Genetic Variation in the Endangered Endemic Species Cycas fairylakea (Cycadaceae) in China and Implications for Conservation

Cycas fairylakea is an endangered endemic species in China. Genetic diversity within and among four natural populations of this species in China was investigated using amplified fragments length polymorphism (AFLP). A moderate to low level of intraspecific genetic diversity was detected in this species (at population level: P = 39.57 %, H₀ = 0.244; at species level: P = 60.22%, H₀ = 0.356). The among-population component accounted for, respectively, 25.7 and 31.5% of the genetic variation, according to AMOVA and Shannon’s index, indicating most of the genetic variation was found between individuals within populations. All four populations have opposite pyramid age structure, and few coning individuals, which is still decreasing. Possibly because of habitat degradation and environmental pollution, plant diseases and insect pests in the populations were extremely serious, suggesting that the main factors threatening the survival of C. fairylakea populations were not genetic variation, but human activities and the breeding system of this species.     

Patterns of allozyme diversity in several selected rare species in Korea and implications for conservation

Because some endemic plants appear to be adapted to a narrow setof environmental conditions with limited genetic diversity, an analysis ofpopulation genetic structure is necessary to fully evaluate the impact of rarityon genetic variation. Listed as endangered species in Korea, only fewpopulations of Abeliophyllum distichum, Leonticemicrorhyncha, Bupleurum euphorbioides, andBerchemia berchmiaefolia were found. A reduced level ofgenetic variation in B. berchemiaefolia is consistent withthe occurrence of a genetic bottleneck and inbreeding. Leonticemicrorhyncha differed dramatically from other taxa in its observed level of geneticvariation, probably due to its predominant selfing. The level ofallozyme variation maintained by A. distichum was high forendemic species. Compared to species with similar traits, A.distichum maintained a relatively higher genetic diversity, probably dueto floral heteromorphism and preferred outcrossing. Bupleurumeuphorbioides maintained a higher genetic diversity due to outcrossing,but at the individual locus, deficiency of heterozygosity prevailed. Probablyinbreeding between local neighborhoods was frequent because A.distichum and B. euphorbioides were pollinatedby small fly species which might be less effective as a pollen dispersal, andtheir visits were extremely scarce and controlled by the weather conditions.Since much of the species-to-species variation in genetic diversity is due tothe specific ecological and evolutionary history of a species, any managementplan developed should be based on historical changes in the population size anddistribution to better predict the amounts and patterns of genetic diversity.     

Comparisons of within-population genetic variation in sexual and agamospermousAmelanchier (Rosaceae) using RAPD markers

We compared genetic variation of sexualAmelanchier bartramiana and facultatively agamospermous (asexually seed-producing)A. laevis at one site where the two species are sympatric. We analyzed 77 random amplified polymorphic DNA (RAPD) markers in 29A. bartramiana individuals and 76 RAPD markers in 31A. laevis individuals. The two species do not differ significantly in mean genetic variation. However, 22.4% of genetic similarity values betweenA. laevis individuals exceed the highest value ofA. bartramiana and may represent the effect of agamospermy. Variation withinA. laevis may be the result of sexuality, hybridization, polyploidy, and other factors.     

Genetic differentiation among morphological variants of Acacia saligna (Mimosaceae)

This study investigated the pattern of genetic variation in Acacia saligna (Labill.) H.L. Wendl. to facilitate its development as a crop species for dryland salinity management. A. saligna is a morphologically variable species and four main variants are recognized. The genetic structure within A. saligna was investigated in populations across the geographic range of the species using nuclear restriction fragment length polymorphism loci. The analysis identified considerable genetic variation within A. saligna that was genetically structured into three groups. Two of the three groups corresponded to variants recognized in the field study; the third group encompassed the other two variants which, though morphologically different, were not genetically differentiated. The level of genetic differentiation between the groups suggests they may represent different taxa and a taxonomic revision of the species may be required. Identification of different taxa within A. saligna will have implications for the utilization and domestication of the species, as the taxa will need to be evaluated separately to determine their suitability for agroforestry. The high genetic variation between and within groups suggests there is a large genetic base available for breeding improved cultivars.     

RAPD Profiling in Detecting Genetic Variation in Endemic Coelonema (Brassicaceae) of Qinghai-Tibet Plateau of China

Random amplified polymorphic DNA (RAPD) markers were used to measure genetic diversity of Coelonema draboides (Brassicaceae), a genus endemic to the Qilian Mountains of the Qinghai-Tibet Plateau. We sampled 90 individuals in 30 populations of Coelonema draboides from Datong and Huzhu counties of Qinghai Province in P.R. China. A total of 186 amplified bands were scored from the 14 RAPD primers, with a mean of 13.3 amplified bands per primer, and 87% (161 bands) polymorphic bands (PPB) was found. Analysis of molecular variance (AMOVA) shows that a large proportion of genetic variation (84.2%) resides among individuals within populations, while only 15.8% resides among populations. The species shows higher genetic diversity between individuals than other endemic and endangered plants. The RAPDs provide a useful tool for assessing genetic diversity of rare, endemic species and for resolving relationships among populations. The results show that the genetic diversity of this species is high, possibly allowing it to adapt more easily to environmental variations. The main factor responsible for the high level of differentiation within populations and the low level of diversity among populations is probably the outcrossing and long-lived nature of this species. Some long-distance dispersal, even among far separated populations, is also a crucial determinant for the pattern of genetic variation in the species. This distributive pattern of genetic variation of C. draboides populations provides important baseline data for conservation and collection strategies for the species. It is suggested that only populations in different habitats should be studied and protected, not all populations, so as to retain as much genetic diversity as possible.     

Genetic differentiation of theErigeron species (Asteraceae) in the Alps: A case of unusual allozymic uniformity

Allozyme variation was studied in all nine diploidErigeron species known from the Alps:E. alpinus, E. neglectus, E. polymorphus, E. candidus, E. uniflorus, E. atticus, E. gaudinii, E. acer, andE. angulosus. A total of 248 individuals from 24 natural populations was investigated using starch gel electrophoresis. Seven enzymes and 13 loci were assessed. Genetic variation within populations was low with the proportion of polymorphic loci ranging from 0.0–0.385, and average number of alleles per polymorphic locus from 2.0–2.5. In general, 70–100% of the genetic variation was attributed to between population differences. Mean genetic identities for pair-wise comparisons of populations averaged 0.893 within species, and 0.890 among species. Interspecific genetic variation of populations usually did not exceed intraspecific variation. It was concluded that theErigeron species from the Alps may have arisen by recent speciation probably during the epoches of glaciation. Morphological and ecological differences between species seem to be based on few gene loci.     

中国东部沿海水仙归化群体的遗传多样性

为揭示我国东部归化水仙(Narcissus tazetta var. chinensis)的群体遗传多样性,利用2个叶绿体基因mat K和trn H-psb A片段对采自沪、浙、闽的5个代表群体的49株水仙进行了评估。结果表明,双基因联合序列的总长为1443bp,共定义6个单倍型,各归化群体的遗传多样性水平为DLSYPTDNJD=ZZCMD。AMOVA分析表明,群体内变异为遗传变异的主要来源(91.98%),群体间的遗传分化较低(Fst=0.080 22)。群体物种水平上的谱系结构不显著(Nst=0.020Gst=0.031;P0.05)。Mantel检验表明水仙群体间的遗传距离与地理距离呈显著的线性相关(r=0.929,P=0.02 0.05)。中性检验和错配分布分析结果均暗示水仙群体背离了快速扩张模型的假设。单倍型分布的中介网络图结合系统发育NJ树均将所有群体划分为2大分支。因此,我国东部沿海水仙归化群体整体遗传多样性水平较低,各群体间遗传分化较弱,遗传变异主要来自群体内,物种近期未经历扩张事件,可能是基因流受海岛隔离、自身生物学特征、生境异质性与及人为干扰的综合作用影响。     

Evolution of Dystaenia takesimana (Apiaceae), endemic to Ullung Island, Korea

Dystaenia (Apiaceae) consists of two species, one distributed in Japan (D. ibukiensis), and the other endemic to Ullung Island, Korea (D. takesimana). In comparison with representative outgroup taxa in Ligusticum, Seseli, Angelica, and Osmorhiza, Dystaenia is shown to be monophyletic based on sequences from chloroplast trnL-F intron and spacer regions confirming previously published results using ITS sequences. Loss of one large part of trn L-F in D. takesimana strongly suggests that this species evolved from D. ibukiensis rather than the reverse. AFLP analysis within and among twelve populations (six from each species; total 126 individuals) using three primer combinations reveals 130 reliable fragments. Neighbour-joining analysis shows the two species to be distinct populational systems. Levels of overall genetic variation as measured by Shannon Diversity are significantly higher in D. takesimana. Geographic structuring of genetic variation occurs within D. ibukiensis but not within D. takesimana, suggesting that the Ullung species exists as a single population. It is hypothesised that after a founder-effect reduction of genetic variation, anagenetic speciation may have occurred in D. takesimana by gradual morphological divergence accompanied by accumulation of genetic variation through mutation, recombination and drift.     

Variation of isozyme patterns among Arachis species

The genus Arachis contains a large number of species and undescribed taxa with patterns of genetic variation that are little understood. The objectives of this investigation were to estimate genetic diversity among species of Arachis by utilizing electrophoretic techniques and to establish the potential for use of isozymes as markers for germplasm introgression. One-hundred-and-thirteen accessions representing six of the seven sections of the genus were analyzed for isozyme variation of 17 enzymes. Section Rhizomatosae species were not included because they produce very few seeds. Seeds were macerated and the crude extract was used for starch-gel electrophoretic analyses. Although the cultivated species has few polymorphic isozymes, the diploid species are highly variable and two-to-six bands were observed for each isozyme among accessions. Because of the large number of isozyme differences between A. hypogaea and A. batizocoi (the presumed donor of the B genome), this species can no longer be considered as a progenitor of the cultivated peanut. Seed-to-seed polymorphisms within many accessions were also observed which indicate that germplasm should be maintained as bulk seed lots, representative of many individuals, or as lines from individual plants from original field collections. The area of greatest interspecific genetic diversity was in Mato Grosso, Brazil; however, the probability of finding unique alleles from those observed in A. hypogaea was greatest in north, north-central, south and southeast Brazil. The large number of polymorphic loci should be useful as genetic markers for interspecific hybridization studies.     

Low genetic diversity and significant population structuring in the relict Amentotaxus argotaenia complex (Taxaceae) based on ISSR fingerprinting

Amentotaxus, a genus of the Taxaceae, represents an ancient lineage that has long existed in Eurasia. All Amentotaxus species experienced frequent population expansion and contraction over periodical glaciations in Tertiary and Quaternary. Among them, Amentotaxus argotaenia complex consists of three morphologically alike species, A. argotaenia, Amentotaxus yunnanensis, and Amentotaxus formosana. This complex is distributed in the subtropical region of mainland China and Taiwan where many Pleistocene refugia have been documented. In this study, genetic diversity and population structuring within and between species were investigated based on the inter-simple sequence repeats (ISSR) fingerprinting. Mean genetic diversity within populations was estimated in three ways: (1) the percentage of polymorphic loci out of all loci (P) (2) Neis unbiased expected heterozygosity (He), and (3) Shannons index of phenotypic diversity. For a total of 310 individuals of 15 populations sampled from the three species, low levels of ISSR genetic variation within populations were detected, with P=4.66–16.58%, He=0.0176–0.0645 and Hpop=0.0263–0.0939, agreeing with their seriously threatened status. AMOVA analyses revealed that the differences between species only accounted for 27.38% of the total variation, whereas differences among populations and within populations were 57.70 and 14.92%, respectively, indicating substantial isolation between the patch-like populations. A neighbor-joining tree identified a close affinity between A. yunnanensis and A. formosana. Genetic drift due to small population size, plus limited current gene flow, resulted in significant genetic structuring. Low levels of intrapopulational genetic variation and considerable interpopulational divergence were also attributable to demographic bottlenecks during and/or after the Pleistocene glaciations.     

‘Good-genes’ and ‘compatible-genes’ effects in an Alpine whitefish and the information content of breeding tubercles over the course of the spawning season

Abies ziyuanensis is a highly endangered fir species endemic to South China. Unlike other Abies species that are distributed in areas with cold climates, A. ziyuanensis is restricted to several isolated island-like localities at subtropical mountains. In this study, we used dominant amplified fragment length polymorphism (AFLP) and co-dominant simple sequence repeats (SSR) markers to infer the genetic structure of A. ziyuanensis. Seven populations consisting of 139 individuals were sampled across their whole distribution. A. ziyuanenesis has a relatively low level of genetic variation, with a mean genetic diversity per population (He) of 0.136 (AFLP) and 0.337 (SSR), which is lower than that of other reported endemic species based on the same kind of marker. We observed high population differentiation, with Gst = 0.482 (AFLP) and Fst = 0.250 (SSR), among the seven populations. AMOVA also detected significant differentiation among populations (Φst (AFLP) = 0.550 and Φst (SSR) = 0.289) and among regions (Φct (AFLP) = 0.139 and Φct (SSR) = 0.135) in both marker types. Both ongoing evolutionary forces (e.g., genetic drift resulting from small population size) and historical events (e.g., population contraction and fragmentation during and after the Quaternary glacial cycles) may have contributed to the genetic structure in A. ziyuanensis.     

Genetic diversity and structure in the rare Colorado endemic plant <Emphasis Type="Italic">Physaria bellii</Emphasis> Mulligan (Brassicaceae)

Physaria bellii (Brassicaceae) is a rare, outcrossing perennial endemic to shale and sandstone outcrops along the Front Range of northern Colorado, USA. This species is locally abundant, but ranked G2/S2—imperiled because of threats to its habitat and a small number of populations—according to NatureServe’s standardized ranking system. Leaf tissue from ten populations was analyzed with ISSR (Inter-Simple Sequence Repeat) markers to discern the amount of genetic diversity and degree of population subdivision in P. bellii. Genetic diversity was moderate (0.22) and a moderately high degree of population structure was found (F _ST calculated using two algorithms ranged from 0.17 to 0.24). An AMOVA partitioned most of the variation among individuals within populations (76%), and the remainder among populations (24%). Results from a Principal Coordinates analysis were consistent with the geographic distribution of populations. A Mantel test of the correlation between genetic and geographic distances was highly significant (P < 0.001). The pattern of variation thus appears to be distributed along a gradient, and efforts to conserve this species should involve preserving enough populations so that gene flow between populations is not interrupted.     

Use of single-primer DNA amplifications in genetic studies of peanut (Arachis hypogaea L.)

A recent approach to detecting genetic polymorphism involves the amplification of genomic DNA using single primers of arbitrary sequence. When separated electrophoretically in agarose gels, the amplification products give banding patterns that can be scored for genetic variation. The objective of this research was to apply these techniques to cultivated peanut (Arachis hypogaea L.) and related wild species to determine whether such an approach would be feasible for the construction of a genetic linkage map in peanut or for systematic studies of the genus. Two peanut cultivars, 25 unadapted germplasm lines of A. hypogaea, the wild allotetraploid progenitor of cultivated peanut (A. monticola), A. glabrata (a tetraploid species from section Rhizomatosae), and 29 diploid wild species of Arachis were evaluated for variability using primers of arbitrary sequence to amplify segments of genomic DNA. No variation in banding pattern was observed among the cultivars and germplasm lines of A. hypogaea, whereas the wild Arachis species were uniquely identified with most primers tested. Bands were scored (+/–) in the wild species and the PAUP computer program for phylogenetic analysis and the HyperRFLP program for genetic distance analysis were used to generate dendrograms showing genetic relationships among the diploid Arachis species evaluated. The two analyses produced nearly identical dendrograms of species relationships. In addition, approximately 100 F₂ progeny from each of two interspecific crosses were evaluated for segregation of banding patterns. Although normal segregation was observed among the F₂ progeny from both crosses, banding patterns were quite complex and undesirable for use in genetic mapping. The dominant behavior of the markers prevented the differentiation of heterozygotes from homozygotes with certainty, limiting the usefulness of arbitrary primer amplification products as markers in the construction of a genetic linkage map in peanut.     

Isolated populations or isolated taxa? A case study in narrowly-distributed snapdragons (Antirrhinum sect. Sempervirentia) using RAPD markers

RAPD fingerprinting was used to study species boundaries in narrowly distributed endemic species in Antirrhinum section Sempervirentia. Based on RAPD data, similarity values within species were relatively high but pair-wise similarity values among species were low. Partitioning of the overall RAPD variation using AMOVA showed that most of the variation was found among species (58.06%), whereas the remaining phenotypic diversity was distributed among populations (25.18%) and among individuals within populations (16.76%). Comparison of the matrices of geographical distances and phenetic distances (1-Dice index) among populations using the Mantel test showed a moderate, but statistically significant correlation (r=0.588, P < 0.01), suggesting that isolation by distance is responsible for the distribution of genetic variation among Antirrhinum populations. Phenetic relationships among Antirrhinum samples based on a Dice similarity matrix showed a clear taxonomic pattern, confirming the grouping of individuals within their own populations and the clustering of populations within species. Individuals of A. charidemi, A. valentinum and A. subbaeticum, from subsection Valentina, made up a discrete group, whereas the samples belonging to subsection Sempervirentia (A. petegasii, A. sempervirens, A. microphyllum, A. pulverulentum) clustered together. RAPD data are entirely congruent with the subsection classification scheme proposed by Fernández Casas (1997) in section Sempervirentia. However, A. subbaeticum, treated as a synonym of A. valentinum by Fernández-Casas (1997), showed an unique RAPD profile characterized by the highest number of fixed species-specific markers found in section Sempervirentia. Thus, although A. valentinum appeared the most closely related species to A. subbaeticum, molecular data suggested that this species merits taxonomic distinction.     

Potential genetic consequences of a recent bottleneck in the Amur tiger of

The Amur tiger, Panthera tigris altaica, is a highly endangered felid whose range and population size has been severely reduced in recent times. At present, the wild population is estimated at 490 individuals, having rebounded from the 20–30 tigers remaining following a severe bottleneck in the 1940's. The current study presents preliminary data on the patterns and levels of genetic variation in the mitochondrial DNA control region using DNA extracted from non-invasively sampled faecal material, collected throughout the entire range of P. t. altaica in the Russian Far East. Analysis of 82 scat samples representing at least 27 individuals revealed extremely low levels of CR haplotype diversity, characterized by a single widespread haplotype (96.4%) and two rare variants, each differing by a single step within the hypervariable I (2.4%) and central conserved regions (1.2%), respectively. A comparison with previous data on cytochrome bvariation in 14 captive individuals revealed a potentially greater amount of genetic variation represented in captivity relative to that found in the wild population. The extremely low levels of mitochondrial DNA variation in the wild population is discussed in light of the demographic processes that might have shaped these patterns as well as the potential bias introduced through analysis of fecal samples. These results highlight the continuing need to assess levels of genetic variation even in recovering populations that are increasing in number and underscore the important role that captive breeding programs may play in preserving remnant genetic diversity of endangered species.     

Hybridization and genetic variation in Danish populations of European crab apple (Malus sylvestris)

The aim of the present study was to investigate the genetic variation in Danish populations of the endangered European crab apple (Malus sylvestris). Special emphasis was given to hybridization between the wild species and its cultivated relative Malus ×domestica. A total of 178 wild individuals from four Danish populations were studied along with a reference sample of 29 old cultivars. The genetic variation within and among samples was studied at ten microsatellite marker loci. Additionally, a morphological analysis was carried out to identify hybrids and test for correspondence between phenotypic and genotypic indices of hybridization. From application of ordination and a model-based cluster analysis to the molecular data, two clusters were identified consisting of wild and cultivated individuals, respectively. This indicates that pronounced admixture between the two species is not present. At the population level, a high correspondence was found between geographic isolation from M. ×domestica and genotypic and morphological indices of hybridization. As expected, isolated populations appeared less affected by hybridization than poorly isolated populations. Isolated “pure” M. sylvestris populations could thus be identified. However, morphological and molecular evidences of hybridization were found to be divergent at the individual level. This is suggestive of some historical introgression into the M. sylvestris gene pool and indicates that relying exclusively on either morphological or molecular characters as diagnostic markers in studies of hybridization between M. ×domestica and M. sylvestris might lead to fallible results. Combined application of genetic and morphological markers is therefore recommended.     

Wheat phylogeny determined by RFLP analysis of nuclear DNA. 2. Wild tetraploid wheats

Intra- and inter-specific variations in the nuclear DNA of Triticum dicoccoides Körn. (2n = 28, genome constitution AABB) and T. araraticum Jakubz. (2n = 28, AAGG), wild species, respectively, of the Emmer and Timopheevi group, were studied by restriction fragment length polymorphism (RFLP) analysis. Total DNAs of 32 T. dicoccoides and 24 T. araraticum accessions, collected from throughout the distribution areas of these species, were treated with two 6-bp cutters and hybridized with 30 nuclear DNA clones as probes to detect RFLPs. A total of 167 hybrid bands were observed per accession. All the enzyme-probe combinations showed RFLPs between accessions. The average genetic distance between the T. dicoccoides accessions was 0.0135 ± 0.0031 and that between the T. araraticum accessions 0.0036 ± 0.0015, indicative of about a four-fold intraspecific variation in T. dicoccoides as compared to T. araraticum in terms of genetic distance. No significant genetic differentiation was found for the geographical populations of these species, the genetic distance between the two species being 0.0482 ± 0.0022. The interspecific divergence corrected for intraspecific divergence was 0.0395, about three times that for T. dicoccoides and 11 times that for T. araraticum. The results show that in the wild state the Emmer and Timopheevi groups are clearly differentiated and that T. dicoccoides has much greater variation than T. araraticum, suggesting a relatively recent origin for the latter and therefore a diphyletic origin for these species.Contribution from the Laboratory of Genetics, Faculty of Agriculture, Kyoto University, Japan, No. 535     

Allozyme polymorphisms, outerossing rates, and hybridization of South AmericanNothofagus

Electrophoretically detectable genetic variation was used to describe the genetic structure of three South American species ofNothofagus: the widespreadN. betuloides andN. dombeyi, and the geographically restrictedN. nitida. Although the widespread species possess higher levels of genetic variation, the three species have more genetic variation within than among populations. These results are consistent with the theoretical expectations for woody, presumably highly outerossed species with wind-borne seeds.Estimates of outcrossing rates from progeny arrays yielded slightly higher average t-values forN. nitida (1.158) andN. dombeyi (range 0.873–1.045) than forN. betuloide (0.878). Hierarchical analysis of population structure revealed values of F_IS and FIT that were positive and significantly different from zero at most loci and for each species. The levels of inbreeding detected by F-statistics indicate some degree of self-fertilization and/or population substructuring into discrete family groups. Reduced seed vagility and regeneration of natural stands after disturbance by a few remnant individuals would probably generate the recruitment of related seedlings underneath parent trees.The analysis of a putative hybrid population betweenN. nitida andN. betuloides indicated that individuals clearly segregated intonitida-like orbetuloides- like individuals. The reduced outcrossing rate ofbetuloides-like individuals from the hybrid site (t=0.585) is interpreted in concert with low pollen availability and the increased probability of selfing and/or hybridization inNothofagus.     

Genetic spatial clustering: significant implications for conservation of wild soybean (Glycine soja: Fabaceae)

Knowledge of spatial patterns of genetic variation within populations of wild relative species has significant implications with respect to sampling strategies for ex situ and in situ conservation. To study spatial genetic structure of wild soybean (Glycine soja Sieb. et Zucc.) at the fine scale, three natural populations in northern China were analyzed using inter-simple sequence repeat (ISSR) fingerprints for estimating kinship coefficients. A regression analysis of kinship coefficients against spatial distances revealed that individuals occurring close together tended to be more genetically related. The Sp statistic further indicated a comparable spatial pattern among the three wild soybean populations with similar Sp values (mean = 0.0734, varied from 0.0645 to 0.0943) detected across the three populations. Genetic patches were on average ca. 20 m in size, and the effective neighborhood sizes varied between 10 and 15 m. The spatial genetic structure evident in the wild soybean populations may be attributed to the restricted seed dispersal and predominant inbreeding mating system of this species. The detection of family structure in the populations of wild soybean has a significant implication for the effective conservation of the important genetic resources.