Mating system and genetic diversity of a rare desert legume Ammopiptanthus nanus （Leguminosae）

Ammopiptanthus nanus is an endangered evergreen shrub endemic to the deserts of central Asia and plays an important role in delaying further desertification. We examined allozyme variation and AFLP diversity in A. nanus populations and investigated the mating system of this species using progeny arrays assayed for polymorphic allozyme loci. Mating system analysis in the Keyi＇eryongke＇er population showed low levels of outcrossing, and strong inbreeding depression. Low levels of genetic variation were detected at both population （allozyme, Pp=14.0%, A=l.14, He=0.031; AFLP, Pp=14.5%, Shannon＇s information index I=0.063） and species （allozyme, Pp=21.1%, A=1.21, He=0.040; AFLP, Pp=20.9%, I=0.083） levels; while moderate genetic differentiation existed among populations, as indicated by allozymes （Gsa-=0.081） and AFLP （GST=0.151-0.193）. Founder effect, bottlenecks in evolutionary history, the mixed mating system and co-ancestry may have influenced the level of genetic diversity in A. nanus. Markers of both types provide new insights for conservation management, indicating that the Biao＇ertuokuoyi and Keyi＇eryongke＇er populations should be given priority for in situ conservation and regarded as seed sources for ex situ conservation.     

Genetic diversity and geographic differentiation in endangered Ammopiptanthus (Leguminosae) populations in desert regions of northwest China as revealed by ISSR analysis

• Background and Aims The desert legume genus Ammopiptanthuscomprises two currently endangered species, A. mongolicus andA. nanus. Genetic variability and genetic differentiation betweenthe two species and within each species were examined. • Methods Inter-simple sequence repeat (ISSR) marker datawere obtained and analysed with respect to genetic diversity,structure and gene flow. • Key Results Despite the morphological similarity betweenA. mongolicus and A. nanus, the two species are geneticallydistinct from each other, indicated by 63 % species-specificbands. Low genetic variability was detected for both populationlevel (Shannon indices of diversity H_pop = 0·106, percentageof polymorphic loci P = 18·55 % for A. mongolicus; H_pop= 0·070, P = 12·24 % for A. nanus) and specieslevel (H_sp = 0·1832, P = 39·39 % for A. mongolicus;H_sp = 0·1026, P = 25·89 % for A. nanus). Moderategenetic differentiation was found based on different measures(AMOVA _ST and Hickory ^B) in both A. mongolicus (0·3743–0·3744)and A. nanus (0·2162–0·2369). • Conclusions The significant genetic difference betweenthe two species might be due to a possible vicariant evolutionaryevent from a single common ancestor through the fragmentationof their common ancestor's range. Conservation strategies forthese two endangered species are proposed.     

Genetic diversity in Cytisus aeolicus Guss. (Leguminosae), a rare endemite of the Italian flora

ABSTRACT

Cytisus aeolicus Guss, is an endemic plant restricted to the isles of Vulcano, Stromboli and Alicudi in the Aeolian archipelago. All known populations were assayed for genetic variability using enzyme polymorphism. Allozyme variation at 16 loci coding for 10 enzyme systems was examined. The great majority of loci turned out to be monomorphic or fixed heterozygous. The observed genetic depauperation is indicative of historical factors, such as the bottleneck effect associated with migration, and the founder effect in population re-establishment. The low genetic diversity is largely partitioned within rather than among populations, indicating that extant populations have not been established long enough for divergence to have occurred. The genetic uniformity, combined with the scattered distribution of small populations, and the negative shift of population size point to a critically endangered species. Strategies for conservation are considered.     

新疆沙冬青抗冻蛋白的提取分离及其热滞活性测定

抗冻蛋白被发现于低温环境下生存的生物体中,它能够吸附在冰晶表面并改变冰晶的生长形态,对于增加植物的抗低温胁迫能力有重要作用。用纤维素DE-52离子柱层析提取分离出了冬季新疆沙冬青(Ammopiptanthus nanus)叶片中的抗冻蛋白(AFPs)。差示扫描量热法(DSC)测定结果表明,当蛋白浓度为20mg/ml时,抗冻蛋白的热滞活性(THA)为0·46℃。经SDS-PAGE电泳分析,此抗冻蛋白的分子量为119·24kDa。     

Genetic diversity of the endangered Chinese endemic herb Primulina tabacum (Gesneriaceae) revealed by amplified fragment length polymorphism (AFLP)

Primulina tabacum Hance, is a critically endangered perennial endemic to limestone area in South China. Genetic variability within and among four extant populations of this species was assessed using AFLP markers. We expected a low genetic diversity level of this narrowly distributed species, but our results revealed that a high level of genetic diversity remains, both at population level (55.5% of markers polymorphic, H _E = 0.220, I _S = 0.321), and at species level (P = 85.6% of markers polymorphic, H _E = 0.339, I _S = 0.495), probably resulting from its refugial history and/or breeding system. High levels of genetic differentiation among populations was apparent based on Nei’s genetic diversity analysis (G _st=0.350). The restricted gene flow between populations is a potential reason for the high genetic differentiation. The population genetic diversity of P. tabacum revealed here has clear implications for conservation and management. To maintain present levels of genetic diversity, in situ conservation of all populations is necessary.     

Comparison of genetic diversity between narrowly endemic shrub Menziesia goyozanensis and its widespread congener M. pentandra (Ericaceae)

Menziesia goyozanensis, a Japanese kindof fool's huckleberry, is a shrub speciesendemic only to Mt. Goyo in northern Japan.This species is listed as `criticallyendangered' in the Red Data Book of wild plantsin Japan, and only a single population has yetbeen discovered. The allozyme diversity andRAPD variation of Menziesia pentandrawere eight- and threefold greater than those ofM. goyozanensis, respectively, althoughwe examined only five populations representinga part of the distribution of M.pentandra. Because the environment surroundingthe population of M. goyoznaensis seemsto be stable at present, it is not necessary toplan an urgent ecological conservation practicefor the species. However, considering that thehabitat of M. goyozanensis is very smalland large vegetative disturbances occasionallyoccur in cool temperate forests, in which thespecies dwells, ex situ conservation isessential.     

Patterns of allozyme variation in relation to population size of the threatened plantMegaleranthis saniculifolia (Ranunculaceae) in Korea

Patterns of variation at 27 allozyme loci were investigated in the endangered endemic plantMegaleranthis saniculifolia. Levels of allozyme variation (A = 1.47,P = 40%,He = 0.088) were also compared with other endemic plant species. Genetic divergence between populations was very high (G ^st = 0.271 ), with moderate to high interpopulation differentiation, which probably arose through historical bottlenecks in a landscape of habitat fragmentation and/or human influence. The percentage of polymorphic loci, heterozygosity, and mean number of alleles per locus were positively related to population size, probably due to the stochastic loss of rare alleles in the smaller populations. Individuals in the small and marginal populations (TB, KD, and CJ) showed higher proportions of fixed loci. These ecologically marginal populations were typically more distant from the nearest neighboring population and were more genetically distinct from one another. The genetic structure of the current population ofM. saniculifolia is probably the result of local extinctions of intervening populations. This, in turn, is due to the Pleistocene climatic change and increased habitat destruction. A positive association appears to exist between genetic diversity and population size. Although these small population sizes are more sensitive to stochastic events, securing a certain number of individuals from the three larger populations (SB, JB, and TG) could be accomplished as part of a conservation strategy. In addition, it is important to prioritize populations in different regions in order to limit population declines caused by large-scale environmental catastrophes.     

Conservation of rare species with island-like distributions: A case study of Lasthenia conjugens (Asteraceae) using population genetic structure and the distribution of rare markers

Californian vernal pools, a patchy, island-like habitat, are endangered as a result of habitat destruction. Conservation of the remaining vernal pool habitat is essential for the persistence of several endangered species. We present the first study examining DNA-level genetic diversity within and among populations of a vernal pool plant species. We investigated genetic variation across eight populations of the US federally endangered vernal pool endemic Lasthenia conjugens (Asteraceae) using intersimple sequence repeat (ISSR) markers. Genetic diversity within the species was high (Nei's gene diversity estimate was 0.37), with moderate differentiation among populations (Bayesian F _ST analog of 0.124). Using an amova analysis, we found that the majority of the genetic variation (84%) was distributed within populations. There is a significant relationship between geographical distance and pairwise genetic differentiation as measured by the Bayesian estimate θ^B. The alternative hypotheses of historic geological processes within the Central Valley and contemporary gene flow are discussed as explanations of the data. Because of the vulnerability of the populations, we calculated a probability of loss for rare alleles (fragments) in the populations. Calculations show that sampling only one of the eight populations for ex-situ conservation or restoration will capture approximately 54% of the sampled rare fragments. We believe that one of the sampled populations has become extinct since it was sampled. When removing this population from the above-mentioned calculations, sampling one population will capture only 41.3% of the sampled rare fragments. We recommend sampling strategies for future conservation and restoration efforts of L. conjugens.     

Population genetic analysis of Elliottiaracemosa (Ericaceae), a rare Georgia shrub

Genetic and genotypic diversity found within populations of threatened plant species can have important implications for their conservation and management. In this study we describe genetic and genotypic diversity found within 10 populations of the endemic shrub Elliottiaracemosa (Ericaceae), the Georgia plume. E. racemosa is a threatened species known from fewer than 50 locations, all within the state of Georgia, USA. Seedset is limited to nonexistent in some E. racemosa populations and sexual recruitment has not been documented. However, the species is known to spread vegetatively via root-sprouts. Twenty-one allozyme loci were resolved for E. racemosa, nine of which were polymorphic. Compared with other woody taxa, E. racemosa has low genetic (i.e. allelic) diversity within populations (H_ep = 0.063) and at the species level (H_es = 0.091). Most of the genetic variation (82%) was found within populations, and genetic identities between populations were high (mean I = 0.96). However, genotypic diversity (i.e. the number of multilocus genotypes) differed markedly among populations. Two of the 10 populations consisted almost entirely of single multilocus genotypes, whereas more than 20 multilocus genotypes (in samples of 48 stems) were detected at three sites. Sites in which few multilocus genotypes were detected have low seedset, suggesting that the lack of clonal diversity limits reproduction in some populations of this reportedly self-incompatible species.     

Reproductive biology and population genetic structure of Kandelia candel (Rhizophoraceae), a viviparous mangrove species

The pollination biology, mating system, and population genetic structure of Kandelia candel were investigated. Field observations on its pollination and reproductive biology suggested that this species is pollinator dependent for fruit set, and bee activities can lead to substantial geitonogamous selfing. Quantitative analysis of the mating system parameters was performed using progeny arrays assayed for allozyme markers. Multilocus outcrossing rates (t(m)) were estimated to be 0.697 ± 0.091 and 0.797 ± 0.062 in two populations. In comparison to other plant species with mixed-mating system, the level of allozyme variation was very low in the 13 populations sampled along the coastlines of Hong Kong. At the species level, the proportion of polymorphic loci was 20%, number of alleles per locus was 1.2, and heterozygosity was 0.0362. The total gene diversity was primarily distributed within populations (H(S )= 0.0339), and the coefficient of genetic differentiation among populations was low (G(ST )= 0.064). This pattern of population genetic structure suggests that gene flow, primarily in the form of water-dispersed seedlings in viviparous mangrove species, is not as limited as previously thought. However, microgeographic pattern in allele frequency at the marker loci could still be detected between the western and eastern coastal populations.     

扩增片段长度多态性实验技术及在动物遗传多样性研究中的应用

扩增片段长度多态性(AFLP)是一种有效的分子遗传标记方法,具有经济、简便、模板需要量少、重复性高、结果可靠等优点。目前AFLP在动物方面的应用还不是很多,处于初级阶段,主要用于鉴定分类关系、种群遗传多样性分析、遗传连锁图谱构建等方面。     

Allozyme and RAPD analysis of the genetic diversity and geographic variation in wild populations of the American chestnut (Fagaceae)

Genetic variation among 12 populations of the American chestnut (Castanea dentata) was investigated. Population genetic parameters estimated from allozyme variation suggest that C. dentata at both the population and species level has narrow genetic diversity as compared to other species in the genus. Average expected heterozygosity was relatively low for the population collected in the Black Rock Mountain State Park, Georgia (He = 0.096 +/- 0.035), and high for the population in east central Alabama (He = 0.196 +/- 0.048). Partitioning of the genetic diversity based on 18 isozyme loci showed that ~10% of the allozyme diversity resided among populations. Cluster analysis using unweighted pair-group method using arithmetric averages of Rogers' genetic distance and principal components analysis based on allele frequencies of both isozyme and RAPD loci revealed four groups: the southernmost population, south-central Appalachian populations, north-central Appalachian populations, and northern Appalachian populations. Based on results presented in this study, a conservation strategy and several recommendations related to the backcross breeding aimed at restoring C. dentata are discussed.     

A population genetic study of the endangered plant species Limonium dufourii (Plumbaginaceae) based on amplified fragment length polymorphism (AFLP)

Limonium dufourii ( Plumbaginaceae ) is a triploid species with obligate apomictic reproduction and is endemic to the East Mediterranean coast of Spain, where it is present in only six populations, most of which have a very low number of individuals. Genetic variation and population structure in this species was studied using amplified fragment length polymorphisms (AFLPs) as markers, using the same individuals as in a previous study with random amplified polymorphic DNA (RAPD). Three different primers provided 252 bands of which 51 were polymorphic among the 152 individuals analysed. Those polymorphic bands were able to define 65 different phenotypes, of which all but two were present in only one population. The comparative analyses of data from AFLPs with those from RAPDs show a high degree of concordance. Additionally, and given the nature of these markers, we propose the estimation of nucleotide divergences from AFLP patterns. Relationships among the different AFLP patterns and the estimates of population genetic parameters obtained with this evolutionary distance are in good agreement with previous results. These analyses show that substantial genetic variability and differentiation exist within and among populations of L. dufourii . Their higher reproducibility and the possibility of obtaining estimates of nucleotide divergence make AFLPs a much better DNA fingerprinting technique.     

Genetic diversity and pedigree for Sophora toromiro (Leguminosae): a tree extinct in the wild

Sophora toromiro was endemic to the Pacific island of Rapa Nui (Easter Island) and is extinct in the wild. The species has survived in private and botanic gardens. The species is the subject of an international study to support its conservation ex situ and eventual re-introduction. As a contribution to this study the genetic diversity of the surviving stocks and herbarium samples has been assessed by means of random amplified polymorphic DNA (RAPD) and inter-simple sequence repeat (inter-SSR) analysis. The analysis indicated low levels of genetic diversity suggesting a very small number of founders. In addition, misidentified trees were identified allowing their removal from the conservation project.     

Conservation of genetic diversity in the endangered plant Eriogonum ovalifolium var. vineum (Polygonaceae)

The purpose of his research was to describethe organization of genetic variation in thefederally endangered plant taxon Eriogonumovalifolium var. vineum using allozymes. Such information can help prioritize sites andmanagement choices for capturing andmaintaining genetic variation and can reducethe number of populations necessary to committo conservation, thus reducing costs andconflicts with competing land uses. Information on genetic diversity patterns alsoprovides insight into evolutionary anddemographic history of a taxon which canprovide means to assess future risk of erosionof diversity. Similar to other Eriogonumovalifolium varieties, E. ovalifoliumvar. vineum is diverse atboth the taxon (proportion of polymorphic loci[P] = 0.55 and alleles per locus [A] =5.45 [SD = 2.5]) and population (P = 0.56 [SD =0.11] and A = 2.68 [SD = 0.35]) levels. Gene diversity (H _e) withinpopulations averaged 0.19 (SD = 0.03). Wefound some evidence for limited clonalreproduction within populations. Populationswere moderately differentiated from one another(_S = 0.14) and showed moderatedeviations from Hardy-Weinberg equilibriumwithin populations (f = 0.14 and F =0.19). Mean Nei's genetic distance (D)among all pairs of populations was 0.02. Populations with high levels of multiplemeasures of genetic diversity, high levels ofdifferentiation, and low levels of apparentinbreeding are suggested as conservationpriorities.     

Historical and biological determinants of genetic diversity in the highly endemic triploid sea lavender Limonium dufourii (Plumbaginaceae)

Microsatellite markers were used to evaluate the genetic diversity and population genetic structure in the critically endangered Limonium dufourii (Plumbaginaceae), a highly endemic triploid species from the coasts of eastern Spain. Sixty-five alleles from 13 microsatellite regions were amplified in a sample of 122 individuals collected from the six extant populations. Microsatellite patterns were consistent with the triploid nature of L. dufourii. Alleles were unambiguously assigned to two different parental subgenomes in this hybrid species and the greater contribution of the diploid parental subgenome was confirmed. Eleven, 25 and 26 multilocus genotypes were recorded from the haploid, diploid and from the combined information of both subgenomes, respectively. Genetic diversity was mostly distributed among populations (72.06% of the total genetic variation). Genotypes from Marjal del Moro populations grouped into two highly structured clusters (88.41% of the total variance). The observed patterns of distribution of genetic diversity are interpreted to result from multiple hybridization events and isolation between populations. Threats to this species are mainly anthropogenic (urbanization and tourism pressure), although stochastic risks cannot be ignored. Therefore, in order to preserve extant genetic variation of L. dufourii, in situ strategies such as the preservation of its habitat are a high priority. Several recommendations in order to assist ex situ measures to guarantee the success of conservation strategies and maintain the relationships between individuals and populations are proposed.     

Mating system parameters and population genetic structure in pioneer populations of Prosopis velutina (Leguminosae)

This study describes the genetic structure and provides estimates of mating system parameters in three natural populations of Prosopis velutina Woot. in southeastern Arizona. F statistics derived from isozyme data revealed the presence of both interpopulation and intrapopulation genetic differentiation. This population structure is discussed in relation to the ecological history of these populations that invaded the grasslands sites from adjacent riparian areas within the last 90 years, and possible modes of seed dispersal. The multilocus estimation program MLT of Ritland (Journal of Heredity 8: 235–237, 1990) was used to provide estimates of t_s, t_m, and the fixation index (F) of the maternal parents. Average estimates of t_s ranged from 0.591 to 0.912. Estimates of t_m ranged from 0.609 to 1.004, and averaged 0.758. The difference between t_m and t_s, which provides a measure of biparental inbreeding, averaged 0.058. This last result, together with significant heterogeneity found in pollen allele frequencies, suggests that a family structure has developed in the populations. A negative F value for one of the populations suggests selection against homozygotes between the seedling stage and maturity.     

Clonal diversity in the rare Calamagrostis porteri ssp. insperata (Poaceae): comparative results for allozymes and random amplified polymorphic DNA (RAPD) and intersimple sequence repeat (ISSR) markers

Four populations of the rare, highly clonal grass Calamagrostis porteri ssp. insperata were examined using allozymes and the two polymerase chain reaction (PCR)-based markers, random amplified polymorphic DNA (RAPD) and intersimple sequence repeat (ISSR) bands. Only one of the 15 allozyme loci was variable and two alleles were detected, both of which were found in two populations, while only one genotype was detected in the other two populations. ISSR and RAPD markers detected more genotypes within populations than did allozymes. ISSR markers detected more diversity than RAPD markers in three of the four populations examined. In one population, no RAPD diversity was found whereas eight different genotypes were found among the 10 plants with ISSR markers. This diversity is present despite rare flowering, no documented occurrence of seed set in natural populations and very low seed set with experimental pollinations, all of which suggest that sexual reproduction rarely occurs. The subspecies is self-compatible, but seed initiation is lower in selfed ovules; also, there is high embryo abortion regardless of pollen source. Variation detected by RAPD and ISSR primers may reflect higher levels of sexual reproduction in the past, very rare sexual reproduction in extant populations, somatic mutations, or a combination of the three. Although the PCR-based markers identify several multilocus genotypes within populations, it is not known whether these all represent distinct genets generated by sexual reproduction or result from somatic mutations in the old, perennial and highly clonal plants.     

Genetic diversity and hierarchical population structure of a rare autotetraploid plant,Aster kantoensis (Asteraceae)

We sampled 17 populations of a rare autotetraploid Aster kantoensis (Asteraceae) from three river systems located in central Japan, and studied them for allelic variation at 22 enzyme loci. There was no significant correlation between the actual population size and three genetic diversity parameters, suggesting that the effective population size was very small even for the large populations, i.e., even large populations may still have a high probability of being of recent origin and remain influenced by the founder effect. Compared to other autotetraploid species, the total genetic variation of A. kantoensis is small. The number of alleles and gene diversity of a population were not significantly different among the river systems, although the percentage of polymorphic loci was different. Genetic differentiation among river systems was larger than between populations within the river systems, thereby indicating that gene flow between river systems is small, especially between the Kinu River system and Tama or Sagami River systems.     

High genetic diversity in a rare and endangered sunflower as compared to a common congener

Determining the genetic structure of isolated or fragmented species is of critical importance when planning a suitable conservation strategy. In this study, we use nuclear and chloroplast SSRs (simple sequence repeats) to investigate the population genetics of an extremely rare sunflower, Helianthus verticillatus Small, which is known from only three locations in North America. We investigated levels of genetic diversity and population structure compared to a more common congener, Helianthus angustifolius L., using both nuclear and chloroplast SSRs. We also investigated its proposed hybrid origin from Helianthus grosseserratus Martens and H. angustifolius. Twenty-two nuclear SSRs originating from the cultivated sunflower (Helianthus annuus L.) expressed sequence tag (EST) database, and known to be transferable to H. verticillatus and its putative parental taxa, were used in this study thereby allowing for statistical control of locus-specific effects in population genetic analyses. Despite its rarity, H. verticillatus possessed significantly higher levels of genetic diversity than H. angustifolius at nuclear loci and equivalent levels of chloroplast diversity. Significant levels of population subdivision were observed in H. verticillatus but of a magnitude comparable to that of H. angustifolius. Inspection of multilocus genotypes also revealed that clonal spread is highly localized. Finally, we conclude that H. verticillatus is not of hybrid origin as it does not exhibit a mixture of parental alleles at nuclear loci, and it does not share a chloroplast DNA haplotype with either of its putative parents.