Phylogeography of <Emphasis Type="Italic">Ceriops tagal</Emphasis> (Rhizophoraceae) in Southeast Asia: the land barrier of the Malay Peninsula has caused population differentiation between the Indian Ocean and South China Sea

The genetic structure of mangrove species is greatly affected by their geographic history. Nine natural populations of Ceriops tagal were collected from Borneo, the Malay Peninsula, and India for this phylogeographic study. Completely different haplotype compositions on the east versus west coasts of the Malay Peninsula were revealed using the atpB-rbcL and trnL-trnF spacers of chloroplast DNA. The average haplotype diversity (Hd) of the total population was 0.549, nucleotide diversity (θ) was 0.030, and nucleotide difference (π) was 0.0074. The cladogram constructed by the index of population differentiation (G _ST) clearly separated the South China Sea populations from the Indian Ocean populations. In the analysis of the minimum spanning network, the Indian Ocean haplotypes were all derived from South China Sea haplotypes, suggesting a dispersal route of C. tagal from Southeast Asia to South Asia. The Sunda Land river system and surface currents might be accountable for the gene flow directions in the South China Sea and Bay of Bengal, respectively. The historical geography not only affected the present genotype distribution but also the evolution of C. tagal. These processes result in the genetic differentiation and the differentiated populations that should be considered as Management Units (MUs) for conservation measurements instead of random forestation, which might lead to gene mixing and reduction of genetic variability of mangrove species. According to this phylogeographic study, populations in Borneo, and east and west Malay Peninsula that have unique genotypes should be considered as distinct MUs, and any activities resulting in gene mixing with each other ought to be prevented.     

Strong genetic differentiation of the East-Himalayan <Emphasis Type="Italic">Megacodon stylophorus</Emphasis> (Gentianaceae) detected by Inter-Simple Sequence Repeats (ISSR)

Megacodon stylophorus (Clarke) Smith is a perennial alpine herb endemic to the species-rich eastern Himalayan region. Its populations are locally scattered as isolated patches throughout this region. Genetic variation within and among six populations of this species was assessed using ISSR fingerprinting with 13 primers. High levels of genetic diversity exist within species (P = 69.83%, H_T = 0.1949 and H_sp = 0.3047), while the within-population diversity is low (P = 11.21%, H_E = 0.0532 and H_pop = 0.0792). Extraordinarily high levels of genetic differentiation were detected among populations based on various statistics, including Neis genetic diversity analysis (72.7%), Shannons diversity index (74.01%) and AMOVA (80.70%). That is, populations shared low levels of genetic identity (I = 0.8203 ± 0.0430). This genetic structure was probably due to severe genetic drift of the small-sized patchy populations resulting from postglacial habitat fragmentations. The observed genetic structure of the populations implies that as many populations as possible should be considered for any in situ and ex situ conservation practice on this species.     

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.     

A cline of allozyme variation inAbies mariesii

Genetic variation at 22 allozyme loci was examined for 1,003 trees from 11 isolated natural populations ofAbies mariesii covering all except the southernmost region of its geographic range. Genetic diversity within species (H _es=0.063) was low compared to many other long-lived woody species. Most of the genetic variation is found within populations (G _ST=0.144) despite their isolated distribution. Genetic distance between populations was positively correlated with geographic distance. Genetic diversity within populations was generally low (meanH _ep=0.054), but varied across populations in a clinal fashion such that genetic variation decreased with increasing latitude. These genetic characteristics may reflect the distribution history of this species.     

Population genetic structure of three tree species in the mangrove genus Ceriops (Rhizophoraceae) from the Indo West Pacific

Ceriops is a viviparous mangrove with widespread species Ceriops decandra and C. tagal, and an endemic species C. australis. Genetic diversity of the three species was screened in 30 populations collected from 23 locations in the Indo West Pacific (IWP) using Inter-simple sequence repeats (ISSR) and sequences of partial nuclear gene (G3pdh) and chloroplast DNA (trnV-trnM). At the species level, the total gene diversity (Ht) revealed by ISSRs was 0.270, 0.118, and 0.089 in C. decandra, C. tagal, and C. australis, respectively. A total of six haplotypes of G3pdh and five haplotypes of trnV-trnM were recognized among the three species. Only C. decandra was detected containing more than one haplotype from each sequence data set (four G3pdh haplotypes and three trnV-trnM haplotypes). At the population level, genetic diversity of Ceriops was relatively low inferred from ISSRs (He = 0.028, 0.023, and 0.053 in C. decandra, C. tagal, and C. australis, respectively). No haplotype diversity within population was detected from any of the three species. Cluster analysis based on ISSRs identified three major geographical groups in correspond to the East Indian Ocean (EIO), South China Sea (SCS), and North Australia (NA) in both C. decandra and C. tagal. The cladogram from DNA sequences also detected the same three geographical groups in C. decandra. Analysis of molecular variance (AMOVA) revealed that most of the total variation was accounted for by differentiation between the three major geographical regions of both C. decandra and C. tagal. The significant genetic structure may result from the geological events in these regions during the recent Pleistocene glaciations. This study also provided insights into the phylogenetics of Ceriops. Yelin Huang and Fengxiao Tan contributed equally to this work.     

Allozyme variation in the diploid (A genome) populations ofScilla scilloides (Hyacinthaceae)

Allozyme variation at eleven loci encoding seven enzyme systems were examined in 20 populations of diploid (genome AA, 2n = 16)Scilla scilloides in China. In comparison with the average species of seed plants studied, populations of this species display a high amount of genetic variation (A = 2.0, P = 58.6%, H_o = 0.172, and H_e = 0.185). Allozyme variation pattern revealed predominant outcrossing within populations and considerable differentiation (F_ST = 0.314) among populations as well as between the subtropic and temperate regions. The wide distribution, long existence and outcrossing are presumably the main factors responsible for the high genetic diversity within populations. But the gravity dispersal of seeds and pollination by small insects set limits to the increase of genetic variation within populations and promote differentiation between populations and regions. In addition, allozyme variation does not distinguishS. scilloides var.albo-viridis and suggests that subtropic populations may be considered as a genetic entity.     

Allozyme Diversity and Population Genetic Structure of Pinus densata Master in Northwestern Yunnan, China

We investigated the levels and patterns of genetic diversity of Pinus densata Master in Yunnan. Horizontal starch-gel electrophoresis was performed on macrogametophytes collected from nine populations in northwestern Yunnan, China. Compared with other gymnosperm species, P. densata has higher mean values for all measures of genetic diversity. Allozyme polymorphism (0.99 criterion) was 97.0% and 71.4% at the species and population levels, respectively. The average number of alleles per locus was 3.1 and 2.0 at the species and population levels. Mean expected heterozygosity was substantially higher in P. densata than average values investigated for other gymnosperms both at the population (H _ep = 0.174±0.031) and at the species (H _es = 0.190) levels. Of the total genetic variation, less than 12% was partitioned among populations (G _ST = 0.112). Our allozyme survey supports the suggestion that the observed higher diversity in P. densata may be attributed partly to its hybrid origin between two genetically distinct species, P. yunnanensis and P. tabulaeformis. In addition, we suggest that introgression would give rise to the increase in genetic diversity occurring in P. densata.     

Genetic diversity and structure of the endemic Caesalpinia hintonii complex (Leguminosae: Caesalpinioideae) in Mexico

The Caesalpinia hintonii complex is formed by five endemic species (C. hintonii, C. laxa, C. macvaughii, C. melanadenia and C. epifanioi) occurring in central Mexico. This species complex is under incipient genetic divergence as by-product of local adaptations in reproductive and morphological traits to different habitats. We estimate the genetic variation and structure of populations of this species complex to assess the extent of genetic differentiation among populations and related species along its geographic distribution. Estimations of genetic diversity and structure were done based on ten enzymes and 18 loci. Mean number of alleles per locus ranged from 1.5 to 1.9. Polymorphic loci ranged from 42.1 to 68.4. Observed (H_o: range 0.191–0.275) and expected (H_e: range 0.205–0.317) heterozygosities in this complex were higher compared with other endemic and legume species. Neis genetic diversity estimates showed that most genetic variation was found within (H_S = 0.325) rather than among populations (D_ST = 0.085). Populations of the species C. hintonii showed a considerable genetic differentiation (F_ST = 0.207). The results of genetic diversity and structure within and among populations are in accord with the great morphological differentiation described for this species complex.     

Genetic differentiation in Thai populations of the rare species Afgekia sericea Craib (Leguminosae) revealed by RAPDPCR assays

Genetic diversity within and among populations was investigated using RAPD-PCR assays in a rare species, Afgekia sericea Craib. Two hundred and sixty-nine individuals were sampled from nine geographically isolated populations from northeastern Thailand. This study includes 73 RAPD markers. Within population polymorphism as measured by percentage of polymorphic RAPDs, varied between 38.4% and 60.3%. Genetic variability was measured using Shannon's information index and partitioned into between- and within-population components. Overall, genetic variation among A. sericea populations was high (H_sp-H_pop)/H_sp=48.2%. The genetic diversity for the species (H_sp) was 43.2%. Mean of within-population values (H_pop) for all populations was 0.224. The total genetic diversity was explained by high variation among populations (mean Gst=0.426), which is consistent with low gene flow among populations (Nm=0.35). High between-population genetic variation observed in this study could be explained by limited migration through seed and/or pollen dispersal among populations. Conservation strategies of A. sericea are discussed in the context of these results. This revised version was published online in July 2006 with corrections to the Cover Date.     

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.     

Genetic diversity among geographically separated populations of Nepenthes mirabilis

The distribution of Nepenthes mirabilis ranges from Northeast (NE) to South (S) Thailand. Eleven individuals from NE, S and Suen Jatujak market in Bangkok, Central (C) Thailand, were collected and divided into four populations according to their geographical areas. These four populations were analyzed to determine a genetic diversity profile using thirteen inter-simple sequence repeat markers. The individuals produced 75.18% polymorphic banding profiles. The Shannon’s index was used to estimate genetic diversity. Total genetic diversity (H _T) and inter-population genetic diversity (H _S) were 0.854 and 0.678, respectively. The degree of genetic differentiation (G _ST) of the species populations is 0.206, whereas the gene flow (Nm) among all the various geographical area populations is 1.016. Both the dendrogram and the results of the Shannon’s diversity index suggest great genetic diversity. These results support the broad range of distribution sites of Nepenthes mirabilis, which would require high genetic diversity to adapt to the environmental variations that can be found between NE, C, and S Thailand. ANOVA shows that the genetic diversity in each population is not significantly different (P > 0.05). Mantel tests reveal that geographical distance is an important factor for affecting the genetic distances among populations.     

Detection of Low Genetic Variation in a Critically Endangered Chinese Pine, Pinus squamata, Using RAPD and ISSR Markers

With only 32 individuals in the northeastern corner of Yunnan Province, China, Pinus squamata is one of the most endangered conifers in the world. Using two classes of molecular markers, RAPD and ISSR, its very low genetic variation was revealed. Shannon's index of phenotypic diversity (I) was 0.030, the mean effective number of alleles per locus (A_e) was 1.032, the percentage of polymorphic loci (P) was 6.45, and the expected heterozygosity (H_e) was 0.019 at the species level based on RAPD markers. The results of ISSR were consistent with those detected by RAPD but somewhat higher (I = 0.048, A_e = 1.042, P = 12.3, H_e = 0.029). The genetic variation of the subpopulation on the southwest-facing slope was much higher than that of the subpopulation on the northeast-facing slope, which may be attributed to the more diverse environment on the southwest-facing slope. The genetic differentiation between the two subpopulations was very low. The between-subpopulation variabilities, Φ_ST, calculated from RAPD and ISSR data were 0.011 and 0.024. Because of the lack of fossil records and geological historical data, it was difficult to explain the extremely low genetic diversity of the species. We postulate that this ancient pine might have experienced strong bottlenecks during its long evolutionary history, which caused the loss of genetic variation. Genetic drift and inbreeding in post-bottlenecked small populations may be the major forces that contribute to low genetic diversity. Human activities such as logging may have accelerated the loss of genetic diversity in P. squamata.     

Low levels of genetic variation inPhenakospermum guyannense (Strelitziaceae), a widespread bat-pollinated Amazonian herb

Phenakospermum guyannense is a monotypic, arborescent, long-lived monocot that is widespread in Amazonian South America. This outcrossing species is pollinated primarily by phyllostomid bats. Given these life-history characteristics,P. guyannense is expected to exhibit high levels of genetic variation and gene flow. We used isozyme electrophoresis and randomly amplified polymorphic DNA (RAPD) to characterize genetic variation in populations ofP. guyannense from French Guiana. Both measures detected a surprisingly low level of genetic variation, with only five out of twenty (25%) allozyme loci polymorphic (P), 1.35 alleles per locus (A), and an expected heterozygosity (H_e) of 0.090 at the species level. Isozymic genetic variation was even lower within populations (P = 17.5, A = 1.24, H_e = 0.074), and was corroborated by a RAPD assay that used 26 arbitrary primers (P = 3.61, A = 1.04, H_e = 0.014). Although overall levels of variation were low, the detectable variation was distributed as would be expected for an outcrossing species with extensive gene flow (mean G_ST = 0.230). We suspect thatP. guyannense is depauperate in genetic variation because of a series of bottlenecks that affected the species over this portion of its range.     

Genetic diversity of four esterase loci in natural populations of Hordeum spontaneum C. Koch from Jordan

Summary The diversity of four esterase loci was studied electrophoretically in 690 individual spikes representing 12 populations of wild barley (Hordeum spontaneum C. Koch.) collected from central, peripheral and marginal regions of its distribution in Jordan. A minimum of 6, 10, 5 and 5 alleles were observed at the Est-1, Est-2, Est-4 and Est-5 loci, respectively. Est-2 and Est-4 were the most diverse loci (H_c=0.53±0.05 and 0.46±0.07, respectively). Est-5 was intermediate (H_c=0.33+0.07) and Est-1 was the lowest (H_c=0.22±0.04). Polymorphism was highest in the central populations (H_e=0.52±0.04), followed by the peripheral (H_e=0.40±0.05) and the marginal (H_e=0.22±0.05) populations. Average allelic diversity between (G_st=0.49) and within (H_s=0.51) populations reflects a high allelic differentiation among these populations. Log-linear analyses revealed that four two-locus terms and two three-locus terms were significantly associated (P<0.05). Geographical distances between populations were not significantly correlated with Nei's genetic similarity index (r=0.16; P<0.19). It is postulated that diversifying selection is a major factor in the population genetic differentiation of these esterase 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。     

Genetic Diversity and Geographic Differentiation in the Threatened Species Dysosma pleiantha in China as Revealed by ISSR Analysis

Dysosma pleiantha, an important threatened medicinal plant species, is restricted in distribution to southeastern China. The species is capable of reproducing both sexually and asexually. In this study, inter-simple sequence repeat marker data were obtained and analyzed with respect to genetic variation and genetic structure. The extent of clonality, together with the clonal and sexual reproductive strategies, varied among sites, and the populations under harsh ecological conditions tended to have large clones with relatively low clonal diversity caused by vegetative reproduction. The ramets sharing the same genotype show a clumped distribution. Across all populations surveyed, average within-population diversity was remarkably low (e.g., 0.111 for Nei’s gene diversity), with populations from the nature reserves maintaining relatively high amounts of genetic diversity. Among all populations, high genetic differentiation (AMOVA: Φ_ST = 0.500; Nei’s genetic diversity: G _ST = 0.465, Bayesian analysis: Φ_B = 0.436) was detected, together with an isolation-by-distance pattern. Low seedling recruitment due to inbreeding, restricted gene flow, and genetic drift are proposed as determinant factors responsible for the low genetic diversity and high genetic differentiation observed.     

Genetic variation and population structure in Korean populations of sand dune speciesSalsola komarovi (Chenopodiaceae)

Salsola komarovi lljin is a herbaceous annual native to the sand dunes and beaches of Japan, northern China, Sakhalln and Korea. Starch-gel electrophoresis was conducted on leaves and stems collected from 300 plants in eight Korean populations. The mean number of alleles per locus (A _p=1.51), mean expected heterozygosity (He _p=0.116), and total genetic diversity (H _T=0.279) were comparable with those for species with similar life history and ecological traits. A general conformance of genotype frequencies to Hardy-Weinberg expectations (meanF _IS=−0.030) indicates thatS. komarovi is an outcrossing species. Slightly more than 20% of the genetic variation was found among populations (F _ST=0.204). In addition, significant differences in allele frequency were detected between populations at all 11 polymorphic loci (P<0.001). Nei's genetic identities range from 0.885 to 0.985 with a mean of 0.942. However, indirect estimates of the number of migrant per generation (0.97, calculated fromF _ST and 0.31, calculated from seven private alleles) indicate that the levels of gene flow is low among Korean populations. Although the species maintains a moderate level of genetic variation within populations, the small, isolated natural populations of the species have been severely destructed by human activities, particularly in summer season. If this is true, conservation efforts should be focused on those populations that currently maintain the most genetic diversity (e.g., populations of Cheju Island and coast of the southwestern Korean Peninsula).     

Demographic bottlenecks and low gene flow in remnant populations of the critically endangered <Emphasis Type="Italic">Berchemiella wilsonii</Emphasis> var<Emphasis Type="Italic">. pubipetiolata</Emphasis> (Rhamnaceae) inferred from microsatellite markers

Berchemiella wilsonii var. pubipetiolata (Rhamnaceae) is an endangered plant with only four remnant populations in eastern China. Population genetic information is essential for understanding population history and formulating conservation strategies for this species. Thirteen microsatellite loci were used to investigate genetic variation and population structure of the four remnant populations. Moderate levels of expected heterozygosity (H _E = 0.466–0.543) and low allelic diversity (A = 3.1–3.6 and A _R = 2.2–2.4, respectively) were observed within populations. Bottleneck tests found three out of four populations to deviate from mutation-drift equilibrium under the two-phase model (TPM), suggesting a recent population decline, which is congruent with known demographic history. The evolutionary history of the species seems dominated by genetic drift rather than gene flow. Low historical gene flow was inferred from several different approaches and N _m ranged from 0.582 by the private allele method to 0.783 by the coalescent method. Contemporary gene flow was also found to be even lower for only one first generation migrant was detected with individual-based assignment analysis. Restricted pollen and seed dispersal as well as a recent decline in population size associated with habitat fragmentation may have contributed to low levels of historical and contemporary gene flow, and resulted in a high genetic differentiation. Under this scenario, Berchemiella wilsonii var. pubipetiolata populations are expected to display more pronounced population genetic structure in the future as a result of increased inbreeding and genetic drift.     

Genetic Diversity and Genetic Structure of an Endangered Species, Trillium tschonoskii

The genetic diversity and genetic structure of Trillium tschonoskii (Maxim) were investigated using amplified fragment length polymorphism markers. Eight primer combinations were carried out on 105 different individuals sampled from seven populations. Of the 619 discernible DNA fragments generated, 169 (27.3%) were polymorphic. The percentage of polymorphic bands within populations ranged from 4.52 to 10.50. Genetic diversity (H_E) within populations ranged from 0.0130 to 0.0379, averaging 0.0536 at the species level. Genetic differentiation among populations was detected based on Nei's genetic diversity analysis (53.03%) and analysis of molecular variance (AMOVA) (52.43%). AMOVA indicated significant genetic differentiation among populations (52.43% of the variance) and within populations (47.57% of the variance) (p < 0.0002). Gene flow was low (0.4429) among populations. Species breeding system and limited gene flow among populations are plausible reasons for the high genetic differentiation observed for this species. We propose an appropriate strategy for conserving the genetic resources of T. tschonoskii in China.