Misidentification of Neosartorya pseudofischeri as Aspergillus fumigatus in a lung transplant patient.

We present a case of disseminated Neosartorya pseudofischeri infection in a bilateral lung transplant patient with cystic fibrosis. The organism was originally misidentified from respiratory specimens as Aspergillus fumigatus using colonial and microscopic morphology. DNA sequencing subsequently identified the organism correctly as N. pseudofischeri.

Exploring the chemodiversity and biological activities of the secondary metabolites from the marine fungus Neosartorya pseudofischeri.

The production of fungal metabolites can be remarkably influenced by various cultivation parameters. To explore the biosynthetic potentials of the marine fungus, Neosartorya pseudofischeri, which was isolated from the inner tissue of starfish Acanthaster planci, glycerol-peptone-yeast extract (GlyPY) and glucose-peptone-yeast extract (GluPY) media were used to culture this fungus. When cultured in GlyPY medium, this fungus produced two novel diketopiperazines, neosartins A and B (1 and 2), together with six biogenetically-related known diketopiperazines,1,2,3,4-tetrahydro-2, 3-dimethyl-1,4-dioxopyrazino[1,2-a]indole (3), 1,2,3,4-tetrahydro-2-methyl-3-methylen e-1,4-dioxopyrazino[1,2-a]indole (4), 1,2,3,4-tetrahydro-2-methyl-1,3,4-trioxopyrazino[1,2-a] indole (5), 6-acetylbis(methylthio)gliotoxin (10), bisdethiobis(methylthio)gliotoxin (11), didehydrobisdethiobis(methylthio)gliotoxin (12) and N-methyl-1H-indole-2-carboxamide (6). However, a novel tetracyclic-fused alkaloid, neosartin C (14), a meroterpenoid, pyripyropene A (15), gliotoxin (7) and five known gliotoxin analogues, acetylgliotoxin (8), reduced gliotoxin (9), 6-acetylbis(methylthio)gliotoxin (10), bisdethiobis(methylthio) gliotoxin (11) and bis-N-norgliovictin (13), were obtained when grown in glucose-containing medium (GluPY medium). This is the first report of compounds 3, 4, 6, 9, 10 and 12 as naturally occurring. Their structures were determined mainly by MS, 1D and 2D NMR data. The possible biosynthetic pathways of gliotoxin-related analogues and neosartin C were proposed. The antibacterial activity of compounds 2-14 and the cytotoxic activity of compounds 4, 5 and 7-13 were evaluated. Their structure-activity relationships are also preliminarily discussed.

Isavuconazole activity against Aspergillus lentulus, Neosartorya udagawae, and Cryptococcus gattii, emerging fungal pathogens with reduced azole susceptibility.

Isavuconazole is an extended-spectrum triazole with in vitro activity against a wide variety of fungal pathogens. Clinical isolates of molds Aspergillus lentulus and Neosartorya udagawae and yeast Cryptococcus gattii VGII (implicated in the outbreak in the Pacific Northwest, North America) exhibit reduced susceptibilities to several azoles but higher susceptibilities to isavuconazole.

Invasive sino-orbital mycosis in an aplastic anemia patient caused by Neosartorya laciniosa.

We report the first case of Neosartorya laciniosa invasive sinusitis involving the orbit in an immunocompromised male with aplastic anemia. Treatment included surgical debridement with enucleation of the eye and combination voriconazole and micafungin therapy followed by voriconazole alone. The fungus was identified using sequencing of partial benA and calmodulin genes.

Secondary metabolites from a culture of the fungus Neosartorya pseudofischeri and their in vitro cytostatic activity in human cancer cells.

Four known (1, 2, 3, and 6) and three new compounds including a 1,4-diacetyl-2,5-dibenzylpiperazine derivative (4), a quinazolinone-containing indole derivative (5), and a new ester of 2,4-dihydroxy-6-methylbenzoic acid (7) were isolated from the fungus Neosartorya pseudofischeri S. W. Peterson. Compound 2 displayed in vitro growth inhibitory activity that ranged between the activities of etoposide and carboplatin, chosen as reference compounds, in six distinct cancer cell lines. Compound 1 displayed less activity than 2. Computer-assisted phase-contrast microscopy-related analysis revealed that 2 displayed cytostatic, not cytotoxic, effects in human U373 glioblastoma and A549 non-small cell lung cancer apoptosis-resistant cells with marked inhibition of mitotic rates. Cancer cells in the remaining phases of the cell cycle were unchanged. Flow cytometry analysis further confirmed that 2 does not induce apoptotic features in U373 or A549 cancer cells. Thus, 2 represents a novel chemical scaffold from which derivatives for anticancer cytostatic compounds can be derived.

Uncommon Neosartorya udagawae fungus as a causative agent of severe corneal infection.

We report the first documented case of a posttraumatic fungal keratitis caused by Neosartorya udagawae. The patient was empirically treated with fluconazole until a corneal scraping grew an Aspergillus fumigatus-like fungus, and itraconazole therapy was then established. A sequence-based approach assigned the isolate to the species. Five months after completion of antifungal therapy, endophthalmitis occurred and orbital exenteration was necessary.

Biodegradation of petroleum hydrocarbons by Neosartorya sp. BL4.

A new petroleum hydrocarbon-degrading fungus, isolated from an oil contaminant soil, was identified as Neosartorya (teleomorph of Aspergillus) sp. This isolate was able to degrade total petroleum hydrocarbons (TPHs) without a lag phase, but degradation rates decreased with increasing initial TPH concentrations (5,000-20,000 mg L(-1)). The TPH degradation by the isolate showed a substrate inhibition behavior with an inhibition constant (K(i)) of 1,860 mg L(-1). Dual lag phase of TPH degradation indicated the ability to adapt its metabolic activity to utilize different types of hydrocarbons as an electron donor. Initially n-alkanes were rapidly removed without lag phase in the whole range of substrate and heavy molecular weight alkanes (HMWAs; C23-C24) and low molecular weight alkanes (LMWAs C9-C15) out of n-alkane hydrocarbons were degraded rapidly, whereas the removal of mid molecular weight alkanes (MMWAs; C16-C22) was relatively slower. Relatively slow degradation of MMWAs is probably caused by biotransformation of HMWAs or non-alkane hydrocarbons to MMWAs.

A rapid method for identifying Byssochlamys and Hamigera.

Heat-resistant fungi, genera Byssochlamys, Talaromyces, Neosartorya, and Hamigera, contribute significantly to the spoilage of heat-processed acidic foods, due to the formation of heat-resistant ascospores. Here, we first evaluated the differences in the beta-tubulin gene between Byssochlamys and Hamigera and developed specific primers to identify the Byssochlamys species fulva, nivea, and spectabilis, and Hamigera. Using primers designed for B. fulva and B. nivea (B1F/1R), specific PCR products were detected for B. fulva and B. nivea, as well as B. langunculariae and B. zollerniae, two closely related species. Similarly, the Pae4F/4R-1 and H2F/2R primers produced specific PCR products for B. spectabilis and Hamigera, respectively. Using these three primer sets, strains involved in acidic food spoilage and environmental contamination were not detected. The detection limits of all primer sets were 1 ng of DNA by PCR and 10 pg of DNA by nested PCR. Each PCR assay was specific, even if the sample was contaminated 1,000-fold by other fungal DNA. Thus, this method has proved to possess an extremely high degree of specificity.

Invasive aspergillosis due to Neosartorya udagawae.

BACKGROUND: Invasive aspergillosis (IA) is most commonly caused by the morphospecies Aspergillus fumigatus. However, genetic-based methods indicate that organisms phenotypically identified as A. fumigatus actually constitute a mold complex, designated Aspergillus section fumigati subgenus fumigati. METHODS: Multilocus sequencing and analysis was performed on fungi identified as A. fumigatus from the clinical culture collection maintained at the National Institutes of Health from 2000 through 2008, with a focus on the internal transcribed spacer 1 and 2 regions of ribosomal DNA (rDNA), beta-tubulin, and rodlet A genes. We reviewed the medical records, radiology, and histopathology of corresponding patients. To confirm identification of Neosartorya udagawae isolates, mating studies were performed with reference strains. Antifungal susceptibility testing was performed by broth microdilution and read at 48 hours. RESULTS: Thirty-six cases of infection attributed to A. fumigatus were identified; 4 were caused by N. udagawae (3 in patients with chronic granulomatous disease and 1 in a patient with myelodysplastic syndrome). Disease due to N. udagawae was chronic, with a median duration of 35 weeks, compared with a median duration of 5.5 weeks for patients with chronic granulomatous disease who had infection due to A. fumigatus sensu stricto (P < .05 , Mann-Whitney U test). Infection spread across anatomical planes in a contiguous manner and was refractory to standard therapy. Two of the 4 patients died. N. udagawae demonstrated relatively higher minimum inhibitory concentrations to various agents, compared with those demonstrated by contemporary A. fumigatus sensu stricto isolates. CONCLUSIONS: To our knowledge, this is the first report documenting infection due to N. udagawae. Clinical manifestations were distinct from those of typical IA. Fumigati-mimetics with inherent potential for antifungal resistance are agents of IA. Genetic identification of molds should be considered for unusual or refractory IA.

Allergic fungal rhinosinusitis caused by Neosartorya hiratsukae from India.

We report here a case of allergic fungal rhinosinusitis caused by Neosartorya hiratsukae. This fungus was never previously been isolated from any case with fungal rhinosinusitis. The identification of this agent was confirmed by comparing the nucleotide sequence of the ITS region of ribosomal DNA with that in the GenBank DNA database. Identification of N. hiratsukae on the basis of colony morphology and microscopic feature may be difficult due to similarities with a few Aspergillus species. Scanning electron microscopy or DNA sequence analysis is essential for the accurate identification.

Heat activation of Neosartorya and Talaromyces ascospores and enhancement by organic acids.

Neosartorya and Talaromyces are typical fungi capable of producing heat resistant ascospores responsible for the spoilage of processed fruit products. In this study, the heat activation rates of Neosartorya and Talaromyces ascospores were investigated in several suspending media at various heating temperatures. Ascospores were dispersed in pH 3.5 McIlvain buffer, organic acid/alcohol-supplemented McIlvain buffer and grape juice (pH 3.5, 5.0 degrees Brix) prior to heat treatments. In McIlvain buffer, the number of germinating ascospores increased logarithmically with longer exposure to heating at an test temperatures. Heat activation rates (k values) accelerated with increasing temperature. The calculated activation energy (Ea) values were similar among ascospores from the same genus, but the Ea of the test Neosartorya spp. were greater than that of the test Talaromyces spp. Greater k values were calculated from acetate-supplemented McIlvain buffer and grape juice. Similarly, normal- and branched-chain fatty acids were shown to enhance the heat activation rate of the ascospores in McIlvain buffer systems. These results could assist the food industry in designing adequate thermal processes for food products against the heat resistant fungi.

Identification of two marine fungi and evaluation of their antivirus and antitumor activities.

OBJECTIVE: To identify two marine fungi and evaluate the inhibitory effects of their crude extracts on Tobacco mosaic virus and two tumor cell lines. METHODS: Crude extracts was obtained by extracting with MeOH and evaporated in vacuo. The extracts was water-soluble fraction which was dissolved in water, and the other fraction was water insoluble. The fungi were identified by morphology and Internal Transcribed Spcer (ITS) rDNA molecular methods. The inhibitory effect on Tobacco mosaic virus was evaluated by indirect enzyme linked immunosorbent assay, and the anti-tumor activity was tested by methyl thiazolyl tetrazolium method. RESULTS: The fungi were identified as Penicillium oxalicum and Neosartorya fischeri. There crude extracts inhibited Tobacco Mosaic Virus and two tumor cell lines. The active fraction named 0312F1 inhibited Tobacco Mosaic Virus and tumor cell lines and was water-soluble. The fraction named 1008F1 inhibited Tobacco Mosaic Virus and was insoluble in water, whereas the fraction inhibited tumor cell lines was water-soluble. CONCLUSION: The active fraction named 0312F1 inhibited Tobacco Mosaic Virus was different from that named 1008F1 inhibited Tobacco Mosaic Virus. The active fraction named 0312F1 inhibited tumor cell lines was the same as that named 1008F1. Furthermore, the inhibitory activity of water-soluble fraction named 0312F1 against BEL-7404 cell line was much higher than that against SGC-7901 cell lines, whereas the inhibitory activity of active fraction named 1008F1 against SGC-7901 cell line was much higher.

Neuroprotective Secondary Metabolite Produced by an Endophytic Fungus, Neosartorya fischeri JS0553, Isolated from Glehnia littoralis.

Roots of Glehnia littoralis have been used to heal stroke as a traditional medicine. Even though many studies on this plant have been conducted, the secondary metabolites produced by its endophytes and their bioactivities have not been investigated thus far. Therefore, a new meroditerpenoid named sartorypyrone E (1) and eight known compounds (2-9) were isolated from extracts of cultured Neosartorya fischeri JS0553, an endophyte of G. littoralis. The isolated metabolites were identified using spectroscopic methods and chemical reaction, based on a comparison to literature data. Relative and absolute stereochemistries of compound 1 were also elucidated. To identify the protective effects of isolated compounds (1-9) in HT22 cells against glutamate-induced cytotoxicity, we assessed inhibition of cell death, intracellular reactive oxygen species (ROS) accumulation, and calcium ion (Ca2+) influx. Among the isolates, compound 8, identified as fischerin, showed significant neuroprotective activity on glutamate-mediated HT22 cell death through inhibition of ROS, Ca2+ influx, and phosphorylation of mitogen-activated protein kinase, including c-Jun N-terminal kinase, extracellular signal-regulated kinase, and p38. The results suggested that the metabolites produced by the endophyte N. fischeri JS0553 might be related to the neuroprotective activity of its host plant, G. littoralis.

Screening of thermotolerant and thermophilic fungi aiming ß-xylosidase and arabinanase production.

Plant cell wall is mainly composed by cellulose, hemicellulose and lignin. The heterogeneous structure and composition of the hemicellulose are key impediments to its depolymerization and subsequent use in fermentation processes. Thus, this study aimed to perform a screening of thermophilic and thermotolerant filamentous fungi collected from different regions of the São Paulo state, and analyze the production of ß-xylosidase and arabinanase at different temperatures. These enzymes are important to cell wall degradation and synthesis of end products as xylose and arabinose, respectively, which are significant sugars to fermentation and ethanol production. A total of 12 fungal species were analyzed and 9 of them grew at 45 °C, suggesting a thermophilic or thermotolerant character. Additionally Aspergillus thermomutatus anamorph of Neosartorya and A. parasiticus grew at 50 °C. Aspergillus niger and Aspergillus thermomutatus were the filamentous fungi with the most expressive production of ß-xylosidase and arabinanase, respectively. In general for most of the tested microorganisms, ß-xylosidase and arabinanase activities from mycelial extract (intracellular form) were higher in cultures grown at high temperatures (35-40 °C), while the correspondent extracellular activities were favorably secreted from cultures at 30 °C. This study contributes to catalogue isolated fungi of the state of São Paulo, and these findings could be promising sources for thermophilic and thermotolerant microorganisms, which are industrially important due to their enzymes.

Method for identifying heat-resistant fungi of the genus Neosartorya.

Species of the genus Neosartorya are heat-resistant fungi that cause the spoilage of heat-processed acidic foods due to the formation of heat-resistant ascospores, and they produce mycotoxins, such as fumitremorgins and gliotoxin. Their anamorphs are phylogenetically and morphologically very close to Aspergillus fumigatus, which has never been reported as a spoilage agent in heat-processed food products. Therefore it is important to discriminate between the species of Neosartorya and A. fumigatus in the food industry. In the present study, we examined ß-tubulin and calmodulin genes to identify Neosartorya and A. fumigatus at the species level and found a region for specifically detecting these species. We succeeded in developing the PCR method of differentiating and identifying Neosartorya and A. fumigatus using specific primer sets. Moreover, we developed specific primer sets to identify Neosartorya species, N. fischeri, N. glabra, N. hiratsukae, N. pseudofischeri, and N. spinosa-complex, which are important in food spoilage; these fungi vary in heat resistance and productivity of mycotoxins, depending on the species. PCR using these primer sets did not detect other fungi involved in food spoilage and environmental contamination. These identification methods are rapid and simple with extremely high specificity.

Sinonasal and sino-orbital aspergillosis in 23 cats: aetiology, clinicopathological features and treatment outcomes.

Aetiology, clinicopathological findings and treatment outcomes were documented in 23 cats (1.5-13 years of age) with sinonasal (SNA, n=6) or sino-orbital (SOA, n=17) aspergillosis. Cases recruited retrospectively and prospectively were included if fungal hyphae were identified on cytological or histological examination and the fungal pathogen was identified by PCR and DNA sequencing (ITS1 or ITS1-5.8S-ITS2 regions, rDNA gene cluster). Fungal culture was positive in 22/23 cases. In cases of SNA, the fungal pathogen was Aspergillus fumigatus (n=4), Neosartorya fischeri or A. lentulus (n=1) or a non-speciated Neosartorya spp. (n=1). In all cases of SOA (n=17), the fungal pathogen was identified as Neosartorya spp. Nine cats had brachycephalic conformation. Cats with SNA were more likely to be infected with A. fumigatus and had a better prognosis than cats with SOA.

First evidence of mineralization of petroleum asphaltenes by a strain of Neosartorya fischeri.

A fungal strain isolated from a microbial consortium growing in a natural asphalt lake is able to grow in purified asphaltenes as the only source of carbon and energy. The asphaltenes were rigorously purified in order to avoid contamination from other petroleum fractions. In addition, most of petroporphyrins were removed. The 18S rRNA and ß-tubulin genomic sequences, as well as some morphologic characteristics, indicate that the isolate is Neosartorya fischeri. After 11 weeks of growth, the fungus is able to metabolize 15.5% of the asphaltenic carbon, including 13.2% transformed to CO(2) . In a medium containing asphaltenes as the sole source of carbon and energy, the fungal isolate produces extracellular laccase activity, which is not detected when the fungus grow in a rich medium. The results obtained in this work clearly demonstrate that there are microorganisms able to metabolize and mineralize asphaltenes, which is considered the most recalcitrant petroleum fraction.