Ethmalosa fimbriata, is a West African pelagic species still abundant in most habitats of its distribution range and thought to be only recently affected by anthropogenic pressure (habitat destruction or fishing pressure). Its presence in a wide range of coastal habitats characterised by different hydrodynamic processes, represents a case study useful for evaluating the importance of physical structure of the west African shoreline on the genetic structure of a small pelagic species. To investigate this question, the genetic diversity of E. fimbriata was assessed at both regional and species range scales, using mitochondrial (mt) and nuclear DNA markers. Whereas only three panmictic units were identified with mtDNA at the large spatial scale, nuclear genetic markers (EPIC: exon-primed intron-crossing) indicated a more complex genetic pattern at the regional scale. In the northern-most section of shad’s distribution range, up to 4 distinct units were identified. Bayesian inference as well as spatial autocorrelation methods provided evidence that gene flow is impeded by the presence of deep-water areas near the coastline , such as the Cap Timiris and the Kayar canyons in Mauritania and Senegal, respectively. The added discriminatory power provided by the use of EPIC markers proved to be essential to detect the influence of more subtle, contemporary processes  acting within the glacial refuges identified previously by mtDNA.The bonga shad,  The spatial expansion of fisheries and its nefarious impact on marine ecosystems throughout the second half of the twentieth century is now well established . From itSardina pilchardus) was genetically depleted as a probable consequence of intensive fishing in the recent past. Also considering sardine, Chlaida et al. [Ethmalosa fimbriata] [Sarotherodon melanotheron] [Trachurus trecae] [Sardinella aurita] [E. fimbriata]  ; [Sarothotheron] ,29; [Tra trecae] ) or mate aurita] ; [E. fimmbriata] ). Only ombriata] .E. fimbriata, Clupeidae), a euryhaline species distributed from Mauritania to Angola, Durand et al. . The sies first , then ced et al. .M  among population clusters previously determined using TESS and/or BARRIER. MIGRATE-n v3.4.4 used the Brownian mutation model and mutation was considered to be constant for all loci. We used maximum likelihood (ML) implemented in MIGRATE-n to infer the various parameters and the 95% confidence intervals  for each chain. One of every 20 reconstructed genealogies was sampled for both the short and long chains. We used an adaptive heating scheme with 4 concurrent chains; the analyses were run on a cluster computer using one master and 6 compute nodes. Assuming a average mutation rate of 10-5 per locus per generation, average Θ estimates were translated to estimates of average effective population sizes (i.e. Ne = Θ/4μ) for each population cluster previously defined by TESS and/or BARRIER. Mutation rates of microsatellite loci are traditionally assumed to range from 10-3 to 10-5 per generation .The program MIGRATE-n v3.4.4  was usedervals  ,67). FS. FSM , but, butM .Allelic richness and observed levels of heterozygosity were quite variable among these loci, ranging from 5 alleles at the B in ANG . Locus GG sample . No signG sample . MicrochFST = 0.032; p < 0.001) or the data set corrected for locus Gapdh-slow . In contrast with the lack of population differentiation found within the northern biogeographic unit using mtDNA variability ] against geographical distance  for the Rousset  . Assuming µ =10-5, these results translated to average values of Ne ranging from 2150 (95% CI: 1932.5-2582.5) for the southern ANG population of E. fimbriata to 2600 (95% CI: 2435-2827.5) individuals for the population cluster grouping samples from GUI to SAL. The inferred number of migrants between putative A and B clusters (i.e. MA→B~MB→A) are reported in M[SAL+GAM+CAS+GUI]→MAU = 4.116) than the reciprocal estimate (MMAU→[SAL+GAM+CAS+GUI] = 1.866), suggesting that the central population cluster might be the origin of the northern peripheral population as suggested by mtDNA , SEN, and MAU.The occurrence of weakly differentiated samples from SEN and ICA+CON that are geographically disjunct and belong to previously identified distinct mtDNA clades see  is diffiSalvelinus fontinalis) in the Northwest Atlantic, while significant IBD was found among populations located at the northern and southern margins of the distributional range. This may have occurred because of a lower local effective population sizes in these areas compared to the central population. However, estimates of nuclear effective population sizes were not significantly different in samples distributed over the distribution range of the Bonga shad used in this study . Reduction in Ne was not demonstrated using the M-ratio, indicating relative demographic population stability in the Bonga shad. The cause of observed IBD cannot be due only to differential susceptibility of nuclear and mtDNA markers to genetic drift and selection. Sex-biased dispersal and/or biased reproductive behaviour  acting at different spatial or temporal scales could be alternative or complementary explanations to contrasted IBD patterns at nuclear and mtDNA markers. Biological data on sex-ratio, sex-specific dispersal and homing (degree of population closeness) are few and often dramatically distinct in the Bonga shad. For example, Blay and Eyeson )Microsatellite data used in this manuscript: available on request.


Larimichthys polyactis) in the Northwest Pacific marginal seas. Among these loci, we detected at least two microsatellites, anonymous H16 and HSP27 to be clearly under diversifying selection in outlier tests. Sequence cloning and analysis revealed that H16 was located in the intron of BAHCC1 gene. Landscape genetic analysis showed that H16 mutations were significantly associated with temperature, which further supported the diversifying selection at this locus. These marker types presented different patterns of population structure: (i) mitochondrial DNA phylogeny showed no evidence of genetic divergence and demonstrated only one glacial linage; (ii) population differentiation using putatively neutral microsatellites presented a pattern of high gene flow in the L. polyactis. In addition, several genetic barriers were identified; (iii) the population differentiation pattern revealed by loci under diversifying selection was rather different from that revealed by putatively neutral loci. The results above suggest local adaptation in the small yellow croaker. In summary, population genetic studies based on different marker types disentangle the effects of demographic history, migration, genetic drift and local adaptation on population structure and also provide valuable new insights for the design of management strategies in L. polyactis. The genetic differentiation of many marine fish species is low. Yet local adaptation may be common in marine fish species as the vast and changing marine environment provides more chances for natural selection. Here, we used anonymous as well as known protein gene linked microsatellites and mitochondrial DNA to detect the population structure of the small yellow croaker ( In many marine fish species, especially migratory species, the signal of population differentiation is weak and hard to detect due to high levels of gene flow. It is traditionally difficult to delineate the population structure or detect the population subdivision lacking clear barriers to gene flow within ocean basins . EspeciaIn recent years, a growing number of studies have revealed low but statistically significant differentiation among populations of many marine fishes using microsatellites, a highly variable and presumably neutral genetic marker -11. MitoGadus morhua) between the North Sea and the Baltic Sea [Platichthys ﬂesus) were grouped according to environmental similarities using Hsc70 locus [PanI locus. White et al. [Coryphaenoides rupestris) at locus Crup7 was related to depth, suggesting that directional selection presumably acted on a linked locus. Several recent studies suggested directional differentiated microsatellites may have formed due to selection acting on their linked loci, which were related to environmental factors such as temperature, salinity and depth or water pressure [Several previous studies have reported local selection in some marine fishes. For example, the haemoglobin locus showed differentiation along latitudinal cline in Atlantic cod , is an important fishery species endemic to the Northwest Pacific, inhabiting coastal waters across the Bohai Sea, the Yellow Sea and the East China Sea [L. polyactis are subjected to a high fishing pressure and now approaches overexploited [L. polyactis will be feeding in the nearby waters. Several studies on genetic structure of L. polyactis in our studied areas obtained inconsistent or even conflicting results. Although studies using RAPD and AFLP [L. polyactis [L. polyactis has hindered the stock management and conservation to some extent. Considering the low sampling resolution in these studies  [The small yellow croaker, hina Sea . As one xploited -32. Thisxploited . The avexploited -33. An uxploited . This fixploited . The ovexploited . After sand AFLP -38 all solyactis . Such dilow Sea) -39, a moL. polyactis across its distribution range along the Pacific coast of China using anonymous and known functional genes linked microsatellites, and mtDNA. Our purpose was to evaluate the levels and patterns of genetic variation and differentiation. Further, we aimed to find the interacting effects of historical and recent gene flow on shaping the population structure of L. polyactis. We also tried to detect the footprints of local adaptation considering large effective population size and varying environmental variables in the distribution areas of this species. Among the microsatellites, we detected evidence of local adaptation across the distribution range of L. polyactis. Using different marker types, we studied the patterns of population structure and disentangled the effects of demographic history, migration, genetic drift and natural selection from environmental factors on population structure of L. polyactis.In this study, we analyzed a large sample of The species is not protected by Chinese law or by any of the countries where and when sampling was performed. It is a commercially harvested species in China and the other Northeast Asian countries.Samples were collected from 32 localities  along the coast of China across the species’ distribution range from 2007 to 2010 . All theFirstly, all sampled individuals were genotyped using 22 of our newly developed putatively neutral microsatellite loci, namely, H5, H16, H22, H26, H31, H33, H37, H38, H43, H44, H47, H51, H54, H65, H70, H71, H75, H80, H82, H84, H106 and H111 . BesidesCOI) and Cytochrome b (Cytb) genes were targeted for sequencing using the following primer sets: 5ʹ-cgttataatcttctttatag-3ʹ and 5ʹ-gtgtcatgaaggacaatgtc-3ʹ for COI, and 5ʹ-gcctgatgaaactttggctc-3ʹ and 5ʹ-caaggactccgcctagtttg-3ʹ for Cytb, respectively. PCR amplification was performed in a 50-μL volume containing 0.5μM of each primer, 0.2mM dNTP, 1.5mM MgCl2, 1×PCR buffer, 1U Taq DNA polymerase (Fermentas), and 20ng template genomic DNA. PCR program was as follows: 5 min at 94°C followed by 35 cycles of 30 s at 94°C, 30 s at 55°C, and 60 s at 72°C with a final extension of 5 min at 72°C. PCR products were then purified and used as templates for sequencing in both directions with primers above. Sequencing work was also conducted on an ABI PRISM 3730 DNA automated sequencer. Sequences have been deposited in GenBank with accession no.JN243355-JN243749 and JN250600-JN250994.Two mtDNA fragments of Cytochrome c Oxidase subunit I (O), expected heterozygosities (HE), inbreeding coefficient (FIS) and allele richness  were calculated in FSTAT version 2.9.3.2 [ST values with 95% confidence interval (CI) were also estimated using FSTAT version 2.9.3.2 [Genetic variation indices including the number of alleles (A), observed heterozygosities . Furthermore, we used the method of Beaumont & Nichols [ST across all populations and under a model of 10 groups each consisting of 50 populations. We performed this test based on 100 000 simulated loci and by using the software Arlequin 3.5 [For microsatellites neutrality, we firstly used a Bayesian simulation-based test implemented in BAYESCAN  to test  Nichols  implemen Nichols . This me Nichols . Therefo Nichols  that is  Nichols  under a ST matrices acquired by computing pairwise Wright’s F-statistics. In this analysis, we estimated two types of FST matrices: only the neutral loci and the outlier loci under diversifying selection detected in outlier tests. The significance of the tests were computed by a permutation with 10 000 replicates using ARLEQUIN 3.5 [5 iterations after a 105 burn-in length with K ranging from 1 to 8. The locations were included as priors considering rather low genetic divergence in this species. The most likely structures were evaluated using K and ΔK methods [The population structure was assessed using pairwise FQUIN 3.5 . In ordeQUIN 3.5 . We alsoQUIN 3.5 . This pr methods . The eff methods ,55. The ST matrices, generated by bootstrapping over loci. As this landscape genetics approach highly favors a sufficient spatial resolution of sampling, it was considered well appropriate for our data sets.The population structure for neutral loci was also investigated using a landscape genetics method in the software BARRIER . This prST values for each population and then relates them to environmental factors using a generalized linear model. In our study, we assessed seven environmental variables including sampling temperature and salinity, latitude and longitude, as well as historical average temperature in April, in May and in April and May retrieved from weather.sina.com.cn (shown in 7 (= 128) alternative regression models. This method calculates the posterior probabilities for each of these models using a reversible jump MCMC approach. We performed this analysis under default parameters. Based on the results of GESTE, we also tested the level of correlation between pairwise FST and pairwise differences of environmental factors of the highest posterior probability in outlier tests using Mantel test. As the environmental factors above are considered highly related to the behavior of spawning migration for marine fishes [In order to elucidate the effects of different environmental factors on population structuring, we used the hierarchical Bayesian approach of Foll and Gaggiotti , implemeshown in , which le fishes -61, the Demographic history of population size variations was investigated in the form of heterozygote excess based on neutral loci using the program Bottleneck version 1.2.02 . BecauseDicentrarchus labrax). Based on sequence homology among European seabass (GenBank accession no.CBN81236), zebrafish  and orange-spotted grouper (unpublished data from genome sequencing program of our laboratory), we designed primers to amplify the corresponding sequence and aimed to confirm that H16 locus is also linked with BAHCC1 gene in L. polyactis. PCR was conducted using Takara LA kit ver. 2.1. Amplified fragments were ligated into PMD-18T vector (Takara) after purification using the E.Z.N.A. Gel Extraction Kit  and were further sequenced in both directions.In outlier tests, we detected one anonymous locus, H16 showed much higher level of genetic divergence (see results). Blast against GenBank revealed that this locus showed more than 90% of sequence identity with the intron sequence of BAH domain and coiled-coil containing protein 1 (BAHCC1) gene of European seabass  and nucleotide diversity (π) showed no differences among geographical groups but significant homogeneity across samples  and was highly significant (P < 0.001). However, the overall FST value for locus H16 (FST = 0.092) was much higher than the other loci, indicating diversifying selection at this locus. Outlier tests based on BAYESCAN showed that four loci H16, H65, HSP27 and H51 were under diversifying selection under the decisive criterion of Jeffrey’s interpretation that loci with Log 10 (Bayes factors) > 2 were considered as outliers  and was statistically significant (P < 0.001). Nevertheless, this value was markedly lower than that of the directional selected loci H16  and HSP27 . In PCA plotting, we didn’t observe a clear geographical pattern of population structure based on neutral loci .The overall neutral Fral loci . The sigral loci . Besides samples . Excludi samples , suggestST analysis at locus H16 revealed a clear separation between samples from NECS and CECS , human (Homo sapiens) and some other species , and so on [ Our results here likely indicate that diversifying selection in nd so on , suggestL. polyactis using putatively neutral and diversifying selected microsatellites and mtDNA analysis. The joint application of these different marker types not only disentangled the effects of different factors shaping the genetic structure, but also suggested unique evolutionary history of L. polyactis.The population structuring of marine fishes has been considered to be the results of interaction among environmental factors, behaviors, genetic drift and demographic history. In our work, we provided different and comprehensive insights into the genetic structuring of L. polyactis across distribution range using mtDNA, suggesting only one refuge for L. polyactis during Pleistocene glaciations, the ancient East China Sea. The Bohai Sea and the Yellow Sea were exposed due to low sea level during this period. Considering the long-term glacial period and frequent interglacial periods, immediate gene flow during this period might have blurred the existence of multiple glacial refugia. However, phylogenetic studies on marine species in the studied areas found multiple lineages coexisting in the current Yellow Sea and East China Sea and no evidence of interglacial gene flows blurring genetic lineage divergence [L. polyactis could have been shaped by relatively recent events but not by glacial events.Multiple allopatric glacial refugia are necessary for population divergence during Pleistocene . In our vergence ,70. In tL. polyactis. Additionally, several less well supported genetic barriers were also identified in the Bohai Sea and the northern Yellow Sea, suggesting high genetic heterogeneity for the populations in these areas. Importantly, we detected some evidence of panmixia for L. polyactis in the East China Sea, revealing that L. polyactis in this area is an independent migratory group as suggested by Liu [In our study, inconsistent results between neutral and diversifying selected microsatellites reflect the interaction effects of recent gene flow or migration, genetic drift and local adaptation on population structure. Based on neutral data, we found high gene flow and weak population structure, as well as a weak pattern of IBD across samples. Nevertheless, the landscape approach might provide some evidence of geographical structure, which reflects the effects of migration routes on population structure. On one hand, the program BARRIER identified one strong genetic discontinuity being placed between the BS and the NYS. This barrier shows little difference with the boundary between the northern and the central migratory groups partitioned by fishery research based on migration routes ,34. The d by Liu  and IkedCynoscion regalis), Atlantic herring (Clupea harengus) and Atlantic Bluefin Tuna (Thunnus thynnus), indicating the influences of migration routes on population structuring by blocking gene flow [L. polyactis spawn eggs typically at waters near shore and juveniles are also prone to live in inshore waters and shallow estuaries [Many studies have revealed population structuring in migratory marine fishes such as weakfish  and the ocean currents in studied areas began to form since then [L. polyactis differentiation because the physiological activities of migratory marine fishes are very sensitive to temperature changes [L. polyactis may have initially differentiated under selection of temperature between NECS and CECS, and then recolonized the Bohai Sea and the Yellow Sea. As a result, these two seas seemed like the secondary contact zone for L. polyactis. However, the mechanism of maintaining such pattern of divergence was still unclear. The adaptive differentiation for samples in the Bohai Sea and the Yellow Sea may have resulted from the physiological behavior of native environments adaptation and the higher strength of selection than the migration rates [During the last glacial maximum (LGM), the ancient East China Sea was lowered into a trough due to the low sea level and the shallow shelf ,93, whicnce then . The sounce then . These tnce then . Among t changes . Seen fron rates .L. polyactis recolonizing the Yellow Sea and the Bohai Sea by providing suitable conditions [ST and BARRIER analyses.As the formation of the southward China Coastal Cold Current was after 14ka BP , it is unditions . This exL. polyactis likely have formed after recolonizing the Bohai Sea and the Yellow Sea at about 6ka BP. As this period was sufficient to built up significant genetic differentiation considering the relatively short generation time of L. polyactis (about one year), the low differentiation for neutral microsatellites is most likely due to high effective migration rates rather than genetic drift [Further, the modern migration routes of ic drift . In addiic drift .L. polyactis differentiation from environmental factors, which can help understand the adaptive evolutionary processes [L. polyactis.Effective stock management and conservation are hindered by the lacking of knowledge of the demographic history, the patterns of gene flow and the adaptation to environmental factors . In partrocesses . Last buL. polyactis. At the same time, the adaptive differentiation provides novel insights into the population structure and evolutionary process of L. polyactis and serves as important information for designating stock boundaries and evolutionary units for L. polyactis management and conservation. Although the mechanism of local adaptation is currently unclear, our study adds to the understanding of the evolutionary pattern of migratory marine fish species, which can help predict future variation of biodiversity and establish conservation strategies.Our work demonstrates that joint application of different genetic marker types to study population dynamics provides a comprehensive understanding of the interplay among different evolutionary forces. Our results showed that different patterns of population structure using mitochondrial DNA, putatively neutral and outlier microsatellites disentangle the effects of demographic history, migration, genetic drift and environmental factors on the population divergence of Table S1Summary of the microsatellites linked to functional genes.(DOCX)Click here for additional data file.Table S2Larimichthys polyactis.Summary statistics at 25 microsatellite loci across 32 samples of (XLSX)Click here for additional data file.Table S3Genetic diversity of microsatellites and mitochondrial DNA for each sample.(DOCX)Click here for additional data file.Table S4Values for pairwise differentiation  and associated P values . (a) FST values based on 20 neutral microsatellites loci. (b) FST values based on locus H16. (c) FST values based on locus HSP27. (d) ΦST values based on mtDNA. Differentiation with P < 0.0001 (correction for multiple tests of 496 comparisons at 0.05 levels) was highlighted in bold. Inter-population differentiation is highlighted with light shading.(XLSX)Click here for additional data file.Table S5Primer sequences used for BAHCC1 gene cloning.(DOCX)Click here for additional data file.Table S6Demographic statistics for mitochondrial DNA.(DOCX)Click here for additional data file.Table S7Sampling location and Genebank accession number for each sample.(DOCX)Click here for additional data file.Figure S1ΔK values calculated according to Evanno et al.(2005) for 20 and 22 loci data sets, respectively. (DOCX)Click here for additional data file.Figure S2Sequence alignment of BAHCC1 protein sequences among several species.(DOCX)Click here for additional data file.Figure S3Larimichthys polyactis) based on mitochondrial DNA haplotypes.Phylogenetic tree of the small yellow croaker ( Bootstrap supports of more than 50% from 1000 replicates are shown. The Large yellow croaker (Larimichthys crocea) is used as outgroup (GenBank no. EU339149).(DOCX)Click here for additional data file.


Spinturnix myoti and Spinturnix bechsteini) with very similar life histories. Their respective hosts, the greater mouse-eared bat (Myotis myotis) and the Bechstein’s bat (Myotis bechsteinii) have social systems that differ in several substantial features, such as group size, mating system and dispersal patterns.The population genetic structure of a parasite, and consequently its ability to adapt to a given host, is strongly linked to its own life history as well as the life history of its host. While the effects of parasite life history on their population genetic structure have received some attention, the effect of host social system has remained largely unstudied. In this study, we investigated the population genetic structure of two closely related parasitic mite species (S. myoti we found high levels of genetic diversity and very little pairwise differentiation, whereas in S. bechsteini we observed much less diversity, strongly differentiated populations and strong temporal turnover. These differences are likely to be the result of the differences in genetic drift and dispersal opportunities afforded to the two parasites by the different social systems of their hosts.We found that the two mite species have strongly differing population genetic structures. In Our results suggest that host social system can strongly influence parasite population structure. As a result, the evolutionary potential of these two parasites with very similar life histories also differs, thereby affecting the risk and evolutionary pressure exerted by each parasite on its host. In the evolutionary arms race between hosts and their parasites, the interacting species may use immunological, physiological and behavioural adaptations ,2. HostsRelative dispersal rates of hosts and parasites have been studied in a number of systems. For instance, in the black-legged kittiwake and a parasitic tick, parasite populations were found to be much more spatially structured than those of their hosts . This di. The mite species (Spinturnix myoti and Spinturnix bechsteini) are closely related and have indistinguishable life histories  [et al. [Amplification of kx et al. using thkx et al.. MicrosaF288850] . Amplifi .The datasets for The authors declare that they have no competing interests.All authors designed the research and participated in sample collection; JvS and NB performed the molecular genetic studies and data analysis; and JvS supported by all authors wrote the paper. All authors read and approved the final manuscript.Supplementary analyses to: The effect of host social system on parasite population genetic structure: comparative population genetics of two ectoparasitic mites and their bat hosts.Click here for file


Hangul (Cervus elaphus hanglu), the eastern most subspecies of red deer, is now confined only to the mountains in the Kashmir region of Jammu & Kashmir State of India. It is of great conservation significance as this is the last and only hope for Asiatic survivor of the red deer species in India. Wild population of free ranging hangul deer inhabiting in and around Dachigam National Park was genetically assessed in order to account for constitutive genetic attributes of hangul population using microsatellite markers.ID sibs) was 0.009 (9 in 1000). Altogether, 49 different alleles were observed with mean (± s.e.) allelic number of 8.17 ± 1.05, ranging from 5 to 11 per locus. The observed heterozygosity ranged between 0.08 and 0.83, with mean 0.40 ± 0.11 and the inbreeding coefficient ranged between −0.04 and 0.87 with mean 0.38 ± 0.15. Majority of loci (5/6) were found to be informative . All loci deviated from Hardy-Weinberg equilibrium except Ca-38 (P > 0.05) and none of the pairs of loci showed significant linkage disequilibrium except the single pair of Ca-30 and Ca-43 (P < 0.05).In a pool of 36 multi-locus genotypes, 30 unique individuals were identified based on six microsatellite loci. The estimated cumulative probability of identity assuming all individuals were siblings (Phangul population is significantly inbred and exhibited a low genetic diversity in comparison to other deer populations of the world. We suggest prioritizing the potential individuals retaining high heterozygosity for ex situ conservation and genetic monitoring of the hangul population should be initiated covering the entire distribution range to ensure the long term survival of hangul. We speculate further ignoring genetics attributes may lead to a detrimental effect which can negatively influence the reproductive fitness and survivorship of the hangul population in the wild.The preliminary findings revealed that  Hangul or Kashmir red deer (Cervus elaphus hanglu), one of the four eastern most distributed subspecies of red deer found in the Indian sub-continent, is of great conservation significance since it is the only Asiatic survivor of the red deer inhabiting the broadleaf forest and temperate grassland habitats of Zanskar mountain range in Jammu and Kashmir [hangul was widely distributed in the mountains of Kashmir Himalayas, Chenab valley and some parts of Chamba district in Himachal Pradesh [hangul deer have shrunken dramatically due to environmental and anthropogenic pressures, mainly in the form of habitat loss due to deforestation, degradation and encroachment [ca. 1000 km2) encompassing the Dachigam National Park and its adjoining protected areas [Hangul is considered as least concern (LC) under IUCN red data list of threatened species but it is placed under Schedule-I of the Indian Wildlife (Protection) Act, 1972 and Jammu & Kashmir Wildlife (Protection) Act, 1978. It has also been listed among the top 15 conservation priority species by Government of India [hangul population by adopting scientifically robust methods since 2004. The information on the ecology of hangul is inadequate and only a few short terms ecological studies have been conducted majority of them dealing with habitat use, feeding habits, distribution and conservation assessment surveys [hangul population is lacking while similar studies have been conducted on other sympatric red deer species [ Kashmir . Histori Pradesh . Over tioachment ,3. The ced areas . The ested areas . The ested areas . Hangul  species .hangul population of Dachigam National Park using the pneumatic shed hair samples collected opportunistically during the hangul population estimation exercise carried out in March, 2011. We took advantage of the microsatellite markers that have been cloned for Chital deer (Cervus axis) for analyzing genetic diversity of hangul population [Genetic characterization of a species is considered necessary to formulate breeding policies and prioritize the conservation action plans in an effective and meaningful way ,14. Therpulation  since crpulation -18.hangul also shed their pneumatic hairs during onset of spring which act as an insulator by protecting them from severe cold during winter. We participated in the hangul population estimation exercise carried out in March, 2011 by the Department of Wildlife Protection, Jammu & Kashmir and collected individual tuft of pneumatic hair samples (n = 84). The samples were stored in parchment paper envelope in dry condition till shipment to the laboratory for genetic analysis. The locations of the collected samples were recorded using geographical positioning system and plotted on the map using ArcGis 9.3 [Like the other red deer species, the cGis 9.3   and Mplex 2  using Qiagen Multiplex PCR Kit . PCR reactions were set up in a 15 μl of reaction volume containing 7.5 μl of 2× Qiagen Multiplex PCR Master Mix, 0.30 μl of 10 μM of each primer pair, 1 μl of Q solution (supplied with kit), 2 μl of DNA elutant (approx. 20 ng) and remaining RNase-free water to make the final reaction volume 15 μl. PCR amplification were performed on ABI 9600 Fast Thermocycler . The amplification conditions were 15 min initial heat-activation of Hot Start Taq DNA polymerase at 95°C, followed by 35 cycles of denaturation at 94°C for 30 s, annealing at specific temperature (Ta- 60°C for Mplex 1 and 58°C for Mplex 2) for 90 s and extension at 72°C for 60 s with a final extension at 60°C for 30 min. Fluorescence based fragment analysis was performed on ABI 3130 Genetic Analyzer  and allele scoring was done manually using GeneMapper software version 3.7 .A set of seven microsatellites which were originally isolated from Chital deer and tested for cross amplification in several other cervids were selected for the present study . The genID) and siblings (PID sibs) was calculated using identity analysis module in GenAlEx version 6.5 [Inevitable genotyping errors due to allelic drop out (ADO) and false alleles (FA) if left unaccounted, may lead to erroneous results in individual identification, population assignment, kinship and census studies ,21. We tsion 6.5 .a) and effective alleles (Ne), observed (Ho) and expected (He) heterozygosity using POPGENE version 1.32 software [FIS was calculated according to Weir and Cockerham (W&C) [The per locus diversity was quantified by estimating the numbers of observed (Nsoftware . The polsoftware . For thesoftware  using thsoftware . The unbam (W&C)  using GEam (W&C) .Of the total samples (n = 84) collected, 35 samples could not be used for the genetic analysis as these hair samples were either devoid of roots or too few in numbers to go for DNA extraction. Seven samples did not amplify while six samples did not produce consensus genotypes after duplicate genotyping. We observed fuzzy profile for locus Ca-60 even after repeating several times and this ambiguity was found to be consistent with all the samples forcing us to remove this locus from the initial set. Thirty six samples produced consensus genotypes after duplicate genotyping and analyzed for further analysis.1) and FA (ϵ2) error rates were not significant for any of the loci except for locus Ca-30 that exhibited considerably high ADO error rate i.e. 0.65 (Table ID sibs) was 0.009 (9 in 1000), while locus wise probability of matching genotypes among unrelated individuals (PID) and siblings (PID sibs) varied from 0.30-0.04 to 0.60-0.30, respectively . The observed heterozygosity in the population ranged between 0.08 and 0.83, with mean 0.40 ± 0.11 and expected heterozygosity ranged between 0.41 and 0.86 with mean 0.66 ± 0.07. Thus the population showed a low genetic diversity when compared to the other cervids - reindeer (0.733), red deer (0.554), roe deer  indicated heterozygote deficiency due to inbreeding with mean inbreeding coefficient to be 0.38 ± 0.15. This estimate was considerably higher than the reported estimates in other cervids - Vietnamese sika deer  and we postulate heterozygote deficit and presence of null alleles could be the reason of HWE departure [P < 0.05). Heterozygosity of locus Ca-30 was found to be offensive and the extremely low magnitude of observed heterozygosity ‘close to zero’ was duly supported by heterozygote deficiency due to large allele drop out and the presence of considerably high frequency of null allele , Chitall.[et al.), Vietnal.[et al.), musk dl.[et al.). The FIon et al.) and musl.[et al.. Howeverl.[et al.. We compeparture . Mutatioe.g. dominant males. In polygynous species such as hangul, the dominant males have their specific territories leading to meta-populations which certainly show allelic frequency shift over generations. Logically, a single dominant male may easily increase a particular set of alleles among his progenies as it will be preferred for mating by many of the female hangul deer in harems. Bebié and McElligott [Higher observed than expected heterozygosity for loci Ca-38 and Ca-42 remained cryptic and this may be attributed with the facts that the contribution of individual breeding animals is not equal to the next generation, Elligott  also proElligott ,42. The hangul population is considerably inbred and has a low genetic diversity when compared to other deer populations of the world. In particular, such losses of genetic diversity, reduced viability and fecundity due to inbreeding (inbreeding depression) are of concern to the long term survival of the species. We remained conservative in making strong inferences due to small sample size and involving only a few number of microsatellites. However, we present the constitutive genetic attributes of hangul population of Dachigam National Park that needs to be followed by the inclusion of more microsatellites for population monitoring of hangul in this area. We recommend the entire hangul population in and around Dachigam National Park should be investigated further in combination with a good sampling strategy to investigate species biology  and to prioritize the potential individuals retaining high heterozygosity for conservation breeding program.This preliminary study showed that the The authors declare that they have no competing interests.Mukesh conceived the idea, participated in its design and coordinated in writing the manuscript. LKS and SAC collected samples. Mukesh carried out PCR assays, genotyping, data analysis and wrote the manuscript. SS, SPG, LKS and SAC assisted in improving the MS with their comments. VPK and NM assisted in wet lab experiments. All authors read and approved the final manuscript.Authors’ biographyMukesh (Ph.D.) - Young Scientist affiliate at Wildlife Institute of India (WII), Dehradun and engaged in the conservation genetics of Cheer Pheasant in Himachal Pradesh, India. Lalit K. Sharma (Ph.D.) - Biodiversity Communication Expert at Indian Council of Forestry Research and Education (ICFRE), Dehradun, India. Samina A. Charoo (Ph.D.) – Research Officer at Department of Wildlife Protection, Jammu & Kashmir, India. Ved P. Kumar and Nipun Mohan (M.Sc.) are research scholars at WII. S.P. Goyal (Ph.D.) and S. Sathyakumar (Ph.D.) are senior wildlife biologists at WII and involved in strengthening wildlife conservation in India for over 25 years through teaching, training and research.


As WES/WGS increases in sensitivity, however, family history will be critical for prioritizing variant analysis and adding perspective (pretest probability) to the interpretation of susceptibility genes and findings “incidental” to the clinical indication for testing. : a review of general genetic principles, modes of inheritance, family/individual specific risk assessment, an in depth discussion of the diagnosis and natural history, potential testing options, and case management for the condition occurring within the family or for which they are at risk  approaches have evolved from these educational approaches. Genetic counselors and geneticists are trained in patient education and will continue to find roles in developing interactive educational content across many of these venues. I encourage research on the effectiveness of these approaches, and clinicians may need to have multiple educational approaches available to address the varied learning styles of patients.Genetic counselors and clinical geneticists have traditionally served as the “genetic experts,” in medicine, often in a consulting role despite being a primary medical specialty. In the past decade, genetic counselor roles have expanded significantly from the original prenatal and pediatric genetic counselor roles. A significant minority have taken on “laboratory genetic counselor” roles (National Society of Genetic Counselors One thing that separates genetic counselors from other health professionals with expertise in genetics is their stated focus on the psychosocial adaption to genetic conditions or genetic risk (Biesecker and Peters We have moderately good data about the psychosocial impact of learning carrier status (Lewis et al. Given the psychosocial implications of learning that one carries a highly penetrant condition that may have limited medical actionability, and given the worldwide history of eugenics and stigma associated with genetic conditions, there has been a strong focus on individual autonomy and non-directiveness around genetic testing decisions and future medical management (Weil et al. The profession of genetic counseling has undergone many transitions since its inception over 40 years ago; it has expanded from a primarily pediatric and obstetric focus at a time when genetic testing did not even exist, into multiple medical specialties that have access to rapidly changing genetic tests. The unique skills and roles of clinical geneticists and genetic counselors will become even more paramount, and genomic counseling will evolve in ways that preserve the central tenets of values-based decision making for patients while also promoting patient health outcomes.


Enhydris subtaeniata  using sequence data from three mitochondrial fragments . We sampled E. subtaeniata from seven locations in three river drainage basins that encompassed most of this species’ range. Genetic diversities were typically low within locations but high across locations. Moreover, each location had a unique suite of haplotypes not shared among locations, and pairwise φST values (0.713–0.998) were highly significant between all location pairs. Relationships among phylogroups were well resolved and analysis of molecular variance (AMOVA) revealed strong geographical partitioning of genetic variance among the three river drainage basins surveyed. The genetic differences observed among the populations of E. subtaeniata were likely shaped by the Quaternary landscapes of Indochina and the Sunda Shelf. Historically, the middle and lower Mekong consisted of strongly dissected river valleys separated by low mountain ranges and much of the Sunda Shelf consisted of lowland river valleys that served to connect faunas associated with major regional rivers. It is thus likely that the contemporary genetic patterns observed among populations of E. subtaeniata are the result of their histories in a complex terrain that created abundant opportunities for genetic isolation and divergence yet also provided lowland connections across now drowned river valleys.During the Cenozoic, Southeast Asia was profoundly affected by plate tectonic events, dynamic river systems, fluctuating sea levels, shifting coastlines, and climatic variation, which have influenced the ecological and evolutionary trajectories of the Southeast Asian flora and fauna. We examined the role of these paleogeographic factors on shaping phylogeographic patterns focusing on a species of semiaquatic snake,  Despite being only 4% of the Earth's land surface, Southeast Asia is a global biodiversity hotspot with 20–25% of the planet's animal and plant species . During Previous studies have identified various geographic features of the region that have likely influenced geographic variation in the distribution of terrestrial, freshwater, and marine species of this region, as well as the patterns of genetic diversity and population structure. In particular, Indochina has a complex geological history, with a number of potentially important physiographic events occurring in geologically recent times (Quaternary) that certainly affected many taxa  and the Chao Phraya populations, indicative of the historical connection between these two river systems. Oryzias minutillus, that correlated with river systems in Thailand and provide evidence of past historical connections between the Chao Phraya and Mekong systems. Henicorhynchus siamensis, from the Middle and Lower Mekong and suggest that the Khone Falls  from three mitochondrial genes. We sampled across the three major river drainages that encompass most of this species’ current range and used phylogenetic and population genetic approaches to evaluate several hypotheses about the effects of Quaternary geological and climatic processes on diversification. Population genetic analyses were used to assess the amount of genetic divergence and population genetic structure and we applied known mutation rates to estimate the timing of lineage divergences, and compared these with the timing of geological and climatic events in Indochina during the Quaternary. We then used phylogenetic methods to evaluate the relationships among major evolutionary lineages and the degree of concordance between phylogroups and the geographical configuration of river drainage basins. Finally, isolation-by-distance analyses were used to evaluate whether genetic divergences reflected geographical distances among populations. Our findings indicate that the historical biogeography of Indochina has strongly influence contemporary population structure and these findings are evaluated in the context of previous molecular studies in the region.This study aimed to improve our understanding of the effects of historical biogeography in Indochina by examining phylogeographic patterns for E. subtaeniata (n = 48) from three river basins encompassing most of this species’ range (n = 26); the Lower Mekong as the Mekong and tributaries below the Khone Falls to the South China sea (n = 7); and the Chao Phraya in Thailand (n = 15). We collected snakes from seven locations in these three river basins: Ban Mai and Ban Nong Pueng (Khon Kaen region) from the Chi tributary of the Middle Mekong; Ban Badan from the Mun tributary of the Middle Mekong, and Dong Khanthung from the Mekong proper; U. Minh Thong and Tonle Sap in the Lower Mekong, and Bung Boraphet in the Chao Phraya . Three mitochondrial fragments were selected for analyses based on their previous usefulness for evaluating phylogenetic and phylogeographic patterns in snakes : ATPase  primers  and prot primers , and visFST and φST values were calculated for the ten possible comparisons among these five locations: φST using the Tamura–Nei model of sequence evolution. Two Analyses of Molecular Variance (AMOVAs) were performed. The first AMOVA included the five locations with sample sizes >5, in order to evaluate genetic divergence among sampled locations. The second hierarchical AMOVA included all seven sampled locations grouped into three drainage basins, in order to evaluate our biogeographic hypotheses  implemented in TCS 1.13 , and maxtaeniata  for whictaeniata . Both MP–1 million–1 years (95% highest posterior densities [HPD] 0.0059–0.013) for mitochondrial cytochrome b and ND1 genes, respectively, for a semiaquatic snake. These estimated evolutionary rates correspond to 1.6–2.2% sequence divergences between a pair of lineages per million years (95% HPD 1.2–2.6%), typical of the mitochondrial molecular clock (MLclock) (clock hypothesis using a likelihood ratio test that compared the likelihood scores of the unconstrained best ML tree (MLbest) and an alternative tree with an enforced MLclock that was constructed in PAUP* (clock– MLbest] and tested against a chi-square distribution with df = number of taxa minus two) did not reject the clock hypothesis . As such, we used the MLclock estimates of E. subtaeniata phylogroups.MLclock) . We testin PAUP* . The likE. subtaeniata tends to be confined to aquatic habitats, we also tested whether distances through suitable habitats (contemporary drainage basins) correlated better with genetic divergence. ST/1 –φST for Mantel tests, however φST values can be unreliable for small sample sizes. As such, we used both φST/1 –φST and corrected “p” distances as genetic distances for the Mantel tests.Mantel tests of correlations between genetic and geographic distance matrices, implemented in the computer program Isolation by Distance Web Service (IBDWS) , were usb—1122 bp) with 123 variable sites  that described 18 putative haplotypes among 48 individuals (h = 0.88) but within-location haplotype diversities were generally low (0–0.42)  and highly significant between all location pairs (n≥ 5) when considering haplotype frequencies alone , while 95.9% of the genetic variation was attributed to differences among the five locations when sequence divergences among haplotypes were taken into account . Hierarchical AMOVA accounting for sequence divergences among haplotypes revealed strong  population subdivision at all levels, with 48.9% of genetic variation partitioned among the three drainage basins (φCT = 0.489), and 47.5% of genetic variation partitioned among locations within regions (φSC = 0.929), with an overall φST of 0.964.The final alignment comprised 2785 bp  . The two(0–0.42) . Each saon pairs . Strong E. subtaeniata was strongly supported by parsimony bootstrap values and many intraspecific clades also were well resolved  ranged from 0.80 to 0.85 , suggestp” distances. These tests demonstrated a strong and statistically significant correlation between genetic distances and geographic distance along drainage basins  and the strong geographical structuring of haplotypes into local populations  would likely prove interesting and perhaps further clarify barriers to gene flow. However, the collection sites for E. subtaeniata in Indochina are perhaps best understood in the context of the Quaternary terrain associated with the Upper and Lower Mekong, the Chao Phraya River, and the Sunda Shelf. The landscapes of both the Khorat Basin of the Upper Mekong and the Great Lake Basin (Tonle Sap) of the Lower Mekong consisted of strongly dissected river valleys separated by numerous low mountain ranges in the early and mid Quaternary (E. subtaeniata populations in Indochina is primarily a product of their histories in a terrain that imposed abundant opportunities for genetic isolation and independent evolution, as well as lowland connections across now drowned river valleys.The substantial genetic differences among the populations of aternary . This hi


Margarella antarctica. To previously published data from five populations spanning a 1500 km transect along the length of the Western Antarctic Peninsula, we added new AFLP data for four populations separated by up to 6 km within Ryder Bay, Adelaide Island. Overall, we found a nonlinear isolation-by-distance pattern, suggestive of weaker population structure within Ryder Bay than is present over larger spatial scales. Nevertheless, significantly positive Fst values were obtained in all but two of ten pairwise population comparisons within the bay following Bonferroni correction for multiple tests. This is in contrast to a previous study of the broadcast spawner Nacella concinna that found no significant genetic differences among several of the same sites. By implication, the topshell's direct-developing lifestyle may constrain its ability to disperse even over relatively small geographic scales.Understanding the relationship between life-history variation and population structure in marine invertebrates is not straightforward. This is particularly true of polar species due to the difficulty of obtaining samples and a paucity of genomic resources from which to develop nuclear genetic markers. Such knowledge, however, is essential for understanding how different taxa may respond to climate change in the most rapidly warming regions of the planet. We therefore used over two hundred polymorphic Amplified Fragment Length Polymorphisms (AFLPs) to explore population connectivity at three hierachical spatial scales in the direct developing Antarctic topshell  Marine species exhibit extreme heterogeneity in dispersal ability as estimated from genetic data Although several recent meta-analytical and comparative studies lend support to these theoretical expectations In addition to the above factors, our ability to draw general conclusions in respect of the relationship between life history and population structure is also hindered by a bias in the literature towards species from low latitudes Antarctica provides an unparalleled opportunity to undertake studies of the origins and maintenance of biological diversity, both at the levels of species and populations Consistent with predictions based on life history, several Antarctic brooding species have been found to be so highly structured as to invoke cryptic speciation et al. Nacella concinna and the brooding topshell Margarella antarctica sampled at five locations along the Antarctic Peninsula , each of which was in turn sub-sampled three times to provide a fine-scale geographic perspective An intriguing aspect of the above study was the finding that M. antarctica populations within Ryder Bay more structured than those of N.concinna as predicted by differences in their life history? Also, does life history influence the pattern of microgeographic population structure, or is this shaped in both species by the same underlying processes? To address these questions, we expanded our previous macrogeographic M. antarctica dataset to incorporate 174 additional individuals sampled from four sites over a 1–6 km spatial scale within Ryder Bay. All 414 individuals were genotyped at 228 polymorphic AFLP loci.The above studies raise additional questions in a comparative context. For example, are M. antarctica samples were collected by SCUBA divers during the austral summer of 1999 from the shallow sublittoral zone off Rothera Point (East Beach), Leonie Island (Leonie North North East) and from Anchorage North and Trolval on Anchorage Island  as described in detail by Hoffman et al. enalex v6 The program AMargarella antarctica populations spanning the Antarctic Peninsula Fst = 0.0872, P<0.0001). Restricting the analysis to the five populations sampled within Ryder Bay, global Fst was lower (0.0181) but still highly significant (P<0.0001), indicating the presence of fine-scale population genetic structuring. The relationship between geographic and genetic distance was highly significant overall , with a power regression fitting the data considerably better (r2 = 0.811) than linear, logarithmic or second order polynomial regressions . Restricting the analysis to populations within Ryder Bay, the isolation-by-distance relationship remained positive but failed to reach statistical significance .To a previous dataset comprising five Fst values for each of the pairwise population comparisons within Ryder Bay ranged from 0.003 to 0.034 .to 0.034 . All butM. antarctica to allow a comparison over three hierarchical spatial scales with the broadcast-spawning limpet N. concinna. Consistent with expectations, M. antarctica populations were structured throughout Ryder Bay, with the overall magnitude of genetic differentiation being greater than previously found in equivalent comparisons involving N. concinna. This lends further support to the notion that life history could be an important determinant of population structure in many benthic marine species.Although studies of the population genetic structure of marine species are commonplace, relatively few have explicitly compared direct with indirect-developers, particularly over multiple geographic scales and using the same methodologies with samples from the same sites. Consequently, we extended previous work on the brooding top shell M. antarctica sampled only five populations from the Western Antarctic Peninsula r2 = 0.811 versus 0.661, Because the previous study of Fst was still significantly non-zero  and around twenty times larger than the equivalent value for N. concinna based on the same five populations . Such a marked difference could potentially reflect the influence of Antarctic conditions on the two species' life histories. For example, larval lifetimes tend to be longer in species adapted to cold climates Laternula elliptica can live for up to 36 years Adamussium colbecki for over 100 years Despite population structure being relatively weak within Ryder Bay, global M. antarctica and N. concinna not only differed in the strength of population structure, but also in the way in which genetic variation was partitioned within Ryder Bay. All but two pairwise comparisons in M. antarctica were statistically significant following Bonferroni correction for multiple tests, suggesting that population structure although weak may extend across much if not all of the bay. In contrast, most of the N. concinna populations from Ryder Bay were indistinguishable from one another, with only three populations from Anchorage Island revealing significant genetic differences A related observation is that M. antarctica is benthic topology. Although Ryder Bay is only 15–20 km across, it attains a maximum depth of over 500 metres M. antarctica to disperse over soft sediments is greatly diminished relative to hard substrata such as bed rock or loose rubble. This hypothesis would be amenable to testing using a ‘landscape genetics’ approach if sufficiently fine-scale data could be collected on depth and substrate distribution across Ryder Bay. It is noteworthy in this context that the only comparison in which we did not obtain a significant Fst value prior to Bonferroni correction involved the two geographically closest M. antarctica populations, Trolval and Anchorage North, which were situated less than one kilometre apart on the coast of Anchorage Island. These two sites are separated by rocky coast and around 100 m of soft sediment at depths of 30–40 m (L. Peck pers. obs). However both sites are on the north side of Anchorage Island where steep to vertical rocky slopes descend to beyond 300 m. It is therefore possible that gene transfer between these two sites could take place along the rocky continuum beyond 40 m.Another factor that could potentially contribute to population structure in This study was made feasible through the use of AFLPs because these markers are capable of generating large numbers of genome-wide distributed bands in virtually any organism, including diverse Antarctic marine taxa M. antarctica and N. concinna individuals from Ryder Bay was fairly similar at 189 and 155 respectively, and the smaller panel is still reasonably large. Classical analyses of population structure also assume that all loci are selectively neutral, but some studies have reported conflicting results for different types of marker, suggesting that this may not always be the case As with any type of genetic marker, AFLPs can also be difficult to compare across species boundaries because different subsets of bands will be generated and these will invariably differ both in number and polymorphic information content. However, we believe this is unlikely to account for the contrasting strength and patterns of genetic structure observed in this study, since the number of informative bands obtained for Elsewhere, several factors other than life-history have also been invoked to explain the population structure of marine species. For example, Galarza et al. Finally, although our study design allowed us to control for technical factors such as the sampling scheme, choice of genetic marker and the genotyping and scoring protocols, we were not able to include further replicates at the same spatial scale from elsewhere that would help disassociate our findings from any conditions that could be specific to Ryder Bay. Detailed lichenographic studies suggest that the retreat of ice has been uneven across the bay, leaving behind a patchwork of habitats of varying ages M. antarctica to include a microgeographic component. Overall, a strong but nonlinear isolation-by-distance pattern was detected, indicating relatively weak population structure over scales of 1–6 km. Nevertheless, this was consistent across the bay and greater in magnitude than previously documented for the broadcast spawner N. concinna. By implication, life history variation, specifically protected versus broadcast reproductive modes, may impact the population structure of benthic marine species over multiple geographic scales. This could have implications for understanding how Antarctic marine invertebrates may respond to climate change, since dispersal is a key means by which locally extirpated populations might be replenished from adjacent locations.This study extended previous work on the direct developing top shell


Limanda limanda, a species used as a bioindicator of marine contaminants. Firstly, the spatiotemporal genetic structure of dab around UK waters (39 samples across 15 sites for four years: 2005–2008) is evaluated with 16 microsatellites. Two temporally stable groups are identified corresponding to the North and Irish Seas (average between basin Bioindicators are species for which some quantifiable aspect of its biology, a biomarker, is assumed to be sensitive to ecosystem health. However, there is frequently a lack of information on the underlying genetic and environmental drivers shaping the spatiotemporal variance in prevalence of the biomarkers employed. Here, we explore the relative role of potential variables influencing the spatiotemporal prevalence of biomarkers in dab,  Human activities often result in the transformation or pollution of habitats with high levels of contaminants. Aquatic environments are particularly threatened as they often serve as intentional or unintentional repositories of chemical contaminants (Islam and Tanaka Limanda limanda) is a key European bioindicator sentinel species  and expected heterozygosity (HS), locus-specific fixation indexes (FIS), deviations from Hardy–Weinberg equilibrium (HWE) and linkage equilibrium were tested in GENEPOP V4.0 . Global and pairwise amova) ; second, Mantel tests of correlation between pairwise amovas to test whether sexes were structured within samples (variance due to sexes within samples); and finally, the OSx-statistic . Two levels of biomarkers are considered here, each composed of several indicators and collectively encompassing a range of sensitivities to contaminant exposureAll contaminant and disease data were collected on behalf of the Clean Safe Seas Evidence Group and obtained via the Marine Environment Monitoring and Assessment National database (MERMAN) , epidermal papilloma (EP), skin ulceration (U) and skin hyperpigmentation (HYP), among 100 individuals were recorded in each of 35 samples , providing a more detailed examination of age, population and exposure on liver lesions, particularly neoplastic processes.Sex and age were available for individuals in the liver pathology exercise, while site age averages were obtained from the 2004 CSEMP assessment for the external analysis Fig. D. We assEstimates of contaminant exposure were obtained from the CSEMP data. Contaminant exposure is evaluated every year for five individuals per station . Heavy metal  and seven polychlorinated biphenyls , were measured from liver tissue. Averages of these values (within site and year) were used as site × year exposure estimates   and DAC5-70 (FIS = 0.371), showed heterozygote deficiencies , which measures the level of population structuring, including all samples and markers, suggested weak structuring of dab samples. There were, however, marked and consistent differences in allele frequencies for some loci among sites. The patterns of genic subdivision indicated that the allele distribution was consistently significantly different from random (P < 0.05) for several loci (38 of 64 loci–year comparisons), further suggesting the existence of genetic structure. Such allelic structure is illustrated in the CA  when comparing samples across sea basins. The pattern between North Sea and Irish Sea was consistent across years. Within the North Sea, NS1 samples were significantly differentiated from some of the near-shore North Sea and English Channel samples in some of the years  genetic differentiation when compared to most North Sea samples and some Irish Sea samples. The K-means clustering analysis suggested that k = 2 best fitted the data for all years, separating all North Sea and English Channel samples in one cluster from those collected in the Irish Sea and Atlantic. The amova (Table P < 0.001). When F-values were corrected for high heterozygosity, differentiation among samples within clusters remained low , the patterns of genetic distances (P < 0.05) with each other  and also displayed by the CA . Detail examination of amovas by loci revealed that one of the markers (DAC4-40) was responsible for most of such variance among sexes within samples, as amovas became nonsignificant after the removal of DAC4-40 , the very high among-sexes F′-values .e Deniel . Both sens Table . Altogetrkers DAC-40 was rStandardizing the data before variable importance analysis did not affect the ranking of the most important variables for each biomarker. However, it allowed distinguishing biomarkers significantly correlated with variables, as evaluated by the conditional inference trees, from those where no significant dependencies between any of the variables and biomarkers could be found. Three of the biomarkers, LY, U and EP, had low standardized scores Fig. , and conr = −0.460; P = 0.038), especially under 4 years of age .The conditional inference tree of the individual liver pathology classification highlighted how age, population structure and contaminant exposure all interacted and had significant effects on liver health Fig. . Age wasHO, FIS, IR and hL) were correlated with any of the biomarkers.None of the measures of genetic diversity at either samples or individual level  independently. Under such a scenario, regional differences in dab morphological traits Bakhsh , and mosNo measured variables  were associated with prevalence of three of the external biomarkers: LY, EP and U, indicating that either: (i) other contaminants not considered here may play a role in the prevalence of these diseases, (ii) studying their prevalence at sample level was not appropriate, and an individual model may be more adequate, (iii) they are not biomarkers of contaminant exposure. In accordance with the latter, Stentiford et al. found thAge and contaminant exposure have recognized effects in dab liver health  are characterized by a high prevalence of skin hyperpigmentation, ulcerations and frequent occurrence of neoplastic processes Cefas . The exiThe low prevalence of diseases in the English Channel may be a direct result of the absence of older fish (>4 years) at these sites, which typically exhibit higher disease prevalence than younger fish elsewhere (Stentiford et al. Although contaminants have been shown to be capable of impacting neutral genetic diversity (Bourret et al. heterogeneity of population-specific responses would require the application of high-throughput population genomic approaches targeting either candidate genes of known function or genomic regions under selection combined with translocation/common garden and exposure experiments (Stinchcombe and Hoekstra Potential for adaptation can be evaluated experimentally by analysing candidate genes on multigenerational selected lines (van Wijk et al. unequivocal adaptation to contaminant exposure in the wild would require the assessment of multigenerational changes at known functional genes, pre- and postexposure, while ratifying no population replacement (Hansen et al. Overall, our data reveal the existence of significant genetic structuring in dab at the regional scale of sea basins. The congruence among microsatellite markers and the observed temporal stability of genetic differentiation re-emphasizes the importance of temporal replicate sampling in elucidating genetic structuring among marine populations. The subtle differentiation found here may not necessarily imply high levels of gene flow among populations (Hauser and Carvalho within the same population boundaries. In doing so, environmental managers would avoid confounding variance potentially due to population-specific inherent susceptibility, or alternatively, resistance to disease, and that which is truly symptomatic of exposure to contaminants. Additionally, population-specific disease-frequency baselines can then be established with evolutionary meaningful geographical boundaries, allowing detection of finer variation in spatial patterns of disease prevalence. Population genetics, biomonitoring and functional genomic analyses should be integrated to further understand the underlying role of genetic variation in biomarker prevalence and heterogeneity in measures of exposure (Cerdá et al. Our study illustrates how genetic markers can elucidate population structure in a key biomonitoring species, delineating boundaries of comparable sampling sites and potential for differential response to environmental stress. It is recommended that biomonitoring programmes integrate the extent and spatiotemporal patterns of genetic structuring within assessments: individuals from target polluted sites should be compared to individuals from reference sites


Aedes polynesiensis is the primary vector of Wuchereria bancrofti in the South Pacific and an important vector of dengue virus. An improved understanding of the mosquito population genetics is needed for insight into the population dynamics and dispersal, which can aid in understanding the epidemiology of disease transmission and control of the vector. In light of the potential release of a Wolbachia infected strain for vector control, our objectives were to investigate the microgeographical and temporal population genetic structure of A. polynesiensis within the Society Islands of French Polynesia, and to compare the genetic background of a laboratory strain intended for release into its population of origin.A. polynesiensis samples collected in French Polynesia from 2005-2008 and introgressed A. polynesiensis and Aedes riversi laboratory strains. Examination of genetic differentiation was performed using F-statistics, STRUCTURE, and an AMOVA. BAYESASS was used to estimate direction and rates of mosquito movement.A panel of eight microsatellite loci were used to genotype FST values, AMOVA, and STRUCTURE analyses suggest low levels of intra-island differentiation from multiple collection sites on Tahiti, Raiatea, and Maupiti. Significant pair-wise FST values translate to relatively minor levels of inter-island genetic differentiation between more isolated islands and little differentiation between islands with greater commercial traffic . STRUCTURE analyses also indicate two population groups across the Society Islands, and the genetic makeup of Wolbachia infected strains intended for release is similar to that of wild-type populations from its island of origin, and unlike that of A. riversi.The observed panmictic population on Tahiti, Raiatea, and Moorea is consistent with hypothesized gene flow occurring between islands that have relatively high levels of air and maritime traffic, compared to that of the more isolated Maupiti and Tahaa. Gene flow and potential mosquito movement is discussed in relation to trials of applied autocidal strategies. Aedes polynesiensis is a day biting pest and the major vector of Wuchereria bancrofti and a secondary vector of Dengue virus in the South Pacific [A. polynesiensis established concurrent with the arrival of man in the South Pacific, approximately 1500-3000 years ago and has spread throughout French Polynesia and other island groups ranging from Fiji to the Tuamotu Archipelago [A. polynesiensis is adapted to ovipositing in both man-made  and natural containers [A. polynesiensis oviposits in include: coconut shells, rock holes, tree-holes, and crab holes generated by the gecardinid crab, Cardisoma carnifex (Herbst). The ability of A. polynesiensis to survive in numerous larval habitats has contributed to its widespread dispersal and the difficulty of control [ Pacific . A. polyhipelago . A. polyntainers . Natural control .A. polynesiensis have been largely unsuccessful, despite the use of insecticides, biological control, and larval habitat removal [A. polynesiensis in French Polynesia is important for defining the scale on which vector control using area-wide techniques is likely to be most effective. More specifically, characterization of the population genetic structure will help to define the level of population structuring, including the potential for cryptic subgroups that may be unaffected by releases of incompatible males [Attempts to control  removal ,4-6. Are removal . Understle males . An imprWolbachia-based incompatible insect technique (IIT) analogous to the sterile insect technique (SIT) has been suggested as an alternative for control of A. polynesiensis [Wolbachia-based IIT strategy, female sterility is artificially sustained by repeated inundative releases of incompatible males, resulting in mosquito population decrease and possibly elimination. A strain (CP) that harbors an incompatible Wolbachia infection has previously been developed for a potential release for field-testing of a Wolbachia-based IIT strategy in French Polynesia [A. riversi (AR) to transfer the maternally inherited endosymbiotic Wolbachia infection from A. riversi into the A. polynesiensis genetic background [A. polynesiensis collected from Maupiti served as the A. polynesiensis genotype donor. As a result of the introgression of the A. polynesiensis genetic background, it is important to ensure the genetic makeup of the CP strain is similar to the indigenous population and different from that of A. riversi prior to potential releases.A esiensis . In the olynesia . The CP olynesia ,9,10. Asckground . A. polyA. polynesiensis using isoenzymes have shown genetic differentiation between populations on islands from different archipelagos of French Polynesia [A. polynesiensis associated with different ecotypes, however, previous studies using isoenzymes have suggested that there is no genetic differentiation associated with habitat differences [Previous population genetic studies of olynesia . A more olynesia . Polynesferences .A. polynesiensis on a microgeographical scale, using samples collected from five islands in the Society Islands of French Polynesia. Vector population structure and movement patterns are relevant to the ecology and evolution of A. polynesiensis, gene flow and mosquito movement [Wolbachia-based population suppression strategy, to that of the indigenous population and A. riversi. Results are discussed in reference to the development of area-wide vector control efforts to control A. polynesiensis.The goal of the present study was to better understand the population genetic structure of movement , insectimovement , diseasemovement , previoumovement ,18, and movement ,19. FurtA. riversi) [2O and homogenized using a mini-bead beater  in a 1.5 ml eppendorf tube containing 100 μl of sterile diH2O and a 2.5 mm glass bead . The homogenate was then used for extraction with a DNeasy tissue kit , following manufacturer's instructions.Adult mosquitoes were collected in 2005, 2007, and 2008. Collection locations are denoted in Figure A. polynesiensis were characterized as part of a previous study . MICRO-. 2.9.3;  was used. 2.9.3; ). GENEPOT values  were cal alleles . To deteK hypothetical populations. Five independent runs were carried out for each value of K to check for consistency. To explore population genetic structure of all samples collected, K was allowed to vary from one to 12 using an admixture model. To examine for intra-island structure, K was allowed to vary for each island: Raiatea, one to nine, Maupiti, one to three, and Tahiti, one to four. For each analysis the burn-in length was set to 100,000 and the run length to 250,000 iterations. Real and simulated data have suggested that it is not straightforward to determine the optimal value of K when complex structure exists [K were used when examining the population genetic structure of all the islands sampled. The first method used was ΔK (a measure of the second order rate of change in the likelihood of K) [K. Differences are the result of 'label switching' of clusters across replicates and 'genuine multimodality'. CLUMPP [K value and does not effect the determination of K. Subsequently, DISTRUCT [Bayesian analysis was implemented in the program STRUCTURE , to infe exists . Hence, od of K) . The sec. CLUMPP  was usedDISTRUCT  was usedFST values and the natural logarithm of the straight-line distance between islands.To determine if geographic distance was influencing the observed pattern of genetic differentiation between main island groups, an isolation by distance analysis was conducted using Mantel tests within the sub-program Isolde (implemented in GENEPOP). Isolation by distance was tested by correlating linearized m, which is defined as the proportion of first generation migrants.As an additional analysis of genetic differentiation between island groups, a hierarchical analysis of population structure was performed, by estimating Nei's genetic distance values . Nei's gFST values were calculated using the ENA null allele correction method [FST also suggests little difference in FST values when comparing all populations with and without the ENA correction .Analyses of the ten-microsatellite loci show all to be polymorphic for at least one collection site Table . Locus An method . AlthougOut of 532 tests for linkage disequilibrium, four were significant after a Bonferroni correction. The four significant tests were between locus AP1 and APT2 in populations, MP, MPA, MOG2, and NF. However, in the remaining populations these loci remained completely unlinked. Hence, we found no evidence of subdivision among populations due to linked loci.FST value  between samples collected in March and July of 2008 on motu Toamaro. However, since FST values were low, samples were not differentiated in further analyses. No evidence of temporal genetic differentiation was suggested by the non-significant pair-wise FST values between March and November (0.030), and July and November (0.006) samples. Low and/or non-significant FST values suggest little evidence of temporal genetic variation between samples collected at different times at Gump Station, Moorea (-0.015). Since little to no differentiation was observed between temporal samples from motu Toamaro and Gump Station Moorea, all collections were included in the subsequent analyses examining for intra- and inter island differentiation.A low but a significant level of differentiation was suggested by the pair-wise P < 0.00001) . As another method to examine for intra-island genetic differentiation, a Bayesian analysis using STRUCTURE was performed. Results indicated sampled populations on Raiatea belonged to one genetic cluster. STRUCTURE was not able to show population substructure between mainland island samples of Raiatea and motu samples as suggested by pair-wise FST tests. Separate analyses using STRUCTURE were conducted to examine for population sub-structuring on Maupiti and Tahiti, demonstrating one genetic cluster each for samples from Maupiti and Tahiti.An AMOVA indicates little difference among populations within islands. Differences between populations within islands accounted for only 4.83% of the variation. However, the variation contributes significantly to analyses (1) Table , which mea Table . In addies Table . To deteFST values translate to a relatively minor level of genetic differentiation, and suggest Maupiti, Tahaa, and Moorea are differentiated from each other and Tahiti, while Raiatea is not significantly differentiated from Tahaa and Tahiti, but differentiated from Maupiti and Moorea  suggestK Figure . In FiguFST values associated with geographic distance, the correlation was not significant, suggesting geographic distance cannot fully explain the observed pattern of genetic structure (P > 0.32)  Figure .m >0.11)  Table . However1) Table .A. polynesiensis in Society Islands of French Polynesia. Results suggest minor levels of genetic variability of A. polynesiensis populations across the Society Islands. It was expected that the structuring of the Society Islands would have resulted in substantial genetic differentiation between different island groups based upon an island model of population differentiation [A. polynesiensis [Aedes taeniorhynchus among the Galapagos Islands attributed to human assisted mosquito movement of mosquitoes [Our study describes the population genetic structure of medically important ntiation . The obsesiensis . Resultssquitoes . The islsquitoes . FurtherFST values were observed when comparing Maupiti, Tahaa, and Moorea to the other islands. Tahaa, while geographically close to Raiatea, does not have an airport or large harbor; instead, it is primarily accessed by small boats. Maupiti has an airport, but it is the most geographically isolated of the sampled sites and receives relatively few flights, therefore may be less affected by human assisted movement of mosquitoes. Furthermore, the airport is located on a motu (population not sampled). Thus, based on the source-sink hypothesis, air introductions of mosquitoes to a motu would have a reduced probability of moving to the main island and other motus. Moorea is geographically close to Tahiti and receives a considerable amount of commercial traffic from Tahiti, suggesting considerable mosquito movement between these two islands as suggested by low pair-wise FST values (0.020) and significant levels of migration (m = 0.152) (Table Minor levels of genetic differentiation according to pair-wise 2) Table . MosquitFST estimates and suggest low levels of differentiation among islands except for Maupiti, which falls into a separate genetic cluster. Maupiti is the most geographically isolated island supporting the existence of two genetic clusters in the natural population. Log likelihood values also confirm the presence of three clusters two of which exist in the natural population and the other the A. riversi laboratory population. The observed clustering pattern also suggests that the CP strain has accumulated an allelic distribution, like that of wild type populations from Maupiti. The observed CP genotype is consistent with the introgression of the A. riversi genetic background into the A. polynesiensis laboratory strain created from a source on Maupiti [Wolbachia-based IIT strategy in French Polynesia. The confirmation that the genetic makeup is like that of the indigenous population, especially collections from Maupiti suggests the strain is genetically compatible with natural populations. Furthermore, the CP strain displays high rates of CI with naturally infected populations [Results from the STRUCTURE analysis agree with  Maupiti . As expeulations ,10, suggulations . In smalA. polynesiensis from different collection localities throughout the Society Islands may help determine the role of genetic differentiation on vector competence. While little population genetic structure was observed within islands, results suggest the presence of two distinct populations on less inhabited islands of Maupiti and Tahaa. Variation in genotype may be responsible for variation in vector competency, but is yet to be determined. Further experimentation examining the vector competence of different genotypes collected from different localities  may have implications for the epidemiology of disease transmission and will contribute to the planning of vector control strategies.The observed results also have epidemiological implications. Previous studies have suggested that variation in vector competence can be associated with geographic location ,18,40. FWolbachia-based IIT strategy [Wolbachia-based strategy is focused on population replacement, where the reproductive advantage afforded by Wolbachia induced CI as a population replacement strategy to drive wanted phenotypes into natural populations [A. polynesiensis is important for the design and interpreting the outcome of area-wide elimination techniques. If the genotype of released individuals is different than natural populations, emigrants and immigrants out of targeted and into control populations can be identified, respectively. These findings also highlight the importance of obtaining baseline population genetic data prior to area-wide control strategies to understand potential re-infestation events after such a strategy. Populations identified as genetic sinks may be better suited as initial release areas, minimizing the risk of released mosquitoes establishing in untargeted areas for IIT and replacement strategies. Data suggests that motu surrounding Maupiti are potential sites for initial releases of cytoplasmically incompatible males as part of an IIT or replacement strategy, since there is little evidence of emigration and immigration to and from the main island.Results presented here are relevant to the design and implementation of area-wide control strategies such as genetic control approaches   and autostrategy ,44-46. Aulations . UnderstWolbachia infected strain ; AMOVA:The authors declare that they have no competing interests.CLB and SLD conceived and designed the experiments. CLB performed the experiments. CLB analyzed the data. CLB and SLD wrote the paper and both authors approved the final version of the manuscript.