower genetic diversity. LD initially generated by genetic drift gradually reduces with time. The genetic drift effects are substantial in historic pig breeds. The development of European and Chinese pig breeds has been different and the study of these differences can provide additional insights on diversity and these processes. It appears that the differences described to date are the result of differences in the number of ancestral stocks as well as the differences in modern breeding systems. Selection is likely to have been stronger in Europe and N America than in Asia. European breeds had smaller effective population sizes, resulting in stronger LD [eN than Hampshire and Duroc breeds, producing different LDs for the same marker-intervals [Domestication in livestock essentially results in two processes: the decline in onger LD . Americantervals , 59. The5.Sus scrofa) and compared them with human and other livestock. It is concluded that genetic diversity is smaller in pig than in human and other livestock. A relatively larger genetic variation within populations (ISF) exists in pig compared with cattle and goat. A majority of genetic variation in pig was distributed within rather than between populations, similar to human and other livestock. A large genetic distance exists between Asian and European populations. Asian breeds harbour much larger variability than European breeds. For the different genetic diversity structures, the inter-site associations described by LD were much stronger in pig (1 - 3 cM) than in human (~ 30 kb) and cattle (~ 100 kb), but less than in sheep (~ 10 cM). LD varied largely among different populations in pig and sheep, but a little in cattle. This review confirms that, strong breeding and selection programs have occurred in pig for a relatively long time, which resulted in the relatively lower heterozygosity and higher ISF and larger LD, compared with human and cattle. This also implies that a smaller effective population size and less informative markers would be needed for pig whole genome association studies (at least for European populations). To some extent, this difference might arise from the joint effects of migration, selection, and drift during the process of pig domestication. In this review, we summarized genomic diversity within and between sites in pig (


Bashania fangiana) clonal populations with two different genet ages (≤30 years versus >70 years) at Wolong National Natural Reserve, Sichuan province, China. We generated AFLP fingerprints for 96 leaf samples, collected at 30 m intervals in the two populations, using ten selective primer pairs. A total of 92 genotypes were identified from the both populations. The mean proportion of distinguishable genotypes (G/N) was 0.9583 (0.9375 to 0.9792) and Simpson's index of diversity (D) was 0.9982 (0.9973 to 0.9991). So, two B. fangiana populations were multiclonal and highly diverse. The largest single clone may occur over a distance of about 30 m. Our results demonstrated that the genotypic diversity and genet density of B. fangiana clonal population did not change significantly (47 versus 45) with genet aging and low partitioned genetic differentiation was between the two populations (Gst = 0.0571). The analysis of molecular variance consistently showed that a large proportion of the genetic variation (87.79%) existed among the individuals within populations, whereas only 12.21% were found among populations. In addition, the high level of genotypic diversity in the two populations implies that the further works were needed to investigate the reasons for the poor seed set in B. fangiana after flowering.Amplified fragment length polymorphism (AFLP) fingerprints were used to reveal genotypic diversity of dwarf bamboo ( Populus tremuloides  Pteridium aquilinum (clone size up to 20 ha) et al. Low recruitment rates from seeds have been reported to be common features in clonal populations even if seed production is abundant Bashania fangiana is a perennial monocarpic species, which has both types of sympodial and monopodial rhizome systems in a genet. It grows up to 2.0 m in height, vigorously extend rhizomes, and often form dense populations in the mixed coniferous and broad-leaved and subalpine coniferous forest understoreys, north-western Sichuan province, China B. fangiana flowers once about 50–60 years or even more seldom B. fangiana often flowers so simultaneously over an extensive area and then dies. After mass death, its populations recover mainly by the development of seedling cohorts. When the seedlings reach full size stage, they extend rhizomes and produce culms, resulting in dense populations again B. fangiana is one of the staple food bamboos for the giant panda (Ailuropoda melanoleuca), which is crucial to maintain its survival and reproduction B. fangiana population, and no information such as number of clones in a population is available. Because rhizomes of B. fangiana grow leptomorphically, different genets are likely to mix with each other, but no genetic identification of culms of B. fangiana stands has been done.Dwarf bamboo B. fangiana in two 3.15 ha plots and to reveal the genetic structure of two B. fangiana populations with different genet ages (≤30 years versus >70 years). We addressed the following questions: (1) how many clones (genets) exist in two populations respectively; and (2) how large an area does a single clone occupy. In the complete absence of seedling recruitment, genotypic diversity  is constrained to the level of the initial cohort and can only decline over time B. fangiana population will decline over genet age.Amplified fragment length polymorphism (AFLP) B. fangiana, one of the staple food bamboos for the giant panda (Ailuropoda melanoleuca) and often forming dense populations in the mixed coniferous and broad-leaved and subalpine coniferous forest understoreys in natural reserve was used as investigated subject, and we confirmed that our studies did not involve endangered or protected species. In addition, no specific permission was required for these locations because our study was the general sampling experiment.This study was carried out at an open field  in Wolong National Natural Reserve, which is located in the southern slope of the Qionglai Mountains, Sichuan province, China. We obtained appropriate permissions from the Forestry Bureau of Wenchuan County, and assistance from the forestry workers for field study. In present study, −1 with 76.66% of annual rainfall occurring between May and September. There are different types of vegatation with a vertical distribution: evergreen broad-leaved forest below 1600 m; mixed evergreen broad-leaved and deciduous broad-leaved forest between 1600 and 2000 m; deciduous broad-leaved forest between 1500 and 2600 m; mixed coniferous and broad-leaved forest between 2000–2600 m; subalpine coniferous forest between 2600 and 3800 m; and interspersed shrublands and meadows above 3800 m. B. fangiana, Fargesia robusta and F. nitida are the main panda grazing species within pandas' habitat in Wolong National Natural Reserve The aim of the Wolong National Natural Reserve is to conserve giant pandas and their habitat. It is in the subtropical area with an average annual temperature 8.42°C; −1.34°C in January and 17.06°C in July. Precipitation averages 884.24 mm yearTsuga chinensis, Betula albo-sinensis. B. fangiana forms dense population in the forest understorey. So, the light, soil and topographic conditions et al seemed to be uniform in the research plots. One plot (N1) was established after the mass-flowered of B. fangiana in 1983 and its genet age was not more than 30 years old. The other (N2) was at least more than 70 years old . Within each plot, 48 leaf samples were collected at each intersection point on a 30×30 m grid and immediatly preserved in silica gel to prevent DNA degradation.In september 2012, two rectangular sampling plots (150 m×210 m) were chosen near the Wuyipeng Research Station in Wolong National Natural Reserve. The two plots belonging to mixed coniferous and broad-leaved forest, were dominated by MseI(M-) and EcoRI(E-) primers used in the selective amplification were identified as the most informative and reliable from a set of 256 primer pairs during a preliminary investigation  according to the manufacturer's instructions. Then, DNA samples were suspended in TE buffer. AFLP analysis consists of four steps: restriction digestion, adaptor ligation, preselective amplification and selective amplification and was conducted essentially as described by Vos et al. tigation . The amptigation , and thei(Ni-1)]/[N(N-1)]}, where Ni is the number of samples with genotype i and N is the total number of samples; and (iii) genotypic evenness min)/(Dmax-Dmin), Where Dmin = [(G-1)(2N-G)]/[N(N-1)] and Dmax = [N(G-1)]/[G(N-1)]. D ranges 0 in a population composed of a single clone to 1 in a population where every sampled individual represents a different clone. E ranges from 0 in a population where all samples represent different clones, or where one clone dominates and the other clones are represented by a single sample, to 1 in a population where all clones are represented by the same number of samples. All three measures of genotypic diversity were based on the 96 samples collected at each intersection point on the 30×30 m grid in the two populations.Samples showing the same multilocus AFLP phenotype were considered to be the same genotype. To determine the genotypic diversity  in the two populations with different genet ages, three measures were calculated following Ellstrand & Roose We also used POPGENE 1.31 to calculate four genetic diversity indices within each population, that are commonly used in the literature: the number of polymorphic loci, the percentage of polymorphic loci (PPB), the Nei's B. fangiana ramets samples. The observed numbers of bands by AFLP analysis were 202 using the ten selective primer pairs, each primer pair produced an average of 20.2 distinct bands. Excluding commom bands, ten primer pairs yielded a total of 111 polymorphic bands among these 96 samples. 107 were polymorphic in the N1 ramet population, and 91 were polymorphic in the N2 ramet population  with identical AFLP fingerprints were detected. The clonal structure of the two populations is depicted in B. fangiana was 0.9583 and 0.9982, respectively (N1 = 0.1463(±0.1750) and hN2 = 0.1229(±0.1704); IN1 = 0.2286(±0.2555) and IN2 = 0.1920(±0.2494), respectively. So, there was no great difference in genotypic diversity and genetic diversity between N1 and N2 populations.Among 96 ectively . The parThe results from our analysis of molecular variance (AMOVA) showed that a large proportion of the genetic variation (87.79%) existed among individuals within the populations, and that 12.21% resided among the populations. The genetic differentiation among the populations was significant (P<0.001) . ConsistB. fangiana population first clustered into one group, then clustered with other population. Only a small number of ramets  did not gathered together with other ramets in the same population (N1), which suggesting larger genetic variation compared with others in the population. Results of the UPGMA analysis also revealed that the high genetic similarity were shared between the two populations  were higher than the average values found in other clonal plants  B. fangiana is confirmed by the fact that both of the sampled populations were multiclonal, compared to an average of 62% in Ellstrand and Roose B. fangiana species had the levels of percentage of polymorphic loci (P = 54.95%) and Nei's gene diversity (h = 0.1428) similar to the average values calculated for populations of 120 wind pollinated outcrossing taxa  B. fangiana is monocarpic, often flower simultaneously and die subsequently over intervals of several decades B. fangiana is more like a typical ISR species in the ISR-RSR spectrum of Eriksson.Many clonal plant populations usually can be as diverse as those of non-clonal plant species Sasa senanensis were incurred by the environmental heterogeneity B. fangiana with a obligatory xenogamy breeding system, the genotypic diversity of its seedlings establishing a new population is expected to be high compared with other less out-crossed species. So, the maintenance of genetic diversity through time could be fostered by the particular reproductive system of B. fangiana. Bengtsson B. fangiana can only reproduction by asexual after established. Therefore, founder effects in B. fangiana might be an important mechanism for maintaining in a high level of neutral genetic variability.The genetic and genotypic diversity could be influenced by a number of processes  The alternative explanation is that the high level of genetic diversity is also maintaining through clonal growth. Clonal growth could retard the loss of genetic diversity within populations by several mechanisms In addition, the higher genetic diversity in our study might result from the higher power of resolution of the DNA-based molecular marker (AFLP), and sampling scheme  with which ramets are sampled also influences these estimates of genotypic diversity.B. fangiana by low Gst value 0.0571, which was supported by the AMOVA analysis that partitioned genetic variation among populations was only 12.21%. Many factors can determine the genetic structure of plant populations, including the reproductive biology B. fangiana populations might result from its extensive longevity, obligate xenogamy reproductive system and the large gene flow (Nm = 8.2639). UPGMA dendrogram and PCA analysis further exhibited that low partitioned genetic variation existed between the two populations which implied the high genetic similarity were shared between the two populations. This outcome could have reflected the possibility that the examined populations of B. fangiana still represented the abundant genetic variation of their ancestors which was supported by the high level of genetic diversity in the two populations. In fact, this tendency is reinforced by the naturally long vegetative phase and low seed set over the life cycle, which are typical features of woody bamboos Small genetic differentiation was detected in B. fangiana populations are not dominated by one clone or have any dominate genotype and we did not observe any mixed distribution of multiple clones. The largest size of a single clone might occurs a distance of about 30 m at least, which was consistent with a excavation study showing that the annual rhizome increments of B. fangiana was about 45 cm B. fangiana may be an obligate xenogamy plant B. fangiana was very high, but the seed setting rate was rare after flowering B. fangiana may not caused by the low level of genotypic diversity and other reason may need to be responsible for it. The pistil and stamen are not mature at the same time or the disadvantage of its own physical/physiological structure and morphology may contribute to the seed sterility B. fangiana, the further study may be needed.Table S1Binary matrix of AFLPs result for empirical AFLP analysis.(XLS)Click here for additional data file.


Liposcelis bostrychophila and L. entomophila (Psocoptera: Liposcelididae) are found throughout the world and are often associated with humans, food stores and habitations. These insects have developed high levels of resistance to various insecticides in grain storage systems. However, the population genetic structure and gene flow of psocids has not been well categorized, which is helpful to plan appropriate strategies for the control of these pests.The psocids Cytb) and ITS (ITS1-5.8S-ITS2) regions. In total, 177 individual L. bostrychophila and 272 individual L. entomophila were analysed. Both Cytb and ITS sequences showed high genetic diversity for the two species with haplotype diversities ranged from 0.154±0.126 to 1.000±0.045, and significant population differentiation  was also detected among populations investigated. A Mantel test indicated that for both species there was no evidence for isolation-by-distance (IBD). The neutrality test and mismatch distribution statistics revealed that the two species might have undergone population expansions in the past.The two species were sampled from 15 localities in China and analyzed for polymorphisms at the mitochondrial DNA (L. bostrychophila and L. entomophila displayed high genetic diversity and widespread population genetic differentiation within and between populations. The significant population differentiation detected for both psocids may be mainly due to other factors, such as genetic drift, inbreeding or control practices, and less by geographic distance since an IBD effect was not found.Both  Ne) Genetic variability is considered to be the foundation of evolution, and it can be affected by a variety of factors, such as mutation rates, effective population size, and gene flow Cytb) gene Mitochondrial DNA (mtDNA), due to its accelerated rate of evolution, simple maternally inherited, and lacking of recombination, has been used as a powerful and efficient way to study genetic diversity in insect populations at both the inter- and intra-specific levels, especially for cytochrome b (Liposcelis bostrychophila Badonnel and L. entomophila (Enderlein) are found throughout the world and are often associated with humans, food stores and habitations L. bostrychophila and L. entomophila are similar in morphology and often commix within the same ecosystems. As the closest free living relatives of parasitic lice, these two species can also transmit harmful microorganisms, including fungi and bacteria L. bostrychophila, L. entomophila has been found to be more commonly and more widely distributed in grain storage systems in China. Unlike the sexual reproduction undertaken by L. entomophila, L. bostrychophila is an obligatory parthenogenetic species and displays considerable adaptability to deal with local or temporary situations L. bostrychophila, parthenogenesis can maintain heterozygosity but with the loss of variation L. bostrychophila also shows a considerable degree of morphological and physiological variation both between and within clones The small, wingless psocids, L. bostrychophila has been investigated using allozymes L. decolor and cross-amplified in five other Liposcelis species of economic importance L. decolor in Australia L. entomophila. As little is known of the population genetics of L. bostrychophila or L. entomophila in China, we undertook a genetic study to investigate the genetic diversity, the population genetic structure and population demographic history of the two psocids using mitochondrial Cytb and ITS sequences.On the other hand, numerous studies have been conducted on psocid physiology, biochemistry, and molecular biology in China and worldwide No specific permits were required for the described field studies, and no specific permissions were required for these locations/activities. We confirm that these locations are not privately-owned or protected in any way and the field studies did not involve endangered or protected species.L. bostrychophila and 11 populations of L. entomophila were sampled at grain storage facilities from 15 locations in eight provinces of China from 2008 to 2010, covering three Chinese Animal Fauna Regions  (Approximately nine populations of n (SCR)] . All speCytb and ITS regions were amplified by PCR. A 485 bp fragment of the mitochondrial Cytb gene was amplified using the primer pairs CBF1 (5′-TATGTACTACCATGAGGACAAATATC-3′) and CBR1 (5′- ATTACACCTCCTAATTTATTAGGAAT-3′) 5′-CCGATTGAACGATTTAGTGAGGTCTT-3′) and 28SR1 (5′-TGCTTAAATTCAGCGGGTATTCTCG-3′) 2+, 10×PCR reaction buffer, ddH2O and 2 unit of Taq DNA polymerase . PCR reactions were performed on a Bio-RAD S1000™ Thermal Cycler  and consisted of an initial denaturation step at 94°C for 4 min, followed by 35 cycles of 94°C for 30 s, with annealing temperatures of 46°C (Cytb) or 60°C (ITS) for 50 s and 72°C for 1 min, with a final 10 min extension at 72°C. PCR products were visualized on 1.0% agarose gels under UV light and purified using a gel band purification kit .The Cytb genes were directly sequenced using PCR purified products from both forward and reverse PCR primers. For ITS sequences, the purified products were ligated into pGEM-T Easy vectors  followed by ampicillin selection. All the sequences were read on an ABI 3730 automated DNA sequencer . Sequences were assembled using Bioedit 7.0.9.0 Cytb), and JN828813–JN828948 (ITS).n), haplotype diversities (h), and nucleotide diversities (π) for the populations of each species were estimated using the software DnaSP 5.10.00 rg) and sum of square deviations (SSD)] were performed using Arlequin 3.1 rg value represents a population which has experienced sudden expansion DFSFs, Tajima's D are nearly zero in a constant-size population; significantly negative values signify a sudden expansion in population size, whereas significantly positive values indicate processes such as a population subdivision or recent population bottleneck The number of haplotypes  version 1.52 Pairwise Cytb haplotypes and ITS haplotypes of the two psocids, L. bostrychophila and L. entomophila, were performed with a Bayesian phylogenetic inference framework and maximum-likelihood (ML) method, respectively. L. decolor (JF440956 and JF742536), was used as an outgroup due to its close relationship with L. bostrychophila and L. entomophila. First, the coding region Cytb sequences were translated into amino acids for confirmation in MEGA 5.01 Cytb and ITS haplotypes sequence alignments were conducted with Clustal X Multiple Alignment Cytb sequences, transversions and transitions versus sequence divergence were plotted, to evaluate the possibility of sequence saturation in DAMBE 5.2.34 Cytb haplotypes, three partitioned Bayesian analyses were carried out and the best-fit model for 1st, 2nd, and 3rd codon position was HKY+I, TPM3uf+G, HKY+G, respectively. The TPM3uf+I+G substitution model was utilized for the ITS haplotypes data set. Phylogenetic trees for Cytb haplotypes data set were reconstructed using MrBayes v3.12 http://www.atgc-montpellier.fr/phyml/) Phylogenetic relationships between the Traditional phylogenetic methods assume that ancestral nodes are no longer present in the data set and that the evolution of the data set follows a bifurcating pattern. Intraspecific data sets usually have features, including the persistence of ancestral haplotypes, the existence of multiple descendant haplotypes and often low levels of sequence variation Cytb gene was aligned and analyzed from 251 individuals (including 103 L. bostrychophila and 148 L. entomophila) and no insertions or deletions were detected in any of the sequences. The aligned sequences of 10 populations of L. entomophila contained 55 segregating sites with 60 different polymorphisms representing 28 singleton sites and 27 parsimony informative sites. In comparison, the aligned sequences of eight populations of L. bostrychophila had 76 segregating sites with 78 different polymorphisms representing 37 singleton sites and 39 parsimony informative sites.A 433 bp fragment of the L. entomophila populations and 40 haplotypes in L. bostrychophila populations  and the lowest was 0.257   and the lowest was the SZ population (0.154) , and the nucleotide diversity of L. bostrychophila (π = 0.02801±0.00117) was higher than that of L. entomophila (π = 0.00778±0.00071), but the difference was not significant . However, the two clades (Clade Lb1 and Clade Lb2)  . However, the nucleotide diversity of L. bostrychophila (π = 0.00875±0.00341) was not significantly higher than that of L. entomophila either (π = 0.00360±0.00185) . For the CCR, both the haplotype and nucleotide diversities of L. bostrychophila  were higher than those of L. entomophila , but not significant .In total, 39 haplotypes were detected in ulations . Among L . The ove (0.154) . The hapade Lb2)  were conL. bostrychophila formed to a single well-supported clade designated as clade Lb, while all the haplotypes corresponding to L. entomophila formed another single well-supported clade designated as clade Le  . AMOVA showed that 75.14% of the variation was among populations and only 24.86% was partitioned within populations. When performing a three-level AMOVA to test for structure between SWCR and CCR, we also detected a high genetic structure  between these two groups with a large portion of total genetic structure (67.15% was partitioned among groups). A small portion of the total genetic structure (15.88%) was apportioned among populations and was also significant within a region . For L. entomophila, a two-level AMOVA test for structure among populations showed high genetic structure  with slightly more of the total genetic structure apportioned among populations (56.65%). When performing a three-level AMOVA to test for genetic structure among groups, we found that population genetic structure was significant  with a little more portion (36.84%) of total genetic structure. Thus, both of these two species had significant geographic structure, but compared to L. entomophila, L. bostrychophila had more significant geographic structure. For SAMOVA analysis, the K value increased from 2 to 5 for L. bostrychophila and L. entomophila, respectively. The FCT was significant and highest at 5 groups for both species  and L. entomophila  populations.AMOVA results showed that there was significant genetic differentiation of lations) . For L.  species . The ManL. bostrychophila, pairwise FST ranged from 0.000 to 0.982 and most of the values were statistically significant  were shown in The complete ITS1-5.8 S-ITS2 sequences varied from 764 to 780 bp and 774 to 782 bp for L. bostrychophila were sequenced for ITS and 59 haplotypes were detected. The difference among the haplotypes was due to 227 polymorphic sites, of which 161 were parsimony informative sites. One hundred and twenty four individuals of L. entomophila were also sequenced for ITS and 77 haplotypes were detected and the difference among these haplotypes was due to 155 polymorphic sites, of which 97 were parsimony informative sites. The numbers of haplotype (n), diversities of haplotype (h) and nucleotide (π) for each population of L. bostrychophila and L. entomophila were summarized in L. bostrychophila individuals (0.990) was higher than all of L. entomophila individuals (0.975) and the nucleotide diversity of L. bostrychophila (π = 0.04081±0.00194) was also higher than that of L. entomophila (π = 0.02569±0.00188).In total, 74 individuals of L. bostrychophila, all of the 59 haplotypes were divided into two clusters (CladeLbI-1 and CladeLbI-2) with high support values (98 and 97) and not arranged according to our predefined geographic regions  was within populations. For SAMOVA analysis, the K value also increased from 2 to 5 for these two species  indicated that there was no significant correlation between the standardized genetic distance [FST/(1−FST)] and the log of the geographical distance for L. bostrychophila  and L. entomophila  populations, indicating that isolation by distance might not be a factor in the gene flow of L. bostrychophila and L. entomophila. Pairwise estimates of FST among L. bostrychophila and L. entomophila populations revealed that the genetic differentiation existed between a large portion of L. bostrychophila and L. entomophila populations , nuclear makers were more sensitive to detect gene flows among the interbreeding populations FST values also provide evidence for population differentiation for both psocids. Intriguingly, the high level of gene flow was also characterized in populations of both species. Although the dispersal ability of psocids seems to be very limited due to their small body size and wingless nature, psocids have great colonization potential and may migrate by human-mediated transport or through air current FST values and high level of gene flows can be explained by high frequency of inbreeding within a population. The Mantel test revealed a negative non-significant correlation between geographic and genetic distance for both species, which is consistent with previous studies of genetic structure of L. bostrychophilaL. bostrychophila and L. entomophila. It is probable that the genetic differentiation is caused by other factors, such as genetic drift, inbreeding or control practices, and less by geographic distance.In the present study, the significant genetic structure was detected among the examined populations according to our predefined regions by two-level and three-level AMOVA analyses using profiles . The SAMprofiles . The lacL. bostrychophila and L. entomophila. Especially for L. bostrychophila, a high overall genetic variability, as also observed in other populations analysed using either allozymes L. bostrychophila was higher than that of L. entomophila although the levels of nucleotide diversity were highly variable among different populations for each species. Here, in the comparison of genetic diversities between two psocids, we did not imply the reproduction mode was the key factor in shaping genetic diversity. Actually, high haplotype diversity at a gene locus within populations is determined by many other factors, including large population size, environmental heterogeneity, life-history traits, and origin and ages of the species Echmepteryx hageni (bark lice) was reported when comparing to its sexual populations L. bostrychophila (0.041) and L. entomophila (0.026) inferred from ITS sequences were very high compared to other reported ITS sequences nucleotide diversity of invasive insects, such as Stomoxys calcitrans and Cimex lectulariusL. bostrychophila and L. entomophila might explain why these species has a broad tolerance to environmental and habitat stresses and the fast mutational processes inherent in individuals, as well as populations, can enable these two Liposcelis species to successfully adapt to complex environments.Sequences from mtDNA and nuclear DNA were characterized by very high levels of genetic diversity in asexually and sexually reproducing psocids, L. bostrychophila is that this pest is a primarily parthenogenetic psocid occasionally undergoes sexual reproduction, which even at very low frequencies can generate substantial diversity, and that the species is panmictic L. bostrychophila populations (about 17 generations per year).Some of the main mechanisms of mutational, ecological and evolutionary change in parthenogenetic insects have been reviewed recently L. bostrychophila is parthenogenetic as a result of being infected by a Wolbachia-like rickettsial bacterium, which is transovarially transmitted Wolbachia infection has affected the mitochondrial genetic diversity of the host insects and usually reduces the genetic diversity Wolbachia infection on mtDNA variation in psocids.In addition, many insects are ubiquitously associated with diverse endosymbiotic microorganisms. It is known that some psocids can be infected by endosymbionts Fs is strongly affected by population expansion or selective sweeps. In various neutrality tests, Fu's Fs is a particularly powerful test of population growth and it is much more powerful than mismatch distribution in detecting signals of population growth L. bostrychophila populations combined, Fu's Fs and Tajima's D values were not significantly negative except significantly negative Fu's Fs values inferred from ITS, suggesting population expansion in total populations. Due to significantly negative Fu's Fs and Tajima's D values for all L. entomophila populations combined , we speculate that the populations might have undergone population expansion in the past. Evidence from the mismatch distribution statistics and approximate star-like haplotypes median-joining networks also support the hypothesis that L. bostrychophila and L. entomophila have experienced a historical increase in population size after a period of having a small effective population. The pattern of genetic variability with high haplotype diversity but relatively low nucleotide diversity in Cytb gene also suggests the population has experienced population expansion rg and SSD failed to reject the null hypothesis of population expansion , dispersal of source populations and the resultant genetic structure among populations within China is useful for proposing successful integrated pest management tactics for these pests of grain storage systems. However, we realized that the small sized samples used in this study and populations examined here do not represent the true distribution of these two psocids in China. Also, only mtDNA gene and ITS gene were used as the molecular marker and their short length contains limited information for population genetic analyses. Nuclear genes, such as microsatellite DNA need to be examined to gain a deeper understanding of the population genetics and invasion biology of these two psocids. Indeed, microsatellite loci have already been isolated for L. bostrychophila and L. entomophilaA thorough understanding of Table S1Fixation indices corresponding to groups of populations inferred by SAMOVA analysis for two species.(DOC)Click here for additional data file.Table S2Pairwise FST values  and gene flow  among populations for L. bostrychophila using ITS sequences.(DOC)Click here for additional data file.Table S3Pairwise FST values  and gene flow  among populations for L. entomophila using ITS sequences.(DOC)Click here for additional data file.


Chickens represent an important animal genetic resource for improving farmers’ income in Africa. The present study provides a comparative analysis of the genetic diversity of village chickens across a subset of African countries. Four hundred seventy-two chickens were sampled in 23 administrative provinces across Cameroon, Benin, Ghana, Côte d’Ivoire, and Morocco. Geographical coordinates were recorded to analyze the relationships between geographic distribution and genetic diversity. Molecular characterization was performed with a set of 22 microsatellite markers. Five commercial lines, broilers and layers, were also genotyped to investigate potential gene flow. A genetic diversity analysis was conducted both within and between populations.(P < 0.01) were identified, matching with Major Farming Systems  described by the FAO.High heterozygosity levels, ranging from 0.51 to 0.67, were reported for all local populations, corresponding to the values usually found in scavenging populations worldwide. Allelic richness varied from 2.04 for a commercial line to 4.84 for one population from Côte d’Ivoire. Evidence of gene flow between commercial and local populations was observed in Morocco and in Cameroon, which could be related to long-term improvement programs with the distribution of crossbred chicks. The impact of such introgressions seemed rather limited, probably because of poor adaptation of exotic birds to village conditions, and because of the consumers’ preference for local chickens. No such gene flow was observed in Benin, Ghana, and Côte d’Ivoire, where improvement programs are also less developed. The clustering approach revealed an interesting similarity between local populations found in regions sharing high levels of precipitation, from Cameroon to Côte d’Ivoire. Restricting the study to Benin, Ghana, and Côte d’Ivoire, did not result in a typical breed structure but a south-west to north-east gradient was observed. Three genetically differentiated areas Local chickens form a highly variable gene pool constituting a valuable resource for human populations. Climatic conditions, farming systems, and cultural practices may influence the genetic diversity of village chickens in Africa. A higher density of markers would be needed to identify more precisely the relative importance of these factors. Knowledge-based management of animal genetic resources (AnGR) is critical to answer the current agricultural, socio-economic, and environmental challenges. Consequently, characterization of AnGR constitutes one of the priorities of the FAO global plan of action for AnGR , in partVillage poultry make a significant contribution to poverty alleviation and household food security in many developing countries . About 1The aim of the present study was to provide a large-scale analysis of the genetic diversity of local chickens in several countries from the central, western and northern parts of the African continent, in order to address questions important for further conservation strategies. These questions deal with (i) the amount of genetic diversity found within these populations, (ii) the search for a possible correlation between the genetic structure and agroecological distribution, and (iii) the detection of a possible gene flow between local populations and commercial lines.The sampling design involved 5 countries . Blood samples were drawn from the wing vein of 472 local adult chickens. Samples representative of Benin, Côte d’Ivoire, and Ghana chicken populations were collected throughout each country Table . These pGenomic DNA was extracted from blood using the Qiagen® Kit  at the biotechnology laboratory of the University of Ghana  or using the NaOH extraction protocol at the IAV (Moroccan samples). A total of 601 individuals, including 472 local African chickens and 129 individuals from commercial lines, were genotyped for 22 microsatellite loci from the AvianDiv panel . PCR ampr ≥ 0.2 could be considered to be potentially problematic for calculations. The allele frequencies, number of alleles, observed (Ho), non-biased expected (He) heterozygosity, and F-Statistics [4.05.2[4.07[Ar) was computed with the rarefaction method using FSTAT 2.9.3.2[Administrative provinces were considered as sampling units for performing preliminary estimations of genetic polymorphism. The presence of null alleles was tested using FreeNA : loci wiatistics  were ests [4.05.2. GENEPOP05.2[4.07 was used05.2[4.07. AllelicT 2.9.3.2. Test siDR; [DR distance, a NeighborNet [4.8[The matrix of Reynolds distances  was comDR; . RegardighborNet  network rNet [4.8.t-test procedures.Clustering approaches were performed, in a first step, on the 27 populations using a Bayesian clustering procedure implemented in STRUCTURE , consideDAS) [1.2.28[0.7[Potential genetic introgression from commercial lines to local populations was investigated by computing the individual allele shared distance (DAS)  matrix f) [1.2.28 and TREE.2.28[0.7 respectiAmongst the 22 markers, 188 alleles were identified, with the number of alleles per marker ranging from 2 to 23  to 0.665 (MAR-Mek) with a mean value of 0.560 (±0.078), according to the genetic diversity indices of the studied populations  values rose from 2.04 (WEL-A) to 4.84 (CIV-SCo) with a mean value of 3.93 (±0.73). Fis ranged from −0.081 (BEN-Oué) to 0.131 (MAR-Mek). After sequential Bonferroni correction, 6 populations showed a significant deficit of heterozygotes for 1 or 2 loci, and one population exhibited 1 locus with heterozygote excess. Only two locus x population combinations out of 616 were identified with a potentially null allele : MCW037 x CAM-Est and MCW330 x BEL-C. However, excluding these two loci had very minor effects on Fis and He , suggesting that null alleles are not the main cause of significant Fis values. Hence we chose to conserve all 22 loci. Testing population differentiation, 112 pairs of populations were found as non-significantly differentiated out of the 378 tests performed  found in low frequency in some regions, may also indicate commercial introgression from broiler lines [Since introgression from commercial lines may constitute a major issue for conservation and management of local chicken populations ,11, we tW locus  found inW locus ) and CôtW locus ) or in oW locus ), while W locus ,54) or MCameroon . In CameAs in other places in Africa, several cooperating programs  were set up in these countries in order to improve poultry production, some of these involving the introduction of exotic cockerels for genetic improvement. Differences in the temporal framework of these programs could be, at least partially, responsible for the contrasting patterns of genetic introgression observed in our study. On the one hand, long-term operations took place in Morocco and Cameroon. In Morocco , the fir1 females from exotic cocks are not as successful as the local hens in natural reproduction and chick rearing. Furthermore, commercial chicks have been selected with an optimum feeding system, and may not satisfy their nutritional needs in scavenging conditions. According to some authors [Yet, taking into account the duration of exotic cocks introduction, genetic introgression appears finally relatively low even in Morocco and Cameroon. Limited impact of introductions may be explained by a poor adaptation of the genotypes introduced to local conditions or by the consumer’s preference. Indeed, commercial lines have been counter-selected against broodiness, and then females have lost the ability to incubate their eggs naturally, whereas this ability is particularly important for self propagation of village chickens in harsh environments. Although the genetic determinism of broodiness is still under study , it is e authors ,61, ther authors -59,62,63 authors ,54,64 su authors .P < 0.01 for RCFS/C-RCFS comparison and P < 0.001 for the two other ones). Within the RCFS, according to surveys conducted in Benin [P = 0.059) was observed between animals of northern and southern part of Benin, according to the Structure analysis restricted to Benin, Ghana, and Côte d’Ivoire . The number of markers used may be too limited to ensure detection of a genetic differentiation on a quantitative trait such as growth.Usually, scavenging chicken populations do not exhibit a typical breed structure ,15,45,65in Benin , the weiin Benin  showed tHuman settlements and migrations may constitute another driving factor for the genetic structure of chicken populations. Although not well documented, there is a lot of movement of chickens between these countries as a result of long history of trade and migration. In particular, a large part of the human groups living in the south-east part of Côte d’Ivoire and south-west part of Ghana belong to the same ethnic group, namely the Akan, originating from Ghana ,69. AmonThese agroecological, cultural and demographic factors could result in genetic drift, gene flows or adaptation phenomena, and therefore explain the genetic structure pattern found across these countries. On the basis of the present data, it is, however, difficult to assess to what extent each of these factors has impacted genetic differentiation of these chicken populations.African local chickens form a highly variable gene pool which constitutes a valuable resource for human populations. There is a large number of driving forces playing either for or against population differentiation of local chickens. Molecular studies bring complementary information to social surveys and phenotypic data, and allows to set up an integrated program of characterization and conservation of indigenous populations, as recommended by the FAO ,71. In tThe authors declare that they have no competing interests.XR, IAKY, BBK, CVYG, KB and MTB conceived the project; IAKY, BBK, CVYG, ROA, NEL, JCF, KB, XR and BB collected the samples and data; XR and GL analysed the data; XR, GL and MTB led the writing, and all the authors participated in the discussion. All authors read and approved the final manuscript.A), number of private alleles (Ap) and allelic richness (Ar).Summary of polymorphic measures for microsatellite markers. For each and over all populations or within each African country’s population or commercial line, the following information are given: allele range, number of alleles (Click here for fileDR) and levels of significance of genic differentiation among the 28 chicken populations. DR values are above the diagonal and levels of significance are below the diagonal. For population codes, see Table Pairwise genetic distances , for K = 2-16, using Q-matrix averaged overall 100 runs.Click here for fileLn(P(D)) and (b) similarity function G’ according to the number of cluster K (K = 1 to 16). STRUCTURE analysis involving all 28 populations . Evolution of (a) likelihood Click here for fileLn(P(D)) according to the number of cluster K (K = 1 to 6).STRUCTURE analysis restricted to the 18 African local chicken populations from Ghana, Benin and Côte d’Ivoire. Evolution of likelihood Click here for fileR distance.Neighbor-Net for the complete dataset , based on Reynolds DClick here for fileDAS distance among 242 samples representing local chickens from Benin (n = 113) and 5 commercial lines (n = 129). red: Benin; orange: BS-D; dark green: BS-C; dark blue: Bel-C; light green: Wel-A; light blue: BD-B; black star indicates individuals with qi > 0.2, for clusters specific to commercial lines .Individual neighbor-joining dendrogram based on Click here for fileDAS distance among 250 samples representing local chickens from Côte d’Ivoire (n = 121) and 5 commercial lines (n = 129). red: Côte d’Ivoire; orange: BS-D; dark green: BS-C; dark blue: Bel-C; light green: Wel-A; light blue: BD-B; black star indicates individuals with qi > 0.2, for clusters specific to commercial lines .Individual neighbor-joining dendrogram based on Click here for fileDAS distance among 241 samples representing local chickens from Ghana (n = 112) and 5 commercial lines (n = 129). red: Ghana; orange: BS-D; dark green: BS-C; dark blue: Bel-C; light green: Wel-A; light blue: BD-B; black star indicates individuals with qi > 0.2, for clusters specific to commercial lines .Individual neighbor-joining dendrogram based on Click here for fileDAS distance among 214 samples representing local chickens from Cameroon (n = 85) and 5 commercial lines (n = 129). red: Cameroon; orange: BS-D; dark green: BS-C; dark blue: Bel-C; light green: Wel-A; light blue: BD-B; black star indicates individuals with qi > 0.2, for clusters specific to commercial lines .Individual neighbor-joining dendrogram based on Click here for fileDAS distance among 175 samples representing local chickens from Morocco (n = 46) and 5 commercial lines (n = 129). red: Morocco; orange: BS-D; dark green: BS-C; dark blue: Bel-C; light green: Wel-A; light blue: BD-B; black star indicates individuals with qi > 0.2, for clusters specific to commercial lines .Individual neighbor-joining dendrogram based on Click here for file


Ho) of >0.7 and moderately polymorphic in Tripura population (Ho > 0.6). However, both the population showed least polymorphism at TH01 locus (Ho < 0.6). A comparison between Chakma and Tripura population data revealed statistically significant differences in allele frequency distribution for most of the loci. A similar comparison with the mainstream Bengali population using previously published data from this lab also showed significant difference in allele frequency with these two tribal populations.Allele frequencies of ten autosomal STR loci, D3S1358, vWA, D16S539, D2S1338, D8S1179, D21S11, D18S51, D19S433, TH01, and FGA were investigated in Chakma and Tripura tribal populations of Bangladesh. In both the populations, all loci were in Hardy-Weinberg equilibrium except for FGA locus in Chakma and D21S11 in Tripura. All the loci were highly polymorphic in Chakma population with an observed heterozygosity ( Short tandem repeats (STRs) are highly polymorphic sequences of nucleotides, which are abundant in eukaryotic genome. They form approximately 3% of the total human genome and occur on average in every 10 000 nucleotides . STRs reIn this study, we report the allele frequency data from Chakma and Tripura tribal population of Bangladesh. Besides the forensic uses, we find this study a very powerful adjunct for further regional and worldwide meta analysis.Liquid blood samples were collected from randomly selected 109 Chakma and 58 Tripura individuals in Bangladesh. There are four ethnic groups in Bangladesh; the Dravidian, proto-Australian, Mongolian, and Bengalis. The Bengalis are by far of all, constituting 98% of the population of the country. The Chakmas and Tripuras tribal groups along with Marmas, Garos, and Manipuris belong to the Mongoloid stock and are Sino-Tibetan in origin. They live in Chittagong Hill Tract, Khagrachari, and Bandarban districts along the eastern border of Bangladesh adjoining Myanmar and Indian states of Mizoram and Tripura. The Chittagong Hill Tracts host 13 main tribes of which Chakmas are largest single tribes in Bangladesh. According to the 2001 population census, there were about 300 000 Chakmas and 90 000 Tripuras. The Dravidian element of population is represented mainly by Oraons, a tribe of central India in origin. The Australoid group includes Khasias and Santals, mainly labourers in the tea garden at the Sylhet district.DNA was extracted using the Chelex-100 method as described by Walsh et al. . ExtractlSTR SGM Plus PCR amplification kit . The PCR reaction was carried out in a GenAmp PCR System 2720 (Appliled Biosystems). Thermal cycling parameters were setup according to the manufacturer's protocol.Approximately 1-2 ng of template DNA was used for each PCR amplification process. Ten autosomal STR loci namely D3S1358, vWA, D16S539, D2S1338, D8S1179, D21S11, D18S51, D19S433, TH01, and FGA were coamplified using AmpFavant Genetic Analyzer (Applied Biosystems) using POP-4 polymer and data collection software ver. 1.1. Data were sized using GeneScan Software version 3.7 and internal GeneScan-500 ROX size standard. Genotype of each locus was determined after comparison with allelic ladder using Genotyper software version 3.7 NT.PCR-amplified fragments were separated and analyzed on ABI Prism 3100-Allele frequencies at each locus and statistical parameters of forensic efficiency were calculated by using PowerStat Microsoft Excel Workbook template . PossibllSTR SGM Plus PCR amplification kit (Applied Biosystems). Approximately 10% of samples from both the populations were regenotypes to ensure reproducibility and accuracy. All genotype results were in full concordance.Positive control DNA and allelic ladder provided in AmpF−12 (1 in 2.86 × 1011) and 1.03 × 10−11 (1 in 9.65 × 1010), respectively, for Chakma and Tripura populations. The combined power of exclusion of paternity (PE) for the Chakma and Tripura were calculated to be 0.99997 and 0.9998, respectively. Significant deviation from Hardy-Weinberg equilibrium was observed for FGA locus in Chakma and D21S11 in Tripura population. Both the populations showed least polymorphism at TH01 locus (PIC <0.6). This may be due to inbreeding and lack of admixture, which is a characteristic feature of these populations. Although both the populations achieved combined probability of match (PM) in the order of 1010 to 1011, it would be reasonable to implement a “θ correction” while using this data in forensic casework, as recommended by the National Research Council (1996) [Allele frequencies of Chakma and Tripura tribal groups are shown in Tables 2 1 in 2.6 × 1011 The present study was undertaken from the standpoint of compiling own-population genetic database, owing to the fact that population and subpopulation differences at STR loci of forensic interest are growing up. In this context, we embarked on compiling the allele frequency database of two ethnolinguistic populations of Bangladesh. The obtained high combined PM and PE values demonstrated that STR loci employed in SGM Plus PCR amplification kit would be highly efficient for personal identification and paternity cases in spite of some population substructuring. Besides forensic benefits, we find this study very useful for regional and worldwide human population meta-analysis.