Genetic diversity of Astragalus nitidiflorus, a critically endangered endemic of SE Spain,and implications for its conservation

Inter-simple sequence repeats (ISSR) markers were used to assess the genetic diversity and population structure in five populations of Astragalus nitidiflorus, a critically endangered species endemic to southeast Spain. Eight primers amplified 78 bands with 40 (51.3%) being polymorphic. Statistical results indicated a low genetic diversity at the population and species level, with percentages of polymorphic bands (PPB) ranging from 28.2 to 37.2% (an average of 31.8%), and means of gene diversity (H_E) of 0.129 and 0.171 respectively. The Shannon’s index (SI) ranged from 0.160 to 0.214 at the population level and was 0.260 at the species level. A low level of genetic differentiation among populations was detected, based on the Shannon’s information index (0.297), the coefficient of genetic differentiation between populations (G_ST = 0.2418) and AMOVA analysis (Φ_ST = 0.255). The estimated gene flow (Nm) was 0.789. The high genetic connectivity found among populations of A. nitidiflorus is an evidence of a recent habitat fragmentation. In addition, a bottleneck event in the past has been revealed, with a subsequent reduction of population size and a loss of genetic variation. Based on these results, the conservation strategy of A. nitidiflorus was proposed.     

Assessing genetic diversity of populations of topmouth culter (Culter alburnus) in China using AFLP markers

Genetic diversity of five wild populations and a cultured population of topmouth culter (Culter alburnus) was investigated using amplified fragment length polymorphism (AFLP). A total of 373 reproducible bands amplified with seven AFLP primer combinations were obtained from 163 fish. The percentage of polymorphic loci ranged widely from 37.0% to 69.2% within distinct populations. The cultured population appeared to have a lower level of polymorphism (37.0%), gene diversity (0.121 ± 0.188) and Shannon's Information index (0.183 ± 0.263) than the wild populations. Analysis of molecular variance (AMOVA) revealed that average F_ST value overall loci was 0.2671, and the percentage of variation within population (73.29%) was larger than among populations (26.71%) (P < 0.01). The six populations were clustered into two major clades with UPGMA. The results from analysis of population pairwise gene flow indicated moderate gene flow among populations. Our study indicated that the genetic diversity of the cultured population was reduced compared with the wild populations. Geographic isolation, habitat, and artificial selection all may have played important roles in population differentiation. The information may be beneficial to future broodstock selection and defining conservation management for the different populations of topmouth culter.     

Genetic structure and diversity of the Diachasmimorpha longicaudata species complex in Thailand: SSCP analysis of mitochondrial 16S rDNA and COI DNA sequences

Genetic variation among 20 populations of Diachasmimorpha longicaudata (Ashmead) (Hymenoptera: Braconidae) in Thailand was investigated using single strand conformation polymorphism (SSCP) analysis of mitochondrial DNA sequences. From a total 641 individual parasitoids, seven distinct haplotypes containing a total of 32 polymorphic sites were observed from cytochrome oxidase subunit I (COI) sequences along with five distinct haplotypes containing a total 16 polymorphic sites from 16S rDNA sequences. Values obtained through pairwise F_ST comparisons and analysis of molecular variance (AMOVA) indicated significant genetic differentiation among D. longicaudata populations in Thailand. Congruent relationships showing separation of these populations into three groups were obtained from Neighbor joining and Bayesian phylogenetic tree analyses along with the use of haplotype networks. This SSCP analysis of populations of the D. longicaudata species complex is the first report using molecular population genetic methods to analyze the structure of this parasitoid species in Thailand. This may provide useful information for release of parasitoid strains to maximize their benefit in biological control programs.     

Population structure and genetic diversity of Stipa grandis P. Smirn,a dominant species in the typical steppe of northern China

Random amplified polymorphic DNA (RAPD) markers were used to characterize genetic heterogeneity within and among five populations of Stipa grandis in the Xilingol Plateau. Estimates of the percentage of polymorphic bands, Shannon's diversity information index and Nei's gene diversity index were comparatively high in the five populations, and the Population GSM was found to have the highest genetic diversity among all populations. An analysis of molecular variance indicated that the majority of variation existed within populations (74.12%), and that there was significant differentiation among populations (Φ_ST = 25.88%, P < 0.001). Genetic distance (Φ_ST) ranged from 0.198 to 0.310 and the differentiation between pair-wise populations was significant when individual pairs of populations were compared. Based on the Φ_ST values, gene flow (Nm) was estimated and was found to vary from 0.556 to 1.013 between pair-wise populations and 0.7412 among populations. The results of UPGMA cluster analysis and nonmetric multi-dimensional scaling analysis indicated that most variation occurred within populations and that genetic differentiation had happened between populations. These findings are important for a better understanding of the adaptive strategy of S. grandis in northern China and will be useful for conservation managers to work out an effective strategy to protect this important species.     

Analysis of microsatellite loci variations in herring (Clupea pallasii marisalbi) from the White Sea

The genetic diversity among spawning groups of herring from different parts of the White Sea was assessed using ten microsatellite loci. All loci were polymorphic with the expected heterozygosity estimates varying in the range of 12.7–94.1% (mean was 59.5%). The degree of genetic differentiation displayed by White Sea herring was statistically significant (θ = 2.03%). The level of pairwise genetic differentiation F _ST was 0–0.085, and it was statistically significant in most of the comparison pairs between the herring samples. A hierarchical analysis of molecular variance (AMOVA) revealed the statistically significant differentiation of White Sea herring. 96.59% genetic variation was found within the samples and 3.41% variation was found among the populations. The main component of interpopulation diversity (1.85%) falls at the differences between two ecological forms of herring, spring- and summer-spawning. Within the spring-spawning form, the presence of local stocks in Kandalaksha Bay, Onega Bay, and Dvina Bay was demonstrated.     

Complex genetic structures of Sardinella zunasi in the Northwest Pacific detected by AFLP markers

Seventy-one Sardinella zunasi individuals collected from 5 locations in the Northwest Pacific were used in an AFLP analysis to examine the population genetic structure of S. zunasi. Using 4 selective primer combinations, 414 putative loci were detected, including 340 polymorphic loci (82.13%). Both AMOVA and pairwise F_ST analysis detected significant population genetic structure between Chinese and Japanese populations, supporting separate stocks in this species. The UPGMA trees also detected significant spatial structure within S. zunasi. Pleistocene glaciations are hypothesized to be responsible for the genetic differentiation. The significant genetic differentiation found among S. zunasi populations might also be due to geographic isolation, because a pattern of isolation-by-distance was observed. The weaker but significant structures between the Japanese Aichi and Kagawa populations might be impacted by the semi-enclosed coastal topography there. No significant genetic structure was detected among the Chinese populations, perhaps because of S. zunasi’s migratory behavior and ocean current patterns.     

Comparison of the extent of genetic variation of Vallisneria natans and its sympatric congener V. spinulosa in lakes of the middle-lower reaches of the Yangtze River

Vallisneria natans and Vallisneria spinulosa are two morphologically very similar and sympatrically dominant submerged macrophytes in lakes of the middle-lower reaches of the Yangtze River. Genetic variation was compared based on a total of 196 individuals from six V. natans populations and 201 individuals from seven V. spinulosa populations. Using eight ISSR primers, a total of 139 and 129 DNA fragments were generated with 121 being polymorphic in V. natans and 99 in V. spinulosa. The two species maintained higher genetic variation both at the species and population levels in comparison with other aquatic macrophytes. A higher level of genetic diversity among populations was found in V. natans than in V. spinulosa: the percentage of polymorphic loci (PPL) in V. natans was 52-62% vs. 38-47% in V. spinulosa; gene diversity (H) was 0.21 in V. natans vs. 0.17 in V. spinulosa.Both an analysis of molecular variance (AMOVA) and F-estimation (F_ST) showed that most of the total genetic variation resided within populations of both species (AMOVA: 85% and 80%; F_ST: 0.132 and 0.202), indicating low genetic differentiation between populations. Principal coordinates analysis (PCA) indicated evident gene flow between populations of both species. The outcrossing reproductive mode and pervasive gene flow might have played important roles in maintaining high genetic diversity and in shaping low population differentiation of the two Vallisneria species, while the extent of clonal growth might account for the different levels of population divergence between them.     

Genetic Polymorphism of Microsatellite DNA in Two Populations of Northern Sheatfish (Silurus soldatovi)

In this article, population variations and genetic structures of two populations of northern sheatfish (Silurus soldatovi) were analyzed using 24 microsatellite loci enriched from southern catfish (S. meriaionalis Chen) by magnetic beads. Gene frequency (P), observed heterozygosity (Ho), expected heterozygosity (He), polymorphism information contents (PIC), and number of effective alleles (Ne) were determined. One population was wild, ripe individuals collected from Heilongjiang River (HNS); the other was cultured fry collected from Songhuajiang River (SNS). The Hardy-Weinberg equilibrium (HWE) was tested by the genetic departure index (d). The coefficient of gene differentiation G_ST and Φ_ST by AMOVA (Analysis of Molecular Variety) was imputed using Arlequin software in this study. In addition, a phylogenetic tree was constructed by UPGMA method based on the pairwise Nei's standard distances using PHYLIP. A total of 1 357 fragments with sizes ranging between 102 bp and 385 bp were acquired by PCR amplifications. The average number of alleles of the two populations was 8.875. Results indicated that these microsatellite loci were highly polymorphic and could be used as genetic markers. The mean values of the parameters P, Ho, He, PIC, and Ne were 0.165, 0.435, 0.758, 0.742, and 5.019 for HNS and 0.147, 0.299, 0.847, 0.764, and 5.944 for SNS, respectively. Although there were differences, there were no significant differentiations except for the locus HLJcf37. These populations to a certain extent deviated from HWE, such as excessive and deficient heterozygote numbers. The value of G_ST was 0.078 and above 98% of the variation were differences among individuals within the population, so the variation between populations was insignificant. Cluster analysis also showed that the relationships among individuals were very close. In conclusion, the microsatellite markers that were developed through this study are useful for genetic analysis and the genetic culture that was proposed in this study has no significant impact on S. soldatovi.     

Genetic diversity and population structure of the red stingray,Dasyatis akajei inferred by AFLP marker

To examine population genetic diversity and variation of the red stingray Dasyatis akajei, samples from 1 freshwater region and 6 coastal localities within its distribution range were analyzed by using amplified fragment length polymorphism (AFLP) technology. A total of 207 loci were identified by 4 primer combinations from 87 individuals, 174 of which were polymorphic (84.1%). A high level of Nei's gene diversity was observed with the overall value of 0.230 ± 0.179. No significant genealogical clusters associated with sampling sites were revealed on the UPGMA tree. Both analysis of molecular variance (AMOVA, Φ_ST = 0.085, P = 0.00) and pairwise F_ST indicated significant genetic differentiation among four marine samples. However, no particular genetic differentiation was detected between marine and the limited sampling freshwater populations (AMOVA, Φ_CT = 0.056, P > 0.05). Except for the TZ vs. WZ (5.193), the gene flow estimates (N_m) demonstrated the effective immigrants were 1.918-2.976, suggesting low level dispersal between pairwise marine populations. Species-specific habits (demersal and sluggish habits) are probably responsible for the population structuring.     

Genetic Diversity Within and Among Lepidium draba Populations from Eastern Anatolia Based on RAPD Analysis

Genetic variation and structure of six natural populations of Lepidium draba L. from Eastern Anatolia were assessed using random amplified polymorphic DNA (RAPD) markers. For RAPD analysis, 12 primers generated 218 reproducible bands across the six populations analyzed, of which 73 bands (33.3%) were polymorphic. The mean Nei’s gene diversity value for all six populations was 0.1771. Shannon’s information index varied with population (0.2278–0.3082), averaging 0.2608. Analysis of molecular variance (AMOVA) showed that genetic diversity was greater within populations (58.66%) than among populations (30.68%). In addition, the variation between groups was 10.33%. The genetic differentiation among populations (G _ST) was 0.3210, indicating that most genetic diversity occurs within populations. Gene flow (Nm) was low, at only 0.5288.     

Microsatellite analysis reveals the genetic structure and gene flow of the aquatic quillwort Isoetes sinensis, a critically endangered species in China

Isoetes sinensis is a critically endangered aquatic quillwort, occurring in two fragmented sites of China (Xiuning county of Anhui; Jiande county of Zhejiang). During a five-year period (2004-2009), the areas and sizes of the two populations diminished dramatically due to intensive human activities. Genetic structure of the species was investigated using simple sequence repeat makers (SSRs). For seven nuclear microsatellites, high levels of genetic diversity were found within populations (H_E = 0.324-0.447). Strong genetic differentiation was detected between populations (G_ST = 0.376), while weak genetic differentiation was found within populations (G_ST = 0.026-0.080). Notably, in contrast to the source-sink model suggested by previous study (Chen et al., 2009), the migration pattern of I. sinensis along the Xin’an River is best explained by the linear symmetrical non-adjacent flow model (LSNF), which indicates that intensive human activities of recent years have greatly affected the gene exchange pattern among I. sinensis subpopulations.     

AFLP markers suggest low population genetic differentiation of the black rockfish Sebastes schlegelii

AFLP markers were used to investigate the population genetic differentiation among populations of Sebastes schlegelii from China and Japan. A total of 405 bands were amplified for 180 individuals from 9 populations by 3 pairs of selective primers. S. schlegelii exhibited high Nei's gene diversity with an average value of 0.29 for all populations. No significant genealogical branches or clusters corresponding to sampling localities were detected by UPGMA tree. The results of AMOVA analysis and pairwise F_ST values showed no genetic divergence among different geographic populations. The result of STRUCTURE demonstrated all populations of S. schlegelii examined in the study shared one single gene pool.     

Genetic structure of South Australian Pyrenophora teres populations as revealed by microsatellite analyses

The aim of this study was to determine the genetic structure of South Australian field populations of the barley net blotch pathogens, Pyrenophora teres f. sp. teres (PTT) and P. teres f. sp. maculata (PTM), using microsatellite DNA markers. Three PTT populations (76 isolates total) and two PTM populations (43 isolates total) were sampled from separate fields during a single growing season. The results showed that of the 20 microsatellite loci examined, 17 (85 %) were polymorphic within the PTT and PTM populations. In total, 120 distinct alleles were identified of which only 11 (9 %) were shared between the two population types. Nei’s measure of gene diversity across the PTT and PTM populations was similar at 0.38 and 0.40, respectively, and also much higher than previously reported from studies in which other types of molecular markers were used. The coefficient of genetic differentiation among both populations was the same (G_ST = 0.03) and the low and insignificant estimates of F_ST, as indicated by θ, between populations of the same type (PTT: θ < 0.008, PTM: θ = 0.014) indicated that isolates sampled from different areas within the same field were genetically similar. In contrast, high and significant genetic differentiation was observed among and between populations of different type (G_ST = 0.42, θ > 0.567). The high number of unique multilocus haplotypes observed within the PTT (84 %) and PTM (100 %) populations, combined with a 1:1 distribution of both mating types, suggested that sexual reproduction was predominant among these populations. However, tests for multilocus associations showed that both PTT and PTM populations were in significant linkage disequilibrium. Although the levels of disequilibrium were low, an asexual reproductive component could not be excluded.     

Population genetic structure and conservation implications of Ceriops decandra in Malay Peninsula and North Australia

Comprehensive information of mangrove genetic resources is requisite for developing strategies for their effective conservation and sustainable use. Genetic diversity within and among populations of a widespread viviparous mangrove Ceriops decandra was determined using inter-simple sequence repeat (ISSR). Ten natural populations were collected from Malay Peninsula and North Australia. At the species level, high genetic variation was detected (P = 72%, H_E = 0.253, and I = 0.379). The estimate of G_ST was 0.882, indicating a high level of genetic differentiation among populations. When populations were grouped according to geographic regions, i.e., East Malaya, West Malaya, Southmost Malaya, and North Australia, AMOVA suggested that most of the total variation (87%) was accounted for by differentiation between regions, with only 4% accounting for variation among populations within regions, and a further 9% partitioned among individuals within a population. A UPGMA dendrogram based on genetic distance revealed a deep split between populations from the eastern Indian Ocean and all others from the western Pacific Ocean, which may result from the historical lowering of sea level at these regions during the recent Pleistocene glaciations. An understanding of the genetic structure of C. decandra provides insight for the conservation and management of this species.     

Contrasting population makeup of two intertidal gastropod species that differ in dispersal opportunities

This study of population structure of two intertidal monodontine species: Diloma nigerrima and Diloma subrostrata, revealed the level of genetic connectedness among populations. Despite their markedly different geographic ranges (D. nigerrima is a geographically widespread species, inhabiting both Chile and New Zealand, including its subantarctic islands, whereas D. subrostrata is endemic to New Zealand), both species are believed to possess short-lived lecithotrophic larval stages. Polymorphic DNA microsatellite sequences were used to reveal the level of genetic connectedness among populations, thus inferring the two species' relative effective dispersal abilities. For each species F statistics, AMOVA values and the strength of the relationship between geographic and genetic distance were calculated. We observed a higher within-species level of genetic variation (Φ_ST = 0.099 vs. Φ_ST = 0.016) and a higher proportion of variance (11.15% vs. 0%) among populations of D. nigerrima than of D. subrostrata. A larger fraction of significant F_ST values was observed among D. nigerrima population pairs (65%) than among D. subrostrata population pairs (33%). Significant correlation between genetic and geographic distance was observed for D. nigerrima but not for D. subrostrata, but this relationship was not consistent among pairwise D. nigerrima population comparisons and PCA analysis confirms that, for each species, population structure does not follow a consistent pattern of increasing with geographic distance. The lack of population structure among D. subrostrata populations is probably due to its ubiquitous distribution, meaning little opportunity exists for genetic structure. D. nigerrima, by contrast has a patchier distribution, which allows for greater opportunities for genetic differentiation to occur. We argue that, despite the probable short larval stage in this species, the lack of geographical pattern in the genetic structure found in D. nigerrima is best explained by occasional dispersal over relatively short distances around the coast of New Zealand, over longer distances from New Zealand to the subantarctic islands and even across the Pacific Ocean from New Zealand to Chile.     

High genetic differentiation and variation as revealed by ISSR marker in Pseudotaxus chienii (Taxaceae), an old rare conifer endemic to China

Pseudotaxus chienii (Taxaceae) is an endangered conifer species endemic to China, with a historically disjunct distribution pattern. Eleven populations sampled throughout its range were examined using inter-simple sequence repeats (ISSR) markers. Twenty primers generated 242 bands, and each detected polymorphic loci, with 73.14% of polymorphic loci overall. The estimate for θ^B was 0.5306, whereas G_ST and Φ_ST values were 0.6146 and 0.6401, respectively. 50.62% of the total diversity based on Shannon's index of phenetic diversity was attributed to among populations, which was consistent with the θ^B estimation. Compared with other conifers, remarkable genetic differentiation occurred among P. chienii populations. A Mantel test indicated that pairwise values of θ^B were significantly related with geographical distances between populations (r = 0.676, P = 0.001). In terms of the above results, conservation strategies and maintenance plans of P. chienii were recommended.     

Genetic diversity of Aegiceras corniculatum (Myrsinaceae) revealed by amplified fragment length polymorphism (AFLP)

Aegiceras corniculatum is a cryptoviviparous mangrove tree distributed in the Indo-West Pacific. The genetic structure of 13 populations of A. corniculatum from South China, Malay Peninsula, Sri Lanka, and North Australia, was assessed by amplified fragment length polymorphism (AFLP) markers. Our results showed a relatively high level of genetic variation at the species level (P = 92%, H_E = 0.294 and H_s = 0.331 ± 0.001). The value of G_ST was 0.698, suggesting significant genetic differentiation among populations. At the population level, however, genetic diversity was low (P = 24%, H_E = 0.086 and H_s = 0.127 ± 0.001). When populations were grouped according to geographic regions, i.e., South China, Malay Peninsula and Sri Lanka, it was inferred from analysis of molecular variance (AMOVA) that about half the total variation (49%) was accounted for differentiation between regions. A UPGMA dendrogram based on genetic distance also revealed five major clades corresponding to geographical regions within the distribution of A. corniculatum, although the precise relationships among the clades were not fully concordant with expected geographical delineations and need further study.     

Microsatellite diversity and population genetic structure of yellowcheek, Elopichthys bambusa (Cyprinidae) in the Yangtze River

Yellowcheek carp (Elopichthys bambusa) is the only species of genus Elopichthys. It is widely distributed in Chinese freshwaters but currently its populations have declined to threatening level. We examined the genetic diversity and population structure of E. bambusa in the Yangtze River basin. A total of nine polymorphic microsatellite markers were employed to study five populations occurring in middle and lower reaches of the river. The results revealed low-to-moderate genetic diversity. The number of alleles per locus varied between 3 and 8 with an average of 4.8. Observed heterozygosity ranged from 0.15 to a maximum of 1.00. Significant deviations (P < 0.01) from Hardy–Weinberg equilibrium were observed for all the tested locus-population combinations with clear heterozygosity deficits. AMOVA indicated that majority of the variance lies within populations (93.81%) than among the populations (7.05%). Pairwise F_ST and unbiased genetic distance pointed out significant differentiation among the samples from populations with different connections to the Yangtze River. In the UPGMA dendrogram, clustering pattern of populations indicated that most of the populations are reproductively isolated due to anthropogenic interventions. Clustering of PYL and DTL populations shows ongoing gene flow through the mainstream. The recent hydrological alterations and overfishing are major factors shaping the current genetic structure. These results can be helpful for effective management and sustainable conservation of E. bambusa populations.     

Patterns of Genetic Variation in Swertia przewalskii, an Endangered Endemic Species of the Qinghai-Tibet Plateau

Swertia przewalskii Pissjauk. (Gentianaceae) is a critically endangered and endemic plant of the Qinghai-Tibet Plateau in China. RAPD and ISSR analyses were carried out on a total of 63 individuals to assess the extent of genetic variation in the remaining three populations. Percentage of polymorphic bands was 94% (156 bands) for RAPD and 96% (222 bands) for ISSR. A pairwise distance measure calculated from the RAPD and ISSR data was used as input for analysis of molecular variance (AMOVA). AMOVA indicated that a high proportion of the total genetic variation (52% for RAPD and 56% for ISSR) was found among populations; pairwise Φ _ST comparisons showed that the three populations examined were significantly different (p < 0.001). Significant genetic differentiation was found based on different measures (AMOVA and Hickory θ_B) in S. przewalskii (0.52 on RAPD and 0.56 on ISSR; 0.46 on RAPD and 0.45 on ISSR). The differentiation of the populations corresponded to low average gene flow (0.28 based on RAPD and 0.31 based on ISSR), whereas genetic distance-based clustering and coalescent-based assignment analyses revealed significant genetic isolation among populations. Our results indicate that genetic diversity is independent of population size. We conclude that although sexual reproduction and gene flow between populations of S. przewalskii are very limited, they have preserved high levels of genetic diversity. The main factors responsible for the high level of difference among populations are the isolation and recent fragmentation under human disturbance.     

Genetic structure in the paleoendemic and endangered Petagnaea gussonei (Spreng.) Rauschert (Saniculoideae, Apiaceae) and implications for its conservation

Our investigation aims to understand the genetic structure and evolutionary history of Petagnaea gussonei, an ancient and endangered species belonging to the Saniculoideae subfamily (Apiaceae). It is paleoendemic to Sicily, with a small number of populations in the Nebrodi Mountains. A total of seven chloroplast microsatellite repeat loci and 12 AFLP primer combinations were used to screen 115 individuals corresponding to 17 populations. The ratio of seed to pollen flow was also calculated using the modified Ennos equation. A relatively high level of genetic diversity was detected with AFLPs (e.g., 0.045 < H < 0.278), and a moderate variation was also found using cpSSRs (0 < Hk < 0.667). Two different haplotypes (B and W) were identified, with five populations being monomorphic for haplotype B. There was no genetic differentiation on the basis of haplotypic frequency (G _ST) and similarity (R _ST), and no phylogeographic structure was detected among the populations. AFLP values also confirmed that the populations are not very genetically differentiated. The principal component analysis based on pairwise genetic differences showed three groupings without a geographical correlation. The AMOVA analysis indicates that the amount of variation is higher within populations (82 %) than among populations (18 %). Results of the pollen flow/seed flow ratio indicated positive values for each population, indicating that gene flow by seed is not more efficient than by pollen. Instead, the total pollen/seed flow for all population presents a negative value, suggesting that pollen dispersal does not appear to be more effective over the long range for gene flow than seed dispersal. This differentiation level supports the hypothesis that the fragmentation and isolation of the residual populations is in progress. This phenomenon is due not only to post-ice age climate changes, but also to direct and indirect anthropic actions.