Rampant drift in artificially fragmented populations of the endangered tidewater goby (Eucyclogobius newberryi)

Habitat fragmentation and its genetic consequences are a critically important issue in evaluating the evolutionary penalties of human habitat modification. Here, we examine the genetic structure and diversity in naturally subdivided and artificially fragmented populations of the endangered tidewater goby (Eucyclogobius newberryi), a small fish restricted to discrete coastal lagoons and estuaries in California, USA. We use five naturally fragmented coastal populations from a 300‐ km spatial scale as a standard to assess migration and drift relative to eight artificially fragmented bay populations from a 30‐ km spatial scale. Using nine microsatellite loci in 621 individuals, and a 522‐base fragment of mitochondrial DNA control region from 103 individuals, we found striking differences in the relative influences of migration and drift on genetic variation at these two scales. Overall, the artificially fragmented populations exhibited a consistent pattern of higher genetic differentiation and significantly lower genetic diversity relative to the naturally fragmented populations. Thus, even in a species characterized by habitat isolation and subdivision, further artificial fragmentation appears to result in substantial population genetic consequences and may not be sustainable.     

Genetic analysis and conservation of 31 surviving individuals of a rare New Zealand tree, Metrosideros bartlettii (Myrtaceae)

Metrosideros bartlettii (Myrtaceae) is a distinctive and extremely rare tree, endemic to New Zealand, first discovered in 1975. Prior to this study, a total of 19 adult individuals of the species had been reported; these are located in three small forest remnants in the far north of the North Island of New Zealand. Here we describe a total of 31 adult M. bartlettii at the three sites, including 12 individuals newly discovered by us. We analyse the genetic diversity of the species, using microsatellites to examine the chloroplast genome and amplified fragment length polymorphisms (AFLPs) to monitor nuclear variation. The results clearly demonstrate that M. bartlettii is a unique species, distinct from its two closest relatives M. robusta and M. excelsa. Analysis of genetic diversity within the 31 remaining individuals of M. bartlettii showed an average heterozygosity (< H >) of 0.18 and a proportion of polymorphic genes (< P >) of 0.44. Population structure, as shown by 286 AFLP loci, varied between the three geographical sites; the site with fewest individuals, containing two trees, showed some separation from the populations at the other two locations. These two latter sites, by contrast, had highly overlapping AFLP population diversity profiles. The implications of these results for conservation of the species are discussed.     

Genetic variation in remnant populations of Dalbergia nigra (Papilionoideae), an endangered tree from the Brazilian Atlantic Forest

BACKGROUND AND AIMS: Dalbergia nigra, known as Brazilian rosewood, is an endangered tree species restricted to the Brazilian Atlantic Forest and has been intensively logged for five centuries due to its high-quality wood. The objective of the present study was to assess the genetic variation and structure in adults and saplings of the species from a large reserve of the Atlantic Forest, the Rio Doce State Park, and from two small surrounding fragments, one better preserved and another with a high degree of anthropogenic disturbance. METHODS: Analyses of genetic variation and structure were conducted by studying allozyme markers. Seven putative enzymatic loci were resolved, five of them being polymorphic. KEY RESULTS: The mean numbers of alleles per locus (A) were 1.93 and 1.73, while the percentages of polymorphic loci (P) were 93 and 73 % for adults and saplings, respectively. Saplings from the fragment with high anthropogenic disturbance exhibited the lowest values of A and P. The fragment that constitutes a conservation area exhibited genetic variation similar to the population from the large reserve. The observed (H(o)) and expected (H(e)) heterozygosities were not significantly different among the three populations. Only sapling populations showed F(ST) values (divergence among populations) significantly different from zero over all studied loci. The fragment with high anthropogenic disturbance exhibited considerable genetic divergence in relation to the above-cited populations. CONCLUSIONS: The evaluated populations displayed mean levels of genetic variation intermediate to those expected for narrow and widespread species. The results suggest that fragments with similar area and geographical distance from a large protected reserve can exhibit different levels of genetic variation, depending on the degree of anthropogenic disturbance. The considerable genetic variation in the protected fragment points to the importance of adequate conservation of small fragments for the preservation of genetic variation in D. nigra.     

Genetic diversity in Nothofagus alessandrii (Fagaceae), an endangered endemic tree species of the coastal maulino forest of Central Chile

BACKGROUND AND AIMS: The endemic tree Nothofagus alessandrii (Fagaceae) has been historically restricted to the coastal range of Region VII of central Chile, and its forests have been increasingly destroyed and fragmented since the end of the 19th century. In this study, the patterns of within- and among-population genetic diversity in seven fragments of this endangered narrowly endemic tree were examined. METHODS: Allozyme electrophoresis of seven loci of N. alessandrii was used to estimate genetic diversity, genetic structure and gene flow. KEY RESULTS: High levels of genetic diversity were found as shown by mean expected heterozygosity (H(e) = 0.182 +/- 0.034), percentage of polymorphic loci (P(p) = 61.2 %), mean number of alleles per locus (A = 1.8) and mean number of alleles per polymorphic locus (A(p) = 2.3). Genetic differentiation was also high (G(ST) = 0.257 and Nm = 0.7). These values are high compared with more widespread congeneric species. CONCLUSIONS: Despite its endemic status and restricted geographical range N. alessandrii showed high levels of genetic diversity. The observed patterns of diversity are explained in part by historical processes and more recent human fragmentation.     

中国特有濒危树种毛红椿核心和边缘居群的遗传多样性

选用8对多态的微卫星引物, 研究了毛红椿(Toona ciliata var. pubescens)核心居群和边缘居群的遗传多样性。研究结果显示, 边缘居群的观察等位基因数和有效等位基因数并不比核心居群低, 甚至更高。普通广布和稀有地方等位基因在所有居群中均有分布, 而普通地方和稀有广布等位基因分别仅在5个和3个居群中有分布。从4种类型等位基因总数上看, 边缘居群高于核心居群, 特别是边缘居群的稀有地方等位基因数量明显多于核心居群。边缘居群的平均期望杂合度和观察杂合度都高于核心居群。核心居群间的基因分化系数为0.1520, 边缘居群间为0.3045, 边缘居群与核心居群的遗传分化差异达到显著水平。核心居群间基因流大于1, 而边缘居群间基因流小于1, 说明核心居群间基因交流频繁, 边缘居群由于片段化和地形影响, 基因流较小。Mantel检验结果显示, 居群间遗传距离与地理距离的相关性不显著, 说明地理距离对毛红椿居群遗传分化的影响不显著。     

Isolation and characterization of microsatellite loci in an endangered tree species, Afzelia xylocarpa (Kurz) Craib (Caesalpinioideae)

Afzelia xylocarpa is listed as an endangered species on the International Union for the Conservation of Nature and Natural Resources World list of Threatened Trees, due to overexploitation for its valuable timber and habitat loss. We isolated eight polymorphic microsatellite loci from A. xylocarpa using dual suppression polymerase chain reaction technique. These loci provide microsatellite markers with high polymorphism, as the number of alleles ranged from two to seven, and the estimate of gene diversity was between 0.285 and 0.795. The markers are now available for more detailed investigation of population genetic structure and gene flow among A. xylocarpa populations.     

Genetic variation in a tropical tree species influences the associated epiphytic plant and invertebrate communities in a complex forest ecosystem

Genetic differences among tree species, their hybrids and within tree species are known to influence associated ecological communities and ecosystem processes in areas of limited species diversity. The extent to which this same phenomenon occurs based on genetic variation within a single tree species, in a diverse complex ecosystem such as a tropical forest, is unknown. The level of biodiversity and complexity of the ecosystem may reduce the impact of a single tree species on associated communities. We assessed the influence of within-species genetic variation in the tree Brosimum alicastrum (Moraceae) on associated epiphytic and invertebrate communities in a neotropical rainforest. We found a significant positive association between genetic distance of trees and community difference of the epiphytic plants growing on the tree, the invertebrates living among the leaf litter around the base of the tree, and the invertebrates found on the tree trunk. This means that the more genetically similar trees are host to more similar epiphyte and invertebrate communities. Our work has implications for whole ecosystem conservation management, since maintaining sufficient genetic diversity at the primary producer level will enhance species diversity of other plants and animals.     

Development of SCAR markers distinguishing pure seedlings of the endangered species Morus boninensis from M. boninensis × M. acidosa hybrids for conservation in Bonin (Ogasawara) Islands

To support the recovery of Morusboninensis Koidz. (Moraceae), a criticallyendangered species distributed in the BoninIslands of Japan, we need to distinguishbetween pure M. boninensis and hybrid(M. boninensis × M. acidosa Griff.)seedlings within populations derived from M. boninensis mother trees. Therefore, wedeveloped three co-dominant SCAR (SequenceCharacterized Amplified Region) markers todistinguish between M. boninensis, M. acidosa and their hybrids. All threemarkers could clearly identify the species andthe hybrids, although we observed one case of aputative null allele or a backcross at onelocus (Mos0251-1). Hence, we suggest that pureand hybrid seedlings could be unambiguouslyidentified by the simultaneous use of the threemarkers. When we investigated a seedlingpopulation produced by a single mother treegrowing in the Iguma-bay population ofHaha-jima Island, the proportion of embryosformed by pollination from M. acidosapollen was about 13%. All hybrid seedlingsidentified according to the molecular markersshowed incisions in their leaves, amorphological character that has never beenobserved in pure M. boninensisindividuals. Our results suggest that M.boninensis can potentially be multiply byexcluding hybrids from the seedling pools,using incisions on the leaves as morphologicalmarkers to identify them.     

Pollination and reproduction of Psychotria homalosperma,an endangered distylous tree endemic to the oceanic Bonin (Ogasawara) Islands,Japan

Distyly is usually rare or not observed in species thriving on oceanic islands. The rarity of this breeding system is probably because of the difficulty of colonization for distylous plants and the paucity of pollinators on oceanic islands. However, the endemic and endangered tree Psychotria homalosperma has maintained its distylous nature in the oceanic Bonin Islands, Japan. To understand how the distylous breeding system of P. homalosperma has been maintained on these islands and to characterize the reproduction systems, we studied the pollination and reproductive biology of this species. Specifically, we observed current flower visitors and estimated their effects on plant reproduction. We also examined the floral traits and floral volatiles of P. homalosperma to infer its original pollinators, because plant–pollinator relationships in the Bonin Islands have recently been disrupted by anthropological activities. Finally, we examined the fruit set and pollen tube growth in the stigmas under hand and open pollination. Although original pollinators were presumed to be moths with long proboscises, the introduced honeybee, Apis mellifera, was the most common flower visitor. The honeybee carried pollen grains only unidirectionally, from the short‐ to long‐styled morphs, because it could not reach the hidden stigmas of the short‐styled flowers, and long‐styled flowers set fruits 1.7–38 times more than short‐styled ones. This case study indicates that the instability of pollinator fauna can cause distylous species to be rare on oceanic islands.     

Genetic differentiation within and among island populations of the endangered plant Aster miyagii (Asteraceae), an endemic to the Ryukyu Islands

Genetic diversity was examined at 17 putative allozyme loci in 18 populations of the insular endemic plant Aster miyagii (Asteraceae). This species is geographically restricted to only three islands of the Ryukyu Islands and is on the federal list of threatened plants. Genetic differentiation within an island is small, suggesting that gene flow among populations on the same island is sufficiently large to prevent divergence. By contrast, genetic differentiation among islands is large, especially between Amamioshima Island and the other two islands, suggesting that gene flow between the islands is highly restricted. Two unique alleles are nearly fixed in populations on Amamioshima Island, which is the southernmost island of the three. Comparatively, genetic diversity is the smallest on Amamioshima Island. This genetic paucity on Amamioshima Island is probably a result of a population bottleneck at colonization or the small effective population size on this island. Genetic diversity at the species level of A. miyagii is larger than those of the species with a similar life history and of the congeneric widespread species, suggesting that the species has an old origin as an insular endemic species.     

Development and characterization of microsatellite markers for Hibiscus glaber Matsum. ex Nakai, an endemic tree species of the oceanic Bonin Islands, Japan

Polymorphic microsatellite markers were developed for Hibiscus glaber, an endemic tree of the Bonin Islands. Eighty-seven of the 208 sequences from an enriched library were unique and containing microsatellites. Ten loci were proved to be highly polymorphic among 78 individuals from the Nishi-jima Island. Total exclusionary powers for the first and the second parents were 99.989% and 99.999%, respectively. Nine loci also amplified single fragment from genomic DNA of H. tiliaceus, a related and widespread congener. Our markers can be reliably used for the estimation of current gene flow within/among populations of the two woody Hibiscus species.     

Isolation and characterization of microsatellite loci in Psilopeganum sinense Hemsl (Rutaceae), an endangered herb endemic to Yangtze River valley

Twenty-nine primer pairs flanking microsatellite repeats were designed from an AC-enriched genomic library of Psilopeganum sinense and tested using 24 individuals derived from a natural population. A total of 11 microsatellite loci were found polymorphic. The number of alleles per locus varied from two to eight, with an average value of 3.7. The ranges of observed and expected heterozygosities were 0.000-0.750 and 0.365-0.800, respectively. These microsatellite loci have been directly applied to the ongoing conservation genetic studies of P. sinense.     

Genetic diversity in an endangered alpine plant, Eryngium alpinum L. (Apiaceae), inferred from amplified fragment length polymorphism markers

Eryngium alpinum L. is an endangered species found across the European Alps. In order to obtain base-line data for the conservation of this species, we investigated levels of genetic diversity within and among 14 populations from the French Alps. We used the amplified fragment length polymorphism (AFLP) technique with three primer pairs and scored a total of 62 unambiguous, polymorphic markers in 327 individuals. Because AFLP markers are dominant, within-population genetic structure (e.g. FIS) could not be assessed. Analyses based either on the assumption of random-mating or on complete selfing lead to very similar results. Diversity levels within populations were relatively high (mean Nei's expected heterozygosity = 0.198; mean Shannon index = 0.283), and a positive correlation was detected between both genetic diversity measurements and population size (Spearman rank correlation: P = 0. 005 and P = 0.002, respectively). Moreover, FST values and exact tests of differentiation revealed high differentiation among populations (mean pairwise FST = 0.40), which appeared to be independent of geographical distance (nonsignificant Mantel test). Founder events during postglacial colonizations and/or bottlenecks are proposed to explain this high but random genetic differentiation. By contrast, we detected a pattern of isolation by distance within populations and valleys. Predominant local gene flow by pollen or seed is probably responsible for this pattern. Concerning the management of E. alpinum, the high genetic differentiation leads us to recommend the conservation of a maximum number of populations. This study demonstrates that AFLP markers enable a quick and reliable assessment of intraspecific genetic variability in conservation genetics.     

Across the great divide: genetic forensics reveals misidentification of endangered cutthroat trout populations

Accurate assessment of species identity is fundamental for conservation biology. Using molecular markers from the mitochondrial and nuclear genomes, we discovered that many putatively native populations of greenback cutthroat trout (Oncorhynchus clarkii stomias) comprised another subspecies of cutthroat trout, Colorado River cutthroat trout (Oncorhynchus clarkii pleuriticus). The error can be explained by the introduction of Colorado River cutthroat trout throughout the native range of greenback cutthroat trout in the late 19th and early 20th centuries by fish stocking activities. Our results suggest greenback cutthroat trout within its native range is at a higher risk of extinction than ever before despite conservation activities spanning more than two decades.     

Genetic diversity of Narcissus tortifolius,an endangered endemic species from Southeastern Spain

In this work, we analyzed inter-simple sequence repeat markers from 10 populations (298 individuals) spanning the whole distribution range of the endemic Narcissus tortifolius. We assessed genetic variation levels and distribution by estimates of genetic diversity, analysis of molecular variance (AMOVA), principal coordinates and Bayesian methods. N. tortifolius showed moderate genetic diversity at intrapopulation level and low genetic differentiation of populations. In general, Almerian populations showed slightly higher levels of genetic diversity than Murcian populations. Our results indicate that habitat fragmentation has not caused genetic depauperation in N. tortifolius but did reveal moderate genetic differentiation. Indeed, principal coordinate analysis and Mantel test revealed a slight tendency to separate populations into two groups (Murcian vs. Almerian). A recent isolation event of populations, together with the perennial nature of this species could be the main reasons for this low to moderate differentiation. Our findings could be used to establish management guidelines for the conservation of this rare species.     

Microsatellite analysis reveals interpopulation differentiation and gene flow in the endangered tree Changiostyrax dolichocarpa (Styracaceae) with fragmented distribution in central China

Polymorphic simple sequence repeat (SSR) markers were used to investigate the impact of habitat fragmentation on the population structure and gene flow of Changiostyrax dolichocarpa, a critically endangered tree in central China. Intrapopulation genetic diversity, population structure and gene flow in the five extant populations of this species were analysed by eight SSR markers. Intrapopulation genetic diversity results suggest that C. dolichocarpa remnants maintained a relatively high degree of genetic diversity despite severe fragmentation. Low genetic differentiation among populations was found based on Wright's F(ST) and amova analysis. Both the F(ST)-based estimate and private allele method revealed high historical gene flow among the remnant populations. Recent immigrants, detected by assignment tests, tend to decrease from the grandparent generation to the current generation. The potentially highly restricted current gene flow among fragments may render the fragmented populations of C. dolichocarpa at a higher risk of local extinction several generations after fragmentation. Both in situ and ex situ conservation management for the remnant populations of C. dolichocarpa are therefore urgently needed to rescue remaining genetic diversity.     

Genetic differentiation and diversity of Acacia koa populations in the Hawaiian Islands

Acacia koa A. Gray (koa) is a leguminous tree endemic to the Hawaiian Islands and can be divided into morphologically distinguishable groups of A. koaia Hillebrand, A. koa and populations that are intermediate between these extremes. The objectives of this investigation were to distinguish among divergent groups of koa at molecular levels, and to determine genetic diversity within and among the groups. Phylogenetic analyses using the ITS/5.8S rDNA and trnK intron sequences did not separate the representative koa types into distinct clusters. An unweighted pair group method with arithmetic mean cluster analysis and principal coordinate analysis, based on allele profiles of 12 microsatellite loci for 215 individual koa samples, separated the population into three distinct clusters consistent with their morphology, A. koaia, A. koa and intermediate forms. There was an average of 8.8 alleles per polymorphic locus (AP) among all koa and koaia individuals. The intermediate populations had the highest genetic diversity (H′ = 1.599), AP (7.9) and total number of unique alleles (21), whereas A. koaia and A. koa showed similar levels of genetic diversity (H′ = 0.965 and 0.943, respectively). No correlation was observed between geographic distance and genetic distance as determined by a Mantel test (r = 0.027, P = 0.91). The data presented here support previous recommendations that morphological variation within koa should be recognized at the subspecific level rather than as distinct species.     

Genetic profiling of the critically endangered palm species Medemia argun using newly developed chloroplast DNA markers

Background: Medemia argun is a rare wild palm tree species. Its global existence is assumed to include the main population of about 1000 trees in the Nubian Desert of Sudan and some scattered individuals in southern Egypt. The species had previously been assumed to be extinct, but then reported to be extant about 20 years ago.

Aims: To assess genetic variation and explore population genetic structure of M. argun, through development and analysis of microsatellite markers.

Methods: The genome sequence mining approach was applied in order to identify microsatellites in the chloroplast genome of Bismarckia nobilis, a species closely related of M. argun. A set of 49 markers were designed, and their characteristics are now provided. Seven chloroplast DNA markers were developed for use in the genetic characterisation of this threatened species.

Results: Five markers were found polymorphic in M. argun, which enabled the assessment of the genetic diversity of the species. Significant genetic differentiation was observed among generations and collection sites, accompanied by low genetic variation. The seven markers developed were polymorphic among the wild relatives Hyphaene thebaica and Borassus aethiopum.

Conclusions: This is the first study to report molecular markers for M. argun. Our results suggest that the genetic consequences of population fragmentation in M. argun are beginning to be evident.