Pancreatic lipase inhibitory activity of monogalactosyldiacylglycerols isolated from the freshwater microalga <Emphasis Type="Italic">Chlorella sorokiniana</Emphasis>

Chemical investigation of the freshwater microalga Chlorella sorokiniana led to the isolation of a monogalactosyldiacylglycerol (MGDG)-rich fraction possessing dose-dependent inhibitory activity against pancreatic lipase activity. The MGDG-rich fraction contains 12 MGDGs identified by LC/HRMS analysis. Among them, three MGDGs were new compounds, namely, (2S)-1-O-(7Z,10Z-hexadecadienoyl)-2-O-(7Z,10Z,13Z-hexadecatrienoyl)-3-O-β-D-galactopyranosylglycerol (1), (2S)-1-O-linoleoyl-2-O-(7Z,10Z-hexadecadienoyl)-3-O-β-D-galactopyranosylglycerol (6), and (2S)-1-O-oleoyl-2-O-(7Z,10Z-hexadecadienoyl)-3-O-β-D-galactopyranosylglycerol (8). The major galactolipids were isolated by semipreparative HPLC and tested for their effect toward pancreatic lipase inhibitory activity. All the tested MGDGs showed significant reduction of pancreatic lipase activity indicating possible beneficial use for management of lipase-related disorders such as obesity.     

Specificity of oxidation of linoleic acid homologs by plant lipoxygenases

The lipoxygenase-catalyzed oxidation of linoleic acid homologs was studied. While the linoleic acid oxidation by maize 9-lipoxygenase (9-LO) specifically produced (9S)-hydroperoxide, the dioxygenation of (11Z,14Z)-eicosadienoic (20:2) and (13Z,16Z)-docosadienoic (22:2) acids by the same enzyme lacked regio- and stereospecificity. The oxidation of 20:2 and 22:2 by 9-LO afforded low yields of racemic 11-, 12-, 14-, and 15-hydroperoxides or 13- and 17-hydroperoxides, respectively. Soybean 13-lipoxygenase-1 (13-LO) specifically oxidized 20:2, 22:2, and linoleate into (ω6S)-hydroperoxides. Dioxygenation of (9Z,12Z)-hexadecadienoic acid (16:2) by both 9-LO and 13-LO occurred specifically, affording (9S)- and (13S)-hydroperoxides, respectively. The data are consistent with the “pocket theory of lipoxygenase catalysis” (i.e. with the penetration of a substrate into the active center with the methyl end first). Our findings also demonstrate that the distance between carboxyl group and double bonds substantially determines the positioning of substrates within the active site.     

Developmental regulators in <Emphasis Type="Italic">Aspergillus fumigatus</Emphasis>

The filamentous fungus Aspergillus fumigatus is the most prevalent airborne fungal pathogen causing severe and usually fatal invasive aspergillosis in immunocompromised patients. This fungus produces a large number of small hydrophobic asexual spores called conidia as the primary means of reproduction, cell survival, propagation, and infectivity. The initiation, progression, and completion of asexual development (conidiation) is controlled by various regulators that govern expression of thousands of genes associated with formation of the asexual developmental structure conidiophore, and biogenesis of conidia. In this review, we summarize key regulators that directly or indirectly govern conidiation in this important pathogenic fungus. Better understanding these developmental regulators may provide insights into the improvement in controlling both beneficial and detrimental aspects of various Aspergillus species.     

Revision of Jaegeria (Compositae-Heliantheae)

The genusJaegeria consists of a diploid-polyploid complex of eight species found in moist habitats from Mexico to South America. The distributions, habitats, generic affinities, and tribal placement are discussed. Chromosome numbers were determined for all species exceptJ. axillaris, which is known only from Colombia.J. glabra andJ. pedunculata are diploid (2n = 18) ;J. hirta, J. macrocephala, J. gracilis, andJ. crassa are tetraploid (2n = 36) ;J. bellidiflora is octoploid (2n = 72). One Galápagos Island endemic,J. gracilis, is quite similar to the common mainland species,J. hirta, but the other endemic,J. crassa, is quite distinct from any other species.J. bellidijlora is most similar morphologically toJ. glabra;J. macrocephala is allied toJ. pedunculata. J. axillaris is somewhat distinct from the other species.     

Effects of transgenic expression of <Emphasis Type="Italic">Brevibacterium linens</Emphasis> methionine gamma lyase (MGL) on accumulation of <Emphasis Type="Italic">Tylenchulus semipenetrans and key aminoacid contents</Emphasis> in <Emphasis Type="Italic">Carrizo citrange</Emphasis>

Alpha-dioxygenases (α-DOX) catalyzing the primary oxygenation of fatty acids to oxylipins were recently found in plants. Here, the biological roles of the pepper α-DOX (Ca-DOX) gene, which is strongly induced during non-host pathogen infection in chili pepper, were examined. Virus-induced gene silencing demonstrated that down-regulation of Ca-DOX enhanced susceptibility to bacterial pathogens and suppressed the hypersensitive response via the suppression of pathogenesis-related genes such as PR4, proteinase inhibitor II and lipid transfer protein (PR14). Ca-DOX-silenced pepper plants also exhibited more retarded growth with lower epidermal cell numbers and reduced cell wall thickness than control plants. To better understand regulation of Ca-DOX, transgenic Arabidopsis plants harboring the β-glucuronidase (GUS) reporter gene driven from a putative Ca-DOX promoter were generated. GUS expression was significantly induced upon avirulent pathogen infection in transgenic Arabidopsis leaves, whereas GUS induction was relatively weak upon virulent pathogen treatment. After treatment with plant hormones, early and strong GUS expression was seen after treatment of salicylic acid, whereas ethylene and methyl jasmonate treatments produced relatively weak and late GUS signals. These results will enable us to further understand the role of α-DOX, which is important in lipid metabolism, defense responses, and growth development in plants.     

Molecular phylogenetics and anti-<Emphasis Type="Italic">Pythium</Emphasis> activity of endophytes from rhizomes of wild ginger congener, <Emphasis Type="Italic">Zingiber zerumbet</Emphasis> Smith

Zingiber zerumbet, a perennial rhizomatous herb exhibits remarkable disease resistance as well as a wide range of pharmacological activities. Towards characterizing the endophytic population of Z. zerumbet rhizomes, experiments were carried out during two different growing seasons viz., early-June of 2013 and late-July of 2014. A total of 34 endophytes were isolated and categorized into 11 morphologically distinct groups. Fungi were observed to predominate bacterial species with colonization frequency values ranging from 12.5 to 50 %. Among the 11 endophyte groups isolated, molecular analyses based on ITS/16S rRNA gene sequences identified seven isolate groups as Fusarium solani, two as F. oxysporum and one as the bacterium Rhizobium spp. Phylogenetic tree clustered the ITS sequences from Z. zerumbet endophytes into distinct clades consistent with morphological and sequence analysis. Dual culture assays were carried out to determine antagonistic activity of the isolated endophytes against Pythium myriotylum, an economically significant soil-borne phytopathogen of cultivated ginger. Experiments revealed significant P. myriotylum growth inhibition by F. solani and F. oxysporum isolates with percentage of inhibition (PoI) ranging from 45.17 ± 0.29 to 62.2 ± 2.58 with F. oxysporum exhibiting higher PoI values against P. myriotylum. Using ZzEF8 metabolite extract, concentration-dependent P. myriotylum hyphal growth inhibition was observed following radial diffusion assays. These observations were confirmed by scanning electron microscopy analysis wherein exposure to ZzEF8 metabolite extract induced hyphal deformities. Results indicate Z. zerumbet endophytes as promising resources for biologically active compounds and as biocontrol agents for soft rot disease management caused by Pythium spp.     

Four new species of <Emphasis Type="Italic">Tubeufia</Emphasis> (Tubeufiaceae,Tubeufiales) from Thailand

Tubeufia was named for a sexual morph taxon. However, several asexual morph species have been accommodated in this genus as well. In our study, four new species of Tubeufia, viz. T. filiformis, T. latispora, T. laxispora and T. mackenziei, are described and illustrated. The phylogenetic placement of the new species is confirmed by analysis of combined ITS, LSU and TEF1α sequence data. A key to Tubeufia species is provided and the new species are compared with similar fungal taxa.     

GPH1 is involved in glycerol accumulation in the three-dimensional networks of the nematode-trapping fungus <Emphasis Type="Italic">Arthrobotrys oligospora</Emphasis>

Turgor is very important for the invasive growth of fungal pathogens. Glycerol, a highly osmotic solvent, is considered to play an important role in turgor generation. The nematophagous fungus Arthrobotrys oligospora mainly lives as a saprophyte. In the presence of nematodes, A. oligospora enters the parasitic stage by forming three-dimensional networks (traps) to capture nematodes. In A. oligospora, we found that glycerol accumulated during nematode-induced trap formation. We demonstrated that deleting gph1, which encodes glycogen phosphorylase, decreased the glycerol content, compared with that of a wild-type strain. Although the number of traps induced by nematodes was not affected in the Δgph1 mutant, the capture rate was lower. Meanwhile, deleting gph1 also affected the growth rate and conidiation capacity of the fungus. These results indicate that glycerol derived from GPH1 is essential for the full virulence of A. oligospora against nematodes.     

Chemical and molecular identification of the invasive termite <Emphasis Type="Italic">Zootermopsis nevadensis</Emphasis> (Isoptera: Archotermopsidae) in Japan

Numerous termite species have been introduced outside their native ranges by human transport, and some have become invasive. The dampwood termite Zootermopsis nevadensis (Hagen), which is native to western North America, has been introduced to and become established in Kawanishi City, Hyogo Prefecture, Japan. Zootermopsis nevadensis is subdivided into two subspecies based on cuticular hydrocarbon (CHC) phenotypes: Z. nevadensis nevadensis and Z. nevadensis nuttingi (Haverty and Thorne). Here, we identified Z. nevadensis in Japan as hybrids between the two subspecies. Chemical analysis showed the presence of 7,15-dimethylhenicosane and 5,17-dimethylhenicosane in the CHCs of Z. nevadensis in Japan, corresponding to the CHC phenotype of Z. n. nevadensis. Conversely, all mitochondrial cytochrome c oxidase subunit I sequences of Z. nevadensis in Japan were identical to sequences from Z. n. nuttingi and hybrids between the two subspecies from a native hybrid zone in California, USA. In addition, phylogenetic analysis showed that Z. nevadensis in Japan formed a clade with Z. n. nuttingi and hybrids between the two subspecies. Our results show discordance between the chemical and genetic features of Z. nevadensis in Japan, indicating that individuals of Z. nevadensis in Japan are hybrids between the two subspecies.     

A taxonomic and phylogenetic study of the <Emphasis Type="Italic">Golovinomyces biocellatus</Emphasis> complex (Erysiphales,Ascomycota) using asexual state morphology and rDNA sequence data

The Golovinomyces biocellatus complex consists of powdery mildew (Erysiphales) species restricted to hosts of the family Lamiaceae. Previous authors used minor morphological features of the sexual state and host range data to split the complex. The data, however, were not sufficient to define a convincing species concept. Our taxonomic study is based on molecular phylogenetic and asexual state morphology data. For morphological studies, mainly features of the asexual morph (conidiophores, conidia, germination patterns) were studied using light and scanning electron microscopy. Detailed line drawings of asexual state features are provided. For phylogenetic analyses, two markers (rDNA: ITS, LSU) of 64 specimens were applied. The phylogeny resulted in two major clades. Clade I consists of specimens with Lamiaceae hosts and three specimens of Verbena. Clade II consists of two sister groups, the first (IIa) with Salvia spp. and the second (IIb) with Lycopus europaeus (the type host G. biocellatus) and Glechoma. Clades I and IIb and two subclades of IIa with Salvia hosts are characterized by specific morphological traits (differences in conidiophore length, conidial shape, width, and germination patterns). Based on these data, we suggest to consider specimens of clades I (including specimens on Verbena) and IIb and the two subclades of IIa as distinct species, namely G. monardae, G. biocellatus, G. salviae, and G. neosalviae sp. nov. A key for the identification of species based on asexual state features is provided. The results are discussed with respect to host range, jumps, co-evolutionary aspects, and distribution patterns.     

<Emphasis Type="Italic">Helicocollum</Emphasis>, a new clavicipitalean genus pathogenic to scale insects (Hemiptera) in Thailand

A new genus pathogenic to scale insects (Coccidae, Hemiptera), Helicocollum, is described from Thailand. It includes the three species, H. chanthaburiensis, H. krabiensis and H. surathaniensis. Phylogenetic inferences on the basis of partial sequences of the large subunit of the nuclear ribosomal DNA, translation elongation factor 1-α and RNA polymerase II subunit 1 place the new genus in the Clavicipitaceae, although morphological features of the phialides resemble Hirsutella in the Ophiocordycipitaceae. Within the Clavicipitaceae, Helicocollum is closely related to Aschersonia, Moelleriella, Regiocrella, and Samuelsia, all known as pathogens of scale insects. Helicocollum differs from other scale insect pathogens in having phialides with helical necks and synnematous or sporodochial conidiomata, but pycnidia. Morphological characteristics are evaluated and compared with other asexual morphs producing twisted, undulated phialidic necks or occurring on scale insects.     

Phylogenetic characterization of two novel <Emphasis Type="Italic">Kamalomyces</Emphasis> species in Tubeufiaceae (Tubeufiales)

Two novel species of bambusicolous fungi in the genus Kamalomyces, collected from northern Thailand, are described and illustrated herein. Kamalomyces bambusicola and K. thailandicus spp. nov. are typical of the genus Kamalomyces (Tubeufiaceae, Tubeufiales) and are morphologically distinct from known species with respect to their size of ascomata, asci and ascospores, ascospore septation and peridium structure, including the subiculum comprising hyphae on the host surface. Morphological examination reveals that the asexual morph of K. bambusicola is associated with its sexual morph in a subiculum forming dictyochlamydosporous conidia, which are similar to the asexual morph of Chlamydotubeufia. Phylogenetic analyses of combined LSU, ITS and TEF1-α sequence data also support these two species as distinct and confirm their phylogenetic affinities within the Tubeufiaceae. In particular, Kamalomyces shares a close phylogenetic relationship to Helicoma.     

Stress factor-induced changes in the activity of antioxidant protective mechanisms in the wild type strain of <Emphasis Type="Italic">Neurospora crassa</Emphasis> and in its photoreceptor complex mutants

The effect of stress factors (changes in oxygen content, temperature, and illumination) on superoxide dismutase (SOD) and catalase activity, as well as on the content of thiol and disulfide groups, in low-molecular-weight compounds and proteins of Neurospora crassa mycelium was studied in the wild type strain and white collar-(wc-1) and white collar-2 (wc-2) mutants. Environmental stress factors induced the activation of both SOD and catalase, as well as an increase in the thiol level in the wild type strain of Neurospora crassa. In the wc-1 and wc-2 mutants, an increase in catalase activity and in the total thiol level was revealed; however, activation of superoxide dismutase was not observed. A decrease in the formation of disulfide bonds in the proteins of wc-1 and wc-2 mutants (as compared with the wild type strain) was recorded. These results indicate disrupted transduction of stress factor signals that promotes reactive oxygen species (ROS) formation in the WCC mutants.     

An efficient method for the sequential production of lipid and carotenoids from the Chlorella Growth Factor-extracted biomass of <Emphasis Type="Italic">Chlorella vulgaris</Emphasis>

Efficient methodology for simultaneous extraction of multiple bioactive compounds from microalgae still remains a major challenge. The present study provides a method for the sequential production of three major products: Chlorella Growth Factor (CGF, a nucleotide-peptide complex enriched with vitamins, minerals, and carbohydrates), lipid, and carotenoids from Chlorella vulgaris biomass in an economically feasible manner. After protein-rich CGF was extracted, the spent biomass was found to contain 12% lipid and 3% carotenoids when extracted individually, compared to that of the un-utilized (fresh) biomass (lipid, 14%; carotenoids, 4%). When extracted simultaneously using conventional methods, the yield of lipid from “CGF and carotenoids-extracted biomass,” and carotenoids from “CGF and lipid-extracted biomass” were significantly reduced (50%). However, simultaneous extraction using different solvent mixtures such as hexane:methanol:water and pentane:methanol:water mixture-augmented lipid yield by 38.5% and carotenoids by 14%, and additionally retained chlorophyll and its derivatives. Column chromatographic approach yielded sequential production of lipid (18%), lutein (9%) with better yields as well as without chlorophyll interference. Different geometric isomers of lutein all-E-(trans)-(3R,3′R,6′R)-β,ε-carotene-3,3′diol, 9Z(cis)-(3R,3′R,6′R)-β,ε-carotene-3,3′diol, and 13Z(cis)-(3R,3′R,6′R)-β,ε-carotene-3,3′diol were purified by HPLC and elucidated by CD, UV, NMR, FT-IR, and Mass spectra. In conclusion, the study provides an efficient and economically viable methodology for sequential production of lipid and lutein along with its geometrical isomers without chlorophyll influence and yield loss from the protein-rich CGF-extracted spent biomass of marine microalga, Chlorella vulgaris.     

Characterization of the velvet regulators in <Emphasis Type="Italic">Aspergillus flavus</Emphasis>

Fungal development and secondary metabolism are closely associated via the activities of the fungal NK-kB-type velvet regulators that are highly conserved in filamentous fungi. Here, we investigated the roles of the velvet genes in the aflatoxigenic fungus Aspergillus flavus. Distinct from other Aspergillus species, the A. flavus genome contains five velvet genes, veA, velB, velC, velD, and vosA. The deletion of velD blocks the production of aflatoxin B1, but does not affect the formation of sclerotia. Expression analyses revealed that vosA and velB mRNAs accumulated at high levels during the late phase of asexual development and in conidia. The absence of vosA or velB decreased the content of conidial trehalose and the tolerance of conidia to the thermal and UV stresses. In addition, double mutant analyses demonstrated that VosA and VelB play an inter-dependent role in trehalose biosynthesis and conidial stress tolerance. Together with the findings of previous studies, the results of the present study suggest that the velvet regulators play the conserved and vital role in sporogenesis, conidial trehalose biogenesis, stress tolerance, and aflatoxin biosynthesis in A. flavus.     

Omega-6-derived oxylipin changes in serum of patients with hepatitis B virus-related liver diseases

Introduction

Chronic hepatitis B virus (HBV) infection is the main etiologic risk factor for hepatocellular carcinoma (HCC). Early studies indicated that the increase of omega-6-derived oxylipins may be involved in the pathogenesis of HBV-related HCC, yet their changes during the distinct clinical phases of chronic HBV infection remain unclear. To fill this gap, in this study we investigated the omega-6-derived oxylipin profiles in patients with three major clinical stages of chronic HBV infection (chronic hepatitis B, liver cirrhosis, and HCC).

Methods

Eighteen omega-6-derived oxylipins were quantified in serum samples of 34 patients with chronic hepatitis B, 46 patients with HBV-related liver cirrhosis, 38 patients with HBV-related HCC, and 50 healthy controls using liquid chromatography tandem mass spectrometry.

Results

Seven oxylipins were found to be altered in patients with HBV-related liver diseases, including 9,10-dihydroxyoctadecenoic acid (9,10-DiHOME), 12,13-DiHOME, 14,15-dihydroxyeicosatrienoic acid (14,15-DiHETrE), 13-hydroxyoctadecadienoic acid (13-HODE), 12-hydroxyeicosatetraenoic acid (12-HETE), 11-HETE, and thromboxane B₂ (TXB₂). Of these, three oxylipins derived from the cytochrome P450 (CYP450) pathways including 9,10-DiHOME, 12,13-DiHOME, and 14,15-DiHETrE were found to be associated with the levels of α-fetoprotein (AFP), a tumor marker. In combination with AFP, age, and gender, a combination of these seven differential oxylipins could significantly enhance the prediction of HBV-related liver diseases, particularly for liver cirrhosis (p?<?0.05).

Conclusion

This study for the first time shows the correlations between CYP450-derived oxylipins and the progression of chronic HBV infection, and sheds a new light on the surveillance of HBV-related live diseases using oxylipins.

     

A new species of <Emphasis Type="Italic">Cosmospora</Emphasis> and the first record of sexual state of <Emphasis Type="Italic">C. lavitskiae</Emphasis>

A new fungicolous species, Cosmospora inonoticola, is described and illustrated based on a specimen collected from northeast China. It is characterized by the occurrence of fruitbodies of Inonotus sp.; perithecia nonstromatic, subglobose to pyriform, not collapsing when dry; clavate asci containing (3–)6(?8) spores; ascospores ellipsoidal to broadly ellipsoidal, pale brown, finely warted, bicellular, conspicuously constricted at septum; and producing a verticillium-like asexual state. A multi-locus analysis of the combined ITS, rpb1 and tub sequences confirmed its taxonomic position, and revealed the distinction from its morphologically similar species including the type species, C. coccinea, which occurs also on Inonotus. A specimen with sexual state of C. lavitskiae is reported for the first time, and a reference specimen is indicated.