Phylogenetic relationships among Phytopythium species,and re-evaluation of Phytopythium fagopyri comb. nov., recovered from damped-off buckwheat seedlings in Japan

We examined the phylogenetic relationships among Phytopythium species using the rDNA ITS region, the LSU rDNA region, and the mitochondrial coxI and coxII genes. The genus was resolved into three monophyletic clades (1–3). Clade 1 was the largest clade, composed of 12 known species. Clades 2 contained two known and one new species candidate and clade 3 contained two known species. Three isolates in clade 2 (FP1, HonMa, and a strain designated as P. helicoides CBS293.35) formed a monophyletic group with high bootstrap support. This monophyletic group was distinct from P. helicoides sensu stricto. All three isolates came from damped-off buckwheat seedlings. The isolates were morphologically identical with one another and were characterized by globose, sub-globose, or pyriform sporangia with apical papillae; internally or internally nested proliferating sporangia; simple sympodia; coiling antheridial stalks; and wavy, sessile, or clavate antheridial cells. The isolates grew at temperatures between 15 °C and 40 °C, and the optimum temperature was 30 °C, with a radial growth rate of 20 mm/24 h. The phylogenetic and morphological analyses indicated that these isolates belong to a distinct species, which was previously under the genus Pythium, named here Phytopythium fagopyri comb. nov.     

Phenotypic and genotypic characteristics of Arcanobacterium haemolyticum isolated from clinical samples in a Danish hospital

Six Arcanobacterium haemolyticum strains isolated from six patients of two hospitals in Denmark were identified phenotypically, also including matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF MS) analysis, and by genotypic methods. The latter were performed by sequencing 16S rDNA and glyceraldehyde 3-phosphate dehydrogenase encoding gene gap and by amplification of an A. haemolyticum specific region of 16S–23S rDNA intergenic spacer region and 23S rDNA. The six A. haemolyticum strains were further investigated for the presence of seven potential virulence genes encoding arcanolysin, phospholipase D, hemolysin A, CAMP factor family protein, collagen binding protein, neuraminidase A and neuraminidase H which appeared to be present in two (seven virulence genes), two (six virulence genes) and two strains (four virulence genes), respectively. The phenotypic and genotypic properties described in the present study might help to reliably identify and further characterize A. haemolyticum isolated from human patients, a species which seems to be of increasing importance.     

5.8S rDNA variability in Allium species belonging to the third evolutionary group

Sequence analysis of 5.8S rDNA in 67 accessions of the subgenus Allium and six other subgenera belonging to the third evolutionary group of Allium genus (Friesen et al., 2006) was performed. Nucleotide substitutions in 5.8S rDNA sequences of Allium accessions were identified and studied for the first time. The probable secondary structure of 5.8S rRNA was constructed. It was shown that mutations in 5.8S rDNA do not involve conserved motifs, and they did not significantly affect the secondary structure of the RNA molecule in Allium accessions.     

Characterization of the rDNA repeat units in the MitchellPetunia genome

The principal rDNA repeating unit in the MitchellPetunia genome has a length of 8.5 kb. In addition there is a major variant of length 9.7 kb, and two minor variants of 9.3 kb and 10.4 kb. The size heterogeneity of the rDNA repeating units results from length differences in the non-transcribed spacer regions. These differences may reflect simple insertions into the non-transcribed spacer region of the major ‘short’ repeat; however, additional sequence changes have occurred since the ‘short’ repeat is characterized by restriction endonuclease cleavage sites which are absent in the longer variant units.     

A genetic and ultrastructural study of three clones of Porphyridium purpureum (Bory de Saint-Vincent, 1797) Drew et Ross, 1965 (Rhodophyta) from the marine microalgae collection of the Zhirmunsky institute of marine biology

In this paper, we present the results of a comparative genetic and ultrastructural study of three clones of the microalga Porphyridium purpureum (Rhodophyta) from the culture collection of marine microalgae of the Zhirmunsky Institute of Marine Biology. All clones, which have different geographical origins, showed a high similarity in terms of the ultramicroscopic structure and the nucleotide sequences of the nuclear ribosomal DNA genes (18S rDNA, ITS1-5.8S rDNA-ITS2, D1-D2 region of 28S rDNA). The obtained data are very helpful for the certification of two strains of P. purpureum that were isolated for the first time in the practice of Russian algological research.     

A new holomorphic species of Mariannaea and epitypification of M. samuelsii

A new holomorphic species, Mariannaea dimorpha, is described and illustrated. The verticillate conidiophores, phialidic conidiogenous cells and aseptate microconidia that form imbricate chains indicate that it belongs to the genus Mariannaea. Sequence analyses of the combined internal transcribed spacer (ITS) and partial large subunit ribosomal DNA (28S rDNA) confirmed its taxonomic position and revealed its distinction from any morphologically similar species. In addition, the teleomorph of M. samuelsii is reported for the first time and its epitype is designated.     

Five new species of the highly diverse genus Plagiostoma (Gnomoniaceae,Diaporthales) from Japan

Members of the genus Plagiostoma (Gnomoniaceae, Diaporthales) are plant pathogenic and endophytic microfungi that inhabit woody and herbaceous plants in temperate regions of the northern hemisphere. In this study, pure cultures were isolated from specimens of Plagiostoma collected in Japan. Regions of the β-tubulin and tef-1α genes and the complete ITS regions 1 and 2, including the 5.8S rDNA, were sequenced and analyzed phylogenetically. Genealogical concordance phylogenetic species recognition and genealogical non-discordance methods were used to define species. Phylogenetic analyses revealed five previously unknown species of Plagiostoma, which are described and illustrated. These species are associated with host plants in the genera Acer (Sapindaceae) and Salix (Salicaceae).     

Identification of hypervariable regions within the 16S–23S rRNA intergenic spacer region of Flavobacterium columnare and its application in assigning genomovar group to an individual strain

Flavobacterium columnare is an important bacterial pathogen of fish with a wide genetic variability within the species. This intra-species diversity has been termed as genomovars and genomovar groups on the basis of Restriction Fragment Length Polymorphisms of 16S rDNA and 16S–23S rDNA intergenic spacer region (ISR), respectively. In this study, we demonstrate the source of genetic heterogeneity in the F. columnare by sequence analysis of ISR. The length of ISR sequences of different genomovars varied from 553 to 592 nucleotides, while the similarity among sequences ranged from 76.1 to 92.6%. A common ISR structure with tRNA^Ala and tRNA^Ile embedded within the sequence was identified in all the genomovars of F. columnare. The results show that strains of F. columnare can be categorized into five genomovar groups based on the heterogeneity of the ISR sequences. Of these, strains belonging to Genomovar I and II can be sub-divided into two groups each; while strains of Genomovar III belong to one group. Sequence similarity between genomovar groups was lower for ISR (76.1–92.6%) as compared to 16S rDNA (96.1–99.4%) indicating its ability to resolve closely related groups within the genomovars of F. columnare. The main source of variation between the genomovar groups is the presence of three hyper variable regions (V1, V2, and V3) in the ISR. Of the three, V3 was found to be the most heterogeneous region and was found to be useful in assigning a genomovar group to an individual strain of F. columnare.     

One-step integration of multiple genes into the oleaginous yeast Yarrowia lipolytica

Yarrowia lipolytica is an unconventional yeast, and is generally recognized as safe (GRAS). It provides a versatile fermentation platform that is used commercially to produce many added-value products. Here we report a multiple fragment assembly method that allows one-step integration of an entire β-carotene biosynthesis pathway (~11 kb, consisting of four genes) via in vivo homologous recombination into the rDNA locus of the Y. lipolytica chromosome. The highest efficiency was 21 %, and the highest production of β-carotene was 2.2 ± 0.3 mg per g dry cell weight. The total procedure was completed in less than one week, as compared to a previously reported sequential gene integration method that required n weeks for n genes. This time-saving method will facilitate synthetic biology, metabolic engineering and functional genomics studies of Y. lipolytica.     

Transcription and processing of rRNA in higher plant cells (Daucus carota,Petroselinum crispum,Acer pseudoplatanus)

Actively dividing cells from parsley (Petroselinum crispum) and carrot (Daucus carota) (bothApiaceae) andAcer pseudoplatanus (Aceraceae) were used to detect the primary gene product of the rDNA in plant cells. Parsley and carrot cells were labelled with [³²P] orthophosphate. In both cases only one high molecular weight rRNA precursor was present on polyacrylamide gels under non-denaturing conditions. Its molecular weight did not exceed 2.5 × 10⁶ daltons. The component emerged from the heterogenous material after a labelling period of 5–10 min. In parsley cells 45 min after onset of incubation labelled mature rRNA (25S and 18S) arrived in the cytoplasm. InAcer pseudoplatanus (incubation period 60 min) two rapidly labelled components did emerge from polyacrylamide gels; their molecular weights were 2.3 and 3.2^? 3.4 × 10⁶ daltons. After electrophoresis under denaturing conditions the larger component was no longer present, thus indicating that it was an aggregate of different RNA molecules. The molecular weights of the rRNA precursors ofD. carota andP. crispum determined after electrophoresis in formamide gels were about 2.1 × 10⁶ daltons. From these results we have no evidence for the existence of rRNA precursors exceeding the molecular weight of 2.5 × 10⁶ daltons.     

On the taxonomy of the genus Systenostrema Hazard & Oldacre, 1975 (Microspora,Thelohaniidae), with description of two new species

The paper treats the taxonomy of the genus Systenostrema Hazard & Oldacre, 1975, starting with an ultrastructural investigation of two new species, parasitic in larvae of the dragonflies Aeshna grandis and Libellula quadrimaculata, collected in Sweden. The two species are identical in pathology and presporal stages, but differ in the shape of spores and sporophorous vesicles, the fine structure of the spores, and numerical characters. The new species, which are named S. alba and S. candida, are compared to the octosporoblastic microsporidia parasitic in Odonata. An emended diagnosis of the genus Systenostrema is given, together with a taxonomic summary. The new combinations S. trichostegiae for Thelohania trichostegiae Baudoin, 1969 and Amblyospora capillata for T. capillata Larsson, 1983 are established.     

Physical localization of rRNA genes by fluorescence in situ hybridization (FISH) and analysis of spacer length variants of 45S rRNA (slvs) genes in some species of genus Sesbania

The somatic chromosome numbers 2n = 12 in S. macrantha, S. coerulescens and 2n = 24 in S. simplicuscula were determined, additionally the contradictory chromosome numbers of S. bispinosa (2n = 12, 13, 14, 24) and S. pachycarpa (2n 12, 14) were determined as 2n = 24 and 2n = 14, respectively. The number of 5S rDNA sites in chromosome pair 1 was highly conserved in all the diploid and tetraploid species studied irrespective of their geographic distribution, suggesting that all diploid and tetraploid species/cytotypes of Sesbania analyzed in the present study are in close proximity. Cytogenetic mapping of the 45S multi-gene family was also carried out using fluorescence in situ hybridization. 45S rDNA was consistently located on short or long arms of two sub-metacentric chromosome pairs except on one chromosome pair in S. macrantha and on three chromosome pairs in S. bispinosa and S. cannabina. Out of these nine species, we observed the homogenization of intergenic spacer in six species and find only one spacer length variant (slv) located on one to three chromosome loci. However, three of the species were observed to have two slvs located on two different chromosomes. The species were grouped as per their evolutionary relationship on the basis of the results of the present study.     

A study of Schizoblastosporion starkeyi-henricii isolates from northern and southern hemispheres of the Earth

A species Schizoblastosporion starkeyi-henricii is common in peats and peaty soils. Fifteen strains of this species isolated from Europe and East Falkland Island were found homogenous in cultural, morphological and physiological characters, with the exception of one strain that was sensitive to elevated osmotic pressure. Analyses of the D1/D2 region of the 26S rDNA and internal transcribed spacers (ITS) confirm that these strains and also Asian isolates are conspecific. Our results show that ecological factors but not geographic distances determine the global distribution of yeast fungi.     

A new species of Backusella (Mucorales) from a Cerrado reserve in Southeast Brazil

Cerrado (Brazilian savanna) areas present a high number of endemic and unknown species and consequently have been included among the world’s biodiversity hotspots. After a survey of zygomycetes from Cerrado areas of the “Reserva Biológica de Mogi Guaçu” (RBMG) in São Paulo State, we have found a Mucor-like fungus that produces very large sporangiospores and clusters of giant cells visible to the naked-eye. This isolate was characterized morphologically, based on partial LSU (28S) and complete ITS rDNA sequences. Maximum likelihood phylogenetic analyses support its inclusion within the recently emended genus Backusella.     

The Indigo of Commerce in Colonial North America

It can be demonstrated that the Indigo of Commerce in Colonial North America consisted of three species in the genusIndigofera. One of these was a native plant,I. Caroliniana Mill, while the other two were introduced.Indigofera tinctoria L. (French Indigo), an Old World species, andI. Suffruticosa Mill. (Guatemala Indigo), a New World species, were both introduced into South Carolina in the late 17th and early 18th centuries. Their cultivation flourished until the American Revolution. Neither of the introduced species has become naturalized in the Carolinas.     

Paenibacillus nicotianae sp. nov., isolated from a tobacco sample

A Gram-stain positive, facultative anaerobic endospore-forming bacterium, designated strain YIM h-19^T, was isolated from a tobacco sample. Cells were observed to be motile rods by means of peritrichous flagella and colonies were observed to be convex, yellow, circular and showed catalase-positive and oxidase-negative reactions. Strain YIM h-19^T was able to grow at 4–45 °C, pH 6.0–8.0 and 0–3 % NaCl (w/v). The predominant respiratory quinone was identified as MK-7. Major fatty acids were identified as anteiso-C_15:0, anteiso-C_17:0 and C_16:0. The polar lipids were found to be phosphatidylethanolamine, diphosphatidylglycerol, phosphatidylglycerol and two unidentified polar lipids. The genomic DNA G+C content was determined to be 54 mol%. 16S rRNA gene sequence analysis showed the strain YIM h-19^T was most closely related to Paenibacillus hordei RH-N24^T and Paenibacillus hunanensis FeL05^T with similarities of 98.30 and 94.64 % respectively. However, DNA–DNA hybridization data indicated that the isolate represented a novel genomic species with the genus Paenibacillus. All data from genotypic and phenotypic analyses support the conclusion that strain YIM h-19^T represents a novel species of the genus Paenibacillus, for which the name Paenibacillus nicotianae sp. nov. is proposed. The type strain is YIM h-19^T (=CGMCC1.12819^T = NRRL B-59112^T).