Genetic variability and the effect of habitat fragmentation in spotted suslik Spermophilus suslicus populations from two different regions

Endangered species worldwide exist in remnant populations, often within fragmented landscapes. Although assessment of genetic diversity in fragmented habitats is very important for conservation purposes, it is usually impossible to evaluate the amount of diversity that has actually been lost. Here, we compared population structure and levels of genetic diversity within populations of spotted suslik Spermophilus suslicus, inhabiting two different parts of the species range characterized by different levels of habitat connectivity. We used microsatellites to analyze 10 critically endangered populations located at the western part of the range, where suslik habitat have been severely devastated due to agriculture industrialization. Their genetic composition was compared with four populations from the eastern part of the range where the species still occupies habitat with reasonable levels of connectivity. In the western region, we detected extreme population structure (F _ST = 0.20) and levels of genetic diversity (Allelic richness ranged from 1.45 to 3.07) characteristic for highly endangered populations. Alternatively, in the eastern region we found significantly higher allelic richness (from 5.09 to 5.81) and insignificant population structure (F _ST = 0.03). As we identified a strong correlation between genetic and geographic distance and a lack of private alleles in the western region, we conclude that extreme population structure and lower genetic diversity is due to recent habitat loss. Results from this study provide guidelines for conservation and management of this highly endangered species.     

Allozyme variation and population genetic structure of common wild rice Oryza rufipogon Griff. in China

In order to determine the genetic diversity and genetic structure of populations in common wild rice Oryza rufipogon, an endangered species, allozyme diversity was analyzed using 22 loci in 607 individuals of 21 natural populations from the Guangxi, Guangdong, Hainan, Yunnan, Hunan, Jiangxi and Fujian provinces in China. The populations studied showed a moderate allozyme variability (A=1.33, P=22.7%, Ho=0.033 and He=0.068), which was relatively high for the genus Oryza. The levels of genetic diversity for Guangxi and Guangdong were significantly higher than those for the other regions, and thus South China appeared to be the center of genetic diversity of O. rufipogon in China. A moderate genetic differentiation (F_ST=0.310, I=0.964) was found among the populations studied. Interestingly, the pattern of population differentiation does not correspond to geographic distance. An estimate of the outcrossing rate (t=0.324) suggests that the species has a typical mixed-mating system. The deficit of heterozygotes (F=0.511) indicates that some inbreeding may have taken place in outcrossing asexual populations because of intra-clone outcrossing events and ”isolation by distance” as a result of human disturbance. In order to predict the long-term genetic survival of fragmented populations, further studies on gene flow among the remaining populations and the genetic effects of fragmentation are proposed. Finally, some implications for the conservation of endangered species are suggested. Received: 22 June 1999 / Accepted: 20 December 1999     

Genetic diversity and population structure of the endangered insect species Carabus variolosus in its western distribution range: Implications for conservation

Genetic variation was assessed in the endangered species C. variolosus (Coleoptera: Carabidae), which is listed in the European Habitats and Species Directive. Twelve populations from the north-western margin of its range in Germany and France to the border region between Slovenia and Croatia were investigated for variation at 16 allozyme loci. In general, genetic diversity was rather low as indicated by a mean allelic richness of 1.3 alleles per locus, a mean gene diversity (H _E ) of 0.071 and a mean proportion of polymorphic loci (P _95% ) of 16%. Genetic diversity did not change directionally from the margin towards the core of the range. Very high differentiation between populations (overall F _ST  = 0.465), the results of a Mantel test, and poor accordance between geographical and genetic distance suggest a high degree of isolation of the island-like distributed populations. High F _ST values and genetic distance measures, even between geographically close populations within the same drainage, confirm a very low dispersal power of this habitat specialist of headwater areas and swamps in woodlands, even in comparison with other flightless beetles. It is concluded that the majority of populations are demographically independent. Hierarchical F-statistics indicate that almost half of the genetic variance is found between regions and populations. Therefore, we recommend that conservation efforts consider every population as a management unit and aim to maintain as many populations as possible over as many regions as possible.     

Conservation genetics of the annual hemiparasitic plant <Emphasis Type="Italic">Melampyrum sylvaticum</Emphasis> (Orobanchaceae) in the UK and Scandinavia

Melampyrum sylvaticum is an endangered annual hemiparasitic plant that is found in only 19 small and isolated populations in the United Kingdom (UK). To evaluate the genetic consequences of this patchy distribution we compared levels of diversity, inbreeding and differentiation from ten populations from the UK with eight relatively large populations from Sweden and Norway where the species is more continuously distributed. We demonstrate that in both the UK and Scandinavia, the species is highly inbreeding (global F _IS = 0.899). Levels of population differentiation were high (F’_ST = 0.892) and significantly higher amongst UK populations (F’_ST = 0.949) than Scandinavian populations (F’_ST = 0.762; P < 0.01). The isolated populations in the UK have, on average, lower genetic diversity (allelic richness, proportion of loci that are polymorphic, gene diversity) than Scandinavian populations, and this diversity difference is associated with the smaller census size and population area of UK populations. From a conservation perspective, the naturally inbreeding nature of the species may buffer the species against immediate effects of inbreeding depression, but the markedly lower levels of genetic diversity in UK populations may represent a genetic constraint to evolutionary change. In addition, the high levels of population differentiation suggest that gene flow among populations will not be effective at replenishing lost variation. We thus recommend supporting in situ conservation management with ex situ populations and human-mediated seed dispersal among selected populations in the UK.     

High population differentiation in the rock-dwelling land snail (<Emphasis Type="Italic">Trochulus caelatus</Emphasis>) endemic to the Swiss Jura Mountains

Understanding patterns of genetic structure is fundamental for developing successful management programmes for isolated populations of threatened species. Trochulus caelatus is a small terrestrial snail endemic to calcareous rock cliffs in the Northwestern Swiss Jura Mountains. Eight microsatellite loci were used to assess the effect of habitat isolation on genetic population structure and gene flow among nine populations occurring on distinct cliffs. We found a high genetic differentiation among populations (mean F _ST = 0.254) indicating that the populations are strongly isolated. Both allelic richness and effective population size were positively correlated with the size of the cliffs. Our findings support the hypothesis that T. caelatus survived on ice-free cliffs during the Pleistocene glacier advancements from the Alps. Due to the establishment of beech and pine forest under recent, temperate climate conditions, dispersal between cliffs is no longer possible for rock-dwelling snails such as T. caelatus. Our results provide basic data for developing a conservation action plan for this endangered gastropod species.     

Genetic diversity of the endangered endemic Agave victoriae-reginae (Agavaceae) in the Chihuahuan Desert

Long-lived perennials are a species-rich, ecologically important component of the North American deserts, yet we know little about their genetic structure, information important for their conservation. Agave victoriae-reginae is an endemic of the Chihuahuan Desert of northern Mexico that is endangered by collection for the ornamental trade. We examined levels and patterns of variation at ten polymorphic allozyme loci in ten populations representing the range of the species. Levels of genetic variation (mean H_e= 0.335) and differentiation (mean F_ST = 0.236) were high. Phenetic clustering suggested the existence of at least three distinct groups of populations. If this pattern of variation is representative of other long-lived desert perennials, it may explain the species richness of this group and will pose a real challenge to gene conservation efforts.     

Species‐level view of population structure and gene flow for a critically endangered primate (Varecia variegata)

Lemurs are among the world's most threatened mammals. The critically endangered black‐and‐white ruffed lemur (Varecia variegata), in particular, has recently experienced rapid population declines due to habitat loss, ecological sensitivities to habitat degradation, and extensive human hunting pressure. Despite this, a recent study indicates that ruffed lemurs retain among the highest levels of genetic diversity for primates. Identifying how this diversity is apportioned and whether gene flow is maintained among remnant populations will help to diagnose and target conservation priorities. We sampled 209 individuals from 19 sites throughout the remaining V. variegata range. We used 10 polymorphic microsatellite loci and ~550 bp of mtDNA sequence data to evaluate genetic structure and population dynamics, including dispersal patterns and recent population declines. Bayesian cluster analyses identified two distinct genetic clusters, which optimally partitioned data into populations occurring on either side of the Mangoro River. Localities north of the Mangoro were characterized by greater genetic diversity, greater gene flow (lower genetic differentiation) and higher mtDNA haplotype and nucleotide diversity than those in the south. Despite this, genetic differentiation across all sites was high, as indicated by high average F_ST (0.247) and Φ_ST (0.544), and followed a pattern of isolation‐by‐distance. We use these results to suggest future conservation strategies that include an effort to maintain genetic diversity in the north and restore connectivity in the south. We also note the discordance between patterns of genetic differentiation and current subspecies taxonomy, and encourage a re‐evaluation of conservation management units moving forward.     

Population genetics of the endangered Wattled Curassow (Crax globulosa,Cracidae, Aves) of the Colombian–Peruvian Amazon using DNA microsatellites and ND2 mitochondrial sequences

The Wattled Curassow (Crax globulosa, Cracidae, Aves) is a large bird living in the Western Amazon basin and a critically endangered species in the Colombian and in the Peruvian Amazon. We carried out the first population genetics analysis of this species employing six nuclear microsatellite markers and sequences of the mtND2 gene. The main results are as follows. (1) The levels of gene diversity were high for the overall population as well as for each of the three islands for both microsatellites and mtDNA. (2) A small amount of genetic differentiation among populations was found with both types of markers (F_ST = 0.027 for microsatellites and N_ST = 0.17 for mitochondrial sequences). (3) Using microsatellites, the Geneclass 2.0 software detected a low correct assignment of individuals to their respective populations. The Structure software only detected one gene pool for the entire area studied. These results are relevant for conservation efforts of this critically endangered species.     

Population structure and genetic diversity of Rana dalmatina in the Iberian Peninsula

The increasing fragmentation of natural habitats may strongly affect patterns of dispersal and gene flow among populations, and thus alter evolutionary dynamics. We examined genetic variation at twelve microsatellite loci in the Agile frog (Rana dalmatina) from 22 breeding ponds in the Iberian Peninsula, the southwest limit of its range, where populations of this species are severely fragmented and are of conservation concern. We investigated genetic diversity, structure and gene flow within and among populations. Diversity as observed heterozygosities ranged from 0.257 to 0.586. The mean number of alleles was 3.6. Just one population showed a significant F _IS value. Four populations show evidence of recent bottlenecks. Strong pattern of structure was observed due to isolation by distance and to landscape structure. The average degree of genetic differentiation among populations was F _ST = 0.185. Three operational conservation units with metapopulation structure were identified. Additionally, there are some other isolated populations. The results reinforce the view that amphibian populations are highly structured even in small geographic areas. The knowledge of genetic structure pattern and gene flow is fundamental information for developing programmes for the preservation of R. dalmatina at the limits of its geographic distribution.     

Analysis of F ST outliers at allozyme loci in Pacific salmon: implications for natural selection

Natural selection has been invoked to explain the observed geographic distribution of allozyme allele frequencies for a number of teleost species. The effects of selection on allozyme loci in three species of Pacific salmon were tested. A simulation-based approach to estimate the null distribution of population differentiation (F _ST) and test for F _ST outliers was used. This approach showed that a majority of allozyme loci conform to neutral expectations predicted by the simulation model, with relatively few F _ST outliers found. No consistent F _ST outlier loci were found across species. Analysis of population sub-groups based on geography and genetic identity reduced the number of outlier loci for some species, indicating that large geographic groups may include genetically divergent populations and/or that there is geographic heterogeneity in selection pressure upon allozyme loci. Two outlier allozyme loci found in this analysis, lactate dehydrogenase-B and malic enzyme, have been shown to be influenced by selection in other teleost species. This approach is also useful in identifying allozyme loci (or other genetic markers) that meet assumptions for population genetic study.     

Genetic variation of wild litchi (<Emphasis Type="Italic">Litchi chinensis</Emphasis> Sonn. subsp. <Emphasis Type="Italic">chinensis</Emphasis>) revealed by microsatellites

Understanding the amount and distribution of genetic diversity in natural populations can inform the conservation strategy for the species in question. In this study, genetic variation at eight nuclear microsatellite loci was used to investigate genetic diversity and population structure of wild litchi (Litchi chinensis Sonn. subsp. chinensis). Totally 215 individuals were sampled, representing nine populations of wild litchi. All eight loci were polymorphic, with a total of 51 alleles. The expected heterozygosity in the nine populations ranged from 0.367 to 0.638 with an average value of 0.526. Inbreeding within wild litchi populations was indicated by a strong heterozygote defect. Significant bottleneck events were detected in the populations from Yunnan and Vietnam, which could be responsible for lower levels of genetic diversity in these populations. Measures of genetic differentiation (F _ST = 0.269) indicated strong differentiation among wild litchi populations. Significant correlation was found between genetic differentiation and geographical distance (r = 0.655, P = 0.002), indicating a strong isolation by distance in these populations. Bayesian clustering suggested genetic separation among three regional groups, namely, the western group, the central group and the eastern group. Some conservation strategies for wild litchi populations were also proposed based on our results.     

Evidence of recent population bottlenecks and inbreeding in British populations of Bechstein’s bat, Myotis bechsteinii

We investigated the population genetics of seven maternity roosts of Bechstein’s bats widely distributed across the south of England. Across all of the populations sampled, two mitochondrial DNA microsatellite loci were fixed for single haplotypes. Genetic diversity across eight nuclear microsatellite loci was similar in all seven populations, with a mean He of 0.727. However, six of the populations showed substantial homozygote excess, with F _IS estimates greater than zero, indicative of recent inbreeding. Bottleneck tests also implied that six of the populations have experienced recent declines. Genetic differentiation among the populations was low, with a mean intersite F _ST estimate of 0.041. There was no significant isolation by distance using allele frequency-based criteria (F _ST and genetic distances), however, a weak correlation was found using the allele size-based R _ST criterion. Assignment tests were unable to distinguish the seven sampling sites as distinct clusters. Mean intra-roost relatedness (r) was 0.079, indicative of recent inbreeding relative to German populations. All but one of the bats had one or more half or full siblings in its maternity roost. In addition, family relationships of individuals within a colony were significantly commoner than family relationships among four proximal roosts <8 km apart. The results are discussed in the context of conservation requirements for this rare British bat.     

Genetic diversity in fragmented populations of the critically endangered spider orchid Caladenia huegelii: implications for conservation

The Orchidaceae is characterised by a diverse range of life histories, reproductive strategies and geographic distribution, reflected in a variety of patterns in the population genetic structure of different species. In this study, the genetic diversity and structure was assessed within and among remnant populations of the critically endangered sexually deceptive orchid, Caladenia huegelii. This species has experienced severe recent habitat loss in a landscape marked by ancient patterns of population fragmentation within the Southwest Australian Floristic Region, a global biodiversity hotspot. Using seven polymorphic microsatellite loci, high levels of within-population diversity (mean alleles/locus = 6.73; mean H _E = 0.690), weak genetic structuring among 13 remnant populations (F _ST = 0.047) and a consistent deficit of heterozygotes from Hardy–Weinberg expectation were found across all populations (mean F _IS = 0.22). Positive inbreeding coefficients are most likely due to Wahlund effects and/or inbreeding effects from highly correlated paternity and typically low fruit set. Indirect estimates of gene flow (Nm = 5.09 using F _ST; Nm = 3.12 using the private alleles method) among populations reflects a historical capacity for gene flow through long distance pollen dispersal by sexually deceived wasp pollinators and/or long range dispersal of dust-like orchid seed. However, current levels of gene flow may be impacted by habitat destruction, fragmentation and reduced population size. A genetically divergent population was identified, which should be a high priority for conservation managers. Very weak genetic differentiation indicates that the movement and mixing of seeds from different populations for reintroduction programs should result in minimal negative genetic effects.     

Genetic diversity and differentiation of <Emphasis Type="Italic">Fagus orientalis</Emphasis> Lipsky in Hyrcanian forests revealed by nuclear and chloroplast microsatellite markers

Oriental beech (Fagus orientalis Lipsky) is a widespread monoecious and wind-pollinated tree species. It is one of the major components of the Hyrcanian forests of Iran and it is of both ecological and economical importance. Twelve beech stands were surveyed at 9 chloroplast (cp) and 6 nuclear (n) polymorphic microsatellite loci (simple sequence repeats, SSR) to provide information on distribution of genetic diversity within and among populations and on gene conservation and silvicultural management of this species. High levels of genetic differentiation were detected for the chloroplast genome (F _ST = 0.80 and R _ST = 0.95), in sharp contrast to the nuclear genome (F _ST = 0.06, R _ST = 0.05). The analysis of molecular variance (AMOVA) showed that 48% of the total cpSSR variation was attributable to differences among regions and 30% to differences among populations within regions, suggesting multiple origins of beech populations in Hyrcanian forests. Nuclear SSRs confirmed the presence of significant differentiation among populations and among geographic regions, even if, as expected, this was less pronounced than that found with cpSSRs (based on AMOVA, differences among regions and among populations within regions each contribute 5% to total nSSR variance). A highly significant correlation between genetic (nSSRs) and geographic distances (R ² = 0.522) was estimated, thus showing an isolation by distance effect. The application of spatial analysis of molecular variance (SAMOVA) using both marker data allowed identification of genetically homogeneous groups of populations. Possible applications of these results for the certification of provenances and/or seed lots and for designing conservation programs are presented and discussed.     

Signatures of diversifying selection at EST‐SSR loci and association with climate in natural Eucalyptus populations

Understanding the environmental parameters that drive adaptation among populations is important in predicting how species may respond to global climatic changes and how gene pools might be managed to conserve adaptive genetic diversity. Here, we used Bayesian F_ST outlier tests and allele–climate association analyses to reveal two Eucalyptus EST‐SSR loci as strong candidates for diversifying selection in natural populations of a southwestern Australian forest tree, Eucalyptus gomphocephala (Myrtaceae). The Eucalyptus homolog of a CONSTANS‐like gene was an F_ST outlier, and allelic variation showed significant latitudinal clinal associations with annual and winter solar radiation, potential evaporation, summer precipitation and aridity. A second F_ST outlier locus, homologous to quinone oxidoreductase, was significantly associated with measures of temperature range, high summer temperature and summer solar radiation, with important implications for predicting the effect of temperature on natural populations in the context of climate change. We complemented these data with investigations into neutral population genetic structure and diversity throughout the species range. This study provides an investigation into selection signatures at gene‐homologous EST‐SSRs in natural Eucalyptus populations, and contributes to our understanding of the relationship between climate and adaptive genetic variation, informing the conservation of both putatively neutral and adaptive components of genetic diversity.     

Signatures of adaptation and genetic structure among the mainland populations of Pinus radiata (D. Don) inferred from SNP loci

Insights into the relative contributions of locus specific and genome-wide effects on population genetic diversity can be gained through separation of their resulting genetic signals. Here we explore patterns of adaptive and neutral genetic diversity in the disjunct natural populations of Pinus radiata (D. Don) from mainland California. A first-generation common garden of 447 individuals revealed significant differentiation of wood phenotypes among populations (P _ST), possibly reflecting local adaptation in response to environment. We subsequently screened all trees for genetic diversity at 149 candidate gene single nucleotide polymorphism (SNP) loci for signatures of adaptation. Ten loci were identified as being possible targets of diversifying selection following F _ST outlier tests. Multivariate canonical correlation performed on a data set of 444 individuals identified significant covariance between environment, adaptive phenotypes and outlier SNP diversity, lending support to the case for local adaptation suggested from F _ST and P _ST tests. Covariation among discrete sets of outlier SNPs and adaptive phenotypes (inferred from multivariate loadings) with environment are supported by existing studies of candidate gene function and genotype–phenotype association. Canonical analyses failed to detect significant correlations between environment and 139 non-outlier SNP loci, which were applied to estimate neutral patterns of genetic differentiation among populations (F _ST 4.3 %). Using this data set, significant hierarchical structure was detected, indicating three populations on the mainland. The hierarchical relationships based on neutral SNP markers (and SSR) were in contrast with those inferred from putatively adaptive loci, potentially highlighting the independent action of selection and demography in shaping genetic structure in this species.     

Population genetics of the corallivorous gastropod Drupella cornus at Ningaloo Reef,Western Australia

A precipitous increase in the abundance of the corallivorous snail Drupella cornus at Ningaloo Reef, Western Australia, raises fundamental questions about the population structure and genetics of this species. We examined genetic heterogeneity at ten polymorphic allozyme loci among samples of adult D. cornus from nine sites along 180 km of Ningaloo Reef, plus two sites from the Abrolhos Islands and one from the Dampier Archipelago, spanning a total distance of 1170 km. Variations in allelic frequencies were small (average F_ST=0.007), indicating that a high degree of planktonic dispersal is the norm. Nevertheless, some heterogeneity among samples was found at four of the loci. This heterogeneity occurred within Ningaloo Reef and did not increase with geographic distance. The local heterogeneity was not a function of habitat type but seemed to be associated with stage of outbreak. However, all outbreak populations came from within Ningaloo Reef and the non-outbreak populations were from outside Ningaloo Reef proper. Our results show peculiarities in the genetic structure of D. cornus on Ningaloo Reef, but the causes are not understood. Correspondence to: K. Holborn     

Low genetic diversity and limited genetic structure across the range of the critically endangered Mexican howler monkey (Alouatta palliata mexicana)

Genetic diversity provides populations with the possibility to persist in ever-changing environments, where selective regimes change over time. Therefore, the long-term survival of a population may be affected by its level of genetic diversity. The Mexican howler monkey (Alouatta palliata mexicana) is a critically endangered primate restricted to southeast Mexico. Here, we evaluate the genetic diversity and population structure of this subspecies based on 83 individuals from 31 groups sampled across the distribution range of the subspecies, using 29 microsatellite loci. Our results revealed extremely low genetic diversity (H_O = 0.21, H_E = 0.29) compared to studies of other A. palliata populations and to other Alouatta species. Principal component analysis, a Bayesian clustering method, and analyses of molecular variance did not detect strong signatures of genetic differentiation among geographic populations of this subspecies. Although we detect small but significant F_ST values between populations, they can be explained by a pattern of isolation by distance. These results and the presence of unique alleles in different populations highlight the importance of implementing conservation efforts in multiple populations across the distribution range of A. p. mexicana to preserve its already low genetic diversity. This is especially important given current levels of population isolation due to the extreme habitat fragmentation across the distribution range of this primate.     

Extremely Low Levels of Allozyme Variation in Southern Korean Populations of the Two Rare and Endangered Lithophytic or Epiphytic <Emphasis Type="Italic">Bulbophyllum Drymoglossum</Emphasis> and <Emphasis Type="Italic">Sarcanthus Scolopendrifolius</Emphasis> (Orchidaceae): Implications for Conservation

Allozyme variation was investigated in two local populations of Bulbophyllum drymoglossum and three populations Sarcanthus scolopendrifolius, two rare and endangered lithophytes and epiphytes from South Korea. Genetic diversity was extremely low within populations (mean H _e = 0.011 for B. drymoglossum; 0.002 for S. scolopendrifolius). Among the putative screened 21 loci, we found only one polymorphic locus for each species. Only one polymorphic locus, detected just one population of each species, revealed significantly high degree of population differentiation between and among populations (F _ST = 0.253 for B. drymoglossum and F _ST = 0.899 for S. scolopendrifolius). These results suggest that genetic drift (consequence of a very small effective population size), coupled with a limited gene flow would play a major role in shaping population genetics of these species in South Korea. The current status of both species (small population sizes, spatially isolated populations, and highly localized habitats) in addition to the extremely low levels of genetic diversity and reckless collection of endangered orchids by plant sellers, significantly threaten the long-term survival of these species in Korea. Conservation of the two species requires both in situ strategies, by introducing of genets to increase effective population sizes by minimizing adverse effects (e.g., outbreeding depression and genetic swamping by non-native genotypes), and ex situ strategies, such as collection of genets from clonal ramets.     

Population genetic structure of the European ground squirrel in the Czech Republic

The European ground squirrel (Spermophilus citellus) is considered an endangered species with declining numbers throughout Europe, most pronounced at the western margin of its distribution area. Being extinct in Germany and Poland, the western margin of its distribution is in the Czech Republic. Here, landscape fragmentation has restricted the ground squirrels into few and very isolated localities where local extinctions still occur. In the present study we analysed European ground squirrels from six Czech and one Slovak localities using five microsatellite loci as genetic marker. The results show a strong genetic differentiation among the investigated populations (mean value of F _ST = 0.16) and high levels of inbreeding (values of F _IS ranged from 0.34 to 0.90). High level of inbreeding is generally considered to affect the viability of each population, which could lead to extinction. One of the most important factors is the lack of migration due to the large distances between the populations and the presence of migration barriers. Based on the results obtained we recommend a few suggestions for a conservation management of this species.