Electrolyte leakage differences between poikilohydrous and homoiohydrous species of Gesneriaceae

Comparative analysis of the electrolyte efflux, as a screening test of the membrane tolerance to water stress, was carried out in poikilohydric plants Ramonda serbica Panč. and Ramonda nathaliae Panč. & Petrov. and homoiohydric plant Saintpaulia ionantha Wendl. from the same family Gesneriaceae. Water stress was induced by PEG 600. The high degree of solute leakage in the East-African drought-intolerant Saintpaulia ionantha points to the loss of membrane integrity. In contrast, Balkan endemites Ramonda serbica and R. nathaliae show high resistance to water stress due to the specific constitutional drought tolerance mechanisms. This revised version was published online in July 2006 with corrections to the Cover Date.     

Genetic variation and conservation assessment of Chinese populations of Magnolia cathcartii (Magnoliaceae), a rare evergreen tree from the South-Central China hotspot in the Eastern Himalayas

Nine natural populations of the rare evergreen tree Magnolia cathcartii (Magnoliaceae) were sampled across its natural range, and amplified fragment length polymorphism (AFLP) markers were used to assess genetic variation within and among populations. Three ex situ populations were also surveyed to determine whether conservation plantings include the entire genetic diversity of the species. Genetic diversity within the natural populations was very low (0.122 for Nei’s gene diversity), and the southeast populations had the highest diversity. The ex situ populations had a lower diversity than the mean diversity for all populations, and none of the ex situ populations reached the levels of diversity found in their source populations. Genetic differentiation was high among natural populations (G _st = 0.247), and an isolation-by-distance pattern was detected. Habitat fragmentation, restricted gene flow, and geitonogamy are proposed to be the primary reasons for the low genetic diversity and high genetic differentiation. More protection is needed, especially for the southeast populations, which possess the highest numbers of unique alleles according to AFLP fragment analyses. The ex situ program was a good first step towards preserving this species, but the current ex situ populations preserve only a limited portion of its genetic diversity. Future ex situ efforts should focus on enhancing the plantings with individuals from southeastern Yunnan.     

Genetic diversity in the US ex situ populations of the endangered Puerto Rican Boa,Chilabothrus inornatus

The Puerto Rican Boa (Chilabothrus inornatus) was placed on the US Endangered Species List in 1970. Progress has been made since to clarify the recovery status of this species, though the design of a new recovery plan must include information regarding genetic variation within and among populations of this species. While measures of genetic diversity in wild populations of this species are finally becoming available, relative genetic diversity represented in ex situ populations is unknown, which hampers efforts to develop an ex situ species management plan. Here, we provide an analysis of genetic diversity in US public and private collections (zoos and breeders) using mitochondrial sequence data and five highly polymorphic nuclear microsatellite loci. We analyzed 50 boas from the US ex situ population and determined overall genetic diversity and relatedness among these individuals. We then compared these data to mitochondrial and microsatellite data obtained from 176 individuals from wild populations across the native range of the species. We found little inbreeding and a large amount of retained genetic diversity in the US ex situ population of C. inornatus relative to wild populations. Genetic diversity in the ex situ population is similar to that found in wild populations. Ours is only the second explicit attempt to characterize genetic diversity at the molecular level in ex situ populations of boid snakes. We anticipate that these results will inform current breeding strategies as well as offer additional information that will facilitate the continuation of ex situ conservation breeding or management in boas.     

Comparative molecular studies on the genetic diversity of an ex situ garden collection and its source population of the critically endangered polish endemic plant <Emphasis Type="Italic">Cochlearia polonica</Emphasis> E. Fröhlich

Cochlearia polonica (Brassicaceae) is a narrow endemic plant extinct in the wild, known only from one transplanted population in southern Poland. The suitable habitats for this species are confined to shallow water courses and springs on sandy ground. During the second half of twentieth century the natural populations of C. polonica declined due to anthropogenic effects (primarily due to a major change of ground water levels). Consequently, 14 individuals were transplanted in the 1970 s into a secondary locality of similar habitat conditions of Centuria river. In the late 1980s five individuals from the transplanted population were used to establish an artificial ex situ site in the Botanical Garden of the Polish Academy of Sciences in Warsaw. In order to investigate the effect of 18 years of ex situ conservation, the level of genetic diversity and genetic structure of the ex situ conserved botanical garden population and its source population were sampled and analyzed using inter simple sequence repeat markers. The percentage values of polymorphic bands and Nei’s gene diversity indicated average genetic variation at species level, whereas at the population level it was relatively low, especially for the garden population. The results suggested differences in genetic composition of the analyzed populations and implied homogenization of the genetic structure of the ex situ conserved population. Ex situ conservation resulted in a decrease of the species’ genetic diversity, implying that the artificial population only partly represents the primary genetic variability found in the source population.     

Genetic diversity in natural populations of the endangered Neotropical orchid Telipogon peruvianus

Telipogon peruvianus is a highly restricted, sexually deceptive, Neotropical orchid species, endemic to the southern Peruvian Andes. It is only known from two localities, which are affected by anthropogenic disturbance. Here, we investigated whether the restricted distribution of T. peruvianus has led to low genetic diversity and inbreeding, thus threatening its survival. We isolated 10 novel microsatellite loci specific for T. peruvianus (and that also cross-amplified in related species) from two adjacent populations. We found that genetic diversity within populations was only moderately reduced, along with some evidence of inbreeding. We found low levels of genetic differentiation, suggesting connectivity by pollen/seed flow between the two populations. Effective population size was comparable to the real number of flowering individuals in the populations and we did not detect the signature of recent bottlenecks. Taken together, these results show that, despite increasing anthropogenic pressure, the two investigated populations of T. peruvianus still host valuable genetic diversity that should be preserved through appropriate conservation strategies.     

Effect of Yangtze River on population genetic structure of the relict plant Parrotia subaequalis in eastern China

Parrotia subaequalis (Hamamelidaceae) is a Tertiary relic species endemic in eastern China. We used inter‐simple sequence repeat (ISSR) markers to access genetic diversity and population genetic structure in natural five populations of P. subaequalis. The levels of genetic diversity were higher at species level (H = 0.2031) but lower at population level (H = 0.1096). The higher genetic diversity at species levels might be attributed to the accumulation of distinctive genotypes which adapted to the different habitats after Quaternary glaciations. Meanwhile, founder effects on the early stage, and subsequent bottleneck of population regeneration due to its biological characteristics, environmental features, and human activities, seemed to explain the low population levels of genetic diversity. The hierarchical AMOVA revealed high levels (42.60%) of among‐population genetic differentiation, which was in congruence with the high levels of Nei's genetic differentiation index (G_ST = 0.4629) and limited gene flow (N_m = 0.5801) among the studied populations. Mantel test showed a significant isolation‐by‐distance, indicating that geographic isolation has a significant effect on genetic structure in this species. Unweighted pair‐group method with arithmetic average clustering, PCoA, and Bayesian analyses uniformly recovered groups that matched the geographical distribution of this species. In particular, our results suggest that Yangtze River has served as a natural barrier to gene flow between populations occurred on both riversides. Concerning the management of P. subaequalis, the high genetic differentiation among populations indicates that preserving all five natural populations in situ and collecting enough individuals from these populations for ex situ conservation are necessary.     

Genetic diversity and structure of the endangered lady's slipper orchid Cypripedium japonicum Thunb. (Orchidaceae) in Japan

Cypripedium japonicum Thunb. (Orchidaceae), once a common perennial herb, is now designated as endangered throughout most of its distribution due to habitat destruction and fragmentation, and the impacts of horticultural collection. We investigated the genetic characteristics of this species for conservation purposes, using microsatellite markers to examine the genetic diversity and structure of 15 native and 5 ex situ populations in Japan. The results imply that although allelic variation is low in Japanese C. japonicum, sexual reproduction by seed, as well as clonal propagation, may occur in some populations. Both native and ex situ populations were found to be genetically differentiated, indicating that some populations may have experienced recent population declines, genetic fragmentation, or bottlenecks. The degree of genetic drift from the putative ancestral population, inferred through STRUCTURE analysis, was more pronounced in northern populations than in southern populations. Some of the ex situ conserved populations exhibited a low degree of differentiation from ancestral native populations. Our results imply that conservation of C. japonicum in Japan is best supported by maintaining individual populations and their unique genetic characteristics.     

Model‐based conservation planning of the genetic diversity of Phellodendron amurense Rupr due to climate change

Climate change affects both habitat suitability and the genetic diversity of wild plants. Therefore, predicting and establishing the most effective and coherent conservation areas is essential for the conservation of genetic diversity in response to climate change. This is because genetic variance is a product not only of habitat suitability in conservation areas but also of efficient protection and management. Phellodendron amurense Rupr. is a tree species (family Rutaceae) that is endangered due to excessive and illegal harvesting for use in Chinese medicine. Here, we test a general computational method for the prediction of priority conservation areas (PCAs) by measuring the genetic diversity of P. amurense across the entirety of northeast China using a single strand repeat analysis of twenty microsatellite markers. Using computational modeling, we evaluated the geographical distribution of the species, both now and in different future climate change scenarios. Different populations were analyzed according to genetic diversity, and PCAs were identified using a spatial conservation prioritization framework. These conservation areas were optimized to account for the geographical distribution of P. amurense both now and in the future, to effectively promote gene flow, and to have a long period of validity. In situ and ex situ conservation, strategies for vulnerable populations were proposed. Three populations with low genetic diversity are predicted to be negatively affected by climate change, making conservation of genetic diversity challenging due to decreasing habitat suitability. Habitat suitability was important for the assessment of genetic variability in existing nature reserves, which were found to be much smaller than the proposed PCAs. Finally, a simple set of conservation measures was established through modeling. This combined molecular and computational ecology approach provides a framework for planning the protection of species endangered by climate change.     

Genetic Diversity and Structure of Sinopodophyllum hexandrum (Royle) Ying in the Qinling Mountains,China

Sinopodophyllum hexandrum is an important medicinal plant whose genetic diversity must be conserved because it is endangered. The Qinling Mts. are a S. hexandrum distribution area that has unique environmental features that highly affect the evolution of the species. To provide the reference data for evolutionary and conservation studies, the genetic diversity and population structure of S. hexandrum in its overall natural distribution areas in the Qinling Mts. were investigated through inter-simple sequence repeats analysis of 32 natural populations. The 11 selected primers generated a total of 135 polymorphic bands. S. hexandrum genetic diversity was low within populations (average H_e = 0.0621), but higher at the species level (H_e = 0.1434). Clear structure and high genetic differentiation among populations were detected by using the unweighted pair group method for arithmetic averages, principle coordinate analysis and Bayesian clustering. The clustering approaches supported a division of the 32 populations into three major groups, for which analysis of molecular variance confirmed significant variation (63.27%) among populations. The genetic differentiation may have been attributed to the limited gene flow (N_m = 0.3587) in the species. Isolation by distance among populations was determined by comparing genetic distance versus geographic distance by using the Mantel test. Result was insignificant (r = 0.212, P = 0.287) at 0.05, showing that their spatial pattern and geographic locations are not correlated. Given the low within-population genetic diversity, high differentiation among populations and the increasing anthropogenic pressure on the species, in situ conservation measures were recommended to preserve S. hexandrum in Qinling Mts., and other populations must be sampled to retain as much genetic diversity of the species to achieve ex situ preservation as a supplement to in situ conservation.     

Low genetic diversity in small peripheral populations of a rare European tree <Emphasis Type="Italic">(Sorbus torminalis)</Emphasis> dominated by clonal reproduction

Analysis of nuclear microsatellites in six small populations of Sorbus torminalis on its northern distribution limit was carried out to evaluate the dominating mode of reproduction as well as genetic diversity within and among populations. Within populations there was a low genetic diversity with heterozygote excess in five populations. Genetic differentiation was high among populations and the amount of clonally arisen recruits was 94–100%. This suggests that clonal growth, small population size and a possible self-incompability system maintain heterozygosity, although some genetic drift would also be expected. The results are discussed with respect to potential conservation strategies for S. torminalis in the study region.     

Novel Chloroplast Microsatellite (cpSSR) Markers for Genetic Diversity Assessment of Cultivated and Wild Hevea Rubber

Understanding the genetic diversity and population structure of Hevea is of great importance for managing its conservation and for utilization of rubber genetic resources. In this study, we investigated the genetic diversity and population structure of eight populations of Hevea rubber genotypes from Malaysia (MY), India (IN), Sri Lanka (LK), Indonesia (ID), France (FR), Thailand (TH), Brazil (BR), and China (CN), in addition to individual primary clones, using 10 nuclear and 11 polymorphic novel chloroplast microsatellite markers (nSSRs and cpSSRs, respectively). The BR population exhibited the greatest genetic diversity (H _e ) for both nSSR (0.841) and haploid genetic diversity (cpSSR; 0.207) markers. Bayesian analysis was applied to infer genetic structure based on nSSR data using STRUCTURE software, and a neighbor-joining tree was used to construct an unrooted phylogram based on Nei’s genetic distance that clustered these Hevea genotypes into three groups. Six polymorphic cpSSR markers produced 13 alleles and eight chlorotypes. Seven chlorotypes, A, B, C, D, E, F, and H were detected among Brazilian populations from Acre (AC), Rondônia (RO), and Mato Grosso (MT) locations. The G and H chlorotypes were found in Asiatic genotypes or Wickham clones and only one genotype, 15 AC-F-7 38-62 from AC location. These results provide valuable data for in situ or ex situ conservation and utilization of germplasm collections for breeding programs.

     

Genetic diversity and structure of Cordyceps sinensis populations from extensive geographical regions in China as revealed by inter-simple sequence repeat markers

Cordyceps sinensis is one of the most valuable medicinal caterpillar fungi native to China. However, its productivity is extremely limited and the species is becoming endangered. The genetic diversity of eighteen C. sinensis populations across its major distributing regions in China was evaluated by inter-simple sequence repeat (ISSR) markers. A total of 141 markers were produced in 180 individuals from the 18 populations, of which 99.3% were polymorphic. The low average of Shannon (0.104) and Nei index (0.07) of the 18 populations indicates that there are little genetic variations within populations. For all 18 populations, estimates of total gene diversity (H_T), gene diversity within populations (H_S), coefficient of genetic differentiation (G_ST), and gene flow (Nm) were 0.170, 0.071, 0.583, and 0.357, respectively. This pattern suggests that the genetic diversity of C. sinensis is low and most of the ISSR variations are found among populations with little gene exchange. The 18 populations are divided into five groups based on the genetic distance and the grouping pattern matches with the geographic distribution along the latitudinal gradient. The five groups show obvious difference in the G_ST and Nm values. Therefore, the genetic diversification of C. sinensis populations may be determined by geographic isolation and the combined effects of life history characters and the interaction with host insect species. The information illustrated by this study is useful for selecting in situ conservation sites of C. sinensis.     

Population structure and genetic diversity of wild <Emphasis Type="Italic">Helianthus</Emphasis> species from Mozambique

The production of sunflower suffered a major decline in Mozambique after its independence in 1975. Civil war, human activities and environmental damage subjected the species to an ecological stress contributing to reduce the number and size of wild populations. As this reduction is often related to a loss of genetic variation we estimated the genetic diversity within and among populations of wild Helianthus from five districts of Mozambique using RAPD markers. The 44 accessions studied grouped into four major clusters exhibiting structured variability with regard to geographic origin. A high level of genetic diversity (He = 0.350 and I = 0.527) was retained at the population level. The genetic variation among populations was high (59.7%), which is consistent with low gene flow (Nm = 0.338). The proportion of total genetic diversity residing among these populations should be kept in mind to devise different conservation strategies in order to preserve these populations. Currently wild Helianthus genetic resources present in Maputo and Sofala are on the edge of extinction mainly due to excessive urbanization. Therefore, conservation of what remains of this plant genetic diversity is essential for sustainable utilization and can be useful for breeding programs.     

Conservation genetics of Sinai’s remnant populations of Moringa peregrina, an economically valuable medicinal plant

Moringa peregrina is an economically valuable tree of Egyptian deserts. It is used medicinally, provides a highly nutritious supplement to Bedouin diets, provides fodder for livestock, and is used for fire wood. M. peregrina seeds have been a source of high-quality oil for cosmetics and perfumes since antiquity. Due to unmanaged grazing and over-collection, M. peregrina has become one of the most endangered tree species in the Egyptian desert ecosystem. A long-term conservation program is urgently needed to maintain or increase the number and size of M. peregrina populations. Ten populations harboring a total of 130 adult M. peregrina were sampled from three disjunct Wadis in South Sinai (W. Me’ar, W. Fieran and W. Zaghra). Open-pollinated seedlings were electrophoretically analyzed to address two basic questions: (1) how is genetic diversity distributed within and among populations within these three Wadis; and (2) what is the mating system of this species. M. peregrina has a mixed mating system with a selfing rate up to 16% and has limited genetic diversity within and significant genetic differentiation among its populations, the majority of which occurs among Wadis. Direct protection is urgently needed to decrease genetic deterioration within M. peregrina populations and to improve their ability to maintain or improve their population numbers. The priority of in situ conservation should be to conserve a few large well-distributed populations representing different Wadis. Ex situ germplasm collections should be made across the species’ range to ensure a representative sample of its genetic variation. Seed orchards designed to maximize cross-fertilization among unrelated individuals should be established to generate propagules to supplement natural populations.     

Genetic structure and diversity in Ramonda myconi (Gesneriaceae): effects of historical climate change on a preglacial relict species

The importance of the Mediterranean Basin as a long-term reservoir of biological diversity has been widely recognized, although much less effort has been devoted to understanding processes that allow species to persist in this area. Ramonda myconi (Gesneriaceae) is a Tertiary relict plant species restricted to the NE Iberian Peninsula. We used RAPD and chloroplast markers to assess the patterns of genetic structure in eight mountain regions covering almost the full species range, to identify the main historical processes that have shaped its current distribution and to infer the number and location of putative glacial refugia. While no cpDNA variation was detected, the species had relatively high levels of RAPD variation. Maximum levels of diversity were found within populations (71%), but there was also a significant differentiation between geographical regions (20%) and among populations within regions (9%). A spatial AMOVA identified three main groups of populations, corresponding to previously recognized centers of endemism and species richness. In addition, we found a marked geographical pattern of decreasing genetic diversity and increasing population differentiation from west to east. Our results support a complex phylogeographic scenario in the Iberian Peninsula of "refugia-within-refugia" and suggest that the higher diversity observed in western regions might be associated with prolonged and more stable climatic conditions in this area during the Quaternary.     

Genetic diversity in populations of Sphagnum capillifolium from the mountains of Bulgaria,and their possible refugial role

Abstract

Genetic diversity in eight populations of Sphagnum capillifolium from different Bulgarian mountains was investigated by means of isozyme electrophoresis. High levels of allelic diversity were found (H_S = 0.119), comparable to earlier estimates for northern European populations (H_S = 0.116). Strong differentiation among populations and a low number of widespread genotypes suggest a high degree of isolation and restricted gene flow between populations, which is consistent with generally small and scattered populations. The large proportion of distinguishable genotypes (mean 0.498) suggests high levels of out-crossing either currently or in the past. Introgression between S. capillifolium and S. rubellum, a species not found in Bulgaria, was suggested by the occurrence of rubellum-alleles in five populations from different mountains. This could be explained by an ancient hybridization event in a sympatric population. Based on (1) the high genetic diversity, (2) the fairly wide distribution of alien alleles, and (3) the isolated distribution of populations even within one mountain, a possible survival of S. capillifolium in the Balkan area during the Quaternary ice periods is hypothesized.     

High levels of genetic structuring as a result of population fragmentation in the tropical tree species <Emphasis Type="Italic">Caesalpinia echinata</Emphasis> Lam.

We have investigated levels of genetic diversity within and among seven remnant populations of Caesalpinia echinata Lam., an endangered species found as fragmented populations in three major areas around the coastal regions of Brazil. Using amplified fragment length polymorphism (AFLP) genetic markers, we detected levels of within-population genetic diversity ranging from 0.092 to 0.163, with the lowest values generally being found in the smallest populations. Estimates of between-population genetic differentiation were strongly correlated with geographical distance (r = 0.884, p < 0.001), which,along with a neighbour-joining phylogenetic analysis, strongly suggested high levels of genetic isolation by distance. Over half (62%) of the total genetic diversity was partitioned between populations, further highlighting the genetic distinctness of individual populations. Taken together, these results suggest that fragmentation has led to an increase in population differentiation between fragments of C. echinata. These formations will be of great value in the development of conservation plans for species exhibiting high levels of genetic differentiation due to fragmentation, such as indication of conservation unit size, which populations should be chosen as priority in conservation plans and which samples should be introduced in areas with a low number of individuals of brazilwood.     

Genetic structure of silver pomfret (Pampus argenteus (Euphrasen, 1788)) in the Arabian Sea,Bay of Bengal,and South China Sea as indicated by mitochondrial COI gene sequences

The silver pomfret (Pampus argenteus) is a commercially important species native to the Indo‐western Pacific area. In the present study, the genetic structure of five P. argenteus populations from the Arabian Sea, Bay of Bengal, and South China Sea were investigated using sequences of the mitochondrial cytochrome c oxidase subunit I (COI) gene. All populations showed high levels of haplotype diversity (h) and low levels of nucleotide diversity (π). Phylogenetic trees indicated that the haplotypes could be divided into three clusters, indicating the geographic distribution of the five populations. Analysis of molecular variance (amova ) indicated the intra‐region variation among these populations to be highly significant. The average pairwise differences and F_st values among the three regions were also found to be significant. These results suggest that P. argenteus in these regions has strong geographic genetic structure. The present results provide new information for the genetic assessment, fishery management, and conservation of this species.     

Human-mediated processes affecting distribution and genetic structure of Squalidus multimaculatus,a freshwater cyprinid with small spatial range

Endemic species typically have a narrow niche breadth, and are likely more vulnerable to extinction than more widespread taxa. Squalidus multimaculatus is a small cyprinid endemic to the Korean Peninsula, and its reported geographical range was restricted to several small rivers located along the southeast coast. Several populations of S. multimaculatus have supposedly been subject to a variety of recent anthropogenic actions. Here, we analyzed the pattern of genetic diversity within and among populations of S. multimaculatus using nine microsatellite loci to quantify the relative contributions of human-mediated processes to the contemporary distribution and genetic structure. Overall, low levels of genetic diversity were exhibited in the populations of S. multimaculatus. Genetic differentiations among populations were not completely represented by their geographical proximity, likely resulting from the low intrapopulation genetic variability and anthropogenic transplants. The most conspicuous outcome of the anthropogenic activities was the introgression of alleles from a closely related species, S. gracilis majimae. Our study showed that anthropogenic transplanting, even with only a small number of individuals, can challenge our conservation goal to maintain the species integrity that has long been shaped in evolutionary processes.     

Taxonomic Status,Phylogenetic Affinities and Genetic Diversity of a Presumed Extinct Genus,Paraisometrum W.T. Wang (Gesneriaceae) from the Karst Regions of Southwest China

Background

The karst regions in South China have an abundance of endemic plants that face high extinction risks. The Chinese Gesneriaceae endemic Paraisometrum mileense ( = Oreocharis mileensis), was presumed extinct for 100 years. After its re-discovery, the species has become one of five key plants selected by the Chinese forestry government to establish a new conservation category for plants with extremely small populations. For conservation purposes, we studied the phylogenetic and population genetic status of P. mileense at the three only known localities in Guangxi, Guizhou and Yunnan.

Methodology/Principal Findings

We collected 64 samples (52 species) of Oreocharis and 8 samples from three provinces of P. mileense and generated molecular phylogenies, and inferred that P. mileense represents a relatively isolated and derived taxonomic unit within Oreocharis. Phylogeographic results of 104 samples of 12 populations of P. mileense indicated that the populations in Yunnan have derived from those in Guangxi and Guizhou. Based on AFLP data, the populations were found to harbor low levels of genetic diversity (He = 0.118), with no apparent gradient across the species’ range, a restricted gene flow and significant isolation-by-distance with limited genetic differentiation among the populations across the three provinces (F _ST = 0.207, P<0.001). The 10 populations in Yunnan were found to represent two distinct lineages residing at different altitudes and distances from villages.

Conclusion/Significance

The low levels of genetic diversity found in P. mileense are perhaps a consequence of severe bottlenecks in the recent past. The distribution of the genetic diversity suggests that all populations are significant for conservation. Current in situ and ex situ measures are discussed. Further conservation actions are apparently needed to fully safeguard this conservation flagship species. Our work provides a model of an integrated study for the numerous endemic species in the karst regions with extremely small populations.