Chemical and Genetic Studies on Hybrid of Ligularia subspicata and Ligularia cyathiceps Collected in Yunnan Province of China

Two morphologically ambiguous Ligularia samples (samples A and B), and samples with morphology of Ligularia subspicata (sample C), Ligularia lamarum (sample D), or Ligularia cyathiceps (sample E), were collected at Tianchi Pond, Shangrila County, Yunnan Province, China. Analysis of the nucleotide sequence of the internal transcribed spacers (ITSs) in the nuclear ribosomal RNA gene cluster indicated that not only sample B but also sample D was a hybrid of L. cyathiceps and L. lamarum/L. subspicata. Although the morphology of sample A suggested that it was also a hybrid, the ITS sequence of sample A was that of L. cyathiceps. Twenty compounds were isolated from the five samples, and the structures of two new compounds 7 and 14 were determined. Furanoeremophilanes typical of L. lamarum/L. subspicata were detected not only in samples C and D, but also in samples A and B. These results indicate that the ability to produce root chemicals can spread through introgression.     

Chemotaxonomic and Micromorphological Traits of Satureja montana L. and S. subspicata Vis. (Lamiaceae)

Satureja montana and S. subspicata are used as spice, pepper substitute, for preparing tea, juice, and as a medicine. Fourteen populations (seven per species) of Satureja montana L. and S. subspicata Vis . growing in Croatia were examined to determine the chemical composition of the essential oil (analyzed by GC‐FID and GC/MS), the content of macroelements (Na, K, Ca, Mg) and trace elements (B, Fe, Cu, Mn, Zn, Al, Pb, Cr, Cd, Ni, Hg, As) analyzed by ICP‐AES, antioxidant compounds (analyzed by UV/VIS spectrophotometer), and the types and distribution of trichomes (analyzed by scanning electron microscopy). The main constituents of the essential oil were carvacrol and thymol in S. montana (all populations belong to one phenol chemotype), while α‐eudesmol, β‐eudesmol, and spathulenol dominated in S. subspicata (three chemotypes could be distinguished). Both species possess considerably higher quantities of Ca and Mg, and moderate concentrations of K and Na, while Hg and As levels were below the limit of quantification. Non‐glandular trichomes, peltate trichomes, and three types of capitate trichomes were observed on leaves, stem, calyx, and corolla.     

Chemical and Genetic Differentiation of Ligularia hodgsonii in Japan and China

The chemical composition of the root extract and neutral base sequences of L. hodgsonii samples collected in Japan and China were examined. From the Japanese and the Chinese samples, 6β‐(angeloyloxy)furanoeremophilan‐15‐oic acid ( 2 ) and 3β‐acetoxy‐6β‐[(2‐methylbutanoyl)oxy]furanoeremophilan‐10β‐ol ( 3 ) were isolated as the major component, respectively. The sequence of the internal transcribed spacers (ITSs) of the ribosomal RNA gene was different in the Japanese and the Chinese samples. In contrast to the significant genetic difference, chemical diversity was limited to the positions of O‐functionalities on the furanoeremophilane skeleton, supporting our hypothesis that the production of furanoeremophilanes is ecologically advantageous.     

中国橐吾属(菊科-千里光族)的分类学研究(四):康定橐吾和Senecio ficariifolius的名实订正(英文)

将康定橐吾(Ligularia kangtingensis S.W.Liu)处理为细茎橐吾[L.hookeri(C.B.Clarke)Hand.-Mazz.]的异名。Senecio ficariifolius H.Lév.Vaniot长期以来被处理为细茎橐吾的异名,但我们检查模式材料后发现其头状花序盘状,与头状花序辐射状的细茎橐吾明显不同,而与头状花序盘状的贵州橐吾[L.leveillei(Vaniot)Hand.-Mazz.]没有区别,故将其处理为贵州橐吾的异名。     

Isolation and toxigenicity of Aspergillus fumigatus from moldy silage

Thirty-nine silage samples were collected from various siloson Terceira Island in the Azores. Samples were examined for the presence of total fungi, and isolates of Aspergillus fumigatus were analyzed for their ability to produce fumitremorgens B and C, fumigaclavines B and C, and gliotoxin. Thirty-four silage samples (87%) were contaminated with fungi, and A. fumigatus was isolated from 27 samples (69%). Samples that were taken from the surface of silos had significantly higher populations of both total fungi and A. fumigatus than did samples taken from the middle of silos. Analysis of 27 A. fumigatus isolates (one representing each positive sample) showed that 59.3% produced fumitremorgen B; 33.3% produced fumitremorgen C; 29.6% produced fumigaclavine B; 7.4% produced fumigaclavine C; and 11.1% produced gliotoxin. Fifty-two percent of the isolates produced multiple toxins, and 25.9% did not produce any of these toxins. Gliotoxin and fumigaclavine C were always produced in combination with other toxins. Because of the demonstrated potential of these A. fumigatus isolates to producemycotoxins, it is important to properly construct and manage silos to prevent their contamination with A. fumigatus.This revised version was published online in October 2005 with corrections to the Cover Date.     

Symbiodinium diversity in the soft coral Heteroxenia sp. and its nudibranch predator Phyllodesmium lizardensis

We examined the diversity of the photosynthetic dinoflagellate, Symbiodinium, over a 2-year period in two invertebrates from Australia’s Northern Great Barrier Reef: the nudibranch Phyllodesmium lizardensis and an octocoral of the genus Heteroxenia. In years one and two, we used denaturing gradient gel electrophoresis with internal transcribed spacer 2 (ITS2) region amplicons and identified two nearly identical genotypes of clade C (C64 and a variant) in all samples of each species. We examined the secondary structure of both sequences and found that each had predicted ∆G values within the range of stable free energy values for Symbiodinium ITS2 sequences. In year two, we also used real-time quantitative polymerase chain reaction assays (qPCR) with clade-specific internal transcribed spacer 1 primers to determine whether there were cryptic clades (A, B, and/or D) associated with either host in addition to clade C. qPCR revealed that clades B, C, and D were present in all animals of both species and that all but two nudibranch samples also harbored clade A. These findings suggest that there may be more flexibility in this host/symbiont interaction than has previously been assumed.

     

Habitat-specific composition of morphotypes with low genetic diversity in the green algal genus Klebsormidium (Streptophyta) isolated from biological soil crusts in Central European grasslands and forests

Terrestrial filamentous green algae of the widely distributed, cosmopolitan genus Klebsormidium (Klebsormidiophyceae, Streptophyta) are typical components of biological soil crusts (BSCs). These communities occur in all climatic zones and on all continents, where soil moisture is limited or where there has been disturbance. BSCs form water-stable aggregates that have important ecological roles in primary production, nitrogen fixation, nutrient cycling, water retention and stabilization of soils. Although available data on Klebsormidium are limited, its functional importance in BSCs is regarded as high. Therefore, in the present study Klebsormidium strains were isolated from BSCs sampled from various grassland and forest plots of different land use intensities in Central Europe, as provided by the Biodiversity Exploratories, and its intraspecific genetic diversity was evaluated. Previous phylogenetic analyses revealed a relationship between sequence similarity and habitat preference with a higher genetic diversity than expected from a morphological classification. We isolated and sequenced 75 Klebsormidium strains. The molecular phylogeny based on the ITS regions showed that all strains belong to either the previously described clade B/C or clade E. This classification was supported by morphological characteristics: strains assigned to clade B/C were identified as Klebsormidium cf. flaccidum or Klebsormidium cf. dissectum, and strains from clade E as K. nitens or Klebsormidium cf. subtile. Within one clade the strains showed low sequence divergences. These minor differences were independent of the sampling region and land use intensity. Interestingly, most of the strains assigned to clade E were isolated from forest sites, whereas strains from clade B/C occurred equally in grassland and forest sites. Therefore, it is reasonable to assume that habitat with its microenvironmental conditions, and not biogeography, controls genetic diversity in Klebsormidium.     

Chemical Constituents in Hybrids of Ligularia tongolensis and L. cymbulifera: Chemical Introgression in L. tongolensis

Two samples with morphologies intermediate between Ligularia tongolensis and L. cymbulifera were collected in Desha, Sichuan Province, and one, in Pachahai, Yunnan Province, P. R. China. The DNA sequencing confirmed that the samples were hybrids of the two species. Tetradymol ( 1 ), the major compound of L. cymbulifera not found in L. tongolensis, was isolated from the hybrid samples collected at both locations, while furanoeremophilan‐15‐oic acid derivative 4 , a compound characteristic to L. tongolensis, was found in the Pachahai hybrid but not in the Desha hybrids. Thus, the chemical consequence of hybridization can be variable. In addition, analysis of L. tongolensis samples at Pachahai indicated that introgression has been a mechanism of generating chemical diversity in the plant. Eleven compounds including three new ones were isolated.     

Laurencia majuscula var. elegans (Rhodophyta,Rhodomelaceae) is reinstated to specific rank as L. elegans

Laurencia majuscula has recently been determined to be conspecific with Laurencia dendroidea. To determine the correct taxonomic placement of its only non‐typical variety, L. majuscula var. elegans, mitochondrial COI‐5P and chloroplast rbcL gene sequences were analyzed and its phylogenetic position was inferred. Laurencia majuscula var. elegans consistently formed a well‐supported clade distinctly separate from the L. dendroidea clade. Molecular analyses revealed that L. majuscula var. elegans generally shows two distinct forms; a long‐branched form, and a compact form. Often the compact form has been identified as a different species based on morphology alone. Included in these analyses were topotype sequences of two species with which L. majuscula var. elegans is most often misidentified; Laurencia nidifica and Laurencia mcdermidiae. Laurencia nidifica formed a well‐supported clade separate from L. majuscula var. elegans. Laurencia mcdermidiae, however, formed a well‐defined clade sister to the L. majuscula var. elegans clade. Morphological comparisons were also made. It is concluded that L. majuscula var. elegans is morphologically distinct from L. dendroidea, L. nidifica, and L. mcdermidiae. It is proposed that L. majuscula var. elegans be reinstated to species level as L. elegans.     

Evidence that clade A and clade B head lice live in sympatry and recombine in Algeria

Pediculus humanus L. (Psocodea: Pediculidae) can be characterized into three deeply divergent lineages (clades) based on mitochondrial DNA. Clade A consists of both head lice and clothing lice and is distributed worldwide. Clade B consists of head lice only and is mainly found in North and Central America, and in western Europe and Australia. Clade C, which consists only of head lice, is found in Ethiopia, Nepal and Senegal. Twenty‐six head lice collected from pupils at different elementary schools in two localities in Algiers (Algeria) were analysed using molecular methods for genotyping lice (cytochrome b and the multi‐spacer typing (MST) method. For the first time, we found clade B head lice in Africa living in sympatry with clade A head lice. The phylogenetic analysis of the concatenated sequences of these populations of head lice showed that clade A and clade B head lice had recombined, suggesting that interbreeding occurs when lice live in sympatry.     

Sesquiterpenes from Lactarius blennius

Six sesquiterpene lactones were isolated from Lactarius blennius. The structures of two new sesquiterpenes, blennin A and blennin B were determinated by spectroscopic methods and the structure of the seco-compound, blennin C, is revised. The two known furan sesquiterpenes and lactarorufin A were also identified.     

Genetic Diversity and Chemical Polymorphism of Some Thymus Species

To ascertain whether there are chemical and genetic relationships among some Thymus species and also to determine correlation between these two sets of data, the essential‐oil composition and genetic variability of six populations of Thymus including: T. daenensis ?elak. (two populations), T. fallax Fisch . & C.A.Mey ., T. fedtschenkoi Ronniger , T. migricus Klokov & Des .‐Shost ., and T. vulgaris L. were analyzed by GC and GC/MS, and also by randomly amplified polymorphic DNA (RAPD). Thus, 27 individuals were analyzed using 16 RAPD primers, which generated 264 polymorphic scorable bands and volatiles isolated by distillation extraction were subjected to GC and GC/MS analyses. The yields of oils ranged from 2.1 to 3.8% (v/w), and 34 components were identified, amounting to a total percentage of 97.8–99.9%. RAPD Markers allowed a perfect distinction between the different species based on their distinctive genetic background. However, they did not show identical clustering with the volatile‐oil profiles.     

Chlamydomonas neoplanoconvexa nom. nov. and its unique phylogenetic position within Volvocales (Chlorophyceae)

Chlamydomonas Ehrenb. is a unicellular volvocalean genus consisting of 400–600 species, most of which are known solely based on microscopy. In this study, a newly isolated strain of Chlamydomonas neoplanoconvexa (M. O. P. Iyengar) Nakada nom. nov. (≡Chlamydomonas planoconvexa M. O. P. Iyengar non J. W. G. Lund) was examined using light microscopy and combined 18S rRNA, rbcL and psaB gene phylogeny. The C. neoplanoconvexa strain was quite similar to Chlamydomonas perpusilla (Korshikov) Gerloff in terms of its small fusiform vegetative cells (～10 µm in length), parietal chloroplast containing a single pyrenoid, and nucleus posterior to the pyrenoid. However, C. neoplanoconvexa was distinguished from C. perpusilla based on the morphology of the papillae and pyrenoids. The two species belong to the clade Caudivolvoxa in the order Volvocales, but are distantly related to each other. Chlamydomonas perpusilla was shown to be sister to Chlorogonium Ehrenb. (clade Chlorogonia, within Caudivolvoxa), while C. neoplanoconvexa represented a second basal lineage within Caudivolvoxa, next to the clade Characiosiphonia. Although the morphology of C. neoplanoconvexa was not particularly outstanding, its unique phylogenetic position will encourage further investigation of this species and its uncultivated relatives.     

Large subunit rDNA sequences from Alexandrium catenella strains isolated during harmful algal blooms in Hong Kong

The partial sequences of the large subunit (24S) ribosomal RNA gene(LsurRNA) of three Alexandrium catenella strains isolatedduring the 1998 and 1989 red tide incidents in Hong Kong were compared,togetherwith the sequences of other Asian A. catenella speciescomplex. The sequences of two A. catenella strainsisolatedin 1998 were identical, despite being isolated at different time (March andApril) and different locations (eastern and southern Hong Kong). In cladisticanalysis, the two A. catenella HK98A & B strains, theA. catenella HK1989 strain, the Chinese strain clade andtheKorean/Japanese A. catenella form a clade distinct fromother A. catenella strains. Interestingly, the twoA. catenella HK98A & B strains only form a clade withtheA. catenella HK1989 strain/ChineseA. catenella (CCMP1493) with low bootstrap values (49%).     

Isolation and Characterization of an Anther-Specific Polygalacturonase Gene, BcMF16, in Brassica campestris ssp. chinensis

Many genes in the genic male sterile A/B line (Bajh97-01A/B) of Chinese cabbage pak choi (Brassica campestris L. subsp. chinensis Makino) are expressed differentially, and some play critical roles in the formation of pollen walls. In this study, one of these genes, Brassica campestris Male Fertility 16 (BcMF16), has been isolated and characterized. The BcMF16 gene shares approximately 85% nucleotide sequence homology with two exopolygalacturonase (EC3.2.1.67) genes of Arabidopsis thaliana. Cluster analysis of polygalacturonase peptides indicate that BcMF16 belongs to the pollen polygalacturonase clade. Quantitative real-time PCR analysis has revealed that BcMF16 is specifically expressed in reproductive tissues of the fertile line of genic male sterile A/B line of Chinese cabbage pak choi, and that expression levels dramatically increased during later stages of pollen development. In situ hybridization has demonstrated that BcMF16 is specifically and transiently expressed in both tapetum and pollen following microspore separation at the tetrad stage.     

Composition and Chemical Variability of Enantia polycarpa Engl. & Diels Leaf Essential Oil from Côte d'Ivoire

The composition of Enantia polycarpa Engl . & Diels leaf essential oil has been investigated for the first time using a combination of chromatographic and spectroscopic techniques. The compositions of 52 leaf essential oil samples have been subjected to statistical analysis, hierarchical cluster analysis (HCA) and principal component analysis (PCA). Four groups were differentiated, of which the compositions were dominated by β‐elemene and germacrene B (Group III, 22/52 samples); germacrene D (Group I, 16/52 samples); β‐cubebene (Group IV, 8/52 samples) and by germacrene B and germacrene D (Group II, 6/52 samples). A special attention was brought to the quantification of the thermolabile components, germacrene A, germacrene B and germacrene C, as well as that of their rearranged compounds, β‐elemene, γ‐elemene and δ‐elemene. ¹³C‐NMR data of β‐cubebene have been provided.     

Albertania and Egbenema gen. nov. from Nigeria and the United States,expanding biodiversity in the Oculatellaceae (cyanobacteria)

Knowledge of the tropical terrestrial cyanobacterial flora from the African continent is still limited. Of 31 strains isolated from soil and subaerial samples collected in Lagos State, Nigeria, three were found to be in the Oculatellaceae, including two species in a new genus. Subsequently, isolates from microbial mats in White Sands National Park in New Mexico, United States, and from a rock near the ocean in Puerto Rico, United States, were found to belong to the new genus as well. Cyanobacterial isolates were characterized microscopically, sequenced for the 16S rRNA gene and associated ITS region, and phylogenetically analyzed. Egbenema gen. nov., with three new species, as well as two new species of Albertania were differentiated from all other Oculatellaceae. Both genera belong to a supported clade within the Oculatellaceae that includes Trichotorquatus and Komarkovaea. The two new species of Albertania, A. egbensis and A. latericola, were from the same sample, but were evolutionarily separate based on 16S rRNA gene phylogenies, percent identity below the 98.7% threshold, and ITS rRNA percent dissimilarity >7.0%. Egbenema aeruginosum gen. et sp. nov. was phylogenetically separated from Trichotorquatus and Albertania but was in a clade with other strains belonging to Egbenema. The two Egbenema strains from the United States are here named Egbenema epilithicum sp. nov. and Egbenema gypsiphilum sp. nov. Our results support the hypothesis that further species discoveries of novel cyanobacteria will likely be made in soils and subaerial habitats, as these habitats continue to be studied, both in tropical and temperate biomes.     

Characterization of Leptospira species isolated from soil collected in Japan

Leptospira were isolated from soil obtained from Hokkaido, the northernmost island, to Okinawa, the southernmost island, of Japan using sulfamethoxazole, trimethoprim, amphotericin B, fosfomycin, and 5‐ fluorouracil. Fifty of 132 soil samples (37.9%) were culture‐positive. On the basis of 16S‐rDNA sequences, 12 of the isolated Leptospira were classified into a pathogenic species clade that is closely associated with L. alstonii and L. kmetyi. Nine isolates were classified as intermediate species and were found to be similar to L. licerasiae. Twenty‐seven isolates were classified as non‐pathogenic species, of which 23 were found to be related to L. wolbachii. Non‐pathogenic Leptospira are commonly distributed in environmental soil.     

Phycicoccus ochangensis sp. nov., isolated from soil of a potato cultivation field

Two novel, Gram-positive, motile, coccal bacteria, strains L1b-b9^T and B5a-b5, were isolated from a potato cultivation field in Ochang, Korea. These isolates grew at 10–45°C, pH 5.0–10.0, and in the presence of 8% (w/v) NaCl. The diagnostic diamino acid in the cell-wall peptidoglycan was meso-diaminopimelic acid. The major menaquinone was MK-8(H₄) and the main cellular fatty acids were iso-C_14:0, iso-C_15:0, and anteiso-C_15:0. Polar lipids in strain L1b-b9^T consisted of diphosphatidylglycerol, phosphatidylglycerol, phosphatidylinositol, and an unknown glyco-amino lipid. The G+C content of genomic DNA was 73.6 mol%. A phylogenetic analysis based on 16S rRNA gene sequences showed that strains L1b-b9^T and B5a-b5 shared 99.36% similarity and formed a robust clade with the type species of the genus Phycicoccus. Strain L1b-b9^T is related most closely to Phycicoccus cremeus V2M29^T (97.52% 16S rRNA gene sequence similarity). On the basis of phylogenetic characteristics, the name Phycicoccus ochangensis sp. nov. is proposed for strain LIb-b9^T (=KCTC 19694^T =JCM 17595^T).     

Phylogenetic relationships of Pterocarya (Juglandaceae) with an emphasis on the taxonomic status of Iranian populations using ITS and trnH-psbA sequence data

Pterocarya fraxinifolia (Lam.) Spach., a relict tree species of the Juglandaceae family, is native to the Great Caucasus, Anatolia, and to the Hyrcanian forests of the southern Azerbaijan and Northern Iran. In this study, the phylogenetic relationship of the species, sampled in selected Iranian populations, and the global biogeography of the genus Pterocarya were addressed. Leaves were collected from 8 to 10 trees from three geographically isolated habitats. The samples were analyzed with nuclear (internal transcribed spacer [ITS] regions) and chloroplast (trnH-psbA) DNA markers. The obtained results were compared and analyzed with the data registered in NCBI GenBank. It is reported that the ITS regions varied from 644 to 652 for Pterocarya genus, but we did not observe polymorphisms for Iranian Pterocarya. The phylogenetic tree divided the Pterocarya genus in three clades: clade 1 grouping exclusively the samples P. fraxinifolia, clearly separated from the East Asiatic taxa; clade 2 that includes the species P. hupehensis and P. macroptera; clade 3 clustering P. stenoptera and P. tonkinensis. Although the Iranian Pterocarya samples and P. fraxinifolia from the Caucasus were in the same clade, they presented two different secondary structures. The Iranian populations showed the maximum genetic distance with P. stenoptera and P. tonkinensis. Our analysis demonstrates that the traditional division of all the six species sampled throughout their distribution area as well as the phylogeny of the genus Pterocarya needs to be reviewed.