樟科濒危植物思茅木姜子遗传多样性的ISSR分析

本文采用ISSR标记对中国特有且仅在云南南部狭域分布的樟科濒危植物思茅木姜子(Litseaszemaois)现存8个居群的遗传多样性进行了研究。从96条引物中筛选出了10条,对103个个体进行了扩增,共扩增出77条条带,其中多态性条带为67条。分析结果表明:(1)思茅木姜子的遗传多样性水平很高。在物种水平上,多态位点百分率PPB=87.01%,平均每个位点的有效等位基因数Ne=1.4006,Nei’s基因多样度指数H=0.2466,Shannon多样性信息指数Hsp=0.3826;在居群水平上,PPB=37.99%,Ne=1.2500,H=0.1418,Shannon多样性信息指数Hpop=0.2088。(2)居群间的遗传分化较低。基于Nei’s遗传多样性分析得出的居群间遗传分化系数Gst=0.3700;Shannon’s居群分化系数((Hsp–Hpop)/Hsp)为0.45。AMOVA分析显示:思茅木姜子的遗传变异主要存在于居群内,占总变异的72.99%,居群间的遗传变异占27.01%,表明思茅木姜子属于异交种。(3)两两居群间的Nei’s遗传一致度(I)的范围为0.8233–0.9761。经Mantel检测,居群间的遗传距离和地理距离之间不存在显著的正相关关系(r=0.0925,P=0.6931)。我们推断人类活动的干扰和生境的片断化是导致思茅木姜子濒危现状的主要因素。考虑到目前其遗传多样性水平虽然很高,但各居群个体数量很少,因此应该对思茅木姜子各居群的所有个体实施及时的就地保护;而遗传变异大部分存在于居群内的个体间,所以在迁地保护时应在各居群内大量采样。     

中国樟科木姜子属植物纪要

描述一新种即沧源木姜子Litsea cangyuanensis J．Li et H．W．Li;一中国新纪录种白叶木姜子L．albescens（Hook．f．）D．G．Long;3个新异名：大叶木姜子L．chunii Cheng var．longipedicellata Yang f．latifolia Yang归并到高山木姜子L．chunii Cheng,狭叶桂北木姜子M．subcoriacea Yang et P．H．Huang val．stenophylla Yang et P．H．Huang归并入桂北木姜子L．subcoriacea Yang et P．H．Huang,狭叶华南木姜子L．greenmaniana Allen val．angustifolia Yang et P．H．Huang归入华南木姜子L．greenmaniana Allen;思茅木姜子L．szemaois（H．Liou）J．Li et H．W．Li作为新等级提升为种;并对秃净木姜子L．kingii Hook．f．与山鸡椒L．cubeba（Lour．）Pers．两者间的差异进行澄清,对台湾产的黄肉树应采用L．hypophaea Hayata和广西产的蜂窝木姜子应采用L．foveola Kosterm．进行了讨论。一个新名称：L．sinoglobosa J．Li et H．W．Li替代同名的L．globosa Yang et P．H．Huang。     

中国樟科木姜子属植物纪要*

描述一新种即沧源木姜子Litsea cangyuanensis J. Li et H. W. Li; 一中国新纪录种白叶木姜子L1albescens
( Hook. f. ) D. G. Long; 3 个新异名: 大叶木姜子L1 chunii Cheng var1longipedicellata Yang f1 latif olia Yang 归并到高
山木姜子L1 chunii Cheng, 狭叶桂北木姜子M1 subcoriacea Yang et P. H. Huang var. stenophylla Yang et P. H.
Huang 归并入桂北木姜子L1 subcoriacea Yang et P. H. Huang, 狭叶华南木姜子L1greenmaniana Allen var1 angustifolia
Yang et P. H. Huang 归入华南木姜子L1greenmaniana Allen; 思茅木姜子L1szemaois ( H. Liou) J. Li et H. W. Li
作为新等级提升为种; 并对秃净木姜子L1 kingii Hook. f. 与山鸡椒L1 cubeba ( Lour . ) Pers. 两者间的差异进
行澄清, 对台湾产的黄肉树应采用L1 hypophaea Hayata 和广西产的蜂窝木姜子应采用L1f oveola Kosterm. 进
行了讨论。一个新名称: L1 sinoglobosa J. Li et H. W. Li 替代同名的L1 globosa Yang et P. H. Huang 。     

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

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

Genetic 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.     

A new species of Litsea (Lauraceae) from India

Litsea beei, a new species of Lauraceae from India is described and illustrated. The new species is allied to the Burmese-Malayan species L. myristicaefolia.     

Inter-simple sequence repeat (ISSR) analysis of genetic variation of Chondrus crispus populations from North Atlantic

ISSR analysis was used to investigate genetic variations of 184 haploid and diploid samples from nine North Atlantic Chondrus crispus Stackhouse populations and one outgroup Yellow Sea Chondrus ocellatus Holmes population. Twenty-two of 50 primers were selected and 163 loci were scored for genetic diversity analysis. Genetic diversity varied among populations, percentage of polymorphic bands (PPB) ranged from 27.0 to 55.8%, H (Nei's genetic diversity) ranged from 0.11 to 0.20 and I (Shannon's information index) ranged from 0.16 to 0.30. Estimators PPB, H and I had similar values in intra-population genetic diversity, regardless of calculation methods. Analysis of molecular variance (AMOVA) apportioned inter-population and intra-population variations for C. crispus, showing more genetic variance (56.5%) occurred in intra-population, and 43.5% variation among nine populations. The Mantel test suggested that genetic differentiation between nine C. crispus populations was closely related with geographic distances (R = 0.78, P = 0.002). Results suggest that, on larger distance scale (ca. >1000 km), ISSR analysis is useful for determining genetic differentiations of C. crispus populations including morphologically inseparable haploid and diploid individuals.     

On the neutrality of molecular genetic markers: pedigree analysis of genetic variation in fragmented populations

Many studies employ molecular markers to infer ecological and evolutionary processes, assuming that variation found at genetic loci offers a reliable representation of stochastic events in natural populations. Increasingly, evidence emerges that molecular markers might not always be selectively neutral. However, only a few studies have analysed how deviations from neutrality could affect estimates of genetic variation, using populations with known genealogy. We monitored changes in allozyme variation over eight generations in captive metapopulations of the butterfly Bicyclus anynana. Population demography was recorded by individually marking 35 000 butterflies and constructing pedigrees. We designed a computer program that simulated the inheritance of founder allozyme alleles in butterfly pedigrees. We thus tested whether the observed transmission of allozyme alleles could be explained by random genetic drift alone, or whether there was evidence for positive or negative selection. This analysis showed that in the smallest metapopulations the loss of allozyme variation exceeded the neutral rate. Possibly, linkage disequilibria between deleterious mutations and marker alleles resulted in background selection and a faster erosion of allozyme variation. In larger metapopulations, one locus (MDH) showed a significant heterozygote excess and smaller than expected loss in heterozygosity, observations consistent with (associative) overdominance. This study demonstrates that the neutrality of molecular markers cannot always be assumed, particularly in small populations with a high mutation load.     

ISSR variation in the endemic and endangered plant Cycas guizhouensis (Cycadaceae)

BACKGROUND AND AIMS: Cycas guizhouensis (Cycadaceae) is a rare and endangered species endemic to the southwest of China. An investigation was undertaken into the genetic variation of wild populations. METHODS: ISSR markers were used to determine the genetic variation within and between 12 extant populations of this species. KEY RESULTS: Low genetic diversity (at population level, P = 14.21 %, H(E) = 0.0597; at species level, P = 35.90 %, H(T) = 0.1082) and a high degree of differentiation among populations (G(ST) = 0.4321) were detected. CONCLUSIONS: This genetic structure is considered to be due to the combined effects of slow biochemical evolution, genetic drift, inbreeding and limited gene flow between populations. Based on these findings, strategies are proposed for the genetic conservation and management of the species.     

Evidence for ancient genetic subdivision among recently fragmented populations of the endangered shrub Grevillea caleyi (Proteaceae)

The genetic effects of population fragmentation cannot be interpreted without understanding the underlying pattern of genetic variation resulting from historic population processes. We used AFLP markers to determine genetic structure and distribution of genetic diversity among populations of an endangered Australian shrub Grevillea caleyi (Proteaceae). Populations that occurred historically on four ridges have new been fragmented to varying degrees, producing some large, relatively pristine populations and very small populations consisting of fewer than 10 adult plants. We found marked population genetic structure (65.9% of genetic variation was among populations) and a significant relationship between genetic and geographic distance (rm=0.564, P=0.004). However, only 14% of overall genetic differentiation was attributable to variation among ridges, compared with 52% among populations within ridges. Moreover, genetic diversity within samples of plants did not vary with either population size or degree of isolation. Thus, the present genetic structure of populations is probably almost entirely the product of historical events. Fine-scale structuring within populations prior to fragmentation may have been caused by limited seed and pollen dispersal, despite a complex suite of (mostly avian) pollinators, and a mixed mating system that allows a large amount of selfing. The combined effects of adult longevity and a soil-stored seed bank may have buffered the recently fragmented populations against the effects of dramatic reductions in numbers of adult plants.     

High population differentiation and genetic variation in the endangered Mexican pine Pinus Rzedowskii (Pinaceae)

Pinus rzedowskii is an endangered pine species from Michoaca´n (central Me´xico), which has been previously reported from only three localities. Classified within the subgenus Strobus, it exhibits intermediate morphological characters between subgenera Strobus and Pinus. We analyzed genetic aspects that could shed light on the evolution and conservation of this species. The genetic structure of nine populations was examined using 14 isozyme loci. Pinus rzedowskii has a relatively high level of genetic variation with 46.8% of the loci assayed being polymorphic, a total of 35 alleles, and a mean heterozygosity per population of 0.219. We calculated Wright's F_ST statistic to estimate gene flow indirectly and to evaluate whether or not there was genetic structuring among populations. We found a marked differentiation among populations (F_ST = 0.175) and significant inbreeding (F_IS = 0.247). No pattern of isolation by distance was found. We also constructed a dendrogram based on a genetic distance matrix to obtain an overview of the possible historical relationships among populations. Finally, we found a convex relationship between the genetic distance among populations and the number of ancestral lineages, suggesting that demographically this species has not been at risk recently. Although endangered, with small and fragmented populations, P. rzedowskii shows higher levels of genetic variation than other conifer species with larger populations or similar conservation status.     

Genetic variation within the endangered quillwort Isoëtes hypsophila (Isoetaceae) in China as evidenced by ISSR analysis

The genetic diversity of 56 individuals of Isoëtes hypsophila Hand.-Mazz. from China was investigated by ISSR. Twelve primers were screened from 65 primers, and a total of 119 DNA fragments were scored, of these, 82% were polymorphic bands, which indicated that high levels of genetic variation existed in the natural populations. Genetic diversity varied greatly among populations with the percentage of polymorphic band (PPB) values ranging from 8 to 35%. An analysis of molecular variance (AMOVA) was used to apportion the variation between regions, among populations within regions, and within populations. Results indicated that most of the variance (85%) occurred between Yunnan and Sichuan. The variances among populations within regions and within populations, however, were only 5 and 10%, respectively. In the among-population analysis, the larger part of genetic variation (77%) resided among populations, and less (23%) presented differences within populations. UPGMA cluster analysis showed that there was no distinct genetic differentiation between populations from Sichuan province. A number of causes including limited gene flow, genetic drift and inbreeding might have led to these observed genetic profiles of I. hypsophila.     

Structural studies of an arabinoxylan isolated from Litsea glutinosa (Lauraceae)

A water-soluble arabinoxylan (D-xylose and L-arabinose in the molar ratio 1.0:3.4) was isolated from the mucilaginous bark of Litsea glutinosa (Lauraceae). The results of methylation analysis, partial hydrolysis, and 1H- and 13C-n.m.r. spectroscopy indicated a backbone of (1----4)-linked beta-D-xylopranosyl residues substituted at both positions 2 and 3 with side chains composed of either single or (1----3)-linked arabinofuranosyl residues. Both alpha-L- and beta-L-arabinofuranosyl residues were present. It is possible that side chains composed of two beta-L-arabinofuranosyl residues are attached mainly at O-2 of some xylosyl residues.     

Evolutionary history and genetic connectivity across highly fragmented populations of an endangered daisy

Conservation management can be aided by knowledge of genetic diversity and evolutionary history, so that ecological and evolutionary processes can be preserved. The Button Wrinklewort daisy (Rutidosis leptorrhynchoides) was a common component of grassy ecosystems in south-eastern Australia. It is now endangered due to extensive habitat loss and the impacts of livestock grazing, and is currently restricted to a few small populations in two regions >500 km apart, one in Victoria, the other in the Australian Capital Territory and nearby New South Wales (ACT/NSW). Using a genome-wide SNP dataset, we assessed patterns of genetic structure and genetic differentiation of 12 natural diploid populations. We estimated intrapopulation genetic diversity to scope sources for genetic management. Bayesian clustering and principal coordinate analyses showed strong population genetic differentiation between the two regions, and substantial substructure within ACT/NSW. A coalescent tree-building approach implemented in SNAPP indicated evolutionary divergence between the two distant regions. Among the populations screened, the last two known remaining Victorian populations had the highest genetic diversity, despite having among the lowest recent census sizes. A maximum likelihood population tree method implemented in TreeMix suggested little or no recent gene flow except potentially between very close neighbours. Populations that were more genetically distinctive had lower genetic diversity, suggesting that drift in isolation is likely driving population differentiation though loss of diversity, hence re-establishing gene flow among them is desirable. These results provide background knowledge for evidence-based conservation and support genetic rescue within and between regions to elevate genetic diversity and alleviate inbreeding.Subject terms: Ecological genetics, Population genetics     

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

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

濒危植物绵刺8个种群遗传多样性的AFLP分析

利用8对AFLP引物对我国绵刺的8个种群240份材料的基因组DNA进行扩增,得到大小在65～530bp之间的397条清晰显带,其中296(74.56%)条呈多态性,平均每对AFLP引物得到37条多态性带;用PopGen32软件将AFLP多态性数据进行分析,不同种群的Nei's基因多样性指数和Shannon信息指数变化范围分别在0.0845～0.1779和0.1280～0.2377之间,其中遗传多样性最高为上沙窝种群,最低为银根种群,可将上沙窝种群作为种质遗传中心之一进行保护.绵刺遗传变异有68.31%存在种群内,31.69%种群之间,说明变异主要存在于种群内部.8个种群的平均遗传距离为0.1341,按UPGMA进行聚类分析,结果表明绵刺种群具有明显的地域相关性和遗传类型趋同性,说明不同的种群可能有共同的起源,随机遗传漂变不是影响绵刺种群遗传多样性的主要过程.建议在迁地保护和取样时,不仅要在每个种群中取足够多的个体,而且要在尽可能多的种群中取样,最大限度地保护绵刺的遗传多样性,为进一步地系统演化研究奠定基础.     

秦岭秀雅杜鹃野生种群遗传多样性和遗传分化的AFLP分析

利用AFLP分子标记技术,对秦岭地区7个秀雅杜鹃野生种群的遗传多样性和遗传分化进行研究.结果表明:M52E41、M62E46和M64E94等3个引物组合共扩增出182条DNA片段,其中151条是多态的,多态位点比率为83.1％.7个秀雅杜鹃种群所检测的多态位点百分率(PPL)、Nei的基因多样性指数(h)和Shannon信息指数(I)的变化趋势一致,其排序为眉县种群＞柞水种群＞镇安种群＞户县种群＞宁强种群＞南郑种群＞周至种群.POPGENE分析表明,秀雅杜鹃在物种水平(PPL=91.22％,I=0.7217,h=0.5095)和种群水平(PPL=77.56％,I=0.6409,h=0.4725)都具有较高的遗传多样性.种群间的遗传分化系数(Gst)为7.26％,说明有92.74％的变异存在于种群内部.AMOVA分析表明,在总的遗传变异中,85.3％的变异发生在种群内,14.7％的变异发生在种群间,说明秀雅杜鹃种群的遗传变异主要存在于种群内部.UPGMA聚类分析表明,种群间的遗传距离与种群间的地理距离没有明显的相关性.最后提出了秦岭地区秀雅杜鹃种质资源的保护策略.     

Spatial isolation and genetic differentiation in naturally fragmented plant populations of the Swiss Alps

Aims The effect of anthropogenic landscape fragmentation on the genetic diversity and adaptive potential of plant populations is a major issue in conservation biology. However, little is known about the partitioning of genetic diversity in alpine species, which occur in naturally fragmented habitats. Here, we investigate molecular patterns of three alpine plants (Epilobium fleischeri, Geum reptans and Campanula thyrsoides) across Switzerland and ask whether spatial isolation has led to high levels of population differentiation, increasing over distance, and a decrease of within-population variability. We further hypothesize that the contrasting potential for long-distance dispersal (LDD) of seed in these species will considerably influence and explain diversity partitioning.Methods For each study species, we sampled 20–23 individuals from each of 20–32 populations across entire Switzerland. We applied Random Amplified Polymorphic Dimorphism markers to assess genetic diversity within (Nei's expected heterozygosity, H e; percentage of polymorphic bands, P p) and among (analysis of molecular variance, Φ st) populations and correlated population size and altitude with within-population diversity. Spatial patterns of genetic relatedness were investigated using Mantel tests and standardized major axis regression as well as unweighted pair group method with arithmetic mean cluster analyses and Monmonier's algorithm. To avoid known biases, we standardized the numbers of populations, individuals and markers using multiple random reductions. We modelled LDD with a high alpine wind data set using the terminal velocity and height of seed release as key parameters. Additionally, we assessed a number of important life-history traits and factors that potentially influence genetic diversity partitioning (e.g. breeding system, longevity and population size).Important findings For all three species, we found a significant isolation-by-distance relationship but only a moderately high differentiation among populations (Φ st : 22.7, 14.8 and 16.8%, for E. fleischeri, G. reptans and C. thyrsoides, respectively). Within-population diversity (H e : 0.19–0.21, P p : 62–75%) was not reduced in comparison to known results from lowland species and even small populations with <50 reproductive individuals contained high levels of genetic diversity. We further found no indication that a high long-distance seed dispersal potential enhances genetic connectivity among populations. Gene flow seems to have a strong stochastic component causing large dissimilarity between population pairs irrespective of the spatial distance. Our results suggest that other life-history traits, especially the breeding system, may play an important role in genetic diversity partitioning. We conclude that spatial isolation in the alpine environment has a strong influence on population relatedness but that a number of factors can considerably influence the strength of this relationship.     

准噶尔无叶豆片断化居群的遗传变异及克隆多样性

准噶尔无叶豆(Eremosparton songoricum)是豆科无叶豆属小半灌木, 既能开花结实进行有性繁殖, 又可以靠根茎进行无性克隆繁殖, 为国家三级保护植物, 在中国仅片断化分布于新疆古尔班通古特沙漠局部区域。本文采用ISSR分子标记对采自古尔班通古特沙漠腹地及边缘的7个准噶尔无叶豆自然居群共148个个体进行了遗传变异和克隆多样性分析。8个引物共扩增出84个位点, 其中77个为多态性位点, 物种水平上的多态位点百分比PPB为91.67％, Nei's基因多样性指数I为0.3192, Shannon信息指数H为0.3540; 居群水平上的多态位点百分比PPB为58.45％, Nei's基因多样性指数I为0.2248, Shannon信息指数H为0.3270。居群间的遗传分化系数G_ST为0.2978。AMOVA分析表明, 有31.88%的遗传变异存在于居群间, 表明居群间存在显著的遗传分化。克隆多样性分析表明, 居群水平上, 居群G的Simpson多样性指数和均匀度指数最高, 分别为0.9400和0.9885; 居群E最低, 分别为0.8457和0.9021。物种水平上, Simpson多样性指数为0.9858, 均匀度指数为0.9673。本研究结果表明, 和其他荒漠植物相比, 准噶尔无叶豆表现出较高的遗传变异水平和克隆多样性, 这主要与该物种兼性生殖的繁育方式及多克隆起源有关; 而居群间产生了显著的遗传分化则主要由于人为干扰引起的生境片断化和居群减小而导致的基因交流障碍所致。遗传变异水平和遗传结构的研究将为分析准噶尔无叶豆致濒原因及进化潜力提供重要资料, 对该物种保护具有指导意义。