Molecular characterization of Chlorella cultures of the National Institute for Environmental Studies culture collection with description of Micractinium inermum sp. nov., Didymogenes sphaerica sp. nov., and Didymogenes soliella sp. nov. (Chlorellaceae,Trebouxiophyceae)

Chlorella Beijerinck (Chlorellaceae, Trebouxiophyceae) strains from the collection of the National Institute for Environmental Studies (NIES) were characterized using gene sequence data. The misidentification of a number of strains was rectified. Chlorella vulgaris Beijerinck NIES‐2173 was reclassified as C. sorokiniana Shihira et Krauss. Chlorella sp. NIES‐2171 was described as a new species in the genus Micractinium Fresinius, M. inermum Hoshina et Fujiwara. Chlorella sorokiniana NIES‐2167 and Chlorella sp. NIES‐2330 were found to be phylogenetically related to Didymogenes Schmidle. We propose these two strains be transferred to the genus Didymogenes and given new names: D. sphaerica Hoshina et Fujiwara and D. soliella Hoshina et Fujiwara. Taxonomic decisions were primarily based on small subunit‐internal transcribed spacer ribosomal DNA phylogeny for genus assignment and ITS2 sequence‐structure to determine species autonomy. Our findings suggest that this strategy is the most effective way to use the species concept among autosporic coccoids.     

Phylogenetic reticulation in subtribe Helianthinae

Incongruence between phylogenetic estimates based on nuclear and chloroplast DNA (cpDNA) markers was used to infer that there have been at least two instances of chloroplast transfer, presumably through wide hybridization, in subtribe Helianthinae. One instance involved Simsia dombeyana, which exhibited a cpDNA restriction site phenotype that was markedly divergent from all of the other species of the genus that were surveyed but that matched the restriction site pattern previously reported for South American species of Viguiera. In contrast, analysis of sequence data from the nuclear ribosomal DNA internal transcribed spacer (ITS) region showed Simsia to be entirely monophyletic and placed samples of S. dombeyana as the sister group to the relatively derived S. foetida, a result concordant with morphological information. A sample of a South American species of Viguiera was placed by ITS sequence data as the sister group to a member of V. subg. Amphilepis, which was consistent with cpDNA restriction site data. Samples of Tithonia formed a single monophyletic clade based on ITS sequence data, whereas they were split between two divergent clades based on cpDNA restriction site analysis. The results suggested that cpDNA transfer has occurred between taxa diverged to the level of morphologically distinct genera, and highlight the need for careful and complete assessment of molecular data as a source of phylogenetic information.     

Using DNA‐based techniques to identify hybrids among linear‐leaved Potamogeton plants collected in China

Abstract It is well known that interspecific hybrids occur in the genus Potamogeton. The linear‐leaved Potamogeton species commonly have highly variable morphological characteristics. Their hybrids often show similar vegetative characters to their parental species and their identification based solely on morphology is not always conclusive. In order to clarify whether there are any hybrids from the linear‐leaved Potamogeton plants collected in China, we used internal transcribed spacers (ITS) of nuclear ribosomal DNA and chloroplast rbcL gene sequences to identify the hybrids. Using ITS sequence additivity, we identified four hybrids, namely P. orientalis (P. pusillus×P. oxyphyllus), P. pusillus×P. berchtoldii, P. foliosus×P. octandrus, and P. cristatus×P. octandrus. The latter three hybrids should be considered as new hybrids in Potamogeton. The maternal parents of the four hybrids were confirmed using chloroplast rbcL gene sequences.     

Species delimitation of Chinese hop‐hornbeams based on molecular and morphological evidence

Species delimitation through which infers species boundaries is emerging as a major work in modern systematics. Hop‐hornbeam species in Ostrya (Betulaceae) are well known for their hard and heavy woods. Five species were described in China and their interspecific delimitations remain unclear. In this study, we firstly explored their distributions in all recorded field sites distributed in China. We then selected 110 samples from 22 natural populations of five species from this genus and one type specimen of O. yunnanensis, for molecular barcoding analyses. We sequenced four chloroplast (cp) DNA fragments (trnH–psbA, trnL–trnF, rps16, and trnG) and the nuclear internal transcribed spacer (ITS) region for all samples. Sequence variations of Ostrya from four cpDNA fragments identified three groups that showed no correspondence to any morphological delimitation because of the incomplete lineage sorting and/or possible interspecific introgression in the history. However, phylogenetic analyses of ITS sequence variations discerned four species, O. japonica, O. rehderiana, O. trichocarpa, and O. multinervis while O. yunnanensis nested within O. multinervis. Morphological clustering also discerned four species and showed the complete consistency with molecular evidence. Moreover, our phylogenetic analyses‐based ITS sequence variations suggested that O. trichocarpa comprised an isolated lineage different from the other Eurasian ones. Based on these results, hop‐hornbeams in China should be treated as four separate species. Our results further highlight the importance of ITS sequence variations in delimitating and discerning the closely related species in plants.     

Phylogenetic analysis of the genus Hexachlamys (Myrtaceae) based on plastid and nuclear DNA sequences and their taxonomic implications

Myrtaceae are one of the most species‐rich families of flowering plants in the Neotropics. They include several complex genera and species; Hexachlamys is one of the complex genera. It has not been recognized as a distinct genus and has been included in Eugenia, based on morphological grounds. Therefore, molecular systematic studies may be useful to understand and to help to solve these relationships. Here, we performed a molecular phylogenetic analysis using plastid and nuclear data in order to check the inclusion of Hexachlamys in Eugenia. Plastid (accD, rpoB, rpoC1, trnH‐psbA) and nuclear (ITS2) sequence data were analysed using Bayesian and maximum parsimony methods. The trees constructed using ITS2 and trnH‐psbA were the best able to resolve the relationships between species and genera, revealing the non‐monophyly of Hexachlamys. The molecular phylogenetic analyses were in agreement with previous morphological revisions that have included Hexachlamys in Eugenia. These results reinforce the importance of uniting knowledge and strategies to understand better issues of delimitation of genera and species in groups of plants with taxonomic problems. © 2013 The Linnean Society of London, Botanical Journal of the Linnean Society, 2013, 172 , 532–543.     

Genotyping of a Miso and Soy Sauce Fermentation Yeast,Zygosaccharomyces rouxii,Based on Sequence Analysis of the Partial 26S Ribosomal RNA Gene and Two Internal Transcribed Spacers

We analyzed sequences of the D1D2 domain of the 26S ribosomal RNA gene (26S rDNA sequence), the internal transcribed spacer 1, the 5.8S ribosomal RNA gene, and the internal transcribed spacer 2 (the ITS sequence) from 46 strains of miso and soy sauce fermentation yeast, Zygosaccharomyces rouxii and a closely related species, Z. mellis, for typing. Based on the 26S rDNA sequence analysis, the Z. rouxii strains were of two types, and the extent of sequence divergence between them was 2.6%. Based on the ITS sequence analysis, they were divided into seven types (I–VII). Between the type strain (type I) and type VI, in particular, a 12% difference was detected. The occurrence of these nine genotypes with a divergence of more than 1% in these two sequences suggests that Z. rouxii is a species complex including novel species and hybrids. Z. mellis strains were of two types (type α and type β) based on the ITS sequence. Z. rouxii could clearly be distinguished from Z. mellis by 26S rDNA and ITS sequence analyses, but not by the 16% NaCl tolerance, when used as the sole key characteristic for differentiation between the two species.     

The ITS region of groundwater amphipods: length,secondary structure and phylogenetic information content in Crangonyctoids and Niphargids

The phylogenetic analysis of groundwater amphipods is challenging due to the lack of suitable morphological characters. However, molecular phylogenies based on the 18S and 28S nuclear genes of two Crangonyctoidea species endemic to Iceland, Crymostygius thingvallensis and Crangonyx islandicus, support the taxonomy of these species on the basis of morphological characters. Molecular analyses suggest that the genus Crangonyx is paraphyletic, with the species that is found in Eurasia being highly divergent genetically from the species present in North America and Iceland. Studies of the phylogenetic relationships within the genus Niphargus also warrant further work. The nuclear ITS2 region has recently been proposed as a barcoding marker for plants and animals. In addition, ITS2 has been used to build phylogenies at high taxonomic levels by including its secondary structure. In this study, we want to evaluate the applicability of the ITS region for this group of species and describe its characteristics. The taxonomy of C. thingvallensis, as well as the paraphyly of the genus Crangonyx, is supported herein by phylogenies based on the ITS2 variation. The secondary structure and the length of the ITS2 sequences of the Crangonyctoidea and the Niphargidae species studied are highly variable and are characterized by duplications. The ITS2 sequence of Niphargus plateaui is the longest metazoan sequence deposited in the ITS2 database so far. Although saturation was observed in the nucleotide variation of this marker, the addition of the secondary structure information for the reconstruction of the phylogeny did not add support to the phylogenetic trees. The ITS1 region, which is known to be more variable than ITS2 and bears a large duplication within C. islandicus, was found to be less useful for phylogenetic reconstruction.     

Species distinctions among closely related strains of Eustigmatophyceae (Stramenopiles) emphasizing ITS2 sequence-structure data: Eustigmatos and Vischeria

ABSTRACT

There is an increasing interest in the Eustigmatophyceae, a class of stramenopile microalgae, because they offer a variety of high-value health-beneficial compounds, e.g. polyunsaturated fatty acids (PUFAs), while concomitantly producing high biomass. Clarification of the taxonomy of these organisms at the species level is important in order to achieve reproducible results and constant yields of valuable compounds in their exploitation. Here the distinction of the, so far exclusively, morphologically defined species of the genera Eustigmatos and Vischeria was tested. Distinctions inferred from almost full 18S and ITS2 rRNA as well as plastid-encoded rbcL gene sequences were evaluated following a morphological investigation. The ITS2 secondary-structure-based phylogenies separated independent lineages (species) with long internal branches. This recommends ITS2 as a promising marker for a DNA metabarcoding approach (culture-independent biodiversity assessment). In contrast, the 18S V4 region which is commonly used in metabarcoding was almost invariant, whereas the almost full length sequences distinguished eight groups/types of strains. Monophyly of the species was supported by shared ITS2 secondary structure features, making them distinct from other eustigmatophyte lineages in concordance with phylogenetic analyses. No groups of strains were congruently supported by all three markers. Consequently, the previous distinction of two genera on the basis of morphology cannot be retained and the species should be accommodated in a single genus, Vischeria. Taxonomic changes among the species with the definition of epitypes, on the basis of cryopreserved strains, are recommended. Two findings point to a more complex evolutionary history of the species. The rbcL and nuclear markers resulted in disparate groupings of strains. In three species divergent intragenomic ITS2 paralogues were revealed. Therefore, a still broader taxon sampling, in conjunction with a deep sequencing approach, is needed for a more comprehensive understanding of the complex evolution of eustigmatophyte species.     

Identification of species in the angiosperm family Apiaceae using DNA barcodes

Apiaceae (Umbelliferae) is a large angiosperm family that includes many medicinally important species. The ability to identify these species and their adulterants is important, yet difficult to do so because of their subtle fruit morphological differences and often lack of diagnostic features in preserved specimens. Moreover, dried roots are often the official medical organs, making visual identification to species almost impossible. DNA barcoding has been proposed as a powerful taxonomic tool for species identification. The Consortium for the Barcode of Life (CBOL) Plant Working Group has recommended the combination of rbcL+matK as the core plant barcode. Recently, the China Plant BOL Group proposed that the nuclear ribosomal DNA internal transcribed spacer (ITS), as well as a subset of this marker (ITS2), be incorporated alongside rbcL+matK into the core barcode for seed plants, particularly angiosperms. In this study, we assess the effectiveness of these four markers plus psbA‐trnH as Apiaceae barcodes. A total of 6032 sequences representing 1957 species in 385 diverse genera were sampled, of which 211 sequences from 50 individuals (representing seven species) were newly obtained. Of these five markers, ITS and ITS2 showed superior results in intra‐ and interspecific divergence and DNA barcoding gap assessments. For the matched data set (173 samples representing 45 species in five genera), the ITS locus had the highest identification efficiency (73.3%), yet ITS2 also performed relatively well with 66.7% identification efficiency. The identification efficiency increased to 82.2% when using an ITS+psbA‐trnH marker combination (ITS2+psbA‐trnH was 80%), which was significantly higher than that of rbcL+matK (40%). For the full sample data set (3052 ITS sequences, 3732 ITS2 sequences, 1011 psbA‐trnH sequences, 567 matK sequences and 566 rbcL sequences), ITS, ITS2, psbA‐trnH, matK and rbcL had 70.0%, 64.3%, 49.5%, 38.6% and 32.1% discrimination abilities, respectively. These results confirm that ITS or its subset ITS2 be incorporated into the core barcode for Apiaceae and that the combination of ITS/ITS2+psbA‐trnH has much potential value as a powerful, standard DNA barcode for Apiaceae identification.     

A MORPHOLOGICAL AND MOLECULAR STUDY OF AUSTRAL SARGASSUM (FUCALES,PHAEOPHYCEAE) SUPPORTS THE RECOGNITION OF PHYLLOTRICHA AT GENUS LEVEL,WITH FURTHER ADDITIONS TO THE GENUS SARGASSOPSIS1

Sargassum subgenus Phyllotricha currently includes seven species restricted to Australian and New Zealand coasts. A recent study of Cystoseira and other Sargassaceae genera based on mitochondrial 23S DNA and chloroplast‐encoded psbA sequences resulted in the most widely distributed species of subgenus Phyllotricha, Sargassum decurrens, being transferred to the reinstated monospecific Sargassopsis Trevisan. The fate of the residual six Phyllotricha species, however, was not considered. The present study examines these Phyllotricha species, alongside other Sargassum subgenera, Sargassopsis, Sirophysalis trinodis (formerly Cystoseira trinodis) and the New Zealand endemic Carpophyllum Greville, using morphological evidence and the molecular phylogenetic markers cox3, ITS‐2 and the rbcL–S spacer. Our results suggest both the genus Sargassum and Sargassum subgenus Phyllotricha are polyphyletic as currently circumscribed. Four S. subgen. Phyllotricha species, i.e. S. sonderi, S. decipiens, S. varians and S. verruculosum, form a monophyletic group sister to the genus Carpophyllum, and S. peronii is genetically identical to S. decurrens with regard to all three loci. We propose the resurrection of the genus Phyllotricha Areschoug, with type species Phyllotricha sonderi, and include the new combinations Phyllotricha decipiens, Phyllotricha varians and Phyllotricha verruculosum. Sargassum peronii, S. heteromorphum and S. kendrickii are transferred to Sargassopsis and Sargassum peronii is considered a synonym of Sargassopsis decurrens.     

CONSPECIFICITY AND INDO-PACIFIC DISTRIBUTION OF SYMBIODINIUM GENOTYPES (DINOPHYCEAE) FROM GIANT CLAMS

We previously reported the occurrence of genetically‐diverse symbiotic dinoflagellates (zooxanthellae) within and between 7 giant clam species (Tridacnidae) from the Philippines based on the algal isolates' allozyme and random amplified polymorphic DNA (RAPD) patterns. We also reported that these isolates all belong to clade A of the Symbiodinium phylogeny with identical 18S rDNA sequences. Here we extend the genetic characterization of Symbiodinium isolates from giant clams and propose that they are conspecific. We used the combined DNA sequences of the internal transcribed spacer (ITS)1, 5.8S rDNA, and ITS2 regions (rDNA‐ITS region) because the ITS1 and ITS2 regions evolve faster than 18S rDNA and have been shown to be useful in distinguishing strains of other dinoflagellates. DGGE of the most variable segment of the rDNA‐ITS region, ITS1, from clonal representatives of clades A, B, and C showed minimal intragenomic variation. The rDNA‐ITS region shows similar phylogenetic relationships between Symbiodinium isolates from symbiotic bivalves and some cnidarians as does 18S rDNA, and that there are not many different clade A species or strains among cultured zooxanthellae (CZ) from giant clams. The CZ from giant clams had virtually identical sequences, with only a single nucleotide difference in the ITS2 region separating two groups of isolates. These data suggest that there is one CZ species and perhaps two CZ strains, each CZ strain containing individuals that have diverse allozyme and RAPD genotypes. The CZ isolated from giant clams from different areas in the Philippines (21 isolates, 7 clam species), the Australian Great Barrier Reef (1 isolate, 1 clam species), Palau (8 isolates, 7 clam species), and Okinawa, Japan (1 isolate, 1 clam species) shared the same rDNA‐ITS sequences. Furthermore, analysis of fresh isolates from giant clams collected from these geographical areas shows that these bivalves also host indistinguishable clade C symbionts. These data demonstrate that conspecific Symbiodinium genotypes, particularly clade A symbionts, are distributed in giant clams throughout the Indo‐Pacific.     

Using DNA-based techniques to identify hybrids among linear-leaved Potamogeton plants collected in China

Itis well known that interspecific hybrids occur in the genus Potamogeton. The linear-leaved Potamogeton species commonly have highly variable morphological characteristics. Their hybrids often show similar vegetative characters to their parental species and their identification based solely on morphology is not always conclusive. In order to clarify whether there are any hybrids from the linear-leaved Potamogeton plants collected in China, we used internal transcribed spacers (ITS) of nuclear ribosomal DNA and chloroplast rbcL gene sequences to identify the hybrids. Using ITS sequence additivity, we identified four hybrids, namely R orientalis (Rpusillus × P.oxyphyllus),P.pusillus × P. berchtoldii, P. foliosus × P. octandrus, and P. cristatus × P. octandrus. The latter three hybrids should be considered as new hybrids in Potamogeton. The maternal parents of the four hybrids were confirmed using chloroplast rbcL gene sequences.     

TRUE IDENTITY OF THE EUROPEAN FRESHWATER ULVA (CHLOROPHYTA,ULVOPHYCEAE) REVEALED BY A COMBINED MOLECULAR AND MORPHOLOGICAL APPROACH1

A set of 18 freshwater and morphologically similar marine samples of Ulva were collected from inland and coastal waters throughout Europe to assess their taxonomic identity and invasive potential. An additional 11 specimens were obtained from herbaria. The material was studied using a combination of classical morphological methods and molecular techniques; the latter included sequencing of the nuclear internal transcribed spacer (ITS) region (ITS1‐5.8S‐ITS2) and the chloroplast RUBISCO LSU (rbcL) gene and comparison of the ITS2 secondary structure predictions. Based on classical methods, all the specimens could be determined as U. flexuosa Wulfen and could be further divided into three groups matching three infraspecific taxa. This pattern was generally well supported by molecular phylogenetic analyses. All sequenced samples formed a monophyletic lineage within Ulva, showing a putative synapomorphy in the ITS2 secondary structure. The individual subspecies corresponded to phylogenetic clusters within this lineage. In freshwater habitats, the dominant taxon was U. flexuosa subsp. pilifera, but subsp. paradoxa was also occasionally recorded. In marine habitats, only U. flexuosa subsp. flexuosa and subsp. paradoxa were located. These findings support the view that U. flexuosa subsp. pilifera is primarily a freshwater alga that probably dominates in Europe. As confirmed by the study of herbarium specimens, U. flexuosa should be regarded as indigenous, although it has a tendency to form blooms under certain conditions. Besides clarifying the identity of prevailing European freshwater Ulva, the study provides novel data concerning the distribution and morphological plasticity within the U. flexuosa complex.     

CRYPTIC DIVERSITY AND PHYLOGENETIC RELATIONSHIPS WITHIN THE MASTOCARPUS PAPILLATUS SPECIES COMPLEX (RHODOPHYTA,PHYLLOPHORACEAE)1

Mastocarpus papillatus (C. Agardh) Kütz. is a common intertidal red alga occurring along the west coast of North America from Baja California to Alaska. Sequencing of both the chloroplast‐encoded rbcL gene and the nuclear ribosomal internal transcribed spacer (ITS) regions of ～200 specimens from California to Alaska revealed that M. papillatus is actually a complex of at least five species. All five species have high bootstrap support in phylogenetic analyses of both genetic regions, and in the case of the ITS marker, the species also have distinctive patterns of indels. Three of the species are localized in the mid‐ to upper intertidal, whereas two of the species occur in the low intertidal. The species also have different geographic ranges that overlap in the Vancouver Island area of British Columbia. No distinctive, reliable morphological differences were observed among the species. Although a variety of names are available for species in the complex, it is not yet clear which name goes with which species. As part of the survey, I also sequenced other species of Mastocarpus in the northeast Pacific region, and I provide new distribution records for M. jardinii ( J. Agardh) J. A. West and for a nonpapillate and probably undescribed species of Mastocarpus.     

Molecular definition and the ubiquity of species in the genus Naegleria

To investigate the variability within species of the genus Naegleria, the ITS1,5.8S and ITS2 rDNA were sequenced of several strains of N. lovaniensis and its Western Australian variants, N. australiensis, N. fowleri, N. andersoni, N. jamiesoni, N. tihangensis, N. pringsheimi, N. pagei, N. gruberi sensu lato and a Naegleria lineage that lost a group I intron from the SSUrDNA twintron. As a result, it is possible to define a molecular species within the Naegleria genus. In addition, one strain of each different allozyme cluster was sequenced to investigate whether they belong to described species or should be treated as distinct new species. This leads to the proposal of eleven new species. The sequencing results from those Naegleria spp. of which several strains are available indicate that these species are ubiquitous. The only exception might be the species represented by the WA variants. However, there are still many Naegleria spp. for which only one strain has been isolated, hence, it is important that the search for more isolates should be continued worldwide.     

MOLECULAR DELINEATION OF SPECIES AND SPECIES RELATIONSHIPS IN THE RED ALGAL AGAROPHYTES GRACILARIOPSIS AND GRACILARIA (GRACILARIALES)1

Delineation of species in the economically important agarophyte genera Gracilaria and Gracilariopsis has proven extremely difficult using available morphological characteristics. In this study, we examine the usefulness of two transcribed spacers for molecular systematic studies of these genera. The polymerase chain reaction was used to amplify the internal transcribed spacers (ITSs) and the intervening 5.8S ribosomal DNA of the nuclear ribosomal repeat region. In addition, a plastid spacer region and flanking regions of coding genes were amplified from the RUBISCO operon. Both regions were sequenced for individuals and populations of Gracilariopsis lemaneiformis (Bory) Dawson, Acleto, et Foldvik to determine the usefulness of these spacers in delimiting populations. These studies reveal that there is as much variation among individuals of a population as there is between individuals of geographically separate populations. In addition, the ITS spacer regions were compared between different species of Gracilariopsis and Gracilaria. The nuclear ITS spacer region is conserved at a species level in both genera and provides phylogenetically informative characters that can be used to examine species interrelationships among relatively closely related taxa. However, because of the difficulties of aligning this entire region among species from the two genera, the ITS region is not useful for examining intergenera relationships. ITS interspecies sequence comparisons indicate that Gracilariopsis lemaneiformis from California is significantly different from G. lemaneiformis from China and that a species of Gracilariopsis from Peru is more closely related to G. lemaneiformis from North Carolina than it is to the other Gracilariopsis species examined. In addition, these studies indicate that Gracilaria chilensis Bird, McLachlan, et Oliveira from New Zealand and Gracilaria tenuistipitata Chang et Xia from southeast Asia are as closely related as are Gracilaria verrucosa (Hudson) Papenfuss, G. pacifica Abbott, and Gracilaria robusta Kylin. Phylogenetic analysis of aligned plastid spacer sequences from Gracilaria and Gracilariopsis taxa provide similar conclusions about species relationships.     

Sequence analysis of the ribosomal internal transcribed spacers region in spider mites (Prostigmata: Tetranychidae) occurring in citrus orchards in Eastern Spain: use for species discrimination

Tetranychus urticae is a polyphagous mite which is an important pest of citrus worldwide. This mite can be found feeding on many plant species occurring in the citrus agrosystem moving from weeds to trees. Because field samples consist of a mixture of different Tetranychidae species, as a first step necessary to further implement population characterisation of T. urticae, species‐discriminating criteria based on molecular techniques are needed. In this study, the nucleotide variation of the internal transcribed spacers (ITS) 1 and 2 and the intergenic 5.8S fragment of nuclear rDNA of T. urticae, Tetranychus turkestani, Tetranychus evansi, Tetranychus ludeni and Panonychus citri have been determined. Results demonstrate that for these species, the rDNA ITS2 regions are much more conserved than the corresponding rDNA ITS1. The high homogeneity of the ITS2 sequence observed among the specimens of T. urticae obtained from the same ecoregion makes this DNA sequence an excellent tool for species discrimination. ITS sequences differentiate not only species but also specimens from different geographical origin. Furthermore, polymerase chain reaction–restriction fragment length polymorphism analysis of the ITS2 proved adequate for a quick screening of high numbers of field samples.     

GENETIC DIVERSITY AND INTROGRESSION IN TWO CULTIVATED SPECIES (PORPHYRA YEZOENSIS AND PORPHYRA TENERA) AND CLOSELY RELATED WILD SPECIES OF PORPHYRA (BANGIALES,RHODOPHYTA)1

We investigated the genetic variations of the samples that were tentatively identified as two cultivated Porphyra species (Porphyra yezoensis Ueda and Porphyra tenera Kjellm.) from various natural populations in Japan using molecular analyses of plastid and nuclear DNA. From PCR‐RFLP analyses using nuclear internal transcribed spacer (ITS) rDNA and plastid RUBISCO spacer regions and phylogenetic analyses using plastid rbcL and nuclear ITS‐1 rDNA sequences, our samples from natural populations of P. yezoensis and P. tenera showed remarkably higher genetic variations than found in strains that are currently used for cultivation. In addition, it is inferred that our samples contain four wild Porphyra species, and that three of the four species, containing Porphyra kinositae, are closely related to cultivated Porphyra species. Furthermore, our PCR‐RFLP and molecular phylogenetic analyses using both the nuclear and plastid DNA demonstrated the occurrence of plastid introgression from P. yezoensis to P. tenera and suggested the possibility of plastid introgression from cultivated P. yezoensis to wild P. yezoensis. These results imply the importance of collecting and establishing more strains of cultivated Porphyra species and related wild species from natural populations as genetic resources for further improvement of cultivated Porphyra strains.     

Molecular phylogeography of Carapichea ipecacuanha,an amphitropical shrub that occurs in the understory of both semideciduous and evergreen forests

The medicinal shrub Carapichea ipecacuanha (ipecac) is an amphitropic species with three disjunct areas of distribution. In the Brazilian Atlantic and Amazonian ranges, the species was associated mostly with the understory of seasonal semideciduous forests, whereas in the Central American–Colombian range, the species occurred in the understory of moist evergreen forests. We examined the phylogeographic structure of ipecac using chloroplast trnT‐trnL and nuclear internal transcribed spacer (ITS) sequences from 120 and 46 specimens, respectively. To complement existing data on root alkaloid profiles, we used high‐performance liquid chromatography to assess the levels of emetine and cephaeline in 33 specimens from the two Brazilian ranges. The three ranges shared neither nuclear nor chloroplast haplotypes. The phylogeographic structures showed an uneven distribution of genetic diversity, sharp breaks and high levels of genetic differentiation among ranges. Our results suggest that the extant populations are descendents of at least four distinct ancestral lineages. The Atlantic ipecacs showed higher levels of genetic diversity than ipecacs from the other two ranges; it is likely that they derive from two ancestral lineages, with long‐term persistence in that region. The Amazonian ipecacs were monomorphic with respect to the ITS and cpDNA sequences, which supports the view that there was a recent expansion from a single parental source after a strong genetic bottleneck. The existence of a fourth distinct lineage is apparent from the high levels of genetic and chemical differentiation that we identified in the Central American–Columbian ipecacs.     

Resurrection in Carapa (Meliaceae): a reassessment of morphological variation and species boundaries using multivariate methods in a phylogenetic context

The taxonomy of the amphi‐Atlantic tree genus Carapa (Meliaceae) has long been controversial. Of the three species currently recognized in the genus, two are known to present substantial morphological variation that has been used in the past to distinguish several taxa, most of which are currently placed in synonymy. Here, a combination of field observations, univariate analyses of leaf, floral and seed characters and principal coordinate analyses of floral characters in the context of a molecular phylogenetic analysis was used to investigate the patterns of variation and delimit morphological species anew in the genus. These results support the recognition of 27 species in Carapa, of which 16 are previously described and 11 are new. In general, phylogenetically related species occurred in the same geographical area, but were morphologically distinct. © 2011 The Linnean Society of London, Botanical Journal of the Linnean Society, 2011, 165 , 186–221.