The genetic diversity and spatial genetic structure of the Corso‐Sardinian endemic Ferula arrigonii Bocchieri (Apiaceae)

Corsica and Sardinia represent major hotspots of plant diversity in the Mediterranean area and are priority regions for conservation due to their high number of endemic plant species. However, information supporting human decision‐making on the conservation of these species is still scarce, especially at the genetic level. In this work, the first assessment is reported of the species‐wide spatial genetic structure and diversity of Ferula arrigonii Bocchieri, a Corso‐Sardinian endemic located in a few coastal sites and on small islands. Nine populations covering the entire natural range of the species were investigated by means of AFLP (amplified fragment length polymorphism) markers. Results indicate that this species is characterised by high levels of genetic polymorphism (92% polymorphic fragments) and of genetic diversity (H_w = 0.317) and by relatively low differentiation among populations (F_st = 0.057). PCoA, Bayesian analysis and neighbour‐joining clustering were also employed to investigate the genetic structure of this species. Three genetically distinct groups were detected, although with considerable overlap between populations.     

Genetic diversity and structure of Pinus dabeshanensis revealed by expressed sequence tag-simple sequence repeat (EST-SSR) markers

Assessing patterns of genetic variation in rare endangered species is critical for developing both in situ and ex situ conservation strategies. Pinus dabeshanensis Cheng et Law is an endangered species endemic to the Dabieshan Mountains of eastern China. To obtain fundamental information of genetic diversity, population history, effective population size, and gene flow in this species, we explored patterns of genetic variation of natural populations, in addition to an ex situ conserved population, using expressed sequence tag-simple sequence repeats (EST-SSR) markers. Our results revealed moderate levels of genetic diversity (e.g., H_E = 0.458 vs. H_E = 0.423) and a low level of genetic differentiation (F_ST = 0.028) among natural and conserved populations relative to other conifers. Both contemporary and historical migration rates among populations were high. Bayesian coalescent-based analyses suggested that 3 populations underwent reductions in population size ca. 10,000 yr ago, and that two populations may have experienced recent genetic bottlenecks under the TPM. Bayesian clustering revealed that individuals from the ex situ population were largely assigned to the ‘red’ cluster. Additionally, our results identified private alleles in the natural populations but not in the ex situ population, suggesting that the ex situ conserved population insufficiently represents the genetic diversity present in the species. Past decline in population size is likely to be due to Holocene climate change. Based on the genetic information obtained for P. dabeshanensis, we propose some suggestions for the conservation and efficient management of this endangered species.     

Genetic diversity and population structure of Swertia tetraptera (Gentianaceae), an endemic species of Qinghai-Tibetan Plateau

Swertia tetraptera Maxim is an annual alpine herb endemic to the Qinghai-Tibetan Plateau (QTP). Its populations are locally scattered as isolated patches throughout this region. Genetic variation within and among thirty-four populations of this species was assessed using ISSR fingerprinting with 10 primers. High levels of genetic diversity exist within species (P = 98.9%, I = 0.3475; He = 0.2227), while the within-population diversity is low (P = 32.7%, I = 0.177; He = 0.12). High levels of genetic differentiation were detected among populations based on various statistics, including Nei’s genetic diversity analysis (G_ST = 0.4608), Bayesian analysis (θ^B = 0.476) and AMOVA (F_ST = 0.57). That is, populations shared low levels of genetic identity (I = 0.2622–0.0966). This genetic structure was probably due to severe genetic drift, breeding system and limited gene flow. The observed genetic structure of the populations implies that different populations across the distribution range of the species should be sampled to maintain high genetic diversity when a conservation strategy is implemented.     

Low mtDNA Cytb diversity and shallow population structure of Eleutheronema tetradactylum in the East China Sea and the South China Sea

Eleutheronema tetradactylum is an economically important fish species in China water. To investigate the genetic diversity and describe population structure of it, an 1151 base pair (bp) fragment of the mitochondrial DNA Cytb sequence was analyzed in 120 individuals from four populations in the East China Sea and the South China Sea. A total of 16 haplotypes were defined by 24 variable nucleotide sites. High level of haplotype diversity and low nucleotide diversity were observed in all populations. The results of AMOVA detected that 89.44% of the genetic variation occurred within populations. Significant genetic differentiations were detected among populations (0.05097, P < 0.05), but no large-scale regional differences were detected. Analysis of neutral evolution and mismatch distribution suggested no recent population expansion happened. The present results provided new information for genetic assessment, fishery management and conservation of this species.     

Genetic diversity and structure of the noxious alien grass Praxelis clematidea in southern China

Praxelis clematidea (Asteraceae), a plant species native to South America, is a noxious weed in southern China. We examined the genetic variation and population structure of 12 populations (76 individuals) of P. clematidea from Fujian, Guangdong, and Hainan Provinces in China using inter-simple sequence repeat (ISSR) analysis. From an initial set of 69 ISSR primers, 10 were selected which yielded 80 reproducible bands. Polymorphic bands (P) were 100%, Shannon's information index (I) was 0.4226, and Nei's gene diversity (H) was 0.2791. We infer that the high levels of genetic diversity exhibited by P. clematidea may have contributed to its invasiveness. Gene flow among populations was 2.4930, which has led to homogenization. The coefficient of population differentiation (Gst = 0.1671) indicated low levels of genetic variation among populations and high levels of genetic polymorphism within populations. There was a negative correlation between population elevation and genetic diversity, while there was a significant positive correlation between genetic distance and geographic distance based on a Mantel test (r = 0.5820, P < 0.01). Some populations from different provinces clustered together in principal coordinate and UPGMA analyses indicating that human-mediated events may have contributed to the dispersal of the species.     

Genetic diversity of the endangered Chinese endemic plant Monimopetalum chinense revealed by amplified fragment length polymorphism (AFLP)

Monimopetalum chinense Rehd. is an endangered woody vine endemic to eastern China. Using amplified fragment length polymorphism (AFLP) markers, we examined levels of genetic variation within and among eleven populations located across the species’ distribution. Although modest levels of heterozygosity were detected, other measures of genetic diversity registered relatively high levels of variability, both at the species level (P = 91.0%, H_E = 0.232, I_S = 0.365) and at the population level (P = 53.0%, H_E = 0.155, I_S = 0.239). Populations also exhibited high levels of genetic differentiation (Nei’s genetic diversity analysis, G_ST = 0.330), corresponding to isolation-by-distance and hierarchical population structure. These results indicate that, despite low levels of gene flow, populations of M. chinense still harbor substantial amounts of genetic diversity. Management plans for the species should include measures that ensure genetic diversity remains high within and among extant populations.     

Genetic diversity and population structure of endangered endemic Paeonia jishanensis in China and conservation implications

Paeonia jishanensis, one of the most important ancestral species of cultivated tree peonies, is an endangered ornamental and medicinal plant endemic to China. A total of 236 individuals of P. jishanensis from 10 extant populations were analysed using 21 EST-SSR markers to assess their genetic diversity and population structure. Moderate genetic diversity levels (H_E = 0.340) and high genetic differentiation among populations (F_ST = 0.335) were revealed. Combining the results of STRUCTURE, PCoA and neighbour-joining analyses, the 10 populations were divided into four genetic groups that were significantly related to their geographical origins, which was further supported by hierarchical AMOVA showing the highest variation of 17.9% among groups. The Mantel test showed a significant positive correlation between genetic distance and geographic distance among populations (r = 0.873, P < 0.0001). The genetic structure of P. jishanensis may be due to limited gene flow hindered by vicariance and its breeding system: facultative vegetative reproduction. This study carries significant implications for the conservation and utilization of this endangered species.     

Low genetic diversity and significant structuring in the endangered Mentha cervina populations and its implications for conservation

Eighteen populations of the endangered aromatic and medicinal plant Mentha cervina (Lamiaceae) were sampled across its natural range, in the western half of the Iberian Peninsula, and inter-simple sequence repeats (ISSRs) markers were used to assess genetic diversity and population structure. M. cervina populations exhibited a relatively low genetic diversity (percentage of polymorphic loci PPB = 14.2–58.3%, Nei's genetic diversity H_e = 0.135–0.205, Shannon's information index I = 0.08 − 0.33). However, the genetic diversity at species level was relatively high (PPB = 98.3%; H_e = 0.325; I = 0.23). The results of the analysis of molecular variance indicated very structured populations, with 50% of the variance within populations, 44% among populations and 6% between regions defined by hydrographic basins, in line with the gene differentiation coefficient (G_ST = 0.532). A Mantel test did not find significant correlation between genetic and geographic distance matrices (r = 0.064), indicating that isolation by distance is not shaping the present genetic structure. The levels and patterns of genetic diversity in M. cervina populations were assumed to result largely from a combination of evolutionary history and its unique biological traits, such as breeding system, low capacity of dispersion, small effective size and habitat fragmentation. The high genetic differentiation among populations indicates the necessity of conserving the maximum possible number of populations. The results also provide information to select sites for ex situ conservation. Optimal harvesting strategies, cultivation and tissue culture should also be developed as soon as possible to guarantee sustainable use of the species under 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.     

Genetic diversity and structure of natural Pinus tabulaeformis populations in North China using amplified fragment length polymorphism (AFLP)

Chinese pine (Pinus tabulaeformis carr.), endemic to China, is a conifer species with extensive and fragmented distribution in North China. In this study, the genetic diversity and structure of 20 natural populations of this species were investigated using amplified fragment length polymorphism (AFLP) markers. A total of 445 fragments were revealed with 8 pairs of primers, 379 (85.17%) of which were polymorphic. A moderate level of genetic diversity was detected at the species level (Shannon's information index I = 0.356, Nei's gene diversity H_E = 0.271) and at the population level (I = 0.219, H_E = 0.206). Most of genetic variation was within populations while a considerable level of genetic differentiation was detected (G_ST = 0.352, Ф_ST = 0.304). The high differentiation could be attributed to the complex and fragmented habitats, and a limited gene flow among populations (N_m = 0.572). The Mantel test indicated that there was significant correlation (r = 0.455, P < 0.001) between Nei's genetic distance and geographical distance among all the populations. The results suggested that proper countermeasures should be taken to prevent the habitat further deterioration and maintain the genetic diversity of this species.     

Genetic variation and clonal diversity in populations of Nelumbo nucifera (Nelumbonaceae) in central China detected by ISSR markers

To obtain accurate estimates of population structure for purposes of conservation planning for wild lotus (Nelumbo nucifera Gaertn.) in central China, genetic diversity among and within six populations, and clonal diversity within another two populations of the species were analyzed. The genetic diversity was high (percentage of polymorphic bands, PPB = 90.0%; Shannon's information index, I = 0.383 ± 0.234) at the species level, but low within individual study populations (PPB = 35.8%; Shannon's information index I = 0.165 ± 0.241). The mean coefficient of gene differentiation (G_st) was 0.570, indicating that 43.0% of the genetic diversity resided within the population. Analysis of molecular variance (AMOVA) indicated that 50.47% of the genetic diversity among the study populations was attributed to geographical location while 12.3% was attributed to differences in their habitats. An overall value of mean estimated number of gene flow (N_m = 0.377) indicated that there was limited gene flow among the sampled populations. The level of clonal diversity found within the populations was considerably high (Simpson's diversity index, D = 0.985) indicating that clonal diversity contributes to a major extent to the overall genetic variation in the genetic structure of N. nucifera. On the basis of the high G_st and D values detected in this study we recommend that any future conservation plans for this species should be specifically designed to include those representative populations with the highest genetic variation for both in situ conservation and germplasm collection expeditions.     

Genetic variability of the narrow endemic Rhamnus persicifolia Moris (Rhamnaceae) and its implications for conservation

Rhamnus persicifolia Moris is an endemic small tree belonging to the Rhamnus cathartica group, growing along mountainous streams of Central-Eastern Sardinia (Italy). ISSR markers were used to detect the genetic diversity within and among six populations representative of the species distribution range. In spite of the limited distribution of this endemic taxon, fairly high levels of genetic diversity were detected. Percentage of polymorphic bands (PPB), gene diversity (H_S and H_T) and Shannon information measure (Sh) were calculated both at population (PPB = 30.70%, H_S = 0.1105, Sh = 0.1646) and at species level (PPB = 68.42%, H_T = 0.2066, Sh = 0.3139).The existence of a spatial distribution of genetic diversity in R. persicifolia was revealed by a low gene flow, a fairly high level of genetic differentiation (G_ST = 0.4583) among populations and a positive correlation between genetic and geographic distances (Mantel test, r = 0.71, p = 0.016). The spatial genetic structure was also confirmed with BAPS analysis. Our results show that a certain level of isolation by distance and sex-ratio bias may explain the distribution of genetic diversity among populations.Conservation measures are suggested on the basis of the genetic diversity detected, by implementing an integrated in situ and ex situ conservation program for each population, in order to ensure effective protection for this endemic species.     

Assessing the genetic diversity and population structure of Culter alburnus in China based on mitochondrial 16S rRNA and COI gene sequences

The genetic diversity and population genetic structure of Culter alburnus were investigated using mitochondrial cytochrome c oxidase subunit I (COI) and ribosomal 16S subunit (16S rRNA) gene sequences. A total of 89 individuals from four localities were included in the analysis. Overall, 12 polymorphic sites were observed and 10 haplotypes were defined. The C. alburnus populations were characterized by high haplotype diversity (0.587 ± 0.047) and low nucleotide diversity (0.00197 ± 0.00073). Pairwise fixation index (F_ST) analysis indicated significant genetic differentiation among different populations. The hierarchical analysis of molecular variance (AMOVA) analysis also showed significant genetic divergence (φ_ST = 0.3792, P < 0.01) among these populations. The present results suggest that subdivisions exist among four C. alburnus populations, and should be considered as different management unit for effective conservation and management purposes. This study provides new information for genetic assessment and will be crucial for establishing fisheries management and strategies for this species.     

Genetic diversity of natural populations of Larix principis-rupprechtii in Shanxi Province,China

Prince Rupprecht's Larch (Larix principis-rupprechtii Mayr.) is one of dominant components of middle and high elevation forests in North China. Shanxi Province is well known as “the Hometown of Prince Rupprecht's Larch” in China. In this study, six natural populations of this species across Shanxi were selected to investigate the genetic variation of the species using amplified fragment length polymorphism (AFLP) markers. Results showed that in comparison with some other species of Larix, higher genetic diversity was revealed at the species level for L. principis-rupprechtii (percentage of polymorphic loci PPL = 71.9%, Nei's gene diversity H_E = 0.225, Shannon information index I = 0.341). Most of genetic variation existed within populations (80.5%), while the genetic differentiation among populations was significant (p < 0.001) and higher (G_st = 0.194) than most other species of Larix. The differentiation can be attributed to the limited gene flow (N_m = 1.035) among populations, which could be due to the spatial isolation and habitat fragmentation. The six populations can be divided into three groups based on the Nei's genetic distances between populations (from 0.033 to 0.076). There was no significant correlation (r = 0.268, p > 0.05) between genetic distance and geographic distance among populations. The measures for in-situ or ex-situ conservation should be taken to preserve the genetic diversity of this species.     

Genetic diversity analysis and conservation of the endangered Chinese endemic herb Dendrobium officinale Kimura et Migo (Orchidaceae) based on AFLP

Dendrobium officinale is a critically endangered perennial herb endemic to China. Determining the levels of genetic diversity and patterns of population genetic structure of this species would assist in its conservation and management. Data of 12 populations were used to assess its genetic diversity and population structure, employing the method of amplified fragment length polymorphism (AFLP). A high level of genetic diversity was detected (H (E) = 0.269) with POPGENE. As revealed by AMOVA analysis, there was moderate variation between pairs of populations with Phi(ST) values ranging from 0.047 to 0.578 and on average 26.97% of the genetic variation occurred among populations. Three main clusters were shown in UPGMA dendrogram using TFPGA, which is consistent with the result of principal coordinate ananlysis (PCO) using NTSYS. Keeping a stable environment is critical for the in situ conservation and management of this rare and endangered plant, and for ex situ conservation it is important to design an integrated germplasm bank.     

Mitochondrial genetic diversity and population structure of a vulnerable freshwater fish,rock carp (Procypris rabaudi) in upper Yangtze River drainage

Rock carp (Procypris rabaudi (Tchang)) is an endemic species in the upper Yangtze River drainage. The genetic variability and population structure of eight populations (Mudong (MD), Hejiang (HJ), Wanzhou (WZ), Xishui (XS), Nantan (NT), Cangxi (CX), Tongjiang (TJ) and Wulong (WL)) were investigated based on mitochondrial DNA control region. According to the river dimensions, MD, HJ, WZ, WL and CX were defined as large river populations, while TJ, XS and NT were defined as small. The total genetic diversity level of rock carp was moderate (h = 0.858, π = 0.0079). Genetic differentiations among populations in small rivers were greater than those among populations in large rivers, and small rivers showed significant differentiation from large rivers. Analysis from the coalescent-based method showed that there were asymmetric gene flows among four large river populations (CX, HJ, MD and WZ). Demographic analyses suggested that the species as a whole and the WZ population experienced population expansion, while the HJ, NT, and MD populations experienced bottlenecks. The analysis of genetic diversity and population structure suggested that the WL, TJ, CX, NT and XS populations should be protected and managed separately, and that the HJ, MD and WZ populations should be considered as a genetic management unit.     

Lack of genetic structure in endangered large yellow croaker Larimichthys crocea from China inferred from mitochondrial control region sequence data

The large yellow croaker Larimichthys crocea is an important commercial marine fish species in China. However, information about the population structure of this species is limited. In the present study, mitochondrial DNA (mtDNA) control region was sequenced from four populations of the yellow croaker in the southern Yellow Sea, East China Sea and South China Sea to investigate the genetic structure of this species. A total of 54 haplotypes were detected among 62 individuals of large yellow croaker. High levels of population genetic diversity were observed. Among the four populations, the haplotype diversity was between 0.9895 ± 0.0193 (Xiamen) and 1.0000 ± 0.0524 (Lvsi, Zhoushan). The nucleotide diversity ranged from 0.0208 ± 0.0108 (Xuwen) to 0.0246 ± 0.0138 (Lvsi). The results of AMOVA detected no significant differences among populations. The conventional F_ST statistics were negative and insignificant values. These indicated lack of population genetic structure throughout the Yellow Sea, East China Sea and South China Sea, and random mixing of individuals among the samples. Biological characters of large yellow croaker and lack of physical barrier in the studied area could be the reasons for lack of genetic structure in this species.     

Genetic diversity and population structure of the endemic Azorean juniper,Juniperus brevifolia (Seub.) Antoine,inferred from SSRs and ISSR markers

Juniperus brevifolia is an important woody species endemic of the Azores archipelago (Portugal), found from coastal to mountain environments. Due to colonization and grazing pressure this species has suffered fragmentation, leading to extinction in one island and being threatened in others. The genetic diversity and population structure of J. brevifolia populations was studied to provide guidelines for restoration and conservation programmes. Nuclear Single Sequence Repeats (nSSR) from Juniperus communis and Juniperus przewalskii and Inter Simple Sequence Repeats (ISSR) were tested and results compared to test the transferability of the microsatellites to J. brevifolia. Samples from ten populations over three islands, divided as coastal, mid-altitude and mountain were analysed. Both marker systems revealed results statistically and strongly correlated with each other, and not dependent on population sample size. We observed positive fixation indexes, moderate to high levels of genetic diversity (h = 0.415 for nSSR and h = 0.245 for ISSR), low to moderate φ_pt genetic differentiation (0.070 for nSSR and 0.129 for ISSR) and high gene flow (N_m > 2.432). Regarding Nei's genetic distance the coastal communities clustered together reflecting the phenotypic plasticity, but no specific clustering was observed regarding φ_pt values. Therefore no populations with substantial genetic differentiation were identified, once the diversity is mostly observed within populations. However it is advised the continuous monitoring of J. brevifolia.     

Genetic diversity of the traditional Chinese medicinal plant Ypsilandra thibetica (Melanthiaceae): Applications for conservation

Twelve microsatellite markers were developed to determine the genetic diversity and genetic structure of Ypsilandra thibetica, represented by a total of 90 individuals from six natural populations. All twelve microsatellite loci were polymorphic, and the results indicated that a high genetic diversity was present within populations (mean R_S = 4.996; mean H_E = 0.615), with high levels of genetic structure (mean F_ST = 0.165; mean F_IS = 0.692) among populations. This pattern is likely attributable to consanguineous mating, and this hypothesis is supported by a low relatedness coefficient. Our study suggested that environment factors might restrict gene flow among populations. In addition, physical distances between populations were not related to genetic distances, implying that ancestral populations might have been distributed over a wider area. These results suggest that Y. thibetica should be a high priority for conservation managers.     

Genetic diversity of endangered Manglietia patungensis assessed by inter simple sequence repeat and sequence-related amplified polymorphism markers

Manglietia patungensis Hu is an endangered plant native to China. Knowledge of its genetic diversity and structure would aid its conservation. This study assessed nine natural populations of M. patungensis using two methods: inter simple sequence repeat (ISSR) and sequence-related amplified polymorphism (SRAP) markers. Using 10 ISSR primer pairs, 334 bands were generated, and 10 SRAP primer pairs generated 276 bands. The percent of polymorphic bands (91.32% and 93.48%), Nei's genetic diversity (0.3448 and 0.3323), and Shannon's information index (0.5075 and 0.4935) revealed a high level of genetic diversity at the species level. Total heterozygosity was 0.3439 by ISSR and 0.3281 by SRAP. The mean heterozygosity was 0.2323 by ISSR and 0.2521 by SRAP. The coefficient of genetic differentiation among natural populations was 0.3245 by ISSR and 0.2316 by SRAP. These data indicated higher levels of genetic diversity of M. patungensis within, rather than among, populations. Estimates of gene flow among natural populations were 1.0411 and 1.0589, which implied a certain amount of gene exchange among populations. A Mantel test revealed no significant correlation between genetic and geographic distance. ISSR and SRAP markers are both effective for genetic diversity research in M. Patungensis. Based on these results, conservation of M. patungensis should be performed both in situ and ex situ.