濒危物种山红树居群遗传结构的RAPD分析

利用RAPD分子标记检测了云南省3个山红树居群的遗传多样性和居群遗传结构。10个引物共检测到54个位点, 其中多态位点11个, 占20 37%。与其他的濒危物种相比, 山红树居群内的遗传多样性很低, 居群间的遗传分化很大(78 65%)。大量经济植物的种植加上人为的破坏, 使山红树的生境遭到严重破坏, 数量大为减少, 可能导致了山红树遗传多样性的丧失、居群间较高的遗传分化。基于以上结果, 探讨了山红树进一步的迁地保护措施。     

Reduced Genetic Diversity and Effective Population Size in an Endangered Atlantic Salmon (Salmo Salar) Population from Maine, USA

Atlantic salmon (Salmo salar) populations in Maine, USA, are listed as a Distinct Population Segment under the U.S. Endangered Species Act due to reduced spawning runs and juvenile densities. Whenever possible, optimal conservation strategies for endangered populations should incorporate both present and historical knowledge of genetic variation. We assayed genetic diversity at seven microsatellite loci and at the mitochondrial ND1 gene in an endangered wild population of Atlantic salmon captured from the Dennys River from 1963 to 2001 using DNA’s extracted from archival scale and tissue samples. We examined temporal trends of genetic diversity, population structure, and effective population size (Ne). Overall temporal trends of diversity and Ne show significant reductions from 1963 to 2001 raising the possibility that current restoration efforts may be impacted by historical loss of diversity potentially critical to adaptation. Although our results suggest genetic stability in this population from 1963 to 1981, significant differentiation was observed for both the 1995 and 2001 samples compared with all other temporal samples. The presence of an ND1 mtDNA haplotype in this population, historically observed only in European and Newfoundland stocks, may represent previously unrecognized local wild diversity or, alternatively, may represent introgression from non-native fish.     

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.     

Population Genetic Structure of Endemic and Endangered Yellow Catfish, Horabagrus brachysoma, Using Allozyme Markers

The allozyme variation and population genetic structure of Horabagrus brachysoma in three natural populations from the southern part of the Western Ghats region, India, were investigated by polyacrylamide gel electrophoresis. Variations at 14 loci from 14 enzyme systems were analyzed. The allozyme analysis revealed a high level of genetic variation in this species, with an average of observed alleles per locus of 2.357 and observed heterozygosity of 0.178. The positive value of the fixation index (FIS=0.507) implied a significant deficiency of heterozygosity at the population level. The highly significant probability (P<0.0001) for the overall loci suggested that the three sample sets were not part of the same gene pool.     

Analysis of the Genetic Structure of Endangered Bovine Breeds from the Western Pyrenees Using Dna Microsatellite Markers

In the Western Pyrenees, three out of four native cattle breeds are in grave danger of extinction. Genetic variation of all four breeds was assessed by analyzing 478 animals using 11 microsatellite markers. A moderate/high within-breed variability was found, a favorable factor to consider when planning conservation and improvement programs. Interestingly, the only selected commercial breed, the Pirenaica, showed depressed heterozygosity levels and a low average number of alleles, perhaps explainable by intensive human selection exacerbated by a bottleneck effect. The Pirenaica also exhibited pronounced genetic differences and was the largest contributor of diversity among the breeds from the Western Pyrenees. Among endangered cattle breeds from this region, our results highlight the singularity of the Betizu. Geographic isolation among herds may be responsible for the large F(IS) value found in the Betizu breed. Lastly, our study suggests that the use of highly selected breeds may be one of the causes of distortion in phylogenetic analyses.     

珍稀濒危植物安徽羽叶报春遗传多样性的RAPD分析

利用RAPD分子标记对安徽特有濒危物种安徽羽叶报春（Primula merrilliana）6个自然居群的134个个体的遗传多样性进行了研究。从100个随机引物中筛选出12个RAPD引物,扩增共得到158条带,其中129个多态性位点（PPL）。POPGENE分析显示安徽羽叶报春具有较丰富的遗传变异（PPL=81.65%,He=0.2515,Ho=0.3849）。Nei′s基因多样性指数计算的居群间遗传分化系数（GST=0.5511）与Shannon信息指数（54.48%）基本一致。生境的片段化和基因流障碍可能是导致居群间遗传分化显著的主要原因。针对安徽羽叶报春的居群遗传变异提出了相应的保护措施：保护好自然生境和现有的居群及个体;加强居群间的基因流动;在迁地保护过程中,在尽可能多的居群中采样,以提高栽培居群的遗传多样性。     

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.     

The use of random amplified polymorphic DNA (RAPD) for hybrid detection in Scirpus from the river Schelde (Belgium)

Along the unique freshwater tidal zone of the river Schelde (Belgium), plants of Scirpus species occur primarily in small and fragmented populations. The majority of these are native Scirpus lacustris, S. pungens, S. triqueter, S. tabernaemontani and intermediate morphological forms. The distribution area of S. triqueter is even restricted to this tidal habitat. However, several cultivated S. tabernaemontani strains have recently been introduced. The latter species is often used to stabilize riverbanks. To determine the existing genetic diversity among these species, stems from plants from 44 different locations were subjected to random amplified polymorphic DNA (RAPD) analysis, using 22 decanucleotide primers. Data analysis of the amplified DNA fragments enabled us to unambiguously differentiate among these Scirpus taxa. Hybridization between S. triqueter and S. tabernaemontani was documented, and the studied hybrids were always genetically more similar to S. triqueter than to S. tabernaemontani . Among the introduced clones, at least two different origins could be distinguished. Several of the introduced S. tabernaemontani clones were clearly different from individuals of the same species that were native to this region. As there were different hybrid genotypes owing to different hybrid events or to introgression, this complex should be safeguarded from genetic pollution. New S. tabernaemontani genotypes with markers not previously observed in that hybrid complex of the unique tidal freshwater habitat should be avoided in replantation projects.     

Amphibian Body Impressions from the Mississippian of Pennsylvania,USA

Temnocorpichnus isaacleai is a new ichnogenus and ichnospecies of temnospondyl amphibian body impression from the Mississippian (Visean) Mauch Chunk Formation of eastern Pennsylvania, United States. The shovel-shaped head, robust limbs, relatively short trunk and smooth integument diagnose the ichnotaxon and readily distinguish it from Hermundurichnus and Sauropleura, the only other named body impressions of Paleozoic tetrapods. Temnocorpichnus is a compound ichnogenus in which the footprint ichnogenus Batrachichnus is a behaviorally distinct component. The temnospondyl identity of Temnocorpichnus adds to the sparse and earliest records of temnspondyls, which are of Visean age. The smooth integument of the ichnogenus does not support the presence of ventral scales or armor in the earliest temnospondyls, but body proportions of the Mauch Chunk body impressions indicate a relatively terrestrial temnospondyl not matched by any taxon now known from bones. Three closely associated impressions of Temnocorpichnus on a single bedding plane suggest some sort of gregarious behavior in Mississippian temnospondyls and may support speculation that internal fertilization and associated courtship behavior evolved independently in one group of amphibians more than 300 million years ago.     

珍稀濒危植物阜康阿魏的遗传多样性及遗传结构

明确珍稀濒危小种群阜康阿魏(Ferula fukanensis)的遗传多样性和遗传结构,是对其制定有效的保护和管理策略的基础和前提。本研究基于10对多态性好且可以稳定扩增的SSR引物对来源于3个居群87个珍稀濒危植物阜康阿魏的遗传多样性和遗传结构进行分析。结果显示：小种群的阜康阿魏具有相对较高的遗传多样性,居群间Nei’s基因多样性指数(h_S)为0.514,总的Nei’s基因多样性指数(h_T)为0.516,观测杂合度(H_o)为0.881,期望杂合度(H_e)为0.512,香农信息指数(I)为0.836,多态位点百分率(P_PB)为100%,遗传分化程度较低(F_st=0.007),95.9%的变异发生在居群内,遗传距离与地理距离无显著相关性,66.7%的居群遭遇了遗传瓶颈。结果表明,阜康阿魏遗传变异丰富,有较高的进化潜力。结合该种野外种群现状,建议建立保护区,开展就地保护,并加强引种和人工繁育等迁地保护措施,辅助阜康阿魏保育。本研究可为阜康阿魏植物资源保护提供理...     

Genetic Structure of an Endangered Portuguese Semiferal Pony Breed, the Garrano

The present study intends to survey the genetic variability of an endangered semiferal Portuguese native pony breed, the Garrano. Thirteen microsatellite markers were examined in 277 animals born in 1998, belonging to eight subpopulations corresponding to eight northern Portuguese geographic regions. Mean heterozygosity (H_o) in the Garrano breed was 0.732, ranging from 0.531 to 0.857 across subpopulations. Allelic frequencies and diversity differed significantly between regions, suggesting the existence of genetic differentiation within the breed confirmed by the population differentiation estimator F_ST. Allele sharing genetic distance (Dps) was used to determine the relationship between the analyzed subpopulations, some of which are diverging significantly from the others. Relationships among six Iberian horse breeds (including the Garrano) were assessed through the restricted maximum likelihood method, which clusters the Garrano with another Celtic pony, the Spanish Asturcon, traditionally bred in geographical continuity with the Garrano. Results reveal a statistically significant deficit of heterozygotes within the Garrano breed (F_IT = 0.031, p < 0.05). Regular monitoring should therefore be implemented so that the effect of genetic drift within subpopulations, enhanced by inbreeding, may be successfully minimized.     

ISSR and SRAP Markers Reveal Genetic Diversity and Population Structure of an Endangered Slipper Orchid,Paphiopedilum micranthum (Orchidaceae)

Paphiopedilum micranthum is an endangered pink slipper orchid mainly distributed in the limestone areas of southwestern China. Wild populations of this species have been seriously threatened by excessive collections, rampant smuggling for export, and habitat destruction. We used 15 ISSR markers and 11 SRAP markers to investigate the genetic diversity and structure of 15 natural populations. A high degree of diversity was observed at the species level (ISSR: PPB=9166%, He=03839; SRAP: PPB=9929%, He=02806). Certain degree of genetic differentiation among populations (ISSR: Gst=02577; SRAP: Gst=02383) was detected maybe caused by low gene flow (ISSR: Nm=07201; SRAP: Nm=07991). Consistent with the results of Principal Coordinate Analysis, the UPGMA dendrogram analysis divided the 15 populations into two main clades. In addition to geographic distance, the difference in elevation was another natural factor contributing to this differentiation. Knowledge about genetic diversity and structure gained from our study will be beneficial for the development of reasonable and efficient strategies to conserve this endangered species.     

中国西部麦红吸浆虫种群遗传结构的RAPD分析(双翅目:瘿蚊科)

对采自我国西北春麦区武威市 ( 38°N,1 0 3°E)、皋兰县 ( 36.5°N,1 0 4°E)及冬麦区长安县 ( 34.2°N,1 0 9°E)的麦红吸浆虫 Sitodiplosis mosellana ( Gehin)种群和皋兰县的麦黄吸浆虫 Contarinia tritici( Kirby)种群用 4种引物 S1 8、S61、S50、S83进行了RAPD测定分析。结果表明 ,引物共产生 1 0 6个 RAPD标记 ,没有任何一个引物能单独区分所有个体 ,但四种引物扩增结果联合则可。引物在麦红吸浆虫的 3个地理种群之间没有产生各自的特征带谱 ,但在 2个种之间可扩增出鉴别图谱。聚类分析显示 :同种同地理种群的个体间遗传相似程度高 ,变异范围小 ;同种不同地理种群间的遗传相似程度变异范围大 ,部分与种群内个体间的遗传相似程度重叠 ;不同种之间的遗传相似程度最低。在麦红吸浆虫种群间 ,长安与皋兰的种群遗传关系最近 ,而与武威种群较远。以近缘属种麦黄吸浆虫作参照分析表明 ,麦红吸浆虫不同地理种群之间表现为种下的遗传差异。     

Comparison of microsatellites, single-nucleotide polymorphisms (SNPs) and composite markers derived from SNPs in linkage analysis

There is growing evidence that a map of dense single-nucleotide polymorphisms (SNPs) can outperform a map of sparse microsatellites for linkage analysis. There is also argument as to whether a clustered SNP map can outperform an evenly spaced SNP map. Using Genetic Analysis Workshop 14 simulated data, we compared for linkage analysis microsatellites, SNPs, and composite markers derived from SNPs. We encoded the composite markers in a two-step approach, in which the maximum identity length contrast method was employed to allow for recombination between loci. A SNP map 2.3 times as dense as a microsatellite map (approximately 2.9 cM compared to approximately 6.7 cM apart) provided slightly less information content (approximately 0.83 compared to approximately 0.89). Most inheritance information could be extracted when the SNPs were spaced < 1 cM apart. Comparing the linkage results on using SNPs or composite markers derived from them based on both 3 cM and 0.3 cM resolution maps, we showed that the inter-SNP distance should be kept small (< 1 cM), and that for multipoint linkage analysis the original markers and the derived composite markers had similar power; but for single point linkage analysis the resulting composite markers lead to more power. Considering all factors, such as information content, flexibility of analysis method, map errors, and genotyping errors, a map of clustered SNPs can be an efficient design for a genome-wide linkage scan.     

Low Genetic Diversity and Strong Geographical Structure of the Critically Endangered White-Headed Langur (Trachypithecus leucocephalus) Inferred from Mitochondrial DNA Control Region Sequences

Many Asian colobine monkey species are suffering from habitat destruction and population size decline. There is a great need to understand their genetic diversity, population structure and demographic history for effective species conservation. The white-headed langur (Trachypithecus leucocephalus) is a Critically Endangered colobine species endemic to the limestone karst forests in southwestern China. We analyzed the mitochondrial DNA (mtDNA) control region sequences of 390 fecal samples from 40 social groups across the main distribution areas, which represented one-third of the total extant population. Only nine haplotypes and 10 polymorphic sites were identified, indicating remarkably low genetic diversity in the species. Using a subset of 77 samples from different individuals, we evaluated genetic variation, population structure, and population demographic history. We found very low values of haplotype diversity (h = 0.570 ± 0.056) and nucleotide diversity (π = 0.00323 ± 0.00044) in the hypervariable region I (HVRI) of the mtDNA control region. Distribution of haplotypes displayed marked geographical pattern, with one population (Chongzuo, CZ) showing a complete lack of genetic diversity (having only one haplotype), whereas the other population (Fusui, FS) having all nine haplotypes. We detected strong population genetic structure among habit patches (Φ _ST = 0.375, P < 0.001). In addition, the Mantel test showed a significant correlation between the pairwise genetic distances and geographical distances among social groups in FS (correlation coefficient = 0.267, P = 0.003), indicting isolation-by-distance pattern of genetic divergence in the mtDNA sequences. Analyses of demographic history suggested an overall stable historical population size and modest population expansion in the last 2,000 years. Our results indicate different genetic diversity and possibly distinct population history for different local populations, and suggest that CZ and FS should be considered as one evolutionarily significant unit (ESU) and two management units (MUs) pending further investigation using nuclear markers.     

Genetic Connectivity among Populations of an Endangered Snake Species from Southeastern Australia (Hoplocephalus bungaroides, Elapidae)

For endangered species that persist as apparently isolated populations within a previously more extensive range, the degree of genetic exchange between those populations is critical to conservation and management. A lack of gene flow can exacerbate impacts of threatening processes and delay or prevent colonization of sites after local extirpation. The broad-headed snake, Hoplocephalus bungaroides, is a small venomous species restricted to a handful of disjunct reserves near Sydney, Australia. Mark-recapture studies have indicated low vagility for this ambush predator, suggesting that gene flow also may be low. However, our analyses of 11 microsatellite loci from 163 snakes collected in Morton National Park, from six sites within a 10-km diameter, suggest relatively high rates of gene flow among sites. Most populations exchange genes with each other, with one large population serving as a source area and smaller populations apparently acting as sinks. About half of the juvenile snakes, for which we could reliably infer parentage, were collected from populations other than those in which we collected their putative parents. As expected from the snakes' reliance on rocky outcrops during cooler months of the year, most gene flow appears to be along sandstone plateaux rather than across the densely forested valleys that separate plateaux. The unexpectedly high rates of gene flow on a landscape scale are encouraging for future conservation of this endangered taxon. For example, wildlife managers could conserve broad-headed snakes by restoring habitats near extant source populations in areas predicted to be least affected by future climate change.     

Phytoremediation of Wastewater Containing Lead (Pb) in Pilot Reed Bed Using Scirpus Grossus

Phytoremediation is a technology to clean the environment from heavy metals contamination. The objectives of this study are to threat Pb contaminated wastewater by using phytoremediation technology and to determine if the plant can be mention as hyperaccumulator. Fifty plants of Scirpus grossus were grown in sand medium and 600 L spiked water in various Pb concentration (10, 30 and 50 mg/L) was exposed. The experiment was conducted with single exposure method, sampling time on day-1, day-14, day-28, day-42, day-70, and day-98. The analysis of Pb concentration in water, sand medium and inside the plant tissue was conducted by ICP-OES. Water samples were filtered and Pb concentration were directly analyzed, Pb in sand samples were extracted by EDTA method before analyzed, and Pb in plant tissues were extracted by wet digestion method and analyzed. The results showed that on day-28, Pb concentration in water decreased 100%, 99.9%, 99.7%, and the highest Pb uptake by plant were 1343, 4909, 3236 mg/kg for the treatment of 10, 30, and 50 mg/L respectively. The highest BC and TF were 485,261 on day-42 and 2.5295 on day-70 of treatment 30 mg/L, it can be mentioned that Scirpus grossus is a hyperaccumulator.     

Genetic Analysis of Indian Lentil (Lens culinaris Medikus) Cultivars and Landraces Using RAPD and STMS Markers

Molecular analysis of 29 lentil (Lens culinaris) cultivars and landraces of Indian origin was carried out using twenty RAPD and ten cross-species STMS primers. A total of 97 markers (72 RAPD and 25 STMS) were amplified of which 42.3% were polymorphic. Genetic similarity among the cultivars and landraces was 89.7%. The observed results indicated low level of genetic diversity in the studied material. UPGMA cluster analysis for the combined data of RAPD and STMS revealed two broad clusters — Cluster I with three landraces and Cluster II containing all remaining landraces and cultivars except Precoz. Germplasm line Precoz was found to be the most distinct in individual as well as combined analyses. All cultivars and landraces except K-75 and L4076 could be discriminated from one another using combined data for the two techniques. Germplasm lines Precoz, L830 and cultivars L4147 and JL3 were quite distinct and could be potential germplasm resource.     

Evaluation of Genetic Variation in the Clown Knifefish, Chitala chitala, Using Allozymes, RAPD, and Microsatellites

Twenty-seven enzyme systems, six random amplified polymorphic DNA (RAPD) primers, and two microsatellite loci were tested to determine intraspecific divergence in the natural population of the endangered Indian featherback fish, Chitala chitala, for the first time. The 262 samples of C. chitala were collected from six riverine locations in India: the Satluj, Ganga (Ghagra, Bhagirathi, and Brahmaputra), Mahanadi, and Narmada river systems. The analysis revealed population subdivisions, with an F_ST value from 0.1235 (95% confidence 0.0868–0.1621) for RAPD and a combined F_ST of 0.0344 (95% confidence 0.0340–0.0350) for microsatellite loci. An analysis of 38 allozyme loci did not reveal any polymorphism in the samples from any of the riverine localities; a possible explanation for this could be that the ancestors of Chitala could have faced a population reduction in prehistoric periods, as low allozyme variation is also reported for other species of Chitala from south Asia.     

Microhabitat Types Promote the Genetic Structure of a Micro-Endemic and Critically Endangered Mole Salamander (Ambystoma leorae) of Central Mexico

The reduced immigration and emigration rates resulting from the lack of landscape connectivity of patches and the hospitality of the intervening matrix could favor the loss of alleles through genetic drift and an increased chance of inbreeding. In order for isolated populations to maintain sufficient levels of genetic diversity and adapt to environmental changes, one important conservation goal must be to preserve or reestablish connectivity among patches in a fragmented landscape. We studied the last known population of Ambystoma leorae, an endemic and critically threatened species. The aims of this study were: (1) to assess the demographic parameters of A. leorae and to distinguish and characterize the microhabitats in the river, (2) to determine the number of existing genetic groups or demes of A. leorae and to describe possible relationships between microhabitats types and demes, (3) to determine gene flow between demes, and (4) to search for geographic locations of genetic discontinuities that limit gene flow between demes. We found three types of microhabitats and three genetically differentiated subpopulations with a significant level of genetic structure. In addition, we found slight genetic barriers. Our results suggest that mole salamander’s species are very sensitive to microhabitat features and relatively narrow obstacles in their path. The estimates of bidirectional gene flow are consistent with the pattern of a stepping stone model between demes, where migration occurs between adjacent demes, but there is low gene flow between distant demes. We can also conclude that there is a positive correlation between microhabitats and genetic structure in this population.