Analysis of genetic variation in grass carp (Ctenopharyngodon idellus) from native and colonized regions using ISSR markers

The grass carp (Ctenopharyngodon idella), a freshwater species native to China, have been introduced in more than 100 countries. In this paper, inter simple sequence repeat (ISSR) markers were used to evaluate genetic variation within and among grass carp populations sampled from its native (China) and colonized habitats (USA, Hungary, and Japan). Twenty-two ISSR primers were used to generate 207 bands, of which 82 (39.61 %) were polymorphic. The highest genetic diversity was observed in the Yangtze River population, whereas the lowest diversity was found in the Tone River population (Japan). Compared with colonized populations, the native populations possess approximately twice as much genetic diversity. The AMOVA analysis showed a relatively high level of genetic variation within populations. Significant genetic differences were revealed both among the native populations and between pooled native and colonized populations.     

RPB2 sequences reveal a close phylogenetic relationship between tetraploid Hordelymus and diploid Hordeum species in Triticeae (Poaceae)

The origin of Hordelymus genome has been debated for years, and no consensus conclusion was reached. In this study, we sequenced and analyzed the RPB2 (RNA polymerase subunit II) gene from Hordelymus europaeus (L.) Harz, and its potential diploid ancestor species those were suggested in previous studies. The focus of this study was to examine the phylogenetic relationship of Hordelymus genomes with its potential donor Hordeum, Psathyrostachys, and Taeniatherum species. Two distinguishable copies of sequences were obtained from H. europaeus. The obvious difference between the two copies of sequences is a 24 bp indel (insertion/deletion). Phylogenetic analysis showed a strong affinity between Hordeum genome and Hordelymus with 85% bootstrap support. These results suggested that one genome in tetraploid H. europaeus closely related to the genome in Hordeum species. Another genome in H. europaeus is sister to the genomes in Triticeae species examined here, which corresponds well with the recently published EF-G data. No obvious relationship was found between Hordelymus and either Ta genome donor, Taeniatherum caput-medusae or Ns genome donor, Psathyrostachys juncea. Our data does not support the presence of Ta and Ns genome in H. europaeus, and further confirms that H. europaeus is allopolyploid.     

Low genetic variation detected within the widespread mangrove species Nypa fruticans (Palmae) from Southeast Asia

Genetic diversity within and among six natural populations of Nypa fruticans from China, Vietnam, and Thailand was assessed using SSR and ISSR analysis. Our results showed an extremely low level of genetic diversity of N. fruticans (at the species level, P = 11.76% and 2.88%, He = 0.0279 and 0.0113, I = 0.0470 and 0.0167 by SSRs and ISSRs, respectively) across a total of 183 individuals. No genetic variation was detected within any population except for the Thailand population by SSRs (P = 11.76%, He = 0.0417; I = 0.0622). The bottlenecks during glacial epochs, founder effects, and propagation pattern may be responsible for the extremely low level of genetic diversity of N. fruticans.     

Genetic diversity of the endangered species Kirengeshoma palmata (Saxifragaceae) in China

In order to investigate the levels of genetic diversity of the endangered species Kirengeshoma palmata (Saxifragaceae), four extant populations were sampled and analyzed using inter-simple sequence repeats (ISSR) markers. We expected a low genetic diversity level, but our results revealed a high level of intraspecific genetic diversity, probably resulting from this species being in a refuge during the last glaciation (at population level: P = 63.25%, A_e = 1.47, H_E = 0.26 and H_O = 0.37; at species level: P = 79.00%, A = 1.5538, H_T = 0.2586 and H_sp = 0.3104). A low level of genetic differentiation among populations was detected based on Nei's genetic diversity analysis (16.69%) and AMOVA (19.36%). Populations shared high levels of genetic identity. Insect pollination and seed dispersal by wind may have facilitated extensive gene flow and are likely responsible for this present structure of genetic variation.     

Genetic diversity and geographical differentiation of Dipteronia Oliv. (Aceraceae) endemic to China as revealed by AFLP analysis

The genus Dipteronia Oliv. endemic to central and southern China consists of two species, Dipteronia sinensis Oliv. and Dipteronia dyeriana Henry, both of them are rare and endangered. AFLP markers were used to characterize the genetic diversity and geographical differentiation of the genus. Eight out of 32 PstI + 3/MseI + 3 selective primer combinations screened were applied to the analysis on 142 individuals of 17 D. sinensis and 4 D. dyeriana populations, respectively. A total of 324 fragments with 316 polymorphic were amplified. The proportion of polymorphic loci (PPB) was 97.53%. The Nei's gene diversity in D. sinensis and D. dyeriana was 0.3319 and 0.3047, respectively. About 43.6% (G_ST = 0.4356) of the genetic variation occurred among the populations, indicating a relatively high genetic differentiation among the populations. Cluster analysis grouped the 21 populations into two groups according to their species delimitation. The populations of D. sinensis were further divided into three subgroups corresponding to their geographical distributions. Correlation analysis revealed a significant correlation (p < 0.05) between geographical distance and genetic distance of these populations, suggesting that the relatively high genetic differentiation among the populations of D. sinensis might be caused by geographical isolation.     

Genetic variation in populations of the threatened seagrass Halophila beccarii (Hydrocharitaceae)

Halophila beccarii (Hydrocharitaceae), a small monoecious seagrass, has been listed as a threatened species. In this study, a total of 106 samples were collected from four Chinese populations located at the northern limit of its distribution range. Using six polymorphic microsatellites, we found low genetic variation in this species, in which the mean number of alleles per locus was 2.8, and 16 multi-locus genotypes were revealed. In the four populations, the mean number of alleles per locus ranged from 1.2 to 2, one to eight genotypes were found, and clonal diversity ranged from 0 to 0.23; observed and expected heterozygosity ranged from 0.17 to 0.40 and from 0.09 to 0.24, respectively. Strong genetic differentiation was found among the populations, and the standardized fixation index (F′_ST) was 0.787. Species traits (i.e., clonal growth) and bottleneck effects due to drastic population fluctuation may contribute to the observed low genetic variation.     

Genetic variation within the endangered mangrove species Sonneratia paracaseolaris (Sonneratiaceae) in China detected by inter-simple sequence repeats analysis

Sonneratia paracaseolaris, is a critically endangered mangrove species in China. Using inter-simple sequence repeats (ISSR) markers, we compared the genetic variation of introduced populations with that of natural populations to check whether the genetic diversity has been conserved. At the species level, genetic diversity was relatively high (P = 81.37%, He = 0.2241, and SI = 0.3501). Genetic variation in introduced populations (P = 75.78%, He = 0.2291, and SI = 0.3500) was more than that in natural populations (P = 70.81%, He = 0.1903, and SI = 0.2980). Based on Nei's G_ST value, more genetic differentiation among natural populations was detected (G_ST = 0.3591). Our data show that the genetic diversity of S. paracaseolaris was conserved in introduced populations to some extent, however, owing to the small natural populations and the threats they encountered, more plants should be planted to enlarge and restore the populations.     

Genetic characterisation of uninucleated cyst-producing Entamoeba spp. from ruminants

Six ssrRNA gene sequences were obtained by PCR amplification of DNA from uninucleated Entamoeba cysts isolated from fresh faeces of sheep, cows, a roe deer and a reindeer. Phylogenetic analysis using sequences of non-, uni-, quadri- and octonucleate cyst-producing Entamoeba spp. for comparison showed that all six isolates formed a separate clade nested within the clade of quadrinucleate cyst producers. The data indicate that Entamoeba bovis can be isolated from ruminant hosts other than cattle, and we suggest that organisms clustering with the sheep and cattle isolates analysed in the present study be named E. bovis.     

Conservation genetics of Heritiera littoralis (Sterculiaceae), a threatened mangrove in China,based on AFLP and ISSR markers

AFLP and ISSR markers were used to determine the genetic variations in eight mangrove and non-mangrove populations of Heritiera littoralis (Sterculiaceae), a threatened species in China. Our results showed a moderate to high level of genetic variation in this species (P = 63.69%, H_T = 0.20 for AFLP; P = 76.07%, H_T = 0.22 for ISSR), and a relatively high level of genetic differentiation among populations (G_ST = 0.24 for AFLP and 0.27 for ISSR). Life history traits, long-distance dispersal of floating seeds, and local environments may play important roles in shaping the genetic diversity and genetic structure of this species. Investigation of the plant’s reproductive capacity showed that the natural seed production of H. littoralis was low, as was the germination rate and the transformation rate from juvenile to adult. H. littoralis is seriously threatened and is in urgent need of conservation in China.     

Genetic diversity of wild populations of Rheum tanguticum endemic to China as revealed by ISSR analysis

Rheum tanguticum is an important but endangered traditional Chinese medicine endemic to China. The wild resources have been declining. Establishing the genetic diversity of the species would assist in its conservation and breeding program. Inter-simple sequence repeats (ISSR) markers were used to assess the genetic diversity and population genetic structure in 13 wild populations of R. tanguticum from Qinghai Province. Thirteen selected primers produced 329 discernible bands, with 326 (92.94%) being polymorphic, indicating high genetic diversity at the species level. The Nei's gene diversity (He) was estimated to be 0.1724 within populations (range 0.1026–0.2104), and 0.2689 at the species level. Analysis of molecular variance (AMOVA) showed that the genetic variation was found mainly within populations (71.02%), but variance among populations was only 28.98%. In addition, Nei's differentiation coefficients (G_st) was found to be high (0.3585), confirming the relatively high level of genetic differentiation among populations. Mantel test revealed a significant correlation between genetic and geographic distances (r = 0.573, P = 0.002), and the unweighted pair-group method using arithmetic average (UPGMA) clustering and Principal coordinates analysis (PCoA) demonstrated similar results. Meanwhile, the genetic diversity of R. tanguticum positively correlated with altitude and annual mean precipitation, but negatively correlated with latitude and annual mean temperature. This result might be an explanation that the natural distribution of R. tanguticum is limited to alpine cold areas. We propose conservation strategy and breeding program for this plant.     

Lack of genetic variation of an invasive clonal plant Eichhornia crassipes in China revealed by RAPD and ISSR markers

Random amplified polymorphic DNAs (RAPDs) and inter-simple sequence repeats (ISSRs) markers were used to analyze genetic structure of six populations of invasive plant Eichhornia crassipes that were sampled from its introduced regions in Southern China. Using 25 RAPD primers and 18 ISSR primers, 172 RAPD bands and 145 ISSR bands were produced respectively. But no polymorphic band was detected either within population or among populations by both markers, indicating the genetic diversity of E. crassipes in Southern China is extremely low, and all populations most likely consist of the same genotype. This study suggested that some other adaptability related factors, other than the genetic diversity, are responsible to the E. crassipes rapid expansion in China.     

Population genetic structure of an aquatic herb Batrachium bungei (Ranuculaceae) in the Hengduan Mountains of China

Inter-simple sequence repeat (ISSR) markers were used to investigate the genetic variation of 14 populations of Batrachium bungei (Ranunculaceae) from the Hengduan Mountains of China, which is a global biodiversity region that is poorly studied. A total of 75 bands with 52 polymorphic loci were generated from the ten primers used in this study. A total of 155 different genotypes or clones were identified among the 442 samples. A low level of genetic diversity within population and high genetic differentiation among populations were revealed in this study. Factors, such as inbreeding, clonal growth, bottlenecks, population isolation and founder events during postglacial recolonizations, might have played important roles in shaping the population structure of B. bungei.     

Genetic diversity and phylogeny in Hystrix (Poaceae, Triticeae) and related genera inferred from Giemsa C-banded karyotypes

The phylogenetic relationships of 15 taxa from Hystrix and the related genera Leymus (NsXm), Elymus (StH), Pseudoroegneria (St), Hordeum (H), Psathyrostachys (Ns), and Thinopyrum (E) were examined by using the Giemsa C-banded karyotype. The Hy. patula C-banding pattern was similar to those of Elymus species, whereas C-banding patterns of the other Hystrix species were similar to those of Leymus species. The results suggest high genetic diversity within Hystrix, and support treating Hy. patula as E. hystrix L., and transferring Hy. coreana, Hy. duthiei ssp. duthiei and Hy. duthiei ssp. longearistata to the genus Leymus. On comparing C-banding patterns of Elymus species with their diploid ancestors (Pseudoroegneria and Hordeum), there are indications that certain chromosomal re-arrangements had previously occurred in the St and H genomes. Furthermore, a comparison of the C-banding patterns of the Hystrix and Leymus species with the potential diploid progenitors (Psathyrostachys and Thinopyrum) suggests that Hy. coreana and some Leymus species are closely related to the Ns genome of Psathyrostachys, whereas Hy. duthiei ssp. duthiei, Hy. duthiei ssp. longearistata and some of the Leymus species have a close relationship with the E genome. The results suggest a multiple origin of the polyploid genera Hystrix and Leymus.     

Molecular phylogeny and maternal progenitor implication in the genus Kengyilia (Triticeae: Poaceae): Evidence from COXII intron sequences

Kengyilia is a perennial genus distributing in central and western Asia. Here, the levels of nucleotide diversity for COXII intron were obtained. The estimates of nucleotide diversity for different genome constitution ranged from θ = 0.00082 and π = 0.00082 for St genome species to π = 0.01227 and θ = 0.01229 for P genome species. Employing COXII intron sequences, the phylogenetic relationships within Kengyilia and between Kengyilia genus and its closely related genera were examined. The Maximum Parsimony analysis demonstrated that Kengyilia species were positioned into two clades corresponding to different maternal genomic donor. Kengyilia stenachyra, Kengyilia grandiglumis, Kengyilia hirsuta, Kengyilia melanthera, Kengyilia thoroldiana, Kengyilia alatavica and Kengyilia zhaosuensis were related to species of Agropyron, while Kengyilia kokonorica, Kengyilia rigidula, Kengyilia nana, Kengyilia mutica, Kengyilia longiglumis, Kengyilia laxiflora and Kengyilia gobicola were close to species of Roegneria and Pseudoroegneria. In addition, other three species of Kengyilia, such as Kengyilia batalinii, Kengyilia tahelacana and Kengyilia kaschgarica, were related to Douglasdeweya deweyi, Pseudoroegneria strigosa and Roegneria tibetica. This result indicated that there had been two phylogenetically divergent maternal donors within Kengyilia. Our new finding will help to understand the evolutionary history of the genus Kengyilia.     

Estimation of genetic variation in Cynodon dactylon accessions using the ISSR technique

Genetic variation in 55 accessions of Cynodon dactylon was estimated using inter-simple sequence repeat (ISSR) markers. The plant materials used in this study originated from 17 countries. A total of 236 ISSR fragments were generated with 14 primers. Fragment sizes ranged from 200 to 3000 bp. All scorable bands were polymorphic in nature and none of the primers used produced monomorphic bands, indicating a high level of genetic variation in this grass. The accessions were found to be clustered into eight major groups through the unweighted pair-group method with arithmetic averages. Genetic similarity coefficients (GSC) among the 55 accessions ranged from 0.52 to 0.95. The results clearly indicate that a high level of variation exists in Cynodon accessions. This study shows that the ISSR technique is a reliable tool for differentiating Cynodon accessions and for determining the genetic relationships among them.     

Phylogenetic relationships among Southeast Asian climbing bamboos (Poaceae: Bambusoideae) and the Bambusa complex

A phylogenetic analysis of Bambusa and allies based on the plastid DNA non-coding regions rps16-trnQ, trnC-rpoB, trnH-psbA and trnD-T, and a partial nuclear GBSSI gene, was carried out. This included representatives from all four Bambusa subgenera (including type species), a group of segregate Southeast Asian genera distinctive by their climbing–scrambling culms (Dinochloa, Holttumochloa, Kinabaluchloa, Maclurochloa, Soejatmia, Sphaerobambos), and two other Bambusinae genera (Dendrocalamus, Gigantochloa). The results do not support the present subgeneric classification of Bambusa. The climbing Southeast Asian genera, all of which include species previously placed in Bambusa, are distinct from the “core Bambusa group” (type species and alliance) and the Bambusa complex generally.     

Genetic diversity analysis and transferability of cereal EST-SSR markers to orchardgrass (Dactylis glomerata L.)

The diversity and genetic relationships among 74 orchardgrass accessions were analyzed using cereal EST-SSRs and orchardgrass SSR markers in order to estimate genetic variability and compare the level of diversity. In total, 190 polymorphic bands were detected with an average of 6.3 alleles per SSR loci. The average polymorphic rate (P) for the species was 84.63%, suggesting a high degree of genetic diversity. The molecular variance analysis (AMOVA) showed that the proportion of variance explained by within- and among-geographical groups diversity was74.87% and 25.13%, respectively. The distinct geographical divergence of orchardgrass was revealed between Americas and Oceania. The ecogeographical conditions such as climate and soil, genetic drift and mating system could be the crucial factors for genetic divergence. Furthermore, the study also indicated that northern Africa, Europe and temperate Asia might be the diversity differentiation center of orchardgrass. The result will facilitate the breeding program and germplasm collection and conservation.     

Genetic diversity in Tunisian rosy garlic populations (Allium roseum L.) as evidenced by chloroplastic DNA analysis: Sequence variation of non-coding region and intergenic spacers

Allium roseum (Alliaceae) is an important medicinal and aromatic plant in the flora of Tunisia. It is widespread in southern Tunisia, where it is commonly used as a therapeutic agent. However, its existence is seriously threatened because of over-collection.