Plastid genomes reveal evolutionary shifts in elevational range and flowering time of Osmanthus (Oleaceae)

Species of Osmanthus are economically important ornamental trees, yet information regarding their plastid genomes (plastomes) have rarely been reported, thus hindering taxonomic and evolutionary studies of this small but enigmatic genus. Here, we performed comparative genomics and evolutionary analyses on plastomes of 16 of the 28 currently accepted species, with 11 plastomes newly sequenced. Phylogenetic studies identified four main lineages within the genus that are here designated the: “Caucasian Osmanthus” (corresponding to O. decorus), “Siphosmanthus” (corresponding to O. sect. Siphosmanthus), “O. serrulatus + O. yunnanensis,” and “Core Osmanthus: (corresponding to O. sect. Osmanthus + O. sect. Linocieroides). Molecular clock analysis suggested that Osmanthus split from its sister clade c. 15.83 Ma. The estimated crown ages of the lineages were the following: genus Osmanthus at 12.66 Ma; “Siphosmanthus” clade at 5.85 Ma; “O. serrulatus + O. yunnanensis” at 4.89 Ma; and “Core Osmanthus: clade at 6.2 Ma. Ancestral state reconstructions and trait mapping showed that ancestors of Osmanthus were spring flowering and originated at lower elevations. Phylogenetic principal component analysis clearly distinguished spring‐flowering species from autumn‐flowering species, suggesting that flowering time differentiation is related to the difference in ecological niches. Nucleotide substitution rates of 80 common genes showed slow evolutionary pace and low nucleotide variations, all genes being subjected to purifying selection.     

Exon primed intron-crossing (EPIC) markers reveal natural hybridization and introgression in Actinidia (Actinidiaceae) with sympatric distribution

Interspecific hybridization is widespread among plants. Understanding the phylogenetic relationships among species is necessary for revealing the potential hybridization events. Actinidia, best known as kiwifruit genus found throughout a wide range in eastern Asian from Indonesia to Siberia. In this study, phylogenetic relationships of Actinidia species with sympatric distributions were investigated using three chloroplast introns (trnL-F, atpB-rbcL and rpl32-trnL) and three Exon primed intron-crossing (EPIC) markers. Chloroplast phylogeny supports non-monophyly of the five species studied excluding Actinidia fulvicoma var. lanata. The non-monophyly was also revealed by EPIC markers. Our results showed EPIC markers are more variable and informative for phylogenetic inference than that of chloroplast markers. The incongruences between loci from the plastid and nuclear DNA phylogenic trees may stem from incomplete lineage sorting or historical introgression hybridization. Incomplete lineage sorting may explain the non-monophyly between Actinidia chrysantha (section Maculatae) and other four species (section Stellatae), and introgression hybridization and high level of interspecific gene flow may explain the non-monophyly among the species of sect. Stellatae. Thus, natural hybridization and introgression may be common in Actinidia with sympatric distribution.     

Molecular phylogeny of Osmanthus (Oleaceae) based on non‐coding chloroplast and nuclear ribosomal internal transcribed spacer regions

Abstract The phylogenetic relationships of Osmanthus Lour. were investigated using the nuclear ribosomal internal transcribed spacer (ITS) regions and non‐coding chloroplast regions (psbA‐trnH, trnL‐F). The two datasets support the conclusion that Osmanthus is polyphyletic, with some species of the subtribe Oleinae nested within Osmanthus. Osmanthus didymopetalus P. S. Green is nested within the clade formed by species of section Osmanthus in two trees. Osmanthus attenuatus P. S. Green, O. yunnanensis P. S. Green, and O. gracilinervis R. L. Lu of traditional section Osmanthus are clearly divergent from other accessions, and do not form a monophyletic group with other Osmanthus accessions. Osmanthus marginatus Hemsl. is embedded in the clade formed by species of section Osmanthus in the ITS tree. In cpDNA trees all species of section Osmanthus are placed in the large clade and all species of section Leiolea formed a group. The taxonomic incongruence among trees for ITS and cpDNA indicate hybridization, as introgression may have occurred among some species of sections Osmanthus and Leiolea. Phylogeny of Osmanthus is discussed in light of molecular and morphological data, and a revised infrageneric classification with three sections (Leiolea, Siphosmanthu, and Osmanthus) is presented. The section Linocieroides is abandoned and united with section Osmanthus.     

Historical isolation versus recent long-distance connections between Europe and Africa in bifid toadflaxes (Linaria sect. Versicolores)

Background

Due to its complex, dynamic and well-known paleogeography, the Mediterranean region provides an ideal framework to study the colonization history of plant lineages. The genus Linaria has its diversity centre in the Mediterranean region, both in Europe and Africa. The last land connection between both continental plates occurred during the Messinian Salinity Crisis, in the late Miocene (5.96 to 5.33 Ma).

Methodology/Principal Findings

We analyzed the colonization history of Linaria sect. Versicolores (bifid toadflaxes), which includes c. 22 species distributed across the Mediterranean, including Europe and Africa. Two cpDNA regions (rpl32-trnL^UAG and trnK-matK) were sequenced from 66 samples of Linaria. We conducted phylogenetic, dating, biogeographic and phylogeographic analyses to reconstruct colonization patterns in space and time. Four major clades were found: two of them exclusively contain Iberian samples, while the other two include northern African samples together with some European samples. The bifid toadflaxes have been split in African and European clades since the late Miocene, and most lineage and speciation differentiation occurred during the Pliocene and Quaternary. We have strongly inferred four events of post-Messinian colonization following long-distance dispersal from northern Africa to the Iberian Peninsula, Sicily and Greece.

Conclusions/Significance

The current distribution of Linaria sect. Versicolores lineages is explained by both ancient isolation between African and European populations and recent events of long-distance dispersal over sea barriers. This result provides new evidence for the biogeographic complexity of the Mediterranean region.     

Molecular phylogeny of the endemic fern genera Cyrtomidictyum and Cyrtogonellum (Dryopteridaceae) from East Asia

Cyrtomidictyum Ching and Cyrtogonellum Ching are two eastern Asian endemic genera whose taxonomic affinities and phylogenetic relationships have long been controversial. The main uncertainty surrounds the separation of the two genera from the species-rich genus Polystichum. Here we present a phylogenetic study focusing on the phylogenetic relationships of these polystichoid ferns. We reconstructed the relationships based on DNA sequence variation in four chloroplast genome regions, rbcL, atpB, and the intergenic spacers (IGS) rps4-trnS and trnL-trnF. Maximum likelihood and Bayesian inference analyses confirm earlier results that were based on less comprehensive taxon sampling and either only a single gene (rbcL) or two IGS (rps4-trnS and trnL-trnF). Cyrtomidictyum is the sister of the clade of polystichoid ferns that includes Cyrtogonellum, Cyrtomium subser. Balansana and three sections of Polystichum. Cyrtogonellum groups with several species of Polystichum, and constitutes the sister taxon to Polystichum sect. Sphaenopolystichum. We support the recognition of Cyrtomidictyum as circumscribed initially, rather than expansion of the genus to include either several Polystichum species or Cyrtogonellum, some Polystichum and Cyrtomium species. The monophyly of Cyrtomidictyum is supported by morphological characters such as once-pinnate leaves, free venation, prolongated leaf apices, and exindusiate sori. Two synapomorphic indels in the chloroplast genome, one 15-bp deletion in rps4-trnS, and one 3-bp insertion in trnL-trnF further differentiate Cyrtomidictyum from other polystichoid ferns. The close affinity of Cyrtogonellum to section Sphaenopolystichum of Polystichum s.s. is highly supported by molecular data. However, no shared morphological characters or molecular indels have been detected, although the distinctness of Cyrtogonellum is shown by a 13-bp insertion in the rps4-trnS alignment.     

Phylogeny and classification of the Cuban species of Elaphoglossum (Dryopteridaceae), with description of Elaphoglossum sect. Wrightiana sect. nov.

Although a worldwide phylogeny of the bolbitidoid fern genus Elaphoglossum is now available, little is known about the phylogenetic position of the 34 Cuban species. We performed a phylogenetic analysis of a chloroplast DNA dataset for atpß-rbcL (including a fragment of the gene atpß), rps4-trnS, and trnL-trnF. The dataset included 79 new sequences of Elaphoglossum (67 from Cuba) and 299 GenBank sequences of Elaphoglossum and its most closely related outgroups, the bolbitidoid genera Arthrobotrya, Bolbitis, Lomagramma, Mickelia, and Teratophyllum. We obtained a well-resolved phylogeny including the seven main lineages recovered in previous phylogenetic studies of Elaphoglossum. The Cuban endemic E. wrightii was found to be an early diverging lineage of Elaphoglossum, not a member of E. sect. Squamipedia where it was previously classified. We propose a new section for this species: E. sect. Wrightiana. The early diverging position of E. wrightii is of particular interest because the species is a root climber (i.e., climbing from the soil on the lower portions of tree trunks and not losing its connection with the soil), a growth habit it shares with its closest bolbitidoid outgroup genera. This suggests that holoepiphytism evolved later in Elaphoglossum, and the primary hemiepiphytism of E. amygdalifolium, which is sister to the rest of the genus, was derived independently from ancestors that were root climbers. Based on our phylogenetic analysis and morphological investigations, the species of Cuban Elaphoglossum were found to occur in E. sects. Elaphoglossum, Lepidoglossa, Polytrichia, Setosa, and Squamipedia.     

Multi-gene region phylogenetic analyses suggest reticulate evolution and a clade of Australian origin among paleotropical woody bamboos (Poaceae: Bambusoideae: Bambuseae)

The paleotropical woody bamboo subtribe Bambusinae has not been satisfactorily circumscribed and remains a major taxonomic problem. Genera such as Holttumochloa, Mullerochloa and Temburongia have not been confidently assigned to this subtribe. The phylogenetic relationships among genera hitherto placed in or near the Bambusinae, together with the related subtribes Racemobambosinae and Melocanninae, were investigated using three chloroplast DNA markers (rps16-trnQ, trnC-rpoB and trnD-T intergenic spacers) and a nuclear DNA marker (granule-bound starch synthase I, GBSSI) for a sample of 51 ingroup species and 2 outgroup species. Phylogenetic analyses revealed four distinct lineages among the members of the currently recognized Bambusinae: (1) the Bambusa-Dendrocalamus-Gigantochloa (BDG) complex, (2) the Holttumochloa-Kinabaluchloa clade, (3) the Dinochloa-Mullerochloa-Neololeba-Sphaerobambos (DMNS) clade and (4) Temburongia simplex. The BDG complex, which comprises the core of the Bambusinae, appears to have a complex evolutionary history as indicated by incongruence between the cpDNA and the nuclear gene topologies. Introgressive hybridization and incomplete lineage sorting are possible underlying causes for this complexity. The distinction of the climbing-scrambling bamboo lineages from the core Bambusinae and Racemobambosinae suggests directions for investigating the possible existence of further subtribes with increased taxon and geographical sampling. Possible biogeographic scenarios associated with the Holttumochloa-Kinabaluchloa clade and the DMNS clade are discussed.     

Phylogenetic relationship of wild and cultivated yam species (Dioscorea spp.) of India inferred from PCR–RFLP analysis of two cpDNA loci

The genus Dioscorea comprises approximately 600 species worldwide and about 50 in India. Dioscorea is economically very useful because of its food and medicinal value. Polymerase chain reaction–restriction fragment length polymorphism (PCR–RFLP) analysis was employed to study interspecific relationship between four cultivated and thirteen wild species belonging to seven taxonomical sections found in India. Restriction digestion analysis was performed for the amplified products of chloroplast DNA (cpDNA) loci psbA–trnH and trnL-F regions. Out of 17 restriction endonucleases tested, 5 endonucleases had restriction sites in psbA–trnH region and 6 endonucleases had restriction sites in trnL-F region. Digestion of amplified products with these restriction endonucleases showed 70 fragments of which 62 fragments (88 %) were polymorphic. Dendrogram obtained from UPGMA cluster analysis of two combined loci shows D. bulbifera of section Opsophyton to be closely placed with species of sect. Enantiophyllum. High similarity values obtained in this study between D. pentaphylla (sect. Botrysicyos) and D. esculenta (sect. Combilium) support their common origin. The average of genetic similarities between the ten species of Enantiophyllum section was 82.3 % showing their closeness. The present study suggests that the less studied New World species, D. floribunda (sect. Heterostemon) could have evolved from old world section, Lasiophyton. The principal component analysis (PCA) obtained from data of the two loci shows D. deltoidea of sect. Stenophora to be an outgroup. Interspecific genetic similarities showed D. deltoidea with low similarity values ranging from 27.5 to 37.5 % compared with other species and was, therefore, the most divergent species. D. deltoidea of section Stenophora shows distinctness because of its primitive characters. This study, therefore, establishes species relationships between wild and cultivated yams of India which would help in yam crop improvement programmes.     

Systematics and evolution of Carex sects. Spirostachyae and Elatae (Cyperaceae)

Carex sect. Spirostachyae comprises 25 species displaying the centre of diversity in Eurasia, while sect. Elatae comprises 22 species displaying the centre of diversity in tropical African mountainous regions. Phylogenetic analysis of the 136 ITS and 108 5′trnK intron sequences of 23 species of sect. Spirostachyae and 20 species of sect. Elatae revealed that neither section is monophyletic. With the exclusion of C. montis-eeka (sect. Spirostachyae) and C. insularis, C. iynx and C. longibrachiata (sect. Elatae), the sects. Spirostachyae and Elatae constitute a monophyletic group of 38 species, 22 of which were previously included in sect. Spirostachyae and 16 of which were in sect. Elatae (considering C. thomasii as C. mannii ssp. thomasii). Two main groups with different edaphic preferences were identified in the core Spirostachyae, in congruence with some morphological features. One group comprises primarily acidophilus species including 11 species of sect. Spirostachyae and 16 species of sect. Elatae. The other group includes 11 basophilic species of sect. Spirostachyae. Incongruence between nuclear and plastid genomes was detected, suggesting hybridization or lineage sorting processes in the evolution of the core Spirostachyae.     

Genetic relationships of <Emphasis Type="Italic">Osmanthus</Emphasis> based on ISSR-PCR

Osmanthus is an important gardening plant, but determination of germplasm resources, number of species and bipship among species of Osmanthus are still not clear. In this article, genetic diversity of Osmanthus was first studied by ISSR molecular marker. Based on 20 primers screened from 74 primers, 361 bands were obtained from 17 species, and 95.29 % bands were polymorphic. Furthermore, it was found that the genetic distance between O. attenuatus and O. heterophyllus was the highest (0.5559), and was the lowest (0.2351) between O. ×fortunei and O. heterophyllus. As shown in the dendrogram obtained by UPGMA method, 17 species of Osmanthus could be divided into group I and group II, the group I was composed of O. matsumuraanus, O. americanus, O. yunnanensis and O. attenuatus, and the other species belonged to the group II that was further divided into two subgroups apparently, which were not consistent with the classic classification system, but mostly similar with traditional viewpoint about bipship among species. Therefore, it was feasible and effective to study genetic diversity among species of Osmanthus by ISSR molecular marker.     

Syngonanthus androgynus,a Striking New Species from South America,its Phylogenetic Placement and Implications for Evolution of Bisexuality in Eriocaulaceae

In the present study, we describe and illustrate a remarkable new species of Syngonanthus from South America (Bolivia, Brazil and Peru). This new species is quickly distinguished from all species in the genus by trimerous and bisexual flowers, a unique set of characteristics in Syngonanthus. Complementary to this study, sequences of 33 species were downloaded from GenBank and four species had sequences newly generated for this study. Molecular phylogenetic analyses based on nuclear ribosomal ITS and the plastid regions psbA-trnH and trnL-F were performed to determine its systematic position. The results have shown S. androgynus closely related to a well-supported clade that has been treated as Syngonanthus sect. Carphocephalus. Floral traits associated with this new plant also were surveyed. Character reconstruction suggests that the bisexual flowers originated independently more than once in the genus. However, trimerous flowers appear to be an ancestral condition of the whole genus.     

Evolution of trnF(GAA) pseudogenes in cruciferous plants

In several studies we used the 5′-trnL(UAA)–trnF(GAA) region of the chloroplast DNA for phylogeographic reconstructions, gene diversity calculations and phylogenetic analyses among the genera Arabidopsis and Boechera. Despite the fact that extensive gene duplications are rare within the chloroplast genome of higher plants, within several genera of the Brassicaceae the anticodon domain of the trnF(GAA) gene exhibit extensive gene duplications with 1–12 tandemly repeated copies in close 5′-proximity of the functional gene. A recent re-examination and additional analysis of trnL(UAA)–trnF(GAA) regions from numerous cruciferous taxa not only reveal extensive trnF gene duplications, but also favour the hypothesis that in cruciferous taxa at least four independent phylogenetic lineages are characterized by these pseudogenes. Among these lineages there is one major clade of taxa carrying pseudogenes indicating an ancient split in crucifer evolution. In two case studies, Boechera and Arabidopsis, intra- and inter-molecular recombinations have been shown to be the reason for the reciprocal exchange of several similar motifs. However, functional constraints might favour two to three or five to six copies as shown for Arabidopsis and Boechera. Herein, we compare the occurrence and distribution of pseudogene copy number in the framework of a comprehensive survey of cpDNA haplotype variation in Boechera, the former genus Cardaminopsis and Arabidopsis thaliana and comment on the value of such kind of mutations in phylogenetic and evolutionary reconstructions.     

Evidence of horizontal gene transfer between land plant plastids has surprising conservation implications

Background and AimsHorizontal gene transfer (HGT) is an important evolutionary mechanism because it transfers genetic material that may code for traits or functions between species or genomes. It is frequent in mitochondrial and nuclear genomes but has not been demonstrated between plastid genomes of different green land plant species.MethodsWe Sanger-sequenced the nuclear internal transcribed spacers (ITS1 and 2) and the plastid rpl16 G2 intron (rpl16). In five individuals with foreign rpl16 we also sequenced atpB-rbcL and trnL_UAA-trnF_GAA.Key ResultsWe discovered 14 individuals of a moss species with typical nuclear ITSs but foreign plastid rpl16 from a species of a distant lineage. None of the individuals with three plastid markers sequenced contained all foreign markers, demonstrating the transfer of plastid fragments rather than the entire plastid genome, i.e. entire plastids were not transferred. The two lineages diverged 165–185 Myr BP. The extended time interval since lineage divergence suggests that the foreign rpl16 is more likely explained by HGT than by hybridization or incomplete lineage sorting.ConclusionsWe provide the first conclusive evidence of interspecific plastid-to-plastid HGT among land plants. Two aspects are critical: it occurred at several localities during the massive colonization of recently disturbed open habitats that were created by large-scale liming as a freshwater biodiversity conservation measure; and it involved mosses whose unique life cycle includes spores that first develop a filamentous protonema phase. We hypothesize that gene transfer is facilitated when protonema filaments of different species intermix intimately when colonizing disturbed early succession habitats.     

Phylogeny and evolution of Dyckia (Bromeliaceae) inferred from chloroplast and nuclear sequences

The genus Dyckia (Bromeliaceae) comprises more than 150 terrestrial or epilithic species with a strongly xeromorphic habit. Most of its members belong to the azonal rock vegetation of Neotropical savannas and forests of Brazil and adjacent countries. Dyckia is relatively species-rich compared with its closest relatives Encholirium (27 species) and Deuterocohnia (17 species). Here, we present the first molecular phylogenetic analysis of Dyckia using DNA sequence data from six plastid loci (matK gene, rps16 intron, petD intron, rpl32-trnL, rps16-trnK and trnD-trnT) and a portion of the nuclear gene phyC. A total of 124 accessions were included, corresponding to 79 taxa from six genera. Phylogenetic trees were generated using parsimony, likelihood and Bayesian methods. DNA sequence variation among Dyckia species turned out to be extremely low, and phylogenies were poorly resolved. The monophyly of Dyckia is supported, whereas evidence is provided that Encholirium is paraphyletic. Based on a dated plastid DNA tree, Dyckia experienced a recent radiation starting around 2.9 million years ago. Four major clades could be identified that roughly correspond to the geographic origin of the samples. A parsimony network based on plastid DNA haplotypes shows a star-like pattern, indicating recent range expansions. Our data are compatible with a scenario where Dyckia and Encholirium diverged in northeastern Brazil, whereas one lineage of Dyckia dispersed to southern Brazil from where a rapid colonization of suitable habitats was initiated. We discuss our results in relation to species delimitation in Dyckia.     

Chloroplast phylogeography of threatened aquatic <Emphasis Type="Italic">Oxalis</Emphasis> (Oxalidaceae): significant inter-population structure,divergent haplotypes and conservation implications

The aquatic lineage consisting of the sister taxa Oxalis dines and O. disticha is confined to a few small vernal pools in the semi-arid Greater Cape Floristic Region of South Africa. All known populations are at risk of extinction due to anthropogenically induced disturbance. To identify priority sites for focused conservation management, the chloroplast intergenic spacer regions psbA-trnH and trnS-trnG were used to determine population structure and genetic diversity in this lineage across its distribution range. Population viability was assessed using flower morph ratios as surrogate for sexual health. Fourteen and four haplotypes were identified from O. disticha and O. dines populations, respectively. Analyses of Molecular Variance indicated an extremely high level of interpopulation differentiation across the entire aquatic lineage as well as within O. disticha and O. dines. Fifty percent of the eighteen haplotypes were confined to single pools, and 84% of populations contained only a single haplotype, even though pool interspacing distance for O. disticha was often less than 5 km. Almost half of O. disticha haplotype diversity was restricted to very small populations. Two O. dines haplotypes were restricted to small populations, with one of these presenting a divergent haplotype sister to the remainder of the aquatic lineage. Flower morph frequency ratio analyses suggested that most populations were reproductively healthy. Low haplotypic diversity within local populations and differentiation between populations are consistent with very low seed-level gene flow and sporadic founder effects. Conservation efforts should be focussed on preserving as many pools as possible with small, genetically distinct populations representing a main concern.     

Phylogeny and biogeography of Chinese Heracleum (Apiaceae tribe Tordylieae) with comments on their fruit morphology

The Sino-Himalayan Mountain Region is a major center of diversity of Heracleum (Apiaceae, Tordylieae, Tordyliinae), yet the phylogenetic relationships among its rich diversity of endemic taxa are largely unknown. In this study, we (1) investigate the monophyly of Heracleum and ascertain the phylogenetic placements of those species of Heracleum from China that fall outside of the group; (2) estimate phylogenetic relationships within Heracleum sensu stricto, particularly among those species occurring in the Hengduan Mountain Region of southwestern China; and (3) reconstruct the biogeographic history of Heracleum sensu stricto. To achieve these objectives, Bayesian inference and maximum-parsimony analyses of the nuclear ribosomal DNA (nrDNA) internal transcribed spacer (ITS) region and five noncoding chloroplast DNA (cpDNA) loci (rps16-trnK, rps16 intron, trnQ-rps16, rpl32-trnL, and psbA-trnH), separately and in combination, were carried out. Fruit morphology of the Chinese Heracleum species was examined. Statistical dispersal-vicariance analysis (S-DIVA) was used to reconstruct the biogeographic history. In total, 114 accessions were considered, which included 88 species representing eight genera of subtribe Tordyliinae, 28 of the 29 species of Heracleum occurring in China, and 4 species of Angelica as outgroups. The genus Heracleum is monophyletic upon the exclusion of nine species of Heracleum from China and the inclusion of the Caucasian monotypic genera Mandenovia and Symphyoloma. These nine excluded species show affinities to Tetrataenium, Semenovia, and Angelica. The species of Heracleum endemic to southwestern China form a moderately to strongly supported clade; however, major discordance between the ITS- and cpDNA-derived phylogenies, lack of resolution in the trees, and polyphyly of several species preclude unequivocal hypotheses of phylogenetic relationship. The distinctly clavate vittae, shorter than the length of the mericarp, is a diagnostic feature for Heracleum sensu stricto. Heracleum originated in the West Palearctic, with three possible migration routes leading to its present-day distribution. Major topological conflicts between the ITS and cpDNA trees may be due to interspecific hybridization and/or incomplete lineage sorting. The taxonomic limits of many Chinese Heracleum species remain unclear, possibly because of recent radiation in the Hengduan Mountains.     

Study on chloroplast DNA diversity of cultivated and wild pears (<Emphasis Type="Italic">Pyrus</Emphasis> L.) in Northern China

Eight pairs of chloroplast DNA (cpDNA) universal primers selected from 34 pairs were used to assess the genetic diversity of 132 pear accessions in Northern China. Among them, six amplified cpDNA fragments showed genetic diversity. A total of 24 variable sites, including 1 singleton variable site and 23 parsimony informative sites, as well as 21 insertion-deletion fragments, were obtained from the combined cpDNA sequences (5309–5535 bp). Two trnL-trnF-487 haplotypes, five trnL-trnF-413 haplotypes, five rbcL haplotypes, six trnS-psbC haplotypes, eight accD-psaI haplotypes and 12 rps16-trnQ haplotypes were identified among the individuals. Twenty-one haplotypes were identified based on the combined fragments. The values of nucleotide diversity (P_i), average number of nucleotide differences (k) and haplotype diversity (H_d) were 0.00070, 3.56408 and 0.7960, respectively. No statistical significance was detected in Tajima’s D test. Remarkably, the important cpDNA haplotypes and their representing accessions were identified clearly in this study. H_19 was considered as one of the ancient haplotypes and was a divergent centre. H_16 was the most common haplotype of the wild accessions. H_2 was the haplotype representing the most pear germplasm resources (46 cultivars and two wild Ussurian Pear accessions), followed by haplotype H_5 (30 cultivars, two wild Ussurian Pear accessions and four sand pears in outgroups) representing the cultivars ‘Dangshan Suli’ and ‘Yali’, which harbour the largest and the second largest cultivation areas in China. More importantly, this study demonstrated, for the first time, the supposed evolution routes of Pyrus based on cpDNA divergence in the background of pear phylogeny in Northern China.     

Morphological and Molecular Phylogenetic Data Reveal a New Species of Primula (Primulaceae) from Hunan,China

A new species of Primulaceae, Primula undulifolia, is described from the hilly area of Hunan province in south-central China. Its morphology and distributional range suggest that it is allied to P. kwangtungensis, both adapted to subtropical climate, having contiguous distribution and similar habitat, growing on shady and moist cliffs. Petioles, scapes and pedicels of them are densely covered with rusty multicellular hairs, but the new species can be easily distinguished by its smaller flowers and narrowly oblong leaves with undulate margins. Molecular phylogenetic analysis based on four DNA markers (ITS, matK, trnL-F and rps16) confirmed the new species as an independent lineage and constitutes a main clade together with P. kwangtungensis, P. kweichouensis, P. wangii and P. hunanensis of Primula sect. Carolinella.