Microbotryum Violaceum Research Articles

Bioactive Aromatic Derivatives from Endophytic Fungus, Cytospora sp

Two new benzyl gamma-butyrolactone analogues, (R)-5-((S)-hydroxy(phenyl)-methyl)dihydrofuran-2(3H)-one (1) and its 6-acetate (2), and a new naphthalenone derivative (8), together with eight additional known aromatic derivatives, (S)-5-((S)-hydroxy(phenyl)-methyl)dihydrofuran-2(3H)-one (3), (S)-5-benzyl-dihydrofuran-2(3H)-one (4), 5-phenyl-4-oxopentanoic acid (5), gamma-oxo-benzenepentanoic acid methyl ester (6), 3-(2,5-dihydro-4-hydroxy-5-oxo-3-phenyl-2-furyl)propionic acid (7), (3R)-5-methylmellein (9), integracins A (10) and B (11) were isolated from Cytospora sp., an endophytic fungus isolated from Ilex canariensis from Gomera. The structures of these compounds were elucidated by detailed spectroscopic analysis, comparison with reported data, and chemical interconversion. The absolute configurations of the new compounds (1, 2, 8) were established on the basis of optical rotation or CD spectra analysis. Preliminary studies showed antimicrobial activity of these compounds against the fungi Microbotryum violaceum, Botrytis cinerea and Septoria tritici, the alga Chlorella fusca, and the bacterium Bacillus megaterium.

Three New Antimicrobial Metabolites from the Endophytic Fungus Phomopsis sp.

AbstractTwo new chromones, phomochromone A and B (1 and 2), and one new natural cyclopentenone derivative, phomotenone (3), together with six known compounds, phomosines A–D (4–7), (1S,2S,4S)‐trihydroxy‐p‐menthane (8), and 5‐methylmellein (9), have been isolated from the endophytic fungus Phomopsis sp. through a bioassay‐guided procedure. The structures of the new compounds were assigned on the basis of the 1H and 13C NMR spectra, DEPT, and 2D COSY, HMQC, HMBC, and NOESY experiments, supported by molecular modelling. The absolute configurations of compounds 1–3 were assigned by TD‐DFT CD calculations and, surprisingly, the two related phomochromones A and B have an opposite configuration at C‐2. Compounds 1–3 show good antifungal (Microbotryum violaceum), antibacterial (Escherichia coli, Bacillus megaterium), and antialgal (Chlorella fusca) activities. Compound 8 shows good antialgal and antibacterial activity.

Diversonol and Blennolide Derivatives from the Endophytic Fungus Microdiplodia sp.: Absolute Configuration of Diversonol

Chemical investigation of the fungal strain Microdiplodia sp. isolated from the shrub Lycium intricatum led to the isolation of four new compounds: a hexahydroxanthone (2), a 2,3-dihydrochroman-4-one (3), a 7-oxoxanthone derivative (4), and a 1,4-oxazepan-7-one (5). The relative configurations of the new compounds were determined by intensive NMR investigations, notably NOESY experiments at different temperatures. The absolute configurations of the well-known fungal metabolite diversonol (1) and of other xanthone derivatives (3, 4) were established by means of TDDFT ECD calculations. Most of the metabolites were biologically active, with antibacterial activity against Legionella pneumophila and/or antifungal activity against Microbotryum violaceum.

Synthesis of Biologically Active Nonnatural Palmarumycin Derivatives

AbstractDiels–Alder reaction of benzoquinone monoacetal 1 with 1‐methoxy‐1,3‐butadiene (4) and dimethyl(1‐methyl‐3‐vinylcyclohex‐3‐enyl)(phenyl)silane (5) gave the expected adducts 2 and 3, whereas with siloxy‐dienes such as 3‐methyl‐1‐(trimethylsiloxy)‐1,3‐butadiene (6) a 1,2‐aldol‐type adduct 9 was isolated. The Diels–Alder adduct 2 was isomerized under mild basic conditions to the olefin 10 and eliminated to the diene 11 under mild acidic or thermal conditions. The olefins 2, 10 and 11 were subjected to epoxidation with m‐chloroperbenzoic acid and tert‐butyl hydroperoxide under mild basic conditions to afford the epoxides 12, 15, and 18 under the former conditions and 13, 14, 16, 17, 19, and 20 under the latter conditions. The olefins 1, 2, 10, 11, and 20 were also transformed into the respective cis‐diols 21a and 23a–26a. From theses diols the corresponding diacetates (b) andacetonides (c) were prepared. The compounds were tested against the Gram‐negative bacterium Escherichia coli, the Gram‐positive bacterium Bacillus megaterium, and the fungus Microbotryum violaceum. Whereas compound 25b was the most active of the diacetates and the epoxide 18 the most active of all substances, all of the compounds were biologically active against the three test organisms, equalling and surpassing those of the natural palmarumycins.

Viburspiran, an Antifungal Member of the Octadride Class of Maleic Anhydride Natural Products

AbstractViburspiran (8), a structurally new member of the maleic anhydride natural products, was isolated, along with seven known compounds 9–15, from the fungal endophyte Cryptosporiopsis sp. The structure of the new compound was elucidated by spectroscopic methods and confirmed by single‐crystal X‐ray analysis. The absolute configuration was established by TDDFT CD calculations by using the solid‐state structure for input geometry. Viburspiran (8) is the first representative of a new class of eight‐membered maleic anhydride natural products, named octadrides and exhibited antifungal activity against Botrytis cinerea and Microbotryum violaceum.

DNA polymorphism in recombining and non-recombining mating-type-specific loci of the smut fungus Microbotryum.

The population-genetic processes leading to the genetic degeneration of non-recombining regions have mainly been studied in animal and plant sex chromosomes. Here, we report population genetic analysis of the processes in the non-recombining mating-type-specific regions of the smut fungus Microbotryum violaceum. M. violaceum has A1 and A2 mating types, determined by mating-type-specific 'sex chromosomes' that contain 1-2 Mb long non-recombining regions. If genetic degeneration were occurring, then one would expect reduced DNA polymorphism in the non-recombining regions of this fungus. The analysis of DNA diversity among 19 M. violaceum strains, collected across Europe from Silene latifolia flowers, revealed that (i) DNA polymorphism is relatively low in all 20 studied loci (π∼0.15%), (ii) it is not significantly different between the two mating-type-specific chromosomes nor between the non-recombining and recombining regions, (iii) there is substantial population structure in M. violaceum populations, which resembles that of its host species, S. latifolia, and (iv) there is significant linkage disequilibrium, suggesting that widespread selfing in this species results in a reduction of the effective recombination rate across the genome. We hypothesise that selfing-related reduction of recombination across the M. violaceum genome negates the difference in the level of DNA polymorphism between the recombining and non-recombining regions, and may possibly lead to similar levels of genetic degeneration in the mating-type-specific regions of the non-recombining 'sex chromosomes' and elsewhere in the genome.

Genetic polymorphism of Microbotryum violaceum s. l. Isolates collected from different plant species on the territory of Russia

The present-day studies in the field of systematics and phylogeny of microorganisms, fungi, in particular, are characterized by a wide use of new approaches and methods of molecular biology. The use of a diversity of genetic markers permits a distinct differentiation of closely related species into individual evolutionary independent lines. It is shown in this work that all Microbotryum violaceum s. l. isolates studied by us are divided into five evolutionary groups according to the host plant.

Psorantin, a unique methylene linked dimer of vismin and kenganthranol E, two anthranoid derivatives from the fruits of Psorospermum aurantiacum (Hypericaceae)

Two prenylated anthranoids, psorantin and kenganthranol E, were isolated from the fruit of Psorospermum aurantiacum together with the known compounds ferruginin B, vismin, vismion D, haronginanthrone, kenganthranol B, kenganthraquinone, 1,7-dihydroxyxanthone, paradisiol, fridelin, fridelinol and betulinic acid. Their structures were determined on the basis of spectral data and by comparison with data reported in the literature as well as with authentic specimens for the known compounds. The structures of the new compounds were determined as bis-[3,8,9-trihydroxy-6-methyl-4,4-bis-(3,3-dimethylallyl)anthracenyl]methane ( 1) and 1,8,10-trihydroxy-6-methyl-4,5-bis-(3,3-dimethylallyl)-2,3-(2,2-dimethylpyrano)anthrone ( 2). Psorantin ( 1) is a dimer of vismin formed through a methylene linkage. The two new compounds when tested against the microbial strains Bacillus megaterium, Escherichia coli, Chlorella fusca and Microbotryum violaceum showed no activity.

Using phylogenies of pheromone receptor genes in the Microbotryum violaceum species complex to investigate possible speciation by hybridization

Several cases of speciation by hybridization have been reported in fungi, mostly involving recent hybridization between closely related species. In the basidiomycete genus Microbotryum by contrast some species were suspected to have arisen by hybridization between moderately distant species. In particular two species, M. lagerheimii and M. silenes-acaulis, had different placements in phylogenetic trees depending on the genes considered. Microbotryum species exhibit bipolar heterothallism, and here we analyzed sequences of the two alternate pheromone receptors to obtain further insights on the occurrence of hybridization. Indeed because mating-type loci are always heterozygous homoploid hybrid speciation should leave a permanent footprint at the mating-type locus by retaining the alternate alleles from their respective parental species. The trees obtained with each of the two pheromone receptors were well resolved, and the species relationships were in agreement with published phylogenies. Fungal pheromone receptor genes of basidiomycetes thus appear useful for phylogenetic studies, although it may not be true for the homobasidiomycetes where duplications of these genes have occurred. Furthermore an incongruence between the phylogenies of the two pheromone receptors was found for one species, M. lagerheimii, as previously observed between other nuclear genes. However additional species analyzed here revealed that the incongruence involved the whole clade including both M. lagerheimii and the Microbotryum species parasitizing Lychnis flos-cucucli. The ancestor of these species thus possibly arose via hybridization between distant ancestral lineages, although further studies should address alternative hypotheses, such as chance events during lineage sorting.

The impact of genome defense on mobile elements in Microbotryum

Repeat induced point mutation (RIP), a mechanism causing hypermutation of repetitive DNA sequences in fungi, has been described as a 'genome defense' which functions to inactivate mobile elements and inhibit their deleterious effects on genome stability. Here we address the interactions between RIP and transposable elements in the Microbotryum violaceum species complex. Ten strains of M. violaceum, most of which belong to different species of the fungus, were all found to contain intragenomic populations of copia-like retrotransposons. Intragenomic DNA sequence variation among the copia-like elements was analyzed for evidence of RIP. Among species with RIP, there was no significant correlation between the frequency of RIP-induced mutations and inferred transposition rate based on diversity. Two strains of M. violaceum, from two different plant species but belonging to the same fungal lineage, contained copia-like elements with very low diversity, as would result from a high transposition rate, and these were also unique in showing no evidence of the hypermutation patterns indicative of the RIP genome defense. In this species, evidence of RIP was also absent from a Class II helitron-like transposable element. However, unexpectedly the absolute repetitive element load was lower than in other strains.

Ring B Aromatic Steroids from an Endophytic Fungus, Colletotrichum sp.

The new (22E,24R)-3-acetoxy-19(10-->6)-abeo-ergosta-5,7,9,22-tetraen-3beta-ol (1) and the known (22E,24R)-19(10-->6)-abeo-ergosta-5,7,9,22-tetraen-3beta-ol (2), two interesting ergosteroids with rare aromatized ring B, together with seven known derivatives, namely (22E,24R)-ergosta-5,7,22-trien-3beta-ol (3), (22E,24R)-ergosta-4,7,22-trien-3-one (4), (22E,24R)-ergosta-4,6,8(14),22-tetraen-3-one (5), (22E,24R)-5alpha,8alpha-epidioxyergosta-6,22-dien-3beta-ol (6), (22E,24R)-ergosta-7,22-dien-3beta,5alpha,6beta-triol (7), (22E,24R)-6-acetoxy-ergosta-7,22-dien-3beta,5alpha,6beta-triol (8), and (22E,24R)-3,6-diacetoxy-ergosta-7,22-dien-3beta,5alpha,6beta-triol (9), were isolated from Colletotrichum sp., an endophytic fungus isolated from Ilex canariensis from Gomera. The structures of these compounds were elucidated by detailed spectroscopic analysis, comparison with reported data, and chemical interconversion. The isolation of these metabolites not only displays a beautiful array of chemical diversity, but also gives insight into the biosynthetic interconnections. Preliminary studies showed antimicrobial activity of these compounds against the fungus Microbotryum violaceum, the alga Chlorella fusca, and the bacteria Escherichia coli and Bacillus megaterium.

Evolutionary Strata in a Small Mating-Type-Specific Region of the Smut Fungus Microbotryum violaceum

DNA sequence analysis and genetic mapping of loci from mating-type-specific chromosomes of the smut fungus Microbotryum violaceum demonstrated that the nonrecombining mating-type-specific region in this species comprises approximately 25% ( approximately 1 Mb) of the chromosome length. Divergence between homologous mating-type-linked genes in this region varies between 0 and 8.6%, resembling the evolutionary strata of vertebrate and plant sex chromosomes.

A SUPERMAN-like Gene is Exclusively Expressed in Female Flowers of the Dioecious Plant Silene latifolia

To elucidate the mechanism(s) underlying dioecious flower development, the present study analyzed a SUPERMAN (SUP) homolog, SlSUP, which was identified in Silene latifolia. The sex of this plant is determined by heteromorphic X and Y sex chromosomes. It was revealed that SlSUP is a single-copy autosomal gene expressed exclusively in female flowers. Introduction of a genomic copy of SlSUP into the Arabidopsis thaliana sup (sup-2) mutant complemented the excess-stamen and infertile phenotypes of sup-2, and the overexpression of SlSUP in transgenic Arabidopsis plants resulted in reduced stamen numbers as well as the suppression of petal elongation. During the development of the female flower in S. latifolia, the expression of SlSUP is first detectable in whorls 2 and 3 when the normal expression pattern of the B-class flowering genes was already established and persisted in the stamen primordia until the ovule had matured completely. In addition, significant expression of SlSUP was detected in the ovules, suggestive of the involvement of this gene in ovule development. Furthermore, it was revealed that the de-suppression of stamen development by infection of the S. latifolia female flower with Microbotryum violaceum was accompanied by a significant reduction in SlSUP transcript levels in the induced organs. Taken together, these results demonstrate that SlSUP is a female flower-specific gene and suggest that SlSUP has a positive role in the female flower developmental pathways of S. latifolia.

Hybrid sterility and inviability in the parasitic fungal species complex Microbotryum

Microbotryum violaceum, the anther-smut fungus, forms a complex of sibling species which specialize on different plants. Previous studies have shown the presence of partial ecological isolation and F1 inviability, but did not detect assortative mating apart from a high selfing rate. We investigated other post-mating barriers and show that F1 hybrid sterility, the inability of gametes to mate, increased gradually with the increasing genetic distance between the parents. F2 hybrids showed a reduced ability to infect the plants that was also correlated with the genetic distance. The host on which the F2 hybrids were passaged caused a selection for alleles derived from the pathogen species originally isolated from that host, but this effect was not detectable for the most closely related species. The post-mating barriers thus remain weak among the closest species pairs, suggesting that premating barriers are sufficient to initiate divergence in this system.

Disease Status and Population Origin Effects on Floral Scent: Potential Consequences for Oviposition and Fruit Predation in A Complex Interaction Between A Plant, Fungus, and Noctuid Moth

In the Silene latifolia-Hadena bicruris nursery pollination system, the Hadena moth is both pollinator and seed predator of its host plant. Floral scent, which differs among S. latifolia individuals and populations, is important for adult Hadena to locate its host. However, the success of moth larvae is strongly reduced if hosts are infected by the anther smut fungus Microbotryum violaceum, a pathogen that is transmitted by flower visitors. There were no qualitative differences between the scent of flowers from healthy and diseased plants. In addition, electroantennographic measurements showed that Hadena responded to the same subset of 19 compounds in samples collected from healthy and diseased plants. However, there were significant quantitative differences in scent profiles. Flowers from diseased plants emitted both a lower absolute amount of floral scent and had a different scent pattern, mainly due to their lower absolute amount of lilac aldehyde, whereas their amount of (E)-beta-ocimene was similar to that in healthy flowers. Dual choice behavioral wind tunnel tests using differently scented flowers confirmed that moths respond to both qualitative and quantitative aspects of floral scent, suggesting that they could use differences in floral scent between healthy and infected plants to discriminate against diseased plants. Population mean fruit predation rates significantly increased with population mean levels of the emission rates of lilac aldehyde per flower, indicating that selection on floral scent compounds may not only be driven by effects on pollinator attraction but also by effects on fruit predation. However, variation in mean emission rates of scent compounds per flower generally could not explain the higher fruit predation in populations originating from the introduced North American range compared to populations native to Europe.

New Bioactive 2,3‐Epoxycyclohexenes and Isocoumarins from the Endophytic Fungus Phomopsis sp. from Laurus Azorica

AbstractSix new metabolites, namely cycloepoxylactone (1a), cycloepoxytriol A (2), cycloepoxytriol B (3) and phomolactones A–C (4–6), were isolated from two cultures of the ethyl acetate soluble fraction of the endophytic fungus Phomopsis sp. (internal strain no. 7233). Their structures were determined by means of spectroscopic data including HREIMS, 1H NMR, 13C NMR, and 2D NMR (HMQC, HMBC, and NOESY). The absolute configurations of 1a, 4 and 5 were determined by circular dichroism and comparison of solution CD spectra with TDDFT‐calculated ones. Preliminary studies showed that cycloepoxylactone (1a) has good antibacterial, antifungal, and algicidal activity against Bacillus megaterium, Microbotryum violaceum, and Chlorella fusca, respectively, whereas cycloepoxytriol B (3) has good algicidal activity against Chlorella fusca.(© Wiley‐VCH Verlag GmbH & Co. KGaA, 69451 Weinheim, Germany, 2009)

Diversity of Antimicrobial Pyrenophorol Derivatives from an Endophytic Fungus, Phoma sp.

AbstractPyrenophorol (1) and (–)‐dihydropyrenophorin (3), two known macrodiolides, were isolated together with four new analogues (2, 4–6) and three ring‐opened derivatives (7–9) from Phoma sp., an endophytic fungus isolated from Lycium intricatum from Gomera. The structures of the new compounds were elucidated by detailed spectroscopic analysis, comparison with reported data, and chemical interconversion. The absolute configurations were determined by chemical correlations and a modified Mosher's method. The diversity of these seven newly discovered metabolites not only extends the pyrenophorol macrocyclic family, but also gives insight into the biosynthetic interconnections, in particular by isolation of the open‐chain precursors. Preliminary studies showed antimicrobial activity of these compounds against the fungus Microbotryum violaceum, the alga Chlorellafusca, and the bacteria Escherichia coli and Bacillus megaterium.(© Wiley‐VCH Verlag GmbH & Co. KGaA, 69451 Weinheim, Germany, 2008)

Spatial and Temporal Heterogeneity Explain Disease Dynamics in a Spatially Explicit Network Model

There is an increasing recognition that individual-level spatial and temporal heterogeneity may play an important role in metapopulation dynamics and persistence. In particular, the patterns of contact within and between aggregates (e.g., demes) at different spatial and temporal scales may reveal important mechanisms governing metapopulation dynamics. Using 7 years of data on the interaction between the anther smut fungus (Microbotryum violaceum) and fire pink (Silene virginica), we show how the application of spatially explicit and implicit network models can be used to make accurate predictions of infection dynamics in spatially structured populations. Explicit consideration of both spatial and temporal organization reveals the role of each in spreading risk for both the host and the pathogen. This work suggests that the application of spatially explicit network models can yield important insights into how heterogeneous structure can promote the persistence of species in natural landscapes.

Bioactive Isocoumarins Isolated from the Endophytic Fungus Microdochium bolleyi

Three new isocoumarin derivatives ( 2- 4) were isolated together with monocerin ( 1) from Microdochium bolleyi, an endophytic fungus from Fagonia cretica, a herbaceous plant of the semiarid coastal regions of Gomera. Compounds 2 and 3 are both 12-oxo epimers of 1, and 4 is a ring-opened derivative of 1. The structures were elucidated by detailed spectroscopic analysis and comparison with reported data. The absolute configurations were determined by a modified Mosher's method. Compounds 1, 3, and 4 showed good antifungal, antibacterial, and antialgal activities against Microbotryum violaceum, Escherichia coli, Bacillus megaterium, and Chlorella fusca. Compound 2 was moderately antifungal and antialgal.

New Mono‐ and Dimeric Members of the Secalonic Acid Family: Blennolides A–G Isolated from the Fungus Blennoria sp.

Blennolides A-G (2-8), seven unusual chromanones, were isolated together with secalonic acid B (1) from Blennoria sp., an endophytic fungus from Carpobrotus edulis. This is the first reported isolation of the blennolides 2 and 3 (hemisecalonic acids B and E), the existence of which as the monomeric units of the dimeric secalonic acids had long been postulated. A compound of the proposed structure 4 (beta-diversonolic ester) will need to be revised, as its reported data do not fit those of the established structure of blennolide C (4). Other monomers, the blennolides D-F (5-7) seem to be derived from blennolides A (2) and B (3) by rearrangement of the hydroaromatic ring. The heterodimer 8, composed of the monomeric blennolide A (2) and the rearranged 11-dehydroxy derivative of blennolide E (6), extends the ergochrome family with an ergoxanthin type of skeleton. The structures of the new compounds were elucidated by detailed spectroscopic analysis and further confirmed by an X-ray diffraction study of a single crystal of 2. The absolute configurations were determined by TDDFT calculations of CD spectra, including the solid-state CD/TDDFT approach. Preliminary studies showed strong antifungal and antibacterial activities of these compounds against Microbotryum violaceum and Bacillus megaterium, respectively. They were also active against the alga Chlorella fusca and the bacterium Escherichia coli.