Atranones A-G, from the toxigenic mold Stachybotrys chartarum

Atranones A-G have been isolated from the toxigenic fungus Stachybotrys chartarum. These compounds contain several unusual features including an enol-lactone as part of a 3,7-dioxabicyclo[3.3.0]octane-2-one ring system fused to an 11-membered ring. Two new dolabellane diterpenes, related in structure to the atranones were also isolated, which suggests a diterpenoid origin for the C24 atranones.     

An antifungal and plant growth promoting metabolite from a sterile dark ectotrophic fungus

A sterile dark ectotrophic fungus isolated from roots of an Australian native grass, Neurachne alopecuroidea produces compound 1 in liquid cultures. The structure of the metabolite was determined by spectroscopic and X-ray diffraction studies. The metabolite shows activity against phytopathogens and plant growth promoting activity, properties that are also expressed in vivo by the ectotrophic fungus.     

The constriction of the trapping rings in Dactylaria brochopaga

The mechanism of ring constriction of a nematophagous fungus has been studied in vitro. The process of ring constriction in vivo normally requires less than one second. However, precise observations of the mechanism are difficult due to this rapidity, as well as the obstruction of the ring by the nematode. A method of initiating ring constriction without the use of an inert mechanical stimulus is described. With this method it is also possible to retard the constriction of the rings so that accurate observations of the entire process can be made. An explanation of the possible physiological mechanism is also presented.     

Anti-tumor effects of dehydroaltenusin, a specific inhibitor of mammalian DNA polymerase alpha

In the screening of selective inhibitors of eukaryotic DNA polymerases (pols), dehydroaltenusin was found to be an inhibitor of pol alpha from a fungus (Alternaria tennuis). We succeeded in chemically synthesizing dehydroaltenusin, and the compound inhibited only mammalian pol alpha with IC50 value of 0.5 microM, and did not influence the activities of other replicative pols such as pols delta and epsilon, but also showed no effect on pol alpha activity from another vertebrate, fish, or from a plant species. Dehydroaltenusin also had no influence on the other pols and DNA metabolic enzymes tested. The compound also inhibited the proliferation of human cancer cells with LD50 values of 38.0-44.4 microM. In an in vivo anti-tumor assay on nude mice bearing solid tumors of HeLa cells, dehydroaltenusin was shown to be a promising suppressor of solid tumors. Histopathological examination revealed that increased tumor necrosis and decreased mitotic index were apparently detected by the compound in vivo. Therefore, dehydroaltenusin could be of interest as not only a mammalian pol alpha-specific inhibitor, but also as a candidate drug for anti-cancer treatment.     

Fusarium proliferatum, an endophytic fungus from Dysoxylum binectariferum Hook.f, produces rohitukine, a chromane alkaloid possessing anti-cancer activity

Rohitukine is a chromane alkaloid possessing anti-inflammatory, anti-cancer and immuno-modulatory properties. The compound was first reported from Amoora rohituka (Meliaceae) and later from Dysoxylum binectariferum (Meliaceae) and Schumanniophyton problematicum (Rubiaceae). Flavopiridol, a semi-synthetic derivative of rohitukine is a potent CDK inhibitor and is currently in Phase III clinical trials. In this study, the isolation of an endophytic fungus, Fusarium proliferatum (MTCC 9690) from the inner bark tissue of Dysoxylum binectariferum Hook.f (Meliaceae) is reported. The endophytic fungus produces rohitukine when cultured in shake flasks containing potato dextrose broth. The yield of rohitukine was 186 μg/100 g dry mycelial weight, substantially lower than that produced by the host tissue. The compound from the fungus was authenticated by comparing the LC–HRMS and LC–HRMS/MS spectra with those of the reference standard and that produced by the host plant. Methanolic extract of the fungus was cytotoxic against HCT-116 and MCF-7 human cancer cell lines (IC₅₀ = 10 μg/ml for both cancer cell lines).     

SEPH, a Cdc7p orthologue from Aspergillus nidulans, functions upstream of actin ring formation during cytokinesis

In the filamentous fungus, Aspergillus nidulans, multiple rounds of nuclear division occur before cytokinesis, allowing an unambiguous identification of genes required specifically for cytokinesis. As in animal cells, both an intact microtubule cytoskeleton and progression through mitosis are required for actin ring formation and contraction. The sepH gene from A. nidulans was discovered in a screen for temperature-sensitive cytokinesis mutants. Sequence analysis showed that SEPH is 42% identical to the serine-threonine kinase Cdc7p from fission yeast. Signalling through the Septation Initiation Network (SIN), which includes Cdc7p and the GTPase Spg1p, is emerging as a primary regulatory pathway used by fission yeast to control cytokinesis. A similar group of proteins comprise the Mitotic Exit Network (MEN) in budding yeast. This is the first direct evidence for the existence of a functional SIN-MEN pathway outside budding and fission yeast. In addition to SEPH, potential homologues were also identified in other fungi and plants but not in animal cells. Deletion of sepH resulted in a viable strain that failed to septate at any temperature. Interestingly, quantitative analysis of the actin cytoskeleton revealed that sepH is required for construction of the actin ring. Therefore, SEPH is distinct from its counterpart in fission yeast, in which SIN components operate downstream of actin ring formation and are necessary for ring contraction and later events of septation. We conclude that A. nidulans has components of a SIN-MEN pathway, one of which, SEPH, is required for early events during cytokinesis.     

Anti-inflammatory activity of 6MFA, a compound obtained from fungus Aspergillus ochraceus

6MFA, an interferon-inducing substance obtained from fungus, Aspergillus ochraceus, has shown anti-inflammatory activity both in acute and chronic animal models of inflammation. It was found that 6MFA was equally effective in inhibiting both exudative as well as granulative phase of inflammation. The compound suppressed also cellular migration during inflammatory process and potentiated significantly the anti-inflammatory activity of indomethacin. The compound was devoid of analgesic or antipyretic activity. The probable mechanism of action of this compound is not fully understood. However, the possibility of triggering the induction of endogenous anti-inflammatory substance(s) along with interferon(s), or interaction of induced interferon(s) directly or indirectly with the prostaglandin system has been attributed.     

Hypoxylon sp., an Endophyte of Persea indica, Producing 1,8-Cineole and Other Bioactive Volatiles with Fuel Potential

An endophytic fungus of Persea indica was identified, on the basis of its anamorphic stage, as Nodulosporium sp. by SEM. Partial sequence analysis of ITS rDNA revealed the identity of the teleomorphic stage of the fungus as Hypoxylon sp. It produces an impressive spectrum of volatile organic compounds (VOCs), most notably 1,8-cineole, 1-methyl-1,4-cyclohexadiene, and tentatively identified (+)-.alpha.-methylene-.alpha.-fenchocamphorone, among many others, most of which are unidentified. Six-day-old cultures of Hypoxylon sp. displayed maximal VOC-antimicrobial activity against Botrytis cinerea, Phytophthora cinnamomi, Cercospora beticola, and Sclerotinia sclerotiorum suggesting that the VOCs may play some role in the biology of the fungus and its survival in its host plant. Media containing starch- or sugar-related substrates best supported VOC production by the fungus. Direct on-line quantification of VOCs was measured by proton transfer mass spectrometry covering a continuous range with optimum VOC production occurred at 6 days at 145 ppmv with a rate of production of 7.65 ppmv/h. This report unequivocally demonstrates that 1,8-cineole (a monoterpene) is produced by a microorganism, which represents a novel and important source of this compound. This monoterpene is an octane derivative and has potential use as a fuel additive as do the other VOCs of this organism. Thus, fungal sourcing of this compound and other VOCs as produced by Hypoxylon sp. greatly expands their potential applications in medicine, industry, and energy production.     

Interaction between the black yeast Aureobasidium pullulans and the gall midge Lasioptera ephedricola in gall formation on the desert shrub Ephedra trifurca

Galls produced by the cecidomynd Lastoptera ephedncola on Ephedia trifurca always have a black ring associated with them while galls produced by the congener L ephedrae never do Black ring material after microscopic examination and culture proved to be Aureobasidium pullulans In addition to lacking black ring material neither L ephedrae galls nor healthy stems consistently yielded Aureobasidium on culture Gall and larva size measurements indicated that continued larval presence is not necessary for gall development, suggesting fungus initiated gall formation However inoculation of healthy stems with Aureobasidium caused lesions hut not galls The mycelium m galls did not appear grazed and neither larvae nor pupae contained Aureobasidium propagules suggesting that larvae do not feed directly on fungi These data also suggest that there is no trans-pupal passage of fungus from larvae or pupae to adults Newly emerged females do not carry fungal propagules suggesting that thcy are not inoculated upon exiting the gall Gall position leaf culture and stem culture data suggest that the fungus is picked up from leaves prior to oviposition     

Antifungal and new metabolites of Myrothecium sp. Z16, a fungus associated with white croaker Argyrosomus argentatus

AIMS: Fungal infection is still a life-threatening risk for the immunocompromised population such as AIDS patients and those who receive treatments with immunosuppressors and/or frequent administrations of wide-spectrum antibiotics, which inevitably lead to the drug resistance and unbalanced microflora populations. The present work was accordingly performed to characterize more potent antifungal metabolites from various cultures of marine fungi residing in white croaker Argyrosomus argentatus. METHODS AND RESULTS: The three most common opportunistic human pathogens Candida albicans (CCCCM ID 00148), Aspergillus niger (CCCCM ACCC 30005) and Trichophyton rubrum (CCCCM ID 00001) were selected as test fungi. A total of 16 cultivable fungal strains were isolated from the variant tissues of Ar. argentatus collected from the Yellow Sea, followed by preliminary antifungal screenings of the EtOAc extracts of the corresponding cultures. As a result, the inhibition of the three target fungi, plus being allergic to isolators' skin, were discerned with the EtOAc extract of the fungus under the isolation number Z16 that was identified subsequently as Myrothecium sp. by a combination of morphological and 18S rDNA finger-typing characteristics. A follow-up bioassay fractionation of the EtOAc extract, in conjunction with spectral analyses [MS, (1)H-NMR, (13)C-NMR, distortionless enhancement by polarization transfer (DEPT), heteronuclear multiple quantum coherence (HMQC) and heteronuclear multiple bond resonance (HMBC)] wherever required, afforded eventually the characterization of a new acid (compound 1: 4,5-ditridecyl-octanedioic acid), three macrocyclic trichothecenes including roridin A (compound 2), verrucarin A (compound 3) and 8beta-acetoxy-roridin H (compound 4), (22E,24R)-cerevisterol (compound 5) and N-phenyl-beta-amino-naphthalene (compound 6). In vitro antifungal tests showed that the three trichothecenes were active against A. niger, T. rubrum and C. albicans with MICs of 31.25, 62.5 and 125 microg ml(-1) for compound 2, 250, 125 and 31.25 microg ml(-1) for compound 3 as well as 125, 62.5 and 125 microg ml(-1) for compound 4 respectively. The MICs of ketoconazole (co-assayed herewith as a positive reference) against A. niger, T. rubrum and C. albicans were 31.25, 250, 31.25 microg ml(-1) respectively. A preliminary structure-activity relationship of the antifungal trichothecenes was highlighted in brief. CONCLUSIONS: The present investigation demonstrated that marine fungus Myrothecium sp. Z16 associated with white croaker (Ar. argentatus), was an efficient producer of a new acid and antifungal trichothecenes, the latter presumably being also the allergic substances in the culture. SIGNIFICANCE AND IMPACT OF THE STUDY: The title marine fungus was investigated to be a resource of new aliphatic acid, and trichothecenes with potent antifungal and dermal toxic actions.     

Volatile Terpenoids of Endophyte-free and Infected Peppermint (<Emphasis Type="Italic">Mentha piperita</Emphasis> L.): Chemical Partitioning of a Symbiosis

The study reports the effects on volatiles of an endophytic fungus inhabiting asymptomatically the leaves of peppermint. By means of headspace solid-phase microextraction (HS-SPME) and gaschromatography-mass spectrometry (GC-MS) terpenoids were sampled in time course from the head space of peppermint leaves and roots. After removal of the mycelium from peppermint tissues, fungal volatiles were analyzed and compared with those of pure fungal cultures. In the presence of the endophyte, the relative amount of all main compounds increased in leaves. Starting from the first 14 d of culture, (−)-menthone and (+)-neomenthol were consistently higher than in control plants. On the contrary, (+)-menthofuran increased only by 28 d of culture. Root volatiles were also dramatically altered by the presence of the fungus, with (+)-pulegone accounting for at least 44% of the total volatile emission. (+)-Pulegone was also the main compound of PGP-HSF mycelium isolated from peppermint roots. The sesquiterpenoid cuparene was found as a novel compound of peppermint leaf headspace and was a main volatile of ex planta and pure culture mycelia. The chemical spectrum of terpenoids and their distribution among peppermint roots, leaves, and mycelia are likely to account for a fine regulation of the mutualism in planta and for the acquisition by the fungus of novel metabolic competences. This work is dedicated to the memory of Prof. Silvano Scannerini.     

Biodegradation of crystal violet by the white rot fungus Phanerochaete chrysosporium

Biodegradation of crystal violet (N,N,N',N',N',N'-hexamethylpararosaniline) in ligninolytic (nitrogen-limited) cultures of the white rot fungus Phanerochaete chrysosporium was demonstrated by the disappearance of crystal violet and by the identification of three metabolites (N,N,N',N',N'-pentamethylpararosaniline, N,N,N',N'-tetramethylpararosaniline, and N,N',N'-trimethylpararosaniline) formed by sequential N-demethylation of the parent compound. Metabolite formation also occurred when crystal violet was incubated with the extracellular fluid obtained from ligninolytic cultures of this fungus, provided that an H2O2-generating system was supplied. This, as well as the fact that a purified ligninase catalyzed N-demethylation of crystal violet, demonstrated that biodegradation of crystal violet by this fungus is dependent, at least in part, upon its lignin-degrading system. In addition to crystal violet, six other triphenylmethane dyes (pararosaniline, cresol red, bromphenol blue, ethyl violet, malachite green, and brilliant green) were shown to be degraded by the lignin-degrading system of this fungus. An unexpected result was the finding that substantial degradation of crystal violet also occurred in nonligninolytic (nitrogen-sufficient) cultures of P. chrysosporium, suggesting that in addition to the lignin-degrading system, another mechanism exists in this fungus which is also able to degrade crystal violet.     

Plant pathotoxins from Alternaria citri: The major toxin specific for rough lemon plants

A pathotype of the fungus Alternaria citri that attacks rough lemon plants produces several toxins in culture which specifically damage rough lemon and Rangpur lime plants. The major toxin produced, Toxin I, was by far the most potent compound (ED₅₀ = 30 ng/ml). Five other minor toxins were active at ED₅₀ levels greater than 1 μg/ml. On the basis of mass, ¹H and ¹³C NMR spectra and decoupling studies of Toxin I and derivative, Toxin I is a 19 carbon polyalcohol with an α-dihydropyrone ring. The γ-dihydropyrone tautomer was less predominant. Culture filtrates of A. citri also contained a biologically inactive, partially analogous, component possessing a tetrahydropyran ring. It probably arises from decarboxylation of Toxin I. Toxin I was highly specific and did not affect nonhost plants at 10 000 times the concentrations affecting rough lemon.     

Trechonolide A,a new withanolide type from Trechonaetes laciniata

The structure of a new withanolide type isolated from Trechonaetes laciniata, having a hemiacetal ring system and a 5-member ring lactone has been elucidated by X-ray analysis as (23R)-5β,6β-epoxy-12β,17β-dihydroxy-1-oxo-12,22-hemiacetal-ergosta-2,24-dien-23,26-olide (trechonolide A). A second compound (trechonolide B) having at C-12 a β-methoxy group, has also been isolated and assumed to be an artifact since, by heating trechonolide A in methanol with a trace of acid, the methoxy derivative was produced.     

PAAN135, a novel rhizospheric fungus associated with Cholistan desert grass <Emphasis Type="Italic">Panicum antidotale,</Emphasis> is a species of <Emphasis Type="Italic">Saccharomycetales</Emphasis> and a new source of cyclo-L-prolylglycine diketopiperazine

A number of fungi have been reported for their ability to make symbiotic relationship with plant roots. This mutual interaction helps fungi produce important metabolites in the rhizospheric region supporting plants for better growth and survival. These biologically important fungal metabolites, especially of antibiotic potential, can be isolated and characterized in vitro for further applications. PAAN135, a fungus isolated from rhizosphere of a perennial grass, Panicum antidotale, from a nearby desert Cholistan exhibited antibacterial characteristics against six indicator strains. Culture filtrate of the fungal isolate was extracted with different solvents to isolate the potential antibiotic compound. The antibiotic-like component produced by the fungus was separated in n-Butanol fraction. It was purified using Column Chromatography and identified as a diketopiperazine, cyclo-L-prolylglycine (cPG), after structural elucidation through spectroscopic studies. The purified compound showed very good antibacterial activity against the six indicator strains used in the present study. The diketopiperazine cPG has been reported in the literature as a neuroprotective and memory facilitating agent. After molecular characterization and phylogenetic analysis PAAN135 fungus could only be classified up to the order level Saccharomycetales. Thus the fungus was placed as a new species under unclassified Saccharomycetales under the phylum Ascomycota and named as Saccharomycetales sp. PAAN135. To our knowledge isolation of the cyclo-L-prolylglycine as a potential antibacterial compound from a new fungal species Saccharomycetales sp. PAAN135 from a Cholistan desert rhizosphere is being reported for the first time.     

Biodegradation of crystal violet by the white rot fungus Phanerochaete chrysosporium.

Biodegradation of crystal violet (N,N,N',N',N',N'-hexamethylpararosaniline) in ligninolytic (nitrogen-limited) cultures of the white rot fungus Phanerochaete chrysosporium was demonstrated by the disappearance of crystal violet and by the identification of three metabolites (N,N,N',N',N'-pentamethylpararosaniline, N,N,N',N'-tetramethylpararosaniline, and N,N',N'-trimethylpararosaniline) formed by sequential N-demethylation of the parent compound. Metabolite formation also occurred when crystal violet was incubated with the extracellular fluid obtained from ligninolytic cultures of this fungus, provided that an H2O2-generating system was supplied. This, as well as the fact that a purified ligninase catalyzed N-demethylation of crystal violet, demonstrated that biodegradation of crystal violet by this fungus is dependent, at least in part, upon its lignin-degrading system. In addition to crystal violet, six other triphenylmethane dyes (pararosaniline, cresol red, bromphenol blue, ethyl violet, malachite green, and brilliant green) were shown to be degraded by the lignin-degrading system of this fungus. An unexpected result was the finding that substantial degradation of crystal violet also occurred in nonligninolytic (nitrogen-sufficient) cultures of P. chrysosporium, suggesting that in addition to the lignin-degrading system, another mechanism exists in this fungus which is also able to degrade crystal violet.     

Biotransformation of glabratephrin, a rare type of isoprenylated flavonoids, by Aspergillus niger

Microbial transformation of glabratephrin, the major isolated compound from Tephrosia purpurea, afforded pseudosemiglabrin. The formation of the transformed compound seems to be performed via ring opening-closure of a five-membered ring causing transformation from a spiro into a fused system. The structure of the transformed compound was determined by comprehensive NMR studies, including DEPT, COSY, HMQC, NOE and MS.     

发酵产鸢尾酮真菌的分离鉴定及生香特性的初步研究

从陈化的鸢尾根状茎分离到产鸢尾酮的丝状真菌,以其作用于新鲜鸢尾根状茎,通过固体发酵产生了鸢尾酮。其中1株Rhizopusoryzae（94Y－01）表现最好。对培养基配方、培养温度、培养时间等发酵条件进行了报道。从发酵物提取得到的浸膏和凝脂,其理化指标分析及香气嗅评与传统鸢尾酮产品类似,经薄层层析和气相色谱－质谱（GC／MS）分析,表明含有鸢尾酮。     

Isolation and identification of antimicrobial agent-producing bacterium from <Emphasis Type="Italic">Taxus baccata</Emphasis> rhizosphere antagonistic against clinically significant microbes

A bacterium identified as Pseudomonas fluorescence was isolated from Taxus baccata rhizosphere. Ethyl acetate extract from its culture filtrate yielded an active antimicrobial compound that was purified by TLC. The active metabolites were resolved by column chromatography on silica gel (60–120 mesh). The compound was further characterized on the basis of spectral data (UV, IR and ¹HNMR), which indicated the presence of an aromatic ring and phenolic functionality. The compound showed significant antimicrobial activity against two-gram positive bacteria (B. subtilis and S. aureus), four-gram negative bacteria (E. coli, K. pneumoniae, S. flexneri and P. aeruginosa), and one pathogenic fungus (Candida albicans). The minimum inhibitory concentration (MIC) of the compound ranged between 75μg to 250 μg/ml.     

发酵产鸢尾酮真菌的分离鉴定及生香特性的初步研究*

从陈化的鸢尾根状茎分离到产鸢尾酮的丝状真菌,以其作用于新鲜鸢尾根状茎,通过固体发酵产生了鸢尾酮。其中1株Rhizopusoryzae（94Y－01）表现最好。对培养基配方、培养温度、培养时间等发酵条件进行了报道。从发酵物提取得到的浸膏和凝脂,其理化指标分析及香气嗅评与传统鸢尾酮产品类似,经薄层层析和气相色谱－质谱（GC／MS）分析,表明含有鸢尾酮。