Chloroplast DNA phylogeny, reticulate evolution, and biogeography of Paeonia (Paeoniaceae)

The coding region of the mat K gene and two intergenic spacers, psb A-trn H and trn L(UAA)-trn F(GAA), of cpDNA were sequenced to study phylogenetic relationships of 32 Paeonia species. In the psb A-trn H intergenic spacer, short sequences bordered by long inverted repeats have undergone inversions that are often homoplasious mutations. Insertions/deletions found in the two intergenic spacers, mostly resulting from slipped-strand mispairing, provided relatively reliable phylogenetic information. The mat K coding region, evolving more rapidly than the trnL-trn F spacer and more slowly than the psb A-trn H spacer, produced the best resolved phylogenetic tree. The mat K phylogeny was compared with the phylogeny obtained from sequences of internal transcribed spacers (ITS) of nuclear ribosomal DNA. A refined hypothesis of species phylogeny of section Paeonia was proposed by considering the discordance between the nuclear and cpDNA phylogenies to be results of hybrid speciation followed by inheritance of cpDNA of one parent and fixation of ITS sequences of another parent. The Eurasian and western North American disjunct distribution of the genus may have resulted from interrruption of the continuous distribution of ancestral populations of extant peony species across the Bering land bridge during the Miocene. Pleistocene glaciation may have played an important role in triggering extensive reticulate evolution within section Paeonia and shifting distributional ranges of both parental and hybrid species.     

Morphological and molecular phylogenetics in the genus Leptopilina (Hymenoptera: Cynipoidea: Eucoilidae)

We have conducted cladistic analyses of the genus Leptopilina , a group of Drosophila parasitoids studied intensively by (behavioural) ecologists. Twenty-three morphological characters were scored in ten Leptopilina and two outgroup species. At the same time, DNA sequences for the second ribosomal internal transcribed spacer (ITS2) were gathered for eight Leptopilina and one outgroup species. Both data sets yielded phylogenetic trees which were largely compatible. A 'total evidence' analysis resulted in a single tree that provides a relatively robust phylogenetic reconstruction of the group, which may serve as a basis for historically interpreting the distribution of ecological and behavioural traits.     

Extensive chloroplast haplotype variation indicates Pleistocene hybridization and radiation of North American Arabis drummondii, A. x divaricarpa, and A. holboellii (Brassicaceae)

Arabis drummondii, A. holboellii and their hybrid A. x divaricarpa are widespread perennials of open habitats in North America. A phylogenetic analysis based on noncoding chloroplast DNA sequences (trnL intron and trnL/F intergenic spacer) resolved A. drummondii as a monophyletic taxon, but found A. holboellii to bear chloroplast haplotypes from highly diverged evolutionary lineages. This raised the question of a possible polyphyletic origin of A. holboellii. Arabis x divaricarpa was found to be of recent and polytopic origin, a result consistent with its presumed hybrid origin. One hundred and three chloroplast haplotypes were detected within 719 Arabis accessions investigated. The majority of chloroplast-types were estimated to have arisen prior to the Wisconsin glaciation. Phylogeographical analysis using nested clade analysis, suggested for A. holboellii (i). past fragmentation events, partitioning genetic variation in several instances between the Sierra Nevada, the Southern Rocky Mountains and the Colorado Plateau on the one hand and the Central to Northern Rockies of the United States and adjacent Cascades on the other; and for both parental species (ii). recolonization of major areas formerly covered by the Wisconsin glaciation by three haplotypes; and (iii). restricted gene flow indicating isolation by distance in areas south of the last glacial maximum. Arabis x divaricarpa is closely codistributed with its parental species and resampled their haplotypes. The highest genetic diversity was found in the Rocky Mountains from Idaho and Montana south to Utah and Colorado. This area was further hypothesized to have played a major role in the origin of both parental species and probably represented an important glacial refugium. However, evidence for glacial refugia was also found in arctic and boreal regions of Alaska and near the Great Lakes. In comparison to nuclear ribosomal internal transcribed spacer data, chloroplast DNA divergence was very high and evidently predated the origin of A. drummondii and possibly A. holboellii. Divergence of major chloroplast lineages dates back to the middle of the Pleistocene at least. Extensive hybridization is the most likely evolutionary factor working on A. holboellii to explain the revealed discrepancy in nuclear DNA and chloroplast DNA diversification.     

A molecular perspective on the genera Paragonimus Braun, Euparagonimus Chen and Pagumogonimus Chen

The status of the genera Euparagonimus Chen, 1963 and Pagumogonimus Chen, 1963 relative to Paragonimus Braun, 1899 was investigated using DNA sequences from the mitochondrial cytochrome c oxidase subunit I (CO1) gene (partial) and the nuclear ribosomal DNA second internal transcribed spacer (ITS2). In the phylogenetic trees constructed, the genus Pagumogonimus is clearly not monophyletic and therefore not a natural taxon. Indeed, the type species of Pagumogonimus, P. skrjabini from China, is very closely related to Paragonimus miyazakii from Japan. The status of Euparagonimus is less obvious. Euparagonimus cenocopiosus lies distant from other lungflukes included in the analysis. It can be placed as sister to Paragonimus in some analyses and falls within the genus in others. A recently published morphological study placed E. cenocopiosus within the genus Paragonimus and probably this is where it should remain.     

Molecular Phylogenetic Relationships Between Prostanoid-Containing Okinawan Soft Coral (<Emphasis Type="Italic">Clavularia viridis</Emphasis>) and Nonprostanoid-Containing <Emphasis Type="Italic">Clavularia</Emphasis> Species Based on Ribosomal ITS Sequence

To study phylogenetic relationships among Okinawan soft corals of the genus Clavularia, the ribosomal internal transcribed spacer sequences of host corals and the 18S rDNA sequences of symbiotic algae were analyzed. The molecular phylogenetic trees of hosts showed that a prostanoid-containing species, Clavularia viridis, is deeply diverged from other species of Clavularia which do not biosynthesize the prostanoids as the main secondary metabolites. Comparison of their trees suggested poor phylogenetic concordance between hosts and symbionts.     

Secondary structure model for the ITS-2 precursor rRNA of strongyloid nematodes of equids: implications for phylogenetic inference

In order to maximise the positional homology in the primary sequence alignment of the second internal transcribed spacer for 30 species of equine strongyloid nematodes, the secondary structures of the precursor ribosomal RNA were predicted using an approach combining an energy minimisation method and comparative sequence analysis. The results indicated that a common secondary structure model of the second internal transcribed spacer of these nematodes was maintained, despite significant interspecific differences (2–56%) in primary sequences. The secondary structure model was then used to refine the primary second internal transcribed spacer sequence alignment. The “manual” and “structure” alignments were both subjected to phylogenetic analysis using three different tree-building methods to compare the effect of using different sequence alignments on phylogenetic inference. The topologies of the phylogenetic trees inferred from the manual second internal transcribed spacer alignment were usually different to those derived from the structure second internal transcribed spacer alignment. The results suggested that the positional homology in the second internal transcribed spacer primary sequence alignment was maximised when the secondary structure model was taken into consideration.     

ITS regions in diploids of Aegilops (Poaceae) and their phylogenetic implications

The nucleotide sequences of the internal transcribed spacer (ITS) of nuclear ribosomal DNA in nine diploid species representing six sections of Aegilops were determined by direct sequencing of PCR-amplified DNA fragments. These sequences were aligned with two ITS sequences of additional species from Genbank. Sequence divergences were estimated using Kimura two-parameter model, and the phylogenetic analyses were performed using the maximum parsimony (MP) and the neighbor-joining (NJ) methods with PAUP and PHYLIP, respectively. The sequence divergences between the diploid species varied from 0.5% to 4.68%. The resulting MP tree and NJ tree showed relatively congruent phylogenetic relationships among these species, except Ae. caudata. Particularly, Ae. speltoides was basal within the two trees. The paraphyletic relationships between Ae. speltoides and two species of Sect. Sitopsis, and between Ae. uniaristata and two species of Sect. Comopyrum were supported strongly. The ITS data suggest that currently recognized sections within Aegilops should be reconsidered.     

Phylogenetic relationships of Australian strongyloid nematodes inferred from ribosomal DNA sequence data

The sequence of the second internal transcribed spacer of the ribosomal DNA was determined for the following strongyloid nematodes: Cylicocyclus insignis, Chabertia ovina, Oesophagostomum venulosum, Cloacina communis, Cloacina hydriformis, Labiostrongylus labiostrongylus, Parazoniolaimus collaris, Macropostrongylus macropostrongylus, Macropostrongylus yorkei, Rugopharynx australis, Rugopharynx rosemariae, Macropostrongyloides baylisi, Oesophagostomoides longispicularis and Paramacropostrongylus toraliformis, and compared with published sequences for species of Strongylus and for Hypodontus macropi. The resultant phylogenetic trees supported current hypotheses based on morphological evidence for the separation of the families Strongylidae and Chabertiidae, but did not support the separation of the endemic Australian genera as a distinctive clade within the Chabertiidae. The implications of this finding for the phylogenetic origins of the Australian strongyloids are discussed.     

Dactylorhiza (Orchidaceae) in European Russia: combined molecular and morphological analysis

Four plastid and two nuclear (internal transcribed spacer [ITS] ribosomal DNA) markers were used in this study of the Dactylorhiza maculata and D. incarnata complexes (Orchidaceae: Orchidinae) to determine diversity and taxonomic distribution of haplotypes, hybridization frequencies, and maternal parentage of hybrids in 125 samples from 78 populations from European Russia and the Caucasus. A morphometric study of all populations revealed significant correspondence between morphological and plastid DNA data. Most D. maculata sensu stricto (s.s.) specimens from Russia have D. fuchsii haplotypes; this could be evidence for introgression or widespread hybridization between these species in northern Russia. Heterogeneity within populations is much higher for ITS data and is strongly correlated with latitude. Both plastid and nuclear data are significantly correlated with distribution along a south-north axis. Several haplotypes and ITS alleles uncommon in western Europe are more widely distributed in Russia, whereas some frequent haplotypes from western Europe are absent.     

ITS sequence variation and concerted evolution in the natural hybrid species Malus toringoides

The internal transcribed spacer (ITS) region of nuclear ribosomal DNA (nrDNA) is one of the most used molecular characters in plant systematics. Our previous studies based on morphological analysis and ITS sequence variation suggested that Malus toringoides (Rehd.) Hughes is derived from hybridization between M. transitoria (Batal.) Schneid. and M. kansuensis (Batal.) Schneid. To further understand the variation pattern of ITS sequences in M. toringoides, and to elucidate the evolutionary processes that affect ITS sequence variation after hybridization, we sampled 99 accessions from multiple populations of the hybrid and parental species, and then obtained totally 254 ITS sequences by cloning and sequencing. Our ITS variation data demonstrates three outcomes of ITS repeats after hybrid speciation. ～ 27–41% of M. toringoides have only M. transitoria type ITS sequence, ～ 40–70% have M. transitoria type ITS sequence plus one or two chimeric ITS sequences generated by recombination between parental ITS sequences, and six accessions retain both parental type ITS sequences. The plausible evolutionary processes that created the observed ITS variations were inferred to be the joint actions of recombination, concerted evolution, pseudogenization and backcrossing. Our study provides further understandings of the variation model of ITS repeats after hybridization as well as the evolution of M. toringoides after its hybrid speciation.     

基于形态和分子数据的山东梨园新成灾害螨柑橘全爪螨的鉴定

【目的】本研究旨在准确鉴定山东梨园一种新成灾害螨,为其进一步研究和有效防控奠定基础。【方法】采用体视显微镜直接观察雌成螨的形态特征,制作雄成螨的侧面观玻片标本,观察其阳茎形态特征,进行形态鉴定。从单头雌成螨中提取基因组总DNA,以螨类特异性引物扩增线粒体细胞色素氧化酶亚基Ⅰ(cytochrome oxidase subunitⅠ, COⅠ)基因及核糖体DNA第一内转录间隔区(internal transcribed spacer 1, ITS1)和第二内转录间隔区(internal transcribed spacer 2, ITS2)基因并测序,所得序列在NCBI网站上进行Blastn比对,下载与其一致性高的相关基因序列构建系统发育树和计算遗传距离,进行分子鉴定。【结果】结果表明,梨树上害螨的雌成螨背部有粗大突起,可确定为全爪螨属Panonychus害螨。雄螨阳茎特征与柑橘全爪螨Panonychus citri的很相似,其COⅠ, ITS1和ITS2基因序列均与柑橘全爪螨的相应序列具有很高的同源性(核苷酸序列一致性在99%以上),基于COⅠ, ITS1和ITS2基因序列计算的其与柑橘...     

Reticulate hybridization of <Emphasis Type="Italic">Alpinia</Emphasis> (Zingiberaceae) in Taiwan

Reticulate hybridization is a complicated and creative mechanism in plant evolution that can cause interference in phylogenetic studies. Based on observations of intermediate morphology, low pollen fertility, and overlapping distributions of putative parent species, Yang and Wang (Proceedings of the cross-strait symposium on floristic diversity and conservation. National Museum of Natural Science, Taichung, Taiwan, pp 183–197, 1998) first proposed reticulate hybridization of Alpinia in Taiwan. In the present study, molecular tools were used to explore relationships between four parental species and their homoploidy hybrids, and the impact of hybridization on phylogeny reconstruction. Based on DNA markers, maternal heritance of the chloroplast genome, and additivity of nuclear ribosomal internal transcribed spacer, the present results provide strong support for the hybridization hypothesis. Co-existence of parental ribotypes within hybrids revealed that these hybridization events were current, while reciprocal and introgressive hybridization were inferred from chloroplast DNA data. Furthermore, iterative hybridizations involving more than two parental species may occur in notorious hybrid zones. Ecological, phenological, and physiological evidence provides insight into why such frequent hybridization occurs in Taiwanese Alpinia. In the phylogenetic tree of the Zerumbet clade reconstructed in this study, the chloroplast sequences from one hybrid species were not grouped into a subclade, implying instability caused by hybridization. Failure to find morphological apomorphies and biogeographical patterns in this clade was likely partially due to reticulate hybridization. Electronic supplementary material  The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

Molecular taxonomic position of the elephant schistosome, Bivitellobilharzia nairi, newly discovered in Sri Lanka

Bivitellobilharzia nairi (Mudaliar and Ramanujachar, 1945) Dutt and Srivastava, 1955 was first recorded in India. A number of adult worm specimens of this schistosome species were recovered from a domestic elephant, which died in 1999 in Sri Lanka. This is the first report of this schistosome from Sri Lanka. In the present study, in order to clarify the phylogenetic relationship with other species of schistosomes, sequences from the second internal transcribed spacer (ITS2) of the ribosomal gene repeat, part of the 28S ribosomal RNA gene (28S), and part of the mitochondrial cytochrome c oxidase subunit 1 (CO1) gene from B. nairi were analyzed. Two intraspecific variations were seen within 13 individuals in the ITS2 region. In the CO1 region of the mitochondrial DNA, there were four haplotypes in the nucleotide sequences and two haplotypes in the amino acid sequences. Phylogenetic analysis using the nuclear DNA showed that B. nairi was basal to all of species of the genus Schistosoma. The 28S tree also showed that the mammalian lineage was monophyletic. However, phylogenetic analysis using the mitochondrial DNA showed that B. nairi was nested within the genus Schistosoma. The taxonomical position for this species as well as the contradiction between the results from the nuclear and mitochondrial genes were discussed.     

Natural Hybrids between Ligularia vellerea and L.subspicata (Asteraceae: Senecioneae)

Three types of intermediate morphological individuals were found in the sympatric distribution areas of Ligularia vellerea and Lsubspicata. These intermediate individuals were hypothesized to be the hybrids of these two species based on the detailed comparison of their morphological characters. To test this hypothesis, we compared the DNA sequence of Lvellerea, Lsubspicata and the intermediate individuals by using the internal transcribed spacer region of nuclear ribosomal and atpB rbcL intergenic spacer region of the chloroplast. Direct sequencing of ITS clearly had additives and then cloned sequencing cloned out two kinds of sequences that were identical to those putative parental species, thus these putative hybrids were approved. With different morphological characters among three types, their plastid donors were all Lsubspicata according to the maternal sequences of atpB rbcL.     

利用三种分子标记研究缘毛类纤毛虫的系统发育地位

为了探讨缘毛类纤毛虫的系统发育地位 ,利用RAPD方法得到了 9种缘毛类纤毛虫、 1种四膜虫和1种喇叭虫的 3个随机引物的电泳带谱 ;测定了 7种缘毛类纤毛虫rRNA基因中的间隔区 1(ITS1)和小亚基核糖体核糖核酸 (SSrRNA)基因序列 ,并构建了相应的系统树。在比较和分析RAPD、ITS1和SSrRNA基因序列在缘毛类纤毛虫系统发育研究中的适用范围的基础上 ,以SSrRNA基因序列为分子标记研究了缘毛类纤毛虫系统发育地位 ,结果表明 :①缘毛亚纲是单系的 ,作为寡膜纲中一个亚纲的分类地位是合理的 ;②缘毛类纤毛虫可能是寡膜纲中较高等的一个类群。     

Intercontinental Mediterranean disjunct mosses: morphological and molecular patterns

This study focused on three species that occur disjunctly between western North America and the Mediterranean region of southern Europe, northern Africa, and western Asia, forming the so-called Madrean-Tethyan distribution pattern. Quantitative morphological characters were measured in New and Old World plants to find any subtle phenotypic differentiation between the disjunct populations. Sequences from the nuclear ribosomal internal transcribed spacer region were obtained from the same populations to assess differentiation at the molecular level and to compare molecular diversity with patterns of morphological similarity among plants. Little or no morphological differentiation existed between New and Old World plants in any of the species, but internal transcribed spacer (ITS) sequences revealed some phylogeographic structure. Patterns of morphological similarity in all three species were incongruent with phylogeographic structure revealed by sequence data. New World populations were more variable than Old World populations at the molecular level in the three species. Despite some evidence for differentiation between disjunct plants, no plausible mutation rate would date the divergence at ≥20 million years ago (MYA), as implied by the Madrean-Tethyan hypothesis. Recent long-distance dispersal is a more likely explanation for intercontinental disjunctions in these species.     

Genomic Characterization of Interspecific Hybrids between the Scallops Argopecten purpuratus and A. irradians irradians

The Peruvian scallop (Argopecten purpuratus) has been introduced to China and has successfully been hybridized with the bay scallop (A. irradians irradians). The F1 hybrids of these two scallops exhibited a large increase in production traits and some other interesting new characteristics. To understand the genetic basis of this heterosis, nuclear gene and partial mtDNA sequences, and genomic in situ hybridization (GISH) were employed to analyze the genomic organization of the hybrids. Amplification of the ribosomal DNA internal transcribed spacer (ITS) showed that the parental ITS sequences were present in all the hybrid individuals, illustrating that the hybrid offspring inherited nuclear DNA from both parents. Sequence analyses of the ITS region further confirmed that the hybrids harbored alleles from their parents; some recombinant variants were also detected, which revealed some alterations in the nuclear genetic material of the hybrids. The analysis of mitochondrial 16S rDNA showed that the hybrids possessed sequences that were identical to the 16S rDNA of the female parents, proving a matrilineal inheritance of mitochondrial genes in scallops. In addition, GISH clearly discriminated between the parental chromosomes and indicated a combination of haploid genomes of duplex parents in the hybrids. The genetic analyses in our study illustrated that the F1 hybrids inherited nuclear material from both parents and cytoplasmic genetic material maternally, and some variations occurred in the genome, which might contribute to a further understanding of crossbreeding and heterosis in scallop species.     

青藏高原黄绿蜜环菌纯培养菌种的分离培养及分子鉴定

首次从采自青藏高原、与高原牧草嵩草属Kobresia草本植物形成外生菌根的黄绿蜜环菌Armillarialuteo-virens子实体中分离获得一组织分离菌株,运用rDNA-ITS和rDNA-IGS-1测序技术对该组织分离菌株是否为黄绿蜜环菌的纯培养菌种进行分子鉴定,并基于黄绿蜜环菌的5.8S/ITS和IGS-1序列进行核酸序列数据库GenBank同源性检索比对、构建系统发育树。结果表明,本研究获得的黄绿蜜环菌子实体组织分离菌株即为其纯培养菌种。基于ITS的系统发育分析表明黄绿蜜环菌与口蘑科内其它属间物种的系统发育关系较远;基于IGS-1的系统发育分析表明黄绿蜜环菌与蜜环菌属内的其它种序列差异较大,系统发育关系较远,而与Lepiota属内的部分种具有较近的系统发育关系。本研究首次基于分子手段对我国青藏高原的黄绿蜜环菌种进行了分离培养、分子鉴定和系统发育分析,为黄绿蜜环菌的科学分类提供了分子依据。     

Sequence differences in the internal transcribed spacers of DNA among four species of hookworm (Ancylostomatoidea: Ancylostoma)

The two ribosomal DNA internal transcribed spacers (1 and 2) of the hookworms Ancylostoma caninum, A. tubaeforme, A. ceylanicum and A. duodenale were sequenced. The sequence lengths were similar among the four species, except that A. ceylanicum had slightly longer (by 5–7 bp) internal transcribed spacer 1 and 2 sequences. The predicted secondary structure of the internal transcribed spacer 2 precursor rRNA was similar for all species, despite interspecific differences in primary sequence ranging from 0.9% to 13.2%. Interspecific differences in internal transcribed spacer 1 sequence ranged from 0.9% to 7.5%. A cladistic analysis of the sequence data, using the human hookworm Necator americanus as the outgroup, provided little resolution of the phylogenetic relationships, except that A. ceylanicum occurred on a branch external to the other three species. Nonetheless, internal transcribed spacers 1 and 2 may provide useful phylogenetic information at higher taxonomic levels within the superfamily Ancylostomatoidea.     

A Phylogenetic Analysis of Heterorhabditis (Nemata: Rhabditidae) Based on Internal Transcribed Spacer 1 DNA Sequence Data

Internal transcribed spacer 1 sequences were used to infer phylogenetic relationships among 8 of the 9 described species and one putative species of the entomopathogenic nematode genus Heterorhabditis. Sequences were aligned and optimized based on pairwise genetic distance and parsimony criteria and subjected to a variety of sequence alignment parameters. Phylogenetic trees were constructed with maximum parsimony, cladistic, distance, and maximum likelihood algorithms. Our results gave strong support for four pairs of sister species, while relationships between these pairs also were resolved but less well supported. The ITS1 region of the nuclear ribosomal repeat was a reliable source of homologous characters for resolving relationships between closely related taxa but provided more tenuous resolution among more divergent lineages. A high degree of sequence identity and lack of autapomorphic characters suggest that sister species pairs within three distinct lineages may be mutually conspecific. Application of these molecular data and current morphological knowledge to the delimitation of species is hindered by an incomplete understanding of their variability in natural populations.