Inferring the history of the polyploid Silene aegaea (Caryophyllaceae) using plastid and homoeologous nuclear DNA sequences

The origin of the rare allotetraploid Silene aegaea was inferred from plastid rps16 intron sequences, homoeologous copies of nuclear ribosomal internal transcribed spacer (ITS) sequences, and an intron from the nuclear gene coding for the second largest subunit of RNA polymerase II (RPB2). The nuclear DNA regions support the S. sedoides and S. pentelica lineages as most closely related to the two S. aegaea paralogues. A few recombinant ITS sequences were found, but as PCR recombination could be demonstrated, no true recombination could be demonstrated. No recombination was found in the RPB2 sequences. Plastid rps16 intron sequences strongly support S. pentelica as the maternal lineage. The strength of the approach of using homoeologous sequences of several loci is demonstrated, and its usefulness for the study of phylogenies of groups including polyploids is emphasized.     

Reticulate phylogenetics and phytogeographical structure of Heliosperma (Sileneae, Caryophyllaceae) inferred from chloroplast and nuclear DNA sequences

The Balkan Peninsula is known to be one of the most diverse and species-rich parts of Europe, but its biota has gained much less attention in phylogenetic and evolutionary studies compared to other southern European mountain systems. We used nuclear ribosomal internal transcribed spacer (ITS) sequences and intron sequences of the chloroplast gene rps16 to examine phylogenetic and biogeographical patterns within the genus Heliosperma (Sileneae, Caryophyllaceae). The ITS and rps16 intron sequences both support monophyly of Heliosperma, but the data are not conclusive with regard to its exact origin. Three strongly supported clades are found in both data sets, corresponding to Heliosperma alpestre, Heliosperma macranthum and the Heliosperma pusillum clade, including all other taxa. The interrelationships among these three differ between the nuclear and the plastid data sets. Hierarchical relationships within the H. pusillum clade are poorly resolved by the ITS data, but the rps16 intron sequences form two well-supported clades which are geographically, rather than taxonomically, correlated. A similar geographical structure is found in the ITS data, when analyzed with the NeighbourNet method. The apparent rate of change within Heliosperma is slightly higher for rps16 as compared to ITS. In contrast, in the Sileneae outgroup, ITS substitution rates are more than twice as high as those for rps16, a situation more in agreement with what has been found in other rate comparisons of noncoding cpDNA and ITS. Unlike most other Sileneae ITS sequences, the H. pusillum group sequences display extensive polymorphism. A possible explanation to these patterns is extensive hybridization and gene flow within Heliosperma, which together with concerted evolution may have eradicated the ancient divergence suggested by the rps16 data. The morphological differentiation into high elevation, mainly widely distributed taxa, and low elevation narrow endemics is not correlated with the molecular data, and is possibly a result of ecological differentiation.     

The origin and number of introductions of the Hawaiian endemic Silene species (Caryophyllaceae)

The Hawaiian endemic Silene are a small group of woody or semiwoody representatives from a large, predominantly herbaceous, species-rich genus. We here investigated the origin and number of introductions of the endemic Hawaiian Silene based on phylogenetic relationships inferred from DNA sequences from both the plastid (the rps16 intron) and the nuclear (ribosomal internal transcribed sequences, ITS, and intron 23 of the RPB2 gene) genomes. Silene antirrhina, a widespread weedy American annual, is strongly supported as sister to a monophyletic group consisting of the Hawaiian Silene, indicating a single colonization event. There are no obvious morphological similarities between S. antirrhina and any of the species of Hawaiian Silene. Our results suggest an American origin for the Hawaiian endemics because that would require only a single trans-ocean dispersal. Two of the Hawaiian endemics (S. struthioloides and S. hawaiiensis) that form a subclade in the analyses have evolved woodiness after introduction to the Hawaiian Islands. Our results contribute to other recent results based on molecular phylogenetics that emphasize the American continent as a source area for the Hawaiian flora and support a striking morphological radiation and evolution of woodiness from a single introduction to the archipelago.     

A phylogenetic analysis of palm subtribe Archontophoenicinae (Arecaceae) based on 14 DNA regions

Subtribe Archontophoenicinae belongs to Areceae, the largest of all palm tribes. It includes 15 species distributed in five genera, all found in the south‐western Pacific Region. Archontophoenicinae are rather homogeneous in morphology, making phylogenetic relationships problematic to reconstruct using morphological characters. In this study we investigated phylogenetic relationships in Archontophoenicinae based on all 15 species of the subtribe, using a combination of nine plastid and five nuclear DNA sequence markers. The plastid regions used were the coding rbcL, matK, ndhF and rpoC1 (exon 2) and the non‐coding rps16 intron, atpF‐atpH, psbK‐psbI, trnL‐trnF and trnQ‐rps16. The nuclear regions used were AG1, BRSC, ITS2, PRK and RPB2, which have all proved useful in palm systematics. We compared the phylogenetic hypotheses resulting from the plastid versus nuclear datasets, and combined both datasets to retrieve as much phylogenetic information as possible. Our results strongly support a clade composed of all species of Archontophoenix, Actinokentia, Chambeyronia and Kentiopsis, but raise the question of whether Actinorhytis, the fifth genus, should remain in Archontophoenicinae. Interspecific relationships in ‘core Archontophoenicinae’ still remain incompletely resolved, despite the gene and taxon sampling being substantially greater than in previous studies, and question the monophyly of the New Caledonian genera Chambeyronia and Kentiopsis. © 2014 The Linnean Society of London, Botanical Journal of the Linnean Society, 2014, 175 , 469–481.     

RAPD patterns,nrDNA ITS sequences and morphological patterns inSilene sectionSedoideae (Caryophyllaceae)

Hierarchical patterns inSilene sect.Sedoideae were investigated using random amplified polymorphic DNA (RAPD), nucleotide sequences of the internal transcribed spacer (ITS) regions of the 18S–28S nuclear ribosomal DNA, and discrete morphological characters. All data sets firmly supported the species recognized. The RAPD data offered the best resolution at the intraspecific level, supporting the current intraspecific classifications ofS. sedoides andS. integripetala. The ITS sequences and the morphological data gave poor resolution within species, and the three data sets disagreed about the relationships among species. The signal from the RAPD data was strongest and remained when the total data set was analysed. The three data sets all support an amphiploid origin ofS. aegaea, with the strongest evidence from the ITS sequences. Incongruences among data sets as well as merits and shortcomings of each are discussed. The robustness of the results can be evaluated using perturbations of data, i.e., bootstrap and jackknife of taxa and characters. These methods should not be taken as methods of statistical inference at the taxonomic level, because unbiased sampling appears impossible. RAPD data, however, come close to being suitable for statistical estimation of hierarchies at the genome level, but several methodological problems have to be solved.     

Molecular data suggest a hybrid origin for the invasive Caulerpa racemosa (Caulerpales,Chlorophyta) in the Mediterranean Sea

Morphological data has provided a basis for the hypothesis that three taxa belonging to the Caulerpa racemosa complex occur in the Mediterranean Sea: var. turbinata–uvifera, var. lamourouxi, and the `invasive variety'. In order to test this hypothesis and to determine the origin of the `invasive variety', the transcribed spacer ITS1–ITS2 and an 18S ribosomal DNA (rDNA) intron were analysed from 16 isolates of Caulerpa racemosa. The `invasive variety' shows intraindividual polymorphism for both types of sequences. The ITS1–ITS2 data confirm that the three morphological varieties of C. racemosa from the Mediterranean Sea are distinct taxonomic units. The 18S intron data suggest that the new `invasive variety' could be a recent hybrid between var. turbinata–uvifera and an unknown tropical strain. Incongruence between the phylogenetic tree computed from ITS1–ITS2 regions and the 18S intron indicates that homogenization processes of concerted evolution have run at different rates.     

Molecular phylogenetics of subfamily Calamoideae (Palmae) based on nrDNA ITS and cpDNA rps16 intron sequence data

Phylogenetic relationships among the 22 genera of the palm subfamily Calamoideae were investigated using DNA sequence data from the nuclear ribosomal internal transcribed spacer (ITS) region and the chloroplast rps16 intron. The rps16 intron displayed low levels of variation, corroborating previous reports that the chloroplast genome of palms is highly conserved. High levels of within-individual polymorphism were identified in the ITS region, indicating that concerted evolution is not effectively homogenizing the ITS repeats. In the majority of cases, multiple clones from individuals resolved as monophyletic. However, the high levels of homoplasy in the ITS dataset, along with generally poor jackknife support for many clades, led to concerns that topologies obtained from these data might be unreliable. Nevertheless, congruence between trees based on ITS data alone and those based on rps16 intron data was high. Simultaneous analyses of both datasets yielded well-resolved topologies with high levels of jackknife support. A number of exciting groups emerged from the analyses: the African rattan clade comprising the endemic African rattan genera Laccosperma, Eremospatha, and Oncocalamus; the Lepidocaryeae-Raphia clade comprising the fan-leaved New World tribe Lepidocaryeae and the African genus Raphia; and the Asian clade comprising all Asian genera except Eugeissona. The position of Eugeissona was variable, although it did not resolve inside any of the three major clades mentioned above.     

Rps3 and rpl16 genes do not overlap in Oenothera mitochondria: GTG as a potential translation initiation codon in plant mitochondria?

Characterization of the Oenothera mitochondrial ribosomal gene cluster rps19-rps3-rpl16 shows the two genes rps3 and rpl16 to be separated by 9 nucleotides. The first codon of rpl16 is a GTG codon for valine and the only potential translational start. This GTG codon is conserved at the same position in maize, Petunia and Marchantia mitochondria, while sequences diverge upstream. These observations suggest that GTG at least at this position may act as translation initiation codon in plant mitochondria. Analysis of RNA editing suggests both genes to code for functional ribosomal proteins in Oenothera mitochondria. A duplication/recombination event at a decanucleotide in the intron of rps3 created a pseudogene missing part of the intron and the 3 exon.     

Intraspecific nucleotide divergence in Saccharomycodes ludwigii,and proposal of Saccharomycodes pseudoludwigii sp. nov,a new apiculate yeast isolated from China

The six synonyms currently accepted under Saccharomycodes ludwigii were investigated for by phenotypic properties, however, the sequence diversity of the rRNA and protein coding genes have not yet been determined. Nine strains including the type strains of synonyms of S. ludwigii deposited in the CBS yeast collection, Westerdijk Fungal Biodiversity Institute, Utrecht, The Netherlands, were analyzed using a multi-locus sequence analysis (MLSA) approach that included sequences of 18S ribosomal RNA (rRNA), the D1/D2 domains of the 26S rRNA, the ITS region (including the 5.8S rRNA) and fragments of genes encoding the largest subunit of the RNA polymerase II (RPB1 and RPB2) and translation elongation factor 1-α (TEF1). Our results showed that the nine strains have identical D1/D2, 18S and RPB2 sequences and similar ITS, RPB1 and TEF1 sequences, which indicated that they are conspecific. In addition, a novel species of Saccharomycodes, S. pseudoludwigii sp. nov. (type CGMCC 2.4526 ^T) that was isolated from fruit and tree bark in China, is proposed. The MycoBank number of this new species is MB 811,650.

     

Evolution of a RNA polymerase gene family in Silene (Caryophyllaceae)-incomplete concerted evolution and topological congruence among paralogues

Four low-copy nuclear DNA intron regions from the second largest subunits of the RNA polymerase gene family (RPA2, RPB2, RPD2a, and RPD2b), the internal transcribed spacers (ITSs) from the nuclear ribosomal regions, and the rps16 intron from the chloroplast were sequenced and used in a phylogenetic analysis of 29 species from the tribe Sileneae (Caryophyllaceae). We used a low stringency nested polymerase chain reaction (PCR) approach to overcome the difficulties of constructing specific primers for amplification of the low copy nuclear DNA regions. Maximum parsimony analyses resulted in largely congruent phylogenetic trees for all regions. We tested overall model congruence in a likelihood context using the software PLATO and found that ITSs, RPA2, and RPB2 deviated from the maximum likelihood model for the combined data. The topology parameter was then isolated and topological congruence assessed by nonparametric bootstrapping. No strong topological incongruence was found. The analysis of the combined data sets resolves previously poorly known major relationships within Sileneae. Two paralogues of RPD2 were found, and several independent losses and incomplete concerted evolution were inferred. The among-site rate variation was significantly lower in the RNA polymerase introns than in the rps16 intron and ITSs, a property that is attractive in phylogenetic analyses.     

The phylogenetic utility of chloroplast and nuclear DNA markers and the phylogeny of the Rubiaceae tribe Spermacoceae

The phylogenetic utility of chloroplast (atpB-rbcL, petD, rps16, trnL-F) and nuclear (ETS, ITS) DNA regions was investigated for the tribe Spermacoceae of the coffee family (Rubiaceae). ITS was, despite often raised cautions of its utility at higher taxonomic levels, shown to provide the highest number of parsimony informative characters, in partitioned Bayesian analyses it yielded the fewest trees in the 95% credible set, it resolved the highest proportion of well resolved clades, and was the most accurate region as measured by the partition metric and the proportion of correctly resolved clades (well supported clades retrieved from a combined analysis regarded as “true”). For Hedyotis, the nuclear 5S-NTS was shown to be potentially as useful as ITS, despite its shorter sequence length. The chloroplast region being the most phylogenetically informative was the petD group II intron.We also present a phylogeny of Spermacoceae based on a Bayesian analysis of the four chloroplast regions, ITS, and ETS combined. Spermacoceae are shown to be monophyletic. Clades supported by high posterior probabilities are discussed, especially in respect to the current generic classification. Notably, Oldenlandia is polyphyletic, the two subgenera of Kohautia are not sister taxa, and Hedyotis should be treated in a narrow sense to include only Asian species.     

Phylogenetic relationships among species of <Emphasis Type="Italic">Leotia</Emphasis> (Leotiales) based on ITS and RPB2 sequences

Thirty-three collections of Leotia were used to investigate inter-and infra-specific relationships in the genus. Collections were obtained from various parts of the world and represent the ascomatal color forms typical in species of the genus. The ITS rDNA and a variable region of the RNA polymerase II (RPB2) gene were sequenced and analyzed using parsimony and maximum likelihood methods. Although ITS and RPB2 tree topologies differed in regard to the position of two clades of L. lubrica and L. atrovirens, no significant conflict between ITS and RPB2 data or trees was found as determined by the partition homogeneity test. RPB2 sequences in general gave results comparable to ITS; the RPB2 sequences were more easily aligned. Phylogenetic analysis of the sequence data indicates that L. viscosa, L. lubrica and L. atrovirens are polyphyletic species. This suggests that ascomatal color in fresh specimens is not a reliable character alone for determining species in this group. Four major well-supported groups were found; these do not fully correspond to the commonly recognized species. Stipe color, in both fresh and dry condition, seems to correlate with the major recognized groups but features of the ascospores, asci and paraphyses prove too variable to be informative. The most basal group of Leotia species, identified as L. atrovirens, differ from all others by having stipes without gel tissue in their outer layers.     

Molecular Evolution of the mtDNA Encoded <Emphasis Type="Italic">rps3</Emphasis> Gene Among Filamentous Ascomycetes Fungi with an Emphasis on the Ophiostomatoid Fungi

The mitochondrial ribosomal protein S3 (rps3) gene within the fungi is extremely diverse in location and organization, some versions of this gene have been incorporated into a group I intron, others appear to have gained large insertions, microsatellite expansions, or have been invaded by homing endonucleases. Among the ascomycetes fungi the group I intron encoded version of rps3 appears to have a rather complex evolutionary history including first the acquisition of rps3 by a group I intron (mL2449), the loss of the mL2499 intron and the establishment of rps3 as a free-standing gene, and the eventual loss of the intron derived version of rps3.     

Polyphyly of the Padus group of Prunus (Rosaceae) and the evolution of biogeographic disjunctions between eastern Asia and eastern North America

Prunus subgenus Padus is a group with a wide distribution in temperate eastern Asia and eastern North America with one species extending to Europe and one to Central America. Phylogenetic relationships of subgenus Padus were reconstructed using sequences of nuclear ribosomal ITS, and plastid ndhF gene, and rps16 intron and rpl16 intron. Prunus subgenus Padus is shown to be polyphyletic. Taxa of subgenus Padus and subgenus Laurocerasus are highly intermixed in both the ITS and the plastid trees. The results support two disjunctions between eastern North America and Eurasia within the Padus group. One disjunction is between Prunus virginiana of eastern North America and P. padus of Eurasia, estimated to have diverged at 2.99 (95 % HPD 0.59–6.15)–4.1 (95 % HPD 0.63–8.59) mya. The other disjunction is between P. serotina and its Asian relatives. The second disjunction may have occurred earlier than the former one, but the age estimate is difficult due to the unresolved phylogenetic position of the P. serotina complex.     

Molecular evolution and genome divergence at <Emphasis Type="Italic">RPB2</Emphasis> gene of the St and H genome in <Emphasis Type="Italic">Elymus</Emphasis> species

Molecular evolution of the second largest subunit of low copy nuclear RNA polymerase II (RPB2) in allotetrploid StH genomic species of Elymus is characterized here. Our study first reported a 39-bp MITE stowaway element insertion in the genic region of RPB2 gene for all tetraploid Elymus St genome and diploid Pseudoroegneria spicata and P. stipifolia St genome. The sequences on 3′-end are highly conserved, with AGTA in all sequences but H10339 (E. fibrosis), in which the AGTA was replaced with AGAA. All 12 Stowaway-containing sequences encompassed a 9 bp conserved TIRs (GAGGGAGTA). Interestingly, the 5′-end sequence of GGTA which was changed to AGTA or deleted resulted in Stowaway excision in the H genome of Elymus sepcies, in which Stowaway excision did not leave footprint. Another two large insertions in all St genome sequences are also transposable-like elements detected in the genic region of RPB2 gene. Our results indicated that these three transposable element indels have occurred prior to polyploidization, and shaped the homoeologous RPB2 loci in St and H genome of Eymus species. Nucleotide diversity analysis suggested that the RPB2 sequence may evolve faster in the polyploid species than in the diploids. Higher level of polymorphism and genome-specific amplicons generated by this gene indicated that RPB2 is an excellent tool for investigating the phylogeny and evolutionary dynamics of speciation, and the mode of polyploidy formation in Elymus species.