An evaluation of species relationships in the Porphyra perforata complex (Bangiales,Rhodophyta) using starch gel electrophoresis

Traditional morphological features have formed the basis for distinguishing species of Porphyra. Among these features are number of cell layers, number of chloroplasts per cell, arrangement of reproductive structures on the thallus, and overall morphology. Chromosome number and chromosome morphology have helped corroborate some species identities. A survey of northeast Pacific species of Porphyra using starch gel electrophoresis of 15 soluble proteins has shown that electrophoretic banding patterns provide a reliable diagnostic tool for species identification. Data from starch gel electrophoresis are presented to confirm the identities of species formerly associated with the Porphyra perforata species-complex in British Columbia and northern Washington. Porphyra abbottae, P. fallax, P. kanakaensis, and P. torta are recognized as distinct species, and Porphyra sanjuanensis is synonymized with P. perforata.     

Further fragmentation of the polyphyletic genus Polyalthia (Annonaceae): molecular phylogenetic support for a broader delimitation of Marsypopetalum

The species-rich genus Polyalthia has previously been shown to be highly polyphyletic, with species represented in at least five different clades. The Polyalthia species that are associated with Marsypopetalum and Trivalvaria (as revealed either by previous phylogenetic studies or inferred on the basis of comparative morphology) were included in a molecular phylogenetic study based on three chloroplast DNA regions (matK, rbcL and trnL-F). Maximum parsimony, maximum likelihood and Bayesian analyses consistently revealed that several Polyalthia species form a well-supported clade with Marsypopetalum pallidum, and that this clade is sister to Trivalvaria. Diagnostic morphological characters for the clades are re-evaluated and shown to be congruent with the molecular phylogeny. Five Polyalthia species (P. crassa, P. littoralis, P. lucida, P. modesta and P. tristis) are accordingly transferred to Marsypopetalum.     

The phylogenetic relationships of the charismatic poster frogs,Phyllomedusinae (Anura,Hylidae)

The leaf or monkey frogs of the hylid subfamily Phyllomedusinae are a unique group of charismatic anurans. We present a molecular phylogenetic analysis that includes 45 of the 60 species of phyllomedusines using up to 12 genes and intervening tRNAs. The aims were to gain a better understanding of the phylogenetic position of Phrynomedusa, test the monophyly and explore the relationships among several putative lineages (Hylomantis, the H. buckleyi Group, Phasmahyla, the four species groups of Phyllomedusa, and the species of Phyllomedusa that remain unassigned to any group), and to examine the implications of our phylogeny for the evolution of several characters in phyllomedusines. The analyses resulted in a well‐supported phylogenetic hypothesis that provides a historical framework for a discussion of the evolution of characters associated with reproductive biology, gliding behaviour, the physiology of waterproofing, and bioactive peptides. Implications include an earlier origin for eggless capsules than for leaf‐folding behaviour during amplexus, two independent origins of gliding, and an earlier origin of reduction in evaporative water loss than uricotelism, which is a result that originally was predicted on the basis of physiology alone. Furthermore, our results support the prediction that bioactive peptides from different peptide families are to be expected in all species of Phyllomedusinae. Hylomantis (as recently redefined) is shown to be paraphyletic and the synonymy of Agalychnis is revised to remedy this problem by including both Hylomantis and Pachymedusa. © The Willi Hennig Society 2009.     

Esterase Variants in Four Species of the Paramecium aurelia Complex1

One hundred eighty-eight stocks of Paramecium primaurelia, P. biaurelia, P. tetraurelia, and P. octaurelia were grown axenically and tested for five esterases, visualized by starch gel electrophoresis, in a search for variant stocks. The five esterases can be distinguished on the bases of their substrate specificity, sensitivity to an inhibitor, and response to different growth conditions. This paper addresses the nature of the electrophoretic change in mobility of the variant stocks in order that species relationships can be more accurately assessed. Crosses carried out in all four species show that single genes determine the differences in mobility between variant and common subtypes. Extracts of variant stocks that gave similar patterns were run against each other, tested for their sensitivity to the inhibitor, and the pattern was compared to that found in extracts of stocks with variant and common subtypes in other species. The majority of the variants in P. primaurelia, P. tetraurelia, and P. octaurelia show an electrophoretic mobility characteristic of a common subtype, or a variant, in another species. The same proportion of variant subtypes as common subtypes have mobilities similar to esterase subtypes found in other species. Of the four species examined in this paper, P. tetraurelia and P. octaurelia appear to be most closely related on the basis of shared esterase subtypes. In P. biaurelia the mobilities of most of the variants are unique, as are the common esterase subtypes in this species. P. biaurelia stands out as having the greatest number of esterase subtypes, with very few of them homologous to subtypes found in other species. This observation supports the idea of greater diversification of stocks within P. biaurelia than for the other three species.     

Analysis of allozyme variability in three Plantago species and a comparison to morphological variability

Summary The level of electrophoretic variability in three Plantago species, P. major, P. coronopus, and P. lanceolata, was analyzed in relation to their breeding systems and compared with their morphological variability. From each species several populations were analyzed. The outcrossing P. lanceolata had the highest level of electrophoretic variability and the lowest population differentiation. The inbreeding P. major showed the opposite: a low level of electrophoretic variability and a high population differentiation. P. coronopus, with an intermediate breeding system, had an intermediate level of variability and differentiation. In comparing the species, it appeared that P. coronopus and P. major showed good concordance in the distribution of both kinds of variability, each having only a slightly higher morphological than electrophoretic differentiation between populations. P. lanceolata showed a higher morphological than electrophoretic differentiation between populations. A comparison of populations, within species, revealed good concordance of electrophoretic and morphological variability only within P. coronopus, while some populations of the other two species had relatively lower morphological variability compared with electrophoretic variability.Grassland Species Research Group Publication No. 182     

Novel morphological and genetic tools to discriminate species among the family Plumatellidae (Phylactolaemata,Bryozoa)

Some species of the freshwater bryozoans (Bryozoa, Phylactolaemata) belonging to the genus Plumatella are remarkably difficult to identify because of the large similarity of superficial architecture of their statoblasts. The examination of statoblasts by scanning electron microscope (SEM) has in fact resolved only some taxonomic questions. In this article, the authors report on novel morphological and molecular traits to discriminate among ten species of Plumatellidae (P. viganoi, P. repens, P. geimermassardi, P. rugosa, P. reticulata, P. casmiana, P. fungosa, P. emarginata, P. vaihiriae, and Hyalinella punctata). The former traits are based on shape, number, and position of annular chamber pores, whereas the latter reside on amplification and sequence analysis of the Internal Transcribed Spacer (ITS) region of nuclear rDNA. The successful amplification of ITS region from statoblasts and zooidial tubules allowed us to sequence this region on all the species investigated. The ITS sequences showed the presence of sufficient and informative polymorphisms to discriminate among morphologically similar species. It is noteworthy that the resulting ITS phylogenetic tree largely corroborated the distinction of at least two groups of freshwater bryozoans inferred on the basis of the annular chamber pore morphology. This study provides innovative approaches to reliably characterize freshwater bryozoans species and gain more insight into their taxonomy, phylogenetic relationship, and biodiversity.     

Phylogenetic relationship of Paenibacillus species based on putative replication origin regions and analysis of an yheCD-like sequence found in this region

To determine the phylogenetic relationship among Paenibacillus species, putative replication origin regions were compared. In the rsmG-gyrA region, gene arrangements in Paenibacillus species were identical to those of Bacillus species, with the exception of an open reading frame (orf14) positioned between gyrB and gyrA, which was observed only in Paenibacillus species. The orf14 product was homologous to the endospore-associated proteins YheC and YheD of Bacillus subtilis. Phylogenetic analysis based on the YheCD proteins suggested that Orf14 could be categorized into the YheC group. In the Paenibacillus genome, DnaA box clusters were found in rpmH-dnaA and dnaA-dnaN intergenic regions, known as box regions C and R, respectively; this localization was similar to that observed in B. halodurans. A phylogenetic tree based on the nucleotide sequences of the whole replication origin regions suggested that P. popilliae, P. thiaminolyticus, and P. dendritiformis are closely related species.     

Molecular Phylogenetic Characterization of Streptomyces Protease Inhibitor Family

We previously found that proteinaceous protease inhibitors homologous to Streptomyces subtilisin inhibitor (SSI) are widely produced by various Streptomyces species, and we designated them ``SSI-like proteins' (Taguchi S, Kikuchi H, Suzuki M, Kojima S, Terabe M, Miura K, Nakase T, Momose H [1993] Appl Environ Microbiol 59:4338–4341). In this study, SSI-like proteins from five strains of the genus Streptoverticillium were purified and sequenced, and molecular phylogenetic trees were constructed on the basis of the determined amino acid sequences together with those determined previously for Streptomyces species. The phylogenetic trees showed that SSI-like proteins from Streptoverticillium species are phylogenetically included in Streptomyces SSI-like proteins but form a monophyletic group as a distinct lineage within the Streptomyces proteins. This provides an alternative phylogenetic framework to the previous one based on partial small ribosomal RNA sequences, and it may indicate that the phylogenetic affiliation of the genus Streptoverticillium should be revised. The phylogenetic trees also suggested that SSI-like proteins possessing arginine or methionine at the P1 site, the major reactive center site toward target proteases, arose multiple times on independent lineages from ancestral proteins possessing lysine at the P1 site. Most of the codon changes at the P1 site inferred to have occurred during the evolution of SSI-like proteins are consistent with those inferred from the extremely high G + C content of Streptomyces genomes. The inferred minimum number of amino acid replacements at the P1 site was nearly equal to the average number for all the variable sites. It thus appears that positive Darwinian selection, which has been postulated to account for accelerated rates of amino acid replacement at the major reaction center site of mammalian protease inhibitors, may not have dictated the evolution of the bacterial SSI-like proteins. Received: 23 August 1996 / Accepted: 20 November 1996     

Species relationships inPhaseolus: Electrophoretic analysis of seed storage proteins

Relationships among storage proteins in seeds from cultivars and primitive accessions of the four economically most important species ofPhaseolus — P. vulgaris, P. coccineus, P. acutifolius andP. lunatus — were studied. Analysis of SDS-polyacrylamide gel electrophoretic patterns of storage seed proteins revealed common characteristics in the major groups of polypeptides forP. vulgaris, P. coccineus andP. acutifolius, while clear differences existed between thesePhaseolus species and P.lunatus.     

Phylogenetic relationships and molecular evolution in uropeltid snakes (Serpentes: Uropeltidae): allozymes and albumin immunology

Multilocus electrophoretic methods and microcomplement fixation comparisons of serum albumin are used to assess phylogenetic relationships among species of uropeltid snakes, to infer aspects of their population biology and biogeography, and to evaluate their relationships to other primitive snakes (Henophidia). There is very good agreement between phylogenetic inferences derived from the electrophoretic data and those derived from the albumin immunological data. Protein variation detected by electrophoresis is relatively high among 17 operational taxonomic units (OTUs) examined. The mean number of alleles per locus (5.1 across all OTUs), levels of polymorphism (25% of loci), and heterozygosity (4–6%), are typical of, or greater than, values reported for other snakes. Species of uropeltids are genetically highly differentiated, as measured by genetic distances (lowest interspecific Nei's unbiased genetic distances, 0.22-0.27 among several Sri Lankan species; 2.3 between Teretrurus of India and other uropeltines). The phylogenetic tree most consistent with both the immunological and electrophoretic data shows uropeltines from Sri Lanka to be monophyletic, but the Indian species are paraphyletic with respect to those from Sri Lanka. Rhinophis travancoricus of India is inferred to be the sister taxon to the Sri Lankan radiation. As the genera are presently understood, neither Rhinophis nor Uropeltis appears to be monophyletic. A biogeographic scenario derived from the phylogenetic hypothesis suggests an early diversification of uropeltids in India, followed by a single invasion into the lowlands of Sri Lanka. Subsequent evolution on Sri Lanka resulted in occupation of montane biotopes. Cylindrophis is the sister group to uropeltines and is considered a member of the Uropeltidae. The immunological data indicate no phylogenetic association between uropeltids and other ‘anilioid’ taxa, specifically Anilius, Loxocemus or Xenopeltis, although we cannot rule out a very remote relationship. We specifically reject the hypothesis that uropeltines and scolecophidians form a clade relative to henophidians. High levels of genetic variation and a trend toward negative F_IS values for polymorphic loci in three populations suggest generally large effective population sizes and outbreeding in these species. The niche-width variation hypothesis for allozyme loci is not supported by the uropeltid data. In comparison to other vertebrates, the relationship between Nei's genetic distance and albumin immunological distance in uropeltids suggests either conservative albumin evolution or strong differentiation at electrophoretic loci.     

Phylogeny of Nematoda and Cephalorhyncha Derived from 18S rDNA

Phylogenetic relationships of nematodes, nematomorphs, kinorhynchs, priapulids, and some other major groups of invertebrates were studied by 18S rRNA gene sequencing. Kinorhynchs and priapulids form the monophyletic Cephalorhyncha clade that is the closest to the coelomate animals. When phylogenetic trees were generated by different methods, the position of nematomorphs appeared to be unstable. Inclusion of Enoplus brevis, a representative of a slowly evolving nematode lineage, in the set of analyzed species refutes the tree patterns, previously derived from molecular data, where the nematodes appear as a basal bilateral lineage. The nematodes seem to be closer to the coelomate animals than was speculated earlier. According to the results obtained, nematodes, nematomorphs, tardigrades, arthropods, and cephalorhynchs are a paraphyletic association of closely related taxa. Received: 1 December 1997 / Accepted: 9 April 1998     

Alcohol dehydrogenase of Biomphalaria glabrata (Mollusca: Pulmonata): Polymorphism,genetic analysis,and interspecific variations

Alcohol dehydrogenase of Biomphalaria glabrata has been characterized by electrophoresis, substrate specificities, and other physicochemical means. It exists as a multiple molecular form possessing a minimum number of three bands in ovotestis, five in digestive gland, and six in albumen gland. Each organ shows characteristic electrophoretic forms which differ in substrate specificities and the response to the organomercurial inhibitor p-hydroxymercuribenzoate. Mercaptoethanol treatment has no effect on any electrophoretic form. Genetic analyses of the electrophoretic variants show that three different loci are responsible for the synthesis of the various electrophoretic forms observed in this species. Different species vary in their electrophoretic patterns. A possible role of alcohol dehydrogenase isozymes in the phylogenetic relationship among three species, B. glabrata, B. tenagophila, and B. straminea, has been discussed.This work was supported by a grant from the Conselho Nacional de Pesquisas, Brazil.     

Revisiting protein heterozygosity in plants��nucleotide diversity in allozyme coding genes of conifer Pinus sylvestris

Allozyme variation has been and continues to be a major source of information on the level of genetic variation among plant species. Deciphering the molecular basis of electrophoretic variation is essential for understanding the forces affecting the protein level variation. In this study, the relationship between allozyme heterozygosity and nucleotide diversity in plants is investigated among and within species. Allozyme and nucleotide diversity in 27 plant species was reviewed. At the multilocus level, the two methods are congruent: a clear correlation between the two measures of genetic diversity among plant species was observed, strengthening the view that effective population size is the major determinant of genome-wide diversity. Nucleotide diversity at six allozyme coding genes (6pgdB, aco, gdh, gotC, mdhA, and mdhB) in conifer Pinus sylvestris was investigated jointly with electrophoretic data. Single non-synonymous charge-changing mutations were found together with electrophoretic alleles that consequently were mutationally unique. Synonymous site nucleotide diversity (point estimate of θ _W—0.009 per bp) and silent site divergence from Pinus pinaster at allozyme coding loci were at comparable levels with other loci in the species. Linkage disequilibrium was extensive compared to earlier estimates from P. sylvestris and other trees, spanning several kilobases. Allozyme coding genes had an excess of closely related haplotypes whose frequency has recently increased possibly as a result of partial selective sweeps or balancing selection, but complex demographic effects cannot be excluded.     

Species-specific proteins of the 50S subunit of the chloroplast ribosome in the genus Nicotiana

Summary The large subunits (50S) of chloroplast ribosomes were isolated from Nicotiana tabacum, a species of the Western Hemisphere, and from N. excelsior and N. gossei, Australian species. Their proteins were compared by two-dimensional gel electrophoresis. A pair of proteins (T₁₂ and T₁₂) observed in N. tabacum has electrophoretic mobilities which differ from those of a similarly migrating, and probably homologous, pair of proteins observed in N. excelsior and N. gossei. The species-specific proteins in N. tabacum differ slightly in electrophoretic mobilities based on both charge and molecular weight from those in N. excelsior and N. gossei. Tryptic digests of radioiodinated proteins reveal that the peptide maps of all six proteins are similar. These results suggest that chemically altered forms of one or more proteins of the 50S chloroplast ribosome subunit may exist in vivo.     

Phylogenetic analysis of six species of pacific abalone (Haliotidae) based on DNA sequences of 16s rRNA and cytochrome c oxidase subunit I mitochondrial genes

Six species of abalones (Haliotidae) are found on the Korean coasts. Identification and characterization of these abalones are usually based on morphologic characters. In this research we compared the partial sequences of the mitochondrial 16S ribosomal RNA and cytochrome c oxidase subunit I genes to identify species using molecular data and to determine their phylogenetic relationships. Sequence alignments and phylogenetic analysis revealed that the 6 species fell into 2 distinct groups which were genetically distant from each other and exhibited little internal phylogenetic resolution. One group included Haliotis discus hannai, H. discus discus, H. madaka, and H. gigantea, while the other group contained H. diversicolor supertexta and H. diversicolor diversicolor. The 16S rRNA sequences were relatively more conserved than to the COI sequences, but both gene sequences provided sufficient phylogenetic information to distinguish among the 6 species of Pacific abalone, and thus could be valuable molecular characters for species identification.     

Geometric morphometrics of the forewing shape and size discriminate Plebeia species (Hymenoptera: Apidae) nesting in different substrates

Historically, studies evaluating morphological diversity in stingless bees (Hymenoptera, Apidae: Meliponini) by geometric morphometrics have been used to successfully discriminate taxa and/or populations. Moreover, the use of geometric morphometrics to evaluate phylogenetic morphological variation among stingless bee species has received less attention. Here, we used geometric morphometrics to assess taxonomic discrimination and putative phylogenetic signals for six diapausing stingless bee species (Plebeia) occurring in southern Brazil. In all, 12 landmarks were captured from forewings of P. droryana, P. saiqui, P. emerina, P. remota, P. nigriceps and P. wittmanni. Our data show that the centroid size of the forewings reliably discriminated, for example, between P. droryana and P. emerina from P. saiqui. Moreover, this trait does not have a significant phylogenetic signal. In turn, we found that the overall accuracy in discriminating between the six Plebeia species according to forewing shape was 84%, while the confusion matrix achieved 71%. Interestingly, our discriminant analysis separated Plebeia species nesting in tree cavities from those nesting under granitic rocks. The latter group has second cubital (landmarks = 5, 6, 7), first medial (landmarks = 2, 3, 8) and first submarginal cells (landmarks = 3, 4, 9, 10) that are larger than those of species nesting in trees. The forewing shape showed a strong phylogenetic signal, therefore suggesting that its variation may be due to an evolutionary history shared between Plebeia species studied here rather than to environmental features. This work sheds light on the value of forewing size and shape attributes in discriminating Plebeia species within same genus. We suggest that landmarks separating different taxonomic groups could be incorporated into dichotomous keys to help in identifying clades of complex resolution.     

Taxonomic heterogeneity of the collection strains of fluorescent pseudomonads

Typing of 13 strains of fluorescent pseudomonads from the Belarusian collection of nonpathogenic microorganisms (BIM) by ERIC-PCR and BOX-PCR revealed high level of genetic heterogeneity in bacteria, most of which have been previously identified as Pseudomonas fluorescens according to the classical scheme. Evaluation of the similarities of the 16S rRNA gene sequences and their phylogenetic analysis excluded affiliation of the bacteria under study within the same species and allowed them to be distributed within three relatively distant clusters of the genus Pseudomonas phylogenetic tree. While eight strains fell into the phylogenetic group of P. fluorescens, only one of them could be identified as P. fluorescens. Four strains clustered within the P. vancouverensis phylogenetic group, formed by new species, which have been described mainly according to the evaluation of genome relationships. One bacterium was related to a stable branch that did not contain any type strains of the known Pseudomonas spp. These results indicate taxonomic heterogeneity of collection strains of the fluorescent pseudomonads and demonstrate the necessity of identification of them considering the requirements of phylogenetic bacterial taxonomy.     

Seed morphology and variation in the genus<Emphasis Type="Italic"> Pachycereus</Emphasis> (Cactaceae)

Seeds of 13 Pachycereus species and two Stenocereus species that have been suggested as closely related were examined with the scanning electron microscope. Quantitative features were evaluated using multivariate analysis in order to identify characters that distinguish them. Several species groups were recognized on the basis of 16 qualitative characters. All species studied are keeled. Stenocereus aragonii and S. eichlamii share with most Pachycereus species large size, glossy appearance, and a flat relief on periclinal cells in the lateral region. Pachycereus gatesii and P. schottii are unique in having the smallest seeds and a deeply impressed hilum-micropylar region. P. hollianus does not exhibit micro-relief on periclinal walls in the lateral region, and P. fulviceps has no expanded testa border. Multivariate analysis showed that four characters, length, breadth, hilum-micropylar region length, and angle, made the greatest contribution to distinguishing among species groups. More than 80% of P. fulviceps, P. hollianus, P. tepamo, P. weberi, and S. eichlamii seeds could be classified correctly using four seed features and the percentage was even higher using just two or three features for P. gatesii, P. grandis, P. militaris, P. pringlei, and P. schottii. Testa appearance, testa cell-pattern, and position relative to the rim of the hilum-micropylar region were found to be potentially informative and should be combined with other sources of data in future phylogenetic analyses.