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Sample records for aspergillus nidulans modified

  1. Presence of histones in Aspergillus nidulans

    PubMed Central

    1976-01-01

    Five major histone proteins have been extracted from chromatin isolated from purified nuclei of the fungus, Aspergillus nidulans. These proteins had chromatographic properties which were similar to reference calf thymus histones and were purified to electrophoretic homegeneity by gel chromatography of Bio-Gel P10, Bio-Gel P60, and Sephadex G-100. Electrophoresis of these proteins in three different systems (urea- starch, urea-acetic acid polyacrylamide, and discontinuous SDS polyacrylamide) showed that the A. nidulans histones H3 and H4 were nearly identical to calf thymus H3 and H4 with respect to net charge and molecular weight criteria, whereas the fungal histones H1, H2a and H2b were similar but not identical to the corresponding calf thymus histones. Amino acid analysis of A. nidulans histones H2a, H2b, and H4 showed them to be closely related to the homologous calf thymus histones. The mobility patterns of A. nidulans ribosomal basic proteins in three different electrophoretic systems were distinctly different from those of the fungal histones. PMID:799641

  2. Sexual origins of British Aspergillus nidulans isolates.

    PubMed Central

    Geiser, D M; Arnold, M L; Timberlake, W E

    1994-01-01

    Aspergillus nidulans is a holomorphic fungus, capable of producing both meiotically and mitotically derived spores. Meiosis may be an evolutionary relic in this species because it is potentially capable of mitotic recombination and because most Aspergilli lack the ability to produce meiotic spores. We tested the null hypothesis that meiosis has been a major factor in the origin of strains of A. nidulans from Great Britain by estimating linkage disequilibrium among restriction fragment length polymorphisms. These strains belong to different heterokaryon compatibility groups and are thus incapable of undergoing mitotic recombination with one another, so any recombination evidenced by linkage equilibrium is assumed to be the result of meiosis. Eleven cosmid clones of known chromosomal origin were used to generate multilocus genotypes based on restriction-pattern differences for each heterokaryon compatibility group. Low levels of genetic variation and little linkage disequilibrium were found, indicating that the heterokaryon compatibility groups represent recently diverged lineages that arose via meiotic recombination. The null hypothesis that loci are independent could not be rejected. Additionally, low levels of electrophoretic karyotype variation were indicative of meiosis. We conclude that although A. nidulans probably propagates in a primarily clonal fashion, recombination events are frequent enough to disrupt the stable maintenance of clonal genotypes. We further conclude that the British heterokaryon compatibility groups arose via recombination and not through novel mutation. Images PMID:7907796

  3. Identification of glucose transporters in Aspergillus nidulans.

    PubMed

    Dos Reis, Thaila Fernanda; Menino, João Filipe; Bom, Vinícius Leite Pedro; Brown, Neil Andrew; Colabardini, Ana Cristina; Savoldi, Marcela; Goldman, Maria Helena S; Rodrigues, Fernando; Goldman, Gustavo Henrique

    2013-01-01

    To characterize the mechanisms involved in glucose transport, in the filamentous fungus Aspergillus nidulans, we have identified four glucose transporter encoding genes hxtB-E. We evaluated the ability of hxtB-E to functionally complement the Saccharomyces cerevisiae EBY.VW4000 strain that is unable to grow on glucose, fructose, mannose or galactose as single carbon source. In S. cerevisiae HxtB-E were targeted to the plasma membrane. The expression of HxtB, HxtC and HxtE was able to restore growth on glucose, fructose, mannose or galactose, indicating that these transporters accept multiple sugars as a substrate through an energy dependent process. A tenfold excess of unlabeled maltose, galactose, fructose, and mannose were able to inhibit glucose uptake to different levels (50 to 80 %) in these s. cerevisiae complemented strains. Moreover, experiments with cyanide-m-chlorophenylhydrazone (CCCP), strongly suggest that hxtB, -C, and -E mediate glucose transport via active proton symport. The A. nidulans ΔhxtB, ΔhxtC or ΔhxtE null mutants showed ~2.5-fold reduction in the affinity for glucose, while ΔhxtB and -C also showed a 2-fold reduction in the capacity for glucose uptake. The ΔhxtD mutant had a 7.8-fold reduction in affinity, but a 3-fold increase in the capacity for glucose uptake. However, only the ΔhxtB mutant strain showed a detectable decreased rate of glucose consumption at low concentrations and an increased resistance to 2-deoxyglucose. PMID:24282591

  4. Apical control of conidiation in Aspergillus nidulans.

    PubMed

    Oiartzabal-Arano, Elixabet; Perez-de-Nanclares-Arregi, Elixabet; Espeso, Eduardo A; Etxebeste, Oier

    2016-05-01

    The infection cycle of filamentous fungi consists of two main stages: invasion (growth) and dispersion (development). After the deposition of a spore on a host, germination, polar extension and branching of vegetative cells called hyphae allow a fast and efficient invasion. Under suboptimal conditions, genetic reprogramming of hyphae results in the generation of asexual spores, allowing dissemination to new hosts and the beginning of a new infection cycle. In the model filamentous fungus Aspergillus nidulans, asexual development or conidiation is induced by the upstream developmental activation (UDA) pathway. UDA proteins transduce signals from the tip, the polarity site of hyphae, to nuclei, where developmental programs are transcriptionally activated. The present review summarizes the current knowledge on this tip-to-nucleus communication mechanism, emphasizing its dependence on hyphal polarity. Future approaches to the topic will also be suggested, as stimulating elements contributing to the understanding of how apical signals are coupled with the transcriptional control of development and pathogenesis in filamentous fungi. PMID:26782172

  5. Genetics of Polyketide Metabolism in Aspergillus nidulans

    PubMed Central

    Klejnstrup, Marie L.; Frandsen, Rasmus J. N.; Holm, Dorte K.; Nielsen, Morten T.; Mortensen, Uffe H.; Larsen, Thomas O.; Nielsen, Jakob B.

    2012-01-01

    Secondary metabolites are small molecules that show large structural diversity and a broad range of bioactivities. Some metabolites are attractive as drugs or pigments while others act as harmful mycotoxins. Filamentous fungi have the capacity to produce a wide array of secondary metabolites including polyketides. The majority of genes required for production of these metabolites are mostly organized in gene clusters, which often are silent or barely expressed under laboratory conditions, making discovery and analysis difficult. Fortunately, the genome sequences of several filamentous fungi are publicly available, greatly facilitating the establishment of links between genes and metabolites. This review covers the attempts being made to trigger the activation of polyketide metabolism in the fungal model organism Aspergillus nidulans. Moreover, it will provide an overview of the pathways where ten polyketide synthase genes have been coupled to polyketide products. Therefore, the proposed biosynthesis of the following metabolites will be presented; naphthopyrone, sterigmatocystin, aspyridones, emericellamides, asperthecin, asperfuranone, monodictyphenone/emodin, orsellinic acid, and the austinols. PMID:24957370

  6. Purification and properties of beta-galactosidase from Aspergillus nidulans.

    PubMed

    Díaz, M; Pedregosa, A M; de Lucas, J R; Torralba, S; Monistrol, I F; Laborda, F

    1996-12-01

    Beta-Galactosidase from mycelial extract of Aspergillus nidulans has been purified by substrate affinity chromatography and used to obtain anti-beta-galactosidase polyclonal antibodies. A. nidulans growing in lactose as carbon source synthesizes one active form of beta-galactosidase which seems to be a multimeric enzyme of 450 kDa composed of monomers with 120 and 97 kDa. Although the enzyme was not released to the culture medium, some enzymatic activity was detected in a cell-wall extract, thus suggesting that it can be an extracellular enzyme. Beta-Galactosidase of A. nidulans is a very unstable enzyme with an optimum pH value of 7.5 and an optimum temperature of 30 degrees C. It was only active against beta-galactoside substrates like lactose and p-nitrophenyl-beta-D-galactoside (PNPG).

  7. The chemical identification and analysis of Aspergillus nidulans secondary metabolites

    PubMed Central

    Sanchez, James F.

    2013-01-01

    Filamentous fungi have long been recognized to be a rich source of secondary metabolites with potential medicinal applications. The recent genomic sequencing of several Aspergillus species has revealed that many secondary metabolite gene clusters are apparently silent under standard laboratory conditions. Several successful approaches have been utilized to upregulate these genes and unearth the corresponding natural products. A straightforward, reliable method to purify and characterize new metabolites therefore should be useful. Details are provided herein on the cultivation of Aspergillus nidulans and the LC/MS analysis of the metabolic profile. Following is an explanation of silica gel chromatography, HPLC, and preparative TLC. Finally, the NMR characterization of previously unknown A. nidulans metabolites is detailed. PMID:23065610

  8. Response of Aspergillus nidulans and Physarum polycephalum to microwave irradiation.

    PubMed

    Mezykowski, T; Bal, J; Debiec, H; Kwarecki, K

    1980-06-01

    The influence of microwaves on genetic processes in Aspergillus nidulans and Physarum polycephalum was investigated. Suspensions of organisms were exposed in the far zone to 2450-MHz waves at 10 mW/cm2 for one hour in both CW and pulsed (1 microsecond, 600 pps) fields. Spores of A. nidulans were irradiated before and during germination. No changes in survival rate or in frequency of morphological mutation were found. Polycephalum under the influence of CW microwaves incorporated 3H-Thymine into DNA at a rate five times that of controls and twice that of thermal controls. The accelerated synthesis may reflect more efficient volume heating by microwaves, or in the presence of microthermal gradients in suspensions, or field-specific influences in concern with focal or volume heating. PMID:7003154

  9. Characterization of melanin pigment produced by Aspergillus nidulans.

    PubMed

    Gonçalves, R C R; Lisboa, H C F; Pombeiro-Sponchiado, S R

    2012-04-01

    Although most of the Ascomycetes present DHN-melanin, some reports suggest that A. nidulans does not produce this type of melanin. In this study, we analyzed the pigment extracted from highly melanized strains (MEL1 and MEL2) of Aspergillus nidulans to determine the type of melanin present in this fungus. Our results showed that the pigment produced by MEL1 and MEL2 mutants possesses physical and chemical properties and UV- and IR-spectra very similar to synthetic DOPA-melanin. The characterization of this pigment in terms of its degradation products indicated the presence of indolic units, which were also found in synthetic DOPA-melanin. The analyses of the elemental composition showed that the pigment extracted from these mutants has a high percentage of nitrogen and, therefore, it cannot be DHN-melanin, which presents only trace of nitrogen. This observation was confirmed in the test with tricyclazole because this inhibitor of DHN-melanin biosynthesis did not suppress pigment production in the MEL1 and MEL2 strains. On the other hand, in a medium containing tropolone, an inhibitor of DOPA-melanin biosynthesis, the dark pigmentation of the colonies was not observed indicating that this compound inhibited melanin production in these strains. Taken together, the results obtained in this study indicate that melanin produced by these mutants is DOPA type, representing the first report on characterization of this type of melanin in A. nidulans.

  10. Regulation of Conidiation by Light in Aspergillus nidulans

    PubMed Central

    Ruger-Herreros, Carmen; Rodríguez-Romero, Julio; Fernández-Barranco, Raul; Olmedo, María; Fischer, Reinhard; Corrochano, Luis M.; Canovas, David

    2011-01-01

    Light regulates several aspects of the biology of many organisms, including the balance between asexual and sexual development in some fungi. To understand how light regulates fungal development at the molecular level we have used Aspergillus nidulans as a model. We have performed a genome-wide expression analysis that has allowed us to identify >400 genes upregulated and >100 genes downregulated by light in developmentally competent mycelium. Among the upregulated genes were genes required for the regulation of asexual development, one of the major biological responses to light in A. nidulans, which is a pathway controlled by the master regulatory gene brlA. The expression of brlA, like conidiation, is induced by light. A detailed analysis of brlA light regulation revealed increased expression after short exposures with a maximum after 60 min of light followed by photoadaptation with longer light exposures. In addition to brlA, genes flbA–C and fluG are also light regulated, and flbA–C are required for the correct light-dependent regulation of the upstream regulator fluG. We have found that light induction of brlA required the photoreceptor complex composed of a phytochrome FphA, and the white-collar homologs LreA and LreB, and the fluffy genes flbA–C. We propose that the activation of regulatory genes by light is the key event in the activation of asexual development by light in A. nidulans. PMID:21624998

  11. Identification and Characterization of Aspergillus Nidulans Mutants Defective in Cytokinesis

    PubMed Central

    Harris, S. D.; Morrell, J. L.; Hamer, J. E.

    1994-01-01

    Filamentous fungi undergo cytokinesis by forming crosswalls termed septa. Here, we describe the genetic and physiological controls governing septation in Aspergillus nidulans. Germinating conidia do not form septa until the completion of their third nuclear division. The first septum is invariantly positioned at the basal end of the germ tube. Block-and-release experiments of nuclear division with benomyl or hydroxyurea, and analysis of various nuclear division mutants demonstrated that septum formation is dependent upon the third mitotic division. Block-and-release experiments with cytochalasin A and the localization of actin in germlings by indirect immunofluorescence showed that actin participated in septum formation. In addition to being concentrated at the growing hyphal tips, a band of actin was also apparent at the site of septum formation. Previous genetic analysis in A. nidulans identified four genes involved in septation (sepA-D). We have screened a new collection of temperature sensitive (ts) mutants of A. nidulans for strains that failed to form septa at the restrictive temperature but were able to complete early nuclear divisions. We identified five new genes designated sepE, G, H, I and J, along with one additional allele of a previously identified septation gene. On the basis of temperature shift experiments, nuclear counts and cell morphology, we sorted these cytokinesis mutants into three phenotypic classes. Interestingly, one class of mutants fails to form septa and fails to progress past the third nuclear division. This class of mutants suggests the existence of a regulatory mechanism in A. nidulans that ensures the continuation of nuclear division following the initiation of cytokinesis. PMID:8150280

  12. Aspergillus nidulans galactofuranose biosynthesis affects antifungal drug sensitivity.

    PubMed

    Alam, Md Kausar; El-Ganiny, Amira M; Afroz, Sharmin; Sanders, David A R; Liu, Juxin; Kaminskyj, Susan G W

    2012-12-01

    The cell wall is essential for fungal survival in natural environments. Many fungal wall carbohydrates are absent from humans, so they are a promising source of antifungal drug targets. Galactofuranose (Galf) is a sugar that decorates certain carbohydrates and lipids. It comprises about 5% of the Aspergillus fumigatus cell wall, and may play a role in systemic aspergillosis. We are studying Aspergillus wall formation in the tractable model system, A. nidulans. Previously we showed single-gene deletions of three sequential A. nidulans Galf biosynthesis proteins each caused similar hyphal morphogenesis defects and 500-fold reduced colony growth and sporulation. Here, we generated ugeA, ugmA and ugtA strains controlled by the alcA(p) or niiA(p) regulatable promoters. For repression and expression, alcA(p)-regulated strains were grown on complete medium with glucose or threonine, whereas niiA(p)-regulated strains were grown on minimal medium with ammonium or nitrate. Expression was assessed by qPCR and colony phenotype. The alcA(p) and niiA(p) strains produced similar effects: colonies resembling wild type for gene expression, and resembling deletion strains for gene repression. Galf immunolocalization using the L10 monoclonal antibody showed that ugmA deletion and repression phenotypes correlated with loss of hyphal wall Galf. None of the gene manipulations affected itraconazole sensitivity, as expected. Deletion of any of ugmA, ugeA, ugtA, their repression by alcA(p) or niiA(p), OR, ugmA overexpression by alcA(p), increased sensitivity to Caspofungin. Strains with alcA(p)-mediated overexpression of ugeA and ugtA had lower caspofungin sensitivity. Galf appears to play an important role in A. nidulans growth and vigor.

  13. Heterologous expression of the Aspergillus nidulans alcR-alcA system in Aspergillus niger.

    PubMed

    Nikolaev, I; Mathieu, M; van de Vondervoort, P; Visser, J; Felenbok, B

    2002-10-01

    The inducible and strongly expressed alcA gene encoding alcohol dehydrogenase I from Aspergillus nidulans was transferred together with the activator gene alcR, in the industrial fungus Aspergillus niger. This latter organism does not possess an inducible alc system but has an endogenously constitutive lowly expressed alcohol dehydrogenase activity. The overall induced expression of the alcA gene was of the same order in both fungi, as monitored by alcA transcription, alcohol dehydrogenase activity and heterologous expression of the reporter enzyme, beta-glucuronidase. However, important differences in the pattern of alcA regulation were observed between the two fungi. A high basal level of alcA transcription was observed in A. niger resulting in a lower ratio of alcA inducibility. This may be due to higher levels of the physiological inducer of the alc regulon, acetaldehyde, from general metabolism in A. niger which differs from that of A. nidulans.

  14. Identification of Developmental Regulatory Genes in Aspergillus Nidulans by Overexpression

    PubMed Central

    Marhoul, J. F.; Adams, T. H.

    1995-01-01

    Overexpression of several Aspergillus nidulans developmental regulatory genes has been shown to cause growth inhibition and development at inappropriate times. We set out to identify previously unknown developmental regulators by constructing a nutritionally inducible A. nidulans expression library containing small, random genomic DNA fragments inserted next to the alcA promoter [ alcA (p) ] in an A. nidulans transformation vector. Among 20,000 transformants containing random alcA (p) genomic DNA fusion constructs, we identified 66 distinct mutant strains in which alcA (p) induction resulted in growth inhibition as well as causing other detectable phenotypic changes. These growth inhibited mutants were divided into 52 FIG (Forced expression Inhibition of Growth) and 14 FAB (Forced expression Activation of brlA) mutants based on whether or not alcA (p) induction resulted in accumulation of mRNA for the developmental regulatory gene brlA. In four FAB mutants, alcA (p) induction not only activated brlA expression but also caused hyphae to differentiate into reduced conidiophores that produced viable spores from the tips as is observed after alcA (p) :: brlA induction. Sequence analyses of the DNA fragments under alcA (p) control in three of these four sporulating strains showed that in two cases developmental activation resulted from overexpression of previously uncharacterized genes, whereas in the third strain, the alcA (p) was fused to brlA. The potential uses for this strategy in identifying genes whose overexpression results in specific phenotypic changes like developmental induction are discussed. PMID:7713416

  15. Identification of developmental regulatory genes in Aspergillus nidulans by overexpression.

    PubMed

    Marhoul, J F; Adams, T H

    1995-02-01

    Overexpression of several Aspergillus nidulans developmental regulatory genes has been shown to cause growth inhibition and development at inappropriate times. We set out to identify previously unknown developmental regulators by constructing a nutritionally inducible A. nidulans expression library containing small, random genomic DNA fragments inserted next to the alcA promoter [alcA(p)] in an A. nidulans transformation vector. Among 20,000 transformants containing random alcA(p) genomic DNA fusion constructs, we identified 66 distinct mutant strains in which alcA(p) induction resulted in growth inhibition as well as causing other detectable phenotypic changes. These growth inhibited mutants were divided into 52 FIG (Forced expression Inhibition of Growth) and 14 FAB (Forced expression Activation of brlA) mutants based on whether or not alcA(p) induction resulted in accumulation of mRNA for the developmental regulatory gene brlA. In four FAB mutants, alcA(p) induction not only activated brlA expression but also caused hyphae to differentiate into reduced conidiophores that produced viable spores from the tips as is observed after alcA(p)::brlA induction. Sequence analyses of the DNA fragments under alcA(p) control in three of these four sporulating strains showed that in two cases developmental activation resulted from overexpression of previously uncharacterized genes, whereas in the third strain, the alcA(p) was fused to brlA. The potential uses for this strategy in identifying genes whose overexpression results in specific phenotypic changes like developmental induction are discussed.

  16. Using Aspergillus nidulans to identify antifungal drug resistance mutations.

    PubMed

    He, Xiaoxiao; Li, Shengnan; Kaminskyj, Susan G W

    2014-02-01

    Systemic fungal infections contribute to at least 10% of deaths in hospital settings. Most antifungal drugs target ergosterol (polyenes) or its biosynthetic pathway (azoles and allylamines), or beta-glucan synthesis (echinocandins). Antifungal drugs that target proteins are prone to the emergence of resistant strains. Identification of genes whose mutations lead to targeted resistance can provide new information on those pathways. We used Aspergillus nidulans as a model system to exploit its tractable sexual cycle and calcofluor white as a model antifungal agent to cross-reference our results with other studies. Within 2 weeks from inoculation on sublethal doses of calcofluor white, we isolated 24 A. nidulans adaptive strains from sectoring colonies. Meiotic analysis showed that these strains had single-gene mutations. In each case, the resistance was specific to calcofluor white, since there was no cross-resistance to caspofungin (echinocandin). Mutation sites were identified in two mutants by next-generation sequencing. These were confirmed by reengineering the mutation in a wild-type strain using a gene replacement strategy. One of these mutated genes was related to cell wall synthesis, and the other one was related to drug metabolism. Our strategy has wide application for many fungal species, for antifungal compounds used in agriculture as well as health care, and potentially during protracted drug therapy once drug resistance arises. We suggest that our strategy will be useful for keeping ahead in the drug resistance arms race. PMID:24363365

  17. Cooperation of Aspergillus nidulans enzymes increases plant polysaccharide saccharification.

    PubMed

    Tramontina, Robson; Robl, Diogo; Maitan-Alfenas, Gabriela Piccolo; de Vries, Ronald P

    2016-07-01

    Efficient polysaccharide degradation depends on interaction between enzymes acting on the main chain and the side chains. Previous studies demonstrated cooperation between several enzymes, but not all enzyme combinations have been explored. A better understanding of enzyme cooperation would enable the design of better enzyme mixtures, optimally profiting from synergistic effects. In this study, we analyzed the cooperation of several enzymes involved in the degradation of xylan, glucan, xyloglucan and crude plant biomass from Aspergillus nidulans by single and combined incubations with their polymeric substrate. Positive effects were observed between most enzymes, although not always to the same extent. Moreover, the tailor made cocktails formulated in this study resulted in efficient release of glucose from plant biomass. This study also serves as an example for the complex cooperation that occurs between enzymes in plant biomass saccharification and how expression in easily-accessible hosts, such as Pichia pastoris, can help in revealing these effects. PMID:26848939

  18. Isolation of a sexual sporulation hormone from Aspergillus nidulans.

    PubMed Central

    Champe, S P; el-Zayat, A A

    1989-01-01

    Psi factor is a substance produced by Aspergillus nidulans that induces premature sexual sporulation. Chromatographic analysis of psi-active extracts showed that psi activity resides in several different forms. Two of the forms, psiA1 and psiB1, have been isolated and have been shown to have closely similar compositions. The most abundant form, psiA1, reacts with alcohols in acidic solution by the addition of one entire molecule of the alcohol. This reaction, which is reversible, suggests that psiA1 may be a lactone whose ring is opened by alcohol addition. At high concentration, psiA1 is antagonistic to the response exhibited by the other forms of psi, but this antagonism is lost by the alcoholic derivatives. At least one unpurified psi species can be converted to psiA1 by acid catalysis. We suggest that psiA1 may be the metabolic precursor of at least some of the other more active psi components and that this conversion during Aspergillus development may be part of the process that triggers sexual sporulation. Images PMID:2661541

  19. Overexpression of two penicillin structural genes in Aspergillus nidulans.

    PubMed

    Fernández-Cañón, J M; Peñalva, M A

    1995-01-01

    We have placed two different penicillin structural genes from Aspergillus nidulans, ipnA (encoding isopenicillin N synthetase, IPNS) and acyA (encoding acyl-CoA:6-aminopenicillanic acid acyltransferase, AAT), under the control of the strong alcA promoter [alcA(p)]. Single copies of these transcriptional fusions were targeted to the same chromosomal location and conditions have been worked out which simultaneously allow induction of the alcA(p) and support penicillin biosynthesis. Transcriptional induction of the chimeric genes alcA(p)::ipnA or alcA(p)::acyA(cdna) in the relevant recombinant strains results in 10-fold higher levels of the ipnA or acyA transcripts than those resulting from transcription of the corresponding endogenous genes. This increase causes a 40-fold rise in IPNS activity or a 8-fold rise in AAT activity. Despite this rise in enzyme levels, forced expression of the ipnA gene results in only a modest increase in levels of exported penicillin, whereas forced expression of the acyA gene reduces penicillin production, showing that neither of these enzymes is rate-limiting for penicillin biosynthesis in A. nidulans. A genomic version of the alcA(p)::acyA fusion in which the acyA gene is interrupted by three small introns, is inducible by threonine to a lesser extent (as determined by both acyA mRNA levels and AAT enzyme levels) than the corresponding cDNA version, suggesting that processing of the introns present in the primary transcript may limit acyA expression.

  20. The Spt-Ada-Gcn5 Acetyltransferase (SAGA) complex in Aspergillus nidulans.

    PubMed

    Georgakopoulos, Paraskevi; Lockington, Robin A; Kelly, Joan M

    2013-01-01

    A mutation screen in Aspergillus nidulans uncovered mutations in the acdX gene that led to altered repression by acetate, but not by glucose. AcdX of A. nidulans is highly conserved with Spt8p of Saccharomyces cerevisiae, and since Spt8p is a component of the Spt-Ada-Gcn5 Acetyltransferase (SAGA) complex, the SAGA complex may have a role in acetate repression in A. nidulans. We used a bioinformatic approach to identify genes encoding most members of the SAGA complex in A. nidulans, and a proteomic analysis to confirm that most protein components identified indeed exist as a complex in A. nidulans. No apparent compositional differences were detected in mycelia cultured in acetate compared to glucose medium. The methods used revealed apparent differences between Yeast and A. nidulans in the deubiquitination (DUB) module of the complex, which in S. cerevisiae consists of Sgf11p, Sus1p, and Ubp8p. Although a convincing homologue of S. cerevisiae Ubp8p was identified in the A. nidulans genome, there were no apparent homologues for Sus1p and Sgf11p. In addition, when the SAGA complex was purified from A. nidulans, members of the DUB module were not co-purified with the complex, indicating that functional homologues of Sus1p and Sgf11p were not part of the complex. Thus, deubiquitination of H2B-Ub in stress conditions is likely to be regulated differently in A. nidulans compared to S. cerevisiae.

  1. Engineering Aspergillus nidulans for heterologous ent-kaurene and gamma-terpinene production.

    PubMed

    Bromann, Kirsi; Toivari, Mervi; Viljanen, Kaarina; Ruohonen, Laura; Nakari-Setälä, Tiina

    2016-07-01

    Terpenes are a large and varied group of natural products with a wide array of bioactivities and applications. The chemical production of industrially relevant terpenes can be expensive and time-consuming due to the structural complexity of these compounds. Here, we studied Aspergillus nidulans as a heterologous host for monoterpene and diterpene production. Previously, we identified a novel diterpene gene cluster in A. nidulans and showed that overexpression of the cluster-specific transcription factor (pbcR) led to ent-pimara-8(14),15-diene (PD) production. We report further characterization of the A. nidulans PD synthase gene (pbcA). In A. nidulans, overexpression of pbcA resulted in PD production, while deletion of pbcA abolished PD production. Overexpression of Fusarium fujikuroi ent-kaurene synthase (cps/ks) and Citrus unshiu gamma-terpinene synthase resulted in ent-kaurene and gamma-terpinene production, respectively. A. nidulans is a fungal model organism and a close relative to other industrially relevant Aspergillus species. A. nidulans is a known producer of many secondary metabolites, but its ability to produce heterologous monoterpene and diterpene compounds has not been characterized. Here, we show that A. nidulans is capable of heterologous terpene production and thus has potential as a production host for industrially relevant compounds. The genetic engineering principles reported here could also be applied to other Aspergilli. PMID:27098256

  2. Characterization and biotechnological application of recombinant xylanases from Aspergillus nidulans.

    PubMed

    Maitan-Alfenas, Gabriela P; Oliveira, Mariana B; Nagem, Ronaldo A P; de Vries, Ronald P; Guimarães, Valéria M

    2016-10-01

    Two xylanases from Aspergillus nidulans, XlnB and XlnC, were expressed in Pichia pastoris, purified and characterized. XlnB and XlnC achieved maximal activities at 60°C and pH 7.5 and at 50°C and pH 6.0, respectively. XlnB showed to be very thermostable by maintaining 50% of its original activity after 49h incubated at 50°C. XlnB had its highest activity against wheat arabinoxylan while XlnC had the best activity against beechwood xylan. Both enzymes were completely inhibited by SDS and HgCl2. Xylotriose at 1mg/ml also totally inibited XlnB activity. TLC analysis showed that the main product of beechwood xylan hydrolysis by XlnB and XlnC was xylotetraose. An additive effect was shown between XlnB and XlnC and the xylanases of two tested commercial cocktails. Sugarcane bagasse saccharification results showed that these two commercial enzymatic cocktails were able to release more glucose and xylose after supplementation with XlnB and XlnC.

  3. Characterization and biotechnological application of recombinant xylanases from Aspergillus nidulans.

    PubMed

    Maitan-Alfenas, Gabriela P; Oliveira, Mariana B; Nagem, Ronaldo A P; de Vries, Ronald P; Guimarães, Valéria M

    2016-10-01

    Two xylanases from Aspergillus nidulans, XlnB and XlnC, were expressed in Pichia pastoris, purified and characterized. XlnB and XlnC achieved maximal activities at 60°C and pH 7.5 and at 50°C and pH 6.0, respectively. XlnB showed to be very thermostable by maintaining 50% of its original activity after 49h incubated at 50°C. XlnB had its highest activity against wheat arabinoxylan while XlnC had the best activity against beechwood xylan. Both enzymes were completely inhibited by SDS and HgCl2. Xylotriose at 1mg/ml also totally inibited XlnB activity. TLC analysis showed that the main product of beechwood xylan hydrolysis by XlnB and XlnC was xylotetraose. An additive effect was shown between XlnB and XlnC and the xylanases of two tested commercial cocktails. Sugarcane bagasse saccharification results showed that these two commercial enzymatic cocktails were able to release more glucose and xylose after supplementation with XlnB and XlnC. PMID:27235731

  4. The fungal phytochrome FphA from Aspergillus nidulans.

    PubMed

    Brandt, Sonja; von Stetten, David; Günther, Mina; Hildebrandt, Peter; Frankenberg-Dinkel, Nicole

    2008-12-12

    The red light-sensing photoreceptor FphA from Aspergillus nidulans is involved in the regulation of developmental processes in response to light. Here we present extended biochemical and spectroscopic characterization of recombinant FphA using a synthetic gene with host-adapted codon usage. The recombinant photosensory domain FphAN753 was shown to display all features of a bona fide phytochrome. It covalently binds biliverdin as chromophore and undergoes red/far-red light-inducible photoconversion with both parent states being protonated. The large N-terminal variable extension of FphA exerts a stabilizing effect on the active Pfr state. Upon substitution of the highly conserved histidine 504, involved in the hydrogen-bonding network of the protein moiety and the chromophore, chromophore attachment and photoreversibility were completely impaired. FphA is a functional sensor histidine kinase with a strong red-light-dependent autophosphorylation activity. Furthermore, intermolecular trans-phosphorylation to the response regulator domain of a second monomer could be demonstrated. Interestingly, co-incubation of FphA and FphA variants led to enhanced autophosphorylation, including the "inactive" Pr form. The latter observed phenomenon might suggest that auto- and trans-phosphorylation activity is modulated by additional interaction partners leading to variable phosphorylation events that trigger a specific output response.

  5. Oxidative stress-induced calcium signalling in Aspergillus nidulans.

    PubMed

    Greene, Vilma; Cao, Hong; Schanne, Francis A X; Bartelt, Diana C

    2002-05-01

    The effects of oxidative stress on levels of calcium ion (Ca(2+)) in Aspergillus nidulans were measured using strains expressing aequorin in the cytoplasm (Aeq(cyt)) and mitochondria (Aeq(mt)). When oxidative stress was induced by exposure to 10-mM H(2)O(2), the mitochondrial calcium response (Ca(mt)(2+)) was greater than the change in cytoplasmic calcium (Ca(c)(2+)). The Ca(mt)(2+) response to H(2)O(2) was dose dependent, while the increase in [Ca(c)(2+)] did not change with increasing H(2)O(2). The increase in both [Ca(c)(2+)] and [Ca(mt)(2+)] in response to oxidative stress was enhanced by exposure of cells to Ca(2+). The presence of chelator in the external medium only partially inhibited the Ca(mt)(2+) and Ca(c)(2+) responses to oxidative stress. Reagents that alter calcium fluxes had varied effects on the Ca(mt)(2+) response to peroxide. Ruthenium red blocked the increase in [Ca(mt)(2+)], while neomycin caused an even greater increase in [Ca(mt)(2+)]. Treatment with ruthenium red and neomycin had no effect on the Ca(c)(2+) response. Bafilomycin A and oligomycin had no effect on either the mitochondrial or cytoplasmic response. Inhibitors of both voltage-regulated calcium channels and intracellular calcium release channels inhibited the Ca(2+)-dependent component of the Ca(mt)(2+) response to oxidative stress. We conclude that the more significant Ca(2+) response to oxidative stress occurs in the mitochondria and that both intracellular and extracellular calcium pools can contribute to the increases in [Ca(c)(2+)] and [Ca(mt)(2+)] induced by oxidative stress.

  6. SAGA complex components and acetate repression in Aspergillus nidulans.

    PubMed

    Georgakopoulos, Paraskevi; Lockington, Robin A; Kelly, Joan M

    2012-11-01

    Alongside the well-established carbon catabolite repression by glucose and other sugars, acetate causes repression in Aspergillus nidulans. Mutations in creA, encoding the transcriptional repressor involved in glucose repression, also affect acetate repression, but mutations in creB or creC, encoding components of a deubiquitination system, do not. To understand the effects of acetate, we used a mutational screen that was similar to screens that uncovered mutations in creA, creB, and creC, except that glucose was replaced by acetate to identify mutations that were affected for repression by acetate but not by glucose. We uncovered mutations in acdX, homologous to the yeast SAGA component gene SPT8, which in growth tests showed derepression for acetate repression but not for glucose repression. We also made mutations in sptC, homologous to the yeast SAGA component gene SPT3, which showed a similar phenotype. We found that acetate repression is complex, and analysis of facA mutations (lacking acetyl CoA synthetase) indicates that acetate metabolism is required for repression of some systems (proline metabolism) but not for others (acetamide metabolism). Although plate tests indicated that acdX- and sptC-null mutations led to derepressed alcohol dehydrogenase activity, reverse-transcription quantitative real-time polymerase chain reaction showed no derepression of alcA or aldA but rather elevated induced levels. Our results indicate that acetate repression is due to repression via CreA together with metabolic changes rather than due to an independent regulatory control mechanism.

  7. The regulation of phosphoenolpyruvate carboxykinase and the NADP-linked malic enzyme in Aspergillus nidulans.

    PubMed

    Kelly, J M; Hynes, M J

    1981-04-01

    It has previously been suggested that the synthesis of phosphoenolpyruvate carboxykinase (EC 4.1.1.32) in Aspergillus nidulans is regulated by a repression-derepression mechanism involving a glycolytic intermediate, and not by induction. Results obtained using compounds that enter the tricarboxylic acid cycle via 2-oxoglutarate, and that can supply both a carbon and a nitrogen source for A. nidulans, suggest it is more likely that the synthesis of phosphoenolpyruvate carboxykinase is inducible, and only weakly regulated by carbon catabolite repression. a similar study of the regulation of the NADP-linked malic enzyme (EC 1.1.1.40) indicates that it too may be inducible.

  8. Characterization of the Aspergillus nidulans septin (asp) gene family.

    PubMed Central

    Momany, M; Zhao, J; Lindsey, R; Westfall, P J

    2001-01-01

    Members of the septin gene family are involved in cytokinesis and the organization of new growth in organisms as diverse as yeast, fruit fly, worm, mouse, and human. Five septin genes have been cloned and sequenced from the model filamentous fungus A. nidulans. As expected, the A. nidulans septins contain the highly conserved GTP binding and coiled-coil domains seen in other septins. On the basis of hybridization of clones to a chromosome-specific library and correlation with an A. nidulans physical map, the septins are not clustered but are scattered throughout the genome. In phylogenetic analysis most fungal septins could be grouped with one of the prototypical S. cerevisiae septins, Cdc3, Cdc10, Cdc11, and Cdc12. Intron-exon structure was conserved within septin classes. The results of this study suggest that most fungal septins belong to one of four orthologous classes. PMID:11238387

  9. Searching for gold beyond mitosis: Mining intracellular membrane traffic in Aspergillus nidulans.

    PubMed

    Peñalva, Miguel A; Galindo, Antonio; Abenza, Juan F; Pinar, Mario; Calcagno-Pizarelli, Ana M; Arst, Herbert N; Pantazopoulou, Areti

    2012-01-01

    The genetically tractable filamentous ascomycete fungus Aspergillus nidulans has been successfully exploited to gain major insight into the eukaryotic cell cycle. More recently, its amenability to in vivo multidimensional microscopy has fueled a potentially gilded second age of A. nidulans cell biology studies. This review specifically deals with studies on intracellular membrane traffic in A. nidulans. The cellular logistics are subordinated to the needs imposed by the polarized mode of growth of the multinucleated hyphal tip cells, whereas membrane traffic is adapted to the large intracellular distances. Recent work illustrates the usefulness of this fungus for morphological and biochemical studies on endosome and Golgi maturation, and on the role of microtubule-dependent motors in the long-distance movement of endosomes. The fungus is ideally suited for genetic studies on the secretory pathway, as mutations impairing secretion reduce apical extension rates, resulting in phenotypes detectable by visual inspection of colonies. PMID:22645705

  10. Gβ-Like CpcB Plays a Crucial Role for Growth and Development of Aspergillus nidulans and Aspergillus fumigatus

    PubMed Central

    Kong, Qing; Wang, Long; Liu, Zengran; Kwon, Nak-Jung; Kim, Sun Chang; Yu, Jae-Hyuk

    2013-01-01

    Growth, development, virulence and secondary metabolism in fungi are governed by heterotrimeric G proteins (G proteins). A Gβ-like protein called Gib2 has been shown to function as an atypical Gβ in Gpa1-cAMP signaling in Cryptococcus neoformans. We found that the previously reported CpcB (cross pathway control B) protein is the ortholog of Gib2 in Aspergillus nidulans and Aspergillus fumigatus. In this report, we further characterize the roles of CpcB in governing growth, development and toxigenesis in the two aspergilli. The deletion of cpcB results in severely impaired cellular growth, delayed spore germination, and defective asexual sporulation (conidiation) in both aspergilli. Moreover, CpcB is necessary for proper expression of the key developmental activator brlA during initiation and progression of conidiation in A. nidulans and A. fumigatus. Somewhat in accordance with the previous study, the absence of cpcB results in the formation of fewer, but not micro-, cleistothecia in A. nidulans in the presence of wild type veA, an essential activator of sexual development. However, the cpcB deletion mutant cleistothecia contain no ascospores, validating that CpcB is required for progression and completion of sexual fruiting including ascosporogenesis. Furthermore, unlike the canonical GβSfaD, CpcB is not needed for the biosynthesis of the mycotoxin sterigmatocystin (ST) as the cpcB null mutant produced reduced amount of ST with unaltered STC gene expression. However, in A. fumigatus, the deletion of cpcB results in the blockage of gliotoxin (GT) production. Further genetic analyses in A. nidulans indicate that CpcB may play a central role in vegetative growth, which might be independent of FadA- and GanB-mediated signaling. A speculative model summarizing the roles of CpcB in conjunction with SfaD in A. nidulans is presented. PMID:23936193

  11. Identification and Characterization of a Novel Diterpene Gene Cluster in Aspergillus nidulans

    PubMed Central

    Bromann, Kirsi; Toivari, Mervi; Viljanen, Kaarina; Vuoristo, Anu; Ruohonen, Laura; Nakari-Setälä, Tiina

    2012-01-01

    Fungal secondary metabolites are a rich source of medically useful compounds due to their pharmaceutical and toxic properties. Sequencing of fungal genomes has revealed numerous secondary metabolite gene clusters, yet products of many of these biosynthetic pathways are unknown since the expression of the clustered genes usually remains silent in normal laboratory conditions. Therefore, to discover new metabolites, it is important to find ways to induce the expression of genes in these otherwise silent biosynthetic clusters. We discovered a novel secondary metabolite in Aspergillus nidulans by predicting a biosynthetic gene cluster with genomic mining. A Zn(II)2Cys6–type transcription factor, PbcR, was identified, and its role as a pathway-specific activator for the predicted gene cluster was demonstrated. Overexpression of pbcR upregulated the transcription of seven genes in the identified cluster and led to the production of a diterpene compound, which was characterized with GC/MS as ent-pimara-8(14),15-diene. A change in morphology was also observed in the strains overexpressing pbcR. The activation of a cryptic gene cluster by overexpression of its putative Zn(II)2Cys6–type transcription factor led to discovery of a novel secondary metabolite in Aspergillus nidulans. Quantitative real-time PCR and DNA array analysis allowed us to predict the borders of the biosynthetic gene cluster. Furthermore, we identified a novel fungal pimaradiene cyclase gene as well as genes encoding 3-hydroxy-3-methyl-glutaryl-coenzyme A (HMG-CoA) reductase and a geranylgeranyl pyrophosphate (GGPP) synthase. None of these genes have been previously implicated in the biosynthesis of terpenes in Aspergillus nidulans. These results identify the first Aspergillus nidulans diterpene gene cluster and suggest a biosynthetic pathway for ent-pimara-8(14),15-diene. PMID:22506079

  12. Reengineering an azaphilone biosynthesis pathway in Aspergillus nidulans to create lipoxygenase inhibitors.

    PubMed

    Somoza, Amber D; Lee, Kuan-Han; Chiang, Yi-Ming; Oakley, Berl R; Wang, Clay C C

    2012-02-17

    Sclerotiorin, an azaphilone polyketide, is a bioactive natural product known to inhibit 15-lipoxygenase and many other biological targets. To readily access sclerotiorin and analogs, we developed a 2-3 step semisynthetic route to produce a variety of azaphilones starting from an advanced, putative azaphilone intermediate (5) overproduced by an engineered strain of Aspergillus nidulans. The inhibitory activities of the semisynthetic azaphilones against 15-lipoxygenase were evaluated with several compounds displaying low micromolar potency. PMID:22296232

  13. Identification and characterization of a novel diterpene gene cluster in Aspergillus nidulans.

    PubMed

    Bromann, Kirsi; Toivari, Mervi; Viljanen, Kaarina; Vuoristo, Anu; Ruohonen, Laura; Nakari-Setälä, Tiina

    2012-01-01

    Fungal secondary metabolites are a rich source of medically useful compounds due to their pharmaceutical and toxic properties. Sequencing of fungal genomes has revealed numerous secondary metabolite gene clusters, yet products of many of these biosynthetic pathways are unknown since the expression of the clustered genes usually remains silent in normal laboratory conditions. Therefore, to discover new metabolites, it is important to find ways to induce the expression of genes in these otherwise silent biosynthetic clusters. We discovered a novel secondary metabolite in Aspergillus nidulans by predicting a biosynthetic gene cluster with genomic mining. A Zn(II)(2)Cys(6)-type transcription factor, PbcR, was identified, and its role as a pathway-specific activator for the predicted gene cluster was demonstrated. Overexpression of pbcR upregulated the transcription of seven genes in the identified cluster and led to the production of a diterpene compound, which was characterized with GC/MS as ent-pimara-8(14),15-diene. A change in morphology was also observed in the strains overexpressing pbcR. The activation of a cryptic gene cluster by overexpression of its putative Zn(II)(2)Cys(6)-type transcription factor led to discovery of a novel secondary metabolite in Aspergillus nidulans. Quantitative real-time PCR and DNA array analysis allowed us to predict the borders of the biosynthetic gene cluster. Furthermore, we identified a novel fungal pimaradiene cyclase gene as well as genes encoding 3-hydroxy-3-methyl-glutaryl-coenzyme A (HMG-CoA) reductase and a geranylgeranyl pyrophosphate (GGPP) synthase. None of these genes have been previously implicated in the biosynthesis of terpenes in Aspergillus nidulans. These results identify the first Aspergillus nidulans diterpene gene cluster and suggest a biosynthetic pathway for ent-pimara-8(14),15-diene.

  14. Small heat shock proteins, phylogeny in filamentous fungi and expression analyses in Aspergillus nidulans.

    PubMed

    Wu, Jianbing; Wang, Mingshuang; Zhou, Liting; Yu, Dongliang

    2016-01-10

    Small heat shock proteins (sHSPs) have been characterized in organisms from all three domains of life and viruses and are involved in a wide range of biological functions. However, the evolution and function of sHSP in Aspergillus species are largely unknown. In the present work, sHSPs were identified in 31 filamentous fungi, including species from Aspergillus, Penicillium, Fusarium and Magnaporthe, as well as Botrytis cinerea and Neurospora crassa. Phylogenetic analysis revealed high level of divergence of sHSPs among filamentous fungi that orthologs could be only found between very closely related species. Strikingly, duplication of shsp genes occurred in genera Penicillium and also Aspergillus nidulans was observed, which might be an important pathway of sHSPs evolution. Expression analysis of shsp genes revealed that sHSPs were involved in response of A. nidulans to various conditions, including cold/heat as well as oxidative and osmotic stresses, and that the recent duplicated sHSPs in A. nidulans had highly similar function. PMID:26403724

  15. Cross-talk between light and glucose regulation controls toxin production and morphogenesis in Aspergillus nidulans

    PubMed Central

    Atoui, A.; Kastner, C.; Larey, C.M.; Thokala, R.; Etxebeste, O.; Espeso, E.A.; Fischer, R.

    2010-01-01

    Light is a major environmental stimulus that has a broad effect on organisms, triggering a cellular response that results in an optimal adaptation enhancing fitness and survival. In fungi, light affects growth, and causes diverse morphological changes such as those leading to reproduction. Light can also affect fungal metabolism, including the biosynthesis of natural products. In this study we show that in Aspergillus nidulans the effect of light on the production of the sterigmatocystin (ST) toxin depends on the glucose concentration. In cultures grown with 1% glucose and exposed to light, ST production was lower than when grown in the dark. This lower ST production coincided with an elevated rate of cellular damage with partial loss of nuclear integrity and vacuolated cytoplasm. However, in cultures grown with 2% glucose these effects were reversed and light enhanced ST production. Glucose abundance also affected the light-dependent subcellular localization of the VeA (velvet) protein, a key regulator necessary for normal light-dependent morphogenesis and secondary metabolism in Aspergilli and other fungal genera. The role of other VeA-associated proteins, particularly the blue light-sensing proteins LreA and LreB (WC-1 and WC-2 orthologs), on conidiation could also be modified by the abundance of glucose. We also show that LreA and LreB, as well as the phytochrome FphA, modulate not only the synthesis of sterigmatocystin, but also the production of the antibiotic penicillin. PMID:20816830

  16. Functional characterization of a xylose transporter in Aspergillus nidulans

    PubMed Central

    2014-01-01

    Background The production of bioethanol from lignocellulosic feedstocks will only become economically feasible when the majority of cellulosic and hemicellulosic biopolymers can be efficiently converted into bioethanol. The main component of cellulose is glucose, whereas hemicelluloses mainly consist of pentose sugars such as D-xylose and L-arabinose. The genomes of filamentous fungi such as A. nidulans encode a multiplicity of sugar transporters with broad affinities for hexose and pentose sugars. Saccharomyces cerevisiae, which has a long history of use in industrial fermentation processes, is not able to efficiently transport or metabolize pentose sugars (e.g. xylose). Subsequently, the aim of this study was to identify xylose-transporters from A. nidulans, as potential candidates for introduction into S. cerevisiae in order to improve xylose utilization. Results In this study, we identified the A. nidulans xtrD (xylose transporter) gene, which encodes a Major Facilitator Superfamily (MFS) transporter, and which was specifically induced at the transcriptional level by xylose in a XlnR-dependent manner, while being partially repressed by glucose in a CreA-dependent manner. We evaluated the ability of xtrD to functionally complement the S. cerevisiae EBY.VW4000 strain which is unable to grow on glucose, fructose, mannose or galactose as single carbon source. In S. cerevisiae, XtrD was targeted to the plasma membrane and its expression was able to restore growth on xylose, glucose, galactose, and mannose as single carbon sources, indicating that this transporter accepts multiple sugars as a substrate. XtrD has a high affinity for xylose, and may be a high affinity xylose transporter. We were able to select a S. cerevisiae mutant strain that had increased xylose transport when expressing the xtrD gene. Conclusions This study characterized the regulation and substrate specificity of an A. nidulans transporter that represents a good candidate for further directed

  17. Induction of apoptosis by sphingoid long-chain bases in Aspergillus nidulans.

    PubMed

    Cheng, Jijun; Park, Tae-Sik; Chio, Li-Chun; Fischl, Anthony S; Ye, Xiang S

    2003-01-01

    Sphingolipid metabolism is implicated to play an important role in apoptosis. Here we show that dihydrosphingosine (DHS) and phytosphingosine (PHS), two major sphingoid bases of fungi, have potent fungicidal activity with remarkably high structural and stereochemical specificity against Aspergillus nidulans. In fact, only naturally occurring DHS and PHS are active. Further analysis revealed that DHS and PHS induce rapid DNA condensation independent of mitosis, large-scale DNA fragmentation, and exposure of phosphatidylserine, all common morphological features characteristic of apoptosis, suggesting that DHS and PHS induce apoptosis in A. nidulans. The finding that DNA fragmentation requires protein synthesis, which implies that an active process is involved, further supports this proposition. The induction of apoptosis by DHS and PHS is associated with the rapid accumulation of reactive oxygen species (ROS). However, ROS are not required for apoptosis induced by DHS and PHS, as scavenging of ROS by a free radical spin trap has no effect. We further demonstrate that apoptosis induced by DHS and PHS is independent of metacaspase function but requires mitochondrial function. Together, the results suggest that DHS and PHS induce a type of apoptosis in A. nidulans most similar to the caspase-independent apoptosis observed in mammalian systems. As A. nidulans is genetically tractable, this organism should be an ideal model system for dissecting sphingolipid signaling in apoptosis and, importantly, for further elucidating the molecular basis of caspase-independent apoptosis. PMID:12482970

  18. Ammonium-induced internalisation of UapC, the general purine permease from Aspergillus nidulans.

    PubMed

    Valdez-Taubas, Javier; Harispe, Laura; Scazzocchio, Claudio; Gorfinkiel, Lisette; Rosa, Alberto L

    2004-01-01

    The Aspergillus nidulans UapC protein is a high-affinity, moderate-capacity, uric acid-xanthine transporter, which also displays a low transport capacity for hypoxanthine, adenine, and guanine. It has been previously shown that a functional UapC-GFP fusion protein localises at the plasma membrane. Here, we demonstrate that ammonium, a preferred nitrogen source, dramatically changes the subcellular distribution of UapC. After addition of ammonium, UapC-GFP is removed from the plasma membrane and is concentrated into the vacuolar compartment. A chimeric gene construct in which an inducible promoter, insensitive to nitrogen repression, drives the expression of UapC-GFP, allowed us to demonstrate that the ammonium-dependent redistribution of UapC can be dissociated from the transcriptional repression of the gene. These results provide further support for the occurrence of endocytosis and the lysosomal-endosomal function of the vacuolar compartment in A. nidulans.

  19. Regulation of the Aspergillus nidulans pectate lyase gene (pelA).

    PubMed Central

    Dean, R A; Timberlake, W E

    1989-01-01

    Aspergillus nidulans pectate lyase was purified from culture filtrates. The enzyme catalyzed a random eliminative cleavage reaction, had an apparent molecular weight of 40,000, and a pl of 4.2. Pectate lyase antisera were produced and used to identify pectate lyase clones in a cDNA expression library. Thirteen of 14 clones identified immunologically cross-hybridized. The identity of the single-copy pectate lyase gene, which we designated pelA, was confirmed in two ways. First, several cDNA clones expressed pectate lyase activity in Escherichia coli. Second, targeted mutation of the gene in A. nidulans resulted in complete loss of enzyme activity. pelA encodes a 1,300-nucleotide mRNA that was present in cells grown with polygalacturonic acid as carbon source but absent from cells grown with glucose or acetate as carbon source. Thus, pectate lyase expression is regulated at the level of mRNA accumulation. PMID:2535502

  20. Multiple Phosphatases Regulate Carbon Source-Dependent Germination and Primary Metabolism in Aspergillus nidulans.

    PubMed

    de Assis, Leandro José; Ries, Laure Nicolas Annick; Savoldi, Marcela; Dinamarco, Taisa Magnani; Goldman, Gustavo Henrique; Brown, Neil Andrew

    2015-03-11

    Aspergillus nidulans is an important mold and a model system for the study of fungal cell biology. In addition, invasive A. nidulans pulmonary infections are common in humans with chronic granulomatous disease. The morphological and biochemical transition from dormant conidia into active, growing, filamentous hyphae requires the coordination of numerous biosynthetic, developmental, and metabolic processes. The present study exhibited the diversity of roles performed by seven phosphatases in regulating cell cycle, development, and metabolism in response to glucose and alternative carbon sources. The identified phosphatases highlighted the importance of several signaling pathways regulating filamentous growth, the action of the pyruvate dehydrogenase complex as a metabolic switch controlling carbon usage, and the identification of the key function performed by the α-ketoglutarate dehydrogenase during germination. These novel insights into the fundamental roles of numerous phosphatases in germination and carbon sensing have provided new avenues of research into the identification of inhibitors of fungal germination, with implications for the food, feed, and pharmaceutical industries.

  1. Protein Kinase C (PkcA) of Aspergillus nidulans Is Involved in Penicillin Production

    PubMed Central

    Herrmann, Martina; Spröte, Petra; Brakhage, Axel A.

    2006-01-01

    The biosynthesis of the β-lactam antibiotic penicillin in the filamentous fungus Aspergillus nidulans is catalyzed by three enzymes that are encoded by the acvA, ipnA, and aatA genes. A variety of cis-acting DNA elements and regulatory factors form a complex regulatory network controlling these β-lactam biosynthesis genes. Regulators involved include the CCAAT-binding complex AnCF and AnBH1. AnBH1 acts as a repressor of the penicillin biosynthesis gene aatA. Until now, however, little information has been available on the signal transduction cascades leading to the transcription factors. Here we show that inhibition of protein kinase C (Pkc) activity in A. nidulans led to cytoplasmic localization of an AnBH1-enhanced green fluorescent protein (EGFP) fusion protein. Computer analysis of the genome and screening of an A. nidulans gene library revealed that the fungus possesses two putative Pkc-encoding genes, which we designated pkcA and pkcB. Only PkcA showed all the characteristic features of fungal Pkc's. Production of pkcA antisense RNA in A. nidulans led to reduced growth and conidiation in Aspergillus minimal medium, while in fermentation medium it led to enhanced expression of an aatAp-lacZ gene fusion, reduced pencillin production, and predominantly cytoplasmic localization of AnBH1. These data agree with the finding that inhibition of Pkc activity prevented nuclear localization of AnBH1-EGFP. As a result, repression of aatA expression was relieved. The involvement of Pkc in penicillin biosynthesis is also interesting in light of the fact that in the yeast Saccharomyces cerevisiae, Pkc plays a major role in maintaining cell integrity. PMID:16598003

  2. Protein kinase C (PkcA) of Aspergillus nidulans is involved in penicillin production.

    PubMed

    Herrmann, Martina; Spröte, Petra; Brakhage, Axel A

    2006-04-01

    The biosynthesis of the beta-lactam antibiotic penicillin in the filamentous fungus Aspergillus nidulans is catalyzed by three enzymes that are encoded by the acvA, ipnA, and aatA genes. A variety of cis-acting DNA elements and regulatory factors form a complex regulatory network controlling these beta-lactam biosynthesis genes. Regulators involved include the CCAAT-binding complex AnCF and AnBH1. AnBH1 acts as a repressor of the penicillin biosynthesis gene aatA. Until now, however, little information has been available on the signal transduction cascades leading to the transcription factors. Here we show that inhibition of protein kinase C (Pkc) activity in A. nidulans led to cytoplasmic localization of an AnBH1-enhanced green fluorescent protein (EGFP) fusion protein. Computer analysis of the genome and screening of an A. nidulans gene library revealed that the fungus possesses two putative Pkc-encoding genes, which we designated pkcA and pkcB. Only PkcA showed all the characteristic features of fungal Pkc's. Production of pkcA antisense RNA in A. nidulans led to reduced growth and conidiation in Aspergillus minimal medium, while in fermentation medium it led to enhanced expression of an aatAp-lacZ gene fusion, reduced pencillin production, and predominantly cytoplasmic localization of AnBH1. These data agree with the finding that inhibition of Pkc activity prevented nuclear localization of AnBH1-EGFP. As a result, repression of aatA expression was relieved. The involvement of Pkc in penicillin biosynthesis is also interesting in light of the fact that in the yeast Saccharomyces cerevisiae, Pkc plays a major role in maintaining cell integrity.

  3. Hypertonic conditions trigger transient plasmolysis, growth arrest and blockage of transporter endocytosis in Aspergillus nidulans and Saccharomyces cerevisiae.

    PubMed

    Bitsikas, Vassilis; Karachaliou, Mayia; Gournas, Christos; Diallinas, George

    2011-01-01

    By using Aspergillus nidulans strains expressing functional GFP-tagged transporters under hypertonic conditions, we noticed the rapid appearance of cortical, relatively static, fluorescent patches (0.5-2.3 μm). These patches do not correspond to transporter microdomains as they co-localize with other plasma membrane-associated molecules, such as the pleckstrin homology (PH) domain and the SsoA t-Snare, or the lipophilic markers FM4-64 and filipin. In addition, they do not show characteristics of lipid rafts, MCCs or other membrane microdomains. Deconvoluted microscopic images showed that fluorescent patches correspond to plasma membrane invaginations. Transporters remain fully active during this phenomenon of localized plasmolysis. Plasmolysis was however associated with reduced growth rate and a dramatic blockage in transporter and FM4-64 endocytosis. These phenomena are transient and rapidly reversible upon wash-out of hypertonic media. Based on the observation that block in endocytosis by hypertonic treatment altered dramatically the cellular localization of tropomyosin (GFP-TpmA), although it did not affect the cortical appearance of upstream (SlaB-GFP) or downstream (AbpA-mRFP) endocytic components, we conclude that hypertonicity modifies actin dynamics and thus acts indirectly on endocytosis. This was further supported by the effect of latrunculin B, an actin depolymerization agent, on endocytosis. We show that the phenomena observed in A. nidulans also occur in Saccharomyces cerevisiae, suggesting that they constitute basic homeostatic responses of ascomycetes to hypertonic shock. Finally, our work shows that hypertonic treatments can be used as physiological tools to study the endocytic down-regulation of transporters in A. nidulans, as non-conditional genetic blocks affecting endocytic internalization are lethal or severely debilitating.

  4. Hypertonic conditions trigger transient plasmolysis, growth arrest and blockage of transporter endocytosis in Aspergillus nidulans and Saccharomyces cerevisiae.

    PubMed

    Bitsikas, Vassilis; Karachaliou, Mayia; Gournas, Christos; Diallinas, George

    2011-01-01

    By using Aspergillus nidulans strains expressing functional GFP-tagged transporters under hypertonic conditions, we noticed the rapid appearance of cortical, relatively static, fluorescent patches (0.5-2.3 μm). These patches do not correspond to transporter microdomains as they co-localize with other plasma membrane-associated molecules, such as the pleckstrin homology (PH) domain and the SsoA t-Snare, or the lipophilic markers FM4-64 and filipin. In addition, they do not show characteristics of lipid rafts, MCCs or other membrane microdomains. Deconvoluted microscopic images showed that fluorescent patches correspond to plasma membrane invaginations. Transporters remain fully active during this phenomenon of localized plasmolysis. Plasmolysis was however associated with reduced growth rate and a dramatic blockage in transporter and FM4-64 endocytosis. These phenomena are transient and rapidly reversible upon wash-out of hypertonic media. Based on the observation that block in endocytosis by hypertonic treatment altered dramatically the cellular localization of tropomyosin (GFP-TpmA), although it did not affect the cortical appearance of upstream (SlaB-GFP) or downstream (AbpA-mRFP) endocytic components, we conclude that hypertonicity modifies actin dynamics and thus acts indirectly on endocytosis. This was further supported by the effect of latrunculin B, an actin depolymerization agent, on endocytosis. We show that the phenomena observed in A. nidulans also occur in Saccharomyces cerevisiae, suggesting that they constitute basic homeostatic responses of ascomycetes to hypertonic shock. Finally, our work shows that hypertonic treatments can be used as physiological tools to study the endocytic down-regulation of transporters in A. nidulans, as non-conditional genetic blocks affecting endocytic internalization are lethal or severely debilitating. PMID:20919858

  5. Heterologous Expression of Fungal Secondary Metabolite Pathways in the Aspergillus nidulans Host System.

    PubMed

    van Dijk, J W A; Wang, C C C

    2016-01-01

    Heterologous expression of fungal secondary metabolite genes allows for the product formation of otherwise silent secondary metabolite biosynthesis pathways. It also allows facile expression of mutants or combinations of genes not found in nature. This capability makes model fungi an ideal platform for synthetic biology. In this chapter a detailed description is provided of how to heterologously express any fungal secondary metabolite gene(s) in a well-developed host strain of Aspergillus nidulans. It covers all the necessary steps from identifying a gene(s) of interest to culturing mutant strains to produce secondary metabolites.

  6. Heterologous Expression of Fungal Secondary Metabolite Pathways in the Aspergillus nidulans Host System.

    PubMed

    van Dijk, J W A; Wang, C C C

    2016-01-01

    Heterologous expression of fungal secondary metabolite genes allows for the product formation of otherwise silent secondary metabolite biosynthesis pathways. It also allows facile expression of mutants or combinations of genes not found in nature. This capability makes model fungi an ideal platform for synthetic biology. In this chapter a detailed description is provided of how to heterologously express any fungal secondary metabolite gene(s) in a well-developed host strain of Aspergillus nidulans. It covers all the necessary steps from identifying a gene(s) of interest to culturing mutant strains to produce secondary metabolites. PMID:27417927

  7. Laser capture microdissection to identify septum-associated proteins in Aspergillus nidulans.

    PubMed

    Zhang, Ying; Fischer, Reinhard; Teichert, Ines; Kück, Ulrich

    2016-01-01

    To spatially resolve genetic differences at the cellular level, the laser-capture microdissection technique was developed. With this method cells can be cut from tissues with a laser beam and analyzed for DNA, RNA or protein composition. Here we adapted the technique to isolate septal microtubule-organizing center (MTOC)-associated proteins in Aspergillus nidulans About 3000 septa were collected and subjected to peptide fingerprinting by mass-spectrometric analysis. We identified the microtubule polymerase AlpA and found it interacts with ApsB specifically at sMTOCs, suggesting that AlpA might be involved in the assembly or the functioning of this protein complex. PMID:26951366

  8. Aspergillus nidulans Ambient pH Signaling Does Not Require Endocytosis.

    PubMed

    Lucena-Agell, Daniel; Galindo, Antonio; Arst, Herbert N; Peñalva, Miguel A

    2015-06-01

    Aspergillus nidulans (Pal) ambient pH signaling takes place in cortical structures containing components of the ESCRT pathway, which are hijacked by the alkaline pH-activated, ubiquitin-modified version of the arrestin-like protein PalF and taken to the plasma membrane. There, ESCRTs scaffold the assembly of dedicated Pal proteins acting downstream. The molecular details of this pathway, which results in the two-step proteolytic processing of the transcription factor PacC, have received considerable attention due to the key role that it plays in fungal pathogenicity. While current evidence strongly indicates that the pH signaling role of ESCRT complexes is limited to plasma membrane-associated structures where PacC proteolysis would take place, the localization of the PalB protease, which almost certainly catalyzes the first and only pH-regulated proteolytic step, had not been investigated. In view of ESCRT participation, this formally leaves open the possibility that PalB activation requires endocytic internalization. As endocytosis is essential for hyphal growth, nonlethal endocytic mutations are predicted to cause an incomplete block. We used a SynA internalization assay to measure the extent to which any given mutation prevents endocytosis. We show that none of the tested mutations impairing endocytosis to different degrees, including slaB1, conditionally causing a complete block, have any effect on the activation of the pathway. We further show that PalB, like PalA and PalC, localizes to cortical structures in an alkaline pH-dependent manner. Therefore, signaling through the Pal pathway does not involve endocytosis.

  9. Genetic evidence for a microtubule-capture mechanism during polarised growth of Aspergillus nidulans.

    PubMed

    Manck, Raphael; Ishitsuka, Yuji; Herrero, Saturnino; Takeshita, Norio; Nienhaus, G Ulrich; Fischer, Reinhard

    2015-10-01

    The cellular switch from symmetry to polarity in eukaryotes depends on the microtubule (MT) and actin cytoskeletons. In fungi such as Schizosaccharomyces pombe or Aspergillus nidulans, the MT cytoskeleton determines the sites of actin polymerization through cortical cell-end marker proteins. Here we describe A. nidulans MT guidance protein A (MigA) as the first ortholog of the karyogamy protein Kar9 from Saccharomyces cerevisiae in filamentous fungi. A. nidulans MigA interacts with the cortical ApsA protein and is involved in spindle positioning during mitosis. MigA is also associated with septal and nuclear MT organizing centers (MTOCs). Super-resolution photoactivated localization microscopy (PALM) analyses revealed that MigA is recruited to assembling and retracting MT plus ends in an EbA-dependent manner. MigA is required for MT convergence in hyphal tips and plays a role in correct localization of the cell-end markers TeaA and TeaR. In addition, MigA interacts with a class-V myosin, suggesting that an active mechanism exists to capture MTs and to pull the ends along actin filaments. Hence, the organization of MTs and actin depend on each other, and positive feedback loops ensure robust polar growth. PMID:26272919

  10. Linkage of Oxidative Stress and Mitochondrial Dysfunctions to Spontaneous Culture Degeneration in Aspergillus nidulans*

    PubMed Central

    Li, Lin; Hu, Xiao; Xia, Yongliang; Xiao, Guohua; Zheng, Peng; Wang, Chengshu

    2014-01-01

    Filamentous fungi including mushrooms frequently and spontaneously degenerate during subsequent culture maintenance on artificial media, which shows the loss or reduction abilities of asexual sporulation, sexuality, fruiting, and production of secondary metabolites, thus leading to economic losses during mass production. To better understand the underlying mechanisms of fungal degeneration, the model fungus Aspergillus nidulans was employed in this study for comprehensive analyses. First, linkage of oxidative stress to culture degeneration was evident in A. nidulans. Taken together with the verifications of cell biology and biochemical data, a comparative mitochondrial proteome analysis revealed that, unlike the healthy wild type, a spontaneous fluffy sector culture of A. nidulans demonstrated the characteristics of mitochondrial dysfunctions. Relative to the wild type, the features of cytochrome c release, calcium overload and up-regulation of apoptosis inducing factors evident in sector mitochondria suggested a linkage of fungal degeneration to cell apoptosis. However, the sector culture could still be maintained for generations without the signs of growth arrest. Up-regulation of the heat shock protein chaperones, anti-apoptotic factors and DNA repair proteins in the sector could account for the compromise in cell death. The results of this study not only shed new lights on the mechanisms of spontaneous degeneration of fungal cultures but will also provide alternative biomarkers to monitor fungal culture degeneration. PMID:24345786

  11. A glucose-derepressed promoter for expression of heterologous products in the filamentous fungus Aspergillus nidulans.

    PubMed

    Hintz, W E; Lagosky, P A

    1993-07-01

    We describe a putative binding sequence (GCGGGGC) for the glucose-responsive repressor protein CreA at two positions upstream of the transcription start site of the alcohol dehydrogenase I (alcA) gene of Aspergillus nidulans. To positively identify the putative binding sites as CreA-specific, the GCGGGGC blocks were mutated at five internal nucleotide positions to GTACTAC and reintroduced into the wild type alcA promoter driving expression of the endogenous alcohol dehydrogenase I gene. This CreA-binding site variant was then transformed into an AlcR constitutive A. nidulans host strain (T2625) and growth was monitored in the presence of the non-metabolized glucose analogue, 2-deoxyglucose. Positive transformants were selected by their ability to grow using ethanol as a carbon source in the presence of 2-deoxyglucose. Similar CreA binding site variant alcA promoters should permit the alcA-driven expression of heterologous genes in A. nidulans in the presence of glucose, the preferred carbon source for biomass accumulation and provides a model for controlling carbon-catabolite regulated expression in other expression systems.

  12. A time course analysis of the extracellular proteome of Aspergillus nidulans growing on sorghum stover

    PubMed Central

    2012-01-01

    Background Fungi are important players in the turnover of plant biomass because they produce a broad range of degradative enzymes. Aspergillus nidulans, a well-studied saprophyte and close homologue to industrially important species such as A. niger and A. oryzae, was selected for this study. Results A. nidulans was grown on sorghum stover under solid-state culture conditions for 1, 2, 3, 5, 7 and 14 days. Based on analysis of chitin content, A. nidulans grew to be 4-5% of the total biomass in the culture after 2 days and then maintained a steady state of 4% of the total biomass for the next 12 days. A hyphal mat developed on the surface of the sorghum by day one and as seen by scanning electron microscopy the hyphae enmeshed the sorghum particles by day 5. After 14 days hyphae had penetrated the entire sorghum slurry. Analysis (1-D PAGE LC-MS/MS) of the secretome of A. nidulans, and analysis of the breakdown products from the sorghum stover showed a wide range of enzymes secreted. A total of 294 extracellular proteins were identified with hemicellulases, cellulases, polygalacturonases, chitinases, esterases and lipases predominating the secretome. Time course analysis revealed a total of 196, 166, 172 and 182 proteins on day 1, 3, 7 and 14 respectively. The fungus used 20% of the xylan and cellulose by day 7 and 30% by day 14. Cellobiose dehydrogenase, feruloyl esterases, and CAZy family 61 endoglucanases, all of which are thought to reduce the recalcitrance of biomass to hydrolysis, were found in high abundance. Conclusions Our results show that A. nidulans secretes a wide array of enzymes to degrade the major polysaccharides and lipids (but probably not lignin) by 1 day of growth on sorghum. The data suggests simultaneous breakdown of hemicellulose, cellulose and pectin. Despite secretion of most of the enzymes on day 1, changes in the relative abundances of enzymes over the time course indicates that the set of enzymes secreted is tailored to the

  13. Screening of medicinal plants for induction of somatic segregation activity in Aspergillus nidulans.

    PubMed

    Ramos Ruiz, A; De la Torre, R A; Alonso, N; Villaescusa, A; Betancourt, J; Vizoso, A

    1996-07-01

    Knowledge about mutagenic properties of plants commonly used in traditional medicine is limited. A screening for genotoxic activity was carried out in aqueous or alcoholic extracts prepared from 13 medicinal plants widely used as folk medicine in Cuba: Lepidium virginicum L. (Brassicaceae): Plantago major L. and Plantago lanceolata L. (Plantaginaceae); Ortosiphon aristatus Blume, Mentha x piperita L., Melissa officinalis L. and Plectranthus amboinicus (Lour.) Spreng. (Lamiaceae); Cymbopogon citratus (DC.) Stapf (Poaceae); Passiflora incarnata L. (Passifloraceae); Zingiber officinale Roscoe (Zingiberaceae); Piper auritum HBK. (Piperaceae); Schinus terebinthifolius Raddi (Anacardeaceae) and Momordica charantia L. (Cucurbitaceae). A plate incorporation assay with Aspergillus nidulans was employed, allowing detection of somatic segregation as a result of mitotic crossing-over, chromosome malsegregation or clastogenic effects. Aspergillus nidulans D-30, a well-marked strain carrying four recessive mutations for conidial color in heterozygosity, which permitted the direct visual detection of segregants, was used throughout this study. As a result, only in the aqueous extract of one of the plants screened (Momordica charantia) a statistical significant increase in the frequency of segregant sectors per colony was observed, and consequently, a genotoxic effect is postulated. PMID:8771452

  14. Screening of medicinal plants for induction of somatic segregation activity in Aspergillus nidulans.

    PubMed

    Ramos Ruiz, A; De la Torre, R A; Alonso, N; Villaescusa, A; Betancourt, J; Vizoso, A

    1996-07-01

    Knowledge about mutagenic properties of plants commonly used in traditional medicine is limited. A screening for genotoxic activity was carried out in aqueous or alcoholic extracts prepared from 13 medicinal plants widely used as folk medicine in Cuba: Lepidium virginicum L. (Brassicaceae): Plantago major L. and Plantago lanceolata L. (Plantaginaceae); Ortosiphon aristatus Blume, Mentha x piperita L., Melissa officinalis L. and Plectranthus amboinicus (Lour.) Spreng. (Lamiaceae); Cymbopogon citratus (DC.) Stapf (Poaceae); Passiflora incarnata L. (Passifloraceae); Zingiber officinale Roscoe (Zingiberaceae); Piper auritum HBK. (Piperaceae); Schinus terebinthifolius Raddi (Anacardeaceae) and Momordica charantia L. (Cucurbitaceae). A plate incorporation assay with Aspergillus nidulans was employed, allowing detection of somatic segregation as a result of mitotic crossing-over, chromosome malsegregation or clastogenic effects. Aspergillus nidulans D-30, a well-marked strain carrying four recessive mutations for conidial color in heterozygosity, which permitted the direct visual detection of segregants, was used throughout this study. As a result, only in the aqueous extract of one of the plants screened (Momordica charantia) a statistical significant increase in the frequency of segregant sectors per colony was observed, and consequently, a genotoxic effect is postulated.

  15. Functional Analysis of Sterol Transporter Orthologues in the Filamentous Fungus Aspergillus nidulans

    PubMed Central

    Bühler, Nicole; Hagiwara, Daisuke

    2015-01-01

    Polarized growth in filamentous fungi needs a continuous supply of proteins and lipids to the growing hyphal tip. One of the important membrane compounds in fungi is ergosterol. At the apical plasma membrane ergosterol accumulations, which are called sterol-rich plasma membrane domains (SRDs). The exact roles and formation mechanism of the SRDs remained unclear, although the importance has been recognized for hyphal growth. Transport of ergosterol to hyphal tips is thought to be important for the organization of the SRDs. Oxysterol binding proteins, which are conserved from yeast to human, are involved in nonvesicular sterol transport. In Saccharomyces cerevisiae seven oxysterol-binding protein homologues (OSH1 to -7) play a role in ergosterol distribution between closely located membranes independent of vesicle transport. We found five homologous genes (oshA to oshE) in the filamentous fungi Aspergillus nidulans. The functions of OshA-E were characterized by gene deletion and subcellular localization. Each gene-deletion strain showed characteristic phenotypes and different sensitivities to ergosterol-associated drugs. Green fluorescent protein-tagged Osh proteins showed specific localization in the late Golgi compartments, puncta associated with the endoplasmic reticulum, or diffusely in the cytoplasm. The genes expression and regulation were investigated in a medically important species Aspergillus fumigatus, as well as A. nidulans. Our results suggest that each Osh protein plays a role in ergosterol distribution at distinct sites and contributes to proper fungal growth. PMID:26116213

  16. The WOPR Domain Protein OsaA Orchestrates Development in Aspergillus nidulans

    PubMed Central

    Alkahyyat, Fahad; Ni, Min; Kim, Sun Chang; Yu, Jae-Hyuk

    2015-01-01

    Orchestration of cellular growth and development occurs during the life cycle of Aspergillus nidulans. A multi-copy genetic screen intended to unveil novel regulators of development identified the AN6578 locus predicted to encode a protein with the WOPR domain, which is a broadly present fungi-specific DNA-binding motif. Multi-copy of AN6578 disrupted the normal life cycle of the fungus leading to enhanced proliferation of vegetative cells, whereas the deletion resulted in hyper-active sexual fruiting with reduced asexual development (conidiation), thus named as osaA (Orchestrator of Sex and Asex). Further genetic studies indicate that OsaA balances development mainly by repressing sexual development downstream of the velvet regulator VeA. The absence of osaA is sufficient to suppress the veA1 allele leading to the sporulation levels comparable to veA+ wild type (WT). Genome-wide transcriptomic analyses of WT, veA1, and ΔosaA veA1 strains by RNA-Seq further corroborate that OsaA functions in repressing sexual development downstream of VeA. However, OsaA also plays additional roles in controlling development, as the ΔosaA veA1 mutant exhibits precocious and enhanced formation of Hülle cells compared to WT. The OsaA orthologue of Aspergillus flavus is able to complement the osaA null phenotype in A. nidulans, suggesting a conserved role of this group of WOPR domain proteins. In summary, OsaA is an upstream orchestrator of morphological and chemical development in Aspergillus that functions downstream of VeA. PMID:26359867

  17. Cloning and analysis of the positively acting regulatory gene amdR from Aspergillus nidulans

    SciTech Connect

    Andrianopoulos, A.; Hynes, M.J. )

    1988-08-01

    The positively acting regulatory gene amdR of Aspergillus nidulans coordinately regulates the expression of four unlinked structural genes involved in acetamide (amdS), omega amino acid (gatA and gabA), and lactam (lamA) catabolism. By the use of DNA-mediated transformation of A.nidulans, the amdR regulatory gene was cloned from a genomic cosmid library. Southern blot analysis of DNA from various loss-of-function amdR mutants revealed the presence of four detectable DNA rearrangements, including a deletion, an insertion, and a translocation. No detectable DNA rearrangements were found in several constitutive amdR/sup c/ mutants. The authors discuss an analysis of the fate of amdR-bearing plasmids in transformants which showed that 10 to 20% of the transformation events were homologous integrations or gene conversions. The authors describe how this phenomenon was exploited in developing a strategy by which amdr/sup c/ and amdR/sup -/ alleles can be readily cloned and analyzed. Examination of the transcription of amdR by Northern blot (RNA blot) analysis revealed the presence of two mRNAs (2.7 and 1.8 kilobases) which were constitutively synthesized at a very low level. In addition, amdR transcription did not appear to depend on the presence of a functional amdR product nor was it altered in amdR/sup c/ mutants.

  18. Characterization of Aspergillus nidulans α-glucan synthesis: roles for two synthases and two amylases.

    PubMed

    He, Xiaoxiao; Li, Shengnan; Kaminskyj, Susan G W

    2014-02-01

    Cell walls are essential for fungal survival and growth. Fungal walls are ∼ 90% carbohydrate, mostly types not found in humans, making them promising targets for anti-fungal drug development. Echinocandins, which inhibit the essential β-glucan synthase, are already clinically available. In contrast, α-glucan, another abundant fungal cell wall component has attracted relatively little research attention because it is not essential for most fungi. Aspergillus nidulans has two α-glucan synthases (AgsA and AgsB) and two α-amylases (AmyD and AmyG), all of which affect α-glucan synthesis. Gene deletion showed that AgsB was the major synthase. In addition, AmyG promoted α-glucan synthesis whereas AmyD had a repressive effect. The lack of α-glucan had no phenotypic impact on solid medium, but reduced conidial adhesion during germination in shaken liquid. Moreover, α-glucan level correlated with resistance to Calcofluor White. Intriguingly, overexpression of agsA could compensate for the loss of agsB at the α-glucan level, but not for phenotypic defects. Thus, products of AgsA and AgsB have different roles in the cell wall, consistent with agsA being mainly expressed at conidiation. These results suggest that α-glucan contributes to drug sensitivity and conidia adhesion in A. nidulans, and is differentially regulated by two synthases and two amylases.

  19. An Acidic Thermostable Recombinant Aspergillus nidulans Endoglucanase Is Active towards Distinct Agriculture Residues

    PubMed Central

    Tavares, Eveline Queiroz de Pinho; Rubini, Marciano Regis; Mello-de-Sousa, Thiago Machado; Duarte, Gilvan Caetano; de Faria, Fabrícia Paula; Ferreira Filho, Edivaldo Ximenes; Kyaw, Cynthia Maria; Silva-Pereira, Ildinete; Poças-Fonseca, Marcio Jose

    2013-01-01

    Aspergillus nidulans is poorly exploited as a source of enzymes for lignocellulosic residues degradation for biotechnological purposes. This work describes the A. nidulans Endoglucanase A heterologous expression in Pichia pastoris, the purification and biochemical characterization of the recombinant enzyme. Active recombinant endoglucanase A (rEG A) was efficiently secreted as a 35 kDa protein which was purified through a two-step chromatography procedure. The highest enzyme activity was detected at 50°C/pH 4. rEG A retained 100% of activity when incubated at 45 and 55°C for 72 h. Purified rEG A kinetic parameters towards CMC were determined as Km = 27.5 ± 4.33 mg/mL, Vmax = 1.185 ± 0.11 mmol/min, and 55.8 IU (international units)/mg specific activity. Recombinant P. pastoris supernatant presented hydrolytic activity towards lignocellulosic residues such as banana stalk, sugarcane bagasse, soybean residues, and corn straw. These data indicate that rEG A is suitable for plant biomass conversion into products of commercial importance, such as second-generation fuel ethanol. PMID:23936633

  20. G-protein coupled receptor-mediated nutrient sensing and developmental control in Aspergillus nidulans.

    PubMed

    Brown, Neil Andrew; Dos Reis, Thaila Fernanda; Ries, Laure Nicolas Annick; Caldana, Camila; Mah, Jae-Hyung; Yu, Jae-Hyuk; Macdonald, Jeffrey M; Goldman, Gustavo Henrique

    2015-10-01

    Nutrient sensing and utilisation are fundamental for all life forms. As heterotrophs, fungi have evolved a diverse range of mechanisms for sensing and taking up various nutrients. Despite its importance, only a limited number of nutrient receptors and their corresponding ligands have been identified in fungi. G-protein coupled receptors (GPCRs) are the largest family of transmembrane receptors. The Aspergillus nidulans genome encodes 16 putative GPCRs, but only a few have been functionally characterised. Our previous study showed the increased expression of an uncharacterised putative GPCR, gprH, during carbon starvation. GprH appears conserved throughout numerous filamentous fungi. Here, we reveal that GprH is a putative receptor involved in glucose and tryptophan sensing. The absence of GprH results in a reduction in cAMP levels and PKA activity upon adding glucose or tryptophan to starved cells. GprH is pre-formed in conidia and is increasingly active during carbon starvation, where it plays a role in glucose uptake and the recovery of hyphal growth. GprH also represses sexual development under conditions favouring sexual fruiting and during carbon starvation in submerged cultures. In summary, the GprH nutrient-sensing system functions upstream of the cAMP-PKA pathway, influences primary metabolism and hyphal growth, while represses sexual development in A. nidulans. PMID:26179439

  1. Multiple Phosphatases Regulate Carbon Source-Dependent Germination and Primary Metabolism in Aspergillus nidulans

    PubMed Central

    de Assis, Leandro José; Ries, Laure Nicolas Annick; Savoldi, Marcela; Dinamarco, Taisa Magnani; Goldman, Gustavo Henrique; Brown, Neil Andrew

    2015-01-01

    Aspergillus nidulans is an important mold and a model system for the study of fungal cell biology. In addition, invasive A. nidulans pulmonary infections are common in humans with chronic granulomatous disease. The morphological and biochemical transition from dormant conidia into active, growing, filamentous hyphae requires the coordination of numerous biosynthetic, developmental, and metabolic processes. The present study exhibited the diversity of roles performed by seven phosphatases in regulating cell cycle, development, and metabolism in response to glucose and alternative carbon sources. The identified phosphatases highlighted the importance of several signaling pathways regulating filamentous growth, the action of the pyruvate dehydrogenase complex as a metabolic switch controlling carbon usage, and the identification of the key function performed by the α-ketoglutarate dehydrogenase during germination. These novel insights into the fundamental roles of numerous phosphatases in germination and carbon sensing have provided new avenues of research into the identification of inhibitors of fungal germination, with implications for the food, feed, and pharmaceutical industries. PMID:25762568

  2. Functional characterization of the putative Aspergillus nidulans poly(ADP-ribose) polymerase homolog PrpA.

    PubMed

    Semighini, Camile P; Savoldi, Marcela; Goldman, Gustavo H; Harris, Steven D

    2006-05-01

    Poly(ADP-ribose) polymerase (PARP) is a highly conserved enzyme involved in multiple aspects of animal and plant cell physiology. For example, PARP is thought to be intimately involved in the early signaling events that trigger the DNA damage response. However, the genetic dissection of PARP function has been hindered by the presence of multiple homologs in most animal and plant species. Here, we present the first functional characterization of a putative PARP homolog (PrpA) in a microbial system (Aspergillus nidulans). PrpA belongs to a group of PARP homologs that includes representatives from filamentous fungi and protists. The genetic analysis of prpA demonstrates that it is an essential gene whose role in the DNA damage response is sensitive to gene dosage. Notably, temporal patterns of prpA expression and PrpA-GFP nuclear localization suggest that PrpA acts early in the A. nidulans DNA damage response. Additional studies implicate PrpA in farnesol-induced cell death and in the initiation of asexual development. Collectively, our results provide a gateway for probing the diverse functions of PARP in a sophisticated microbial genetic system.

  3. Analysis of acetate non-utilizing (acu) mutants in Aspergillus nidulans.

    PubMed

    Armitt, S; McCullough, W; Roberts, C F

    1976-02-01

    Genetic analysis of 119 acetate non-utilizing (acu) mutants in Aspergillus nidulans revealed ten new loci affecting acetate metabolism in addition to the three previously recognized on the basis of resistance to fluoroacetate and acetate non-utilization. The enzyme lesions associated with mutations at seven of the acu loci are described. These are: facA (= acuA), acetyl-CoA synthase; acuD, isocitrate lyase; acuE, malate synthase; acuF, phosphoenolpyruvate carboxykinase; acuG, fructose 1,6-diphosphatase; acuK and acuM, malic enzyme. The acu loci have been mapped and are widely distributed over the genome of A. nidulans. Close linkage has only been found between acuA and acuD (less than 1% recombination). There is no evidence for any pleiotropic mutation in that region affecting the expression of both these genes. Poor induction of the enzymes of the glyoxylate cycle, isocitrate lyase and malate synthase in mutants lacking acetyl-CoA synthase, and also in the other two classes of fluoroacetate-resistant mutants, indicates that the inducer, acetate, may be metabolized to a true metabolic inducer, perhaps acetyl-CoA, to effect formation of the enzymes. There is no evidence of any other class of pleiotropic recessive acu mutations affecting the expression of the acuD and acuE genes, which are therefore thought to be subject to negative rather than positive control.

  4. Functional analysis of alcS, a gene of the alc cluster in Aspergillus nidulans.

    PubMed

    Flipphi, Michel; Robellet, Xavier; Dequier, Emmanuel; Leschelle, Xavier; Felenbok, Béatrice; Vélot, Christian

    2006-04-01

    The ethanol utilization pathway (alc system) of Aspergillus nidulans requires two structural genes, alcA and aldA, which encode the two enzymes (alcohol dehydrogenase and aldehyde dehydrogenase, respectively) allowing conversion of ethanol into acetate via acetyldehyde, and a regulatory gene, alcR, encoding the pathway-specific autoregulated transcriptional activator. The alcR and alcA genes are clustered with three other genes that are also positively regulated by alcR, although they are dispensable for growth on ethanol. In this study, we characterized alcS, the most abundantly transcribed of these three genes. alcS is strictly co-regulated with alcA, and encodes a 262-amino acid protein. Sequence comparison with protein databases detected a putative conserved domain that is characteristic of the novel GPR1/FUN34/YaaH membrane protein family. It was shown that the AlcS protein is located in the plasma membrane. Deletion or overexpression of alcS did not result in any obvious phenotype. In particular, AlcS does not appear to be essential for the transport of ethanol, acetaldehyde or acetate. Basic Local Alignment Search Tool analysis against the A. nidulans genome led to the identification of two novel ethanol- and ethylacetate-induced genes encoding other members of the GPR1/FUN34/YaaH family, AN5226 and AN8390.

  5. Sequencing of Aspergillus nidulans and comparative analysis with A. fumigatus and A. oryzae.

    PubMed

    Galagan, James E; Calvo, Sarah E; Cuomo, Christina; Ma, Li-Jun; Wortman, Jennifer R; Batzoglou, Serafim; Lee, Su-In; Baştürkmen, Meray; Spevak, Christina C; Clutterbuck, John; Kapitonov, Vladimir; Jurka, Jerzy; Scazzocchio, Claudio; Farman, Mark; Butler, Jonathan; Purcell, Seth; Harris, Steve; Braus, Gerhard H; Draht, Oliver; Busch, Silke; D'Enfert, Christophe; Bouchier, Christiane; Goldman, Gustavo H; Bell-Pedersen, Deborah; Griffiths-Jones, Sam; Doonan, John H; Yu, Jaehyuk; Vienken, Kay; Pain, Arnab; Freitag, Michael; Selker, Eric U; Archer, David B; Peñalva, Miguel A; Oakley, Berl R; Momany, Michelle; Tanaka, Toshihiro; Kumagai, Toshitaka; Asai, Kiyoshi; Machida, Masayuki; Nierman, William C; Denning, David W; Caddick, Mark; Hynes, Michael; Paoletti, Mathieu; Fischer, Reinhard; Miller, Bruce; Dyer, Paul; Sachs, Matthew S; Osmani, Stephen A; Birren, Bruce W

    2005-12-22

    The aspergilli comprise a diverse group of filamentous fungi spanning over 200 million years of evolution. Here we report the genome sequence of the model organism Aspergillus nidulans, and a comparative study with Aspergillus fumigatus, a serious human pathogen, and Aspergillus oryzae, used in the production of sake, miso and soy sauce. Our analysis of genome structure provided a quantitative evaluation of forces driving long-term eukaryotic genome evolution. It also led to an experimentally validated model of mating-type locus evolution, suggesting the potential for sexual reproduction in A. fumigatus and A. oryzae. Our analysis of sequence conservation revealed over 5,000 non-coding regions actively conserved across all three species. Within these regions, we identified potential functional elements including a previously uncharacterized TPP riboswitch and motifs suggesting regulation in filamentous fungi by Puf family genes. We further obtained comparative and experimental evidence indicating widespread translational regulation by upstream open reading frames. These results enhance our understanding of these widely studied fungi as well as provide new insight into eukaryotic genome evolution and gene regulation. PMID:16372000

  6. Breaking the Silence: Protein Stabilization Uncovers Silenced Biosynthetic Gene Clusters in the Fungus Aspergillus nidulans

    PubMed Central

    Gerke, Jennifer; Bayram, Özgür; Feussner, Kirstin; Landesfeind, Manuel; Shelest, Ekaterina; Feussner, Ivo

    2012-01-01

    The genomes of filamentous fungi comprise numerous putative gene clusters coding for the biosynthesis of chemically and structurally diverse secondary metabolites (SMs), which are rarely expressed under laboratory conditions. Previous approaches to activate these genes were based primarily on artificially targeting the cellular protein synthesis apparatus. Here, we applied an alternative approach of genetically impairing the protein degradation apparatus of the model fungus Aspergillus nidulans by deleting the conserved eukaryotic csnE/CSN5 deneddylase subunit of the COP9 signalosome. This defect in protein degradation results in the activation of a previously silenced gene cluster comprising a polyketide synthase gene producing the antibiotic 2,4-dihydroxy-3-methyl-6-(2-oxopropyl)benzaldehyde (DHMBA). The csnE/CSN5 gene is highly conserved in fungi, and therefore, the deletion is a feasible approach for the identification of new SMs. PMID:23001671

  7. The 2008 update of the Aspergillus nidulans genome annotation: a community effort.

    PubMed

    Wortman, Jennifer Russo; Gilsenan, Jane Mabey; Joardar, Vinita; Deegan, Jennifer; Clutterbuck, John; Andersen, Mikael R; Archer, David; Bencina, Mojca; Braus, Gerhard; Coutinho, Pedro; von Döhren, Hans; Doonan, John; Driessen, Arnold J M; Durek, Pawel; Espeso, Eduardo; Fekete, Erzsébet; Flipphi, Michel; Estrada, Carlos Garcia; Geysens, Steven; Goldman, Gustavo; de Groot, Piet W J; Hansen, Kim; Harris, Steven D; Heinekamp, Thorsten; Helmstaedt, Kerstin; Henrissat, Bernard; Hofmann, Gerald; Homan, Tim; Horio, Tetsuya; Horiuchi, Hiroyuki; James, Steve; Jones, Meriel; Karaffa, Levente; Karányi, Zsolt; Kato, Masashi; Keller, Nancy; Kelly, Diane E; Kiel, Jan A K W; Kim, Jung-Mi; van der Klei, Ida J; Klis, Frans M; Kovalchuk, Andriy; Krasevec, Nada; Kubicek, Christian P; Liu, Bo; Maccabe, Andrew; Meyer, Vera; Mirabito, Pete; Miskei, Márton; Mos, Magdalena; Mullins, Jonathan; Nelson, David R; Nielsen, Jens; Oakley, Berl R; Osmani, Stephen A; Pakula, Tiina; Paszewski, Andrzej; Paulsen, Ian; Pilsyk, Sebastian; Pócsi, István; Punt, Peter J; Ram, Arthur F J; Ren, Qinghu; Robellet, Xavier; Robson, Geoff; Seiboth, Bernhard; van Solingen, Piet; Specht, Thomas; Sun, Jibin; Taheri-Talesh, Naimeh; Takeshita, Norio; Ussery, Dave; vanKuyk, Patricia A; Visser, Hans; van de Vondervoort, Peter J I; de Vries, Ronald P; Walton, Jonathan; Xiang, Xin; Xiong, Yi; Zeng, An Ping; Brandt, Bernd W; Cornell, Michael J; van den Hondel, Cees A M J J; Visser, Jacob; Oliver, Stephen G; Turner, Geoffrey

    2009-03-01

    The identification and annotation of protein-coding genes is one of the primary goals of whole-genome sequencing projects, and the accuracy of predicting the primary protein products of gene expression is vital to the interpretation of the available data and the design of downstream functional applications. Nevertheless, the comprehensive annotation of eukaryotic genomes remains a considerable challenge. Many genomes submitted to public databases, including those of major model organisms, contain significant numbers of wrong and incomplete gene predictions. We present a community-based reannotation of the Aspergillus nidulans genome with the primary goal of increasing the number and quality of protein functional assignments through the careful review of experts in the field of fungal biology.

  8. The 2008 update of the Aspergillus nidulans genome annotation: a community effort

    PubMed Central

    Wortman, Jennifer Russo; Gilsenan, Jane Mabey; Joardar, Vinita; Deegan, Jennifer; Clutterbuck, John; Andersen, Mikael R.; Archer, David; Bencina, Mojca; Braus, Gerhard; Coutinho, Pedro; von Döhren, Hans; Doonan, John; Driessen, Arnold J.M.; Durek, Pawel; Espeso, Eduardo; Fekete, Erzsébet; Flipphi, Michel; Estrada, Carlos Garcia; Geysens, Steven; Goldman, Gustavo; de Groot, Piet W.J.; Hansen, Kim; Harris, Steven D.; Heinekamp, Thorsten; Helmstaedt, Kerstin; Henrissat, Bernard; Hofmann, Gerald; Homan, Tim; Horio, Tetsuya; Horiuchi, Hiroyuki; James, Steve; Jones, Meriel; Karaffa, Levente; Karányi, Zsolt; Kato, Masashi; Keller, Nancy; Kelly, Diane E.; Kiel, Jan A.K.W.; Kim, Jung-Mi; van der Klei, Ida J.; Klis, Frans M.; Kovalchuk, Andriy; Kraševec, Nada; Kubicek, Christian P.; Liu, Bo; MacCabe, Andrew; Meyer, Vera; Mirabito, Pete; Miskei, Márton; Mos, Magdalena; Mullins, Jonathan; Nelson, David R.; Nielsen, Jens; Oakley, Berl R.; Osmani, Stephen A.; Pakula, Tiina; Paszewski, Andrzej; Paulsen, Ian; Pilsyk, Sebastian; Pócsi, István; Punt, Peter J.; Ram, Arthur F.J.; Ren, Qinghu; Robellet, Xavier; Robson, Geoff; Seiboth, Bernhard; Solingen, Piet van; Specht, Thomas; Sun, Jibin; Taheri-Talesh, Naimeh; Takeshita, Norio; Ussery, Dave; vanKuyk, Patricia A.; Visser, Hans; van de Vondervoort, Peter J.I.; de Vries, Ronald P.; Walton, Jonathan; Xiang, Xin; Xiong, Yi; Zeng, An Ping; Brandt, Bernd W.; Cornell, Michael J.; van den Hondel, Cees A.M.J.J.; Visser, Jacob; Oliver, Stephen G.; Turner, Geoffrey

    2010-01-01

    The identification and annotation of protein-coding genes is one of the primary goals of whole-genome sequencing projects, and the accuracy of predicting the primary protein products of gene expression is vital to the interpretation of the available data and the design of downstream functional applications. Nevertheless, the comprehensive annotation of eukaryotic genomes remains a considerable challenge. Many genomes submitted to public databases, including those of major model organisms, contain significant numbers of wrong and incomplete gene predictions. We present a community-based reannotation of the Aspergillus nidulans genome with the primary goal of increasing the number and quality of protein functional assignments through the careful review of experts in the field of fungal biology. PMID:19146970

  9. Fusarium sambucinum astA gene expressed during potato infection is a functional orthologue of Aspergillus nidulans astA.

    PubMed

    Piłsyk, Sebastian; Natorff, Renata; Gawińska-Urbanowicz, Hanna; Kruszewska, Joanna S

    2015-06-01

    Sulfate assimilation plays a vital role in prototrophic organisms. Orthologues of the alternative sulfate transporter (AstA) gene from Aspergillus nidulans were identified in the fungal plant pathogens Fusarium sambucinum and Fusarium graminearum. By physiological and biochemical analyses, the AstA orthologues were determined to be able to uptake sulfate from the environment. Similarly to astA in A. nidulans, the FsastA gene was found to be regulated by sulfur metabolite repression (SMR) in a sulfur-dependent manner. In contrast, the FgastA transcript was undetectable, however, when the FgastA gene was expressed heterologously in A. nidulans, the translated FgAstA protein acted as a sulfate transporter. Interestingly, F. sambucinum astA expression was remarkably augmented in infected potato tubers, despite the presence abundant sulfate and was found not to be correlated with plant resistance.

  10. Fusarium sambucinum astA gene expressed during potato infection is a functional orthologue of Aspergillus nidulans astA.

    PubMed

    Piłsyk, Sebastian; Natorff, Renata; Gawińska-Urbanowicz, Hanna; Kruszewska, Joanna S

    2015-06-01

    Sulfate assimilation plays a vital role in prototrophic organisms. Orthologues of the alternative sulfate transporter (AstA) gene from Aspergillus nidulans were identified in the fungal plant pathogens Fusarium sambucinum and Fusarium graminearum. By physiological and biochemical analyses, the AstA orthologues were determined to be able to uptake sulfate from the environment. Similarly to astA in A. nidulans, the FsastA gene was found to be regulated by sulfur metabolite repression (SMR) in a sulfur-dependent manner. In contrast, the FgastA transcript was undetectable, however, when the FgastA gene was expressed heterologously in A. nidulans, the translated FgAstA protein acted as a sulfate transporter. Interestingly, F. sambucinum astA expression was remarkably augmented in infected potato tubers, despite the presence abundant sulfate and was found not to be correlated with plant resistance. PMID:25986548

  11. Characterization of the aodA, dnmA, mnSOD and pimA genes in Aspergillus nidulans

    PubMed Central

    Leiter, Éva; Park, Hee-Soo; Kwon, Nak-Jung; Han, Kap-Hoon; Emri, Tamás; Oláh, Viktor; Mészáros, Ilona; Dienes, Beatrix; Vincze, János; Csernoch, László; Yu, Jae-Hyuk; Pócsi, István

    2016-01-01

    Mitochondria play key roles in cellular energy generation and lifespan of most eukaryotes. To understand the functions of four nuclear-encoded genes predicted to be related to the maintenance of mitochondrial morphology and function in Aspergillus nidulans, systematic characterization was carried out. The deletion and overexpression mutants of aodA, dnmA, mnSOD and pimA encoding alternative oxidase, dynamin related protein, manganese superoxide dismutase and Lon protease, respectively, were generated and examined for their growth, stress tolerances, respiration, autolysis, cell death, sterigmatocystin production, hyphal morphology and size, and mitochondrial superoxide production as well as development. Overall, genetic manipulation of these genes had less effect on cellular physiology and ageing in A. nidulans than that of their homologs in another fungus Podospora anserina with a well-characterized senescence. The observed interspecial phenotypic differences can be explained by the dissimilar intrinsic stabilities of the mitochondrial genomes in A. nidulans and P. anserina. Furthermore, the marginally altered phenotypes observed in A. nidulans mutants indicate the presence of effective compensatory mechanisms for the complex networks of mitochondrial defense and quality control. Importantly, these findings can be useful for developing novel platforms for heterologous protein production, or on new biocontrol and bioremediation technologies based on Aspergillus species. PMID:26846452

  12. Characterization of the aodA, dnmA, mnSOD and pimA genes in Aspergillus nidulans.

    PubMed

    Leiter, Éva; Park, Hee-Soo; Kwon, Nak-Jung; Han, Kap-Hoon; Emri, Tamás; Oláh, Viktor; Mészáros, Ilona; Dienes, Beatrix; Vincze, János; Csernoch, László; Yu, Jae-Hyuk; Pócsi, István

    2016-02-05

    Mitochondria play key roles in cellular energy generation and lifespan of most eukaryotes. To understand the functions of four nuclear-encoded genes predicted to be related to the maintenance of mitochondrial morphology and function in Aspergillus nidulans, systematic characterization was carried out. The deletion and overexpression mutants of aodA, dnmA, mnSOD and pimA encoding alternative oxidase, dynamin related protein, manganese superoxide dismutase and Lon protease, respectively, were generated and examined for their growth, stress tolerances, respiration, autolysis, cell death, sterigmatocystin production, hyphal morphology and size, and mitochondrial superoxide production as well as development. Overall, genetic manipulation of these genes had less effect on cellular physiology and ageing in A. nidulans than that of their homologs in another fungus Podospora anserina with a well-characterized senescence. The observed interspecial phenotypic differences can be explained by the dissimilar intrinsic stabilities of the mitochondrial genomes in A. nidulans and P. anserina. Furthermore, the marginally altered phenotypes observed in A. nidulans mutants indicate the presence of effective compensatory mechanisms for the complex networks of mitochondrial defense and quality control. Importantly, these findings can be useful for developing novel platforms for heterologous protein production, or on new biocontrol and bioremediation technologies based on Aspergillus species.

  13. Characterization and localization of the cytoplasmic dynein heavy chain in Aspergillus nidulans.

    PubMed

    Xiang, X; Roghi, C; Morris, N R

    1995-10-10

    Migration of nuclei throughout the mycelium is essential for the growth and differentiation of filamentous fungi. In Aspergillus nidulans, the nudA gene, which is involved in nuclear migration, encodes a cytoplasmic dynein heavy chain. In this paper we use antibodies to characterize the Aspergillus cytoplasmic dynein heavy chain (ACDHC) and to show that the ACDHC is concentrated at the growing tip of the fungal mycelium. We demonstrate that four temperature-sensitive mutations in the nudA gene result in a striking decrease in ACDHC protein. Cytoplasmic dynein has been implicated in nuclear division in animal cells. Because the temperature-sensitive nudA mutants are able to grow slowly with occasional nuclei found in the mycelium and are able to undergo nuclear division, we have created a deletion/disruption nudA mutation and a tightly downregulated nudA mutation. These mutants exhibit a phenotype very similar to that of the temperature-sensitive nudA mutants with respect to growth, nuclear distribution, and nuclear division. This suggests that there are redundant backup motor proteins for both nuclear migration and nuclear division.

  14. Regulation of genes encoding cellulolytic enzymes by Pal-PacC signaling in Aspergillus nidulans.

    PubMed

    Kunitake, Emi; Hagiwara, Daisuke; Miyamoto, Kentaro; Kanamaru, Kyoko; Kimura, Makoto; Kobayashi, Tetsuo

    2016-04-01

    Cellulosic biomass represents a valuable potential substitute for fossil-based fuels. As such, there is a strong need to develop efficient biotechnological processes for the enzymatic hydrolysis of cellulosic biomass via the optimization of cellulase production by fungi. Ambient pH is an important factor affecting the industrial production of cellulase. In the present study, we demonstrate that several Aspergillus nidulans genes encoding cellulolytic enzymes are regulated by Pal-PacC-mediated pH signaling, as evidenced by the decreased cellulase productivity of the palC mutant and pacC deletants of A. nidulans. The deletion of pacC was observed to result in delayed induction and decreased expression of the cellulase genes based on time course expression analysis. The genome-wide identification of PacC-regulated genes under cellobiose-induced conditions demonstrated that genes expressed in a PacC-dependent manner included 82 % of ClrB (a transcriptional activator of the cellulase genes)-regulated genes, including orthologs of various transporter and β-glucosidase genes considered to be involved in cellobiose uptake or production of stronger inducer molecules. Together with the significant overlap between ClrB- and PacC-regulated genes, the results suggest that PacC-mediated regulation of the cellulase genes involves not only direct regulation by binding to their promoter regions but also indirect regulation via modulation of the expression of genes involved in ClrB-dependent transcriptional activation. Our findings are expected to contribute to the development of more efficient industrial cellulase production methods.

  15. Regulation of the acuF gene, encoding phosphoenolpyruvate carboxykinase in the filamentous fungus Aspergillus nidulans.

    PubMed

    Hynes, Michael J; Draht, Oliver W; Davis, Meryl A

    2002-01-01

    Phosphoenolpyruvate carboxykinase (PEPCK) is a key enzyme required for gluconeogenesis when microorganisms grow on carbon sources metabolized via the tricarboxylic acid (TCA) cycle. Aspergillus nidulans acuF mutants isolated by their inability to use acetate as a carbon source specifically lack PEPCK. The acuF gene has been cloned and shown to encode a protein with high similarity to PEPCK from bacteria, plants, and fungi. The regulation of acuF expression has been studied by Northern blotting and by the construction of lacZ fusion reporters. Induction by acetate is abolished in mutants unable to metabolize acetate via the TCA cycle, and induction by amino acids metabolized via 2-oxoglutarate is lost in mutants unable to form 2-oxoglutarate. Induction by acetate and proline is not additive, consistent with a single mechanism of induction. Malate and succinate result in induction, and it is proposed that PEPCK is controlled by a novel mechanism of induction by a TCA cycle intermediate or derivative, thereby allowing gluconeogenesis to occur during growth on any carbon source metabolized via the TCA cycle. It has been shown that the facB gene, which mediates acetate induction of enzymes specifically required for acetate utilization, is not directly involved in PEPCK induction. This is in contrast to Saccharomyces cerevisiae, where Cat8p and Sip4p, homologs of FacB, regulate PEPCK as well as the expression of other genes necessary for growth on nonfermentable carbon sources in response to the carbon source present. This difference in the control of gluconeogenesis reflects the ability of A. nidulans and other filamentous fungi to use a wide variety of carbon sources in comparison with S. cerevisiae. The acuF gene was also found to be subject to activation by the CCAAT binding protein AnCF, a protein homologous to the S. cerevisiae Hap complex and the mammalian NFY complex.

  16. Regulation of the acuF Gene, Encoding Phosphoenolpyruvate Carboxykinase in the Filamentous Fungus Aspergillus nidulans

    PubMed Central

    Hynes, Michael J.; Draht, Oliver W.; Davis, Meryl A.

    2002-01-01

    Phosphoenolpyruvate carboxykinase (PEPCK) is a key enzyme required for gluconeogenesis when microorganisms grow on carbon sources metabolized via the tricarboxylic acid (TCA) cycle. Aspergillus nidulans acuF mutants isolated by their inability to use acetate as a carbon source specifically lack PEPCK. The acuF gene has been cloned and shown to encode a protein with high similarity to PEPCK from bacteria, plants, and fungi. The regulation of acuF expression has been studied by Northern blotting and by the construction of lacZ fusion reporters. Induction by acetate is abolished in mutants unable to metabolize acetate via the TCA cycle, and induction by amino acids metabolized via 2-oxoglutarate is lost in mutants unable to form 2-oxoglutarate. Induction by acetate and proline is not additive, consistent with a single mechanism of induction. Malate and succinate result in induction, and it is proposed that PEPCK is controlled by a novel mechanism of induction by a TCA cycle intermediate or derivative, thereby allowing gluconeogenesis to occur during growth on any carbon source metabolized via the TCA cycle. It has been shown that the facB gene, which mediates acetate induction of enzymes specifically required for acetate utilization, is not directly involved in PEPCK induction. This is in contrast to Saccharomyces cerevisiae, where Cat8p and Sip4p, homologs of FacB, regulate PEPCK as well as the expression of other genes necessary for growth on nonfermentable carbon sources in response to the carbon source present. This difference in the control of gluconeogenesis reflects the ability of A. nidulans and other filamentous fungi to use a wide variety of carbon sources in comparison with S. cerevisiae. The acuF gene was also found to be subject to activation by the CCAAT binding protein AnCF, a protein homologous to the S. cerevisiae Hap complex and the mammalian NFY complex. PMID:11741859

  17. Expression of Aspergillus nidulans phy Gene in Nicotiana benthamiana Produces Active Phytase with Broad Specificities

    PubMed Central

    Oh, Tae-Kyun; Oh, Sung; Kim, Seongdae; Park, Jae Sung; Vinod, Nagarajan; Jang, Kyung Min; Kim, Sei Chang; Choi, Chang Won; Ko, Suk-Min; Jeong, Dong Kee; Udayakumar, Rajangam

    2014-01-01

    A full-length phytase gene (phy) of Aspergillus nidulans was amplified from the cDNA library by polymerase chain reaction (PCR), and it was introduced into a bacterial expression vector, pET-28a. The recombinant protein (rPhy-E, 56 kDa) was overexpressed in the insoluble fraction of Escherichia coli culture, purified by Ni-NTA resin under denaturing conditions and injected into rats as an immunogen. To express A. nidulans phytase in a plant, the full-length of phy was cloned into a plant expression binary vector, pPZP212. The resultant construct was tested for its transient expression by Agrobacterium-infiltration into Nicotiana benthamiana leaves. Compared with a control, the agro-infiltrated leaf tissues showed the presence of phy mRNA and its high expression level in N. benthamiana. The recombinant phytase (rPhy-P, 62 kDa) was strongly reacted with the polyclonal antibody against the nonglycosylated rPhy-E. The rPhy-P showed glycosylation, two pH optima (pH 4.5 and pH 5.5), an optimum temperature at 45~55 °C, thermostability and broad substrate specificities. After deglycosylation by peptide-N-glycosidase F (PNGase-F), the rPhy-P significantly lost the phytase activity and retained 1/9 of the original activity after 10 min of incubation at 45 °C. Therefore, the deglycosylation caused a significant reduction in enzyme thermostability. In animal experiments, oral administration of the rPhy-P at 1500 U/kg body weight/day for seven days caused a significant reduction of phosphorus excretion by 16% in rat feces. Besides, the rPhy-P did not result in any toxicological changes and clinical signs. PMID:25192284

  18. NsdD is a key repressor of asexual development in Aspergillus nidulans.

    PubMed

    Lee, Mi-Kyung; Kwon, Nak-Jung; Choi, Jae Min; Lee, Im-Soon; Jung, Seunho; Yu, Jae-Hyuk

    2014-05-01

    Asexual development (conidiation) of the filamentous fungus Aspergillus nidulans occurs via balanced activities of multiple positive and negative regulators. For instance, FluG (+) and SfgA (-) govern upstream regulation of the developmental switch, and BrlA (+) and VosA (-) control the progression and completion of conidiation. To identify negative regulators of conidiation downstream of FluG-SfgA, we carried out multicopy genetic screens using sfgA deletion strains. After visually screening >100,000 colonies, we isolated 61 transformants exhibiting reduced conidiation. Responsible genes were identified as AN3152 (nsdD), AN7507, AN2009, AN1652, AN5833, and AN9141. Importantly, nsdD, a key activator of sexual reproduction, was present in 10 independent transformants. Furthermore, deletion, overexpression, and double-mutant analyses of individual genes have led to the conclusion that, of the six genes, only nsdD functions in the FluG-activated conidiation pathway. The deletion of nsdD bypassed the need for fluG and flbA∼flbE, but not brlA or abaA, in conidiation, and partially restored production of the mycotoxin sterigmatocystin (ST) in the ΔfluG, ΔflbA, and ΔflbB mutants, suggesting that NsdD is positioned between FLBs and BrlA in A. nidulans. Nullifying nsdD caused formation of conidiophores in liquid submerged cultures, where wild-type strains do not develop. Moreover, the removal of both nsdD and vosA resulted in even more abundant development of conidiophores in liquid submerged cultures and high-level accumulation of brlA messenger (m)RNA even at 16 hr of vegetative growth. Collectively, NsdD is a key negative regulator of conidiation and likely exerts its repressive role via downregulating brlA.

  19. Functional Characterization of Aspergillus nidulans ypkA, a Homologue of the Mammalian Kinase SGK

    PubMed Central

    Colabardini, Ana Cristina; Brown, Neil Andrew; Savoldi, Marcela; Goldman, Maria Helena S.; Goldman, Gustavo Henrique

    2013-01-01

    The serum- and glucocorticoid-regulated protein kinase (SGK) is an AGC kinase involved in signal cascades regulated by glucocorticoid hormones and serum in mammals. The Saccharomyces cerevisiae ypk1 and ypk2 genes were identified as SGK homologues and Ypk1 was shown to regulate the balance of sphingolipids between the inner and outer plasma membrane. This investigation characterized the Aspergillus nidulans YPK1 homologue, YpkA, representing the first filamentous fungal YPK1 homologue. Two conditional mutant strains were constructed by replacing the endogenous ypk1 promoter with two different regulatable promoters, alcA (from the alcohol dehydrogenase gene) and niiA (from the nitrate reductase gene). Both constructs confirmed that ypkA was an essential gene in A. nidulans. Repression of ypkA caused decreased radial growth, a delay in conidial germination, deficient polar axis establishment, intense branching during late stages of growth, a lack of asexual spores, and a terminal phenotype. Membrane lipid polarization, endocytosis, eisosomes and vacuolar distribution were also affected by ypkA repression, suggesting that YpkA plays a role in hyphal morphogenesis via coordinating the delivery of cell membrane and wall constituents to the hyphal apex. The A. nidulans Pkh1 homologue pkhA was also shown to be an essential gene, and preliminary genetic analysis suggested that the ypkA gene is not directly downstream of pkhA or epistatic to pkhA, rather, ypkA and pkhA are genetically independent or in parallel. BarA is a homologue of the yeast Lag1 acyl-CoA-dependent ceramide synthase, which catalyzes the condensation of phytosphingosine with a fatty acyl-CoA to form phytoceramide. When barA was absent, ypkA repression was lethal to the cell. Therefore, there appears to be a genetic interaction between ypkA, barA, and the sphingolipid synthesis. Transcriptional profiling of ypkA overexpression and down-regulation revealed several putative YpkA targets associated with the

  20. Functional characterization of Aspergillus nidulans ypkA, a homologue of the mammalian kinase SGK.

    PubMed

    Colabardini, Ana Cristina; Brown, Neil Andrew; Savoldi, Marcela; Goldman, Maria Helena S; Goldman, Gustavo Henrique

    2013-01-01

    The serum- and glucocorticoid-regulated protein kinase (SGK) is an AGC kinase involved in signal cascades regulated by glucocorticoid hormones and serum in mammals. The Saccharomyces cerevisiae ypk1 and ypk2 genes were identified as SGK homologues and Ypk1 was shown to regulate the balance of sphingolipids between the inner and outer plasma membrane. This investigation characterized the Aspergillus nidulans YPK1 homologue, YpkA, representing the first filamentous fungal YPK1 homologue. Two conditional mutant strains were constructed by replacing the endogenous ypk1 promoter with two different regulatable promoters, alcA (from the alcohol dehydrogenase gene) and niiA (from the nitrate reductase gene). Both constructs confirmed that ypkA was an essential gene in A. nidulans. Repression of ypkA caused decreased radial growth, a delay in conidial germination, deficient polar axis establishment, intense branching during late stages of growth, a lack of asexual spores, and a terminal phenotype. Membrane lipid polarization, endocytosis, eisosomes and vacuolar distribution were also affected by ypkA repression, suggesting that YpkA plays a role in hyphal morphogenesis via coordinating the delivery of cell membrane and wall constituents to the hyphal apex. The A. nidulans Pkh1 homologue pkhA was also shown to be an essential gene, and preliminary genetic analysis suggested that the ypkA gene is not directly downstream of pkhA or epistatic to pkhA, rather, ypkA and pkhA are genetically independent or in parallel. BarA is a homologue of the yeast Lag1 acyl-CoA-dependent ceramide synthase, which catalyzes the condensation of phytosphingosine with a fatty acyl-CoA to form phytoceramide. When barA was absent, ypkA repression was lethal to the cell. Therefore, there appears to be a genetic interaction between ypkA, barA, and the sphingolipid synthesis. Transcriptional profiling of ypkA overexpression and down-regulation revealed several putative YpkA targets associated with the

  1. Combined Expression of Aspergillus nidulans Endoxylanase X24 and Aspergillus oryzae (alpha)-Amylase in Industrial Baker's Yeasts and Their Use in Bread Making

    PubMed Central

    Monfort, A.; Blasco, A.; Prieto, J. A.; Sanz, P.

    1996-01-01

    The Aspergillus nidulans endoxylanase X24 and the Aspergillus oryzae (alpha)-amylase cDNAs were placed under the control of the Saccharomyces cerevisiae actin promoter (pACT1) and introduced into baker's yeast. Bread made with transformants expressing both enzymes (YEpACT-AMY-ACT-X24) showed a 30% increase in volume and reduced firmness in comparison with that produced with a commercial strain. Endoxylanase X24 and (alpha)-amylase seem to act synergistically to improve the quality of bread in terms of volume and density. PMID:16535419

  2. The Aspergillus nidulans ATM Kinase Regulates Mitochondrial Function, Glucose Uptake and the Carbon Starvation Response

    PubMed Central

    Krohn, Nadia Graciele; Brown, Neil Andrew; Colabardini, Ana Cristina; Reis, Thaila; Savoldi, Marcela; Dinamarco, Taísa Magnani; Goldman, Maria Helena S.; Goldman, Gustavo Henrique

    2013-01-01

    Mitochondria supply cellular energy and also perform a role in the adaptation to metabolic stress. In mammals, the ataxia-telangiectasia mutated (ATM) kinase acts as a redox sensor controlling mitochondrial function. Subsequently, transcriptomic and genetic studies were utilized to elucidate the role played by a fungal ATM homolog during carbon starvation. In Aspergillus nidulans, AtmA was shown to control mitochondrial function and glucose uptake. Carbon starvation responses that are regulated by target of rapamycin (TOR) were shown to be AtmA-dependent, including autophagy and hydrolytic enzyme secretion. AtmA also regulated a p53-like transcription factor, XprG, inhibiting starvation-induced XprG-dependent protease secretion and cell death. Thus, AtmA possibly represents a direct or indirect link between mitochondrial stress, metabolism, and growth through the influence of TOR and XprG function. The coordination of cell growth and division with nutrient availability is crucial for all microorganisms to successfully proliferate in a heterogeneous environment. Mitochondria supply cellular energy but also perform a role in the adaptation to metabolic stress and the cross-talk between prosurvival and prodeath pathways. The present study of Aspergillus nidulans demonstrated that AtmA also controlled mitochondrial mass, function, and oxidative phosphorylation, which directly or indirectly influenced glucose uptake. Carbon starvation responses, including autophagy, shifting metabolism to the glyoxylate cycle, and the secretion of carbon scavenging enzymes were AtmA-dependent. Transcriptomic profiling of the carbon starvation response demonstrated how TOR signaling and the retrograde response, which signals mitochondrial dysfunction, were directly or indirectly influenced by AtmA. The AtmA kinase was also shown to influence a p53-like transcription factor, inhibiting starvation-induced XprG-dependent protease secretion and cell death. Therefore, in response to metabolic

  3. Accumulation of stress and inducer-dependent plant-cell-wall-degrading enzymes during asexual development in Aspergillus nidulans.

    PubMed Central

    Prade, R A; Ayoubi, P; Krishnan, S; Macwana, S; Russell, H

    2001-01-01

    Determination and interpretation of fungal gene expression profiles based on digital reconstruction of expressed sequenced tags (ESTs) are reported. A total of 51,524 DNA sequence files processed with PipeOnline resulted in 9775 single and 5660 contig unique ESTs, 31.2% of a typical fungal transcriptome. Half of the unique ESTs shared homology with genes in public databases, 35.8% of which are functionally defined and 64.2% are unclear or unknown. In Aspergillus nidulans 86% of transcripts associate with intermediate metabolism functions, mainly related to carbohydrate, amino acid, protein, and peptide biosynthesis. During asexual development, A. nidulans unexpectedly accumulates stress response and inducer-dependent transcripts in the absence of an inducer. Stress response genes in A. nidulans ESTs total 1039 transcripts, contrasting with 117 in Neurospora crassa, a 14.3-fold difference. A total of 5.6% of A. nidulans ESTs implicate inducer-dependent cell wall degradation or amino acid acquisition, 3.5-fold higher than in N. crassa. Accumulation of stress response and inducer-dependent transcripts suggests general derepression of cis-regulation during terminal asexual development. PMID:11238386

  4. Expression of Drosophila melanogaster xanthine dehydrogenase in Aspergillus nidulans and some properties of the recombinant enzyme.

    PubMed Central

    Adams, Benjamin; Lowe, David J; Smith, Andrew T; Scazzocchio, Claudio; Demais, Stephane; Bray, Robert C

    2002-01-01

    Recent crystal structures of xanthine dehydrogenase, xanthine oxidase and related enzymes have paved the way for a detailed structural and functional analysis of these enzymes. One problem encountered when working with these proteins, especially with recombinant protein, is that the preparations tend to be heterogeneous, with only a fraction of the enzyme molecules being active. This is due to the incompleteness of post-translational modification, which for this protein is a complex, and incompletely understood, process involving incorporation of the Mo and Fe/S centres. The enzyme has been expressed previously in both Drosophila and insect cells using baculovirus. The insect cell system has been exploited by Iwasaki et al. [Iwasaki, Okamoto, Nishino, Mizushima and Hori (2000) J. Biochem (Tokyo) 127, 771-778], but, for the rat enzyme, yields a complex mixture of enzyme forms, containing around 10% of functional enzyme. The expression of Drosophila melanogaster xanthine dehydrogenase in Aspergillus nidulans is described. The purified protein has been analysed both functionally and spectroscopically. Its specific activity is indistinguishable from that of the enzyme purified from fruit flies [Doyle, Burke, Chovnick, Dutton, Whittle and Bray (1996) Eur. J. Biochem. 239, 782-795], and it appears to be more active than recombinant xanthine dehydrogenase produced with the baculovirus system. EPR spectra of the recombinant Drosophila enzyme are reported, including parameters for the Fe/S centres. Only a very weak "Fe/SIII" signal (g(1,2,3), 2.057, 1.930, 1.858) was observed, in contrast to the strong analogous signal reported for the enzyme from baculovirus. Since this signal appears to be associated with incomplete post-translational modification, this is consistent with relatively more complete cofactor incorporation in the Aspergillus-produced enzyme. Thus we have developed a recombinant expression system for D. melanogaster xanthine dehydrogenase, which can be used

  5. Studies of the production of fungal polyketides in Aspergillus nidulans by using systems biology tools.

    PubMed

    Panagiotou, Gianni; Andersen, Mikael R; Grotkjaer, Thomas; Regueira, Torsten B; Nielsen, Jens; Olsson, Lisbeth

    2009-04-01

    Many filamentous fungi produce polyketide molecules with great significance as human pharmaceuticals; these molecules include the cholesterol-lowering compound lovastatin, which was originally isolated from Aspergillus terreus. The chemical diversity and potential uses of these compounds are virtually unlimited, and it is thus of great interest to develop a well-described microbial production platform for polyketides. Using genetic engineering tools available for the model organism Aspergillus nidulans, we constructed two recombinant strains, one expressing the Penicillium griseofulvum 6-methylsalicylic acid (6-MSA) synthase gene and one expressing the 6-MSA synthase gene and overexpressing the native xylulose-5-phosphate phosphoketolase gene (xpkA) for increasing the pool of polyketide precursor levels. The physiology of the recombinant strains and that of a reference wild-type strain were characterized on glucose, xylose, glycerol, and ethanol media in controlled bioreactors. Glucose was found to be the preferred carbon source for 6-MSA production, and 6-MSA concentrations up to 455 mg/liter were obtained for the recombinant strain harboring the 6-MSA gene. Our findings indicate that overexpression of xpkA does not directly improve 6-MSA production on glucose, but it is possible, if the metabolic flux through the lower part of glycolysis is reduced, to obtain quite high yields for conversion of sugar to 6-MSA. Systems biology tools were employed for in-depth analysis of the metabolic processes. Transcriptome analysis of 6-MSA-producing strains grown on glucose and xylose in the presence and absence of xpkA overexpression, combined with flux and physiology data, enabled us to propose an xpkA-msaS interaction model describing the competition between biomass formation and 6-MSA production for the available acetyl coenzyme A.

  6. Characteristics of physiological inducers of the ethanol utilization (alc) pathway in Aspergillus nidulans.

    PubMed Central

    Flipphi, Michel; Kocialkowska, Janina; Felenbok, Béatrice

    2002-01-01

    The ethanol utilization (alc) pathway in Aspergillus nidulans is one of the strongest expressed gene systems in filamentous fungi. The pathway-specific activator AlcR requires the presence of an inducing compound to activate transcription of genes under its control. We have demonstrated recently that acetaldehyde is the sole physiological inducer of ethanol catabolism. In the present study we show that compounds with catabolism related to that of ethanol, i.e. primary alcohols, primary monoamines and l-threonine, act as inducers because their breakdown results in the production of inducing aliphatic aldehydes. Such aldehydes were shown to induce the alc genes efficiently at low external concentrations. When ethanol is mixed with representatives of another class of strong direct inducers, ketones, the physiological inducer, acetaldehyde, prevails as effector. Although direct inducers essentially carry a carbonyl function, not all aldehydes and ketones act as inducers. Structural features discriminating non-inducing from inducing compounds concern: (i) the length of the aliphatic side group(s); (ii) the presence and nature of any non-aliphatic substituent. These characteristics enable us to predict whether or not a given carbonyl compound will induce the alc genes. PMID:11988072

  7. Mutations affecting extracellular protease production in the filamentous fungus Aspergillus nidulans.

    PubMed

    Katz, M E; Flynn, P K; vanKuyk, P A; Cheetham, B F

    1996-04-10

    The extracellular proteases of Aspergillus nidulans are known to be regulated by carbon, nitrogen and sulphur metabolite repression. In this study, a mutant with reduced levels of extracellular protease was isolated by screening for loss of halo production on milk plates. Genetic analysis of the mutant showed that it contains a single, recessive mutation, in a gene which we have designated xprE, located on chromosome VI. The xprE1 mutation affected the production of extracellular proteases in response to carbon, nitrogen and, to a lesser extent, sulphur limitation. Three reversion mutations, xprF1, xprF2 and xprG1, which suppress xprE1, were characterised. Both xprF and xprG map to chromosome VII but the two genes are unlinked. The xprF1, xprF2 and xprG1 mutants showed high levels of milk-clearing activity on medium containing milk as a carbon source but reduced growth on a number of nitrogen sources. Evidence is presented that the xprE1 and xprG1 mutations alter expression of more than one protease and affect levels of alkaline protease gene mRNA.

  8. FacB, the Aspergillus nidulans activator of acetate utilization genes, binds dissimilar DNA sequences.

    PubMed Central

    Todd, R B; Andrianopoulos, A; Davis, M A; Hynes, M J

    1998-01-01

    The facB gene is required for acetate induction of acetamidase (amdS) and the acetate utilization enzymes acetyl-CoA synthase (facA), isocitrate lyase (acuD) and malate synthase (acuE) in Aspergillus nidulans. The facB gene encodes a transcriptional activator with a GAL4-type Zn(II)2Cys6 zinc binuclear cluster DNA-binding domain which is shown to be required for DNA binding. In vitro DNA-binding sites for FacB in the 5' regions of the amdS, facA, acuD and acuE genes have been identified. Mutations in amdS FacB DNA-binding sites affected expression of an amdS-lacZ reporter in vivo and altered the affinity of in vitro DNA binding. This study shows that the FacB Zn(II)2Cys6 cluster binds to dissimilar sites which show similarity in form but not sequence with DNA-binding sites of other Zn(II)2Cys6 proteins. Sequences with homology to FacB sites are found in the 5' regions of genes regulated by the closely related yeast Zn(II)2Cys6 protein CAT8. PMID:9524126

  9. The phosphoproteome of Aspergillus nidulans reveals functional association with cellular processes involved in morphology and secretion.

    PubMed

    Ramsubramaniam, Nikhil; Harris, Steven D; Marten, Mark R

    2014-11-01

    We describe the first phosphoproteome of the model filamentous fungus Aspergillus nidulans. Phosphopeptides were enriched using titanium dioxide, separated using a convenient ultra-long reverse phase gradient, and identified using a "high-high" strategy (high mass accuracy on the parent and fragment ions) with higher-energy collisional dissociation. Using this approach 1801 phosphosites, from 1637 unique phosphopeptides, were identified. Functional classification revealed phosphoproteins were overrepresented under GO categories related to fungal morphogenesis: "sites of polar growth," "vesicle mediated transport," and "cytoskeleton organization." In these same GO categories, kinase-substrate analysis of phosphoproteins revealed the majority were target substrates of CDK and CK2 kinase families, indicating these kinase families play a prominent role in fungal morphogenesis. Kinase-substrate analysis also identified 57 substrates for kinases known to regulate secretion of hydrolytic enzymes (e.g. PkaA, SchA, and An-Snf1). Altogether this data will serve as a benchmark that can be used to elucidate regulatory networks functionally associated with fungal morphogenesis and secretion. All MS data have been deposited in the ProteomeXchange with identifier PXD000715 (http://proteomecentral.proteomexchange.org/dataset/PXD000715).

  10. Mutations in sfdA and sfdB suppress multiple developmental mutations in Aspergillus nidulans.

    PubMed Central

    Kellner, Ellen M; Adams, Thomas H

    2002-01-01

    Conidiophore morphogenesis in Aspergillus nidulans occurs in response to developmental signals that result in the activation of brlA, a well-characterized gene that encodes a transcription factor that is central to asexual development. Loss-of-function mutations in flbD and other fluffy loci have previously been shown to result in delayed development and reduced expression of brlA. flbD message is detectable during both hyphal growth and conidiation, and its gene product is similar to the Myb family of transcription factors. To further understand the regulatory pathway to brlA activation and conidiation, we isolated suppressor mutations that rescued development in strains with a flbD null allele. We describe here two new loci, designated sfdA and sfdB for suppressors of flbD, that bypass the requirement of flbD for development. sfd mutant alleles were found to restore developmental timing and brlA expression to strains with flbD deletions. In addition, sfd mutations suppress the developmental defects in strains harboring loss-of-function mutations in fluG, flbA, flbB, flbC, and flbE. All alleles of sfdA and sfdB that we have isolated are recessive to their wild-type alleles in diploids. Strains with mutant sfd alleles in otherwise developmentally wild-type backgrounds have reduced growth phenotypes and develop conidiophores in submerged cultures. PMID:11805053

  11. Isolation of a gene required for programmed initiation of development by Aspergillus nidulans.

    PubMed Central

    Adams, T H; Hide, W A; Yager, L N; Lee, B N

    1992-01-01

    In contrast to many other cases in microbial development, Aspergillus nidulans conidiophore production initiates primarily as a programmed part of the life cycle rather than as a response to nutrient deprivation. Mutations in the acoD locus result in "fluffy" colonies that appear to grow faster than the wild type and proliferate as undifferentiated masses of vegetative cells. We show that unlike wild-type strains, acoD deletion mutants are unable to make conidiophores under optimal growth conditions but can be induced to conidiate when growth is nutritionally limited. The requirement for acoD in conidiophore development occurs prior to activation of brlA, a primary regulator of development. The acoD transcript is present both in vegetative hyphae prior to developmental induction and in developing cultures. However, the effects of acoD mutations are detectable only after developmental induction. We propose that acoD activity is primarily controlled at the posttranscriptional level and that it is required to direct developmentally specific changes that bring about growth inhibition and activation of brlA expression to result in conidiophore development. Images PMID:1508186

  12. Analysis of fluG mutations that affect light-dependent conidiation in Aspergillus nidulans.

    PubMed Central

    Yager, L N; Lee, H O; Nagle, D L; Zimmerman, J E

    1998-01-01

    Conidiation in Aspergillus nidulans is induced by exposure to red light but can also be induced by blue light in certain mutant strains. We have isolated a mutation in the fluG gene that abolishes responsiveness to red light but does not affect the response to blue light. It has been shown that the veA1 (velvet) mutation allows conidiation to occur in the absence of light. We have identified three other fluG mutations that suppress the veA1 phenotype; these double mutants do not conidiate in the dark. The mutations described here define two new phenotypic classes of fluG alleles that display abnormal responses to light. We have characterized these mutations with respect to their molecular identity and to their effect on fluG transcription. Although it has been shown that fluG is required for the synthesis of an extracellular factor that directs conidiation, we do not detect this factor under conditions that promote conidiation in the veA1 suppressors. Furthermore, extracellular rescue is not observed in fluG deletion strains containing the wild-type veA allele. We propose that a genetic interaction between fluG and veA influences the production of the extracellular signal and regulates the initiation of conidiation. PMID:9691036

  13. Protein expression and subcellular localization of the general purine transporter UapC from Aspergillus nidulans.

    PubMed

    Valdez-Taubas, J; Diallinas, G; Scazzocchio, C; Rosa, A L

    2000-07-01

    The uapC gene of Aspergillus nidulans belongs to a family of nucleobase-specific transporters conserved in prokaryotic and eucaryotic organisms. We report the use of immunological and green fluorescent protein based strategies to study protein expression and subcellular distribution of UapC. A chimeric protein containing a plant-adapted green fluorescent protein (sGFP) fused to the C-terminus of UapC was shown to be functional in vivo, as it complements a triple mutant (i.e., uapC(-) uapA(-) azgA(-)) unable to grow on uric acid as the sole nitrogen source. UapC-GFP is located in the plasma membrane and, secondarily, in internal structures observed as fluorescent dots. A strong correlation was found between cellular levels of UapC-GFP fluorescence and known patterns of uapC gene expression. This work represents the first in vivo study of protein expression and subcellular localization of a filamentous fungal nucleobase transporter.

  14. Organization and Dynamics of the Aspergillus nidulans Golgi during Apical Extension and Mitosis

    PubMed Central

    Pantazopoulou, Areti

    2009-01-01

    Aspergillus nidulans hyphae grow exclusively by apical extension. Golgi equivalents (GEs) labeled with mRFP-tagged PHOSBP domain form a markedly polarized, dynamic network of ring-shaped and fenestrated cisternae that remains intact during “closed” mitosis. mRFP-PHOSBP GEs advance associated with the growing apex where secretion predominates but do not undergo long-distance movement toward the tip that could account for their polarization. mRFP-PHOSBP GEs overlap with the trans-Golgi resident Sec7 but do not colocalize with also polarized accretions of the early Golgi marker GrhAGrh1-GFP, indicating that early and late Golgi membranes segregate spatially. AnSec23-GFP ER exit sites (ERES) are numerous, relatively static foci localizing across the entire cell. However, their density is greatest near the tip, correlating with predominance of early and trans-Golgi elements in this region. Whereas GrhA-GFP structures and ERES reach the apical dome, mRFP-PHOSBP GEs are excluded from this region, which contains the endosome dynein loading zone. After latrunculin-mediated F-actin disruption, mRFP-PHOSBP GEs fragment and, like AnSec23-GFP ERES, depolarize. Brefeldin A transiently collapses late and early GEs into distinct aggregates containing Sec7/mRFP-PHOSBP and GrhA-GFP, respectively, temporarily arresting apical extension. Rapid growth reinitiates after washout, correlating with reacquisition of the normal Golgi organization that, we conclude, is required for apical extension. PMID:19692566

  15. The effect of CreA in glucose and xylose catabolism in Aspergillus nidulans.

    PubMed

    Prathumpai, W; McIntyre, M; Nielsen, J

    2004-02-01

    The catabolism of glucose and xylose was studied in a wild type and creA deleted (carbon catabolite de-repressed) strain of Aspergillus nidulans. Both strains were cultivated in bioreactors with either glucose or xylose as the sole carbon source, or in the presence of both sugars. In the cultivations on single carbon sources, it was demonstrated that xylose acted as a carbon catabolite repressor (xylose cultivations), while the enzymes in the xylose utilisation pathway were also subject to repression in the presence of glucose (glucose cultivations). In the wild type strain growing on the sugar mixture, glucose repression of xylose utilisation was observed; with xylose utilisation occurring only after glucose was depleted. This phenomenon was not seen in the creA deleted strain, where glucose and xylose were catabolised simultaneously. Measurement of key metabolites and the activities of key enzymes in the xylose utilisation pathway revealed that xylose metabolism was occurring in the creA deleted strain, even at high glucose concentrations. Conversely, in the wild type strain, activities of the key enzymes for xylose metabolism increased only when the effects of glucose repression had been relieved. Xylose was both a repressor and an inducer of xylanases at the same time. The creA mutation seemed to have pleiotropic effects on carbohydratases and carbon catabolism.

  16. An Alpha Tubulin Mutation Suppresses Nuclear Migration Mutations in Aspergillus Nidulans

    PubMed Central

    Willins, D. A.; Xiang, X.; Morris, N. R.

    1995-01-01

    Microtubules and cytoplasmic dynein, a microtubule-dependent motor, are required for nuclei to move along the hyphae of filamentous fungi. Nuclear migration in Aspergillus nidulans is blocked by heat-sensitive (hs(-)) mutations in the nudA gene, which encodes dynein heavy chain, and the nudF gene, which encodes a G protein β-subunit-like protein. Hs(-) mutations in the nudC and nudG genes also prevent nuclear migration. We have isolated extragenic suppressor mutations that reverse the hs(-) phenotypes caused by these mutations. Here we show that one nudF suppressor also suppresses hs(-) mutations in nudA, nudC, and nudG and deletions in nudA and nudF. This suppressor mutation is in the tubA alpha tubulin gene, and its characteristics suggest that it destabilizes microtubules. The mutation alters microtubule staining and confers sensitivity to cold and benomyl, two treatments that destabilize microtubules. Treatment with low concentrations of benomyl also suppresses the hs(-) nudA, nudC, nudF, and nudG mutations and the nudA and nudF deletions. Suppression of the hs(-) nudA mutation and the nudA deletion is especially interesting because these strains lack active dynein heavy chain. Together, these results suggest that microtubule destabilization allows nuclei to migrate even in the absence of cytoplasmic dynein motor function. PMID:8601474

  17. Cloning and expression of hemicellulases from Aspergillus nidulans in Pichia pastoris.

    PubMed

    Vasu, Prasanna; Bauer, Stefan; Savary, Brett J

    2012-01-01

    The methylotrophic yeast Pichia pastoris is increasingly used for heterologous expression of high quality proteins in laboratory-scale (milligram) quantities. Commercially available polysaccharide-active enzyme preparations have limited applications in plant cell wall research due to their heterogeneous mix of hydrolytic activities. P. pastoris provides an ideal in vitro expression system for producing monocomponent enzymes, since it lacks endogenous plant cell wall-active enzymes and can perform eukaryotic post-translational modifications (i.e., glycosylation). We have routinely prepared cDNA constructs from Aspergillus nidulans encoding a broad array of hydrolases active on various linkages contained in plant cell wall polysaccharides. The cDNAs were inserted into the pPICZα C shuttle vector (Invitrogen) in-frame with the Saccharomyces cerevisiae α-secretion factor and expressed under the transcriptional control of the highly inducible alcohol oxidase 1 (AOX1) promoter. The enzyme products were efficiently secreted into buffered complex methanol medium (BMMY) as C-terminal his-tagged proteins for simple one-step affinity purification. The insertion of the c-Myc epitope enabled easy immunodetection. Here we present the detailed protocols for primer design, cloning, expression, and activity assays for a representative set of xylan-acting hemicellulases produced in P. pastoris.

  18. Dynamics of Actin Cables in Polarized Growth of the Filamentous Fungus Aspergillus nidulans

    PubMed Central

    Bergs, Anna; Ishitsuka, Yuji; Evangelinos, Minoas; Nienhaus, G. U.; Takeshita, Norio

    2016-01-01

    Highly polarized growth of filamentous fungi requires a continuous supply of proteins and lipids to the hyphal tip. This transport is managed by vesicle trafficking via the actin and microtubule cytoskeletons and their associated motor proteins. Particularly, actin cables originating from the hyphal tip are essential for hyphal growth. Although, specific marker proteins have been developed to visualize actin cables in filamentous fungi, the exact organization and dynamics of actin cables has remained elusive. Here, we observed actin cables using tropomyosin (TpmA) and Lifeact fused to fluorescent proteins in living Aspergillus nidulans hyphae and studied the dynamics and regulation. GFP tagged TpmA visualized dynamic actin cables formed from the hyphal tip with cycles of elongation and shrinkage. The elongation and shrinkage rates of actin cables were similar and approximately 0.6 μm/s. Comparison of actin markers revealed that high concentrations of Lifeact reduced actin dynamics. Simultaneous visualization of actin cables and microtubules suggests temporally and spatially coordinated polymerization and depolymerization between the two cytoskeletons. Our results provide new insights into the molecular mechanism of ordered polarized growth regulated by actin cables and microtubules. PMID:27242709

  19. Potential Link between the NIMA Mitotic Kinase and Nuclear Membrane Fission during Mitotic Exit in Aspergillus nidulans

    PubMed Central

    Davies, Jonathan R.; Osmani, Aysha H.; De Souza, Colin P. C.; Bachewich, Catherine; Osmani, Stephen A.

    2004-01-01

    We have isolated TINC as a NIMA-interacting protein by using the yeast two-hybrid system and have confirmed that TINC interacts with NIMA in Aspergillus nidulans. The TINC-NIMA interaction is stabilized in the absence of phosphatase inhibitors and in the presence of kinase-inactive NIMA, suggesting that the interaction is enhanced when NIMA is not fully activated. TINC is a cytoplasmic protein. TINC homologues and a TINC-like protein (A. nidulans HETC) are conserved in other filamentous fungi. Neither deletion of tinC nor deletion of both tinC and A. nidulans hetC is lethal, but deletion of tinC does produce cold sensitivity as well as osmotic sensitivity. Expression of an amino-terminal-truncated form of TINC (ΔN-TINC) inhibits colony growth in Aspergillus and localizes to membrane-like structures within the cell. Examination of cell cycle progression in these cells reveals that they progress through multiple defective mitoses. Many cells contain large polyploid single nuclei, while some appear to have separated masses of DNA. Examination of the nuclear envelopes of cells containing more than one DNA mass reveals that both DNA masses are contained within a single nuclear envelope, indicating that nuclear membrane fission is defective. The ability of these cells to separate DNA segregation from nuclear membrane fission suggests that this coordination is normally a regulated process in A. nidulans. Additional experiments demonstrate that expression of ΔN-TINC results in premature NIMA disappearance in mitotic samples. We propose that TINC's interaction with NIMA and the cell cycle defects produced by ΔN-TINC expression suggest possible roles for TINC and NIMA during nuclear membrane fission. PMID:15590818

  20. Potential link between the NIMA mitotic kinase and nuclear membrane fission during mitotic exit in Aspergillus nidulans.

    PubMed

    Davies, Jonathan R; Osmani, Aysha H; De Souza, Colin P C; Bachewich, Catherine; Osmani, Stephen A

    2004-12-01

    We have isolated TINC as a NIMA-interacting protein by using the yeast two-hybrid system and have confirmed that TINC interacts with NIMA in Aspergillus nidulans. The TINC-NIMA interaction is stabilized in the absence of phosphatase inhibitors and in the presence of kinase-inactive NIMA, suggesting that the interaction is enhanced when NIMA is not fully activated. TINC is a cytoplasmic protein. TINC homologues and a TINC-like protein (A. nidulans HETC) are conserved in other filamentous fungi. Neither deletion of tinC nor deletion of both tinC and A. nidulans hetC is lethal, but deletion of tinC does produce cold sensitivity as well as osmotic sensitivity. Expression of an amino-terminal-truncated form of TINC (DeltaN-TINC) inhibits colony growth in Aspergillus and localizes to membrane-like structures within the cell. Examination of cell cycle progression in these cells reveals that they progress through multiple defective mitoses. Many cells contain large polyploid single nuclei, while some appear to have separated masses of DNA. Examination of the nuclear envelopes of cells containing more than one DNA mass reveals that both DNA masses are contained within a single nuclear envelope, indicating that nuclear membrane fission is defective. The ability of these cells to separate DNA segregation from nuclear membrane fission suggests that this coordination is normally a regulated process in A. nidulans. Additional experiments demonstrate that expression of DeltaN-TINC results in premature NIMA disappearance in mitotic samples. We propose that TINC's interaction with NIMA and the cell cycle defects produced by DeltaN-TINC expression suggest possible roles for TINC and NIMA during nuclear membrane fission. PMID:15590818

  1. Cloning and Characterization of an Aspergillus nidulans Gene Involved in the Regulation of Penicillin Biosynthesis

    PubMed Central

    Van den Brulle, Jan; Steidl, Stefan; Brakhage, Axel A.

    1999-01-01

    To identify regulators of penicillin biosynthesis, a previously isolated mutant of Aspergillus nidulans (Prg-1) which carried the trans-acting prgA1 mutation was used. This mutant also contained fusions of the penicillin biosynthesis genes acvA and ipnA with reporter genes (acvA-uidA and ipnA-lacZ) integrated in a double-copy arrangement at the chromosomal argB gene. The prgA1 mutant strain exhibited only 20 to 50% of the ipnA-lacZ and acvA-uidA expression exhibited by the wild-type strain and had only 20 to 30% of the penicillin produced by the wild-type strain. Here, using complementation with a genomic cosmid library, we isolated a gene (suAprgA1) which complemented the prgA1 phenotype to the wild-type phenotype; i.e., the levels of expression of both gene fusions and penicillin production were nearly wild-type levels. Analysis of the suAprgA1 gene in the prgA1 mutant did not reveal any mutation in the suAprgA1 gene or unusual transcription of the gene. This suggested that the suAprgA1 gene is a suppressor of the prgA1 mutation. The suAprgA1 gene is 1,245 bp long. Its five exons encode a deduced protein that is 303 amino acids long. The putative SUAPRGA1 protein was similar to both the human p32 protein and Mam33p of Saccharomyces cerevisiae. Analysis of the ordered gene library of A. nidulans indicated that suAprgA1 is located on chromosome VI. Deletion of the suAprgA1 gene resulted in an approximately 50% reduction in ipnA-lacZ expression and in a slight reduction in acvA-uidA expression. The ΔsuAprgA1 strain produced about 60% of the amount of penicillin produced by the wild-type strain. PMID:10583968

  2. Targeting Fungal Genes by Diced siRNAs: A Rapid Tool to Decipher Gene Function in Aspergillus nidulans

    PubMed Central

    Kalleda, Natarajaswamy; Naorem, Aruna; Manchikatla, Rajam V.

    2013-01-01

    Background Gene silencing triggered by chemically synthesized small interfering RNAs (siRNAs) has become a powerful tool for deciphering gene function in many eukaryotes. However, prediction and validation of a single siRNA duplex specific to a target gene is often ineffective. RNA interference (RNAi) with synthetic siRNA suffers from lower silencing efficacy, off-target effects and is cost-intensive, especially for functional genomic studies. With the explosion of fungal genomic information, there is an increasing need to analyze gene function in a rapid manner. Therefore, studies were performed in order to investigate the efficacy of gene silencing induced by RNase III-diced-siRNAs (d-siRNA) in model filamentous fungus, Aspergillus nidulans. Methodology/Principal Findings Stable expression of heterologous reporter gene in A. nidulans eases the examination of a new RNAi-induction route. Hence, we have optimized Agrobacterium tumefaciens-mediated transformation (AMT) of A. nidulans for stable expression of sGFP gene. This study demonstrates that the reporter GFP gene stably introduced into A. nidulans can be effectively silenced by treatment of GFP-d-siRNAs. We have shown the down-regulation of two endogenous genes, AnrasA and AnrasB of A. nidulans by d-siRNAs. We have also elucidated the function of an uncharacterized Ras homolog, rasB gene, which was found to be involved in hyphal growth and development. Further, silencing potency of d-siRNA was higher as compared to synthetic siRNA duplex, targeting AnrasA. Silencing was shown to be sequence-specific, since expression profiles of other closely related Ras family genes in d-siRNA treated AnrasA and AnrasB silenced lines exhibited no change in gene expression. Conclusions/Significance We have developed and applied a fast, specific and efficient gene silencing approach for elucidating gene function in A. nidulans using d-siRNAs. We have also optimized an efficient AMT in A. nidulans, which is useful for stable

  3. Proteolytic activation of both components of the cation stress–responsive Slt pathway in Aspergillus nidulans

    PubMed Central

    Mellado, Laura; Arst, Herbert N.; Espeso, Eduardo A.

    2016-01-01

    Tolerance of Aspergillus nidulans to alkalinity and elevated cation concentrations requires both SltA and SltB. Transcription factor SltA and the putative pseudokinase/protease signaling protein SltB comprise a regulatory pathway specific to filamentous fungi. In vivo, SltB is proteolytically cleaved into its two principal domains. Mutational analysis defines a chymotrypsin-like serine protease domain that mediates SltB autoproteolysis and proteolytic cleavage of SltA. The pseudokinase domain might modulate the protease activity of SltB. Three forms of the SltA transcription factor coexist in cells: a full-length, 78-kDa version and a processed, 32-kDa form, which is found in phosphorylated and unphosphorylated states. The SltA32kDa version mediates transcriptional regulation of sltB and, putatively, genes required for tolerance to cation stress and alkalinity. The full-length form, SltA78kDa, apparently has no transcriptional function. In the absence of SltB, only the primary product of SltA is detectable, and its level equals that of SltA78kDa. Mutations in sltB selected as suppressors of null vps alleles and resulting in cation/alkalinity sensitivity either reduced or eliminated SltA proteolysis. There is no evidence for cation or alkalinity regulation of SltB cleavage, but activation of sltB expression requires SltA. This work identifies the molecular mechanisms governing the Slt pathway. PMID:27307585

  4. Velvet-mediated repression of β-glucan synthesis in Aspergillus nidulans spores

    PubMed Central

    Park, Hee-Soo; Man Yu, Yeong; Lee, Mi-Kyung; Jae Maeng, Pil; Chang Kim, Sun; Yu, Jae-Hyuk

    2015-01-01

    Beta-glucans are a heterologous group of fibrous glucose polymers that are a major constituent of cell walls in Ascomycetes and Basidiomycetes fungi. Synthesis of β (1,3)- and (1,6)-glucans is coordinated with fungal cell growth and development, thus, is under tight genetic regulation. Here, we report that β-glucan synthesis in both asexual and sexual spores is turned off by the NF-kB like fungal regulators VosA and VelB in Aspergillus nidulans. Our genetic and genomic analyses have revealed that both VosA and VelB are necessary for proper down-regulation of cell wall biosynthetic genes including those associated with β-glucan synthesis in both types of spores. The deletion of vosA or velB results in elevated accumulation of β-glucan in asexual spores. Double mutant analyses indicate that VosA and VelB play an inter-dependent role in repressing β-glucan synthesis in asexual spores. In vivo chromatin immuno-precipitation analysis shows that both VelB and VosA bind to the promoter region of the β-glucan synthase gene fksA in asexual spores. Similarly, VosA is required for proper repression of β-glucan synthesis in sexual spores. In summary, the VosA-VelB hetero-complex is a key regulatory unit tightly controlling proper levels of β-glucan synthesis in asexual and sexual spores. PMID:25960370

  5. The penicillin regulator PENR1 of Aspergillus nidulans is a HAP-like transcriptional complex.

    PubMed

    Litzka, O; Papagiannopolous, P; Davis, M A; Hynes, M J; Brakhage, A A

    1998-02-01

    In Aspergillus nidulans, a DNA-binding complex, PENR1, was shown to bind to two CCAAT-box-containing DNA elements located in the promoter regions of the bidirectionally oriented penicillin biosynthesis genes acvA and ipnA, and of the aat promoter. Here, partial purification of PENR1 and western blotting using anti-HAPC sera indicated that the previously identified HAPC protein, which was suggested to be part of the CCAAT-binding complex AnCF, is also part of PENR1. This was confirmed by band shift assays using protein extracts of a delta hapC strain which exhibited no PENR1 DNA-binding activity. Supershift assays and immunoprecipitation analysis using anti-HAPC sera provided evidence that HAPC is part of the PENR1 complex. In delta hapC strains, penicillin production was reduced, as was expression of both an ipnA-lacZ and aat-lacZ gene fusion. Hence, HAPC-containing PENR1 appears to act as an activator on ipnA and aat expression. However, deletion of hapC had little effect on acvA expression during a fermentation run in fermentation medium. Previous results which had shown that specific deletion of the PENR1-binding site between acvA and ipnA resulted in a strong increase of expression of an acvA-uidA gene fusion, together with the present data, suggest the possibility of the existence of a repressor protein that binds close to or overlaps the PENR1-binding site. It is also shown that binding of PENR1 induced bending of a DNA fragment spanning the PENR1-binding site between acvA and ipnA in vitro.

  6. Improvement of Aspergillus nidulans penicillin production by targeting AcvA to peroxisomes.

    PubMed

    Herr, Andreas; Fischer, Reinhard

    2014-09-01

    Aspergillus nidulans is able to synthesize penicillin and serves as a model to study the regulation of its biosynthesis. Only three enzymes are required to form the beta lactam ring tripeptide, which is comprised of l-cysteine, l-valine and l-aminoadipic acid. Whereas two enzymes, AcvA and IpnA localize to the cytoplasm, AatA resides in peroxisomes. Here, we tested a novel strategy to improve penicillin production, namely the change of the residence of the enzymes involved in the biosynthesis. We tested if targeting of AcvA or IpnA (or both) to peroxisomes would increase the penicillin yield. Indeed, AcvA peroxisomal targeting led to a 3.2-fold increase. In contrast, targeting IpnA to peroxisomes caused a complete loss of penicillin production. Overexpression of acvA, ipnA or aatA resulted in 1.4, 2.8 and 3.1-fold more penicillin, respectively in comparison to wildtype. Simultaneous overexpression of all three enzymes resulted even in 6-fold more penicillin. Combination of acvA peroxisomal targeting and overexpression of the gene led to 5-fold increase of the penicillin titer. At last, the number of peroxisomes was increased through overexpression of pexK. A strain with the double number of peroxisomes produced 2.3 times more penicillin. These results show that penicillin production can be triggered at several levels of regulation, one of which is the subcellular localization of the enzymes.

  7. Novel β-1,4-Mannanase Belonging to a New Glycoside Hydrolase Family in Aspergillus nidulans*

    PubMed Central

    Shimizu, Motoyuki; Kaneko, Yuhei; Ishihara, Saaya; Mochizuki, Mai; Sakai, Kiyota; Yamada, Miyuki; Murata, Shunsuke; Itoh, Eriko; Yamamoto, Tatsuya; Sugimura, Yu; Hirano, Tatsuya; Takaya, Naoki; Kobayashi, Tetsuo; Kato, Masashi

    2015-01-01

    Many filamentous fungi produce β-mannan-degrading β-1,4-mannanases that belong to the glycoside hydrolase 5 (GH5) and GH26 families. Here we identified a novel β-1,4-mannanase (Man134A) that belongs to a new glycoside hydrolase (GH) family (GH134) in Aspergillus nidulans. Blast analysis of the amino acid sequence using the NCBI protein database revealed that this enzyme had no similarity to any sequences and no putative conserved domains. Protein homologs of the enzyme were distributed to limited fungal and bacterial species. Man134A released mannobiose (M2), mannotriose (M3), and mannotetraose (M4) but not mannopentaose (M5) or higher manno-oligosaccharides when galactose-free β-mannan was the substrate from the initial stage of the reaction, suggesting that Man134A preferentially reacts with β-mannan via a unique catalytic mode. Man134A had high catalytic efficiency (kcat/Km) toward mannohexaose (M6) compared with the endo-β-1,4-mannanase Man5C and notably converted M6 to M2, M3, and M4, with M3 being the predominant reaction product. The action of Man5C toward β-mannans was synergistic. The growth phenotype of a Man134A disruptant was poor when β-mannans were the sole carbon source, indicating that Man134A is involved in β-mannan degradation in vivo. These findings indicate a hitherto undiscovered mechanism of β-mannan degradation that is enhanced by the novel β-1,4-mannanase, Man134A, when combined with other mannanolytic enzymes including various endo-β-1,4-mannanases. PMID:26385921

  8. Proteolytic activation of both components of the cation stress-responsive Slt pathway in Aspergillus nidulans.

    PubMed

    Mellado, Laura; Arst, Herbert N; Espeso, Eduardo A

    2016-08-15

    Tolerance of Aspergillus nidulans to alkalinity and elevated cation concentrations requires both SltA and SltB. Transcription factor SltA and the putative pseudokinase/protease signaling protein SltB comprise a regulatory pathway specific to filamentous fungi. In vivo, SltB is proteolytically cleaved into its two principal domains. Mutational analysis defines a chymotrypsin-like serine protease domain that mediates SltB autoproteolysis and proteolytic cleavage of SltA. The pseudokinase domain might modulate the protease activity of SltB. Three forms of the SltA transcription factor coexist in cells: a full-length, 78-kDa version and a processed, 32-kDa form, which is found in phosphorylated and unphosphorylated states. The SltA32kDa version mediates transcriptional regulation of sltB and, putatively, genes required for tolerance to cation stress and alkalinity. The full-length form, SltA78kDa, apparently has no transcriptional function. In the absence of SltB, only the primary product of SltA is detectable, and its level equals that of SltA78kDa. Mutations in sltB selected as suppressors of null vps alleles and resulting in cation/alkalinity sensitivity either reduced or eliminated SltA proteolysis. There is no evidence for cation or alkalinity regulation of SltB cleavage, but activation of sltB expression requires SltA. This work identifies the molecular mechanisms governing the Slt pathway. PMID:27307585

  9. Changes of global gene expression and secondary metabolite accumulation during light-dependent Aspergillus nidulans development.

    PubMed

    Bayram, Özgür; Feussner, Kirstin; Dumkow, Marc; Herrfurth, Cornelia; Feussner, Ivo; Braus, Gerhard H

    2016-02-01

    Fungal development and secondary metabolite production are coordinated by regulatory complexes as the trimeric velvet complex. Light accelerates asexual but decreases sexual development of the filamentous fungus Aspergillus nidulans. Changes in gene expression and secondary metabolite accumulation in response to environmental stimuli have been the focus of many studies, but a comprehensive comparison during entire development is lacking. We compared snapshots of transcript and metabolite profiles during fungal development in dark or light. Overall 2.014 genes corresponding to 19% of the genome were differentially expressed when submerged vegetative hyphae were compared to surface development. Differentiation was preferentially asexual in light or preferentially sexual connected to delayed asexual development in dark. Light induces significantly gene expression within the first 24-48h after the transfer to surfaces. Many light induced genes are also expressed in dark after a delay of up to two days, which might be required for preparation of enhanced sexual development. Darkness results in a massive transcriptional reprogramming causing a peak of lipid-derived fungal pheromone synthesis (psi factors) during early sexual development and the expression of genes for cell-wall degradation presumably to mobilize the energy for sexual differentiation. Accumulation of secondary metabolites like antitumoral terrequinone A or like emericellamide start under light conditions, whereas the mycotoxin sterigmatocystin or asperthecin and emodin appear under dark conditions during sexual development. Amino acid synthesis and pool rapidly drop after 72-96h in dark. Subsequent initiation of apoptotic cell-death pathways in darkness happens significantly later than in light. This illustrates that fungal adaptation in differentiation and secondary metabolite production to light conditions requires the reprogramming of one fifth of the potential of its genome.

  10. Expression and specificity profile of the major acetate transporter AcpA in Aspergillus nidulans.

    PubMed

    Sá-Pessoa, Joana; Amillis, Sotiris; Casal, Margarida; Diallinas, George

    2015-03-01

    AcpA has been previously characterized as a high-affinity transporter essential for the uptake and use of acetate as sole carbon source in Aspergillus nidulans. Here, we follow the expression profile of AcpA and define its substrate specificity. AcpA-mediated acetate transport is detected from the onset of conidiospore germination, peaks at the time of germ tube emergence, and drops to low basal levels in germlings and young mycelia, where a second acetate transporter is also becoming apparent. AcpA activity also responds to acetate presence in the growth medium, but is not subject to either carbon or nitrogen catabolite repression. Short-chain monocarboxylates (benzoate, formate, butyrate and propionate) inhibit AcpA-mediated acetate transport with apparent inhibition constants (Ki) of 16.89±2.12, 9.25±1.01, 12.06±3.29 and 1.44±0.13mM, respectively. AcpA is also shown not to be directly involved in ammonia export, as proposed for its Saccharomyces cerevisiae homologue Ady2p. In the second part of this work, we search for the unknown acetate transporter expressed in mycelia, and for other transporters that might contribute to acetate uptake. In silico analysis, genetic construction of relevant null mutants, and uptake assays, reveal that the closest AcpA homologue (AN1839), named AcpB, is the 'missing' secondary acetate transporter in mycelia. We also identify two major short-chain carboxylate (lactate, succinate, pyruvate and malate) transporters, named JenA (AN6095) and JenB (AN6703), which however are not involved in acetate uptake. This work establishes a framework for further exploiting acetate and carboxylate transport in filamentous ascomycetes. PMID:25708319

  11. A putative APSES transcription factor is necessary for normal growth and development of Aspergillus nidulans.

    PubMed

    Lee, Ji-Yeon; Kim, Lee-Han; Kim, Ha-Eun; Park, Jae-Sin; Han, Kap-Hoon; Han, Dong-Min

    2013-12-01

    The nsdD gene encoding a GATA type transcription factor positively controls sexual development in Aspergillus nidulans. According to microarray data, 20 genes that were upregulated by deleting nsdD during various life cycle stages were randomly selected and deleted for functional analysis. None of the mutants showed apparent changes in growth or development compared with those of the wild-type except the AN3154 gene that encodes a putative APSES transcription factor and is an ortholog of Saccharomyces cerevisiae swi4. Deleting AN3154 resulted in retarded growth and development, and the gene was named rgdA (retared growth and development). The rgdA deletion mutant developed a reduced number of conidia even under favorable conditions for asexual development. The retarded growth and development was partially suppressed by the veA1 mutation. The conidial heads of the mutant aborted, showing reduced and irregular shaped phialides. Fruiting body development was delayed compared with that in the wild-type. The mutant did not respond to various nutritional or environmental factors that affected the development patterns. The rgdA gene was expressed at low levels throughout the life cycle and was not significantly affected by several regulators of sexual and asexual development such as nsdD, veA, stuA, or brlA. However, the rgdA gene affected brlA and abaA expression, which function as key regulators of asexual sporulation, suggesting that rgdA functions upstream of those genes.

  12. Localization and function of calmodulin in live-cells of Aspergillus nidulans.

    PubMed

    Chen, Shaochun; Song, Yiju; Cao, Jinling; Wang, Gang; Wei, Hua; Xu, Xushi; Lu, Ling

    2010-03-01

    Calmodulin (CaM) is a small, eukaryotic protein that reversibly binds Ca(2+). Study of CaM localization in genetically tractable organisms has yielded many insights into CaM function. Here, we described the dynamic localization of Aspergillus nidulans CaM (AnCaM) in live-cells by using recombination strains with homologous, single cross-over insertions at the target gene which placed the GFP fused copy under the inducible alcA promoter and the RFP-CaM integration under the native cam promoter. We found that the localization of CaM fusion was quite dynamic throughout the hypha and was concentrated to the active growing sites during germination, hyphal growth, cytokinesis and conidiation. The depletion of CaM by alcA promoter repression induced the explicit abnormalities of germlings with the swollen germ tubes. In addition, the position of highly concentrated GFP-CaM in the extreme apex seemed to determine the hyphal orientation. These data collectively suggest that CaM is constantly required for new hyphal growth. In contrast to this constant accumulation at the apex, GFP-CaM was only transiently localized at septum sites during cytokinesis. Notably, depletion of CaM caused the defect of septation with a completely blocked septum formation indicating that the transient CaM accumulation at the septum site is essential for septation. Moreover, the normal localization of CaM at a hyphal tip required the presence of the functional actin cytoskeleton and the motor protein KipA, which is indispensable for positioning Spitzenkörper. This is the first report of CaM localization and function in live-cells by the site-specific homologous integration in filamentous fungi.

  13. More Than a Repair Enzyme: Aspergillus nidulans Photolyase-like CryA Is a Regulator of Sexual Development

    PubMed Central

    Bayram, Özgür; Biesemann, Christoph; Krappmann, Sven; Galland, Paul

    2008-01-01

    Cryptochromes are blue-light receptors that have presumably evolved from the DNA photolyase protein family, and the genomes of many organisms contain genes for both types of molecules. Both protein structures resemble each other, which suggests that light control and light protection share a common ancient origin. In the genome of the filamentous fungus Aspergillus nidulans, however, only one cryptochrome/photolyase-encoding gene, termed cryA, was identified. Deletion of the cryA gene triggers sexual differentiation under inappropriate culture conditions and results in up-regulation of transcripts encoding regulators of fruiting body formation. CryA is a protein whose N- and C-terminal synthetic green fluorescent protein fusions localize to the nucleus. CryA represses sexual development under UVA350-370 nm light both on plates and in submerged culture. Strikingly, CryA exhibits photorepair activity as demonstrated by heterologous complementation of a DNA repair-deficient Escherichia coli strain as well as overexpression in an A. nidulans uvsBΔ genetic background. This is in contrast to the single deletion cryAΔ strain, which does not show increased sensitivity toward UV-induced damage. In A. nidulans, cryA encodes a novel type of cryptochrome/photolyase that exhibits a regulatory function during light-dependent development and DNA repair activity. This represents a paradigm for the evolutionary transition between photolyases and cryptochromes. PMID:18495868

  14. The Aspergillus nidulans multimeric CCAAT binding complex AnCF is negatively autoregulated via its hapB subunit gene.

    PubMed

    Steidl, S; Hynes, M J; Brakhage, A A

    2001-03-01

    Cis-acting CCAAT elements are frequently found in eukaryotic promoter regions. Many of them are bound by conserved multimeric complexes. In the fungus Aspergillus nidulans the respective complex was designated AnCF (A. nidulans CCAAT binding factor). AnCF is composed of at least three subunits designated HapB, HapC and HapE. Here, we show that the promoter regions of the hapB genes in both A. nidulans and Aspergillus oryzae contain two inversely oriented, conserved CCAAT boxes (box alpha and box beta). Electrophoretic mobility shift assays (EMSAs) using both nuclear extracts and the purified, reconstituted AnCF complex indicated that AnCF binding in vitro to these boxes occurs in a non-mutually exclusive manner. Western and Northern blot analyses showed that steady-state levels of HapB protein as well as hapB mRNA were elevated in hapC and hapE deletion mutants, suggesting a repressing effect of AnCF on hapB expression. Consistently, in a hapB deletion background the hapB-lacZ expression level was elevated compared with the expression in the wild-type. This was further supported by overexpression of hapB using an inducible alcA-hapB construct. Induction of alcA-hapB expression strongly repressed the expression of a hapB-lacZ gene fusion. However, mutagenesis of box beta led to a fivefold reduced expression of a hapB-lacZ gene fusion compared with the expression derived from a wild-type hapB-lacZ fusion. These results indicate that (i) box beta is an important positive cis-acting element in hapB regulation, (ii) AnCF does not represent the corresponding positive trans-acting factor and (iii) that AnCF is involved in repression of hapB.

  15. Characterization of the Aspergillus nidulans aspnd1 gene demonstrates that the ASPND1 antigen, which it encodes, and several Aspergillus fumigatus immunodominant antigens belong to the same family.

    PubMed Central

    Calera, J A; Ovejero, M C; López-Medrano, R; Segurado, M; Puente, P; Leal, F

    1997-01-01

    For the first time, an immunodominant Aspergillus nidulans antigen (ASPND1) consistently reactive with serum samples from aspergilloma patients has been purified and characterized, and its coding gene (aspnd1) has been cloned and sequenced. ASPND1 is a glycoprotein with four N-glycosidically-bound sugar chains (around 2.1 kDa each) which are not necessary for reactivity with immune human sera. The polypeptide part is synthesized as a 277-amino-acid precursor of 30.6 kDa that after cleavage of a putative signal peptide of 16 amino acids, affords a mature protein of 261 amino acids with a molecular mass of 29 kDa and a pI of 4.24 (as deduced from the sequence). The ASPND1 protein is 53.1% identical to the AspfII allergen from Aspergillus fumigatus and 48% identical to an unpublished Candida albicans antigen. All of the cysteine residues and most of the glycosylation sites are perfectly conserved in the three proteins, suggesting a similar but yet unknown function. Analysis of the primary structure of the ASPND1 coding gene (aspnd1) has allowed the establishment of a clear relationship between several previously reported A. fumigatus and A. nidulans immunodominant antigens. PMID:9119471

  16. The CCAAT-binding complex of eukaryotes: evolution of a second NLS in the HapB subunit of the filamentous fungus Aspergillus nidulans despite functional conservation at the molecular level between yeast, A.nidulans and human.

    PubMed

    Tüncher, André; Spröte, Petra; Gehrke, Alexander; Brakhage, Axel A

    2005-09-23

    The heterotrimeric CCAAT-binding complex is evolutionarily conserved in eukaryotic organisms, including fungi, plants and mammals. In the filamentous fungus Aspergillus nidulans, the corresponding complex was designated AnCF (A.nidulans CCAAT-binding factor). AnCF consists of the subunits HapB, HapC and HapE. All three subunits are necessary for DNA binding. HapB contains two putative nuclear localisation signal sequences (NLSs) designated NLS1 and NLS2. Previously, it was shown that only NLS2 was required for nuclear localisation of HapB. Furthermore, HapC and HapE are transported to the nucleus only in complex with HapB via a piggy back mechanism. Here, by using various GFP constructs and by establishing a novel marker gene for transformation of A.nidulans, i.e. the pabaA gene encoding p-aminobenzoic acid synthase, it was shown that the HapB homologous proteins of both Saccharomyces cerevisiae (Hap2p) and human (NF-YA) use an NLS homologous to HapB NLS1 for nuclear localisation in S.cerevisiae. Interestingly, for A.nidulans HapB, NLS1 was sufficient for nuclear localisation in S.cerevisiae. In A.nidulans, HapB NLS1 was also functional when present in a different protein context. However, in A.nidulans, both S.cerevisiae Hap2p and human NF-YA entered the nucleus only when HapB NLS2 was present in the respective proteins. In that case, both proteins Hap2p and NF-YA complemented, at least in part, the hap phenotype of A.nidulans with respect to lack of growth on acetamide. Similarly, A.nidulans HapB and human NF-YA complemented a hap2 mutant of S.cerevisiae. In summary, HapB, Hap2p and NF-YA are interchangeable. Because the A.nidulans hapB mutant was complemented, at least in part, by both the human NF-YA and S.cerevisiae Hap2p this finding suggests that the piggy-back mechanism of nuclear transport found for A.nidulans is conserved in yeast and human.

  17. Bacteria-induced natural product formation in the fungus Aspergillus nidulans requires Saga/Ada-mediated histone acetylation.

    PubMed

    Nützmann, Hans-Wilhelm; Reyes-Dominguez, Yazmid; Scherlach, Kirstin; Schroeckh, Volker; Horn, Fabian; Gacek, Agnieszka; Schümann, Julia; Hertweck, Christian; Strauss, Joseph; Brakhage, Axel A

    2011-08-23

    Sequence analyses of fungal genomes have revealed that the potential of fungi to produce secondary metabolites is greatly underestimated. In fact, most gene clusters coding for the biosynthesis of antibiotics, toxins, or pigments are silent under standard laboratory conditions. Hence, it is one of the major challenges in microbiology to uncover the mechanisms required for pathway activation. Recently, we discovered that intimate physical interaction of the important model fungus Aspergillus nidulans with the soil-dwelling bacterium Streptomyces rapamycinicus specifically activated silent fungal secondary metabolism genes, resulting in the production of the archetypal polyketide orsellinic acid and its derivatives. Here, we report that the streptomycete triggers modification of fungal histones. Deletion analysis of 36 of 40 acetyltransferases, including histone acetyltransferases (HATs) of A. nidulans, demonstrated that the Saga/Ada complex containing the HAT GcnE and the AdaB protein is required for induction of the orsellinic acid gene cluster by the bacterium. We also showed that Saga/Ada plays a major role for specific induction of other biosynthesis gene clusters, such as sterigmatocystin, terrequinone, and penicillin. Chromatin immunoprecipitation showed that the Saga/Ada-dependent increase of histone 3 acetylation at lysine 9 and 14 occurs during interaction of fungus and bacterium. Furthermore, the production of secondary metabolites in A. nidulans is accompanied by a global increase in H3K14 acetylation. Increased H3K9 acetylation, however, was only found within gene clusters. This report provides previously undescribed evidence of Saga/Ada dependent histone acetylation triggered by prokaryotes. PMID:21825172

  18. Comprehensive annotation of secondary metabolite biosynthetic genes and gene clusters of Aspergillus nidulans, A. fumigatus, A. niger and A. oryzae

    PubMed Central

    2013-01-01

    Background Secondary metabolite production, a hallmark of filamentous fungi, is an expanding area of research for the Aspergilli. These compounds are potent chemicals, ranging from deadly toxins to therapeutic antibiotics to potential anti-cancer drugs. The genome sequences for multiple Aspergilli have been determined, and provide a wealth of predictive information about secondary metabolite production. Sequence analysis and gene overexpression strategies have enabled the discovery of novel secondary metabolites and the genes involved in their biosynthesis. The Aspergillus Genome Database (AspGD) provides a central repository for gene annotation and protein information for Aspergillus species. These annotations include Gene Ontology (GO) terms, phenotype data, gene names and descriptions and they are crucial for interpreting both small- and large-scale data and for aiding in the design of new experiments that further Aspergillus research. Results We have manually curated Biological Process GO annotations for all genes in AspGD with recorded functions in secondary metabolite production, adding new GO terms that specifically describe each secondary metabolite. We then leveraged these new annotations to predict roles in secondary metabolism for genes lacking experimental characterization. As a starting point for manually annotating Aspergillus secondary metabolite gene clusters, we used antiSMASH (antibiotics and Secondary Metabolite Analysis SHell) and SMURF (Secondary Metabolite Unknown Regions Finder) algorithms to identify potential clusters in A. nidulans, A. fumigatus, A. niger and A. oryzae, which we subsequently refined through manual curation. Conclusions This set of 266 manually curated secondary metabolite gene clusters will facilitate the investigation of novel Aspergillus secondary metabolites. PMID:23617571

  19. Two separate gene clusters encode the biosynthetic pathway for the meroterpenoids, austinol and dehydroaustinol in Aspergillus nidulans

    PubMed Central

    Lo, Hsien-Chun; Entwistle, Ruth; Guo, Chun-Jun; Ahuja, Manmeet; Szewczyk, Edyta; Hung, Jui-Hsiang; Chiang, Yi-Ming; Oakley, Berl R.; Wang, Clay C. C.

    2012-01-01

    Meroterpenoids are a class of fungal natural products that are produced from polyketide and terpenoid precursors. An understanding of meroterpenoid biosynthesis at the genetic level should facilitate engineering of second-generation molecules and increasing production of first-generation compounds. The filamentous fungus Aspergillus nidulans has previously been found to produce two meroterpenoids, austinol and dehydroaustinol. Using targeted deletions that we created, we have determined that, surprisingly, two separate gene clusters are required for meroterpenoid biosynthesis. One is a cluster of four genes including a polyketide synthase gene, ausA. The second is a cluster of ten additional genes including a prenyltransferase gene, ausN, located on a separate chromosome. Chemical analysis of mutant extracts enabled us to isolate 3,5-dimethylorsellinic acid and ten additional meroterpenoids that are either intermediates or shunt products from the biosynthetic pathway. Six of them were identified as novel meroterpenoids in this study. Our data, in aggregate, allow us to propose a complete biosynthetic pathway for the A. nidulans meroterpenoids. PMID:22329759

  20. Stress tolerances of nullmutants of function-unknown genes encoding menadione stress-responsive proteins in Aspergillus nidulans.

    PubMed

    Leiter, Éva; Bálint, Mihály; Miskei, Márton; Orosz, Erzsébet; Szabó, Zsuzsa; Pócsi, István

    2016-07-01

    A group of menadione stress-responsive function-unkown genes of Aspergillus nidulans (Locus IDs ANID_03987.1, ANID_06058.1, ANID_10219.1, and ANID_10260.1) was deleted and phenotypically characterized. Importantly, comparative and phylogenetic analyses of the tested A. nidulans genes and their orthologs shed light only on the presence of a TANGO2 domain with NRDE protein motif in the translated ANID_06058.1 gene but did not reveal any recognizable protein-encoding domains in other protein sequences. The gene deletion strains were subjected to oxidative, osmotic, and metal ion stress and, surprisingly, only the ΔANID_10219.1 mutant showed an increased sensitivity to 0.12 mmol l(-1) menadione sodium bisulfite. The gene deletions affected the stress sensitivities (tolerances) irregularly, for example, some strains grew more slowly when exposed to various oxidants and/or osmotic stress generating agents, meanwhile the ΔANID_10260.1 mutant possessed a wild-type tolerance to all stressors tested. Our results are in line with earlier studies demonstrating that the deletions of stress-responsive genes do not confer necessarily any stress-sensitivity phenotypes, which can be attributed to compensatory mechanisms based on other elements of the stress response system with overlapping functions.

  1. LAMMER Kinase LkhA Plays Multiple Roles in the Vegetative Growth and Asexual and Sexual Development of Aspergillus nidulans

    PubMed Central

    Kang, Eun-Hye; Kim, Ji-ae; Oh, Hyun-Woo; Park, Hee-Moon

    2013-01-01

    LAMMER kinase plays pivotal roles in various physiological processes in eukaryotes; however, its function in filamentous fungi is not known. We performed molecular studies on the function of the Aspergillus nidulans LAMMER kinase, LkhA, and report its involvement in multiple developmental processes. The gene for LkhA was highly expressed during reproductive organ development, such as that of conidiophores and cleistothecia. During vegetative growth, the patterns of germ tube emergence and hyphal polarity were changed and septation was increased by lkhA deletion. Northern analyses showed that lkhA regulated the transcription of brlA, csnD, and ppoA, which supported the detrimental effect of lkhA-deletion on asexual and sexual differentiation. LkhA also affected expression of cyclin-dependent kinase NimXcdc2, a multiple cell cycle regulator, and StuA, an APSES family of fungal transcription factors that play pivotal roles in multiple differentiation processes. Here, for the first time, we present molecular evidence showing that LAMMER kinase is involved in A. nidulans development by modulating the expression of key regulators of developmental processes. PMID:23516554

  2. The old 3-oxoadipate pathway revisited: new insights in the catabolism of aromatics in the saprophytic fungus Aspergillus nidulans.

    PubMed

    Martins, Tiago M; Hartmann, Diego O; Planchon, Sébastien; Martins, Isabel; Renaut, Jenny; Silva Pereira, Cristina

    2015-01-01

    Aspergilli play major roles in the natural turnover of elements, especially through the decomposition of plant litter, but the end catabolism of lignin aromatic hydrocarbons remains largely unresolved. The 3-oxoadipate pathway of their degradation combines the catechol and the protocatechuate branches, each using a set of specific genes. However, annotation for most of these genes is lacking or attributed to poorly- or un-characterised families. Aspergillus nidulans can utilise as sole carbon/energy source either benzoate or salicylate (upstream aromatic metabolites of the protocatechuate and the catechol branches, respectively). Using this cultivation strategy and combined analyses of comparative proteomics, gene mining, gene expression and characterisation of particular gene-replacement mutants, we precisely assigned most of the steps of the 3-oxoadipate pathway to specific genes in this fungus. Our findings disclose the genetically encoded potential of saprophytic Ascomycota fungi to utilise this pathway and provide means to untie associated regulatory networks, which are vital to heightening their ecological significance.

  3. Enzymatic lesions in methionine mutants of Aspergillus nidulans: role and regulation of an alternative pathway for cysteine and methionine synthesis.

    PubMed Central

    Paszewski, A; Grabski, J

    1975-01-01

    In Aspergillus nidulans the pathway involving cystathionine formation is the main one for homocysteine synthesis. Mutants lacking cystathionine gamma-synthase or beta-cystathionase are auxotrophs suppressible by: (i) mutations in the main pathway of cysteine synthesis (cysA1, cysB1, and cysC1), (ii) mutations causing stimulation of cysteine catabolism (su101), and (iii) mutations in a presumed regulatory gene (suAmeth). A relative shortage of cysteine in the first group of suppressors causes a derepression of homocysteine synthase, the enzyme involved in the alternative pathway of homocysteine synthesis. A similar derepression is observed in the suAmeth strain. Homocysteine synthesized by this pathway serves as precursor for cysteine and methionine synthesis. A mutant with altered homocysteine synthase is a prototroph, indicating that this enzyme is not essential for the fungus. Images PMID:1102536

  4. Secondary Metabolism and Development Is Mediated by LlmF Control of VeA Subcellular Localization in Aspergillus nidulans

    PubMed Central

    Palmer, Jonathan M.; Theisen, Jeffrey M.; Duran, Rocio M.; Grayburn, W. Scott; Calvo, Ana M.; Keller, Nancy P.

    2013-01-01

    Secondary metabolism and development are linked in Aspergillus through the conserved regulatory velvet complex composed of VeA, VelB, and LaeA. The founding member of the velvet complex, VeA, shuttles between the cytoplasm and nucleus in response to alterations in light. Here we describe a new interaction partner of VeA identified through a reverse genetics screen looking for LaeA-like methyltransferases in Aspergillus nidulans. One of the putative LaeA-like methyltransferases identified, LlmF, is a negative regulator of sterigmatocystin production and sexual development. LlmF interacts directly with VeA and the repressive function of LlmF is mediated by influencing the localization of VeA, as over-expression of llmF decreases the nuclear to cytoplasmic ratio of VeA while deletion of llmF results in an increased nuclear accumulation of VeA. We show that the methyltransferase domain of LlmF is required for function; however, LlmF does not directly methylate VeA in vitro. This study identifies a new interaction partner for VeA and highlights the importance of cellular compartmentalization of VeA for regulation of development and secondary metabolism. PMID:23341778

  5. Molecular genetic analysis reveals that a nonribosomal peptide synthetase-like (NRPS-like) gene in Aspergillus nidulans is responsible for microperfuranone biosynthesis.

    PubMed

    Yeh, Hsu-Hua; Chiang, Yi-Ming; Entwistle, Ruth; Ahuja, Manmeet; Lee, Kuan-Han; Bruno, Kenneth S; Wu, Tung-Kung; Oakley, Berl R; Wang, Clay C C

    2012-11-01

    Genome sequencing of Aspergillus species including Aspergillus nidulans has revealed that there are far more secondary metabolite biosynthetic gene clusters than secondary metabolites isolated from these organisms. This implies that these organisms can produce additional secondary metabolites, which have not yet been elucidated. The A. nidulans genome contains 12 nonribosomal peptide synthetase (NRPS), one hybrid polyketide synthase/NRPS, and 14 NRPS-like genes. The only NRPS-like gene in A. nidulans with a known product is tdiA, which is involved in terrequinone A biosynthesis. To attempt to identify the products of these NRPS-like genes, we replaced the native promoters of the NRPS-like genes with the inducible alcohol dehydrogenase (alcA) promoter. Our results demonstrated that induction of the single NRPS-like gene AN3396.4 led to the enhanced production of microperfuranone. Furthermore, heterologous expression of AN3396.4 in Aspergillus niger confirmed that only one NRPS-like gene, AN3396.4, is necessary for the production of microperfuranone.

  6. The Aspergillus nidulans alcA promoter drives tightly regulated conditional gene expression in Aspergillus fumigatus permitting validation of essential genes in this human pathogen.

    PubMed

    Romero, Beatriz; Turner, Geoffrey; Olivas, Israel; Laborda, Fernando; De Lucas, J Ramón

    2003-11-01

    Aspergillus fumigatus causes invasive aspergillosis, a mycosis that is usually fatal in immunocompromised patients. Functional genomics in this fungus will aid the discovery of novel antifungal drugs to treat invasive aspergillosis. However, there is still a need for appropriate molecular genetic tools to facilitate such functional studies. Here, we describe the use of a conditional gene expression system allowing the identification of novel therapeutic targets through validation of essential genes in A. fumigatus. This system is based on the capacity of the Aspergillus nidulans alcA promoter (alcA(p)) to tightly regulate gene expression in this fungus. Conditionally regulated gene expression in A. fumigatus was demonstrated by transcriptional and phenotypic analyses of strains expressing a nuclear migration gene with a terminal phenotype, the A. fumigatus nudC gene, under control of this promoter. This conditional expression system, the first one described in A. fumigatus, will also be useful for investigating the function of essential genes by altering the threonine/glucose ratio in the growth medium.

  7. Protein kinase C overexpression suppresses calcineurin-associated defects in Aspergillus nidulans and is involved in mitochondrial function.

    PubMed

    Colabardini, Ana Cristina; Ries, Laure Nicolas Annick; Brown, Neil Andrew; Savoldi, Marcela; Dinamarco, Taísa Magnani; von Zeska Kress, Marcia Regina; von Zeska, Marcia Regina; Goldman, Maria Helena S; Goldman, Gustavo Henrique

    2014-01-01

    In filamentous fungi, intracellular signaling pathways which are mediated by changing calcium levels and/or by activated protein kinase C (Pkc), control fungal adaptation to external stimuli. A rise in intracellular Ca2+ levels activates calcineurin subunit A (CnaA), which regulates cellular calcium homeostasis among other processes. Pkc is primarily involved in maintaining cell wall integrity (CWI) in response to different environmental stresses. Cross-talk between the Ca2+ and Pkc-mediated pathways has mainly been described in Saccharomyces cerevisiae and in a few other filamentous fungi. The presented study describes a genetic interaction between CnaA and PkcA in the filamentous fungus Aspergillus nidulans. Overexpression of pkcA partially rescues the phenotypes caused by a cnaA deletion. Furthermore, CnaA appears to affect the regulation of a mitogen-activated kinase, MpkA, involved in the CWI pathway. Reversely, PkcA is involved in controlling intracellular calcium homeostasis, as was confirmed by microarray analysis. Furthermore, overexpression of pkcA in a cnaA deletion background restores mitochondrial number and function. In conclusion, PkcA and CnaA-mediated signaling appear to share common targets, one of which appears to be MpkA of the CWI pathway. Both pathways also regulate components involved in mitochondrial biogenesis and function. This study describes targets for PkcA and CnaA-signaling pathways in an A. nidulans and identifies a novel interaction of both pathways in the regulation of cellular respiration.

  8. Protein Kinase C Overexpression Suppresses Calcineurin-Associated Defects in Aspergillus nidulans and Is Involved in Mitochondrial Function

    PubMed Central

    Brown, Neil Andrew; Savoldi, Marcela; Dinamarco, Taísa Magnani; von Zeska, Marcia Regina; Goldman, Maria Helena S.; Goldman, Gustavo Henrique

    2014-01-01

    In filamentous fungi, intracellular signaling pathways which are mediated by changing calcium levels and/or by activated protein kinase C (Pkc), control fungal adaptation to external stimuli. A rise in intracellular Ca2+ levels activates calcineurin subunit A (CnaA), which regulates cellular calcium homeostasis among other processes. Pkc is primarily involved in maintaining cell wall integrity (CWI) in response to different environmental stresses. Cross-talk between the Ca2+ and Pkc-mediated pathways has mainly been described in Saccharomyces cerevisiae and in a few other filamentous fungi. The presented study describes a genetic interaction between CnaA and PkcA in the filamentous fungus Aspergillus nidulans. Overexpression of pkcA partially rescues the phenotypes caused by a cnaA deletion. Furthermore, CnaA appears to affect the regulation of a mitogen-activated kinase, MpkA, involved in the CWI pathway. Reversely, PkcA is involved in controlling intracellular calcium homeostasis, as was confirmed by microarray analysis. Furthermore, overexpression of pkcA in a cnaA deletion background restores mitochondrial number and function. In conclusion, PkcA and CnaA-mediated signaling appear to share common targets, one of which appears to be MpkA of the CWI pathway. Both pathways also regulate components involved in mitochondrial biogenesis and function. This study describes targets for PkcA and CnaA-signaling pathways in an A. nidulans and identifies a novel interaction of both pathways in the regulation of cellular respiration. PMID:25153325

  9. Functional characterisation of the non-essential protein kinases and phosphatases regulating Aspergillus nidulans hydrolytic enzyme production

    PubMed Central

    2013-01-01

    Background Despite recent advances in the understanding of lignocellulolytic enzyme regulation, less is known about how different carbon sources are sensed and the signaling cascades that result in the adaptation of cellular metabolism and hydrolase secretion. Therefore, the role played by non-essential protein kinases (NPK) and phosphatases (NPP) in the sensing of carbon and/or energetic status was investigated in the model filamentous fungus Aspergillus nidulans. Results Eleven NPKs and seven NPPs were identified as being involved in cellulase, and in some cases also hemicellulase, production in A. nidulans. The regulation of CreA-mediated carbon catabolite repression (CCR) in the parental strain was determined by fluorescence microscopy, utilising a CreA::GFP fusion protein. The sensing of phosphorylated glucose, via the RAS signalling pathway induced CreA repression, while carbon starvation resulted in derepression. Growth on cellulose represented carbon starvation and derepressing conditions. The involvement of the identified NPKs in the regulation of cellulose-induced responses and CreA derepression was assessed by genome-wide transcriptomics (GEO accession 47810). CreA::GFP localisation and the restoration of endocellulase activity via the introduction of the ∆creA mutation, was assessed in the NPK-deficient backgrounds. The absence of either the schA or snfA kinase dramatically reduced cellulose-induced transcriptional responses, including the expression of hydrolytic enzymes and transporters. The mechanism by which these two NPKs controlled gene transcription was identified, as the NPK-deficient mutants were not able to unlock CreA-mediated carbon catabolite repression under derepressing conditions, such as carbon starvation or growth on cellulose. Conclusions Collectively, this study identified multiple kinases and phosphatases involved in the sensing of carbon and/or energetic status, while demonstrating the overlapping, synergistic roles of schA and

  10. A single p34cdc2 protein kinase (encoded by nimXcdc2) is required at G1 and G2 in Aspergillus nidulans.

    PubMed

    Osmani, A H; van Peij, N; Mischke, M; O'Connell, M J; Osmani, S A

    1994-06-01

    We have cloned and sequenced a homolog of cdc2 from Aspergillus nidulans that can complement the Schizosaccharomyces pombe cdc2-33 mutation. The gene was deleted and is required for continued nuclear DNA replication but not for mitochondrial DNA replication. Three different temperature-sensitive alleles were generated by reverse genetics. All of the mutations generate the nim phenotype of A. nidulans. The new gene was designated nimXcdc2 as it is not allelic to any of the other nim genes (nimA to nimW) of A. nidulans. Reciprocal shift experiments place an essential function for nimXcdc2 in G1 and G2. Antipeptide antibodies were generated that detect NIMXcdc2, and antisera were also generated to detect NIMEcyclinB. The two p34cdc2 protein species previously detected in A. nidulans, p34 and p37, both precipitate using NIMXcdc2 C-terminus-specific antibodies but only p34 co-precipitates with NIMEcyclinB. Dephosphorylation of denatured p34 converts it to the p37 form, showing p37 to be the non-phosphorylated form of NIMXcdc2. The phosphorylation of p34 is therefore associated with its interaction with NIMEcyclinB. PMID:7962194

  11. Co-cultivation of Aspergillus nidulans Recombinant Strains Produces an Enzymatic Cocktail as Alternative to Alkaline Sugarcane Bagasse Pretreatment.

    PubMed

    Lima, Matheus S; Damasio, André R de L; Crnkovic, Paula M; Pinto, Marcelo R; da Silva, Ana M; da Silva, Jean C R; Segato, Fernando; de Lucas, Rosymar C; Jorge, João A; Polizeli, Maria de L T de M

    2016-01-01

    Plant materials represent a strategic energy source because they can give rise to sustainable biofuels through the fermentation of their carbohydrates. A clear example of a plant-derived biofuel resource is the sugar cane bagasse exhibiting 60-80% of fermentable sugars in its composition. However, the current methods of plant bioconversion employ severe and harmful chemical/physical pretreatments raising biofuel cost production and environmental degradation. Replacing these methods with co-cultivated enzymatic cocktails is an alternative. Here we propose a pretreatment for sugarcane bagasse using a multi-enzymatic cocktail from the co-cultivation of four Aspergillus nidulans recombinant strains. The co-cultivation resulted in the simultaneous production of GH51 arabinofuranosidase (AbfA), GH11 endo-1,4-xylanase (XlnA), GH43 endo-1,5-arabinanase (AbnA) and GH12 xyloglucan specific endo-β-1,4-glucanase (XegA). This core set of recombinant enzymes was more efficient than the alternative alkaline method in maintaining the cellulose integrity and exposing this cellulose to the following saccharification process. Thermogravimetric and differential thermal analysis revealed residual byproducts on the alkali pretreated biomass, which were not found in the enzymatic pretreatment. Therefore, the enzymatic pretreatment was residue-free and seemed to be more efficient than the applied alkaline method, which makes it suitable for bioethanol production.

  12. The formamidase gene of Aspergillus nidulans: regulation by nitrogen metabolite repression and transcriptional interference by an overlapping upstream gene.

    PubMed Central

    Fraser, J A; Davis, M A; Hynes, M J

    2001-01-01

    The ability to utilize formamide as a sole nitrogen source has been found in numerous fungi. We have cloned the fmdS gene encoding a formamidase from Aspergillus nidulans and found that it belongs to a highly conserved family of proteins separate from the major amidase families. The expression of fmdS is primarily regulated via AreA-mediated nitrogen metabolite repression and does not require the addition of exogenous inducer. Consistent with this, deletion analysis of the 5' region of fmdS has confirmed the presence of multiple AreA-binding sites containing a characteristic core GATA sequence. Under carbon starvation conditions the response to nitrogen starvation is eliminated, indicating that the lack of a carbon source may result in inactivation of AreA. Sequence analysis and isolation of cDNAs show that a gene of unknown function lies directly 5' of fmdS with its transcript overlapping the fmdS coding region. Disruption of the 5' gene and analysis of the effects of overexpression of this gene on fmdS expression has shown that expression of this upstream gene interferes with fmdS transcription, resulting in a strong dependence on AreA activation for expression. Therefore the relative position of these two genes is essential for normal regulation of fmdS. PMID:11139496

  13. Characterization of Aspergillus nidulans DidBDid2, a non-essential component of the multivesicular body pathway

    PubMed Central

    Hervás-Aguilar, América; Rodríguez-Galán, Olga; Galindo, Antonio; Abenza, Juan F.; Arst, Herbert N.; Peñalva, Miguel A.

    2010-01-01

    ESCRT-III heteropolymers mediate membrane protein cargo sorting into multivesicular endosomes for subsequent vacuolar degradation. We studied the localization of largely uncharacterized Aspergillus nidulans ESCRT-III using its key structural component Vps32 and the ‘associated’ component DidBDid2. Vps32-GFP localizes to motile early endosomes as reported, but predominates in aggregates often associated with vacuoles due to inability to dissociate from endosomes. DidBDid2 regulating Vps4 (the ATPase disassembling ESCRT-III) is not essential. Consistent with this accessory role, didBΔ is unable to block the MVB sorting of the glutamate transporter AgtA, but increases its steady-state level and mislocalizes a fraction of the permease to the plasma membrane under conditions promoting its vacuolar targeting. didBΔ exacerbates the dominant-negative growth defect resulting from Vps32-GFP over-expression. A proportion of DidB-GFP is detectable in early endosomes colocalizing with RabARab5 and accumulating in nudA1 tips, suggesting that ESCRT-III assembles on endosomes from the early steps of the endocytic pathway. PMID:20362686

  14. High-Affinity Glucose Transport in Aspergillus nidulans Is Mediated by the Products of Two Related but Differentially Expressed Genes

    PubMed Central

    Ventura, Luisa; González, Ramón; Ramón, Daniel; MacCabe, Andrew P.

    2014-01-01

    Independent systems of high and low affinity effect glucose uptake in the filamentous fungus Aspergillus nidulans. Low-affinity uptake is known to be mediated by the product of the mstE gene. In the current work two genes, mstA and mstC, have been identified that encode high-affinity glucose transporter proteins. These proteins' primary structures share over 90% similarity, indicating that the corresponding genes share a common origin. Whilst the function of the paralogous proteins is little changed, they differ notably in their patterns of expression. The mstC gene is expressed during the early phases of germination and is subject to CreA-mediated carbon catabolite repression whereas mstA is expressed as a culture tends toward carbon starvation. In addition, various pieces of genetic evidence strongly support allelism of mstC and the previously described locus sorA. Overall, our data define MstC/SorA as a high-affinity glucose transporter expressed in germinating conidia, and MstA as a high-affinity glucose transporter that operates in vegetative hyphae under conditions of carbon limitation. PMID:24751997

  15. Co-cultivation of Aspergillus nidulans Recombinant Strains Produces an Enzymatic Cocktail as Alternative to Alkaline Sugarcane Bagasse Pretreatment.

    PubMed

    Lima, Matheus S; Damasio, André R de L; Crnkovic, Paula M; Pinto, Marcelo R; da Silva, Ana M; da Silva, Jean C R; Segato, Fernando; de Lucas, Rosymar C; Jorge, João A; Polizeli, Maria de L T de M

    2016-01-01

    Plant materials represent a strategic energy source because they can give rise to sustainable biofuels through the fermentation of their carbohydrates. A clear example of a plant-derived biofuel resource is the sugar cane bagasse exhibiting 60-80% of fermentable sugars in its composition. However, the current methods of plant bioconversion employ severe and harmful chemical/physical pretreatments raising biofuel cost production and environmental degradation. Replacing these methods with co-cultivated enzymatic cocktails is an alternative. Here we propose a pretreatment for sugarcane bagasse using a multi-enzymatic cocktail from the co-cultivation of four Aspergillus nidulans recombinant strains. The co-cultivation resulted in the simultaneous production of GH51 arabinofuranosidase (AbfA), GH11 endo-1,4-xylanase (XlnA), GH43 endo-1,5-arabinanase (AbnA) and GH12 xyloglucan specific endo-β-1,4-glucanase (XegA). This core set of recombinant enzymes was more efficient than the alternative alkaline method in maintaining the cellulose integrity and exposing this cellulose to the following saccharification process. Thermogravimetric and differential thermal analysis revealed residual byproducts on the alkali pretreated biomass, which were not found in the enzymatic pretreatment. Therefore, the enzymatic pretreatment was residue-free and seemed to be more efficient than the applied alkaline method, which makes it suitable for bioethanol production. PMID:27199917

  16. High-affinity glucose transport in Aspergillus nidulans is mediated by the products of two related but differentially expressed genes.

    PubMed

    Forment, Josep V; Flipphi, Michel; Ventura, Luisa; González, Ramón; Ramón, Daniel; Maccabe, Andrew P

    2014-01-01

    Independent systems of high and low affinity effect glucose uptake in the filamentous fungus Aspergillus nidulans. Low-affinity uptake is known to be mediated by the product of the mstE gene. In the current work two genes, mstA and mstC, have been identified that encode high-affinity glucose transporter proteins. These proteins' primary structures share over 90% similarity, indicating that the corresponding genes share a common origin. Whilst the function of the paralogous proteins is little changed, they differ notably in their patterns of expression. The mstC gene is expressed during the early phases of germination and is subject to CreA-mediated carbon catabolite repression whereas mstA is expressed as a culture tends toward carbon starvation. In addition, various pieces of genetic evidence strongly support allelism of mstC and the previously described locus sorA. Overall, our data define MstC/SorA as a high-affinity glucose transporter expressed in germinating conidia, and MstA as a high-affinity glucose transporter that operates in vegetative hyphae under conditions of carbon limitation. PMID:24751997

  17. Co-cultivation of Aspergillus nidulans Recombinant Strains Produces an Enzymatic Cocktail as Alternative to Alkaline Sugarcane Bagasse Pretreatment

    PubMed Central

    Lima, Matheus S.; Damasio, André R. de L.; Crnkovic, Paula M.; Pinto, Marcelo R.; da Silva, Ana M.; da Silva, Jean C. R.; Segato, Fernando; de Lucas, Rosymar C.; Jorge, João A.; Polizeli, Maria de L. T. de M.

    2016-01-01

    Plant materials represent a strategic energy source because they can give rise to sustainable biofuels through the fermentation of their carbohydrates. A clear example of a plant-derived biofuel resource is the sugar cane bagasse exhibiting 60–80% of fermentable sugars in its composition. However, the current methods of plant bioconversion employ severe and harmful chemical/physical pretreatments raising biofuel cost production and environmental degradation. Replacing these methods with co-cultivated enzymatic cocktails is an alternative. Here we propose a pretreatment for sugarcane bagasse using a multi-enzymatic cocktail from the co-cultivation of four Aspergillus nidulans recombinant strains. The co-cultivation resulted in the simultaneous production of GH51 arabinofuranosidase (AbfA), GH11 endo-1,4-xylanase (XlnA), GH43 endo-1,5-arabinanase (AbnA) and GH12 xyloglucan specific endo-β-1,4-glucanase (XegA). This core set of recombinant enzymes was more efficient than the alternative alkaline method in maintaining the cellulose integrity and exposing this cellulose to the following saccharification process. Thermogravimetric and differential thermal analysis revealed residual byproducts on the alkali pretreated biomass, which were not found in the enzymatic pretreatment. Therefore, the enzymatic pretreatment was residue-free and seemed to be more efficient than the applied alkaline method, which makes it suitable for bioethanol production. PMID:27199917

  18. Characterization of an inducible expression system in Aspergillus nidulans using alcA and tubulin-coding genes.

    PubMed

    Waring, R B; May, G S; Morris, N R

    1989-06-30

    Plasmids have been constructed in which expression of a gene can be placed under the control of the inducible promoter of the alcA gene encoding alcohol dehydrogenase I in Aspergillus nidulans. Simplified shuttle vectors carrying pyr4 which complements pyrG89 mutations have also been constructed. These are based on pUC19 and retain alpha-peptide expression. The beta-tubulin genes, tubC and benA, have been placed under the control of alcA and their expression studied. Levels of expression can be assayed phenotypically because increased synthesis of beta-tubulin inhibits vegetative growth. Sensitivity of asexual spore formation to the anti-microtubule drug benomyl provides a means of detecting very low levels of expression of the chimeric genes. Glucose almost completely represses the chimeric genes. Induction is rapid and is maximal within an hour. When a strain carrying seven copies of an alcA::tubC gene fusion was grown under inducing conditions, 6.5% of total sulfate labelled protein consisted of tubC product. Cyclopentanone was the most potent inducer of the chimeric genes on solid media but it also partially inhibited growth. Chimeric alcA::tubC and alcA::benA genes were expressed to very similar levels despite the fact that tubC utilizes many rare codons.

  19. An Aspergillus nidulans GH26 endo-β-mannanase with a novel degradation pattern on highly substituted galactomannans.

    PubMed

    von Freiesleben, Pernille; Spodsberg, Nikolaj; Blicher, Thomas Holberg; Anderson, Lars; Jørgensen, Henning; Stålbrand, Henrik; Meyer, Anne S; Krogh, Kristian B R M

    2016-02-01

    The activity and substrate degradation pattern of a novel Aspergillus nidulans GH26 endo-β-mannanase (AnMan26A) was investigated using two galactomannan substrates with varying amounts of galactopyranosyl residues. The AnMan26A was characterized in parallel with the GH26 endomannanase from Podospora anserina (PaMan26A) and three GH5 endomannanases from A. nidulans and Trichoderma reesei (AnMan5A, AnMan5C and TrMan5A). The initial rates and the maximal degree of enzymatically catalyzed conversion of locust bean gum and guar gum galactomannans were determined. The hydrolysis product profile at maximal degree of conversion was determined using DNA sequencer-Assisted Saccharide analysis in High throughput (DASH). This is the first reported use of this method for analyzing galactomannooligosaccharides. AnMan26A and PaMan26A were found to have a novel substrate degradation pattern on the two galactomannan substrates. On the highly substituted guar gum AnMan26A and PaMan26A reached 35-40% as their maximal degree of conversion whereas the three tested GH5 endomannanases only reached 8-10% as their maximal degree of conversion. α-Galactosyl-mannose was identified as the dominant degradation product resulting from AnMan26A and PaMan26A action on guar gum, strongly indicating that these two enzymes can accommodate galactopyranosyl residues in the -1 and in the +1 subsite. The degradation of α-6(4)-6(3)-di-galactosyl-mannopentaose by AnMan26A revealed accommodation of galactopyranosyl residues in the -2, -1 and +1 subsite of the enzyme. Accommodation of galactopyranosyl residues in subsites -2 and +1 has not been observed for other characterized endomannanases to date. Docking analysis of galactomannooligosaccharides in available crystal structures and homology models supported the conclusions drawn from the experimental results. This newly discovered diversity of substrate degradation patterns demonstrates an expanded functionality of fungal endomannanases, than hitherto

  20. An Aspergillus nidulans GH26 endo-β-mannanase with a novel degradation pattern on highly substituted galactomannans.

    PubMed

    von Freiesleben, Pernille; Spodsberg, Nikolaj; Blicher, Thomas Holberg; Anderson, Lars; Jørgensen, Henning; Stålbrand, Henrik; Meyer, Anne S; Krogh, Kristian B R M

    2016-02-01

    The activity and substrate degradation pattern of a novel Aspergillus nidulans GH26 endo-β-mannanase (AnMan26A) was investigated using two galactomannan substrates with varying amounts of galactopyranosyl residues. The AnMan26A was characterized in parallel with the GH26 endomannanase from Podospora anserina (PaMan26A) and three GH5 endomannanases from A. nidulans and Trichoderma reesei (AnMan5A, AnMan5C and TrMan5A). The initial rates and the maximal degree of enzymatically catalyzed conversion of locust bean gum and guar gum galactomannans were determined. The hydrolysis product profile at maximal degree of conversion was determined using DNA sequencer-Assisted Saccharide analysis in High throughput (DASH). This is the first reported use of this method for analyzing galactomannooligosaccharides. AnMan26A and PaMan26A were found to have a novel substrate degradation pattern on the two galactomannan substrates. On the highly substituted guar gum AnMan26A and PaMan26A reached 35-40% as their maximal degree of conversion whereas the three tested GH5 endomannanases only reached 8-10% as their maximal degree of conversion. α-Galactosyl-mannose was identified as the dominant degradation product resulting from AnMan26A and PaMan26A action on guar gum, strongly indicating that these two enzymes can accommodate galactopyranosyl residues in the -1 and in the +1 subsite. The degradation of α-6(4)-6(3)-di-galactosyl-mannopentaose by AnMan26A revealed accommodation of galactopyranosyl residues in the -2, -1 and +1 subsite of the enzyme. Accommodation of galactopyranosyl residues in subsites -2 and +1 has not been observed for other characterized endomannanases to date. Docking analysis of galactomannooligosaccharides in available crystal structures and homology models supported the conclusions drawn from the experimental results. This newly discovered diversity of substrate degradation patterns demonstrates an expanded functionality of fungal endomannanases, than hitherto

  1. Immobilization and Biochemical Properties of the Enantioselective Recombinant NStcI Esterase of Aspergillus nidulans

    PubMed Central

    Peña-Montes, Carolina; Mondragón-Tintor, María Elena; Castro-Rodríguez, José Augusto; Bustos-Jaimes, Ismael; Navarro-Ocaña, Arturo; Farrés, Amelia

    2013-01-01

    The recombinant NStcI A. nidulans esterase was adsorbed on Accurel MP1000, where protein yield and immobilization efficiency were 42.48% and 81.94%, respectively. Storage stability test at 4°C and RT showed 100% of residual activity after 40 days at both temperatures. The biocatalyst retains more than 70% of its initial activity after 3 cycles of repeated use. Biochemical properties of this new biocatalyst were obtained. Maximum activity was achieved at pH 11 and 30°C, while the best stability was observed with the pH between 9 and 11 at 40°C. NStcI thermostability was increased after immobilization, as it retained 47.5% of its initial activity after 1 h at 60°C, while the free enzyme under the same conditions displayed no activity. NStcI preserved 70% of its initial activity in 100% hexane after 72 h. Enzymatic kinetic resolution of (R,S)-1-phenylethanol was chosen as model reaction, using vinyl acetate as acyl donor. After optimization of reaction parameters, the highest possible conversion (42%) was reached at 37°C, aw of 0.07, and 120 h of bioconversion in hexane with an enantiomeric excess of 71.7%. NStcI has selectivity for (R)-enantiomer. The obtained E value (31.3) is in the range considered useful to resolve enantiomeric mixtures. PMID:23781330

  2. The Set1/COMPASS histone H3 methyltransferase helps regulate mitosis with the CDK1 and NIMA mitotic kinases in Aspergillus nidulans.

    PubMed

    Govindaraghavan, Meera; Anglin, Sarah Lea; Osmani, Aysha H; Osmani, Stephen A

    2014-08-01

    Mitosis is promoted and regulated by reversible protein phosphorylation catalyzed by the essential NIMA and CDK1 kinases in the model filamentous fungus Aspergillus nidulans. Protein methylation mediated by the Set1/COMPASS methyltransferase complex has also been shown to regulate mitosis in budding yeast with the Aurora mitotic kinase. We uncover a genetic interaction between An-swd1, which encodes a subunit of the Set1 protein methyltransferase complex, with NIMA as partial inactivation of nimA is poorly tolerated in the absence of swd1. This genetic interaction is additionally seen without the Set1 methyltransferase catalytic subunit. Importantly partial inactivation of NIMT, a mitotic activator of the CDK1 kinase, also causes lethality in the absence of Set1 function, revealing a functional relationship between the Set1 complex and two pivotal mitotic kinases. The main target for Set1-mediated methylation is histone H3K4. Mutational analysis of histone H3 revealed that modifying the H3K4 target residue of Set1 methyltransferase activity phenocopied the lethality seen when either NIMA or CDK1 are partially functional. We probed the mechanistic basis of these genetic interactions and find that the Set1 complex performs functions with CDK1 for initiating mitosis and with NIMA during progression through mitosis. The studies uncover a joint requirement for the Set1 methyltransferase complex with the CDK1 and NIMA kinases for successful mitosis. The findings extend the roles of the Set1 complex to include the initiation of mitosis with CDK1 and mitotic progression with NIMA in addition to its previously identified interactions with Aurora and type 1 phosphatase in budding yeast.

  3. Effect of cell wall integrity stress and RlmA transcription factor on asexual development and autolysis in Aspergillus nidulans.

    PubMed

    Kovács, Zsuzsanna; Szarka, Máté; Kovács, Szilvia; Boczonádi, Imre; Emri, Tamás; Abe, Keietsu; Pócsi, István; Pusztahelyi, Tünde

    2013-05-01

    The cell wall integrity (CWI) signaling pathway is responsible for cell wall remodeling and reinforcement upon cell wall stress, which is proposed to be universal in fungal cultures. In Aspergillus nidulans, both the deletion of rlmA encoding the RlmA transcription factor in CWI signaling and low concentrations of the cell wall polymer intercalating agent Congo Red caused significant physiological changes. The gene deletion mutant ΔrlmA strain showed decreased CWI and oxidative stress resistances, which indicated the connection between the CWI pathway and the oxidative stress response system. The Congo Red stress resulted in alterations in the cell wall polymer composition in submerged cultures due to the induction of the biosynthesis of the alkali soluble fraction as well as the hydrolysis of cell wall biopolymers. Both RlmA and RlmA-independent factors induced by Congo Red stress regulated the expression of glucanase (ANID_00245, engA) and chitinase (chiB, chiA) genes, which promoted the autolysis of the cultures and also modulated the pellet sizes. CWI stress and rlmA deletion affected the expression of brlA encoding the early conidiophore development regulator transcription factor BrlA and, as a consequence, the formation of conidiophores was significantly changed in submerged cultures. Interestingly, the number of conidiospores increased in surface cultures of the ΔrlmA strain. The in silico analysis of genes putatively regulated by RlmA and the CWI transcription factors AnSwi4/AnSwi6 in the SBF complex revealed only a few jointly regulated genes, including ugmA and srrA coding for UgmA UDP-galactopyranose mutase and SrrA stress response regulator, respectively. PMID:23485399

  4. A mutation in the converter subdomain of Aspergillus nidulans MyoB blocks constriction of the actomyosin ring in cytokinesis.

    PubMed

    Hill, Terry W; Jackson-Hayes, Loretta; Wang, Xiao; Hoge, Brianna L

    2015-02-01

    We have identified a mutant allele of the Aspergillus nidulans homologue of myosin II (myoB; AN4706), which prevents normal septum formation. This is the first reported myosin II mutation in a filamentous fungus. Strains expressing the myoB(G843D) allele produce mainly aberrant septa at 30 °C and are completely aseptate at temperatures above 37 °C. Conidium formation is greatly reduced at 30 °C and progressively impaired with increasing temperature. Sequencing of the myoB(G843D) allele identified a point mutation predicted to result in a glycine-to-aspartate amino acid substitution at residue 843 in the myosin II converter domain. This residue is conserved in all fungal, plant, and animal myosin sequences that we have examined. The mutation does not prevent localization of the myoB(G843D) gene product to contractile rings, but it does block ring constriction. MyoB(G843D) rings at sites of abortive septation disassemble after an extended period and dissipate into the cytoplasm. During contractile ring formation, both wild type and mutant MyoB::GFP colocalize with actin--an association that begins at the pre-ring "string" stage. Down-regulation of wild-type myoB expression under control of the alcA promoter blocks septation but does not prevent actin from aggregating at putative septation sites--the actin rings, however, do not fully coalesce. Both septation and targeting of MyoB are blocked by disruption of filamentous actin using latrunculin B. We propose a model in which myosin assembly at septation sites depends upon the presence of F-actin, but assembly of the actin component of contractile rings depends upon normal levels of myosin only for the final stages of ring compaction.

  5. The Aspergillus nidulans homoaconitase gene lysF is negatively regulated by the multimeric CCAAT-binding complex AnCF and positively regulated by GATA sites.

    PubMed

    Weidner, G; Steidl, S; Brakhage, A A

    2001-02-01

    In beta-lactam-antibiotic-producing fungi, such as Aspergillus (Emericella) nidulans, L-alpha-aminoadipic acid is the branching point of the lysine and penicillin biosynthesis pathways. To obtain a deeper insight into the regulation of lysine biosynthesis genes, the regulation of the A. nidulans lysF gene, which encodes homoaconitase, was studied. Band-shift assays indicated that the A. nidulans multimeric CCAAT-binding complex AnCF binds to two of four CCAAT motifs present in the lysF promoter region. AnCF consists at least of three different subunits, designated HapB, HapC, and HapE. In both a delta hapB and a delta hapC strain, the expression of a translational lysF-lacZ gene fusion integrated in single copy at the chromosomal argB gene locus was two to three-fold higher than in a wild-type strain. These data show that AnCF negatively regulates lysF expression. The results of Northern blot analysis and lysF-lacZ expression analysis did not indicate a lysine-dependent repression of lysF expression. Furthermore, mutational analysis of the lysF promoter region revealed that two GATA sites matching the GATA consensus sequence HGATAR positively affected lysF-lacZ expression. Results of Northern blot analysis also excluded that the global nitrogen regulator AreA is the responsible trans-acting GATA-binding factor.

  6. Aspergillus nidulans alpha-galactosidase of glycoside hydrolase family 36 catalyses the formation of alpha-galacto-oligosaccharides by transglycosylation.

    PubMed

    Nakai, Hiroyuki; Baumann, Martin J; Petersen, Bent O; Westphal, Yvonne; Hachem, Maher Abou; Dilokpimol, Adiphol; Duus, Jens Ø; Schols, Henk A; Svensson, Birte

    2010-09-01

    The alpha-galactosidase from Aspergillus nidulans (AglC) belongs to a phylogenetic cluster containing eukaryotic alpha-galactosidases and alpha-galacto-oligosaccharide synthases of glycoside hydrolase family 36 (GH36). The recombinant AglC, produced in high yield (0.65 g.L(-1) culture) as His-tag fusion in Escherichia coli, catalysed efficient transglycosylation with alpha-(1-->6) regioselectivity from 40 mm 4-nitrophenol alpha-d-galactopyranoside, melibiose or raffinose, resulting in a 37-74% yield of 4-nitrophenol alpha-D-Galp-(1-->6)-D-Galp, alpha-D-Galp-(1-->6)-alpha-D-Galp-(1-->6)-D-Glcp and alpha-D-Galp-(1-->6)-alpha-D-Galp-(1-->6)-D-Glcp-(alpha1-->beta2)-d-Fruf (stachyose), respectively. Furthermore, among 10 monosaccharide acceptor candidates (400 mm) and the donor 4-nitrophenol alpha-D-galactopyranoside (40 mm), alpha-(1-->6) linked galactodisaccharides were also obtained with galactose, glucose and mannose in high yields of 39-58%. AglC did not transglycosylate monosaccharides without the 6-hydroxymethyl group, i.e. xylose, L-arabinose, L-fucose and L-rhamnose, or with axial 3-OH, i.e. gulose, allose, altrose and L-rhamnose. Structural modelling using Thermotoga maritima GH36 alpha-galactosidase as the template and superimposition of melibiose from the complex with human GH27 alpha-galactosidase supported that recognition at subsite +1 in AglC presumably requires a hydrogen bond between 3-OH and Trp358 and a hydrophobic environment around the C-6 hydroxymethyl group. In addition, successful transglycosylation of eight of 10 disaccharides (400 mm), except xylobiose and arabinobiose, indicated broad specificity for interaction with the +2 subsite. AglC thus transferred alpha-galactosyl to 6-OH of the terminal residue in the alpha-linked melibiose, maltose, trehalose, sucrose and turanose in 6-46% yield and the beta-linked lactose, lactulose and cellobiose in 28-38% yield. The product structures were identified using NMR and ESI-MS and five of the 13

  7. Versatile enzyme expression and characterization system for Aspergillus nidulans, with the Penicillium brevicompactum polyketide synthase gene from the mycophenolic acid gene cluster as a test case.

    PubMed

    Hansen, Bjarne G; Salomonsen, Bo; Nielsen, Morten T; Nielsen, Jakob B; Hansen, Niels B; Nielsen, Kristian F; Regueira, Torsten B; Nielsen, Jens; Patil, Kiran R; Mortensen, Uffe H

    2011-05-01

    Assigning functions to newly discovered genes constitutes one of the major challenges en route to fully exploiting the data becoming available from the genome sequencing initiatives. Heterologous expression in an appropriate host is central in functional genomics studies. In this context, filamentous fungi offer many advantages over bacterial and yeast systems. To facilitate the use of filamentous fungi in functional genomics, we present a versatile cloning system that allows a gene of interest to be expressed from a defined genomic location of Aspergillus nidulans. By a single USER cloning step, genes are easily inserted into a combined targeting-expression cassette ready for rapid integration and analysis. The system comprises a vector set that allows genes to be expressed either from the constitutive PgpdA promoter or from the inducible PalcA promoter. Moreover, by using the vector set, protein variants can easily be made and expressed from the same locus, which is mandatory for proper comparative analyses. Lastly, all individual elements of the vectors can easily be substituted for other similar elements, ensuring the flexibility of the system. We have demonstrated the potential of the system by transferring the 7,745-bp large mpaC gene from Penicillium brevicompactum to A. nidulans. In parallel, we produced defined mutant derivatives of mpaC, and the combined analysis of A. nidulans strains expressing mpaC or mutated mpaC genes unequivocally demonstrated that mpaC indeed encodes a polyketide synthase that produces the first intermediate in the production of the medically important immunosuppressant mycophenolic acid. PMID:21398493

  8. Putative PmrA and PmcA are important for normal growth, morphogenesis and cell wall integrity, but not for viability in Aspergillus nidulans.

    PubMed

    Jiang, Hechun; Liu, Feifei; Zhang, Shizhu; Lu, Ling

    2014-11-01

    P-type Ca(2+)-transporting ATPases are Ca(2+) pumps, extruding cytosolic Ca(2+) to the extracellular environment or the intracellular Ca(2+) store lumens. In budding yeast, Pmr1 (plasma membrane ATPase related), and Pmc1 (plasma membrane calcium-ATPase) cannot be deleted simultaneously for it to survive in standard medium. Here, we deleted two putative Ca(2+) pumps, designated AnPmrA and AnPmcA, from Aspergillus nidulans, and obtained the mutants ΔanpmrA and ΔanpmcA, respectively. Then, using ΔanpmrA as the starting strain, the promoter of its anpmcA was replaced with the alcA promoter to secure the mutant ΔanpmrAalcApmcA or its anpmcA was deleted completely to produce the mutant ΔanpmrAΔpmcA. Different from the case in Saccharomyces cerevisiae, double deletion of anpmrA and anpmcA was not lethal in A. nidulans. In addition, deletion of anpmrA and/or anpmcA had produced growth defects, although overexpression of AnPmc1 in ΔanpmrAalcApmcA could not restore the growth defects that resulted from the loss of AnPmrA. Moreover, we found AnPmrA was indispensable for maintenance of normal morphogenesis, especially in low-Ca(2+)/Mn(2+) environments. Thus, our findings suggest AnPmrA and AnPmcA might play important roles in growth, morphogenesis and cell wall integrity in A. nidulans in a different way from that in yeasts.

  9. Novel phacB-Encoded Cytochrome P450 Monooxygenase from Aspergillus nidulans with 3-Hydroxyphenylacetate 6-Hydroxylase and 3,4-Dihydroxyphenylacetate 6-Hydroxylase Activities▿

    PubMed Central

    Ferrer-Sevillano, Francisco; Fernández-Cañón, José M.

    2007-01-01

    Aspergillus nidulans catabolizes phenylacetate (PhAc) and 3-hydroxy-, 4-hydroxy-, and 3,4-dihydroxyphenylacetate (3-OH-PhAc, 4-OH-PhAc, and 3,4-diOH-PhAc, respectively) through the 2,5-dihydroxyphenylacetate (homogentisic acid) catabolic pathway. Using cDNA subtraction techniques, we isolated a gene, denoted phacB, which is strongly induced by PhAc (and its hydroxyderivatives) and encodes a new cytochrome P450 (CYP450). A disrupted phacB strain (ΔphacB) does not grow on 3-hydroxy-, 4-hydroxy-, or 3,4-dihydroxy-PhAc. High-performance liquid chromatography and gas chromatography-mass spectrum analyses of in vitro reactions using microsomes from wild-type and several A. nidulans mutant strains confirmed that the phacB-encoded CYP450 catalyzes 3-hydroxyphenylacetate and 3,4-dihydroxyphenylacetate 6-hydroxylations to generate 2,5-dihydroxyphenylacetate and 2,4,5-trihydroxyphenylacetate, respectively. Both of these compounds are used as substrates by homogentisate dioxygenase. This cytochrome P450 protein also uses PhAc as a substrate to generate 2-OH-PhAc with a very low efficiency. The phacB gene is the first member of a new CYP450 subfamily (CYP504B). PMID:17189487

  10. The Aspergillus nidulans metZ gene encodes a transcription factor involved in regulation of sulfur metabolism in this fungus and other Eurotiales.

    PubMed

    Piłsyk, Sebastian; Natorff, Renata; Sieńko, Marzena; Skoneczny, Marek; Paszewski, Andrzej; Brzywczy, Jerzy

    2015-05-01

    In Aspergillus nidulans, expression of sulfur metabolism genes is activated by the MetR transcription factor containing a basic region and leucine zipper domain (bZIP). Here we identified and characterized MetZ, a new transcriptional regulator in A. nidulans and other Eurotiales. It contains a bZIP domain similar to the corresponding region in MetR and this similarity suggests that MetZ could potentially complement the MetR deficiency. The metR and metZ genes are interrupted by unusually long introns. Transcription of metZ, unlike that of metR, is controlled by the sulfur metabolite repression system (SMR) dependent on the MetR protein. Overexpression of metZ from a MetR-independent promoter in a ΔmetR background activates transcription of genes encoding sulfate permease, homocysteine synthase and methionine permease, partially complementing the phenotype of the ΔmetR mutation. Thus, MetZ appears to be a second transcription factor involved in regulation of sulfur metabolism genes.

  11. Enhancement of Echinocandin B Production by a UV- and Microwave-Induced Mutant of Aspergillus nidulans with Precursor- and Biotin-Supplying Strategy.

    PubMed

    Hu, Zhong-Ce; Peng, Li-Yuan; Zheng, Yu-Guo

    2016-08-01

    Echinocandin B belongs to lipopeptide antifungal antibiotic bearing five types of direct precursor amino acids including proline, ornithine, tyrosine, threonine, and leucine. The objective of this study is to screen over-producing mutant in order to improve echinocandin B production; a stable mutant Aspergillus nidulans ZJB12073, which can use fructose as optimal carbon source instead of expensive mannitol, was selected from thousand isolates after several cycles of UV and microwave irradiation in turn. The results showed that mutant strain ZJB12073 exhibited 1.9-fold improvement in echinocandin B production to 1656.3 ± 40.3 mg/L when compared with the parent strain. Furthermore, the effects of precursor amino acids and some chemicals on echinocandin B biosynthesis in A. nidulans were investigated, respectively. Tyrosine, leucine, and biotin were selected as key factors to optimize the medium employing uniform design method. The results showed that the optimized fermentation medium provided another 63.1 % increase to 2701.6 ± 31.7 mg/L in final echinocandin B concentration compared to that of unoptimized medium.

  12. An ortholog of farA of Aspergillus nidulans is implicated in the transcriptional activation of genes involved in fatty acid utilization in the yeast Yarrowia lipolytica

    SciTech Connect

    Poopanitpan, Napapol; Kobayashi, Satoshi; Fukuda, Ryouichi; Horiuchi, Hiroyuki; Ohta, Akinori

    2010-11-26

    Research highlights: {yields} POR1 is a Yarrowia lipolytica ortholog of farA involved in fatty acid response in A. nidulans. {yields} Deletion of POR1 caused growth defects on fatty acids. {yields} {Delta}por1 strain exhibited defects in the induction of genes involved in fatty acid utilization. -- Abstract: The yeast Yarrowia lipolytica effectively utilizes hydrophobic substrates such as fatty acids and n-alkanes. To identify a gene(s) regulating fatty acid utilization in Y. lipolytica, we first studied homologous genes to OAF1 and PIP2 of Saccharomyces cerevisiae, but their disruption did not change growth on oleic acid at all. We next characterized a Y. lipolytica gene, POR1 (primary oleate regulator 1), an ortholog of farA encoding a transcriptional activator that regulates fatty acid utilization in Aspergillus nidulans. The deletion mutant of POR1 was defective in the growth on various fatty acids, but not on glucose, glycerol, or n-hexadecane. It exhibited slight defect on n-decane. The transcriptional induction of genes involved in {beta}-oxidation and peroxisome proliferation by oleate was distinctly diminished in the {Delta}por1 strains. These data suggest that POR1 encodes a transcriptional activator widely regulating fatty acid metabolism in Y. lipolytica.

  13. The Aspergillus nidulans acuL gene encodes a mitochondrial carrier required for the utilization of carbon sources that are metabolized via the TCA cycle.

    PubMed

    Flipphi, Michel; Oestreicher, Nathalie; Nicolas, Valérie; Guitton, Audrey; Vélot, Christian

    2014-07-01

    In Aspergillus nidulans, the utilization of acetate as sole carbon source requires several genes (acu). Most of them are also required for the utilization of fatty acids. This is the case for acuD and acuE, which encode the two glyoxylate cycle-specific enzymes, isocitrate lyase and malate synthase, respectively, but also for acuL that we have identified as AN7287, and characterized in this study. Deletion of acuL resulted in the same phenotype as the original acuL217 mutant. acuL encodes a 322-amino acid protein which displays all the structural features of a mitochondrial membrane carrier, and shares 60% identity with the Saccharomyces cerevisiae succinate/fumarate mitochondrial antiporter Sfc1p (also named Acr1p). Consistently, the AcuL protein was shown to localize in mitochondria, and partial cross-complementation was observed between the S. cerevisiae and A. nidulans homologues. Extensive phenotypic characterization suggested that the acuL gene is involved in the utilization of carbon sources that are catabolized via the TCA cycle, and therefore require gluconeogenesis. In addition, acuL proves to be co-regulated with acuD and acuE. Overall, our data suggest that AcuL could link the glyoxylate cycle to gluconeogenesis by exchanging cytoplasmic succinate for mitochondrial fumarate.

  14. Mutational Analysis of AREA, a Transcriptional Activator Mediating Nitrogen Metabolite Repression in Aspergillus nidulans and a Member of the “Streetwise” GATA Family of Transcription Factors

    PubMed Central

    Wilson, Richard A.; Arst, Herbert N.

    1998-01-01

    Summary: The transcriptional activator AREA is a member of the GATA family of transcription factors and mediates nitrogen metabolite repression in the fungus Aspergillus nidulans. The nutritional versatility of A. nidulans and its amenability to classical and reverse genetic manipulations make the AREA DNA binding domain (DBD) a useful model for analyzing GATA family DBDs, particularly as structures of two AREA-DNA complexes have been determined. The 109 extant mutant forms of the AREA DBD surveyed here constitute one of the highest totals of eukaryotic transcription factor DBD mutants, are discussed in light of the roles of individual residues, and are compared to corresponding mutant sequence changes in other fungal GATA factor DBDs. Other topics include delineation of the DBD using both homology and mutational truncation, use of frameshift reversion to detect regions of tolerance to mutational change, the finding that duplication of the DBD can apparently enhance AREA function, and use of the AREA system to analyze a vertebrate GATA factor DBD. Some major points to emerge from work on the AREA DBD are (i) tolerance to sequence change (with retention of function) is surprisingly great, (ii) mutational changes in a transcription factor can have widely differing, even opposing, effects on expression of different structural genes so that monitoring expression of one or even several structural genes can be insufficient and possibly misleading, and (iii) a mutational change altering local hydrophobic packing and DNA binding target specificity can markedly influence the behavior of mutational changes elsewhere in the DBD. PMID:9729601

  15. veA-dependent RNA-pol II transcription elongation factor like protein, RtfA, is associated with secondary metabolism and morphological development in Aspergillus nidulans

    PubMed Central

    Ramamoorthy, Vellaisamy; Shantappa, Sourabha; Dhingra, Sourabh; Calvo, Ana M.

    2012-01-01

    In Aspergillus nidulans the global regulatory gene veA is necessary for the biosynthesis of several secondary metabolites, including the mycotoxin sterigmatocystin (ST). In order to identify additional veA-dependent genetic elements involved in regulating ST production, we performed a mutagenesis on a deletion veA (ΔveA) strain to obtain revertant mutants (RM) that regained the capability to produce toxin. Genetic analysis and molecular characterization of one of the revertant mutants, RM3, revealed that a point mutation occurred at the coding region of the rtfA gene, encoding a RNA-pol II transcription elongation factor like protein, similar to Saccharomyces cerevisiae Rtf1. The A. nidulans rtfA gene product accumulates in nuclei. Deletion of rtfA gene in a ΔveA background restored mycotoxin production in a medium-dependent manner. rtfA also affects the production of other metabolites including penicillin. Biosynthesis of this antibiotic decreased in the absence of rtfA. Furthermore, rtfA is necessary for normal morphological development. Deletion of the rtfA gene in wild-type strains (veA+) resulted in a slight decrease in growth rate, drastic reduction in conidiation, and complete loss of sexual development. This is the first study of an Rtf1 like gene in filamentous fungi. We found rtfA putative orthologs extensively conserved in numerous fungal species. PMID:22783880

  16. The selective expression of carbonic anhydrase genes of Aspergillus nidulans in response to changes in mineral nutrition and CO2 concentration.

    PubMed

    Xiao, Leilei; Lian, Bin; Dong, Cuiling; Liu, Fanghua

    2016-02-01

    Carbonic anhydrase (CA) plays an important role in the formation and evolution of life. However, to our knowledge, there has been no report on CA isoenzyme function differentiation in fungi. Two different CA gene sequences in Aspergillus nidulans with clear genetic background provide us a favorable basis for studying function differentiation of CA isoenzymes. Heterologously expressed CA1 was used to test its weathering ability on silicate minerals and real-time quantitative PCR was used to detect expression of the CA1 and CA2 genes at different CO2 concentrations and in the presence of different potassium sources. The northern blot method was applied to confirm the result of CA1 gene expression. Heterologously expressed CA1 significantly promoted dissolution of biotite and wollastonite, and CA1 gene expression increased significantly in response to soluble K-deficiency. The northern blot test further showed that CA1 participated in K-feldspar weathering. In addition, the results showed that CA2 was primary involved in adapting to CO2 concentration change. Taken together, A. nidulans can choose different CA to meet their survival needs, which imply that some environmental microbes have evolved different CAs to adapt to changes in CO2 concentration and acquire mineral nutrition so that they can better adapt to environmental changes. Inversely, their adaption may impact mineral weathering and/or CO2 concentration, and even global change.

  17. A protein kinase C-encoding gene, pkcA, is essential to the viability of the filamentous fungus Aspergillus nidulans.

    PubMed

    Ichinomiya, Masayuki; Uchida, Hirotaka; Koshi, Yukako; Ohta, Akinori; Horiuchi, Hiroyuki

    2007-11-01

    A protein kinase C (PKC)-encoding gene (pkcA) was isolated from the filamentous fungus Aspergillus nidulans. Although we attempted to isolate pkcA deletion mutants, we obtained only heterokaryons that had both DeltapkcA and pkcA(+) nuclei. Conidia produced by the heterokaryon germinated. The germ tubes, however, lysed frequently and no colony formation was observed, indicating that the pkcA gene is essential to the viability of A. nidulans. We constructed conditional mutants (alcA(p)-pkcA mutants) that expressed pkcA under the control of the alcA promoter (alcA(p)). Under alcA(p)-repressing conditions, their colonies were smaller than those of the wild-type strains and their hyphae lysed frequently. These phenotypes were not remedied under moderate- or high-osmolarity conditions; the growth defect deteriorated further under the latter. Under alcA(p)-inducing conditions, the alcA(p)-pkcA mutants also showed growth-sensitivity to cell wall destabilizing agents. These results indicate that pkcA plays an important role in the maintenance of cell integrity.

  18. Depletion of Aspergillus nidulans cotA causes a severe polarity defect which is not suppressed by the nuclear migration mutation nudA2.

    PubMed

    Johns, Sarah Anne; Leeder, Abigail Claire; Safaie, Mehran; Turner, Geoffrey

    2006-06-01

    The Aspergillus nidulans homologue of Neurospora crassa cot-1, cotA, encoding a member of the NDR protein kinase family, has been cloned and expressed under the control of the conditional alcA promoter. Depletion of CotA by repression of the alcA promoter led to a severe growth defect accompanied by loss of polarity. Germlings show greatly enlarged volume of the spores and hyphae, accompanied by an increase in number of nuclei per compartment, though the nucleus/volume ratio is not significantly altered. The depleted CotA phenotype was not suppressed by a nuclear migration mutation nudA2. Double mutants showed an additive, defective phenotype, unlike the suppression of the cot-1 ts mutation by ropy mutations seen in N. crassa, suggesting a different relationship between nuclear migration and the cot signalling pathway in A. nidulans. A functional CotA-GFP fusion protein was found in punctate regions of fluorescence similar to the distribution reported for human NDR2, and as a cap at the hyphal tip.

  19. The Aspergillus nidulans CCAAT-binding factor AnCP/AnCF is a heteromeric protein analogous to the HAP complex of Saccharomyces cerevisiae.

    PubMed

    Kato, M; Aoyama, A; Naruse, F; Tateyama, Y; Hayashi, K; Miyazaki, M; Papagiannopoulos, P; Davis, M A; Hynes, M J; Kobayashi, T; Tsukagoshi, N

    1998-02-01

    The Aspergillus nidulans hapC gene was expressed as a fusion protein with MalE or glutathione-S-transferase (GST) in Escherichia coli, and used for the purification of HapC and the preparation of anti-HapC antiserum. The CCAAT-binding factor AnCP/AnCF contains a component with an approximate molecular mass of 32 kDa that cross-reacts with the antibody. The MalE-HapC fusion protein was able to replace authentic HapC in AnCP when incubated under appropriate conditions. Furthermore, reconstitution experiments with recombinant HapC, yHAP2 and yHAP5 polypeptides showed that all three polypeptides were required for the assembly of a complex capable of binding to CCAAT-containing taaG2 promoter DNA. The relationship between AnCP/AnCF and the Saccharomyces cerevisiae HAP complex is discussed.

  20. Restraint of the G2/M transition by the SR/RRM family mRNA shuttling binding protein SNXAHRB1 in Aspergillus nidulans.

    PubMed

    James, Steven W; Banta, Travis; Barra, James; Ciraku, Lorela; Coile, Clifford; Cuda, Zach; Day, Ryan; Dixit, Cheshil; Eastlack, Steven; Giang, Anh; Goode, James; Guice, Alexis; Huff, Yulon; Humbert, Sara; Kelliher, Christina; Kobie, Julie; Kohlbrenner, Emily; Mwambutsa, Faustin; Orzechowski, Amanda; Shingler, Kristin; Spell, Casey; Anglin, Sarah Lea

    2014-10-01

    Control of the eukaryotic G2/M transition by CDC2/CYCLINB is tightly regulated by protein-protein interactions, protein phosphorylations, and nuclear localization of CDC2/CYCLINB. We previously reported a screen, in Aspergillus nidulans, for extragenic suppressors of nimX2(cdc2) that resulted in the identification of the cold-sensitive snxA1 mutation. We demonstrate here that snxA1 suppresses defects in regulators of the CDK1 mitotic induction pathway, including nimX2(cdc) (2), nimE6(cyclinB), and nimT23(cdc) (25), but does not suppress G2-arresting nimA1/nimA5 mutations, the S-arresting nimE10(cyclinB) mutation, or three other G1/S phase mutations. snxA encodes the A. nidulans homolog of Saccharomyces cerevisiae Hrb1/Gbp2; nonessential shuttling messenger RNA (mRNA)-binding proteins belonging to the serine-arginine-rich (SR) and RNA recognition motif (RRM) protein family; and human heterogeneous ribonucleoprotein-M, a spliceosomal component involved in pre-mRNA processing and alternative splicing. snxA(Hrb) (1) is nonessential, its deletion phenocopies the snxA1 mutation, and its overexpression rescues snxA1 and ΔsnxA mutant phenotypes. snxA1 and a second allele isolated in this study, snxA2, are hypomorphic mutations that result from decreased transcript and protein levels, suggesting that snxA acts normally to restrain cell cycle progression. SNXA(HRB1) is predominantly nuclear, but is not retained in the nucleus during the partially closed mitosis of A. nidulans. We show that the snxA1 mutation does not suppress nimX2 by altering NIMX2(CDC2)/NIME(CYCLINB) kinase activity and that snxA1 or ΔsnxA alter localization patterns of NIME(CYCLINB) at the restrictive temperatures for snxA1 and nimX2. Together, these findings suggest a novel and previously unreported role of an SR/RRM family protein in cell cycle regulation, specifically in control of the CDK1 mitotic induction pathway. PMID:25104516

  1. Restraint of the G2/M Transition by the SR/RRM Family mRNA Shuttling Binding Protein SNXAHRB1 in Aspergillus nidulans

    PubMed Central

    James, Steven W.; Banta, Travis; Barra, James; Ciraku, Lorela; Coile, Clifford; Cuda, Zach; Day, Ryan; Dixit, Cheshil; Eastlack, Steven; Giang, Anh; Goode, James; Guice, Alexis; Huff, Yulon; Humbert, Sara; Kelliher, Christina; Kobie, Julie; Kohlbrenner, Emily; Mwambutsa, Faustin; Orzechowski, Amanda; Shingler, Kristin; Spell, Casey; Anglin, Sarah Lea

    2014-01-01

    Control of the eukaryotic G2/M transition by CDC2/CYCLINB is tightly regulated by protein–protein interactions, protein phosphorylations, and nuclear localization of CDC2/CYCLINB. We previously reported a screen, in Aspergillus nidulans, for extragenic suppressors of nimX2cdc2 that resulted in the identification of the cold-sensitive snxA1 mutation. We demonstrate here that snxA1 suppresses defects in regulators of the CDK1 mitotic induction pathway, including nimX2cdc2, nimE6cyclinB, and nimT23cdc25, but does not suppress G2-arresting nimA1/nimA5 mutations, the S-arresting nimE10cyclinB mutation, or three other G1/S phase mutations. snxA encodes the A. nidulans homolog of Saccharomyces cerevisiae Hrb1/Gbp2; nonessential shuttling messenger RNA (mRNA)-binding proteins belonging to the serine-arginine-rich (SR) and RNA recognition motif (RRM) protein family; and human heterogeneous ribonucleoprotein-M, a spliceosomal component involved in pre-mRNA processing and alternative splicing. snxAHrb1 is nonessential, its deletion phenocopies the snxA1 mutation, and its overexpression rescues snxA1 and ΔsnxA mutant phenotypes. snxA1 and a second allele isolated in this study, snxA2, are hypomorphic mutations that result from decreased transcript and protein levels, suggesting that snxA acts normally to restrain cell cycle progression. SNXAHRB1 is predominantly nuclear, but is not retained in the nucleus during the partially closed mitosis of A. nidulans. We show that the snxA1 mutation does not suppress nimX2 by altering NIMX2CDC2/NIMECYCLINB kinase activity and that snxA1 or ΔsnxA alter localization patterns of NIMECYCLINB at the restrictive temperatures for snxA1 and nimX2. Together, these findings suggest a novel and previously unreported role of an SR/RRM family protein in cell cycle regulation, specifically in control of the CDK1 mitotic induction pathway. PMID:25104516

  2. Arabidopsis and Brachypodium distachyon transgenic plants expressing Aspergillus nidulans acetylesterases have decreased degree of polysaccharide acetylation and increased resistance to pathogens.

    PubMed

    Pogorelko, Gennady; Lionetti, Vincenzo; Fursova, Oksana; Sundaram, Raman M; Qi, Mingsheng; Whitham, Steven A; Bogdanove, Adam J; Bellincampi, Daniela; Zabotina, Olga A

    2013-05-01

    The plant cell wall has many significant structural and physiological roles, but the contributions of the various components to these roles remain unclear. Modification of cell wall properties can affect key agronomic traits such as disease resistance and plant growth. The plant cell wall is composed of diverse polysaccharides often decorated with methyl, acetyl, and feruloyl groups linked to the sugar subunits. In this study, we examined the effect of perturbing cell wall acetylation by making transgenic Arabidopsis (Arabidopsis thaliana) and Brachypodium (Brachypodium distachyon) plants expressing hemicellulose- and pectin-specific fungal acetylesterases. All transgenic plants carried highly expressed active Aspergillus nidulans acetylesterases localized to the apoplast and had significant reduction of cell wall acetylation compared with wild-type plants. Partial deacetylation of polysaccharides caused compensatory up-regulation of three known acetyltransferases and increased polysaccharide accessibility to glycosyl hydrolases. Transgenic plants showed increased resistance to the fungal pathogens Botrytis cinerea and Bipolaris sorokiniana but not to the bacterial pathogens Pseudomonas syringae and Xanthomonas oryzae. These results demonstrate a role, in both monocot and dicot plants, of hemicellulose and pectin acetylation in plant defense against fungal pathogens.

  3. Identification of the novel penicillin biosynthesis gene aatB of Aspergillus nidulans and its putative evolutionary relationship to this fungal secondary metabolism gene cluster.

    PubMed

    Spröte, Petra; Hynes, Michael J; Hortschansky, Peter; Shelest, Ekaterina; Scharf, Daniel H; Wolke, Sandra M; Brakhage, Axel A

    2008-10-01

    The final step of penicillin biosynthesis in the filamentous fungus Aspergillus nidulans is catalysed by isopenicillin N acyltransferase encoded by the aatA gene. Because there is no bacterial homologue, its evolutionary origin remained obscure. As shown here,disruption of aatA still enabled penicillin production. Genome mining led to the discovery of the aatB gene(AN6775.3) which has a similar structure and expression pattern as aatA. Disruption of aatB resulted in a reduced penicillin titre. Surface plasmon resonance analysis and Northern blot analysis indicated that the promoters of both aatA and aatB are bound and regulated by the same transcription factors AnCF and AnBH1f. In contrast to aatA, aatB does not encode a peroxisomal targeting signal (PTS1). Overexpression of a mutated aatB(PTS1) gene in an aatA-disruption strain(leading to peroxisomal localization of AatB)increased the penicillin titre more than overexpression of the wild-type aatB. Homologues of aatA are exclusively part of the penicillin biosynthesis gene cluster,whereas aatB homologues also exist in non-producing fungi. Our findings suggest that aatB is a paralogue of aatA. They extend the model of evolution of the penicillin biosynthesis gene cluster by recruitment of a biosynthesis gene and its cis-regulatory sites upon gene duplication.

  4. The Expression of Sterigmatocystin and Penicillin Genes in Aspergillus nidulans Is Controlled by veA, a Gene Required for Sexual Development

    PubMed Central

    Kato, Naoki; Brooks, Wilhelmina; Calvo, Ana M.

    2003-01-01

    Secondary metabolism is commonly associated with morphological development in microorganisms, including fungi. We found that veA, a gene previously shown to control the Aspergillus nidulans sexual/asexual developmental ratio in response to light, also controls secondary metabolism. Specifically, veA regulates the expression of genes implicated in the synthesis of the mycotoxin sterigmatocystin and the antibiotic penicillin. veA is necessary for the expression of the transcription factor aflR, which activates the gene cluster that leads to the production of sterigmatocystin. veA is also necessary for penicillin production. Our results indicated that although veA represses the transcription of the isopenicillin synthetase gene ipnA, it is necessary for the expression of acvA, the key gene in the first step of penicillin biosynthesis, encoding the delta-(l-alpha-aminoadipyl)-l-cysteinyl-d-valine synthetase. With respect to the mechanism of veA in directing morphological development, veA has little effect on the expression of the known sexual transcription factors nsdD and steA. However, we found that veA regulates the expression of the asexual transcription factor brlA by modulating the α/β transcript ratio that controls conidiation. PMID:14665453

  5. ami1, an orthologue of the Aspergillus nidulans apsA gene, is involved in nuclear migration events throughout the life cycle of Podospora anserina.

    PubMed Central

    Graïa, F; Berteaux-Lecellier, V; Zickler, D; Picard, M

    2000-01-01

    The Podospora anserina ami1-1 mutant was identified as a male-sterile strain. Microconidia (which act as male gametes) form, but are anucleate. Paraphysae from the perithecium beaks are also anucleate when ami1-1 is used as the female partner in a cross. Furthermore, in crosses heterozygous for ami1-1, some crozier cells are uninucleate rather than binucleate. In addition to these nuclear migration defects, which occur at the transition between syncytial and cellular states, ami1-1 causes abnormal distribution of the nuclei in both mycelial filaments and asci. Finally, an ami1-1 strain bearing information for both mating types is unable to self-fertilize. The ami1 gene is an orthologue of the Aspergillus nidulans apsA gene, which controls nuclear positioning in filaments and during conidiogenesis (at the syncytial/cellular transition). The ApsA and AMI1 proteins display 42% identity and share structural features. The apsA gene complements some ami1-1 defects: it increases the percentage of nucleate microconidia and restores self-fertility in an ami1-1 mat+ (mat-) strain. The latter effect is puzzling, since in apsA null mutants sexual reproduction is quite normal. The functional differences between the two genes are discussed with respect to their possible history in these two fungi, which are very distant in terms of evolution. PMID:10835387

  6. Metabolism of D-galactose is dispensable for the induction of the beta-galactosidase (bgaD) and lactose permease (lacpA) genes in Aspergillus nidulans.

    PubMed

    Orosz, Anita; Fekete, Erzsébet; Flipphi, Michel; Karaffa, Levente

    2014-10-01

    In this study, we analyze the expression of the Aspergillus nidulans bgaD-lacpA gene couple (encoding an intracellular beta-galactosidase and a lactose permease) in the presence of D-galactose. This monosaccharide can be catabolized via alternative, independent pathways in this model organism. The inductive capabilities of intermediates of the two alternative routes of D-galactose utilization were addressed in loss-of-function mutants defective in a defined step in one of the two pathways. In a galactokinase (galE9) mutant, the cluster is strongly induced by D-galactose, suggesting that formation of Leloir pathway intermediates is not required. The expression profiles of bgaD and lacpA were similar in wild type, L-arabinitol dehydrogenase (araA1), and hexokinase (hxkA1) negative backgrounds, indicating that intermediates of the oxido-reductive pathway downstream of galactitol are not necessary either. Furthermore, bgaD-lacpA transcription was not induced in any of the tested strains when galactitol was provided as the growth substrate. An hxkA1/galE9 double mutant cannot grow on d-galactose at all, but still produced bgaD and lacpA transcripts upon transfer to d-galactose. We therefore concluded that the physiological inducer of the bgaD-lacpA gene cluster upon growth on D-galactose is the nonmetabolized sugar itself.

  7. A WDR Gene Is a Conserved Member of a Chitin Synthase Gene Cluster and Influences the Cell Wall in Aspergillus nidulans.

    PubMed

    Guerriero, Gea; Silvestrini, Lucia; Obersriebnig, Michael; Hausman, Jean-Francois; Strauss, Joseph; Ezcurra, Inés

    2016-01-01

    WD40 repeat (WDR) proteins are pleiotropic molecular hubs. We identify a WDR gene that is a conserved genomic neighbor of a chitin synthase gene in Ascomycetes. The WDR gene is unique to fungi and plants, and was called Fungal Plant WD (FPWD). FPWD is within a cell wall metabolism gene cluster in the Ascomycetes (Pezizomycotina) comprising chsD, a Chs activator and a GH17 glucanase. The FPWD, AN1556.2 locus was deleted in Aspergillus nidulans strain SAA.111 by gene replacement and only heterokaryon transformants were obtained. The re-annotation of Aspergilli genomes shows that AN1556.2 consists of two tightly linked separate genes, i.e., the WDR gene and a putative beta-flanking gene of unknown function. The WDR and the beta-flanking genes are conserved genomic neighbors localized within a recently identified metabolic cell wall gene cluster in genomes of Aspergilli. The heterokaryons displayed increased susceptibility to drugs affecting the cell wall, and their phenotypes, observed by optical, confocal, scanning electron and atomic force microscopy, suggest cell wall alterations. Quantitative real-time PCR shows altered expression of some cell wall-related genes. The possible implications on cell wall biosynthesis are discussed.

  8. Beyond asexual development: modifications in the gene expression profile caused by the absence of the Aspergillus nidulans transcription factor FlbB.

    PubMed

    Oiartzabal-Arano, Elixabet; Garzia, Aitor; Gorostidi, Ana; Ugalde, Unai; Espeso, Eduardo A; Etxebeste, Oier

    2015-04-01

    In the model fungus Aspergillus nidulans, asexual development is induced from vegetative hyphae by a set of early regulators including the bZIP-type transcription factor FlbB. To determine the range of genes under the influence of the transcriptional activity of FlbB and to characterize their role in fungal development, we sequenced and compared the transcriptomes of a ΔflbB mutant and its isogenic wild-type strain at different developmental stages. Results confirmed the activating role of FlbB on downstream regulators of conidiation such as flbD and brlA. However, FlbB has additional functions beyond the induction of asexual development. Among the changes observed, absence of a functional FlbB caused induction of the dba cluster and synthesis of a secondary metabolite with bactericidal properties. In addition, a new transcriptional target of FlbB was unveiled, urdA, that codes for a putative transcription factor that represses premature sexual development. Taken together, our results indicate that the activators of asexual development simultaneously exert a role on other cellular functions, including an inhibitory effect on the sexual cycle, and reinforce the hypothesis that mutually exclusive metabolic and cellular patterns are associated with different morphogenetic programs.

  9. A WDR Gene Is a Conserved Member of a Chitin Synthase Gene Cluster and Influences the Cell Wall in Aspergillus nidulans

    PubMed Central

    Guerriero, Gea; Silvestrini, Lucia; Obersriebnig, Michael; Hausman, Jean-Francois; Strauss, Joseph; Ezcurra, Inés

    2016-01-01

    WD40 repeat (WDR) proteins are pleiotropic molecular hubs. We identify a WDR gene that is a conserved genomic neighbor of a chitin synthase gene in Ascomycetes. The WDR gene is unique to fungi and plants, and was called Fungal Plant WD (FPWD). FPWD is within a cell wall metabolism gene cluster in the Ascomycetes (Pezizomycotina) comprising chsD, a Chs activator and a GH17 glucanase. The FPWD, AN1556.2 locus was deleted in Aspergillus nidulans strain SAA.111 by gene replacement and only heterokaryon transformants were obtained. The re-annotation of Aspergilli genomes shows that AN1556.2 consists of two tightly linked separate genes, i.e., the WDR gene and a putative beta-flanking gene of unknown function. The WDR and the beta-flanking genes are conserved genomic neighbors localized within a recently identified metabolic cell wall gene cluster in genomes of Aspergilli. The heterokaryons displayed increased susceptibility to drugs affecting the cell wall, and their phenotypes, observed by optical, confocal, scanning electron and atomic force microscopy, suggest cell wall alterations. Quantitative real-time PCR shows altered expression of some cell wall-related genes. The possible implications on cell wall biosynthesis are discussed. PMID:27367684

  10. Modelling and mutational analysis of Aspergillus nidulans UreA, a member of the subfamily of urea/H+ transporters in fungi and plants

    PubMed Central

    Sanguinetti, Manuel; Amillis, Sotiris; Pantano, Sergio; Scazzocchio, Claudio; Ramón, Ana

    2014-01-01

    We present the first account of the structure–function relationships of a protein of the subfamily of urea/H+ membrane transporters of fungi and plants, using Aspergillus nidulans UreA as a study model. Based on the crystal structures of the Vibrio parahaemolyticus sodium/galactose symporter (vSGLT) and of the Nucleobase-Cation-Symport-1 benzylhydantoin transporter from Microbacterium liquefaciens (Mhp1), we constructed a three-dimensional model of UreA which, combined with site-directed and classical random mutagenesis, led to the identification of amino acids important for UreA function. Our approach allowed us to suggest roles for these residues in the binding, recognition and translocation of urea, and in the sorting of UreA to the membrane. Residues W82, Y106, A110, T133, N275, D286, Y388, Y437 and S446, located in transmembrane helixes 2, 3, 7 and 11, were found to be involved in the binding, recognition and/or translocation of urea and the sorting of UreA to the membrane. Y106, A110, T133 and Y437 seem to play a role in substrate selectivity, while S446 is necessary for proper sorting of UreA to the membrane. Other amino acids identified by random classical mutagenesis (G99, R141, A163, G168 and P639) may be important for the basic transporter's structure, its proper folding or its correct traffic to the membrane. PMID:24966243

  11. The intergenic region between the divergently transcribed niiA and niaD genes of Aspergillus nidulans contains multiple NirA binding sites which act bidirectionally.

    PubMed Central

    Punt, P J; Strauss, J; Smit, R; Kinghorn, J R; van den Hondel, C A; Scazzocchio, C

    1995-01-01

    The niaD and niiA genes of Aspergillus nidulans, which code, respectively, for nitrate and nitrite reductases, are divergently transcribed, and their ATGs are separated by 1,200 bp. The genes are under the control of the positively acting NirA transcription factor, which mediates nitrate induction. The DNA binding domain of NirA was expressed as a fusion protein with the glutathione S-transferase of Schistosoma japonicum. Gel shift and footprint experiments have shown that in the intergenic region there are four binding sites for the NirA transcription factor. These sites can be represented by the nonpalindromic consensus 5'CTCCGHGG3'. Making use of a bidirectional expression vector, we have analyzed the role of each of the sites in niaD and niiA expression. The sites were numbered from the niiA side. It appeared that site 1 is necessary for the inducibility of niiA only, while sites 2, 3, and to a lesser extent 4 (which is nearer to and strongly affects niaD) act bidirectionally. The results also suggest that of the 10 binding sites for the AreA protein, which mediates nitrogen metabolite repression, those which are centrally located are physiologically important. The insertion of an unrelated upstream activating sequence into the intergenic region strongly affected the expression of both genes, irrespective of the orientation in which the element was inserted. PMID:7565720

  12. Specific binding sites in the alcR and alcA promoters of the ethanol regulon for the CREA repressor mediating carbon catabolite repression in Aspergillus nidulans.

    PubMed

    Kulmburg, P; Mathieu, M; Dowzer, C; Kelly, J; Felenbok, B

    1993-03-01

    The CREA repressor responsible for carbon catabolite repression in Aspergillus nidulans represses the transcription of the ethanol regulon. The N-terminal part of the CREA protein encompassing the two zinc fingers (C2H2 class family) and an alanine-rich region was expressed in Escherichia coli as a fusion protein with glutathione-S-transferase. Our results show that CREA is a DNA-binding protein able to bind to the promoters of both the specific trans-acting gene, alcR, and of the structural gene, alcA, encoding the alcohol dehydrogenase I. DNase I protection footprinting experiments revealed several specific binding sites in the alcR and in the alcA promoters having the consensus sequence 5'-G/CPyGGGG-3'. The disruption of one of these CREA-binding sites in the alcR promoter overlapping the induction target for the trans-activator ALCR results in a partially derepressed alc phenotype and derepressed alcR transcription, showing that this binding site is functional in vivo. Our data suggest that CREA represses the ethanol regulon by a double lock mechanism repressing both the trans-acting gene, alcR, and the structural gene, alcA.

  13. A pericentrin-related protein homolog in Aspergillus nidulans plays important roles in nucleus positioning and cell polarity by affecting microtubule organization.

    PubMed

    Chen, Peiying; Gao, Rongsui; Chen, Shaochun; Pu, Li; Li, Pin; Huang, Ying; Lu, Ling

    2012-12-01

    Pericentrin is a large coiled-coil protein in mammalian centrosomes that serves as a multifunctional scaffold for anchoring numerous proteins. Recent studies have linked numerous human disorders with mutated or elevated levels of pericentrin, suggesting unrecognized contributions of pericentrin-related proteins to the development of these disorders. In this study, we characterized AnPcpA, a putative homolog of pericentrin-related protein in the model filamentous fungus Aspergillus nidulans, and found that it is essential for conidial germination and hyphal development. Compared to the hyphal apex localization pattern of calmodulin (CaM), which has been identified as an interactive partner of the pericentrin homolog, GFP-AnPcpA fluorescence dots are associated mainly with nuclei, while the accumulation of CaM at the hyphal apex depends on the function of AnPcpA. In addition, the depletion of AnPcpA by an inducible alcA promoter repression results in severe growth defects and abnormal nuclear segregation. Most interestingly, in mature hyphal cells, knockdown of pericentrin was able to significantly induce changes in cell shape and cytoskeletal remodeling; it resulted in some enlarged compartments with condensed nuclei and anucleate small compartments as well. Moreover, defects in AnPcpA significantly disrupted the microtubule organization and nucleation, suggesting that AnPcpA may affect nucleus positioning by influencing microtubule organization.

  14. Comparison of gene expression signatures of diamide, H2O2 and menadione exposed Aspergillus nidulans cultures – linking genome-wide transcriptional changes to cellular physiology

    PubMed Central

    Pócsi, István; Miskei, Márton; Karányi, Zsolt; Emri, Tamás; Ayoubi, Patricia; Pusztahelyi, Tünde; Balla, György; Prade, Rolf A

    2005-01-01

    Background In addition to their cytotoxic nature, reactive oxygen species (ROS) are also signal molecules in diverse cellular processes in eukaryotic organisms. Linking genome-wide transcriptional changes to cellular physiology in oxidative stress-exposed Aspergillus nidulans cultures provides the opportunity to estimate the sizes of peroxide (O22-), superoxide (O2•-) and glutathione/glutathione disulphide (GSH/GSSG) redox imbalance responses. Results Genome-wide transcriptional changes triggered by diamide, H2O2 and menadione in A. nidulans vegetative tissues were recorded using DNA microarrays containing 3533 unique PCR-amplified probes. Evaluation of LOESS-normalized data indicated that 2499 gene probes were affected by at least one stress-inducing agent. The stress induced by diamide and H2O2 were pulse-like, with recovery after 1 h exposure time while no recovery was observed with menadione. The distribution of stress-responsive gene probes among major physiological functional categories was approximately the same for each agent. The gene group sizes solely responsive to changes in intracellular O22-, O2•- concentrations or to GSH/GSSG redox imbalance were estimated at 7.7, 32.6 and 13.0 %, respectively. Gene groups responsive to diamide, H2O2 and menadione treatments and gene groups influenced by GSH/GSSG, O22- and O2•- were only partly overlapping with distinct enrichment profiles within functional categories. Changes in the GSH/GSSG redox state influenced expression of genes coding for PBS2 like MAPK kinase homologue, PSK2 kinase homologue, AtfA transcription factor, and many elements of ubiquitin tagging, cell division cycle regulators, translation machinery proteins, defense and stress proteins, transport proteins as well as many enzymes of the primary and secondary metabolisms. Meanwhile, a separate set of genes encoding transport proteins, CpcA and JlbA amino acid starvation-responsive transcription factors, and some elements of sexual development

  15. The Adenylate-Forming Enzymes AfeA and TmpB Are Involved in Aspergillus nidulans Self-Communication during Asexual Development.

    PubMed

    Soid-Raggi, Gabriela; Sánchez, Olivia; Ramos-Balderas, Jose L; Aguirre, Jesús

    2016-01-01

    Aspergillus nidulans asexual sporulation (conidiation) is triggered by different environmental signals and involves the differentiation of specialized structures called conidiophores. The elimination of genes flbA-E, fluG, and tmpA results in a fluffy phenotype characterized by delayed conidiophore development and decreased expression of the conidiation essential gene brlA. While flbA-E encode regulatory proteins, fluG and tmpA encode enzymes involved in the biosynthesis of independent signals needed for normal conidiation. Here we identify afeA and tmpB as new genes encoding members the adenylate-forming enzyme superfamily, whose inactivation cause different fluffy phenotypes and decreased conidiation and brlA expression. AfeA is most similar to unknown function coumarate ligase-like (4CL-Lk) enzymes and consistent with this, a K544N active site modification eliminates AfeA function. TmpB, identified previously as a larger homolog of the oxidoreductase TmpA, contains a NRPS-type adenylation domain. A high degree of synteny in the afeA-tmpA and tmpB regions in the Aspergilli suggests that these genes are part of conserved gene clusters. afeA, tmpA, and tmpB double and triple mutant analysis as well as afeA overexpression experiments indicate that TmpA and AfeA act in the same conidiation pathway, with TmpB acting in a different pathway. Fluorescent protein tagging shows that functional versions of AfeA are localized in lipid bodies and the plasma membrane, while TmpA and TmpB are localized at the plasma membrane. We propose that AfeA participates in the biosynthesis of an acylated compound, either a p-cuomaryl type or a fatty acid compound, which might be oxidized by TmpA and/or TmpB, while TmpB adenylation domain would be involved in the activation of a hydrophobic amino acid, which in turn would be oxidized by the TmpB oxidoreductase domain. Both, AfeA-TmpA and TmpB signals are involved in self-communication and reproduction in A. nidulans. PMID:27047469

  16. The Adenylate-Forming Enzymes AfeA and TmpB Are Involved in Aspergillus nidulans Self-Communication during Asexual Development.

    PubMed

    Soid-Raggi, Gabriela; Sánchez, Olivia; Ramos-Balderas, Jose L; Aguirre, Jesús

    2016-01-01

    Aspergillus nidulans asexual sporulation (conidiation) is triggered by different environmental signals and involves the differentiation of specialized structures called conidiophores. The elimination of genes flbA-E, fluG, and tmpA results in a fluffy phenotype characterized by delayed conidiophore development and decreased expression of the conidiation essential gene brlA. While flbA-E encode regulatory proteins, fluG and tmpA encode enzymes involved in the biosynthesis of independent signals needed for normal conidiation. Here we identify afeA and tmpB as new genes encoding members the adenylate-forming enzyme superfamily, whose inactivation cause different fluffy phenotypes and decreased conidiation and brlA expression. AfeA is most similar to unknown function coumarate ligase-like (4CL-Lk) enzymes and consistent with this, a K544N active site modification eliminates AfeA function. TmpB, identified previously as a larger homolog of the oxidoreductase TmpA, contains a NRPS-type adenylation domain. A high degree of synteny in the afeA-tmpA and tmpB regions in the Aspergilli suggests that these genes are part of conserved gene clusters. afeA, tmpA, and tmpB double and triple mutant analysis as well as afeA overexpression experiments indicate that TmpA and AfeA act in the same conidiation pathway, with TmpB acting in a different pathway. Fluorescent protein tagging shows that functional versions of AfeA are localized in lipid bodies and the plasma membrane, while TmpA and TmpB are localized at the plasma membrane. We propose that AfeA participates in the biosynthesis of an acylated compound, either a p-cuomaryl type or a fatty acid compound, which might be oxidized by TmpA and/or TmpB, while TmpB adenylation domain would be involved in the activation of a hydrophobic amino acid, which in turn would be oxidized by the TmpB oxidoreductase domain. Both, AfeA-TmpA and TmpB signals are involved in self-communication and reproduction in A. nidulans.

  17. The Adenylate-Forming Enzymes AfeA and TmpB Are Involved in Aspergillus nidulans Self-Communication during Asexual Development

    PubMed Central

    Soid-Raggi, Gabriela; Sánchez, Olivia; Ramos-Balderas, Jose L.; Aguirre, Jesús

    2016-01-01

    Aspergillus nidulans asexual sporulation (conidiation) is triggered by different environmental signals and involves the differentiation of specialized structures called conidiophores. The elimination of genes flbA-E, fluG, and tmpA results in a fluffy phenotype characterized by delayed conidiophore development and decreased expression of the conidiation essential gene brlA. While flbA-E encode regulatory proteins, fluG and tmpA encode enzymes involved in the biosynthesis of independent signals needed for normal conidiation. Here we identify afeA and tmpB as new genes encoding members the adenylate-forming enzyme superfamily, whose inactivation cause different fluffy phenotypes and decreased conidiation and brlA expression. AfeA is most similar to unknown function coumarate ligase-like (4CL-Lk) enzymes and consistent with this, a K544N active site modification eliminates AfeA function. TmpB, identified previously as a larger homolog of the oxidoreductase TmpA, contains a NRPS-type adenylation domain. A high degree of synteny in the afeA-tmpA and tmpB regions in the Aspergilli suggests that these genes are part of conserved gene clusters. afeA, tmpA, and tmpB double and triple mutant analysis as well as afeA overexpression experiments indicate that TmpA and AfeA act in the same conidiation pathway, with TmpB acting in a different pathway. Fluorescent protein tagging shows that functional versions of AfeA are localized in lipid bodies and the plasma membrane, while TmpA and TmpB are localized at the plasma membrane. We propose that AfeA participates in the biosynthesis of an acylated compound, either a p-cuomaryl type or a fatty acid compound, which might be oxidized by TmpA and/or TmpB, while TmpB adenylation domain would be involved in the activation of a hydrophobic amino acid, which in turn would be oxidized by the TmpB oxidoreductase domain. Both, AfeA-TmpA and TmpB signals are involved in self-communication and reproduction in A. nidulans. PMID:27047469

  18. Copy Number Suppressors of the Aspergillus nidulans nimA1 Mitotic Kinase Display Distinctive and Highly Dynamic Cell Cycle-Regulated Locations▿ †

    PubMed Central

    Ukil, Leena; Varadaraj, Archana; Govindaraghavan, Meera; Liu, Hui-Lin; Osmani, Stephen A.

    2008-01-01

    The Aspergillus nidulans NIMA kinase is essential for mitosis and is the founding member of the conserved NIMA-related kinase (Nek) family of protein kinases. To gain insight into NIMA function, a copy number suppression screen has been completed that defines three proteins termed MCNA, MCNB, and MCNC (multi-copy-number suppressor of nimA1 A, B, and C). All display a distinctive and dynamic cell cycle-specific distribution. MCNC has weak similarity to Saccharomyces cerevisiae Def1 within a shared CUE-like domain. MCNC, like Def1, is a cytoplasmic protein with slow mobility during sodium dodecyl sulfate-polyacrylamide gel electrophoresis, and its deletion causes polarization defects and a small colony phenotype. MCNC enters nuclei during mitosis. In contrast, MCNB is a nuclear protein displaying increased nuclear levels as cells progress through interphase but is lost from nuclei at mitosis. MCNB is highly related to the Schizosaccharomyces pombe forkhead transcription factor Sep1 and is likely a transcriptional activator of nimA. Most surprisingly, MCNA, a protein restricted to the aspergilli and pathogenic systemic dimorphic fungi (the Eurotiomycetes), defines a nuclear body located near nucleoli at the nuclear periphery of G2 nuclei. During progression through mitosis, the MCNA body is excluded from nuclei. Cytoplasmic MCNA bodies then diminish during early stages of interphase, and single MCNA bodies are formed within nuclei as interphase progresses. Three sites of MCNA phosphorylation were mapped and mutated to implicate proline-directed phosphorylation in the equal segregation of MCNA during the cell cycle. The data indicate all three MCN proteins likely have cell cycle functions. PMID:18931041

  19. Multiple effects of a commercial Roundup® formulation on the soil filamentous fungus Aspergillus nidulans at low doses: evidence of an unexpected impact on energetic metabolism.

    PubMed

    Nicolas, Valérie; Oestreicher, Nathalie; Vélot, Christian

    2016-07-01

    Soil microorganisms are highly exposed to glyphosate-based herbicides (GBH), especially to Roundup® which is widely used worldwide. However, studies on the effects of GBH formulations on specific non-rhizosphere soil microbial species are scarce. We evaluated the toxicity of a commercial formulation of Roundup® (R450), containing 450 g/L of glyphosate (GLY), on the soil filamentous fungus Aspergillus nidulans, an experimental model microorganism. The median lethal dose (LD50) on solid media was between 90 and 112 mg/L GLY (among adjuvants, which are also included in the Roundup® formulation), which corresponds to a dilution percentage about 100 times lower than that used in agriculture. The LOAEL and NOAEL (lowest- and no-observed-adverse-effect levels) associated to morphology and growth were 33.75 and 31.5 mg/L GLY among adjuvants, respectively. The formulation R450 proved to be much more active than technical GLY. At the LD50 and lower concentrations, R450 impaired growth, cellular polarity, endocytosis, and mitochondria (average number, total volume and metabolism). In contrast with the depletion of mitochondrial activities reported in animal studies, R450 caused a stimulation of mitochondrial enzyme activities, thus revealing a different mode of action of Roundup® on energetic metabolism. These mitochondrial disruptions were also evident at a low dose corresponding to the NOAEL for macroscopic parameters, indicating that these mitochondrial biomarkers are more sensitive than those for growth and morphological ones. Altogether, our data indicate that GBH toxic effects on soil filamentous fungi, and thus potential impairment of soil ecosystems, may occur at doses far below recommended agricultural application rate.

  20. Aspergillus nidulans transcription factor AtfA interacts with the MAPK SakA to regulate general stress responses, development and spore functions.

    PubMed

    Lara-Rojas, Fernando; Sánchez, Olivia; Kawasaki, Laura; Aguirre, Jesús

    2011-04-01

    Fungi utilize a phosphorelay system coupled to a MAP kinase module for sensing and processing environmental signals. In Aspergillus nidulans, response regulator SskA transmits osmotic and oxidative stress signals to the stress MAPK (SAPK) SakA. Using a genetic approach together with GFP tagging and molecular bifluorescence we show that SakA and ATF/CREB transcription factor AtfA define a general stress-signalling pathway that plays differential roles in oxidative stress responses during growth and development. AtfA is permanently localized in the nucleus, while SakA accumulates in the nucleus in response to oxidative or osmotic stress signals or during normal spore development, where it physically interacts with AtfA. AtfA is required for expression of several genes, the conidial accumulation of SakA and the viability of conidia. Furthermore, SakA is active (phosphorylated) in asexual spores, remaining phosphorylated in dormant conidia and becoming dephosphorylated during germination. SakA phosphorylation in spores depends on certain (SskA) but not other (SrrA and NikA) components of the phosphorelay system. Constitutive phosphorylation of SakA induced by the fungicide fludioxonil prevents both, germ tube formation and nuclear division. Similarly, Neurospora crassa SakA orthologue OS-2 is phosphorylated in intact conidia and gets dephosphorylated during germination. We propose that SakA-AtfA interaction regulates gene expression during stress and conidiophore development and that SAPK phosphorylation is a conserved mechanism to regulate transitions between non-growing (spore) and growing (mycelia) states.

  1. Physiological characterization of recombinant Saccharomyces cerevisiae expressing the Aspergillus nidulans phosphoketolase pathway: validation of activity through 13C-based metabolic flux analysis.

    PubMed

    Papini, Marta; Nookaew, Intawat; Siewers, Verena; Nielsen, Jens

    2012-08-01

    Several bacterial species and filamentous fungi utilize the phosphoketolase pathway (PHK) for glucose dissimilation as an alternative to the Embden-Meyerhof-Parnas pathway. In Aspergillus nidulans, the utilization of this metabolic pathway leads to increased carbon flow towards acetate and acetyl CoA. In the first step of the PHK, the pentose phosphate pathway intermediate xylulose-5-phosphate is converted into acetylphosphate and glyceraldehyde-3-phosphate through the action of xylulose-5-phosphate phosphoketolase, and successively acetylphosphate is converted into acetate by the action of acetate kinase. In the present work, we describe a metabolic engineering strategy used to express the fungal genes of the phosphoketolase pathway in Saccharomyces cerevisiae and the effects of the expression of this recombinant route in yeast. The phenotype of the engineered yeast strain MP003 was studied during batch and chemostat cultivations, showing a reduced biomass yield and an increased acetate yield during batch cultures. To establish whether the observed effects in the recombinant strain MP003 were due directly or indirectly to the expression of the phosphoketolase pathway, we resolved the intracellular flux distribution based on (13)C labeling during chemostat cultivations. From flux analysis it is possible to conclude that yeast is able to use the recombinant pathway. Our work indicates that the utilization of the phosphoketolase pathway does not interfere with glucose assimilation through the Embden-Meyerhof-Parnas pathway and that the expression of this route can contribute to increase the acetyl CoA supply, therefore holding potential for future metabolic engineering strategies having acetyl CoA as precursor for the biosynthesis of industrially relevant compounds.

  2. Involvement of Protein Kinase C in the Suppression of Apoptosis and in Polarity Establishment in Aspergillus nidulans under Conditions of Heat Stress

    PubMed Central

    Katayama, Takuya; Uchida, Hirotaka; Ohta, Akinori; Horiuchi, Hiroyuki

    2012-01-01

    The pkcA gene, which encodes a protein kinase C (PKC) in the filamentous fungus Aspergillus nidulans, is essential for its viability. However, little is known about its functions. To address this issue, we constructed and characterized temperature-sensitive mutants of pkcA. The conidia of these mutants swelled slightly and exhibited apoptotic phenotypes at 42°C. The apoptotic phenotypes were suppressed by an osmotic stabilizer. Under these conditions, the conidia swelled extensively and did not form germ tubes. Moreover, polarized distribution of F-actin was not observed. We then utilized deletion mutants of bckA, an ortholog of Saccharomyces cerevisiae bck1 that encodes a mitogen-activated protein (MAP) kinase kinase kinase and functions downstream of PKC in the cell wall integrity pathway. These mutants exhibited apoptotic phenotypes at 42°C, but they did not show defects in polarity establishment under osmotically stabilized conditions. These results suggest that PkcA plays multiple roles during germination under conditions of heat stress. The first of these roles is the suppression of apoptosis induction, while the other involves polarity establishment. The former depends on the MAP kinase cascade, whereas the latter does not. In addition, repolarization, which was observed after depolarization in the wild-type strain and the bckA deletion mutant under conditions of heat stress, was not observed in the pkcA-ts mutant. This suggests that PkcA also plays role in polarity establishment during hyphal growth independent of the MAP kinase cascade under these conditions. PMID:23209763

  3. High-affinity nitrate/nitrite transporters NrtA and NrtB of Aspergillus nidulans exhibit high specificity and different inhibitor sensitivity

    PubMed Central

    Akhtar, Naureen; Karabika, Eugenia; Kinghorn, James R.; Glass, Anthony D.M.; Unkles, Shiela E.

    2015-01-01

    The NrtA and NrtB nitrate transporters are paralogous members of the major facilitator superfamily in Aspergillus nidulans. The availability of loss-of-function mutations allowed individual investigation of the specificity and inhibitor sensitivity of both NrtA and NrtB. In this study, growth response tests were carried out at a growth-limiting concentration of nitrate (1 mM) as the sole nitrogen source, in the presence of a number of potential nitrate analogues at various concentrations, to evaluate their effect on nitrate transport. Both chlorate and chlorite inhibited fungal growth, with chlorite exerting the greater inhibition. The main transporter of nitrate, NrtA, proved to be more sensitive to chlorate than the minor transporter, NrtB. Similarly, the cation caesium was shown to exert differential effects, strongly inhibiting the activity of NrtB, but not NrtA. In contrast, no inhibition of nitrate uptake by NrtA or NrtB transporters was observed in either growth tests or uptake assays in the presence of bicarbonate, formate, malonate or oxalate (sulphite could not be tested in uptake assays owing to its reaction with nitrate), indicating significant specificity of nitrate transport. Kinetic analyses of nitrate uptake revealed that both chlorate and chlorite inhibited NrtA competitively, while these same inhibitors inhibited NrtB in a non-competitive fashion. The caesium ion appeared to inhibit NrtA in a non-competitive fashion, while NrtB was inhibited uncompetitively. The results provide further evidence of the distinctly different characteristics as well as the high specificity of nitrate uptake by these two transporters. PMID:25855763

  4. Multiple effects of a commercial Roundup® formulation on the soil filamentous fungus Aspergillus nidulans at low doses: evidence of an unexpected impact on energetic metabolism.

    PubMed

    Nicolas, Valérie; Oestreicher, Nathalie; Vélot, Christian

    2016-07-01

    Soil microorganisms are highly exposed to glyphosate-based herbicides (GBH), especially to Roundup® which is widely used worldwide. However, studies on the effects of GBH formulations on specific non-rhizosphere soil microbial species are scarce. We evaluated the toxicity of a commercial formulation of Roundup® (R450), containing 450 g/L of glyphosate (GLY), on the soil filamentous fungus Aspergillus nidulans, an experimental model microorganism. The median lethal dose (LD50) on solid media was between 90 and 112 mg/L GLY (among adjuvants, which are also included in the Roundup® formulation), which corresponds to a dilution percentage about 100 times lower than that used in agriculture. The LOAEL and NOAEL (lowest- and no-observed-adverse-effect levels) associated to morphology and growth were 33.75 and 31.5 mg/L GLY among adjuvants, respectively. The formulation R450 proved to be much more active than technical GLY. At the LD50 and lower concentrations, R450 impaired growth, cellular polarity, endocytosis, and mitochondria (average number, total volume and metabolism). In contrast with the depletion of mitochondrial activities reported in animal studies, R450 caused a stimulation of mitochondrial enzyme activities, thus revealing a different mode of action of Roundup® on energetic metabolism. These mitochondrial disruptions were also evident at a low dose corresponding to the NOAEL for macroscopic parameters, indicating that these mitochondrial biomarkers are more sensitive than those for growth and morphological ones. Altogether, our data indicate that GBH toxic effects on soil filamentous fungi, and thus potential impairment of soil ecosystems, may occur at doses far below recommended agricultural application rate. PMID:27068896

  5. Patterns of nucleosomal organization in the alc regulon of Aspergillus nidulans: roles of the AlcR transcriptional activator and the CreA global repressor.

    PubMed

    Mathieu, Martine; Nikolaev, Igor; Scazzocchio, Claudio; Felenbok, Béatrice

    2005-04-01

    We have studied the chromatin organization of three promoters of the alc regulon of Aspergillus nidulans. No positioned nucleosomes are seen in the aldA (aldehyde dehydrogenase) promoter under any physiological condition tested by us. In the alcA (alcohol dehydrogenase I) and alcR (coding for the pathway-specific transcription factor) promoters, a pattern of positioned nucleosomes is seen under non-induced and non-induced repressed conditions. While each of these promoters shows a specific pattern of chromatin restructuring, in both cases induction results in loss of nucleosome positioning. Glucose repression in the presence of inducer results in a specific pattern of partial positioning in the alcA and alcR promoters. Loss of nucleosome positioning depends absolutely on the AlcR protein and it is very unlikely to be a passive result of the induction of transcription. In an alcR loss-of-function background and in strains carrying mutations of the respective AlcR binding sites of the alcA and alcR promoters, nucleosomes are fully positioned under all growth conditions. Analysis of mutant AlcR proteins establishes that all domains needed for transcriptional activation and chromatin restructuring are included within the first 241 residues. The results suggest a two-step process, one step resulting in chromatin restructuring, a second one in transcriptional activation. Partial positioning upon glucose repression shows a specific pattern that depends on the CreA global repressor. An alcR loss-of-function mutation is epistatic to a creA loss-of-function mutation, showing that AlcR does not act by negating a nucleosome positioning activity of CreA.

  6. Genomic clones of Aspergillus nidulans containing alcA, the structural gene for alcohol dehydrogenase and alcR, a regulatory gene for ethanol metabolism.

    PubMed

    Doy, C H; Pateman, J A; Olsen, J E; Kane, H J; Creaser, E H

    1985-04-01

    Our aim was to obtain from Aspergillus nidulans a genomic bank and then clone a region we expected from earlier genetic mapping to contain two closely linked genes, alcA, the structural gene for alcohol dehydrogenase (ADH) and alcR, a positive trans-acting regulatory gene for ethanol metabolism. The expression of alcA is repressed by carbon catabolites. A genomic restriction fragment characteristic of the alcA-alcR region was identified, cloned in pBR322, and used to select from a genomic bank in lambda EMBL3A three overlapping clones covering 24 kb of DNA. Southern genomic analysis of wild-type, alcA and alcR mutants showed that the mutants contained extra DNA at sites near the center of the cloned DNA and are close together, as expected for alcA and alcR. Transcription from the cloned DNA and hybridization with a clone carrying the Saccharomyces cerevisiae gene for ADHI (ADC1) are both confined to the alcA-alcR region. At least one of several species of mature mRNA is about 1 kb, the size required to code for ADH. For all species, carbon catabolite repression overrides control by induction. The overall characteristics of transcription, hybridization to ADC1 and earlier work suggest that alcA consists of a number of exons and/or that the alcA-alcR region represents a cluster of alcA-related genes or sequences.

  7. Specific binding sites for the activator protein, ALCR, in the alcA promoter of the ethanol regulon of Aspergillus nidulans.

    PubMed

    Kulmburg, P; Judewicz, N; Mathieu, M; Lenouvel, F; Sequeval, D; Felenbok, B

    1992-10-15

    ALCR is the specific activator of the Aspergillus nidulans ethanol-utilization pathway, mediating the induction of its own transcription and that of the structural genes alcA and aldA, encoding respectively, alcohol dehydrogenase I and aldehyde dehydrogenase. ALCR is a DNA binding protein in which 6 cysteines are coordinated in a zinc binuclear cluster. This domain was fused to glutathione-S-transferase (GST) and isolated as a GST-ALCR(7-58*) fusion protein from Escherichia coli. Mobility shift assays showed that the ALCR fusion protein binds at sites upstream of the alcA promoter. DNaseI protection footprinting experiments revealed three specific binding sites, two that are direct repeats and one that is an inverted repeat with the same half-site 5'-CCGCA-3'. The half-sites are separated by a variable number of nucleotides in both types of target. The interaction of the ALCR fusion protein with direct and inverted repeats were examined by using interference and protection footprinting assays. In both binding sites, modification of the guanines in the half-sites interfered with the formation of the DNA complex, but the adjacent ones did not. Our results suggest that the ALCR protein makes contact in the major groove of the DNA helix of the half-sites. The functionality of two out of three binding sites of the GST-ALCR protein was demonstrated after their deletion. Therefore, the region encompassing these binding sites is a cis-acting element involved in the full induction of the alcA gene.

  8. Genetic Interaction of Aspergillus nidulans galR, xlnR and araR in Regulating D-Galactose and L-Arabinose Release and Catabolism Gene Expression

    PubMed Central

    Kowalczyk, Joanna E.; Gruben, Birgit S.; Battaglia, Evy; Wiebenga, Ad; Majoor, Eline; de Vries, Ronald P.

    2015-01-01

    In Aspergillus nidulans, the xylanolytic regulator XlnR and the arabinanolytic regulator AraR co-regulate pentose catabolism. In nature, the pentose sugars D-xylose and L-arabinose are both main building blocks of the polysaccharide arabinoxylan. In pectin and arabinogalactan, these two monosaccharides are found in combination with D-galactose. GalR, the regulator that responds to the presence of D-galactose, regulates the D-galactose catabolic pathway. In this study we investigated the possible interaction between XlnR, AraR and GalR in pentose and/or D-galactose catabolism in A. nidulans. Growth phenotypes and metabolic gene expression profiles were studied in single, double and triple disruptant A. nidulans strains of the genes encoding these paralogous transcription factors. Our results demonstrate that AraR and XlnR not only control pentose catabolic pathway genes, but also genes of the oxido-reductive D-galactose catabolic pathway. This suggests an interaction between three transcriptional regulators in D-galactose catabolism. Conversely, GalR is not involved in regulation of pentose catabolism, but controls only genes of the oxido-reductive D-galactose catabolic pathway. PMID:26580075

  9. Genetic Interaction of Aspergillus nidulans galR, xlnR and araR in Regulating D-Galactose and L-Arabinose Release and Catabolism Gene Expression.

    PubMed

    Kowalczyk, Joanna E; Gruben, Birgit S; Battaglia, Evy; Wiebenga, Ad; Majoor, Eline; de Vries, Ronald P

    2015-01-01

    In Aspergillus nidulans, the xylanolytic regulator XlnR and the arabinanolytic regulator AraR co-regulate pentose catabolism. In nature, the pentose sugars D-xylose and L-arabinose are both main building blocks of the polysaccharide arabinoxylan. In pectin and arabinogalactan, these two monosaccharides are found in combination with D-galactose. GalR, the regulator that responds to the presence of D-galactose, regulates the D-galactose catabolic pathway. In this study we investigated the possible interaction between XlnR, AraR and GalR in pentose and/or D-galactose catabolism in A. nidulans. Growth phenotypes and metabolic gene expression profiles were studied in single, double and triple disruptant A. nidulans strains of the genes encoding these paralogous transcription factors. Our results demonstrate that AraR and XlnR not only control pentose catabolic pathway genes, but also genes of the oxido-reductive D-galactose catabolic pathway. This suggests an interaction between three transcriptional regulators in D-galactose catabolism. Conversely, GalR is not involved in regulation of pentose catabolism, but controls only genes of the oxido-reductive D-galactose catabolic pathway.

  10. Isolation of a gene involved in 1,3-beta-glucan synthesis in Aspergillus nidulans and purification of the corresponding protein.

    PubMed Central

    Kelly, R; Register, E; Hsu, M J; Kurtz, M; Nielsen, J

    1996-01-01

    Saccharomyces cerevisiae has two highly homologous genes, FKS1 and FKS2, which encode interchangeable putative catalytic subunits of 1,3-beta-glucan synthase (GS), an enzyme that synthesizes an essential polymer of the fungal cell wall. To determine if GS in Aspergillus species is similar, an FKS homolog, fksA, was cloned from Aspergillus nidulans by cross-hybridization, and the corresponding protein was purified. Sequence analysis revealed a 5,716-nucleotide coding region interrupted by two 56-bp introns. The fksA gene encodes a predicted peptide of 229 kDa, FksAp, that shows a remarkable degree of conservation in size, charge, amino acid identity, and predicted membrane topology with the S. cerevisiae FKS proteins (Fksps). FksAp exhibits 64 and 65% identity to Fks1p and Fks2p, respectively, and 79% similarity. Hydropathy analysis of FksAp suggests an integral membrane protein with 16 transmembrane helices that coincide with the transmembrane helices of the Saccharomyces Fksps. The sizes of the nontransmembrane domains are strikingly similar to those of Fks1p. The region of FksAp most homologous to the Saccharomyces FKS polypeptides is a large hydrophilic domain of 578 amino acids that is predicted to be cytoplasmic. This domain is 86% identical to the corresponding region of Fks1p and is a good candidate for the location of the catalytic site. Antibodies raised against a peptide derived from the FksAp sequence recognize a protein of approximately 200 kDa in crude membranes and detergent-solubilized active extracts. This protein is enriched approximately 300-fold in GS purified by product entrapment. Purified anti-FksAp immunoglobulin G immunodepletes nearly all of the GS activity in crude or purified extracts when Staphylococcus aureus cells are used to precipitate the antibodies, although it does not inhibit enzymatic activity when added to extracts. The purified GS is inhibited by echinocandins with a sensitivity equal to that displayed by whole cells. Thus

  11. In Aspergillus nidulans the Suppressors suaA and suaC Code for Release Factors eRF1 and eRF3 and suaD Codes for a Glutamine tRNA

    PubMed Central

    Liu, Wen; Mellado, Laura; Espeso, Eduardo A.; Sealy-Lewis, Heather M.

    2014-01-01

    In Aspergillus nidulans, after extensive mutagenesis, a collection of mutants was obtained and four suppressor loci were identified genetically that could suppress mutations in putative chain termination mutations in different genes. Suppressor mutations in suaB and suaD have a similar restricted spectrum of suppression and suaB111 was previously shown to be an alteration in the anticodon of a gln tRNA. We have shown that like suaB, a suaD suppressor has a mutation in the anticodon of another gln tRNA allowing suppression of UAG mutations. Mutations in suaA and suaC had a broad spectrum of suppression. Four suaA mutations result in alterations in the coding region of the eukaryotic release factor, eRF1, and another suaA mutation has a mutation in the upstream region of eRF1 that prevents splicing of the first intron within the 5′UTR. Epitope tagging of eRF1 in this mutant results in 20% of the level of eRF1 compared to the wild-type. Two mutations in suaC result in alterations in the eukaryotic release factor, eRF3. This is the first description in Aspergillus nidulans of an alteration in eRF3 leading to suppression of chain termination mutations. PMID:24727290

  12. Use of reporter genes to identify recessive trans-acting mutations specifically involved in the regulation of Aspergillus nidulans penicillin biosynthesis genes.

    PubMed Central

    Brakhage, A A; Van den Brulle, J

    1995-01-01

    Starting from three amino acid precursors, penicillin biosynthesis is catalyzed by three enzymes which are encoded by the following three genes: acvA (pcbAB), ipnA (pcbC), and aat (penDE). To identify trans-acting mutations which are specifically involved in the regulation of these secondary metabolism genes, a molecular approach was employed by using an Aspergillus nidulans strain (AXTII9) carrying acvA-uidA and ipnA-lacZ gene fusions integrated in double copies at the chromosomal argB gene. On minimal agar plates supplemented with X-Gal (5-bromo-4-chloro-3-indolyl-beta-D-galactopyranoside), colonies of such a strain stained blue, which is indicative of ipnA-lacZ expression. After mutagenesis with UV light, colonies were isolated on agar plates with lactose as the carbon source, which produced only a faint blue color or no color at all. Such mutants (named Prg for penicillin regulation) most likely were defective in trans-acting genes. Control experiments revealed that the mutants studied still carried the correct number of gene fusions. In a fermentation run, mutants Prg-1 and Prg-6 exhibited only 20 to 50% of the ipnA-lacZ expression of the wild-type strain and produced only 20 to 30% of the penicillin produced by the wild-type strain. Western blot (immunoblot) analysis showed that these mutants contained reduced amounts of ipnA gene product, i.e., isopenicillin N synthase. Both mutant Prg-1 and mutant Prg-6 also differed in acvA-uidA expression levels from the wild type. Segregation analysis indicated that for both mutants the Prg phenotype resulted from mutation of a single gene. Two different complementation groups, which were designated prgA1 and prgB1, were identified. However, the specific activity of the aat (penDE) gene product, i.e., acyl coenzyme A:6-aminopenicillanic acid acyltransferase, was essentially the same for the mutants as for the wild-type strain, implying that the last step of the penicillin biosynthetic pathway is not affected by the trans

  13. An efficient arabinoxylan-debranching α-L-arabinofuranosidase of family GH62 from Aspergillus nidulans contains a secondary carbohydrate binding site.

    PubMed

    Wilkens, Casper; Andersen, Susan; Petersen, Bent O; Li, An; Busse-Wicher, Marta; Birch, Johnny; Cockburn, Darrell; Nakai, Hiroyuki; Christensen, Hans E M; Kragelund, Birthe B; Dupree, Paul; McCleary, Barry; Hindsgaul, Ole; Hachem, Maher Abou; Svensson, Birte

    2016-07-01

    An α-L-arabinofuranosidase of GH62 from Aspergillus nidulans FGSC A4 (AnAbf62A-m2,3) has an unusually high activity towards wheat arabinoxylan (WAX) (67 U/mg; k cat = 178/s, K m = 4.90 mg/ml) and arabinoxylooligosaccharides (AXOS) with degrees of polymerisation (DP) 3-5 (37-80 U/mg), but about 50 times lower activity for sugar beet arabinan and 4-nitrophenyl-α-L-arabinofuranoside. α-1,2- and α-1,3-linked arabinofuranoses are released from monosubstituted, but not from disubstituted, xylose in WAX and different AXOS as demonstrated by NMR and polysaccharide analysis by carbohydrate gel electrophoresis (PACE). Mutants of the predicted general acid (Glu(188)) and base (Asp(28)) catalysts, and the general acid pK a modulator (Asp(136)) lost 1700-, 165- and 130-fold activities for WAX. WAX, oat spelt xylan, birchwood xylan and barley β-glucan retarded migration of AnAbf62A-m2,3 in affinity electrophoresis (AE) although the latter two are neither substrates nor inhibitors. Trp(23) and Tyr(44), situated about 30 Å from the catalytic site as seen in an AnAbf62A-m2,3 homology model generated using Streptomyces thermoviolaceus SthAbf62A as template, participate in carbohydrate binding. Compared to wild-type, W23A and W23A/Y44A mutants are less retarded in AE, maintain about 70 % activity towards WAX with K i of WAX substrate inhibition increasing 4-7-folds, but lost 77-96 % activity for the AXOS. The Y44A single mutant had less effect, suggesting Trp(23) is a key determinant. AnAbf62A-m2,3 seems to apply different polysaccharide-dependent binding modes, and Trp(23) and Tyr(44) belong to a putative surface binding site which is situated at a distance of the active site and has to be occupied to achieve full activity. PMID:26946172

  14. The SrkA Kinase Is Part of the SakA Mitogen-Activated Protein Kinase Interactome and Regulates Stress Responses and Development in Aspergillus nidulans

    PubMed Central

    Jaimes-Arroyo, Rafael; Lara-Rojas, Fernando; Bayram, Özgür; Valerius, Oliver; Braus, Gerhard H.

    2015-01-01

    Fungi and many other eukaryotes use specialized mitogen-activated protein kinases (MAPK) of the Hog1/p38 family to transduce environmental stress signals. In Aspergillus nidulans, the MAPK SakA and the transcription factor AtfA are components of a central multiple stress-signaling pathway that also regulates development. Here we characterize SrkA, a putative MAPK-activated protein kinase, as a novel component of this pathway. ΔsrkA and ΔsakA mutants share a derepressed sexual development phenotype. However, ΔsrkA mutants are not sensitive to oxidative stress, and in fact, srkA inactivation partially suppresses the sensitivity of ΔsakA mutant conidia to H2O2, tert-butyl-hydroperoxide (t-BOOH), and menadione. In the absence of stress, SrkA shows physical interaction with nonphosphorylated SakA in the cytosol. We show that H2O2 induces a drastic change in mitochondrial morphology consistent with a fission process and the relocalization of SrkA to nuclei and mitochondria, depending on the presence of SakA. SakA-SrkA nuclear interaction is also observed during normal asexual development in dormant spores. Using SakA and SrkA S-tag pulldown and purification studies coupled to mass spectrometry, we found that SakA interacts with SrkA, the stress MAPK MpkC, the PPT1-type phosphatase AN6892, and other proteins involved in cell cycle regulation, DNA damage response, mRNA stability and protein synthesis, mitochondrial function, and other stress-related responses. We propose that oxidative stress induces DNA damage and mitochondrial fission and that SakA and SrkA mediate cell cycle arrest and regulate mitochondrial function during stress. Our results provide new insights into the mechanisms by which SakA and SrkA regulate the remodelling of cell physiology during oxidative stress and development. PMID:25820520

  15. Response regulators SrrA and SskA are central components of a phosphorelay system involved in stress signal transduction and asexual sporulation in Aspergillus nidulans.

    PubMed

    Vargas-Pérez, Itzel; Sánchez, Olivia; Kawasaki, Laura; Georgellis, Dimitris; Aguirre, Jesús

    2007-09-01

    Among eukaryotes, only slime molds, fungi, and plants contain signal transduction phosphorelay systems. In filamentous fungi, multiple sensor kinases appear to use a single histidine-containing phosphotransfer (HPt) protein to relay signals to two response regulators (RR). In Aspergillus nidulans, the RR SskA mediates activation of the mitogen-activated protein kinase SakA in response to osmotic and oxidative stress, whereas the functions of the RR SrrA were unknown. We used a genetic approach to characterize the srrA gene as a new member of the skn7/prr1 family and to analyze the roles of SrrA in the phosphorelay system composed of the RR SskA, the HPt protein YpdA, and the sensor kinase NikA. While mutants lacking the HPt protein YpdA are unviable, mutants lacking SskA (DeltasskA), SrrA (DeltasrrA), or both RR (DeltasrrA DeltasskA) are viable and differentially affected in osmotic and oxidative stress responses. Both RR are involved in osmostress resistance, but DeltasskA mutants are more sensitive to this stress, and only SrrA is required for H(2)O(2) resistance and H(2)O(2)-mediated induction of catalase CatB. In contrast, both RR are individually required for fungicide sensitivity and calcofluor resistance and for normal sporulation and conidiospore viability. The DeltasrrA and DeltasskA sporulation defects appear to be related to decreased mRNA levels of the key sporulation gene brlA. In contrast, conidiospore viability defects do not correlate with the activity of the spore-specific catalase CatA. Our results support a model in which NikA acts upstream of SrrA and SskA to transmit fungicide signals and to regulate asexual sporulation and conidiospore viability. In contrast, NikA appears dispensable for osmotic and oxidative stress signaling. These results highlight important differences in stress signal transmission among fungi and define a phosphorelay system involved in oxidative and osmotic stress, cell wall maintenance, fungicide sensitivity, asexual

  16. Functional analysis of the α-1,3-glucan synthase genes agsA and agsB in Aspergillus nidulans: agsB is the major α-1,3-glucan synthase in this fungus.

    PubMed

    Yoshimi, Akira; Sano, Motoaki; Inaba, Azusa; Kokubun, Yuko; Fujioka, Tomonori; Mizutani, Osamu; Hagiwara, Daisuke; Fujikawa, Takashi; Nishimura, Marie; Yano, Shigekazu; Kasahara, Shin; Shimizu, Kiminori; Yamaguchi, Masashi; Kawakami, Kazuyoshi; Abe, Keietsu

    2013-01-01

    Although α-1,3-glucan is one of the major cell wall polysaccharides in filamentous fungi, the physiological roles of α-1,3-glucan remain unclear. The model fungus Aspergillus nidulans possesses two α-1,3-glucan synthase (AGS) genes, agsA and agsB. For functional analysis of these genes, we constructed several mutant strains in A. nidulans: agsA disruption, agsB disruption, and double-disruption strains. We also constructed several CagsB strains in which agsB expression was controlled by the inducible alcA promoter, with or without the agsA-disrupting mutation. The agsA disruption strains did not show markedly different phenotypes from those of the wild-type strain. The agsB disruption strains formed dispersed hyphal cells under liquid culture conditions, regardless of the agsA genetic background. Dispersed hyphal cells were also observed in liquid culture of the CagsB strains when agsB expression was repressed, whereas these strains grew normally in plate culture even under the agsB-repressed conditions. Fractionation of the cell wall based on the alkali solubility of its components, quantification of sugars, and (13)C-NMR spectroscopic analysis revealed that α-1,3-glucan was the main component of the alkali-soluble fraction in the wild-type and agsA disruption strains, but almost no α-1,3-glucan was found in the alkali-soluble fraction derived from either the agsB disruption strain or the CagsB strain under the agsB-repressed conditions, regardless of the agsA genetic background. Taken together, our data demonstrate that the two AGS genes are dispensable in A. nidulans, but that AgsB is required for normal growth characteristics under liquid culture conditions and is the major AGS in this species.

  17. The amdR product and a CCAAT-binding factor bind to adjacent, possibly overlapping DNA sequences in the promoter region of the Aspergillus nidulans amdS gene.

    PubMed Central

    van Heeswijck, R; Hynes, M J

    1991-01-01

    The amdS gene of Aspergillus nidulans is regulated by a number of positively acting regulatory genes which act additively and independently. Using gel mobility shift assays with crude nuclear extracts we show here that the product of one of these regulatory genes, the amdR gene, binds to DNA fragments containing part of the promoter region of the amdS gene. This confirms the earlier prediction from DNA sequence data that amdR encodes a DNA-binding protein containing a cysteine-rich 'zinc finger' motif. In addition we detected the binding of another previously unidentified protein to an adjacent, possibly overlapping region of the amdS 5' sequence at the site of a consensus 'CCAAT-box' sequence. Replacement of the CCAAT sequence with CCTTT abolished the binding of this protein which we have designated as an A. nidulans 'CCAAT-box' binding factor (AnCF). The 'CCAAT-box' sequence appears to be involved in determining the basal level of transcription of amdS (T.G.Littlejohn and M.J.H., unpublished data). This suggests that AnCF is a transcription factor, and that the 'CCAAT-box' sequences found in the promoters of some filamentous fungal genes function as binding sites for these factors, as in other eucaryotes. Images PMID:2041742

  18. AnCF, the CCAAT Binding Complex of Aspergillus nidulans, Is Essential for the Formation of a DNase I-Hypersensitive Site in the 5′ Region of the amdS Gene

    PubMed Central

    Narendja, Frank M.; Davis, Meryl A.; Hynes, Michael J.

    1999-01-01

    The CCAAT sequence in the amdS promoter of Aspergillus nidulans is recognized by AnCF, a complex consisting of the three evolutionary conserved subunits HapB, HapC, and HapE. In this study we have investigated the effect of AnCF on the chromatin structure of the amdS gene. The AnCF complex and the CCAAT sequence were found to be necessary for the formation of a nucleosome-free, DNase I-hypersensitive region in the 5′ region of the amdS gene. Deletion of the hapE gene results in loss of the DNase I-hypersensitive site, and the positioning of nucleosomes over the transcriptional start point is lost. Likewise, a point mutation in the CCAAT motif, as well as a 530-bp deletion which removes the CCAAT box, results in the loss of the DNase I-hypersensitive region. The DNase I-hypersensitive region and the nucleosome positioning can be restored by insertion of a 35-bp oligonucleotide carrying the CCAAT motif. A DNase I-hypersensitive region has been found in the CCAAT-containing fmdS gene and was also hapE dependent. These data indicate a critical role for the AnCF complex in establishing an open chromatin structure in A. nidulans. PMID:10490592

  19. AnCF, the CCAAT binding complex of Aspergillus nidulans, is essential for the formation of a DNase I-hypersensitive site in the 5' region of the amdS gene.

    PubMed

    Narendja, F M; Davis, M A; Hynes, M J

    1999-10-01

    The CCAAT sequence in the amdS promoter of Aspergillus nidulans is recognized by AnCF, a complex consisting of the three evolutionary conserved subunits HapB, HapC, and HapE. In this study we have investigated the effect of AnCF on the chromatin structure of the amdS gene. The AnCF complex and the CCAAT sequence were found to be necessary for the formation of a nucleosome-free, DNase I-hypersensitive region in the 5' region of the amdS gene. Deletion of the hapE gene results in loss of the DNase I-hypersensitive site, and the positioning of nucleosomes over the transcriptional start point is lost. Likewise, a point mutation in the CCAAT motif, as well as a 530-bp deletion which removes the CCAAT box, results in the loss of the DNase I-hypersensitive region. The DNase I-hypersensitive region and the nucleosome positioning can be restored by insertion of a 35-bp oligonucleotide carrying the CCAAT motif. A DNase I-hypersensitive region has been found in the CCAAT-containing fmdS gene and was also hapE dependent. These data indicate a critical role for the AnCF complex in establishing an open chromatin structure in A. nidulans.

  20. The amdR product and a CCAAT-binding factor bind to adjacent, possibly overlapping DNA sequences in the promoter region of the Aspergillus nidulans amdS gene.

    PubMed

    van Heeswijck, R; Hynes, M J

    1991-05-25

    The amdS gene of Aspergillus nidulans is regulated by a number of positively acting regulatory genes which act additively and independently. Using gel mobility shift assays with crude nuclear extracts we show here that the product of one of these regulatory genes, the amdR gene, binds to DNA fragments containing part of the promoter region of the amdS gene. This confirms the earlier prediction from DNA sequence data that amdR encodes a DNA-binding protein containing a cysteine-rich 'zinc finger' motif. In addition we detected the binding of another previously unidentified protein to an adjacent, possibly overlapping region of the amdS 5' sequence at the site of a consensus 'CCAAT-box' sequence. Replacement of the CCAAT sequence with CCTTT abolished the binding of this protein which we have designated as an A. nidulans 'CCAAT-box' binding factor (AnCF). The 'CCAAT-box' sequence appears to be involved in determining the basal level of transcription of amdS (T.G. Littlejohn and M.J.H., unpublished data). This suggests that AnCF is a transcription factor, and that the 'CCAAT-box' sequences found in the promoters of some filamentous fungal genes function as binding sites for these factors, as in other eucaryotes.

  1. Novel basic-region helix-loop-helix transcription factor (AnBH1) of Aspergillus nidulans counteracts the CCAAT-binding complex AnCF in the promoter of a penicillin biosynthesis gene.

    PubMed

    Caruso, Maria Louise; Litzka, Olivier; Martic, Goran; Lottspeich, Friedrich; Brakhage, Axel A

    2002-10-25

    Cis-acting CCAAT elements are found frequently in eukaryotic promoter regions. Many of the genes containing such elements in their promoters are regulated by a conserved multimeric CCAAT-binding complex. In the fungus Emericella (Aspergillus) nidulans, this complex was designated AnCF (A.nidulans CCAAT-binding factor). AnCF regulates several genes, including the penicillin biosynthesis genes ipnA and aatA. Since it is estimated that the CCAAT-binding complex regulates more than 200 genes, an important question concerns the regulation mechanism that allows so many genes to be regulated by a single complex in a gene-specific manner. One of the answers to this question appears to lie in the interaction of AnCF with other transcription factors. Here, a novel transcription factor designated AnBH1 was isolated. The corresponding anbH1 gene was cloned and found to be located on chromosome IV. The deduced AnBH1 protein belongs to the family of basic-region helix-loop-helix (bHLH) transcription factors. AnBH1 binds in vitro as a homodimer to an, not previously described, asymmetric E-box within the aatA promoter that overlaps with the AnCF-binding site. This is the first report demonstrating that the CCAAT-binding complex and a bHLH transcription factor bind to overlapping sites. Since deletion of anbH1 appears to be lethal, the anbH1 gene was replaced by a regulatable alcAp-anbH1 gene fusion. The analysis of aatAp-lacZ expression in such a strain indicated that AnBH1 acts as a repressor of aatA gene expression and therefore counteracts the positive action of AnCF.

  2. Identification of a major cis-acting DNA element controlling the bidirectionally transcribed penicillin biosynthesis genes acvA (pcbAB) and ipnA (pcbC) of Aspergillus nidulans.

    PubMed Central

    Bergh, K T; Litzka, O; Brakhage, A A

    1996-01-01

    The beta-lactam antibiotic penicillin is produced as a secondary metabolite by some filamentous fungi. In this study, the molecular regulation of the Aspergillus (Emericella) nidulans penicillin biosynthesis genes acvA (pcbAB) and ipnA (pcbC) was analyzed. acvA and ipnA are divergently oriented and separated by an intergenic region of 872 bp. Translational fusions of acvA and ipnA with the two Escherichia coli reporter genes lacZ and uidA enabled us to measure the regulation of both genes simultaneously. A moving-window analysis of the 872-bp intergenic region indicated that the divergently oriented promoters are, at least in part, overlapping and share common regulatory elements. Removal of nucleotides -353 to -432 upstream of the acvA gene led to a 10-fold increase of acvA-uidA expression and simultaneously to a reduction of ipnA-lacZ expression to about 30%. Band shift assays and methyl interference analysis using partially purified protein extracts revealed that a CCAAT-containing DNA element within this region was specifically bound by a protein (complex), which we designated PENR1, for penicillin regulator. Deletion of 4 bp within the identified protein binding site caused the same contrary effects on acvA and ipnA expression as observed for all of the deletion clones which lacked nucleotides -353 to -432. The PENR1 binding site thus represents a major cis-acting DNA element. The intergenic regions of the corresponding genes of the beta-lactam-producing fungi Penicillium chrysogenum and Acremonium chrysogenum also diluted the complex formed between the A. nidulans probe and PENR1 in vitro, suggesting that these beta-lactam biosynthesis genes are regulated by analogous DNA elements and proteins. PMID:8682797

  3. FigA, a putative homolog of low-affinity calcium system member Fig1 in Saccharomyces cerevisiae, is involved in growth and asexual and sexual development in Aspergillus nidulans.

    PubMed

    Zhang, Shizhu; Zheng, Hailin; Long, Nanbiao; Carbó, Natalia; Chen, Peiying; Aguilar, Pablo S; Lu, Ling

    2014-02-01

    Calcium-mediated signaling pathways are widely employed in eukaryotes and are implicated in the regulation of diverse biological processes. In Saccharomyces cerevisiae, at least two different calcium uptake systems have been identified: the high-affinity calcium influx system (HACS) and the low-affinity calcium influx system (LACS). Compared to the HACS, the LACS in fungi is not well known. In this study, FigA, a homolog of the LACS member Fig1 from S. cerevisiae, was functionally characterized in the filamentous fungus Aspergillus nidulans. Loss of figA resulted in retardant hyphal growth and a sharp reduction of conidial production. Most importantly, FigA is essential for the homothallic mating (self-fertilization) process; further, FigA is required for heterothallic mating (outcrossing) in the absence of HACS midA. Interestingly, in a figA deletion mutant, adding extracellular Ca(2+) rescued the hyphal growth defects but could not restore asexual and sexual reproduction. Furthermore, quantitative PCR results revealed that figA deletion sharply decreased the expression of brlA and nsdD, which are known as key regulators during asexual and sexual development, respectively. In addition, green fluorescent protein (GFP) tagging at the C terminus of FigA (FigA::GFP) showed that FigA localized to the center of the septum in mature hyphal cells, to the location between vesicles and metulae, and between the junctions of metulae and phialides in conidiophores. Thus, our findings suggest that FigA, apart from being a member of a calcium uptake system in A. nidulans, may play multiple unexplored roles during hyphal growth and asexual and sexual development.

  4. FigA, a Putative Homolog of Low-Affinity Calcium System Member Fig1 in Saccharomyces cerevisiae, Is Involved in Growth and Asexual and Sexual Development in Aspergillus nidulans

    PubMed Central

    Zhang, Shizhu; Zheng, Hailin; Long, Nanbiao; Carbó, Natalia; Chen, Peiying; Aguilar, Pablo S.

    2014-01-01

    Calcium-mediated signaling pathways are widely employed in eukaryotes and are implicated in the regulation of diverse biological processes. In Saccharomyces cerevisiae, at least two different calcium uptake systems have been identified: the high-affinity calcium influx system (HACS) and the low-affinity calcium influx system (LACS). Compared to the HACS, the LACS in fungi is not well known. In this study, FigA, a homolog of the LACS member Fig1 from S. cerevisiae, was functionally characterized in the filamentous fungus Aspergillus nidulans. Loss of figA resulted in retardant hyphal growth and a sharp reduction of conidial production. Most importantly, FigA is essential for the homothallic mating (self-fertilization) process; further, FigA is required for heterothallic mating (outcrossing) in the absence of HACS midA. Interestingly, in a figA deletion mutant, adding extracellular Ca2+ rescued the hyphal growth defects but could not restore asexual and sexual reproduction. Furthermore, quantitative PCR results revealed that figA deletion sharply decreased the expression of brlA and nsdD, which are known as key regulators during asexual and sexual development, respectively. In addition, green fluorescent protein (GFP) tagging at the C terminus of FigA (FigA::GFP) showed that FigA localized to the center of the septum in mature hyphal cells, to the location between vesicles and metulae, and between the junctions of metulae and phialides in conidiophores. Thus, our findings suggest that FigA, apart from being a member of a calcium uptake system in A. nidulans, may play multiple unexplored roles during hyphal growth and asexual and sexual development. PMID:24376003

  5. Aspergillus nidulans class V and VI chitin synthases CsmA and CsmB, each with a myosin motor-like domain, perform compensatory functions that are essential for hyphal tip growth.

    PubMed

    Takeshita, Norio; Yamashita, Shuichi; Ohta, Akinori; Horiuchi, Hiroyuki

    2006-03-01

    The polarized synthesis of cell wall components such as chitin is essential for the hyphal tip growth of filamentous fungi. The actin cytoskeleton is known to play important roles in the determination of hyphal polarity in Aspergillus nidulans. Previously, we suggested that CsmA, a chitin synthase with a myosin motor-like domain (MMD), was involved in polarized chitin synthesis in a manner dependent on the interaction between the MMD and the actin cytoskeleton. The genome database indicates that A. nidulans possesses another gene encoding another chitin synthase with an MMD. In this study, we characterized this gene, which we designated csmB. The csmB null mutants examined were viable, although they exhibited defective phenotypes, including the formation of balloons and intrahyphal hyphae and the lysis of subapical regions, which were similar to those obtained with csmA null mutants. Moreover, csmA csmB double null mutants were not viable. Mutants in which csmB was deleted and the expression of csmA was under the control of the alcA promoter were viable but severely impaired in terms of hyphal growth under alcA-repressing conditions. We revealed that CsmB with three copies of a FLAG epitope tag localized at the hyphal tips and forming septa, and that the MMD of CsmB was able to bind to actin filaments in vitro. These results suggest that CsmA and CsmB perform compensatory functions that are essential for hyphal tip growth.

  6. The CreA repressor is the sole DNA-binding protein responsible for carbon catabolite repression of the alcA gene in Aspergillus nidulans via its binding to a couple of specific sites.

    PubMed

    Panozzo, C; Cornillot, E; Felenbok, B

    1998-03-13

    Carbon catabolite repression is mediated in Aspergillus nidulans by the negative acting protein CreA. The CreA repressor plays a major role in the control of the expression of the alc regulon, encoding proteins required for the ethanol utilization pathway. It represses directly, at the transcriptional level, the specific transacting gene alcR, the two structural genes alcA and aldA, and other alc genes in all physiological growth conditions. Among the seven putative CreA sites identified in the alcA promoter region, we have determined the CreA functional targets in AlcR constitutive and derepressed genetic backgrounds. Two different divergent CreA sites, of which one overlaps a functional AlcR inverted repeat site, are largely responsible for alcA repression. Totally derepressed alcA expression is achieved when these two CreA sites are disrupted in addition to another single site, which overlaps the functional palindromic induction target. The fact that derepression is always associated with alcA overexpression is consistent with a competition model between AlcR and CreA for their cognate targets in the same region of the alcA promoter. Our results also indicate that the CreA repressor is necessary and sufficient for the total repression of the alcA gene.

  7. The zinc binuclear cluster activator AlcR is able to bind to single sites but requires multiple repeated sites for synergistic activation of the alcA gene in Aspergillus nidulans.

    PubMed

    Panozzo, C; Capuano, V; Fillinger, S; Felenbok, B

    1997-09-01

    The alcA gene which is part of the recently identified ethanol regulon, is one of the most strongly inducible genes in Aspergillus nidulans. Its transcriptional activation is mediated by the AlcR transactivator which contains a DNA-binding domain belonging to the C6 zinc binuclear cluster family. AlcR differs from the other members of this family by several features, the most striking characteristic being its binding to both symmetric and asymmetric DNA sites with the same apparent affinity. However, AlcR is also able to bind to a single site with high affinity, suggesting that unlike the other C6 proteins, AlcR binds as a monomer. In this report, we show that AlcR targets, to be functional in vivo, have to be organized as inverted or direct repeats. In addition, we show a strong synergistic activation of alcA transcription in which the number and the position of the AlcR-binding sites are crucial. The fact that the AlcR unit for in vitro binding is a single site whereas the in vivo functional unit is a repeat opens the question of the mechanism of the strong alcA transactivation. These results show that AlcR displays both in vitro and in vivo a new range of binding specificity and provides a novel example in the C6 zinc cluster protein family.

  8. AnCF, the CCAAT binding complex of Aspergillus nidulans, contains products of the hapB, hapC, and hapE genes and is required for activation by the pathway-specific regulatory gene amdR.

    PubMed

    Steidl, S; Papagiannopoulos, P; Litzka, O; Andrianopoulos, A; Davis, M A; Brakhage, A A; Hynes, M J

    1999-01-01

    CCAAT binding factors (CBFs) positively regulating the expression of the amdS gene (encoding acetamidase) and two penicillin biosynthesis genes (ipnA and aatA) have been previously found in Aspergillus nidulans. The factors were called AnCF and PENR1, respectively. Deletion of the hapC gene, encoding a protein with significant similarity to Hap3p of Saccharomyces cerevisiae, eliminated both AnCF and PENR1 binding activities. We now report the isolation of the genes hapB and hapE, which encode proteins with central regions of high similarity to Hap2p and Hap5p of S. cerevisiae and to the CBF-B and CBF-C proteins of mammals. An additional fungus-specific domain present in HapE was revealed by comparisons with the homologs from S. cerevisiae, Neurospora crassa, and Schizosaccharomyces pombe. The HapB, HapC, and HapE proteins have been shown to be necessary and sufficient for the formation of a CCAAT binding complex in vitro. Strains with deletions of each of the hapB, hapC, and hapE genes have identical phenotypes of slow growth, poor conidiation, and reduced expression of amdS. Furthermore, induction of amdS by omega amino acids, which is mediated by the AmdR pathway-specific activator, is abolished in the hap deletion mutants, as is growth on gamma-aminobutyric acid as a sole nitrogen or carbon source. AmdR and AnCF bind to overlapping sites in the promoters of the amdS and gatA genes. It is known that AnCF can bind independently of AmdR. We suggest that AnCF binding is required for AmdR binding in vivo.

  9. AnCF, the CCAAT Binding Complex of Aspergillus nidulans, Contains Products of the hapB, hapC, and hapE Genes and Is Required for Activation by the Pathway-Specific Regulatory Gene amdR

    PubMed Central

    Steidl, Stefan; Papagiannopoulos, Peter; Litzka, Olivier; Andrianopoulos, Alex; Davis, Meryl A.; Brakhage, Axel A.; Hynes, Michael J.

    1999-01-01

    CCAAT binding factors (CBFs) positively regulating the expression of the amdS gene (encoding acetamidase) and two penicillin biosynthesis genes (ipnA and aatA) have been previously found in Aspergillus nidulans. The factors were called AnCF and PENR1, respectively. Deletion of the hapC gene, encoding a protein with significant similarity to Hap3p of Saccharomyces cerevisiae, eliminated both AnCF and PENR1 binding activities. We now report the isolation of the genes hapB and hapE, which encode proteins with central regions of high similarity to Hap2p and Hap5p of S. cerevisiae and to the CBF-B and CBF-C proteins of mammals. An additional fungus-specific domain present in HapE was revealed by comparisons with the homologs from S. cerevisiae, Neurospora crassa, and Schizosaccharomyces pombe. The HapB, HapC, and HapE proteins have been shown to be necessary and sufficient for the formation of a CCAAT binding complex in vitro. Strains with deletions of each of the hapB, hapC, and hapE genes have identical phenotypes of slow growth, poor conidiation, and reduced expression of amdS. Furthermore, induction of amdS by omega amino acids, which is mediated by the AmdR pathway-specific activator, is abolished in the hap deletion mutants, as is growth on γ-aminobutyric acid as a sole nitrogen or carbon source. AmdR and AnCF bind to overlapping sites in the promoters of the amdS and gatA genes. It is known that AnCF can bind independently of AmdR. We suggest that AnCF binding is required for AmdR binding in vivo. PMID:9858535

  10. The sequence and binding specificity of UaY, the specific regulator of the purine utilization pathway in Aspergillus nidulans, suggest an evolutionary relationship with the PPR1 protein of Saccharomyces cerevisiae.

    PubMed Central

    Suárez, T; de Queiroz, M V; Oestreicher, N; Scazzocchio, C

    1995-01-01

    The uaY gene codes for a transcriptional activator mediating the induction of a number of unlinked genes involved in purine utilization in Aspergillus nidulans. Here we present the complete genomic and cDNA nucleotide sequence of this gene. The gene contains two introns. The derived polypeptide of 1060 residues contains a typical zinc binuclear cluster domain and shows a number of similarities with the PPR1 regulatory gene of Saccharomyces cerevisiae. These similarities are most striking in the putative linker and dimerization regions following the zinc cluster. Gel-shift and DNase I footprinting experiments have been carried out for three genes subject to UaY-mediated induction. The binding sequence is 5'-TCGG-6X-CCGA, which is identical to the proposed PPR1 binding sites. Nevertheless, the identity of the base immediately 3' of the 5'-TCGG sequence clearly affects the affinity of the site. The site upstream of the uapA gene has been shown to be active in vivo. Binding to this site has been analysed by a number of interference techniques. There is an interesting chemical similarity between the co-inducer of the purine utilization pathway (uric acid) and that of the genes of the pyrimidine biosynthetic pathway (dihydroorotic acid) and we show that dihydroorotic acid can act as a poor inducer of at least one activity under UaY control. These striking similarities, together with the unique pattern of regulation of pyrimidine biosynthesis in S. cerevisiae, suggest that PPR1 evolved through recruitment into the pyrimidine biosynthetic pathway of an ancestral gene related to uaY. Images PMID:7729421

  11. Identification of metabolic pathways influenced by the G-protein coupled receptors GprB and GprD in Aspergillus nidulans.

    PubMed

    de Souza, Wagner R; Morais, Enyara Rezende; Krohn, Nadia Graciele; Savoldi, Marcela; Goldman, Maria Helena S; Rodrigues, Fernando; Caldana, Camila; Semelka, Charles T; Tikunov, Andrey P; Macdonald, Jeffrey M; Goldman, Gustavo Henrique

    2013-01-01

    Heterotrimeric G-protein-mediated signaling pathways play a pivotal role in transmembrane signaling in eukaryotes. Our main aim was to identify signaling pathways regulated by A. nidulans GprB and GprD G-protein coupled receptors (GPCRs). When these two null mutant strains were compared to the wild-type strain, the ΔgprB mutant showed an increased protein kinase A (PKA) activity while growing in glucose 1% and during starvation. In contrast, the ΔgprD has a much lower PKA activity upon starvation. Transcriptomics and (1)H NMR-based metabolomics were performed on two single null mutants grown on glucose. We noted modulation in the expression of 11 secondary metabolism gene clusters when the ΔgprB and ΔgprD mutant strains were grown in 1% glucose. Several members of the sterigmatocystin-aflatoxin gene cluster presented down-regulation in both mutant strains. The genes of the NR-PKS monodictyphenone biosynthesis cluster had overall increased mRNA accumulation in ΔgprB, while in the ΔgprD mutant strain the genes had decreased mRNA accumulation. Principal component analysis of the metabolomic data demonstrated that there was a significant metabolite shift in the ΔgprD strain. The (1)H NMR analysis revealed significant expression of essential amino acids with elevated levels in the ΔgprD strain, compared to the wild-type and ΔgprB strains. With the results, we demonstrated the differential expression of a variety of genes related mainly to secondary metabolism, sexual development, stress signaling, and amino acid metabolism. We propose that the absence of GPCRs triggered stress responses at the genetic level. The data suggested an intimate relationship among different G-protein coupled receptors, fine-tune regulation of secondary and amino acid metabolisms, and fungal development.

  12. Purification and physico-chemical characterisation of genetically modified phytases expressed in Aspergillus awamori.

    PubMed

    Martin, Judith A; Murphy, Richard A; Power, Ronan F G

    2006-09-01

    Two heterologous phytases from Aspergillus awamori and Aspergillus fumigatus obtained from submerged cultures of genetically modified fungal strains in addition to two commercially available phytase preparations (Allzyme and Natuphos phytases) were purified to homogeneity using a combination of ultrafiltration, gel filtration and ion exchange. The purified preparations were used in subsequent characterisation studies, in which Western Immunoblot analysis, pH and temperature optima, thermal stability and substrate specificity were assessed. A. fumigatus phyA phytase expressed in A. awamori exhibited activity over a broad pH range together with an increased temperature optimum, and slightly enhanced thermal stability compared to the other phytases tested, and is thus a promising candidate for animal feed applications. This particular phytase retains activity over a wide range of pH values characteristic of the digestive tract and could conceivably be more suited to the increasingly higher feed processing temperatures being utilised today, than the corresponding phytases from Aspergillus niger. PMID:16243522

  13. Positive Regulation in a Eukaryote, a Study of the uaY Gene of ASPERGILLUS NIDULANS: I. Characterization of Alleles, Dominance and Complementation Studies, and a Fine Structure Map of the uaY -oxpA Cluster

    PubMed Central

    Scazzocchio, Claudio; Sdrin, Nicholas; Ong, Gloria

    1982-01-01

    In this paper we characterize genetically a positive eukaryotic regulatory gene: the uaY gene of the ascomycete Aspergillus nidulans. Several steps in the uptake and degradation of purines are under the control of the uaY gene (summarized in Scazzocchio and Gorton 1977). In the present paper 12 uaY- mutations are characterized with respect to their inducibility for adenine deaminase, xanthine dehydrogenase (purine hydroxylase I) and urate oxidase and by the absence of the uric acid-xanthine permease scored in vivo by resistance to 2-thiouric acid. While 10 mutations are uniformly unleaky, two others are almost wild type for the induction of urate oxidase. A fine structure map of the uaY gene shows that the two "leaky" mutations are not clustered. The fine structure mapping unambiguously positions six uaY alleles and provides preliminary but interesting trends regarding the pattern of gene conversion in the uaY gene. The enzyme levels in all uaY-/uaY+ heterozygous diploids are intermediate between the corresponding uaY-/uaY- and uaY+/uaY+ homozygous diploids, suggesting that one functional copy of the uaY gene is able to mediate the complete induction of only one set of structural genes. No complementation was found between any two uaY- alleles. This establishes that the mutations showing either of the phenotypes are alleles in the same gene; it fails to provide evidence for intracistronic complementation. A mutation, oxpA5, causes resistance to the xanthine analogue oxypurinol (4, 6-dihydroxypyrazolo-(3, 4-d)-pyrimidine) and partial constitutivity of adenine deaminase, xanthine dehydrogenase (purine hydroxylase I) and urate oxidase. The constitutive phenotype is suppressed by mutations blocking the synthesis of intracellular inducers. The mutation is recessive and complements fully with the 11 uaY- mutations tested. It maps to the left of all 12 uaY mutations to which it has been crossed. The data indicate that both the resistance and constitutivity arise from one

  14. Nitrogen Metabolism and Growth Enhancement in Tomato Plants Challenged with Trichoderma harzianum Expressing the Aspergillus nidulans Acetamidase amdS Gene

    PubMed Central

    Domínguez, Sara; Rubio, M. Belén; Cardoza, Rosa E.; Gutiérrez, Santiago; Nicolás, Carlos; Bettiol, Wagner; Hermosa, Rosa; Monte, Enrique

    2016-01-01

    Trichoderma is a fungal genus that includes species that are currently being used as biological control agents and/or as biofertilizers. In addition to the direct application of Trichoderma spp. as biocontrol agents in plant protection, recent studies have focused on the beneficial responses exerted on plants, stimulating the growth, activating the defenses, and/or improving nutrient uptake. The amdS gene, encoding an acetamidase of Aspergillus, has been used as a selectable marker for the transformation of filamentous fungi, including Trichoderma spp., but the physiological effects of the introduction of this gene into the genome of these microorganisms still remains unexplored. No evidence of amdS orthologous genes has been detected within the Trichoderma spp. genomes and the amdS heterologous expression in Trichoderma harzianum T34 did not affect the growth of this fungus in media lacking acetamide. However, it did confer the ability for the fungus to use this amide as a nitrogen source. Although a similar antagonistic behavior was observed for T34 and amdS transformants in dual cultures against Rhizoctonia solani, Botrytis cinerea, and Fusarium oxysporum, a significantly higher antifungal activity was detected in amdS transformants against F. oxysporum, compared to that of T34, in membrane assays on media lacking acetamide. In Trichoderma-tomato interaction assays, amdS transformants were able to promote plant growth to a greater extent than the wild-type T34, although compared with this strain the transformants showed similar capability to colonize tomato roots. Gene expression patterns from aerial parts of 3-week-old tomato plants treated with T34 and the amdS transformants have also been investigated using GeneChip Tomato Genome Arrays. The downregulation of defense genes and the upregulation of carbon and nitrogen metabolism genes observed in the microarrays were accompanied by (i) enhanced growth, (ii) increased carbon and nitrogen levels, and (iii) a

  15. Nitrogen Metabolism and Growth Enhancement in Tomato Plants Challenged with Trichoderma harzianum Expressing the Aspergillus nidulans Acetamidase amdS Gene.

    PubMed

    Domínguez, Sara; Rubio, M Belén; Cardoza, Rosa E; Gutiérrez, Santiago; Nicolás, Carlos; Bettiol, Wagner; Hermosa, Rosa; Monte, Enrique

    2016-01-01

    Trichoderma is a fungal genus that includes species that are currently being used as biological control agents and/or as biofertilizers. In addition to the direct application of Trichoderma spp. as biocontrol agents in plant protection, recent studies have focused on the beneficial responses exerted on plants, stimulating the growth, activating the defenses, and/or improving nutrient uptake. The amdS gene, encoding an acetamidase of Aspergillus, has been used as a selectable marker for the transformation of filamentous fungi, including Trichoderma spp., but the physiological effects of the introduction of this gene into the genome of these microorganisms still remains unexplored. No evidence of amdS orthologous genes has been detected within the Trichoderma spp. genomes and the amdS heterologous expression in Trichoderma harzianum T34 did not affect the growth of this fungus in media lacking acetamide. However, it did confer the ability for the fungus to use this amide as a nitrogen source. Although a similar antagonistic behavior was observed for T34 and amdS transformants in dual cultures against Rhizoctonia solani, Botrytis cinerea, and Fusarium oxysporum, a significantly higher antifungal activity was detected in amdS transformants against F. oxysporum, compared to that of T34, in membrane assays on media lacking acetamide. In Trichoderma-tomato interaction assays, amdS transformants were able to promote plant growth to a greater extent than the wild-type T34, although compared with this strain the transformants showed similar capability to colonize tomato roots. Gene expression patterns from aerial parts of 3-week-old tomato plants treated with T34 and the amdS transformants have also been investigated using GeneChip Tomato Genome Arrays. The downregulation of defense genes and the upregulation of carbon and nitrogen metabolism genes observed in the microarrays were accompanied by (i) enhanced growth, (ii) increased carbon and nitrogen levels, and (iii) a

  16. Nitrogen Metabolism and Growth Enhancement in Tomato Plants Challenged with Trichoderma harzianum Expressing the Aspergillus nidulans Acetamidase amdS Gene.

    PubMed

    Domínguez, Sara; Rubio, M Belén; Cardoza, Rosa E; Gutiérrez, Santiago; Nicolás, Carlos; Bettiol, Wagner; Hermosa, Rosa; Monte, Enrique

    2016-01-01

    Trichoderma is a fungal genus that includes species that are currently being used as biological control agents and/or as biofertilizers. In addition to the direct application of Trichoderma spp. as biocontrol agents in plant protection, recent studies have focused on the beneficial responses exerted on plants, stimulating the growth, activating the defenses, and/or improving nutrient uptake. The amdS gene, encoding an acetamidase of Aspergillus, has been used as a selectable marker for the transformation of filamentous fungi, including Trichoderma spp., but the physiological effects of the introduction of this gene into the genome of these microorganisms still remains unexplored. No evidence of amdS orthologous genes has been detected within the Trichoderma spp. genomes and the amdS heterologous expression in Trichoderma harzianum T34 did not affect the growth of this fungus in media lacking acetamide. However, it did confer the ability for the fungus to use this amide as a nitrogen source. Although a similar antagonistic behavior was observed for T34 and amdS transformants in dual cultures against Rhizoctonia solani, Botrytis cinerea, and Fusarium oxysporum, a significantly higher antifungal activity was detected in amdS transformants against F. oxysporum, compared to that of T34, in membrane assays on media lacking acetamide. In Trichoderma-tomato interaction assays, amdS transformants were able to promote plant growth to a greater extent than the wild-type T34, although compared with this strain the transformants showed similar capability to colonize tomato roots. Gene expression patterns from aerial parts of 3-week-old tomato plants treated with T34 and the amdS transformants have also been investigated using GeneChip Tomato Genome Arrays. The downregulation of defense genes and the upregulation of carbon and nitrogen metabolism genes observed in the microarrays were accompanied by (i) enhanced growth, (ii) increased carbon and nitrogen levels, and (iii) a

  17. Cryptic Sexuality in Aspergillus parasiticus and A. flavus

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Ascomycetous fungi of the genus Aspergillus comprise a wide variety of species of biotechnological importance (e.g. A. sojae, A. oryzae, A. niger) as well as pathogens and toxin producers (e.g. A. flavus, A. parasiticus, A. fumigatus, A. nidulans). With the exception of A. nidulans, which is a homot...

  18. Decolourization and detoxification of pulp and paper mill effluent by Emericella nidulans var. nidulans.

    PubMed

    Singhal, Anjali; Thakur, Indu Shekhar

    2009-11-15

    In this study geno-toxicity analysis along with effluent treatment was taken up to evaluate the efficiency of biological treatment process for safe disposal of treated effluent. Four fungi were isolated from sediments of pulp and paper mill in which PF4 reduced colour (30%) and lignin content (24%) of the effluent on 3rd day. The fungal strain was identified as Emericella nidulans var. nidulans (anamorph: Aspergillus nidulans) on the basis of rDNA ITS1 and rDNA ITS2 region sequences. The process of decolourization was optimized by Taguchi approach. The optimum conditions were temperature (30-35 degrees C), rpm (125), dextrose (0.25%), tryptone (0.1%), inoculum size (7.5%), pH (5) and duration (24h). Decolourization of effluent improved by 31% with reduction in colour (66.66%) and lignin (37%) after treatment by fungi in shake flask. Variation in pH from 6 to 5 had most significant effect on decolourization (71%) while variation in temperature from 30 to 35 degrees C had no effect on the process. Treated effluent was further evaluated for geno-toxicity by alkaline single cell gel electrophoresis (SCGE) assay using Saccharomyces cerevisiae MTCC 36 as model organism, indicated 60% reduction. PMID:19586717

  19. Development of RFLP-PCR method for the identification of medically important Aspergillus species using single restriction enzyme MwoI.

    PubMed

    Diba, K; Mirhendi, H; Kordbacheh, P; Rezaie, S

    2014-01-01

    In this study we attempted to modify the PCR-RFLP method using restriction enzyme MwoI for the identification of medically important Aspergillus species. Our subjects included nine standard Aspergillus species and 205 Aspergillus isolates of approved hospital acquired infections and hospital indoor sources. First of all, Aspergillus isolates were identified in the level of species by using morphologic method. A twenty four hours culture was performed for each isolates to harvest Aspergillus mycelia and then genomic DNA was extracted using Phenol-Chloroform method. PCR-RFLP using single restriction enzyme MwoI was performed in ITS regions of rDNA gene. The electrophoresis data were analyzed and compared with those of morphologic identifications. Total of 205 Aspergillus isolates included 153 (75%) environmental and 52 (25%) clinical isolates. A. flavus was the most frequently isolate in our study (55%), followed by A. niger 65(31.7%), A. fumigatus 18(8.7%), A. nidulans and A. parasiticus 2(1% each). MwoI enabled us to discriminate eight medically important Aspergillus species including A. fumigatus, A. niger, A. flavus as the most common isolated species. PCR-RFLP method using the restriction enzyme MwoI is a rapid and reliable test for identification of at least the most medically important Aspergillus species.

  20. KdmB, a Jumonji Histone H3 Demethylase, Regulates Genome-Wide H3K4 Trimethylation and Is Required for Normal Induction of Secondary Metabolism in Aspergillus nidulans.

    PubMed

    Gacek-Matthews, Agnieszka; Berger, Harald; Sasaki, Takahiko; Wittstein, Kathrin; Gruber, Clemens; Lewis, Zachary A; Strauss, Joseph

    2016-08-01

    Histone posttranslational modifications (HPTMs) are involved in chromatin-based regulation of fungal secondary metabolite biosynthesis (SMB) in which the corresponding genes-usually physically linked in co-regulated clusters-are silenced under optimal physiological conditions (nutrient-rich) but are activated when nutrients are limiting. The exact molecular mechanisms by which HPTMs influence silencing and activation, however, are still to be better understood. Here we show by a combined approach of quantitative mass spectrometry (LC-MS/MS), genome-wide chromatin immunoprecipitation (ChIP-seq) and transcriptional network analysis (RNA-seq) that the core regions of silent A. nidulans SM clusters generally carry low levels of all tested chromatin modifications and that heterochromatic marks flank most of these SM clusters. During secondary metabolism, histone marks typically associated with transcriptional activity such as H3 trimethylated at lysine-4 (H3K4me3) are established in some, but not all gene clusters even upon full activation. KdmB, a Jarid1-family histone H3 lysine demethylase predicted to comprise a BRIGHT domain, a zinc-finger and two PHD domains in addition to the catalytic Jumonji domain, targets and demethylates H3K4me3 in vivo and mediates transcriptional downregulation. Deletion of kdmB leads to increased transcription of about ~1750 genes across nutrient-rich (primary metabolism) and nutrient-limiting (secondary metabolism) conditions. Unexpectedly, an equally high number of genes exhibited reduced expression in the kdmB deletion strain and notably, this group was significantly enriched for genes with known or predicted functions in secondary metabolite biosynthesis. Taken together, this study extends our general knowledge about multi-domain KDM5 histone demethylases and provides new details on the chromatin-level regulation of fungal secondary metabolite production. PMID:27548260

  1. KdmB, a Jumonji Histone H3 Demethylase, Regulates Genome-Wide H3K4 Trimethylation and Is Required for Normal Induction of Secondary Metabolism in Aspergillus nidulans

    PubMed Central

    Gacek-Matthews, Agnieszka; Sasaki, Takahiko; Wittstein, Kathrin; Gruber, Clemens; Strauss, Joseph

    2016-01-01

    Histone posttranslational modifications (HPTMs) are involved in chromatin-based regulation of fungal secondary metabolite biosynthesis (SMB) in which the corresponding genes—usually physically linked in co-regulated clusters—are silenced under optimal physiological conditions (nutrient-rich) but are activated when nutrients are limiting. The exact molecular mechanisms by which HPTMs influence silencing and activation, however, are still to be better understood. Here we show by a combined approach of quantitative mass spectrometry (LC-MS/MS), genome-wide chromatin immunoprecipitation (ChIP-seq) and transcriptional network analysis (RNA-seq) that the core regions of silent A. nidulans SM clusters generally carry low levels of all tested chromatin modifications and that heterochromatic marks flank most of these SM clusters. During secondary metabolism, histone marks typically associated with transcriptional activity such as H3 trimethylated at lysine-4 (H3K4me3) are established in some, but not all gene clusters even upon full activation. KdmB, a Jarid1-family histone H3 lysine demethylase predicted to comprise a BRIGHT domain, a zinc-finger and two PHD domains in addition to the catalytic Jumonji domain, targets and demethylates H3K4me3 in vivo and mediates transcriptional downregulation. Deletion of kdmB leads to increased transcription of about ~1750 genes across nutrient-rich (primary metabolism) and nutrient-limiting (secondary metabolism) conditions. Unexpectedly, an equally high number of genes exhibited reduced expression in the kdmB deletion strain and notably, this group was significantly enriched for genes with known or predicted functions in secondary metabolite biosynthesis. Taken together, this study extends our general knowledge about multi-domain KDM5 histone demethylases and provides new details on the chromatin-level regulation of fungal secondary metabolite production. PMID:27548260

  2. rmtA, encoding a putative anginine methyltransferase, regulates secondary metabolism and development in Aspergillus flavus

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Aspergillus flavus is found colonizing numerous oil seed crops such as corn, peanuts, sorghum, treenuts and cotton worldwide, contaminating them with aflatoxin and other harmful potent toxins. In the phylogenetically related model fungus Aspergillus nidulans, the methyltransferase, RmtA, has been de...

  3. Cloning and expression of fungal phytases in genetically modified strains of Aspergillus awamori.

    PubMed

    Martin, Judith A; Murphy, Richard A; Power, Ronan F G

    2003-09-01

    In an effort to produce phytases cost-effectively, and to determine the efficiency of a novel expression system, the genes for Aspergillus awamori ( phyA) phytase and Aspergillus fumigatus ( phyA) phytase (a putative thermostable enzyme) were cloned and overexpressed in A. awamori. Regulation of phytase expression was achieved by separately placing the genes under the transcriptional control of the glucoamylase A ( glaA) promoter of A. awamori. A gene fusion strategy was employed that involved the insertion of a hexapeptide Kex-2 protease cleavage site between the native glucoamylase and heterologous proteins and allowed for the efficient secretion and processing of the resultant chimeric proteins produced in this system by an endogenous Kex-2 protease. The genes for both of the above-mentioned phytases have already been cloned; however, this is the first report of either of the two phytases being fused with the glucoamylase gene, placed under the transcriptional control of the glaA promoter and overexpressed in A. awamori. Following transformation of A. awamori with separate expression vectors (one for each phytase), induction of phytase expression in submerged culture was effected by utilisation of a starch-containing medium. Optimisation of heterologous protein production in small shake-flask cultures involved changes in medium constituents. Maximum phytase expression levels of 200 phytase units (PU) ml(-1) and 62 PU ml(-1) for recombinantly expressed phytases from A. awamori and A. fumigatus, respectively, were obtained in crude fermentation extracts. Subsequent process scale-up to 4 l batch fermentation yielded phytase production levels comparable to those obtained on small scale. The enzyme yields herein reported demonstrate that the expression system developed and the host strain utilised were capable of expressing phytase at levels comparable to, or exceeding, previously reported data. PMID:14513384

  4. RNA sequencing of an nsdC mutant reveals global regulation of secondary metabolic gene clusters in Aspergillus flavus

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The zinc finger transcription factor nsdC is required for both sexual development and aflatoxin production in the saprophytic fungus Aspergillus flavus. While previous work with an nsdC knockout mutant was conducted in Aspergillus nidulans and A. flavus strain 3357, here we demonstrate perturbations...

  5. Verruculogen associated with Aspergillus fumigatus hyphae and conidia modifies the electrophysiological properties of human nasal epithelial cells

    PubMed Central

    Khoufache, Khaled; Puel, Olivier; Loiseau, Nicolas; Delaforge, Marcel; Rivollet, Danièle; Coste, André; Cordonnier, Catherine; Escudier, Estelle; Botterel, Françoise; Bretagne, Stéphane

    2007-01-01

    Background The role of Aspergillus fumigatus mycotoxins in the colonization of the respiratory tract by conidia has not been studied extensively, even though patients at risk from invasive aspergillosis frequently exhibit respiratory epithelium damage. In a previous study, we found that filtrates of A. fumigatus cultures can specifically alter the electrophysiological properties of human nasal epithelial cells (HNEC) compared to those of non pathogenic moulds. Results We fractionated the organic phase of filtrate from 3-day old A. fumigatus cultures using high-performance liquid chromatography. The different fractions were tested for their ability to modify the electrophysiological properties of HNEC in an in vitro primary culture model. The fraction collected between 20 and 30 min mimicked the effects of the whole filtrate, i.e. decrease of transepithelial resistance and increase of potential differences, and contained secondary metabolites such as helvolic acid, fumagillin, and verruculogen. Only verruculogen (10-8 M) had effects similar to the whole filtrate. We verified that verruculogen was produced by a collection of 67 human, animal, plant and environmental A. fumigatus isolates. Using MS-MS analysis, we found that verruculogen was associated with both mycelium and conidia extracts. Conclusion Verruculogen is a secondary metabolite that modifies the electrophysiological properties of HNEC. The role of these modifications in the colonization and invasion of the respiratory epithelium by A. fumigatus on first contact with the epithelium remains to be determined. PMID:17244350

  6. The role of thiol species in the hypertolerance of Aspergillus sp. P37 to arsenic.

    PubMed

    Cánovas, David; Vooijs, Riet; Schat, Henk; de Lorenzo, Víctor

    2004-12-01

    Aspergillus sp. P37 is an arsenate-hypertolerant fungus isolated from a river in Spain with a long history of contamination with metals. This strain is able to grow in the presence of 0.2 M arsenate, i.e. 20-fold higher than the reference strain, Aspergillus nidulans TS1. Although Aspergillus sp. P37 reduces As(V) to As(III), which is slowly pumped out of the cell, the measured efflux of oxyanions is insufficient to explain the high tolerance levels of this strain. To gain an insight into this paradox, the accumulation of acid-soluble thiol species in Aspergillus sp. P37 when exposed to arsenic was compared with that of the arsenic-sensitive A. nidulans TS1 strain. Increasing levels of arsenic in the medium did not diminish the intracellular pool of reduced glutathione in Aspergillus sp. P37, in sharp contrast with the decline of glutathione in A. nidulans under the same conditions. Furthermore, concentrations of arsenic that were inhibitory for the sensitive A. nidulans strain (e.g. 50 mM and above) provoked a massive formation of vacuoles filled with thiol species. Because the major fraction of the cellular arsenic was present as the glutathione conjugate As(GS)3, it is plausible that the arsenic-hypertolerant phenotype of Aspergillus sp. P37 is in part due to an enhanced capacity to maintain a large intracellular glutathione pool under conditions of arsenic exposure and to sequester As(GS)3 in vacuoles. High pressure liquid chromatography analysis of cell extracts revealed that the contact of Aspergillus sp. P37 (but not A. nidulans) with high arsenic concentrations (> or =150 mM) induced the production of small quantities of a distinct thiol species indistinguishable from plant phytochelatin-2. Yet, we argue that phytochelatins do not explain arsenic resistance in Aspergillus, and we advocate the role of As(GS)3 complexes in arsenic detoxification.

  7. The role of thiol species in the hypertolerance of Aspergillus sp. P37 to arsenic.

    PubMed

    Cánovas, David; Vooijs, Riet; Schat, Henk; de Lorenzo, Víctor

    2004-12-01

    Aspergillus sp. P37 is an arsenate-hypertolerant fungus isolated from a river in Spain with a long history of contamination with metals. This strain is able to grow in the presence of 0.2 M arsenate, i.e. 20-fold higher than the reference strain, Aspergillus nidulans TS1. Although Aspergillus sp. P37 reduces As(V) to As(III), which is slowly pumped out of the cell, the measured efflux of oxyanions is insufficient to explain the high tolerance levels of this strain. To gain an insight into this paradox, the accumulation of acid-soluble thiol species in Aspergillus sp. P37 when exposed to arsenic was compared with that of the arsenic-sensitive A. nidulans TS1 strain. Increasing levels of arsenic in the medium did not diminish the intracellular pool of reduced glutathione in Aspergillus sp. P37, in sharp contrast with the decline of glutathione in A. nidulans under the same conditions. Furthermore, concentrations of arsenic that were inhibitory for the sensitive A. nidulans strain (e.g. 50 mM and above) provoked a massive formation of vacuoles filled with thiol species. Because the major fraction of the cellular arsenic was present as the glutathione conjugate As(GS)3, it is plausible that the arsenic-hypertolerant phenotype of Aspergillus sp. P37 is in part due to an enhanced capacity to maintain a large intracellular glutathione pool under conditions of arsenic exposure and to sequester As(GS)3 in vacuoles. High pressure liquid chromatography analysis of cell extracts revealed that the contact of Aspergillus sp. P37 (but not A. nidulans) with high arsenic concentrations (> or =150 mM) induced the production of small quantities of a distinct thiol species indistinguishable from plant phytochelatin-2. Yet, we argue that phytochelatins do not explain arsenic resistance in Aspergillus, and we advocate the role of As(GS)3 complexes in arsenic detoxification. PMID:15364940

  8. PCR-RFLP on β-tubulin gene for rapid identification of the most clinically important species of Aspergillus.

    PubMed

    Nasri, Tuba; Hedayati, Mohammad Taghi; Abastabar, Mahdi; Pasqualotto, Alessandro C; Armaki, Mojtaba Taghizadeh; Hoseinnejad, Akbar; Nabili, Mojtaba

    2015-10-01

    Aspergillus species are important agents of life-threatening infections in immunosuppressed patients. Proper speciation in the Aspergilli has been justified based on varied fungal virulence, clinical presentations, and antifungal resistance. Accurate identification of Aspergillus species usually relies on fungal DNA sequencing but this requires expensive equipment that is not available in most clinical laboratories. We developed and validated a discriminative low-cost PCR-based test to discriminate Aspergillus isolates at the species level. The Beta tubulin gene of various reference strains of Aspergillus species was amplified using the universal fungal primers Bt2a and Bt2b. The PCR products were subjected to digestion with a single restriction enzyme AlwI. All Aspergillus isolates were subjected to DNA sequencing for final species characterization. The PCR-RFLP test generated unique patterns for six clinically important Aspergillus species, including Aspergillus flavus, Aspergillus fumigatus, Aspergillus nidulans, Aspergillus terreus, Aspergillus clavatus and Aspergillus nidulans. The one-enzyme PCR-RFLP on Beta tubulin gene designed in this study is a low-cost tool for the reliable and rapid differentiation of the clinically important Aspergillus species.

  9. Aspergillus fumigatus densities in relation to forest succession and edge effects: implications for wildlife health in modified environments.

    PubMed

    Perrott, John K; Armstrong, Doug P

    2011-09-01

    The hihi (or stitchbird, Notiomystis cincta) is a New Zealand endemic nectivorous forest bird now restricted to one pristine island. Relocation to establish viable hihi populations on other islands has been the main conservation action since the early 1980s. To date, hihi reintroductions to young growth islands have had poor success despite the absence of mammalian predators. It was thought that past failures were due to food limitation, but research suggests that food limitation alone cannot account for their poor survivorship. Post-mortems of dead hihi has shown that aspergillosis caused by Aspergillus fumigatus is a major mortality factor and there is current concern regarding their susceptibility to this fungal disease. In this paper we develop and assess the hypothesis that A. fumigatus limits hihi population viability on modified islands, and suggest that A. fumigatus is a potential indicator species for habitat disturbance. We report that the prevalence of A. fumigatus spores in the soil is much higher in young growth forests and forest edge habitats. Results suggest that hihi mortality rates between islands are potentially due to differential exposure to A. fumigatus spores. We assess relationships between habitat disturbance, A. fumigatus contamination and hihi mortality rates by testing the following predictions: (1) that densities of A. fumigatus spores will be higher on modified islands, (2) that densities of A. fumigatus spores on islands will be correlated with hihi mortality rates and (3) that densities of A. fumigatus spores will be higher at the forest edge than in the interior. We test each of these predictions using soil samples, air samples and samples of nectar from plant species fed on by hihi. PMID:22076057

  10. Negative regulation and developmental competence in Aspergillus

    PubMed Central

    Lee, Mi-Kyung; Kwon, Nak-Jung; Lee, Im-Soon; Jung, Seunho; Kim, Sun-Chang; Yu, Jae-Hyuk

    2016-01-01

    Asexual development (conidiation) in the filamentous fungus Aspergillus nidulans is governed by orchestrated gene expression. The three key negative regulators of conidiation SfgA, VosA, and NsdD act at different control point in the developmental genetic cascade. Here, we have revealed that NsdD is a key repressor affecting the quantity of asexual spores in Aspergillus. Moreover, nullifying both nsdD and vosA results in abundant formation of the development specific structure conidiophores even at 12 h of liquid culture, and near constitutive activation of conidiation, indicating that acquisition of developmental competence involves the removal of negative regulation exerted by both NsdD and VosA. NsdD’s role in repressing conidiation is conserved in other aspergilli, as deleting nsdD causes enhanced and precocious activation of conidiation in Aspergillus fumigatus or Aspergillus flavus. In vivo NsdD-DNA interaction analyses identify three NsdD binding regions in the promoter of the essential activator of conidiation brlA, indicating a direct repressive role of NsdD in conidiation. Importantly, loss of flbC or flbD encoding upstream activators of brlA in the absence of nsdD results in delayed activation of brlA, suggesting distinct positive roles of FlbC and FlbD in conidiation. A genetic model depicting regulation of conidiation in A. nidulans is presented. PMID:27364479

  11. Inducible RNA Interference of brlAβ in Aspergillus nidulans▿

    PubMed Central

    Barton, L. M.; Prade, R. A.

    2008-01-01

    An inducible RNA interference (RNAi) construct composed of inverted repeating alcA promoters flanking the developmental regulatory gene brlAβ was tested in Aspergillus nidulans. On inducing medium, the RNAi strains failed to sporulate and lacked brlAα and brlAβ expression. RNAi was specific for brlAβ, but not brlAα, silencing, indicating brlAα regulation by brlAβ. PMID:18757565

  12. Effect of culture temperature on the heterologous expression of Pleurotus eryngii versatile peroxidase in Aspergillus hosts.

    PubMed

    Eibes, G M; Lú-Chau, T A; Ruiz-Dueñas, F J; Feijoo, G; Martínez, M J; Martínez, A T; Lema, J M

    2009-01-01

    Production of recombinant versatile peroxidase in Aspergillus hosts was optimized through the modification of temperature during bioreactor cultivations. To further this purpose, the cDNA encoding a versatile peroxidase of Pleurotus eryngii was expressed under control of the alcohol dehydrogenase (alcA) promoter of Aspergillus nidulans. A dependence of recombinant peroxidase production on cultivation temperature was found. Lowering the culture temperature from 28 to 19 degrees C enhanced the level of active peroxidase 5.8-fold and reduced the effective proteolytic activity twofold. Thus, a maximum peroxidase activity of 466 U L(-1) was reached. The same optimization scheme was applied to a recombinant Aspergillus niger that bore the alcohol dehydrogenase regulator (alcR), enabling transformation with the peroxidase cDNA under the same alcA promoter. However, with this strain, the peroxidase activity was not improved, while the effective proteolytic activity was increased between 3- and 11-fold compared to that obtained with A. nidulans.

  13. Phosphate solubilizing ability of Emericella nidulans strain V1 isolated from vermicompost.

    PubMed

    Bhattacharya, Satya Sunder; Barman, Soma; Ghosh, Ranjan; Duary, Raj Kumar; Goswami, Linee; Mandal, Narayan C

    2013-10-01

    Phosphorus is one of the key factors that regulate soil fertility. Its deficiencies in soil are largely replenished by chemical fertilizers. The present study was aimed to isolate efficient phosphate solubilizing fungal strains from Eisenia fetida vermicompost. Out of total 30 fungal strains the most efficient phosphate solubilizing one was Emericella (Aspergillus) nidulans V1 (MTCC 11044), identified by custom sequencing of beta-tubulin gene and BLAST analysis. This strain solubilized 13 to 36% phosphate from four different rock phosphates. After three days of incubation of isolated culture with black Mussorie phosphate rock, the highest percentage of phosphate solubilization was 35.5 +/- 1.01 with a pH drop of 4.2 +/- 0.09. Kinetics of solubilization and acid production showed a linear relationship until day five of incubation. Interestingly, from zero to tenth day of incubation, solubility of soil phosphate increased gradually from 4.31 +/- 1.57 to 13.65 +/- 1.82 (mg kg(-1)) recording a maximum of 21.23 +/- 0.54 on day 45 in respect of the V1 isolate. Further, enhanced phosphorus uptake by Phaseolus plants with significant pod yield due to soil inoculation of Emericella nidulans V1 (MTCC 11044), demonstrated its prospect as an effective biofertilizer for plant growth.

  14. Localization of pyruvate carboxylase in organic acid-producing Aspergillus strains.

    PubMed

    Bercovitz, A; Peleg, Y; Battat, E; Rokem, J S; Goldberg, I

    1990-06-01

    The localization of pyruvate carboxylase (cytosolic or mitochondrial) was studied in nine different Aspergillus species (14 strains). In some species (A. aculeatus, A. flavus, A. foetidus, A. nidulans, A. ochraceus, and A. sojae), the pyruvate carboxylase activity could be detected only in the cytosolic fraction of the cells. Pyruvate carboxylase has been found only in the mitochondrial fraction of two strains of Aspergillus wentii. In Aspergillus oryzae and in five strains of Aspergillus niger, pyruvate carboxylase activity was detected both in the mitochondrial fraction and in the cytosol. There was no quantitative or qualitative correlation between the activities of pyruvate carboxylase in the mitochondrial and cytosolic fractions of the cells and the ability of the various Aspergillus strains to accumulate different organic acids.

  15. rtfA, a putative RNA-Pol II transcription elongation factor, is necessary for normal morphological and chemical development in Aspergillus flavus

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The filamentous fungus Aspergillus flavus is an agriculturally important opportunistic plant pathogen that produces potent carcinogenic compounds called aflatoxins. We identified the A. flavus rtfA gene, the ortholog of rtf1 in S. cerevisiae and rtfA in A. nidulans. Interestingly, rtfA has multiple ...

  16. A Genomics Based Discovery of Secondary Metabolite Biosynthetic Gene Clusters in Aspergillus ustus

    PubMed Central

    Pi, Borui; Yu, Dongliang; Dai, Fangwei; Song, Xiaoming; Zhu, Congyi; Li, Hongye; Yu, Yunsong

    2015-01-01

    Secondary metabolites (SMs) produced by Aspergillus have been extensively studied for their crucial roles in human health, medicine and industrial production. However, the resulting information is almost exclusively derived from a few model organisms, including A. nidulans and A. fumigatus, but little is known about rare pathogens. In this study, we performed a genomics based discovery of SM biosynthetic gene clusters in Aspergillus ustus, a rare human pathogen. A total of 52 gene clusters were identified in the draft genome of A. ustus 3.3904, such as the sterigmatocystin biosynthesis pathway that was commonly found in Aspergillus species. In addition, several SM biosynthetic gene clusters were firstly identified in Aspergillus that were possibly acquired by horizontal gene transfer, including the vrt cluster that is responsible for viridicatumtoxin production. Comparative genomics revealed that A. ustus shared the largest number of SM biosynthetic gene clusters with A. nidulans, but much fewer with other Aspergilli like A. niger and A. oryzae. These findings would help to understand the diversity and evolution of SM biosynthesis pathways in genus Aspergillus, and we hope they will also promote the development of fungal identification methodology in clinic. PMID:25706180

  17. Inactivation of the lipoxygenase ZmLOX3 increases susceptibility of maize to Aspergillus spp

    PubMed Central

    Gao, Xiquan; Brodhagen, Marion; Isakeit, Tom; Brown, Sigal Horowitz; Göbel, Cornelia; Betran, Javier; Feussner, Ivo; Keller, Nancy P.; Kolomiets, Michael V.

    2009-01-01

    Plant and fungal lipoxygenases catalyze the oxidation of polyunsaturated fatty acids, creating fatty acid hydroperoxides (oxylipins). Fungal oxylipins are required for normal fungal development and secondary metabolism, and plant host-derived oxylipins interfere with these processes in fungi, presumably by signal mimicry. The maize lipoxygenase gene ZmLOX3 has been implicated previously in seed-Aspergillus interactions, so we tested the interactions of a mutant maize line (lox3–4, in which ZmLOX3 is disrupted) with the mycotoxigenic seed-infecting fungi Aspergillus flavus and Aspergillus nidulans. The lox3–4 mutant was more susceptible than wild type maize to both Aspergillus species. All strains of A. flavus and A. nidulans produced more conidia and aflatoxin (or the precursor sterigmatocystin) on lox3–4 kernels than on wild type kernels, in vitro and under field conditions. Although oxylipins did not differ detectably between A. flavus-infected kernels of the lox3–4 and WT maize, oxylipin precursors (free fatty acids) and a downstream metabolite (jasmonic acid) accumulated to greater levels in lox3–4 than in WT kernels. The increased resistance of the lox3–4 mutant to other fungal pathogens (Fusarium, Colletotrichum, Cochliobolus, and Exserohilum) is in sharp contrast with results described herein for Aspergillus, suggesting that outcomes of lipoxygenase-governed host-pathogen interactions are pathogen-specific. PMID:19132874

  18. Aspergillus mulundensis sp. nov., a new species for the fungus producing the antifungal echinocandin lipopeptides, mulundocandins.

    PubMed

    Bills, Gerald F; Yue, Qun; Chen, Li; Li, Yan; An, Zhiqiang; Frisvad, Jens C

    2016-03-01

    The invalidly published name Aspergillus sydowii var. mulundensis was proposed for a strain of Aspergillus that produced new echinocandin metabolites designated as the mulundocadins. Reinvestigation of this strain (Y-30462=DSMZ 5745) using phylogenetic, morphological, and metabolic data indicated that it is a distinct and novel species of Aspergillus sect. Nidulantes. The taxonomic novelty, Aspergillus mulundensis, is introduced for this historically important echinocandin-producing strain. The closely related A. nidulans FGSC A4 has one of the most extensively characterized secondary metabolomes of any filamentous fungus. Comparison of the full-genome sequences of DSMZ 5745 and FGSC A4 indicated that the two strains share 33 secondary metabolite biosynthetic gene clusters. These shared gene clusters represent ~45% of the total secondary metabolome of each strain, thus indicating a high level intraspecific divergence in terms of secondary metabolism.

  19. Loss of CclA, required for histone 3 lysine 4 methylation, decreases growth but increases secondary metabolite production in Aspergillus fumigatus

    PubMed Central

    Lee, Seul; Dagenais, Taylor R.T.; Andes, David R.; Kontoyiannis, Dimitrios P.

    2013-01-01

    Secondary metabolite (SM) production in filamentous fungi is mechanistically associated with chromatin remodeling of specific SM clusters. One locus recently shown to be involved in SM suppression in Aspergillus nidulans was CclA, a member of the histone 3 lysine 4 methylating COMPASS complex. Here we examine loss of CclA and a putative H3K4 demethylase, HdmA, in the human pathogen Aspergillus fumigatus. Although deletion of hdmA showed no phenotype under the conditions tested, the cclA deletant was deficient in tri- and di-methylation of H3K4 and yielded a slowly growing strain that was rich in the production of several SMs, including gliotoxin. Similar to deletion of other chromatin modifying enzymes, ΔcclA was sensitive to 6-azauracil indicating a defect in transcriptional elongation. Despite the poor growth, the ΔcclA mutant had wild-type pathogenicity in a murine model and the Toll-deficient Drosophila model of invasive aspergillosis. These data indicate that tri- and di-methylation of H3K4 is involved in the regulation of several secondary metabolites in A. fumigatus, however does not contribute to pathogenicity under the conditions tested. PMID:23638376

  20. The Aspergillus Genome Database (AspGD): recent developments in comprehensive multispecies curation, comparative genomics and community resources.

    PubMed

    Arnaud, Martha B; Cerqueira, Gustavo C; Inglis, Diane O; Skrzypek, Marek S; Binkley, Jonathan; Chibucos, Marcus C; Crabtree, Jonathan; Howarth, Clinton; Orvis, Joshua; Shah, Prachi; Wymore, Farrell; Binkley, Gail; Miyasato, Stuart R; Simison, Matt; Sherlock, Gavin; Wortman, Jennifer R

    2012-01-01

    The Aspergillus Genome Database (AspGD; http://www.aspgd.org) is a freely available, web-based resource for researchers studying fungi of the genus Aspergillus, which includes organisms of clinical, agricultural and industrial importance. AspGD curators have now completed comprehensive review of the entire published literature about Aspergillus nidulans and Aspergillus fumigatus, and this annotation is provided with streamlined, ortholog-based navigation of the multispecies information. AspGD facilitates comparative genomics by providing a full-featured genomics viewer, as well as matched and standardized sets of genomic information for the sequenced aspergilli. AspGD also provides resources to foster interaction and dissemination of community information and resources. We welcome and encourage feedback at aspergillus-curator@lists.stanford.edu.

  1. Conidial Hydrophobins of Aspergillus fumigatus

    PubMed Central

    Paris, Sophie; Debeaupuis, Jean-Paul; Crameri, Reto; Carey, Marilyn; Charlès, Franck; Prévost, Marie Christine; Schmitt, Christine; Philippe, Bruno; Latgé, Jean Paul

    2003-01-01

    The surface of Aspergillus fumigatus conidia, the first structure recognized by the host immune system, is covered by rodlets. We report that this outer cell wall layer contains two hydrophobins, RodAp and RodBp, which are found as highly insoluble complexes. The RODA gene was previously characterized, and ΔrodA conidia do not display a rodlet layer (N. Thau, M. Monod, B. Crestani, C. Rolland, G. Tronchin, J. P. Latgé, and S. Paris, Infect. Immun. 62:4380-4388, 1994). The RODB gene was cloned and disrupted. RodBp was highly homologous to RodAp and different from DewAp of A. nidulans. ΔrodB conidia had a rodlet layer similar to that of the wild-type conidia. Therefore, unlike RodAp, RodBp is not required for rodlet formation. The surface of ΔrodA conidia is granular; in contrast, an amorphous layer is present at the surface of the conidia of the ΔrodA ΔrodB double mutant. These data show that RodBp plays a role in the structure of the conidial cell wall. Moreover, rodletless mutants are more sensitive to killing by alveolar macrophages, suggesting that RodAp or the rodlet structure is involved in the resistance to host cells. PMID:12620846

  2. Conidial hydrophobins of Aspergillus fumigatus.

    PubMed

    Paris, Sophie; Debeaupuis, Jean-Paul; Crameri, Reto; Carey, Marilyn; Charlès, Franck; Prévost, Marie Christine; Schmitt, Christine; Philippe, Bruno; Latgé, Jean Paul

    2003-03-01

    The surface of Aspergillus fumigatus conidia, the first structure recognized by the host immune system, is covered by rodlets. We report that this outer cell wall layer contains two hydrophobins, RodAp and RodBp, which are found as highly insoluble complexes. The RODA gene was previously characterized, and DeltarodA conidia do not display a rodlet layer (N. Thau, M. Monod, B. Crestani, C. Rolland, G. Tronchin, J. P. Latgé, and S. Paris, Infect. Immun. 62:4380-4388, 1994). The RODB gene was cloned and disrupted. RodBp was highly homologous to RodAp and different from DewAp of A. nidulans. DeltarodB conidia had a rodlet layer similar to that of the wild-type conidia. Therefore, unlike RodAp, RodBp is not required for rodlet formation. The surface of DeltarodA conidia is granular; in contrast, an amorphous layer is present at the surface of the conidia of the DeltarodA DeltarodB double mutant. These data show that RodBp plays a role in the structure of the conidial cell wall. Moreover, rodletless mutants are more sensitive to killing by alveolar macrophages, suggesting that RodAp or the rodlet structure is involved in the resistance to host cells.

  3. Cold Shock Syndrome in Anacystis nidulans.

    PubMed

    Rao, V S; Brand, J J; Myers, J

    1977-05-01

    The phenomenon of cold shock in Anacystis nidulans has been explored further in terms of loss of viability and immediate and subsequent metabolic effects. Cold shock was observed also in two closely related strains in which unsaturated fatty acid contents are also known to be low and temperature-dependent. Loss of viability was maximum for cells grown at temperatures above 40 C (<10(-4) survivors after 5 min at 0 C) but became negligibly small for cells grown below 34 C. Development of the cold-sensitive condition after transfer 25 --> 39 C was slow and comparable to rate of growth; development of the insensitive condition after transfer 39 --> 25 C was rapid, implying rapid in situ alteration. An immediate metabolic effect, observed as a decrease in rate of photosynthetic O(2) evolution measured at growth temperature, was less severe than loss of viability. Continued light incubation under growth conditions led to slow decay in rate of O(2) evolution accompanied by loss of membrane chlorophyll. The multiple effects which comprise the cold shock syndrome appear to be membrane-related phenomena and thereby provide an experimental probe of normal membrane function.

  4. The histone acetyltransferase GcnE (GCN5) plays a central role in the regulation of Aspergillus asexual development.

    PubMed

    Cánovas, David; Marcos, Ana T; Gacek, Agnieszka; Ramos, María S; Gutiérrez, Gabriel; Reyes-Domínguez, Yazmid; Strauss, Joseph

    2014-08-01

    Acetylation of histones is a key regulatory mechanism of gene expression in eukaryotes. GcnE is an acetyltransferase of Aspergillus nidulans involved in the acetylation of histone H3 at lysine 9 and lysine 14. Previous works have demonstrated that deletion of gcnE results in defects in primary and secondary metabolism. Here we unveil the role of GcnE in development and show that a ∆gcnE mutant strain has minor growth defects but is impaired in normal conidiophore development. No signs of conidiation were found after 3 days of incubation, and immature and aberrant conidiophores were found after 1 week of incubation. Centroid linkage clustering and principal component (PC) analysis of transcriptomic data suggest that GcnE occupies a central position in Aspergillus developmental regulation and that it is essential for inducing conidiation genes. GcnE function was found to be required for the acetylation of histone H3K9/K14 at the promoter of the master regulator of conidiation, brlA, as well as at the promoters of the upstream developmental regulators of conidiation flbA, flbB, flbC, and flbD (fluffy genes). However, analysis of the gene expression of brlA and the fluffy genes revealed that the lack of conidiation originated in a complete absence of brlA expression in the ∆gcnE strain. Ectopic induction of brlA from a heterologous alcA promoter did not remediate the conidiation defects in the ∆gcnE strain, suggesting that additional GcnE-mediated mechanisms must operate. Therefore, we conclude that GcnE is the only nonessential histone modifier with a strong role in fungal development found so far.

  5. Photoinhibition and reactivation of photosynthesis in the cyanobacterium Anacystis nidulans

    SciTech Connect

    Samuelsson, G.; Loenneborg, A.; Rosenqvist, E.; Gustafsson, P.; Oequist, G.

    1985-12-01

    The susceptibility of photosynthesis to photoinhibition and its recovery were studied on cultures of the cyanobacterium Anacystis nidulans. Oxygen evolution and low temperature fluorescence kinetics were measured. Upon exposure to high light A. nidulans showed a rapid decrease in oxygen evolution followed by a quasi steady state rate of photosynthesis. This quasi steady state rate decreased with increasing photon flux density of the photoinhibitory light. Reactivation of photosynthesis in dim light after the photoinhibitory treatment was rapid: 85 to 95% recovery occurred within 2 hours. In the presence of the translation inhibitor, streptomycin (250 micrograms per milliliter), no reactivation occurred. We also found that the damage increased dramatically if the high light treatment was done with streptomycin added. A transcription inhibitor, rifampicin, did not inhibit the reactivation process. Based on these data we conclude that the photoinhibitory damage observed is the net result of a balance between the photoinhibitory process and the operation of the repairing mechanism(s).

  6. Isolation and identification of Aspergillus spp. from brown kiwi (Apteryx mantelli) nocturnal houses in New Zealand.

    PubMed

    Glare, Travis R; Gartrell, Brett D; Brookes, Jenny J; Perrott, John K

    2014-03-01

    Aspergillosis, a disease caused by infection with Aspergillus spp., is a common cause of death in birds globally and is an irregular cause of mortality of captive kiwi (Apteryx spp.). Aspergillus spp. are often present in rotting plant material, including the litter and nesting material used for kiwi in captivity. The aim of this study was to survey nocturnal kiwi houses in New Zealand to assess the levels of Aspergillus currently present in leaf litter. Samples were received from 11 nocturnal kiwi houses from throughout New Zealand, with one site supplying multiple samples over time. Aspergillus was isolated and quantified by colony counts from litter samples using selective media and incubation temperatures. Isolates were identified to the species level by amplification and sequencing of ITS regions of the ribosomal. Aspergillus spp. were recovered from almost every sample; however, the levels in most kiwi houses were below 1000 colony-forming units (CFU)/g of wet material. The predominant species was Aspergillus fumigatus, with rare occurrences of Aspergillus niger, Aspergillus nidulans, and Aspergillus parasiticus. Only one site had no detectable Aspergillus. The limit of detection was around 50 CFU/g wet material. One site was repeatedly sampled as it had a high loading of A. fumigatus at the start of the survey and had two recent clinical cases of aspergillosis diagnosed in resident kiwi. Environmental loading at this site with Aspergillus spp. reduced but was not eliminated despite changes of the litter. The key finding of our study is that the background levels of Aspergillus spores in kiwi nocturnal houses in New Zealand are low, but occasional exceptions occur and are associated with the onset of aspergillosis in otherwise healthy birds. The predominant Aspergillus species present in the leaf litter was A. fumigatus, but other species were also present. Further research is needed to confirm the optimal management of leaf litter to minimize Aspergillus

  7. Isolation and identification of Aspergillus spp. from brown kiwi (Apteryx mantelli) nocturnal houses in New Zealand.

    PubMed

    Glare, Travis R; Gartrell, Brett D; Brookes, Jenny J; Perrott, John K

    2014-03-01

    Aspergillosis, a disease caused by infection with Aspergillus spp., is a common cause of death in birds globally and is an irregular cause of mortality of captive kiwi (Apteryx spp.). Aspergillus spp. are often present in rotting plant material, including the litter and nesting material used for kiwi in captivity. The aim of this study was to survey nocturnal kiwi houses in New Zealand to assess the levels of Aspergillus currently present in leaf litter. Samples were received from 11 nocturnal kiwi houses from throughout New Zealand, with one site supplying multiple samples over time. Aspergillus was isolated and quantified by colony counts from litter samples using selective media and incubation temperatures. Isolates were identified to the species level by amplification and sequencing of ITS regions of the ribosomal. Aspergillus spp. were recovered from almost every sample; however, the levels in most kiwi houses were below 1000 colony-forming units (CFU)/g of wet material. The predominant species was Aspergillus fumigatus, with rare occurrences of Aspergillus niger, Aspergillus nidulans, and Aspergillus parasiticus. Only one site had no detectable Aspergillus. The limit of detection was around 50 CFU/g wet material. One site was repeatedly sampled as it had a high loading of A. fumigatus at the start of the survey and had two recent clinical cases of aspergillosis diagnosed in resident kiwi. Environmental loading at this site with Aspergillus spp. reduced but was not eliminated despite changes of the litter. The key finding of our study is that the background levels of Aspergillus spores in kiwi nocturnal houses in New Zealand are low, but occasional exceptions occur and are associated with the onset of aspergillosis in otherwise healthy birds. The predominant Aspergillus species present in the leaf litter was A. fumigatus, but other species were also present. Further research is needed to confirm the optimal management of leaf litter to minimize Aspergillus

  8. Spectroscopic Evidence for the Two C-H-Cleaving Intermediates of Aspergillus nidulans Isopenicillin N Synthase.

    PubMed

    Tamanaha, Esta; Zhang, Bo; Guo, Yisong; Chang, Wei-Chen; Barr, Eric W; Xing, Gang; St Clair, Jennifer; Ye, Shengfa; Neese, Frank; Bollinger, J Martin; Krebs, Carsten

    2016-07-20

    The enzyme isopenicillin N synthase (IPNS) installs the β-lactam and thiazolidine rings of the penicillin core into the linear tripeptide l-δ-aminoadipoyl-l-Cys-d-Val (ACV) on the pathways to a number of important antibacterial drugs. A classic set of enzymological and crystallographic studies by Baldwin and co-workers established that this overall four-electron oxidation occurs by a sequence of two oxidative cyclizations, with the β-lactam ring being installed first and the thiazolidine ring second. Each phase requires cleavage of an aliphatic C-H bond of the substrate: the pro-S-CCys,β-H bond for closure of the β-lactam ring, and the CVal,β-H bond for installation of the thiazolidine ring. IPNS uses a mononuclear non-heme-iron(II) cofactor and dioxygen as cosubstrate to cleave these C-H bonds and direct the ring closures. Despite the intense scrutiny to which the enzyme has been subjected, the identities of the oxidized iron intermediates that cleave the C-H bonds have been addressed only computationally; no experimental insight into their geometric or electronic structures has been reported. In this work, we have employed a combination of transient-state-kinetic and spectroscopic methods, together with the specifically deuterium-labeled substrates, A[d2-C]V and AC[d8-V], to identify both C-H-cleaving intermediates. The results show that they are high-spin Fe(III)-superoxo and high-spin Fe(IV)-oxo complexes, respectively, in agreement with published mechanistic proposals derived computationally from Baldwin's founding work.

  9. Spectroscopic Evidence for the Two C-H-Cleaving Intermediates of Aspergillus nidulans Isopenicillin N Synthase.

    PubMed

    Tamanaha, Esta; Zhang, Bo; Guo, Yisong; Chang, Wei-Chen; Barr, Eric W; Xing, Gang; St Clair, Jennifer; Ye, Shengfa; Neese, Frank; Bollinger, J Martin; Krebs, Carsten

    2016-07-20

    The enzyme isopenicillin N synthase (IPNS) installs the β-lactam and thiazolidine rings of the penicillin core into the linear tripeptide l-δ-aminoadipoyl-l-Cys-d-Val (ACV) on the pathways to a number of important antibacterial drugs. A classic set of enzymological and crystallographic studies by Baldwin and co-workers established that this overall four-electron oxidation occurs by a sequence of two oxidative cyclizations, with the β-lactam ring being installed first and the thiazolidine ring second. Each phase requires cleavage of an aliphatic C-H bond of the substrate: the pro-S-CCys,β-H bond for closure of the β-lactam ring, and the CVal,β-H bond for installation of the thiazolidine ring. IPNS uses a mononuclear non-heme-iron(II) cofactor and dioxygen as cosubstrate to cleave these C-H bonds and direct the ring closures. Despite the intense scrutiny to which the enzyme has been subjected, the identities of the oxidized iron intermediates that cleave the C-H bonds have been addressed only computationally; no experimental insight into their geometric or electronic structures has been reported. In this work, we have employed a combination of transient-state-kinetic and spectroscopic methods, together with the specifically deuterium-labeled substrates, A[d2-C]V and AC[d8-V], to identify both C-H-cleaving intermediates. The results show that they are high-spin Fe(III)-superoxo and high-spin Fe(IV)-oxo complexes, respectively, in agreement with published mechanistic proposals derived computationally from Baldwin's founding work. PMID:27193226

  10. Aspergillus spinal epidural abscess

    SciTech Connect

    Byrd, B.F. III; Weiner, M.H.; McGee, Z.A.

    1982-12-17

    A spinal epidural abscess developed in a renal transplant recipient; results of a serum radioimmunoassay for Aspergillus antigen were positive. Laminectomy disclosed an abscess of the L4-5 interspace and L-5 vertebral body that contained hyphal forms and from which Aspergillus species was cultured. Serum Aspergillus antigen radioimmunoassay may be a valuable, specific early diagnostic test when systemic aspergillosis is a consideration in an immunosuppressed host.

  11. Identification of Aspergillus Brla Response Elements (Bres) by Genetic Selection in Yeast

    PubMed Central

    Chang, Y. C.; Timberlake, W. E.

    1993-01-01

    The brlA gene of Aspergillus nidulans plays a central role in controlling conidiophore development. To test the hypothesis that brlA encodes a transcriptional regulator and to identify sites of interaction for the BrlA polypeptide, we expressed brlA in Saccharomyces cerevisiae (yeast) strains containing Aspergillus DNA sequences inserted upstream of a minimal yeast promoter fused to the Escherichia coli lacZ gene. Initially, a DNA fragment from the promoter region of the developmentally regulated rodA gene was tested and shown to mediate brlA-dependent transcriptional activation. Two additional DNA fragments were selected from an Aspergillus genomic library by their ability to respond to brlA in yeast. These fragments contained multiple copies of a sequence motif present in the rodA fragment, which we propose to be sites for BrlA interaction and designate brlA response elements (BREs). DNA fragments containing BREs upstream of a minimal Aspergillus promoter were capable of conferring developmental regulation in Aspergillus. Deletion of BREs from the upstream region of rodA greatly decreased its developmental induction. Multiple copies of a synthetic oligonucleotide with the consensus sequence identified among the BREs mediated brlA-dependent transcriptional activation in yeast. The results show that a primary activity of brlA is transcriptional activation and tentatively identify sites of interaction for the BrlA polypeptide. PMID:8417986

  12. Sexual development and cryptic sexuality in fungi: insights from Aspergillus species.

    PubMed

    Dyer, Paul S; O'Gorman, Céline M

    2012-01-01

    Major insights into sexual development and cryptic sexuality within filamentous fungi have been gained from investigations using Aspergillus species. Here, an overview is first given into sexual morphogenesis in the aspergilli, describing the different types of sexual structures formed and how their production is influenced by a variety of environmental and nutritional factors. It is argued that the formation of cleistothecia and accessory tissues, such as Hülle cells and sclerotia, should be viewed as two independent but co-ordinated developmental pathways. Next, a comprehensive survey of over 75 genes associated with sexual reproduction in the aspergilli is presented, including genes relating to mating and the development of cleistothecia, sclerotia and ascospores. Most of these genes have been identified from studies involving the homothallic Aspergillus nidulans, but an increasing number of studies have now in addition characterized 'sex-related' genes from the heterothallic species Aspergillus fumigatus and Aspergillus flavus. A schematic developmental genetic network is proposed showing the inter-relatedness between these genes. Finally, the discovery of sexual reproduction in certain Aspergillus species that were formerly considered to be strictly asexual is reviewed, and the importance of these findings for cryptic sexuality in the aspergilli as a whole is discussed.

  13. Aspergillus Collagen-Like Genes (acl): Identification, Sequence Polymorphism, and Assessment for PCR-Based Pathogen Detection

    PubMed Central

    Tuntevski, Kiril; Durney, Brandon C.; Snyder, Anna K.; LaSala, P. Rocco; Nayak, Ajay P.; Green, Brett J.; Beezhold, Donald H.; Rio, Rita V. M.; Holland, Lisa A.

    2013-01-01

    The genus Aspergillus is a burden to public health due to its ubiquitous presence in the environment, its production of allergens, and wide demographic susceptibility among cystic fibrosis, asthmatic, and immunosuppressed patients. Current methods of detection of Aspergillus colonization and infection rely on lengthy morphological characterization or nonstandardized serological assays that are restricted to identifying a fungal etiology. Collagen-like genes have been shown to exhibit species-specific conservation across the noncollagenous regions as well as strain-specific polymorphism in the collagen-like regions. Here we assess the conserved region of the Aspergillus collagen-like (acl) genes and explore the application of PCR amplicon size-based discrimination among the five most common etiologic species of the Aspergillus genus, including Aspergillus fumigatus, A. flavus, A. nidulans, A. niger, and A. terreus. Genetic polymorphism and phylogenetic analysis of the aclF1 gene were additionally examined among the available strains. Furthermore, the applicability of the PCR-based assay to identification of these five species in cultures derived from sputum and bronchoalveolar fluid from 19 clinical samples was explored. Application of capillary electrophoresis on nanogels was additionally demonstrated to improve the discrimination between Aspergillus species. Overall, this study demonstrated that Aspergillus acl genes could be used as PCR targets to discriminate between clinically relevant Aspergillus species. Future studies aim to utilize the detection of Aspergillus acl genes in PCR and microfluidic applications to determine the sensitivity and specificity for the identification of Aspergillus colonization and invasive aspergillosis in immunocompromised subjects. PMID:24123732

  14. Effects of conidia of various Aspergillus species on apoptosis of human pneumocytes and bronchial epithelial cells.

    PubMed

    Féménia, F; Huet, D; Lair-Fulleringer, S; Wagner, M C; Sarfati, J; Shingarova, L; Guillot, J; Boireau, P; Chermette, R; Berkova, N

    2009-05-01

    Aspergillus species can cause mycoses in human and animals. Previously, we demonstrated that A. fumigatus conidia from a human isolate inhibited apoptosis in human pneumocytes and bronchial epithelial cells. In the current study, we studied the effects of A. fumigatus conidia non-human origin and A. flavus, A. nidulans, A. niger and A. oryzae conidia on human cells apoptosis. Human pneumocytes or bronchial epithelial cells were simultaneously exposed to apoptotic inductors and aspergilli conidia. The cell cultures were analyzed by flow cytometry, immunoblotting, and examination of nuclear morphology. Similar to A. fumigatus conidia, A. flavus conidia inhibited cellular apoptosis while A. nidulans, A. niger and A. oryzae conidia did not affect apoptosis. We further studied the species specificity of conidia: there were no differences in the inhibition of apoptosis by A. fumigatus conidia from either human or bird isolates. In order to determine whether the inhibition of apoptosis by conidia is limited to certain strains, the effect on human cell apoptosis of different A. fumigatus human clinical isolates and A. fumigatus of environmental origin was evaluated. All A. fumigatus isolates inhibited apoptosis; an anti-apoptotic factor was released by conidia. For TNF-induced apoptosis, the anti-apoptotic effect of conidia of all isolates was found to be associated with a reduction of caspase-3 in human cells. The results suggest that suppression of apoptosis may play a role in reducing the efficacy of host defense mechanisms during infection with Aspergillus species. PMID:19117118

  15. Diversity, molecular phylogeny and fingerprint profiles of airborne Aspergillus species using random amplified polymorphic DNA.

    PubMed

    Kermani, Firoozeh; Shams-Ghahfarokhi, Masoomeh; Gholami-Shabani, Mohammadhassan; Razzaghi-Abyaneh, Mehdi

    2016-06-01

    In the present study, diversity and phylogenetic relationship of Aspergillus species isolated from Tehran air was studied using random amplified polymorphic DNA (RAPD)-polymerase chain reaction (RAPD-PCR). Thirty-eight Aspergillus isolates belonging to 12 species i.e. A. niger (28.94 %, 11 isolates), A. flavus (18.42 %, 7 isolates), A. tubingensis (13.15 %, 5 isolates), A. japonicus (10.52 %, 4 isolates), A. ochraceus (10.52 %, 4 isolates), and 2.63 %, 1 isolate from each A. nidulans, A. amstelodami, A. oryzae, A. terreus, A. versicolor, A. flavipes and A. fumigatus were obtained by settle plate method which they were distributed in 18 out of 22 sampling sites examined. Fungal DNA was extracted from cultured mycelia of all Aspergillus isolates on Sabouraud Dextrose Agar and used for amplification of gene fragments in RAPD-PCR using 11 primers. RAPD-PCR data was analyzed using UPGMA software. Resulting dendrogram of combined selected primers including PM1, OPW-04, OPW-05, P160, P54, P10 and OPA14 indicated the distribution of 12 Aspergillus species in 8 major clusters. The similarity coefficient of all 38 Aspergillus isolates ranged from 0.02 to 0.40 indicating a wide degree of similarities and differences within and between species. Taken together, our results showed that various Aspergillus species including some important human pathogenic ones exist in the outdoor air of Tehran by different extents in distribution and diversity and suggested inter- and intra-species genetic diversity among Aspergillus species by RAPD-PCR as a rapid, sensitive and reproducible method. PMID:27116962

  16. Enhanced cellulase producing mutants developed from heterokaryotic Aspergillus strain.

    PubMed

    Kaur, Baljit; Oberoi, H S; Chadha, B S

    2014-03-01

    A heterokaryon 28, derived through protoplast fusion between Aspergillus nidulans and Aspergillus tubingensis (Dal8), was subjected cyclic mutagenesis followed by selection on increasing levels of 2-deoxy glucose (2-DG) as selection marker. The derived deregulated cellulase hyper producing mutant '64', when compared to fusant 28, produced 9.83, 7.8, 3.2, 4.2 and 19.74 folds higher endoglucanase, β-glucosidase, cellobiohydrolase, FPase and xylanase, respectively, under shake cultures. The sequence analysis of PCR amplified β-glucosidase gene from wild and mutant showed nucleotide deletion/substitution. The mutants showed highly catalytic efficient β-glucosidase as evident from low Km and high Vmax values. The expression profiling through zymogram analysis also indicated towards over-expression of cellulases. The up/down regulated expressed proteins observed through SDS-PAGE were identified by Peptide mass fingerprinting The cellulase produced by mutants in conjunction with cellulase free xylanase derived from Thermomyces lanuginosus was used for efficient utilization of alkali treated rice straw for obtaining xylo-oligosaccharides and ethanol.

  17. The Gβ-like protein CpcB is required for hyphal growth, conidiophore morphology and pathogenicity in Aspergillus fumigatus.

    PubMed

    Cai, Zhen-dong; Chai, Yan-fei; Zhang, Cai-yun; Qiao, Wei-ran; Sang, Hong; Lu, Ling

    2015-08-01

    CpcB (cross pathway control B) encodes a yeast Cpc2 and mammalian RACK1 (receptor for activated protein kinase C) ortholog, which is a WD repeat protein with functional homology to the β subunit of heterotrimeric G proteins in Aspergillus fumigatus. Previous study has reported that CpcB governs growth and development in both A. fumigatus and Aspergillus nidulans. However, little is known about the functional identities of CpcB orthologs and their relationships with G protein complexes. In this study, we verified that cytoplasmic AfCpcB acts as a Gβ-like protein ortholog and plays important roles in hyphal growth, conidiophore morphology, cell wall integrity, and virulence in A. fumigatus. Furthermore, double deletion of AfcpcB and AfgpaB (Gα) causes a similar phenotype to AfgpaB mutant with abnormal multiple septa conidiophores but exhibits sparse conidiation with white and fluffy colonies. Thus, the exacerbated conidiation defect suggests that AfcpcB has its own specific function compared to the Gα subunit of AfgpaB or the G-protein complex. In addition, complementation assays using AfcpcB orthologs of A. nidulans and yeasts (Saccharomyces cerevisiae, Schizosaccharomyces pombe, Candida albicans) suggest that all tested fungal AfcpcB orthologs under the A. fumigatus native promoter can largely restore hyphal growth defects in AfcpcB deletion mutant, but only the A. nidulans cpcB ortholog completely rescues the ΔAfcpcB conidiation defect, suggesting that CpcB acts as a Gβ-like protein ortholog in the Aspergilli, but may have unique and important unexplored functions that required for conidiation, which is absent in yeast. PMID:25892048

  18. The alcohol dehydrogenase gene adh1 is induced in Aspergillus flavus grown on medium conducive to aflatoxin biosynthesis.

    PubMed Central

    Woloshuk, C P; Payne, G A

    1994-01-01

    An Aspergillus flavus cDNA library was screened by differential hybridization to isolate clones corresponding to genes that are actively transcribed under culture conditions conducive to aflatoxin biosynthesis. One clone with a 1.28-kb insert was isolated, and its nucleotide sequence was determined. The nucleotide sequence of this clone had 75% DNA identity to those of the alcohol dehydrogenase genes from Aspergillus nidulans, and the putative polypeptide translated from the cDNA sequence had 82% similarity with the amino acid sequences of the A. nidulans proteins. Thus, this gene has been designated adh1. Southern hybridization analysis of genomic DNA from A. flavus indicated that there was one copy of the adh1 gene. Northern (RNA) hybridization analysis indicated that the adh1 transcript accumulated in culture medium conducive to aflatoxin production and the timing of accumulation of adh1 transcripts was similar to that for aflatoxin. Fusion of the promoter region of adh1 to a beta-glucuronidase reporter gene indicated that accumulation of the adh1 transcript was the result of transcriptional activation. These molecular data support previous physiological evidence that showed the importance of carbohydrate metabolism during aflatoxin biosynthesis. Images PMID:8135521

  19. New species of Aspergillus producing sterigmatocystin.

    PubMed Central

    Rabie, C J; Steyn, M; van Schalkwyk, G C

    1977-01-01

    A number of species belonging to the genus Aspergillus were evaluated for their toxicity to ducklings and the ability to produce sterigmatocystin. Three new species capable of producing sterigmatocystin were found, namely, Aspergillus aurantio-brunneus, Aspergillus quadrilineatus, and Aspergillus ustus. All three were toxic to ducklings. The production of sterigmatocystin by Aspergillus rugulosus was confirmed, and the toxicity of Aspergillus stellatus and Aspergillus multicolor is described. PMID:406838

  20. Development in Aspergillus

    PubMed Central

    Krijgsheld, P.; Bleichrodt, R.; van Veluw, G.J.; Wang, F.; Müller, W.H.; Dijksterhuis, J.; Wösten, H.A.B.

    2013-01-01

    The genus Aspergillus represents a diverse group of fungi that are among the most abundant fungi in the world. Germination of a spore can lead to a vegetative mycelium that colonizes a substrate. The hyphae within the mycelium are highly heterogeneous with respect to gene expression, growth, and secretion. Aspergilli can reproduce both asexually and sexually. To this end, conidiophores and ascocarps are produced that form conidia and ascospores, respectively. This review describes the molecular mechanisms underlying growth and development of Aspergillus. PMID:23450714

  1. Eisosome Organization in the Filamentous AscomyceteAspergillus nidulans▿†

    PubMed Central

    Vangelatos, Ioannis; Roumelioti, Katerina; Gournas, Christos; Suarez, Teresa; Scazzocchio, Claudio; Sophianopoulou, Vicky

    2010-01-01

    Eisosomes are subcortical organelles implicated in endocytosis and have hitherto been described only in Saccharomyces cerevisiae. They comprise two homologue proteins, Pil1 and Lsp1, which colocalize with the transmembrane protein Sur7. These proteins are universally conserved in the ascomycetes. We identify in Aspergillus nidulans (and in all members of the subphylum Pezizomycotina) two homologues of Pil1/Lsp1, PilA and PilB, originating from a duplication independent from that extant in the subphylum Saccharomycotina. In the aspergilli there are several Sur7-like proteins in each species, including one strict Sur7 orthologue (SurG in A. nidulans). In A. nidulans conidiospores, but not in hyphae, the three proteins colocalize at the cell cortex and form tightly packed punctate structures that appear different from the clearly distinct eisosome patches observed in S. cerevisiae. These structures are assembled late during the maturation of conidia. In mycelia, punctate structures are present, but they are composed only of PilA, while PilB is diffused in the cytoplasm and SurG is located in vacuoles and endosomes. Deletion of each of the genes does not lead to any obvious growth phenotype, except for moderate resistance to itraconazole. We could not find any obvious association between mycelial (PilA) eisosome-like structures and endocytosis. PilA and SurG are necessary for conidial eisosome organization in ways that differ from those for their S. cerevisiae homologues. These data illustrate that conservation of eisosomal proteins within the ascomycetes is accompanied by a striking functional divergence. PMID:20693301

  2. [Aspergillus insulicola Sp. Nov].

    PubMed

    de Montemayor, L; Santiago, A R

    1975-04-30

    A strain of Aspergillus sp. is described and proposed as a new species under the name "Aspergillus insulicola sp. nov." Montemayor & Santiago, 1973. This strain was isolated from soil samples taken in "Aves Island" during a scientific expedition.--Aves Island, situated at 15 degrees, 40 feet, 42 inches N and 63 degrees, 36 feet, 47 inches W, about 665 Km of the coast of Venezuela, has very special ecological conditions. Due to its smallness: 550 m long and 40 to 120 m across and to its low profile only 3 m over sea level, it is swept by the sea during the periodical storms and hurricanes in the area. It has thus a very interesting fauna and flora. We took a series of soil samples to study its mycological flora. Forty samples were inoculated by dilution method. In this first paper a species is described and proposed as a new species because of its macroscopic and microscopic characteristics, as well as by its biological properties, under the name "Aspergillus insulicola sp. nov.". In its study we have tried to follow as closely as possible the methods recommended by Kennet B. Raper & Dorothy Fenell, world authorities on the genera Aspergillus and Penicillium. The strain is being kept in USB under the number T1, and has been sent to ATCC & CBSC to be incorporated in their collections.

  3. Redox metabolites signal polymicrobial biofilm development via the NapA oxidative stress cascade in Aspergillus

    PubMed Central

    Zheng, He; Kim, Jaekuk; Liew, Mathew; Yan, John K.; Herrera, Oscar; Bok, JinWoo; Kelleher, Neil L.; Keller, Nancy P.; Wang, Yun

    2014-01-01

    Summary Background Filamentous fungi and bacteria form mixed-species biofilms in nature and diverse clinical contexts. They secrete a wealth of redox-active small molecule secondary metabolites, which are traditionally viewed as toxins that inhibit growth of competing microbes. Results Here we report that these “toxins” can act as interspecies signals, affecting filamentous fungal development via oxidative stress regulation. Specifically, in co-culture biofilms, Pseudomonas aeruginosa phenazine-derived metabolites differentially modulated Aspergillus fumigatus development, shifting from weak vegetative growth to induced asexual sporulation (conidiation) along a decreasing phenazine gradient. The A. fumigatus morphological shift correlated with the production of phenazine radicals and concomitant reactive oxygen species (ROS) production generated by phenazine redox cycling. Phenazine conidiation signaling was conserved in the genetic model A. nidulans, and mediated by NapA, a homolog of AP-1-like bZIP transcription factor, which is essential for the response to oxidative stress in humans, yeast, and filamentous fungi. Expression profiling showed phenazine treatment induced a NapA-dependent response of the global oxidative stress metabolome including the thioredoxin, glutathione and NADPH-oxidase systems. Conidiation induction in A. nidulans by another microbial redox-active secondary metabolite, gliotoxin, also required NapA. Conclusions This work highlights that microbial redox metabolites are key signals for sporulation in filamentous fungi, which are communicated through an evolutionarily conserved eukaryotic stress response pathway. It provides a foundation for interspecies signaling in environmental and clinical biofilms involving bacteria and filamentous fungi. PMID:25532893

  4. Translational repression of brlA expression prevents premature development in Aspergillus.

    PubMed Central

    Han, S; Navarro, J; Greve, R A; Adams, T H

    1993-01-01

    The Aspergillus nidulans brlA developmental regulatory locus consists of two overlapping transcription units, brlA alpha and brlA beta, which encode functionally related polypeptides. We used translational fusions between each of the predicted brlA reading frames and the Escherichia coli lacZ gene to test the hypothesis that developmental regulation of brlA alpha and brlA beta expression occurs through different mechanisms. brlA alpha is transcriptionally controlled and a large portion of brlA alpha-directed beta-galactosidase activity is regulated in a brlA-dependent manner. In contrast, brlA beta mRNA is constitutively transcribed but translation of the brlA polypeptide is prevented by the presence of a short open reading frame (microORF) present in the 5' end of brlA beta mRNA. Removing the microORF initiation codon leads to deregulated brlA expression, resulting in an inappropriate activation of development. We propose that one mechanism for developmental induction in A.nidulans involves translational control. Images PMID:8508770

  5. Abundant Respirable Ergot Alkaloids from the Common Airborne Fungus Aspergillus fumigatus†

    PubMed Central

    Panaccione, Daniel G.; Coyle, Christine M.

    2005-01-01

    Ergot alkaloids are mycotoxins that interact with several monoamine receptors, negatively affecting cardiovascular, nervous, reproductive, and immune systems of exposed humans and animals. Aspergillus fumigatus, a common airborne fungus and opportunistic human pathogen, can produce ergot alkaloids in broth culture. The objectives of this study were to determine if A. fumigatus accumulates ergot alkaloids in a respirable form in or on its conidia, to quantify ergot alkaloids associated with conidia produced on several different substrates, and to measure relevant physical properties of the conidia. We found at least four ergot alkaloids, fumigaclavine C, festuclavine, fumigaclavine A, and fumigaclavine B (in order of abundance), associated with conidia of A. fumigatus. Under environmentally relevant conditions, the total mass of ergot alkaloids often constituted >1% of the mass of the conidium. Ergot alkaloids were extracted from conidia produced on all media tested, and the greatest quantities were observed when the fungus was cultured on latex paint or cultured maize seedlings. The values for physical properties of conidia likely to affect their respirability (i.e., diameter, mass, and specific gravity) were significantly lower for A. fumigatus than for Aspergillus nidulans, Aspergillus niger, and Stachybotrys chartarum. The demonstration of relatively high concentrations of ergot alkaloids associated with conidia of A. fumigatus presents opportunities for investigations of potential contributions of the toxins to adverse health effects associated with the fungus and to aspects of the biology of the fungus that contribute to its success. PMID:15933008

  6. Abundant respirable ergot alkaloids from the common airborne fungus Aspergillus fumigatus.

    PubMed

    Panaccione, Daniel G; Coyle, Christine M

    2005-06-01

    Ergot alkaloids are mycotoxins that interact with several monoamine receptors, negatively affecting cardiovascular, nervous, reproductive, and immune systems of exposed humans and animals. Aspergillus fumigatus, a common airborne fungus and opportunistic human pathogen, can produce ergot alkaloids in broth culture. The objectives of this study were to determine if A. fumigatus accumulates ergot alkaloids in a respirable form in or on its conidia, to quantify ergot alkaloids associated with conidia produced on several different substrates, and to measure relevant physical properties of the conidia. We found at least four ergot alkaloids, fumigaclavine C, festuclavine, fumigaclavine A, and fumigaclavine B (in order of abundance), associated with conidia of A. fumigatus. Under environmentally relevant conditions, the total mass of ergot alkaloids often constituted >1% of the mass of the conidium. Ergot alkaloids were extracted from conidia produced on all media tested, and the greatest quantities were observed when the fungus was cultured on latex paint or cultured maize seedlings. The values for physical properties of conidia likely to affect their respirability (i.e., diameter, mass, and specific gravity) were significantly lower for A. fumigatus than for Aspergillus nidulans, Aspergillus niger, and Stachybotrys chartarum. The demonstration of relatively high concentrations of ergot alkaloids associated with conidia of A. fumigatus presents opportunities for investigations of potential contributions of the toxins to adverse health effects associated with the fungus and to aspects of the biology of the fungus that contribute to its success.

  7. Abundant respirable ergot alkaloids from the common airborne fungus Aspergillus fumigatus.

    PubMed

    Panaccione, Daniel G; Coyle, Christine M

    2005-06-01

    Ergot alkaloids are mycotoxins that interact with several monoamine receptors, negatively affecting cardiovascular, nervous, reproductive, and immune systems of exposed humans and animals. Aspergillus fumigatus, a common airborne fungus and opportunistic human pathogen, can produce ergot alkaloids in broth culture. The objectives of this study were to determine if A. fumigatus accumulates ergot alkaloids in a respirable form in or on its conidia, to quantify ergot alkaloids associated with conidia produced on several different substrates, and to measure relevant physical properties of the conidia. We found at least four ergot alkaloids, fumigaclavine C, festuclavine, fumigaclavine A, and fumigaclavine B (in order of abundance), associated with conidia of A. fumigatus. Under environmentally relevant conditions, the total mass of ergot alkaloids often constituted >1% of the mass of the conidium. Ergot alkaloids were extracted from conidia produced on all media tested, and the greatest quantities were observed when the fungus was cultured on latex paint or cultured maize seedlings. The values for physical properties of conidia likely to affect their respirability (i.e., diameter, mass, and specific gravity) were significantly lower for A. fumigatus than for Aspergillus nidulans, Aspergillus niger, and Stachybotrys chartarum. The demonstration of relatively high concentrations of ergot alkaloids associated with conidia of A. fumigatus presents opportunities for investigations of potential contributions of the toxins to adverse health effects associated with the fungus and to aspects of the biology of the fungus that contribute to its success. PMID:15933008

  8. GDP-mannose pyrophosphorylase is essential for cell wall integrity, morphogenesis and viability of Aspergillus fumigatus.

    PubMed

    Jiang, Hechun; Ouyang, Haomiao; Zhou, Hui; Jin, Cheng

    2008-09-01

    GDP-mannose pyrophosphorylase (GMPP) catalyses the synthesis of GDP-mannose, which is the precursor for the mannose residues in glycoconjugates, using mannose 1-phosphate and GTP as substrates. Repression of GMPP in yeast leads to phenotypes including cell lysis, defective cell wall, and failure of polarized growth and cell separation. Although several GMPPs have been isolated and characterized in filamentous fungi, the physiological consequences of their actions are not clear. In this study, Afsrb1, which is a homologue of yeast SRB1/PSA1/VIG9, was identified in the Aspergillus fumigatus genome. The Afsrb1 gene was expressed in Escherichia coli, and recombinant AfSrb1 was functionally confirmed as a GMPP. By the replacement of the native Afsrb1 promoter with an inducible Aspergillus nidulans alcA promoter, the conditional inactivation mutant strain YJ-gmpp was constructed. The presence of 3 % glucose completely blocked transcription of P(alcA)-Afsrb1, and was lethal to strain YJ-gmpp. Repression of Afsrb1 expression in strain YJ-gmpp led to phenotypes including hyphal lysis, defective cell wall, impaired polarity maintenance, and branching site selection. Also, rapid germination and reduced conidiation were documented. However, in contrast to yeast, strain YJ-gmpp retained the ability to direct polarity establishment and septation. Our results showed that the Afsrb1 gene is essential for cell wall integrity, morphogenesis and viability of Aspergillus fumigatus.

  9. Disinfection efficacy of chlorine and peracetic acid alone or in combination against Aspergillus spp. and Candida albicans in drinking water.

    PubMed

    Sisti, Maurizio; Brandi, Giorgio; De Santi, Mauro; Rinaldi, Laura; Schiavano, Giuditta F

    2012-03-01

    The aim of the present study was to evaluate the fungicidal activity of chlorine and peracetic acid in drinking water against various pathogenic Aspergillus spp. and Candida albicans strains. A. nidulans exhibited the greatest resistance, requiring 10 ppm of chlorine for 30 min contact time for a complete inactivation. Under the same experimental conditions, peracetic acid was even less fungicidal. In this case, A. niger proved to be the most resistant species (50 ppm for 60 min for complete inactivation). All Aspergillus spp. were insensitive to 10 ppm even with extended exposure (>5 h). The combination of chlorine and peracetic acid against Aspergillus spp. did not show synergistic effects except in the case of A. flavus. Complete growth inhibition of C. albicans was observed after about 3 h contact time with 0.2 ppm. C. albicans was less sensitive to peracetic acid. Hence the concentrations of chlorine that are usually present in drinking water distribution systems are ineffective against several Aspergillus spp. and peracetic acid cannot be considered an alternative to chlorine for disinfecting drinking water. The combination of the two biocides is not very effective in eliminating filamentous fungi at the concentrations permitted for drinking water disinfection.

  10. Chemodiversity in the genus Aspergillus.

    PubMed

    Frisvad, Jens C; Larsen, Thomas O

    2015-10-01

    Isolates of Aspergillus species are able to produce a large number of secondary metabolites. The profiles of biosynthetic families of secondary metabolites are species specific, whereas individual secondary metabolite families can occur in other species, even those phylogenetically and ecologically unrelated to Aspergillus. Furthermore, there is a high degree of chemo-consistency from isolate to isolate in a species even though certain metabolite gene clusters are silenced in some isolates. Genome sequencing projects have shown that the diversity of secondary metabolites is much larger in each species than previously thought. The potential of finding even further new bioactive drug candidates in Aspergillus is evident, despite the fact that many secondary metabolites have already been structure elucidated and chemotaxonomic studies have shown that many new secondary metabolites have yet to be characterized. The genus Aspergillus is cladistically holophyletic but phenotypically polythetic and very diverse and is associated to quite different sexual states. Following the one fungus one name system, the genus Aspergillus is restricted to a holophyletic clade that include the morphologically different genera Aspergillus, Dichotomomyces, Phialosimplex, Polypaecilum and Cristaspora. Secondary metabolites common between the subgenera and sections of Aspergillus are surprisingly few, but many metabolites are common to a majority of species within the sections. We call small molecule extrolites in the same biosynthetic family isoextrolites. However, it appears that secondary metabolites from one Aspergillus section have analogous metabolites in other sections (here also called heteroisoextrolites). In this review, we give a genus-wide overview of secondary metabolite production in Aspergillus species. Extrolites appear to have evolved because of ecological challenges rather than being inherited from ancestral species, at least when comparing the species in the different

  11. Examining the Evolution of the Regulatory Circuit Controlling Secondary Metabolism and Development in the Fungal Genus Aspergillus

    PubMed Central

    Lind, Abigail L.; Wisecaver, Jennifer H.; Smith, Timothy D.; Feng, Xuehuan; Calvo, Ana M.; Rokas, Antonis

    2015-01-01

    Filamentous fungi produce diverse secondary metabolites (SMs) essential to their ecology and adaptation. Although each SM is typically produced by only a handful of species, global SM production is governed by widely conserved transcriptional regulators in conjunction with other cellular processes, such as development. We examined the interplay between the taxonomic narrowness of SM distribution and the broad conservation of global regulation of SM and development in Aspergillus, a diverse fungal genus whose members produce well-known SMs such as penicillin and gliotoxin. Evolutionary analysis of the 2,124 genes comprising the 262 SM pathways in four Aspergillus species showed that most SM pathways were species-specific, that the number of SM gene orthologs was significantly lower than that of orthologs in primary metabolism, and that the few conserved SM orthologs typically belonged to non-homologous SM pathways. RNA sequencing of two master transcriptional regulators of SM and development, veA and mtfA, showed that the effects of deletion of each gene, especially veA, on SM pathway regulation were similar in A. fumigatus and A. nidulans, even though the underlying genes and pathways regulated in each species differed. In contrast, examination of the role of these two regulators in development, where 94% of the underlying genes are conserved in both species showed that whereas the role of veA is conserved, mtfA regulates development in the homothallic A. nidulans but not in the heterothallic A. fumigatus. Thus, the regulation of these highly conserved developmental genes is divergent, whereas–despite minimal conservation of target genes and pathways–the global regulation of SM production is largely conserved. We suggest that the evolution of the transcriptional regulation of secondary metabolism in Aspergillus represents a novel type of regulatory circuit rewiring and hypothesize that it has been largely driven by the dramatic turnover of the target genes

  12. Aspergillus: sex and recombination.

    PubMed

    Varga, János; Szigeti, Gyöngyi; Baranyi, Nikolett; Kocsubé, Sándor; O'Gorman, Céline M; Dyer, Paul S

    2014-12-01

    The genus Aspergillus is one of the most widespread groups of fungi on Earth, comprised of about 300-350 species with very diverse lifestyles. Most species produce asexual propagula (conidia) on conidial heads. Despite their ubiquity, a sexual cycle has not yet been identified for most of the aspergilli. Where sexual reproduction is present, species exhibit either homothallic (self fertile) or heterothallic (obligate outcrossing) breeding systems. A parasexual cycle has also been described in some Aspergillus species. As in other fungi, sexual reproduction is governed by mating-type (MAT) genes, which determine sexual identity and are involved in regulating later stages of sexual development. Previous population genetic studies have indicated that some supposedly asexual aspergilli exhibit evidence of a recombining population structure, suggesting the presence of a cryptic sexual cycle. In addition, genome analyses have revealed networks of genes necessary for sexual reproduction in several Aspergillus species, again consistent with latent sexuality in these fungi. Knowledge of MAT gene presence has then successfully been applied to induce sexual reproduction between MAT1-1 and MAT1-2 isolates of certain supposedly asexual aspergilli. Recent progress in understanding the extent and significance of sexual reproduction is described here, with special emphasis on findings that are relevant to clinically important aspergilli.

  13. Aspergillus: sex and recombination.

    PubMed

    Varga, János; Szigeti, Gyöngyi; Baranyi, Nikolett; Kocsubé, Sándor; O'Gorman, Céline M; Dyer, Paul S

    2014-12-01

    The genus Aspergillus is one of the most widespread groups of fungi on Earth, comprised of about 300-350 species with very diverse lifestyles. Most species produce asexual propagula (conidia) on conidial heads. Despite their ubiquity, a sexual cycle has not yet been identified for most of the aspergilli. Where sexual reproduction is present, species exhibit either homothallic (self fertile) or heterothallic (obligate outcrossing) breeding systems. A parasexual cycle has also been described in some Aspergillus species. As in other fungi, sexual reproduction is governed by mating-type (MAT) genes, which determine sexual identity and are involved in regulating later stages of sexual development. Previous population genetic studies have indicated that some supposedly asexual aspergilli exhibit evidence of a recombining population structure, suggesting the presence of a cryptic sexual cycle. In addition, genome analyses have revealed networks of genes necessary for sexual reproduction in several Aspergillus species, again consistent with latent sexuality in these fungi. Knowledge of MAT gene presence has then successfully been applied to induce sexual reproduction between MAT1-1 and MAT1-2 isolates of certain supposedly asexual aspergilli. Recent progress in understanding the extent and significance of sexual reproduction is described here, with special emphasis on findings that are relevant to clinically important aspergilli. PMID:25118872

  14. Isolation of Aneuploid-Generating Mutants of ASPERGILLUS NIDULANS, One of Which Is Defective in Interphase of the Cell Cycle

    PubMed Central

    Upshall, A.; Mortimore, I. D.

    1984-01-01

    A method is described for isolating mutants potentially defective in loci involved in mitotic chromosome segregation. Conditional lethal, heat-sensitive (42°) mutants were assayed at a subrestrictive temperature of 37° for an inflated production of colonies displaying phenotypes and behavior patterns of whole chromosome aneuploids. Of 14 mutants, three showed specificity for one disomic phenotype, whereas 11 generated colonies mosaic for different aneuploid phenotypes. This latter group is designated hfa ( high frequency of aneuploid). For ten of the 11 mutants temperature sensitivity and aneuploid production cosegregated, indicating a single mutation in each. These mutations were recessive and nonallelic. Analysis was concentrated on the hfaB3 mutation which is mapped to chromosome VI tightly linked to the methB and tsB loci. The disruptive influence of hfaB3 on mitosis at 37° was shown by (1) ploidy and whole chromosome-type segregation of markers in the breakdown sectors of phenotypically aneuploid colonies obtained from multiply marked homozygous hfaB3 disploids; (2) a high frequency of haploid and nondisjunctional diploid segregants among spontaneous yellow-spored parasexual recombinants taken from green-spored homozygous hfaB3 diploids. The mutation had no effect on meiotic chromosome segregation at 37°. The single interphase nucleus in germlings at 42°, coupled with changes in the mitotic index in temperature exchange experiments, showed hfaB3 to arrest the cell cycle in interphase at restrictive temperature. A conclusion drawn is that the hfaB gene product is required both for entry into mitosis and for normal chromosome segregation in dividing nuclei. PMID:6479583

  15. Metabolomics of Aspergillus fumigatus.

    PubMed

    Frisvad, Jens C; Rank, Christian; Nielsen, Kristian F; Larsen, Thomas O

    2009-01-01

    Aspergillus fumigatus is the most important species in Aspergillus causing infective lung diseases. This species has been reported to produce a large number of extrolites, including secondary metabolites, acids, and proteins such as hydrophobins and extracellular enzymes. At least 226 potentially bioactive secondary metabolites have been reported from A. fumigatus that can be ordered into 24 biosynthetic families. Of these families we have detected representatives from the following families of secondary metabolites: fumigatins, fumigaclavines, fumiquinazolines, trypacidin and monomethylsulochrin, fumagillins, gliotoxins, pseurotins, chloroanthraquinones, fumitremorgins, verruculogen, helvolic acids, and pyripyropenes by HPLC with diode array detection and mass spectrometric detection. There is still doubt whether A. fumigatus can produce tryptoquivalins, but all isolates produce the related fumiquinazolines. We also tentatively detected sphingofungins in A. fumigatus Af293 and in an isolate of A. lentulus. The sphingofungins may have a similar role as the toxic fumonisins, found in A. niger. A further number of mycotoxins, including ochratoxin A, and other secondary metabolites have been reported from A. fumigatus, but in those cases either the fungus or its metabolite appear to be misidentified. PMID:18763205

  16. Previously unknown species of Aspergillus.

    PubMed

    Gautier, M; Normand, A-C; Ranque, S

    2016-08-01

    The use of multi-locus DNA sequence analysis has led to the description of previously unknown 'cryptic' Aspergillus species, whereas classical morphology-based identification of Aspergillus remains limited to the section or species-complex level. The current literature highlights two main features concerning these 'cryptic' Aspergillus species. First, the prevalence of such species in clinical samples is relatively high compared with emergent filamentous fungal taxa such as Mucorales, Scedosporium or Fusarium. Second, it is clearly important to identify these species in the clinical laboratory because of the high frequency of antifungal drug-resistant isolates of such Aspergillus species. Matrix-assisted laser desorption/ionization-time of flight mass spectrometry (MALDI-TOF MS) has recently been shown to enable the identification of filamentous fungi with an accuracy similar to that of DNA sequence-based methods. As MALDI-TOF MS is well suited to the routine clinical laboratory workflow, it facilitates the identification of these 'cryptic' Aspergillus species at the routine mycology bench. The rapid establishment of enhanced filamentous fungi identification facilities will lead to a better understanding of the epidemiology and clinical importance of these emerging Aspergillus species. Based on routine MALDI-TOF MS-based identification results, we provide original insights into the key interpretation issues of a positive Aspergillus culture from a clinical sample. Which ubiquitous species that are frequently isolated from air samples are rarely involved in human invasive disease? Can both the species and the type of biological sample indicate Aspergillus carriage, colonization or infection in a patient? Highly accurate routine filamentous fungi identification is central to enhance the understanding of these previously unknown Aspergillus species, with a vital impact on further improved patient care. PMID:27263029

  17. Aspergillus prosthetic valve endocarditis.

    PubMed Central

    Petheram, I S; Seal, R M

    1976-01-01

    The clinical, laboratory, and histopathological features of seven cases of Aspergillus fumigatus prosthetic valve endocarditis are presented. The exact nature of the lesion, a combination of infective fungal endocarditis and thrombosis on the prosthetic valve, is discussed and the difficulties in clinical diagnosis are emphasized. Helpful indications were sudden unexplained heart failure with the appearance of new murmurs, and emboli to large or medium-sized systemic arteries. Fever and anaemia were inconstant, and in no case was blood culture or precipitin investigation helpful. Spore contamination of operating theatre air was the likely source of infection, and measures taken to overcome this and other predisposing factors are discussed. Since medical diagnosis is usually late and the few reported cures in this condition have included replacement of the prosthesis, early surgical intervention combined with antifungal chemotherapy is advised. Images PMID:788218

  18. Tremorgenic Mycotoxins from Aspergillus Caespitosus

    PubMed Central

    Schroeder, H. W.; Cole, R. J.; Hein, H.; Kirksey, J. W.

    1975-01-01

    Two tremorgenic mycotoxins were isolated from Aspergillus caespitosus, and identified as verruculogen and fumitremorgin B. They were produced at the rate of 172 and 325 mg per kg, respectively, on autoclaved cracked field corn. PMID:1155935

  19. Tremorgenic mycotoxins from Aspergillus caespitosus.

    PubMed

    Schroeder, H W; Cole, R J; Hein, H; Kirksey, J W

    1975-06-01

    Two tremorgenic mycotoxins were isolated from Aspergillus caespitosus, and identified as verruculogen and fumitremorgin B. They were produced at the rate of 172 and 325 mg per kg, respectively, on autoclaved cracked field corn. PMID:1155935

  20. Photorecovery of gamma irradiated cultures of blue-green alga, Anacystis nidulans.

    NASA Technical Reports Server (NTRS)

    Asato, Y.

    1971-01-01

    Evidence is given for photorecovery of Anacystis nidulans after exposures to Co 60 gamma radiation. After irradiation the levels of viable cells were higher in cultures kept in white light than in cultures kept dark for 24 hr. The post-irradiation survival rate increase after 30-min exposures to visible light is demonstrated in cultures irradiated with 35 krad. An increase in survival rates was not observed after exposures to ?red' light.

  1. Effect of barium and nickel on the growth of anacystis nidulans

    SciTech Connect

    Lustigman, L.H.L.B.

    1996-12-31

    Anacystis nidulans is a simple, unicellular, prokaryotic microorganism. Like other cyanobacteria it is an obligate photoautotroph that is similar to gram-negative bacteria in cell wall structure, replication, and ability to harbor plasmids. Cyanobacteria are excellent organisms to serve as models for the investigation of a wide variety of biological problems, including indicators of environmental pollution. There have been several studies on the effects of heavy metals on A. nidulans. Toxic metals are a major water pollution problem. Metals come from natural weathering processes of the earth`s crust, but industrialization and urbanization have led to an increase in contamination of aquatic environments, mainly from industrial discharge, pest or disease control agents applied to plants, urban run-off, mining, soil erosion, sewage effluents, air pollution fallout, and other sources. Among these contaminants are nickel, barium, and their derivatives. This study examined the effects of selected concentrations of nickel chloride and barium chloride on the growth of A. nidulans, with and without the addition of EDTA. 19 refs., 3 figs.

  2. Aspergillus fumigatus and Aspergillosis

    PubMed Central

    Latgé, Jean-Paul

    1999-01-01

    Aspergillus fumigatus is one of the most ubiquitous of the airborne saprophytic fungi. Humans and animals constantly inhale numerous conidia of this fungus. The conidia are normally eliminated in the immunocompetent host by innate immune mechanisms, and aspergilloma and allergic bronchopulmonary aspergillosis, uncommon clinical syndromes, are the only infections observed in such hosts. Thus, A. fumigatus was considered for years to be a weak pathogen. With increases in the number of immunosuppressed patients, however, there has been a dramatic increase in severe and usually fatal invasive aspergillosis, now the most common mold infection worldwide. In this review, the focus is on the biology of A. fumigatus and the diseases it causes. Included are discussions of (i) genomic and molecular characterization of the organism, (ii) clinical and laboratory methods available for the diagnosis of aspergillosis in immunocompetent and immunocompromised hosts, (iii) identification of host and fungal factors that play a role in the establishment of the fungus in vivo, and (iv) problems associated with antifungal therapy. PMID:10194462

  3. Keratitis caused by Aspergillus pseudotamarii.

    PubMed

    Baranyi, Nikolett; Kocsubé, Sándor; Szekeres, András; Raghavan, Anita; Narendran, Venkatapathy; Vágvölgyi, Csaba; Panneer Selvam, Kanesan; Babu Singh, Yendremban Randhir; Kredics, László; Varga, János; Manikandan, Palanisamy

    2013-04-12

    A male patient presented with complaints of redness, pain and defective vision in the left eye. The infiltrate healed completely after two weeks of topical natamycin administration. A polyphasic approach was used to identify the isolate as Aspergillus pseudotamarii, which produced aflatoxins in inducing medium.

  4. Aspergillus infections in cystic fibrosis.

    PubMed

    King, Jill; Brunel, Shan F; Warris, Adilia

    2016-07-01

    Patients with cystic fibrosis (CF) suffer from chronic lung infection and airway inflammation. Respiratory failure secondary to chronic or recurrent infection remains the commonest cause of death and accounts for over 90% of mortality. Bacteria as Staphylococcus aureus, Pseudomonas aeruginosa and Burkholderia cepacia complex have been regarded the main CF pathogens and their role in progressive lung decline has been studied extensively. Little attention has been paid to the role of Aspergillus spp. and other filamentous fungi in the pathogenesis of non-ABPA (allergic bronchopulmonary aspergillosis) respiratory disease in CF, despite their frequent recovery in respiratory samples. It has become more apparent however, that Aspergillus spp. may play an important role in chronic lung disease in CF. Research delineating the underlying mechanisms of Aspergillus persistence and infection in the CF lung and its link to lung deterioration is lacking. This review summarizes the Aspergillus disease phenotypes observed in CF, discusses the role of CFTR (cystic fibrosis transmembrane conductance regulator)-protein in innate immune responses and new treatment modalities. PMID:27177733

  5. Sexual recombination in Aspergillus tubingensis

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Aspergillus tubingensis from section Nigri (Black Aspergilli) is closely related to A. niger and is used extensively in the industrial production of enzymes and organic acids. We recently discovered sexual reproduction in A. tubingensis and in this study, demonstrate that the progeny are products o...

  6. 76 FR 16297 - Aspergillus flavus

    Federal Register 2010, 2011, 2012, 2013, 2014

    2011-03-23

    ... Findings In the Federal Register of March 3, 2010 (75 FR 9596) (FRL-8811-2), EPA issued a notice pursuant..., 2003 (68 FR 41541) (FRL-7311-6). Those health effects data were the basis for establishing the... exemptions for experimental use of Aspergillus flavus AF36 on pistachio (72 FR 28871, May 23, 2007)...

  7. Keratitis caused by Aspergillus pseudotamarii

    PubMed Central

    Baranyi, Nikolett; Kocsubé, Sándor; Szekeres, András; Raghavan, Anita; Narendran, Venkatapathy; Vágvölgyi, Csaba; Panneer Selvam, Kanesan; Babu Singh, Yendremban Randhir; Kredics, László; Varga, János; Manikandan, Palanisamy

    2013-01-01

    A male patient presented with complaints of redness, pain and defective vision in the left eye. The infiltrate healed completely after two weeks of topical natamycin administration. A polyphasic approach was used to identify the isolate as Aspergillus pseudotamarii, which produced aflatoxins in inducing medium. PMID:24432226

  8. EUCAST testing of Isavuconazole susceptibility in Aspergillus: comparison of results for Inoculum standardization using Conidium counting versus optical density.

    PubMed

    Arendrup, Maiken Cavling; Howard, Susan; Lass-Flörl, Cornelia; Mouton, Johan W; Meletiadis, Joseph; Cuenca-Estrella, Manuel

    2014-11-01

    The EUCAST E.DEF9.1 standard recommends standardization of the inoculum concentration by conidium counting using a hemocytometer rather than a spectrophotometer. In this study, we investigated whether the choice of these methods influenced isavuconazole MICs. A blinded collection of 30 molecularly characterized azole-resistant isolates and 10 wild-type Aspergillus fumigatus isolates was shared with four different laboratories. Additionally, each laboratory selected approximately 100 A. fumigatus isolates and 50 isolates each of A. flavus, A. nidulans, A. niger, and A. terreus (1,237 isolates in total). Three laboratories (laboratories 1 to 3) used conidium counting. One laboratory standardized the inoculum using a spectrophotometer (that is, by use of the optical density [OD]) and is referred to as the OD laboratory. Correlation coefficients, intraclass correlation coefficients, and essential agreement were calculated, and 2-log-unit differences were assessed (paired t test). The MIC range for the blinded collection was 0.25 to 16 mg/liter, and a 1-dilution-step difference between the MIC50 and MIC90 across the four laboratories was detected and a 2-dilution-step difference between the modal MICs was detected. Compared to the results for laboratories 1 and 2, a significant correlation was found for the OD laboratory MIC data (correlation coefficients, 0.85 and 0.93, respectively; intraclass correlation coefficients, 0.88 and 0.96, respectively). The number of mutant isolates whose MICs overlapped those of the wild-type isolates was the lowest for the OD laboratory (14/30 [46.7%] mutant isolates), whereas the numbers were 18/30 (60%) isolates for laboratory 1, 17/30 (56.7%) isolates for laboratory 2, and 21/30 (70%) isolates for laboratory 3. For the A. flavus, A. fumigatus, A. nidulans, A. niger, and A. terreus isolates, comparative analysis again defined the MIC distributions from the OD laboratory to be in excellent agreement with those from laboratories 1 and 2

  9. Prevalence of airborne Aspergillus flavus in Khartoum (Sudan) airspora with reference to dusty weather and inoculum survival in simulated summer conditions.

    PubMed

    Abdalla, M H

    1988-12-01

    Khartoum air was scanned for airborne Aspergillus flavus for 12 months using the horizontal gravitational settling method. Frequency of occurrence was related to total fungal catch and dusty weather. The Aspergilli were prevalent (68% of total isolated/plate/month) and A. flavus constituted 31% of the total Aspergilli. In June (hot, dry & dusty) Aspergilli constituted 79% of the total isolates, whilst A. flavus represented 30% from amongst the other Aspergilli. A. flavus, A. niger, A. nidulans (conidial & ascosporic states), A. terreus, Eurotium amstelodami and A. fumigatus, in descending order of prevalence were isolated in June. Other pathogenic or potentially pathogenic forms, isolated, were Cladosporium, Curvularia and Penicillium. Amongst winter isolations A. flavus was sporadic to absent in occurrence. A. flavus spore inocula that underwent hourly intermitted exposure to 45 degrees C, showed a decrease in spore germinability as well as reduced germ length. PMID:3148861

  10. Aspergillus asper sp.nov. and Aspergillus collinsii sp.nov., from Aspergillus section Usti

    Technology Transfer Automated Retrieval System (TEKTRAN)

    In sampling fungi from the built environment, two isolates that could not confidently be placed in described species were encountered. Phenotypic analysis suggested that they belonged in Aspergillus sect. Usti. In order to verify the sectional placement and to assure that they were undescribed rathe...

  11. Quantitative Trait Locus (QTL) Mapping Reveals a Role for Unstudied Genes in Aspergillus Virulence

    PubMed Central

    Christians, Julian K.; Cheema, Manjinder S.; Vergara, Ismael A.; Watt, Cortney A.; Pinto, Linda J.; Chen, Nansheng; Moore, Margo M.

    2011-01-01

    Infections caused by the fungus Aspergillus are a major cause of morbidity and mortality in immunocompromised populations. To identify genes required for virulence that could be used as targets for novel treatments, we mapped quantitative trait loci (QTL) affecting virulence in the progeny of a cross between two strains of A. nidulans (FGSC strains A4 and A91). We genotyped 61 progeny at 739 single nucleotide polymorphisms (SNP) spread throughout the genome, and constructed a linkage map that was largely consistent with the genomic sequence, with the exception of one potential inversion of ∼527 kb on Chromosome V. The estimated genome size was 3705 cM and the average intermarker spacing was 5.0 cM. The average ratio of physical distance to genetic distance was 8.1 kb/cM, which is similar to previous estimates, and variation in recombination rate was significantly positively correlated with GC content, a pattern seen in other taxa. To map QTL affecting virulence, we measured the ability of each progeny strain to kill model hosts, larvae of the wax moth Galleria mellonella. We detected three QTL affecting in vivo virulence that were distinct from QTL affecting in vitro growth, and mapped the virulence QTL to regions containing 7–24 genes, excluding genes with no sequence variation between the parental strains and genes with only synonymous SNPs. None of the genes in our QTL target regions have been previously associated with virulence in Aspergillus, and almost half of these genes are currently annotated as “hypothetical”. This study is the first to map QTL affecting the virulence of a fungal pathogen in an animal host, and our results illustrate the power of this approach to identify a short list of unknown genes for further investigation. PMID:21559404

  12. RmtA, a Putative Arginine Methyltransferase, Regulates Secondary Metabolism and Development in Aspergillus flavus.

    PubMed

    Satterlee, Timothy; Cary, Jeffrey W; Calvo, Ana M

    2016-01-01

    Aspergillus flavus colonizes numerous oil seed crops such as corn, peanuts, treenuts and cotton worldwide, contaminating them with aflatoxin and other harmful potent toxins. In the phylogenetically related model fungus Aspergillus nidulans, the methyltransferase, RmtA, has been described to be involved in epigenetics regulation through histone modification. Epigenetics regulation affects a variety of cellular processes, including morphogenesis and secondary metabolism. Our study shows that deletion of rmtA in A. flavus results in hyperconidiating colonies, indicating that rmtA is a repressor of asexual development in this fungus. The increase in conidiation in the absence of rmtA coincides with greater expression of brlA, abaA, and wetA compared to that in the wild type. Additionally, the rmtA deletion mutant presents a drastic reduction or loss of sclerotial production, while forced expression of this gene increased the ability of this fungus to generate these resistant structures, revealing rmtA as a positive regulator of sclerotial formation. Importantly, rmtA is also required for the production of aflatoxin B1 in A. flavus, affecting the expression of aflJ. Furthermore, biosynthesis of additional metabolites is also controlled by rmtA, indicating a broad regulatory output in the control of secondary metabolism. This study also revealed that rmtA positively regulates the expression of the global regulatory gene veA, which could contribute to mediate the effects of rmtA on development and secondary metabolism in this relevant opportunistic plant pathogen. PMID:27213959

  13. Conidial Dihydroxynaphthalene Melanin of the Human Pathogenic Fungus Aspergillus fumigatus Interferes with the Host Endocytosis Pathway.

    PubMed

    Thywißen, Andreas; Heinekamp, Thorsten; Dahse, Hans-Martin; Schmaler-Ripcke, Jeannette; Nietzsche, Sandor; Zipfel, Peter F; Brakhage, Axel A

    2011-01-01

    Aspergillus fumigatus is the most important air-borne fungal pathogen of humans. The interaction of the pathogen with the host's immune system represents a key process to understand pathogenicity. For elimination of invading microorganisms, they need to be efficiently phagocytosed and located in acidified phagolysosomes. However, as shown previously, A. fumigatus is able to manipulate the formation of functional phagolysosomes. Here, we demonstrate that in contrast to pigmentless pksP mutant conidia of A. fumigatus, the gray-green wild-type conidia inhibit the acidification of phagolysosomes of alveolar macrophages, monocyte-derived macrophages, and human neutrophil granulocytes. Therefore, this inhibition is independent of the cell type and applies to the major immune effector cells required for defense against A. fumigatus. Studies with melanin ghosts indicate that the inhibitory effect of wild-type conidia is due to their dihydroxynaphthalene (DHN)-melanin covering the conidia, whereas the hydrophobin RodA rodlet layer plays no role in this process. This is also supported by the observation that pksP conidia still exhibit the RodA hydrophobin layer, as shown by scanning electron microscopy. Mutants defective in different steps of the DHN-melanin biosynthesis showed stronger inhibition than pksP mutant conidia but lower inhibition than wild-type conidia. Moreover, A. fumigatus and A. flavus led to a stronger inhibition of phagolysosomal acidification than A. nidulans and A. terreus. These data indicate that a certain type of DHN-melanin that is different in the various Aspergillus species, is required for maximal inhibition of phagolysosomal acidification. Finally, we identified the vacuolar ATPase (vATPase) as potential target for A. fumigatus based on the finding that addition of bafilomycin which inhibits vATPase, led to complete inhibition of the acidification whereas the fusion of phagosomes containing wild-type conidia and lysosomes was not affected.

  14. RmtA, a Putative Arginine Methyltransferase, Regulates Secondary Metabolism and Development in Aspergillus flavus

    PubMed Central

    Satterlee, Timothy; Cary, Jeffrey W.; Calvo, Ana M.

    2016-01-01

    Aspergillus flavus colonizes numerous oil seed crops such as corn, peanuts, treenuts and cotton worldwide, contaminating them with aflatoxin and other harmful potent toxins. In the phylogenetically related model fungus Aspergillus nidulans, the methyltransferase, RmtA, has been described to be involved in epigenetics regulation through histone modification. Epigenetics regulation affects a variety of cellular processes, including morphogenesis and secondary metabolism. Our study shows that deletion of rmtA in A. flavus results in hyperconidiating colonies, indicating that rmtA is a repressor of asexual development in this fungus. The increase in conidiation in the absence of rmtA coincides with greater expression of brlA, abaA, and wetA compared to that in the wild type. Additionally, the rmtA deletion mutant presents a drastic reduction or loss of sclerotial production, while forced expression of this gene increased the ability of this fungus to generate these resistant structures, revealing rmtA as a positive regulator of sclerotial formation. Importantly, rmtA is also required for the production of aflatoxin B1 in A. flavus, affecting the expression of aflJ. Furthermore, biosynthesis of additional metabolites is also controlled by rmtA, indicating a broad regulatory output in the control of secondary metabolism. This study also revealed that rmtA positively regulates the expression of the global regulatory gene veA, which could contribute to mediate the effects of rmtA on development and secondary metabolism in this relevant opportunistic plant pathogen. PMID:27213959

  15. 13C-NMR analysis of Aspergillus mutants disturbed in pyruvate metabolism.

    PubMed

    Dijkema, C; Visser, J

    1987-12-10

    The metabolic consequences of two defects in pyruvate metabolism of the hyphal fungus Aspergillus nidulans have been investigated by natural abundance 13C-NMR spectroscopy. A pyruvate dehydrogenase complex (pdh) mutant, grown on acetate, accumulates alanine upon starvation which is derived from mannitol reserves. The L-alanine level increases further upon incubation with the non-permissive substrate D-glucose. L-Glutamate is absent from these spectra as it is required both for the transamination of pyruvate and as a reaction on an impaired energy metabolism in such a pdh-deficient strain. A pyruvate carboxylase (pyc) mutant, grown upon acetate, only starts to accumulate alanine after a long incubation period with D-glucose, due to the long-lasting presence of phosphoenolpyruvate carboxykinase and malic enzyme, which are both induced by growth on acetate. When this strain is grown on D-fructose and L-glutamate, alanine also accumulates within 3 h upon transfer to D-glucose.

  16. Developmental regulators in Aspergillus fumigatus.

    PubMed

    Park, Hee-Soo; Yu, Jae-Hyuk

    2016-03-01

    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.

  17. Developmental regulators in Aspergillus fumigatus.

    PubMed

    Park, Hee-Soo; Yu, Jae-Hyuk

    2016-03-01

    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. PMID:26920882

  18. Atypical Aspergillus parasiticus isolates from pistachio with aflR gene nucleotide insertion identical to Aspergillus sojae

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Aflatoxins are the most toxic and carcinogenic secondary metabolites produced primarily by the filamentous fungi Aspergillus flavus and Aspergillus parasiticus. The toxins cause devastating economic losses because of strict regulations on distribution of contaminated products. Aspergillus sojae are...

  19. Onychomycosis caused by Aspergillus versicolor.

    PubMed

    Veraldi, Stefano; Chiaratti, Anna; Harak, Henry

    2010-07-01

    We report a case of onychomycosis caused by Aspergillus versicolor in a 66-year-old female patient. The infection was characterised clinically by yellowish pigmentation of the nail plate and mild nail bed hyperkeratosis of the first left toe. All other nails were normal. Three direct microscopical examinations of nail samples revealed the presence of hyaline hyphae as well as conidiophores. Pure colonies of A. versicolor were found in three cultures. The patient was successfully treated with oral itraconazole. PMID:19422523

  20. Two metabolites from Aspergillus flavipes.

    PubMed

    Clark, A M; Hufford, C D; Robertson, L W

    1977-01-01

    Two novel fungal metabolites, N-benzoyl-L-phenylalaninol (1a) and asperphenamate (2) were isolated from the culture filtrate and mycelium of Aspergillus flavipes ATCC 11013. N-benzoyl-L-phenylalaninol was identified by direct comparison with an authentic sample. The structure of asperphenamate is proposed as (S)-N-benzoyl-phenylalanine-(S)-2-benzamido-3-phenyl propyl ester, based on chemical and spectroscopic evidence. PMID:875642

  1. Induced Fungal Resistance to Insect Grazing: Reciprocal Fitness Consequences and Fungal Gene Expression in the Drosophila-Aspergillus Model System

    PubMed Central

    Caballero Ortiz, Silvia; Trienens, Monika; Rohlfs, Marko

    2013-01-01

    Background Fungi are key dietary resources for many animals. Fungi, in consequence, have evolved sophisticated physical and chemical defences for repelling and impairing fungivores. Expression of such defences may entail costs, requiring diversion of energy and nutrients away from fungal growth and reproduction. Inducible resistance that is mounted after attack by fungivores may allow fungi to circumvent the potential costs of defence when not needed. However, no information exists on whether fungi display inducible resistance. We combined organism and fungal gene expression approaches to investigate whether fungivory induces resistance in fungi. Methodology/Principal Findings Here we show that grazing by larval fruit flies, Drosophila melanogaster, induces resistance in the filamentous mould, Aspergillus nidulans, to subsequent feeding by larvae of the same insect. Larval grazing triggered the expression of various putative fungal resistance genes, including the secondary metabolite master regulator gene laeA. Compared to the severe pathological effects of wild type A. nidulans, which led to 100% insect mortality, larval feeding on a laeA loss-of-function mutant resulted in normal insect development. Whereas the wild type fungus recovered from larval grazing, larvae eradicated the chemically deficient mutant. In contrast, mutualistic dietary yeast, Saccharomyces cerevisiae, reached higher population densities when exposed to Drosophila larval feeding. Conclusions/Significance Our study presents novel evidence that insect grazing is capable of inducing resistance to further grazing in a filamentous fungus. This phenotypic shift in resistance to fungivory is accompanied by changes in the expression of genes involved in signal transduction, epigenetic regulation and secondary metabolite biosynthesis pathways. Depending on reciprocal insect-fungus fitness consequences, fungi may be selected for inducible resistance to maintain high fitness in fungivore-rich habitats

  2. Palmitoylation of the Cysteine Residue in the DHHC Motif of a Palmitoyl Transferase Mediates Ca2+ Homeostasis in Aspergillus

    PubMed Central

    Zhang, Yuanwei; Zheng, Qingqing; Sun, Congcong; Song, Jinxing; Gao, Lina; Zhang, Shizhu; Muñoz, Alberto; Read, Nick D.; Lu, Ling

    2016-01-01

    Finely tuned changes in cytosolic free calcium ([Ca2+]c) mediate numerous intracellular functions resulting in the activation or inactivation of a series of target proteins. Palmitoylation is a reversible post-translational modification involved in membrane protein trafficking between membranes and in their functional modulation. However, studies on the relationship between palmitoylation and calcium signaling have been limited. Here, we demonstrate that the yeast palmitoyl transferase ScAkr1p homolog, AkrA in Aspergillus nidulans, regulates [Ca2+]c homeostasis. Deletion of akrA showed marked defects in hyphal growth and conidiation under low calcium conditions which were similar to the effects of deleting components of the high-affinity calcium uptake system (HACS). The [Ca2+]c dynamics in living cells expressing the calcium reporter aequorin in different akrA mutant backgrounds were defective in their [Ca2+]c responses to high extracellular Ca2+ stress or drugs that cause ER or plasma membrane stress. All of these effects on the [Ca2+]c responses mediated by AkrA were closely associated with the cysteine residue of the AkrA DHHC motif, which is required for palmitoylation by AkrA. Using the acyl-biotin exchange chemistry assay combined with proteomic mass spectrometry, we identified protein substrates palmitoylated by AkrA including two new putative P-type ATPases (Pmc1 and Spf1 homologs), a putative proton V-type proton ATPase (Vma5 homolog) and three putative proteins in A. nidulans, the transcripts of which have previously been shown to be induced by extracellular calcium stress in a CrzA-dependent manner. Thus, our findings provide strong evidence that the AkrA protein regulates [Ca2+]c homeostasis by palmitoylating these protein candidates and give new insights the role of palmitoylation in the regulation of calcium-mediated responses to extracellular, ER or plasma membrane stress. PMID:27058039

  3. Spiro Fused Diterpene-Indole Alkaloids from a Creek-Bottom-Derived Aspergillus terreus

    PubMed Central

    Cai, Shengxin; Du, Lin; Gerea, Alexandra L.; King, Jarrod B.; You, Jianlan

    2013-01-01

    Four metabolites, teraspiridoles A–D (2–5), formed from the merger of diterpene and modified indole scaffold were obtained from an Aspergillus terreus isolate. The structures and absolute configurations of these natural products were established using NMR, mass spectrometry, Marfey’s method, VCD, and ECD data. Teraspiridole B (3) exhibited weak inhibition of planaria regeneration/survival. PMID:23924243

  4. Workflow to study genetic biodiversity of aflatoxigenic Aspergillus spp. in Georgia, USA

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Peanut seeds were sampled from the entire state of Georgia in 2014. More than 600 isolates of Aspergillus spp. were collected using modified-dichloran rose Bengal (MDRB) medium, 240 of those isolates were fingerprinted with 25 InDel markers within the aflatoxin-biosynthesis gene cluster (ABC). Clust...

  5. Ecophysiological characterization of Aspergillus carbonarius, Aspergillus tubingensis and Aspergillus niger isolated from grapes in Spanish vineyards.

    PubMed

    García-Cela, E; Crespo-Sempere, A; Ramos, A J; Sanchis, V; Marin, S

    2014-03-01

    The aim of this study was to evaluate the diversity of black aspergilli isolated from berries from different agroclimatic regions of Spain. Growth characterization (in terms of temperature and water activity requirements) of Aspergillus carbonarius, Aspergillus tubingensis and Aspergillus niger was carried out on synthetic grape medium. A. tubingensis and A. niger showed higher maximum temperatures for growth (>45 °C versus 40-42 °C), and lower minimum aw requirements (0.83 aw versus 0.87 aw) than A. carbonarius. No differences in growth boundaries due to their geographical origin were found within A. niger aggregate isolates. Conversely, A. carbonarius isolates from the hotter and drier region grew and produced OTA at lower aw than other isolates. However, little genetic diversity in A. carbonarius was observed for the microsatellites tested and the same sequence of β-tubulin gene was observed; therefore intraspecific variability did not correlate with the geographical origin of the isolates or with their ability to produce OTA. Climatic change prediction points to drier and hotter climatic scenarios where A. tubingensis and A. niger could be even more prevalent over A. carbonarius, since they are better adapted to extreme high temperature and drier conditions.

  6. On the relation between phosphate uptake and growth of the cyanobacterium Anacystis nidulans.

    PubMed

    Falkner, G; Wagner, F; Falkner, R

    1994-06-01

    The energy dependent phosphate uptake by the cyanobacterium Anacystis nidulans has been analysed in terms of a "linear energy converter" that describes the interrelationship between phosphate incorporation into the polyphosphate pool and the photosynthetic proton flux at the thylakoid membrane. It is assumed that both processes are coupled in such a way that uptake proceeds with optimal efficiency at the prevailing phosphate concentration in the growth medium. On the basis of this model two important parameters of the uptake system can be calculated: first, a conductivity coefficient that reflects the activity of the uptake system and second, a minimal threshold value of external phosphate at which uptake ceases. The theoretical data are shown to fit the experimentally observed uptake behaviour of cyanobacteria when the same population is subjected to a transition from growth on low to high phosphate concentrations, mimicking a situation that frequently leads in natural waters to an algal bloom. PMID:7987705

  7. Isoindolone-Containing Meroperpenoids from the Endophytic Fungus Emericella nidulans HDN12-249.

    PubMed

    Zhou, Haibo; Sun, Xinhua; Li, Na; Che, Qian; Zhu, Tianjiao; Gu, Qianqun; Li, Dehai

    2016-09-16

    Six isoindolone containing meroterpenoids, emericellolides A-C (1-3) and emeriphenolicins E-G (4-6), were isolated from a plant endophytic fungus Emericella nidulans HDN12-249. Emericellolides A-C (1-3) feature the unprecedented macrolide skeleton composed of an unusual l-glutamate fragment, an isoindolone unit, and a sesquiterpene moiety, while structures of emeriphenolicins E-G (4-6) with two farnesyl groups attached to one isoindolone unit are rare in isoindolone-derived meroterpenoids. These structures including the absolute configurations were established on the basis of MS, NMR, Mo2(AcO)4-induced ECD, Marfey's method, and chemical conversion. Compound 4 exhibited cytotoxicity against HeLa cells with IC50 value of 4.77 μM. PMID:27588428

  8. Cyclopiazonic Acid Biosynthesis of Aspergillus flavus and Aspergillus oryzae

    PubMed Central

    Chang, Perng-Kuang; Ehrlich, Kenneth C.; Fujii, Isao

    2009-01-01

    Cyclopiazonic acid (CPA) is an indole-tetramic acid neurotoxin produced by some of the same strains of A. flavus that produce aflatoxins and by some Aspergillus oryzae strains. Despite its discovery 40 years ago, few reviews of its toxicity and biosynthesis have been reported. This review examines what is currently known about the toxicity of CPA to animals and humans, both by itself or in combination with other mycotoxins. The review also discusses CPA biosynthesis and the genetic diversity of CPA production in A. flavus/oryzae populations. PMID:22069533

  9. Triton X-114 phase fractionation of membrane proteins of the cyanobacterium Anacystis nidulans R2.

    PubMed

    Bricker, T M; Sherman, L A

    1984-11-15

    The thylakoid polypeptides of the cyanobacterium Anacystis nidulans R2 were analyzed by Triton X-114 phase fractionation [C. Bordier (1981) J. Biol. Chem. 256, 1604-1607, as adapted for photosynthetic membranes by T.M. Bricker and L.A. Sherman (1982) FEBS Lett. 149, 197-202]. In this procedure, polypeptides with extensive hydrophobic regions (i.e., intrinsic proteins) form mixed micelles with Triton X-114, and are separated from extrinsic proteins by temperature-mediated precipitation of the mixed Triton X-114-intrinsic protein micelles. The polypeptide pattern after phase fractionation was highly complementary, with 62 of the observed 110 polypeptide components partitioning into the Triton X-114-enriched fraction. Identified polypeptides fractionating into the Triton X-114 phase included the apoproteins for Photosystems I and II, cytochromes f and b6, and the herbicide-binding protein. Identified polypeptides fractioning into the Triton X-114-depleted (aqueous) phase included the large and small subunits of RuBp carboxylase, cytochromes c550 and c554, and ferredoxin. Enzymatic radioiodination of the photosynthetic membranes followed by Triton X-114 phase fractionation allowed direct identification of intrinsic polypeptide components which possess surface-exposed regions susceptible to radioiodination. The most prominent of these polypeptides was a 34-kDa component which was associated with photosystem II. This phase partitioning procedure has been particularly helpful in the clarification of the identity of the membrane-associated cytochromes, and of photosystem II components. When coupled with surface-probing techniques, this procedure is very useful in identifying intrinsic proteins which possess surface-exposed domains. Phase fractionation, in conjunction with the isolation of specific membrane components and complexes, has allowed the identification of many of the important intrinsic thylakoid membrane proteins of A. nidulans R2.

  10. Pulmonary hyalinizing granuloma associated with Aspergillus infection.

    PubMed

    Pinckard, J Keith; Rosenbluth, Daniel B; Patel, Kishor; Dehner, Louis P; Pfeifer, John D

    2003-01-01

    A 38-year-old immunocompetent man with occupational exposure to Aspergillus presented with dyspnea, pleuritic chest pain, and hemoptysis. Chest roentgenograms and computed tomography scans demonstrated multiple pulmonary nodules bilaterally. An initial set of bronchial washing cultures grew Aspergillus fumigatus, serologic testing showed an elevated anti-Aspergillus titer, and immunodiffusion testing was positive for antibody against A. fumigatus and A. niger. There was no microbiologic or serologic evidence of infection by other pathogens, and no clinical or laboratory evidence of autoimmune disease. An open lung biopsy was diagnostic of pulmonary hyalinizing granuloma. This novel association with Aspergillus infection not only expands the spectrum of pathogens linked to pulmonary hyalinizing granuloma but also documents a new pattern of lung disease that can be caused by Aspergillus. PMID:12598920

  11. Conserved Secondary Structures in Aspergillus

    PubMed Central

    McGuire, Abigail Manson; Galagan, James E.

    2008-01-01

    Background Recent evidence suggests that the number and variety of functional RNAs (ncRNAs as well as cis-acting RNA elements within mRNAs ) is much higher than previously thought; thus, the ability to computationally predict and analyze RNAs has taken on new importance. We have computationally studied the secondary structures in an alignment of six Aspergillus genomes. Little is known about the RNAs present in this set of fungi, and this diverse set of genomes has an optimal level of sequence conservation for observing the correlated evolution of base-pairs seen in RNAs. Methodology/Principal Findings We report the results of a whole-genome search for evolutionarily conserved secondary structures, as well as the results of clustering these predicted secondary structures by structural similarity. We find a total of 7450 predicted secondary structures, including a new predicted ∼60 bp long hairpin motif found primarily inside introns. We find no evidence for microRNAs. Different types of genomic regions are over-represented in different classes of predicted secondary structures. Exons contain the longest motifs (primarily long, branched hairpins), 5′ UTRs primarily contain groupings of short hairpins located near the start codon, and 3′ UTRs contain very little secondary structure compared to other regions. There is a large concentration of short hairpins just inside the boundaries of exons. The density of predicted intronic RNAs increases with the length of introns, and the density of predicted secondary structures within mRNA coding regions increases with the number of introns in a gene. Conclusions/Sigificance There are many conserved, high-confidence RNAs of unknown function in these Aspergillus genomes, as well as interesting spatial distributions of predicted secondary structures. This study increases our knowledge of secondary structure in these aspergillus organisms. PMID:18665251

  12. New taxa of Neosartorya and Aspergillus in Aspergillus section Fumigati.

    PubMed

    Hong, Seung-Beom; Shin, Hyeon-Dong; Hong, Joonbae; Frisvad, Jens C; Nielsen, Per V; Varga, János; Samson, Robert A

    2008-01-01

    Three new species of Neosartorya and one new Aspergillus of section Fumigati are proposed using a polyphasic approach based on morphology, extrolite production and partial beta-tubulin, calmodulin, and actin gene sequences. The phylogenetic analyses using the three genes clearly show that the taxa grouped separately from the known species and confirmed the phenotypic differences. Neosartorya denticulata is characterized by its unique denticulate ascospores with a prominent equatorial furrow; N. assulata by well developed flaps on the convex surface of the ascospores which in addition have two distinct equatorial crests and N. galapagensis by a funiculose colony morphology, short and narrow conidiophores and ascospores with two wide equatorial crests with a microtuberculate convex surface. Aspergillus turcosus can be distinguished by velvety, gray turquoise colonies and short, loosely columnar conidial heads. The four new taxa also have unique extrolite profiles, which contain the mycotoxins gliotoxin and viriditoxin in N. denticulate; apolar compounds provisionally named NEPS in N. assulata and gregatins in N. galapagensis. A. turcosus produced kotanins. N. denticulata sp. nov., N. assulata sp. nov., N. galapagensis sp. nov., and A. turcosus sp. nov. are described and illustrated.

  13. New taxa of Neosartorya and Aspergillus in Aspergillus section Fumigati

    PubMed Central

    Hong, Seung-Beom; Shin, Hyeon-Dong; Hong, Joonbae; Frisvad, Jens C.; Nielsen, Per V.; Varga, János

    2007-01-01

    Three new species of Neosartorya and one new Aspergillus of section Fumigati are proposed using a polyphasic approach based on morphology, extrolite production and partial β-tubulin, calmodulin, and actin gene sequences. The phylogenetic analyses using the three genes clearly show that the taxa grouped separately from the known species and confirmed the phenotypic differences. Neosartorya denticulata is characterized by its unique denticulate ascospores with a prominent equatorial furrow; N. assulata by well developed flaps on the convex surface of the ascospores which in addition have two distinct equatorial crests and N. galapagensis by a funiculose colony morphology, short and narrow conidiophores and ascospores with two wide equatorial crests with a microtuberculate convex surface. Aspergillus turcosus can be distinguished by velvety, gray turquoise colonies and short, loosely columnar conidial heads. The four new taxa also have unique extrolite profiles, which contain the mycotoxins gliotoxin and viriditoxin in N. denticulate; apolar compounds provisionally named NEPS in N. assulata and gregatins in N. galapagensis. A. turcosus produced kotanins. N.denticulata sp. nov., N. assulata sp. nov., N. galapagensis sp. nov., and A. turcosus sp. nov. are described and illustrated. Electronic supplementary material The online version of this article (doi:10.1007/s10482-007-9183-1) contains supplementary material, which is available to authorized users. PMID:17610141

  14. Two novel species of Aspergillus section Nigri from indoor air

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Aspergillus collinsii, Aspergillus floridensis, and Aspergillus trinidadensis are described as novel uniseriate species of Aspergillus section Nigri isolated from air samples. To describe the species we used phenotypes from 7-d Czapek yeast extract agar culture (CYA) and malt extract agar culture (M...

  15. Secondary metabolite profiles and antifungal drug susceptibility of Aspergillus fumigatus and closely related species, Aspergillus lentulus, Aspergillus udagawae, and Aspergillus viridinutans.

    PubMed

    Tamiya, Hiroyuki; Ochiai, Eri; Kikuchi, Kazuyo; Yahiro, Maki; Toyotome, Takahito; Watanabe, Akira; Yaguchi, Takashi; Kamei, Katsuhiko

    2015-05-01

    The incidence of Aspergillus infection has been increasing in the past few years. Also, new Aspergillus fumigatus-related species, namely Aspergillus lentulus, Aspergillus udagawae, and Aspergillus viridinutans, were shown to infect humans. These fungi exhibit marked morphological similarities to A. fumigatus, albeit with different clinical courses and antifungal drug susceptibilities. The present study used liquid chromatography/time-of-flight mass spectrometry to identify the secondary metabolites secreted as virulence factors by these Aspergillus species and compared their antifungal susceptibility. The metabolite profiles varied widely among A. fumigatus, A. lentulus, A. udagawae, and A. viridinutans, producing 27, 13, 8, and 11 substances, respectively. Among the mycotoxins, fumifungin, fumiquinazoline A/B and D, fumitremorgin B, gliotoxin, sphingofungins, pseurotins, and verruculogen were only found in A. fumigatus, whereas auranthine was only found in A. lentulus. The amount of gliotoxin, one of the most abundant mycotoxins in A. fumigatus, was negligible in these related species. In addition, they had decreased susceptibility to antifungal agents such as itraconazole and voriconazole, even though metabolites that were shared in the isolates showing higher minimum inhibitory concentrations than epidemiological cutoff values were not detected. These strikingly different secondary metabolite profiles may lead to the development of more discriminative identification protocols for such closely related Aspergillus species as well as improved treatment outcomes.

  16. Secondary metabolite profiles and antifungal drug susceptibility of Aspergillus fumigatus and closely related species, Aspergillus lentulus, Aspergillus udagawae, and Aspergillus viridinutans.

    PubMed

    Tamiya, Hiroyuki; Ochiai, Eri; Kikuchi, Kazuyo; Yahiro, Maki; Toyotome, Takahito; Watanabe, Akira; Yaguchi, Takashi; Kamei, Katsuhiko

    2015-05-01

    The incidence of Aspergillus infection has been increasing in the past few years. Also, new Aspergillus fumigatus-related species, namely Aspergillus lentulus, Aspergillus udagawae, and Aspergillus viridinutans, were shown to infect humans. These fungi exhibit marked morphological similarities to A. fumigatus, albeit with different clinical courses and antifungal drug susceptibilities. The present study used liquid chromatography/time-of-flight mass spectrometry to identify the secondary metabolites secreted as virulence factors by these Aspergillus species and compared their antifungal susceptibility. The metabolite profiles varied widely among A. fumigatus, A. lentulus, A. udagawae, and A. viridinutans, producing 27, 13, 8, and 11 substances, respectively. Among the mycotoxins, fumifungin, fumiquinazoline A/B and D, fumitremorgin B, gliotoxin, sphingofungins, pseurotins, and verruculogen were only found in A. fumigatus, whereas auranthine was only found in A. lentulus. The amount of gliotoxin, one of the most abundant mycotoxins in A. fumigatus, was negligible in these related species. In addition, they had decreased susceptibility to antifungal agents such as itraconazole and voriconazole, even though metabolites that were shared in the isolates showing higher minimum inhibitory concentrations than epidemiological cutoff values were not detected. These strikingly different secondary metabolite profiles may lead to the development of more discriminative identification protocols for such closely related Aspergillus species as well as improved treatment outcomes. PMID:25737146

  17. Genetic diversity of Aspergillus species isolated from onychomycosis and Aspergillus hongkongensis sp. nov., with implications to antifungal susceptibility testing.

    PubMed

    Tsang, Chi-Ching; Hui, Teresa W S; Lee, Kim-Chung; Chen, Jonathan H K; Ngan, Antonio H Y; Tam, Emily W T; Chan, Jasper F W; Wu, Andrea L; Cheung, Mei; Tse, Brian P H; Wu, Alan K L; Lai, Christopher K C; Tsang, Dominic N C; Que, Tak-Lun; Lam, Ching-Wan; Yuen, Kwok-Yung; Lau, Susanna K P; Woo, Patrick C Y

    2016-02-01

    Thirteen Aspergillus isolates recovered from nails of 13 patients (fingernails, n=2; toenails, n=11) with onychomycosis were characterized. Twelve strains were identified by multilocus sequencing as Aspergillus spp. (Aspergillus sydowii [n=4], Aspergillus welwitschiae [n=3], Aspergillus terreus [n=2], Aspergillus flavus [n=1], Aspergillus tubingensis [n=1], and Aspergillus unguis [n=1]). Isolates of A. terreus, A. flavus, and A. unguis were also identifiable by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry. The 13th isolate (HKU49(T)) possessed unique morphological characteristics different from other Aspergillus spp. Molecular characterization also unambiguously showed that HKU49(T) was distinct from other Aspergillus spp. We propose the novel species Aspergillus hongkongensis to describe this previously unknown fungus. Antifungal susceptibility testing showed most Aspergillus isolates had low MICs against itraconazole and voriconazole, but all Aspergillus isolates had high MICs against fluconazole. A diverse spectrum of Aspergillus species is associated with onychomycosis. Itraconazole and voriconazole are probably better drug options for Aspergillus onychomycosis.

  18. [Aspergillus infection and chronic septic granulomatosis].

    PubMed

    Mouy, R; Bremard, C; Fischer, A; Huu Trong, P; Vilmer, E; Griscelli, C

    1985-12-01

    Chronic granulomatous disease of childhood is a hereditary abnormality of phagocytic cells, frequently associated with Aspergillus infections. From 1969 to 1984, 14 of 37 children with chronic granulomatous disease have presented with pulmonary (13 cases) and/or osteo-articular (1 case) aspergillosis. The paucity of symptoms was a characteristic of these infections. Lung lesions extending to the thoracic chest wall carried the bad prognosis. Neither the Aspergillus skin test nor the Aspergillus serology could definitely confirm the diagnosis. Only broncho-alveolar lavage and biopsy with isolation of Aspergillus could confirm the diagnosis. Long-term therapy with amphotericin B alone or associated with other antifungal agents is necessary. For the past 3 years, ketoconazole prophylaxis has been used in 23 children and none of these children has developed aspergillosis.

  19. Aspergillus granuloma of the trigeminal ganglion

    PubMed Central

    Wiles, C M; Kocen, R S; Symon, L; Scaravilli, F

    1981-01-01

    A patient is described with aspergillus flavus granuloma of the trigeminal ganglion. The patient was effectively treated by surgical excision of most of the infected tissue followed by intensive chemotherapy with amphotericin B and flucytosine. Images PMID:6973615

  20. Three new species of Aspergillus section Flavi isolated from almonds and maize in Portugal

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Three new aflatoxin-producing species belonging to Aspergillus section Flavi are described, Aspergillus mottae, Aspergillus sergii and Aspergillus transmontanensis. These species were isolated from Portuguese almonds and maize. An investigation examining morphology, extrolites and molecular data was...

  1. Characterization of the Aspergillus ochraceoroseus aflatoxin/sterigmatocystin biosynthetic gene cluster

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Production of the carcinogenic aflatoxins has been reported from members of Aspergillus section Flavi, Aspergillus section Nidulantes, and a newly proposed section, Aspergillus section Ochraceorosei that consists of Aspergillus ochraceoroseus and A. rambellii. Unlike members of section Flavi, A. oc...

  2. Methylcitrate synthase from Aspergillus fumigatus. Propionyl-CoA affects polyketide synthesis, growth and morphology of conidia.

    PubMed

    Maerker, Claudia; Rohde, Manfred; Brakhage, Axel A; Brock, Matthias

    2005-07-01

    Methylcitrate synthase is a key enzyme of the methylcitrate cycle and required for fungal propionate degradation. Propionate not only serves as a carbon source, but also acts as a food preservative (E280-283) and possesses a negative effect on polyketide synthesis. To investigate propionate metabolism from the opportunistic human pathogenic fungus Aspergillus fumigatus, methylcitrate synthase was purified to homogeneity and characterized. The purified enzyme displayed both, citrate and methylcitrate synthase activity and showed similar characteristics to the corresponding enzyme from Aspergillus nidulans. The coding region of the A. fumigatus enzyme was identified and a deletion strain was constructed for phenotypic analysis. The deletion resulted in an inability to grow on propionate as the sole carbon source. A strong reduction of growth rate and spore colour formation on media containing both, glucose and propionate was observed, which was coincident with an accumulation of propionyl-CoA. Similarly, the use of valine, isoleucine and methionine as nitrogen sources, which yield propionyl-CoA upon degradation, inhibited growth and polyketide production. These effects are due to a direct inhibition of the pyruvate dehydrogenase complex and blockage of polyketide synthesis by propionyl-CoA. The surface of conidia was studied by electron scanning microscopy and revealed a correlation between spore colour and ornamentation of the conidial surface. In addition, a methylcitrate synthase deletion led to an attenuation of virulence, when tested in an insect infection model and attenuation was even more pronounced, when whitish conidia from glucose/propionate medium were applied. Therefore, an impact of methylcitrate synthase in the infection process is discussed.

  3. Construction of a Shuttle Vector for Heterologous Expression of a Novel Fungal α-Amylase Gene in Aspergillus oryzae.

    PubMed

    Yin, Yanchen; Mao, Youzhi; Yin, Xiaolie; Gao, Bei; Wei, Dongzhi

    2015-07-01

    The filamentous fungus Aspergillus oryzae is a well-known expression host used to express homologous and heterologous proteins in a number of industrial applications. To facilitate higher yields of proteins of interest, we constructed the pAsOP vector to express heterologous proteins in A. oryzae. pAsOP carries a selectable marker, pyrG, derived from Aspergillus nidulans, and a strong promoter and a terminator of the amyB gene derived from A. oryzae. pAsOP transformed A. oryzae efficiently via the PEG-CaCl2-mediated transformation method. As proof of concept, green fluorescent protein (GFP) was successfully expressed in A. oryzae transformed by pAsOP-GFP. Additionally, we identified a novel fungal α-amylase (PcAmy) gene from Penicillium sp. and cloned the gene into the vector. After transformation by pAsOPPcAmy, the α-amylase PcAmy from Penicillium sp. was successfully expressed in a heterologous host system for the first time. The α-amylase activity in the A. oryzae transformant was increased by 62.3% compared with the untransformed A. oryzae control. The PcAmy protein produced in the system had an optimum pH of 5.0 and optimum temperature of 30°C. As a cold-adapted enzyme, PcAmy shows potential value in industrial applications because of its high catalytic activity at low temperature. Furthermore, the expression vector reported in this study provides promising utility for further scientific research and biotechnological applications.

  4. Transfer of the Excitation Energy in Anacystis nidulans Grown to Obtain Different Pigment Ratios

    PubMed Central

    Ghosh, A. K.; Govindjee

    1966-01-01

    The blue-green alga, Anacystis nidulans, was grown in lights of different colors and intensities, and its absorption and fluorescence properties were studied. Strong orange light, absorbed mainly by phycocyanin, causes reduction in the ratio of phycocyanin to chlorophyll a; strong red light, absorbed mainly by chlorophyll, causes an increase in this ratio. This confirms the earlier findings of Brody and Emerson (12) on Porphyridum, and of Jones and Myers (8) on Anacystis. Anacystis cultures grown in light of low intensity show, upon excitation of phycocyanin, emission peaks at 600 mμ and 680 mμ, due to the fluorescence of phycocyanin and chlorophyll a, respectively. Changes in the efficiency of energy transfer from phycocyanin to chlorophyll a are revealed by changes in the ratios of these two bands. A decrease in efficiency of energy transfer from phycocyanin to chlorophyll a seems to occur whenever the ratio of chlorophyll a to phycocyanin deviates from the normal. Algae grown in light of high intensity show, upon excitation of phycocyanin, only a fluorescence band at 660 mμ and no band at 680 mμ. This suggests reduced efficiency of energy transfer from phycocyanin to the strongly fluorescent form of chlorophyll a (chlorophyll a2) and perhaps increased transfer to the weakly fluorescent form of chlorophyll a (chlorophyll a1). PMID:5970565

  5. Glycolipid biosynthesis in cyanobacteria. [Anabaena variabilis; Chlorogloeopsis sp. ; Schizothrix calcicola; Anacystis nidulans; Anacystis marina

    SciTech Connect

    Van Dusen, W.J.; Jaworski, J.G.

    1987-05-01

    The biosynthesis of monogalactosyldiacyl-glycerol (MGDG) was studied in five different cyanobacteria. Previous work has shown Anabaena variabilis to synthesize both MGDG and monoglucosyl-diacylglycerol (MG1cDG) with MG1cDG being the precursor of MGDG. They have examined four other cyanobacteria to determine if a similar relationship exists. The cyanobacteria studied were Anabaena variabilis, Chlorogloeopsis sp., Schizothrix calcicola, Anacystis nidulans, and Anacystis marina. Each were grown in liquid culture and lipids were labeled with /sup 14/C)CO/sub 2/ for 20 min., 1.0 hr, 1.0 hr + 10 hr chase. Glycolipids were analyzed by initial separation of MGDG and MG1cDG by TLC followed by further analysis by HPLC. Complete separation of molecular species was obtained isocratically on an ODS column. All of the cyanobacteria labeled 16-C and 18-C fatty acids except for A. marina which labeled only 14-C and 16-C fatty acids. Desaturation of the fatty acids could be observed in the 1.0 hr and chase experiments. All were capable of labeling both MG1cDG and MGDG with the precursor-product relationship being observed. There does not appear to be a direct relationship between the epimerization of the sugar moiety and fatty acid desaturation.

  6. Membrane development in the cyanobacterium, Anacystis nidulans, during recovery from iron starvation

    SciTech Connect

    Pakrasi, H.B.; Goldenberg, A.; Sherman, L.A.

    1985-09-01

    Deprivation of iron from the growth medium results in physiological as well as structural changes in the unicellular cyanobacterium Anacystis nidulans R2. Important among these changes are alterations in the composition and function of the photosynthetic membranes. Room-temperature absorption spectra of iron-starved cyanobacterial cells show a chlorophyll absorption peak at 672 nanometers, 7 nanometers blue-shifted from its normal position at 679 nanometers. Iron-starved cells have decreased amounts of chlorophyll and phycobilins. Their fluorescence spectra (77K) have one prominent chlorophyll emission peak at 684 nanometers as compared to three peaks at 687, 696, and 717 nanometers from normal cells. Chlorophyll-protein analysis of iron-deprived cells indicated the absence of high molecular weight bands. Addition of iron to iron-starved cells induced a restoration process in which new components were initially synthesized and integrated into preexisting membranes; at later times, new membranes were assembled and cell division commenced. Synthesis of chlorophyll and phycocyanins started almost immediately after the addition of iron. The origin of the fluorescence emission at 687 and 696 nanometers is discussed in relation to the specific chlorophyll-protein complexes formed during iron reconstitution. 26 references, 2 figures, 1 table.

  7. Action of Heavy Metals on Hill Activity and O2 Evolution in Anacystis nidulans1

    PubMed Central

    Singh, Devendra P.; Singh, S. P.

    1987-01-01

    Addition of 5 micromolar Cu2+, Cd2+, and Zn2+ was inhibitory to 10 micromolar H2O2-supported Hill activity (dichlorophenolindophenol reduction) and O2 evolution in membrane preparation from Anacystis nidulans. The reversal of Cd2+ and Zn2+ inhibition, in contrast to Cu2+, by exogenously added catalase (EC 1.11.1.6) suggested that the former cations were inhibitory to H2O2 degradation. Ascorbic acid (20 micromolar) supported 27% of the Hill activity which was insensitive to DCMU (10 micromolar) and the remaining activity, attributable to the DCMU sensitive process, was sensitive to inhibition by Cu2+ only. It is suggestive that the action site of Cd2+ and Zn2+ is located between the electron donation sites of H2O2 and ascorbic acid, while that of Cu2+ is located beyond it. Electron donation by reduced glutathione was insensitive to DCMU and Cu2+, indicating that the action site of Cu2+ is prior to its electron donation site. Further, the phenanthroline (10 micromolar) reversal of Cu2+ inhibition of Hill activity suggested a tentative action site of Cu2+ at the level of cytochrome. PMID:16665185

  8. Post-Synthetic Defucosylation of AGP by Aspergillus nidulans α-1,2-Fucosidase Expressed in Arabidopsis Apoplast Induces Compensatory Upregulation of α-1,2-Fucosyltransferases

    PubMed Central

    Pogorelko, Gennady V.; Reem, Nathan T.; Young, Zachary T.; Chambers, Lauran; Zabotina, Olga A.

    2016-01-01

    Cell walls are essential components of plant cells which perform a variety of important functions for the different cell types, tissues and organs of a plant. Besides mechanical function providing cell shape, cell walls participate in intercellular communication, defense during plant-microbe interactions, and plant growth. The plant cell wall consists predominantly of polysaccharides with the addition of structural glycoproteins, phenolic esters, minerals, lignin, and associated enzymes. Alterations in the cell wall composition created through either changes in biosynthesis of specific constituents or their post-synthetic modifications in the apoplast compromise cell wall integrity and frequently induce plant compensatory responses as a result of these alterations. Here we report that post-synthetic removal of fucose residues specifically from arabinogalactan proteins in the Arabidopsis plant cell wall induces differential expression of fucosyltransferases and leads to the root and hypocotyl elongation changes. These results demonstrate that the post-synthetic modification of cell wall components presents a valuable approach to investigate the potential signaling pathways induced during plant responses to such modifications that usually occur during plant development and stress responses. PMID:27448235

  9. Septins AspA and AspC are important for normal development and limit the emergence of new growth foci in the multicellular fungus Aspergillus nidulans

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Septins are cytoskeletal proteins found in fungi, animals and microsporidia where they form multi-septin heteropolymeric complexes that act as scaffolds recruiting and organizing other proteins to ensure normal cell division and development. Here we characterize AspA and AspC, two of the five septin...

  10. Post-Synthetic Defucosylation of AGP by Aspergillus nidulans α-1,2-Fucosidase Expressed in Arabidopsis Apoplast Induces Compensatory Upregulation of α-1,2-Fucosyltransferases.

    PubMed

    Pogorelko, Gennady V; Reem, Nathan T; Young, Zachary T; Chambers, Lauran; Zabotina, Olga A

    2016-01-01

    Cell walls are essential components of plant cells which perform a variety of important functions for the different cell types, tissues and organs of a plant. Besides mechanical function providing cell shape, cell walls participate in intercellular communication, defense during plant-microbe interactions, and plant growth. The plant cell wall consists predominantly of polysaccharides with the addition of structural glycoproteins, phenolic esters, minerals, lignin, and associated enzymes. Alterations in the cell wall composition created through either changes in biosynthesis of specific constituents or their post-synthetic modifications in the apoplast compromise cell wall integrity and frequently induce plant compensatory responses as a result of these alterations. Here we report that post-synthetic removal of fucose residues specifically from arabinogalactan proteins in the Arabidopsis plant cell wall induces differential expression of fucosyltransferases and leads to the root and hypocotyl elongation changes. These results demonstrate that the post-synthetic modification of cell wall components presents a valuable approach to investigate the potential signaling pathways induced during plant responses to such modifications that usually occur during plant development and stress responses. PMID:27448235

  11. Studies on the properties of cellulase enzyme from Aspergillus terreus GN1

    SciTech Connect

    Garg, S.K.; Neelakantan, S.

    1982-03-01

    The cellulase enzyme is produced by cellulolytic microorganisms and is able to degrade cellulose materials. It has assumed greater importance due to its potential use in modifying low-grade roughages, the production of syrups, powering alcohol from cellulosic wastes, and other applications such as single-cell protein production. This paper studies the properties of a cellulase enzyme system from Aspergillus terreus GN1 and the effect of various modulators on its activity. (Refs. 18).

  12. Isolation and characterization of ultraviolet light-sensitive mutants of the blue-green alga Anacystis nidulans.

    NASA Technical Reports Server (NTRS)

    Asato, Y.

    1972-01-01

    Three independently isolated ultraviolet light sensitive (uvs) mutants of Anacystis nidulans were characterized. Strain uvs-1 showed the highest sensitivity to UV by its greatly reduced photoreactivation capacity following irradiation. Pretreatment with caffeine suppressed the dark-survival curve of strain uvs-1, thus indicating the presence of excision enzymes involved in dark repair. Under 'black' and 'white' illumination, strain uvs-1 shows photorecovery properties comparable with wild-type cultures. Results indicate that strains uvs-1, uvs-35, and uvs-88 are probably genetically distinct UV-sensitive mutants.

  13. The Volatome of Aspergillus fumigatus

    PubMed Central

    Calvo, A. M.; Latgé, J. P.

    2014-01-01

    Early detection of invasive aspergillosis is absolutely required for efficient therapy of this fungal infection. The identification of fungal volatiles in patient breath can be an alternative for the detection of Aspergillus fumigatus that still remains problematic. In this work, we investigated the production of volatile organic compounds (VOCs) by A. fumigatus in vitro, and we show that volatile production depends on the nutritional environment. A. fumigatus produces a multiplicity of VOCs, predominantly terpenes and related compounds. The production of sesquiterpenoid compounds was found to be strongly induced by increased iron concentrations and certain drugs, i.e., pravastatin. Terpenes that were always detectable in large amounts were α-pinene, camphene, and limonene, as well as sesquiterpenes, identified as α-bergamotene and β-trans-bergamotene. Other substance classes that were found to be present in the volatome, such as 1-octen-3-ol, 3-octanone, and pyrazines, were found only under specific growth conditions. Drugs that interfere with the terpene biosynthesis pathway influenced the composition of the fungal volatome, and most notably, a block of sesquiterpene biosynthesis by the bisphosphonate alendronate fundamentally changed the VOC composition. Using deletion mutants, we also show that a terpene cyclase and a putative kaurene synthase are essential for the synthesis of volatile terpenes by A. fumigatus. The present analysis of in vitro volatile production by A. fumigatus suggests that VOCs may be used in the diagnosis of infections caused by this fungus. PMID:24906414

  14. ASPERGILLUS LUCHUENSIS , AN INDUSTRIALLY IMPORTANT BLACK ASPERGILLUS IN EAST ASIA

    PubMed Central

    Hong, Seung-Beom; Lee, Mina; Kim, Dae-Ho; Varga, Janos; Frisvad, Jens C.; Perrone, Giancarlo; Gomi, Katsuya; Yamada, Osamu; Machida, Masayuki; Houbraken, Jos; Samson, Robert A.

    2013-01-01

    Aspergilli known as black- and white-koji molds which are used for awamori, shochu, makgeolli and other food and beverage fermentations, are reported in the literature as A. luchuensis, A. awamori, A. kawachii, or A. acidus. In order to elucidate the taxonomic position of these species, available ex-type cultures were compared based on morphology and molecular characters. A. luchuensis, A. kawachii and A. acidus showed the same banding patterns in RAPD, and the three species had the same rDNA-ITS, β-tubulin and calmodulin sequences and these differed from those of the closely related A. niger and A. tubingensis. Morphologically, the three species are not significantly different from each other or from A. niger and A. tubingensis. It is concluded that A. luchuensis, A. kawachii and A. acidus are the same species, and A. luchuensis is selected as the correct name based on priority. Strains of A. awamori which are stored in National Research Institute of Brewing in Japan, represent A. niger (n = 14) and A. luchuensis (n = 6). The neotype of A. awamori (CBS 557.65 =  NRRL 4948) does not originate from awamori fermentation and it is shown to be identical with the unknown taxon Aspergillus welwitschiae. Extrolite analysis of strains of A. luchuensis showed that they do not produce mycotoxins and therefore can be considered safe for food and beverage fermentations. A. luchuensis is also frequently isolated from meju and nuruk in Korea and Puerh tea in China and the species is probably common in the fermentation environment of East Asia. A re-description of A. luchuensis is provided because the incomplete data in the original literature. PMID:23723998

  15. Ribosomal ribonucleic acid and ribosomal precursor ribonucleic acid in Anacystis nidulans.

    PubMed

    Szalay, A; Munsche, D; Wolligiehn, R; Parthier, B

    1972-08-01

    The RNA of the blue-green alga Anacystis nidulans contains three ribosomal RNA species with molecular weights of 0.56x10(6), 0.9x10(6), and 1.1x10(6) if the RNA is extracted in the absence of Mg(2+). The 0.9x10(6)mol.wt. rRNA is extremely slowly labelled in (32)P-incorporation experiments. This rRNA may be a cleavage product of the 1.1x10(6)mol.wt. rRNA from the ribosomes of cells in certain physiological states (e.g. light-deficiency during growth). The cleavage of the 1.1x10(6)mol.wt. rRNA during the extraction procedure can be prevented by the addition of 10mm-MgCl(2). (32)P-pulse-labelling studies demonstrate the rapid synthesis of two ribosomal precursor RNA species. One precursor RNA migrating slightly slower than the 1.1x10(6)mol.wt. rRNA appears much less stable than the other precursor RNA, which shows the electrophoretic behaviour of the 0.7x10(6)mol.wt. rRNA. Our observations support the close relationship between bacteria and blue-green algae also with respect to rRNA maturation. The conversion of the ribosomal precursor RNA species into 0.56x10(6)- and 1.1x10(6)-mol.wt. rRNA species requires Mg(2+) in the incubation medium.

  16. Aspergillus flavus SUMO Contributes to Fungal Virulence and Toxin Attributes.

    PubMed

    Nie, Xinyi; Yu, Song; Qiu, Mengguang; Wang, Xiuna; Wang, Yu; Bai, Youhuang; Zhang, Feng; Wang, Shihua

    2016-09-01

    Small ubiquitin-like modifiers (SUMOs) can be reversibly attached to target proteins in a process known as SUMOylation, and this process influences several important eukaryotic cell events. However, little is known regarding SUMO or SUMOylation in Aspergillus flavus. Here, we identified a novel member of the SUMO family in A. flavus, AfSumO, and validated the existence of SUMOylation in this pathogenic filamentous fungus. We investigated the roles of AfsumO in A. flavus by determining the effects of AfsumO mutations on the growth phenotype, stress response, conidia and sclerotia production, aflatoxin biosynthesis, and pathogenicity to seeds, and we found that SUMOylation plays a role in fungal virulence and toxin attributes. Taken together, these results not only reveal potential mechanisms of fungal virulence and toxin attributes in A. flavus but also provide a novel approach for promising new control strategies of this fungal pathogen. PMID:27532332

  17. The gene for the small subunit of ribulose-1,5-bisphosphate carboxylase/oxygenase is located close to the gene for the large subunit in the cyanobacterium Anacystis nidulans 6301.

    PubMed Central

    Shinozaki, K; Sugiura, M

    1983-01-01

    The gene for the small subunit (SS) of ribulose-1,5-bisphosphate carboxylase/oxygenase from a cyanobacterium, Anacystis nidulans 6301, has been cloned and subjected to sequence analysis. The SS coding region is located close to and downstream from the large subunit (LS) coding region on the same DNA strand. The spacer region between the LS and the SS coding regions contains 93 base pairs (bp), and has no promoter-like sequences. The coding region of A. nidulans SS gene contains 333 bp (111 codons). The deduced amino acid sequence of the A. nidulans SS protein shows 40% homology with those of higher plants. Images PMID:6415615

  18. First case report of isolated aspergillus dacryoadenitis

    PubMed Central

    Acharya, Ishan; Basa, Divya; Kavitha, M

    2016-01-01

    We report a case of isolated Aspergillus dacryoadenitis. A 23-year-old male presented with dull ache, diffuse swelling in superolateral quadrant of the right orbit and proptosis for 4 months. Ocular examination showed conjunctival congestion, discharge in the fornix and palpable lacrimal gland (LG) mass. Routine hematological investigations followed by computed tomography scan of orbits were done. He did not respond to a course of systemic and topical antibiotics. Lateral orbitotomy with extended lid crease incision was performed with excision biopsy of LG. Abundant blackish material was found in the LG intraoperatively. The specimen was sent for histopathological examination (HPE). HPE report showed Aspergillus. Thorough ENT and systemic evaluation ruled out any other site with the fungus. To the best of our knowledge, this is the first case report of Aspergillus infection in LG. PMID:27488157

  19. Polyphasic taxonomy of Aspergillus section Usti

    PubMed Central

    Houbraken, J.; Due, M.; Varga, J.; Meijer, M.; Frisvad, J.C.; Samson, R.A.

    2007-01-01

    Aspergillus ustus is a very common species in foods, soil and indoor environments. Based on chemical, molecular and morphological data, A. insuetus is separated from A. ustus and revived. A. insuetus differs from A. ustus in producing drimans and ophiobolin G and H and not producing ustic acid and austocystins. The molecular, physiological and morphological data also indicated that another species, A. keveii sp. nov. is closely related but distinct from A. insuetus. Aspergillus section Usti sensu stricto includes 8 species: A. ustus, A. puniceus, A. granulosus, A. pseudodeflectus, A. calidoustus, A. insuetus and A. keveii together with Emericella heterothallica. PMID:18490949

  20. Enhanced diversity and aflatoxigenicity in interspecific hybrids of Aspergillus flavus and Aspergillus parasiticus

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Aspergillus flavus and A. parasiticus are two of the most important aflatoxin-producing species that contaminate agricultural commodities worldwide. Both species are heterothallic and undergo sexual reproduction in laboratory crosses. Here, we examine the possibility of interspecific matings betwe...

  1. Taxonomic Characterization and Secondary Metabolite Profiling of Aspergillus Section Aspergillus Contaminating Feeds and Feedstuffs

    PubMed Central

    Greco, Mariana; Kemppainen, Minna; Pose, Graciela; Pardo, Alejandro

    2015-01-01

    Xerophilic fungal species of the genus Aspergillus are economically highly relevant due to their ability to grow on low water activity substrates causing spoilage of stored goods and animal feeds. These fungi can synthesize a variety of secondary metabolites, many of which show animal toxicity, creating a health risk for food production animals and to humans as final consumers, respectively. Animal feeds used for rabbit, chinchilla and rainbow trout production in Argentina were analysed for the presence of xerophilic Aspergillus section Aspergillus species. High isolation frequencies (>60%) were detected in all the studied rabbit and chinchilla feeds, while the rainbow trout feeds showed lower fungal charge (25%). These section Aspergillus contaminations comprised predominantly five taxa. Twenty isolates were subjected to taxonomic characterization using both ascospore SEM micromorphology and two independent DNA loci sequencing. The secondary metabolite profiles of the isolates were determined qualitatively by HPLC-MS. All the isolates produced neoechinulin A, 17 isolates were positive for cladosporin and echinulin, and 18 were positive for neoechinulin B. Physcion and preechinulin were detected in a minor proportion of the isolates. This is the first report describing the detailed species composition and the secondary metabolite profiles of Aspergillus section Aspergillus contaminating animal feeds. PMID:26364643

  2. Taxonomic Characterization and Secondary Metabolite Profiling of Aspergillus Section Aspergillus Contaminating Feeds and Feedstuffs.

    PubMed

    Greco, Mariana; Kemppainen, Minna; Pose, Graciela; Pardo, Alejandro

    2015-09-01

    Xerophilic fungal species of the genus Aspergillus are economically highly relevant due to their ability to grow on low water activity substrates causing spoilage of stored goods and animal feeds. These fungi can synthesize a variety of secondary metabolites, many of which show animal toxicity, creating a health risk for food production animals and to humans as final consumers, respectively. Animal feeds used for rabbit, chinchilla and rainbow trout production in Argentina were analysed for the presence of xerophilic Aspergillus section Aspergillus species. High isolation frequencies (>60%) were detected in all the studied rabbit and chinchilla feeds, while the rainbow trout feeds showed lower fungal charge (25%). These section Aspergillus contaminations comprised predominantly five taxa. Twenty isolates were subjected to taxonomic characterization using both ascospore SEM micromorphology and two independent DNA loci sequencing. The secondary metabolite profiles of the isolates were determined qualitatively by HPLC-MS. All the isolates produced neoechinulin A, 17 isolates were positive for cladosporin and echinulin, and 18 were positive for neoechinulin B. Physcion and preechinulin were detected in a minor proportion of the isolates. This is the first report describing the detailed species composition and the secondary metabolite profiles of Aspergillus section Aspergillus contaminating animal feeds. PMID:26364643

  3. Taxonomic Characterization and Secondary Metabolite Profiling of Aspergillus Section Aspergillus Contaminating Feeds and Feedstuffs.

    PubMed

    Greco, Mariana; Kemppainen, Minna; Pose, Graciela; Pardo, Alejandro

    2015-09-02

    Xerophilic fungal species of the genus Aspergillus are economically highly relevant due to their ability to grow on low water activity substrates causing spoilage of stored goods and animal feeds. These fungi can synthesize a variety of secondary metabolites, many of which show animal toxicity, creating a health risk for food production animals and to humans as final consumers, respectively. Animal feeds used for rabbit, chinchilla and rainbow trout production in Argentina were analysed for the presence of xerophilic Aspergillus section Aspergillus species. High isolation frequencies (>60%) were detected in all the studied rabbit and chinchilla feeds, while the rainbow trout feeds showed lower fungal charge (25%). These section Aspergillus contaminations comprised predominantly five taxa. Twenty isolates were subjected to taxonomic characterization using both ascospore SEM micromorphology and two independent DNA loci sequencing. The secondary metabolite profiles of the isolates were determined qualitatively by HPLC-MS. All the isolates produced neoechinulin A, 17 isolates were positive for cladosporin and echinulin, and 18 were positive for neoechinulin B. Physcion and preechinulin were detected in a minor proportion of the isolates. This is the first report describing the detailed species composition and the secondary metabolite profiles of Aspergillus section Aspergillus contaminating animal feeds.

  4. Platelets enhance activity of antimycotic substances against non-Aspergillus fumigatus Aspergillus species in vitro.

    PubMed

    Perkhofer, Susanne; Trappl, Krista; Striessnig, Barbara; Nussbaumer, Walter; Lass-Flörl, Cornelia

    2011-02-01

    Platelets are known to be part of haemostasis but they are also players in innate host defense. Recently, we observed that platelets attenuate the virulence of Aspergillus spp. in vitro. However, little is known about the antifungal effects of platelets in the presence of antimycotics against non-A. fumigatus Aspergillus species. We therefore investigated whether platelets increase the in vitro activity of amphotericin B, voriconazole, posaconazole and caspofungin against two clinical isolates each of Aspergillus flavus, Aspergillus terreus and Aspergillus niger. The antifungal activity was evaluated by assessing germination percentages, hyphal elongation and hyphal damage by use of XTT. The combination of platelets plus amphotericin B significantly (P < 0.05) enhanced the reduction of germination percentage compared to either substance alone. Among triazoles, voriconazole exhibited significant effects with platelets for all tested aspergilli. Overall, these findings suggest that among the tested antimycotic substances, amphotericin B in combination with platelets has enhancing effects in reducing germination and hyphal elongation in the tested non-A. fumigatus Aspergillus species. These data indicate that platelets act beneficially with antimycotics in an early stage of fungal growth by blocking and/or delaying fungal germination and hyphal elongation; both crucial mechanisms in the development of invasive fungal disease.

  5. Biotransformation of Stypotriol triacetate by Aspergillus niger

    NASA Astrophysics Data System (ADS)

    Areche, Carlos; Vaca, Inmaculada; Labbe, Pamela; Soto-Delgado, Jorge; Astudillo, Luis; Silva, Mario; Rovirosa, Juana; San-Martin, Aurelio

    2011-07-01

    Biological transformation of the meroditerpenoid, stypotriol triacetate ( 1) by the fungi Aspergillus niger, Cunninghamella elegans, Gibberella fujikuroi and Mucor plumbeus was studied. The incubation of 1 with A. niger yielded the new compound 6',14-diacetoxy-stypol-4,5-dione ( 2) whose structure was established by 1H, 13C and 2D NMR and supported by DFT/GIAO.

  6. Interaction between maize seed and Aspergillus flavus

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Aspergillus flavus is an opportunistic fungal pathogen that colonizes maize seeds and contaminates them with aflatoxin. The fungus is localized in the endosperm and aleurone. To investigate the plant microbe interaction, we conducted histological and molecular studies to characterize the internal co...

  7. New taxa in Aspergillus section Usti

    PubMed Central

    Samson, R.A.; Varga, J.; Meijer, M.; Frisvad, J.C.

    2011-01-01

    Based on phylogenetic analysis of sequence data, Aspergillus section Usti includes 21 species, inclucing two teleomorphic species Aspergillus heterothallicus (= Emericella heterothallica) and Fennellia monodii. Aspergillus germanicus sp. nov. was isolated from indoor air in Germany. This species has identical ITS sequences with A. insuetus CBS 119.27, but is clearly distinct from that species based on β-tubulin and calmodulin sequence data. This species is unable to grow at 37 °C, similarly to A. keveii and A. insuetus. Aspergillus carlsbadensis sp. nov. was isolated from the Carlsbad Caverns National Park in New Mexico. This taxon is related to, but distinct from a clade including A. calidoustus, A. pseudodeflectus, A. insuetus and A. keveii on all trees. This species is also unable to grow at 37 °C, and acid production was not observed on CREA. Aspergillus californicus sp. nov. is proposed for an isolate from chamise chaparral (Adenostoma fasciculatum) in California. It is related to a clade including A. subsessilis and A. kassunensis on all trees. This species grew well at 37 °C, and acid production was not observed on CREA. The strain CBS 504.65 from soil in Turkey showed to be clearly distinct from the A. deflectus ex-type strain, indicating that this isolate represents a distinct species in this section. We propose the name A. turkensis sp. nov. for this taxon. This species grew, although rather restrictedly at 37 °C, and acid production was not observed on CREA. Isolates from stored maize, South Africa, as a culture contaminant of Bipolaris sorokiniana from indoor air in Finland proved to be related to, but different from A. ustus and A. puniceus. The taxon is proposed as the new species A. pseudoustus. Although supported only by low bootstrap values, F. monodii was found to belong to section Usti based on phylogenetic analysis of either loci BLAST searches to the GenBank database also resulted in closest hits from section Usti. This species obviously

  8. New taxa in Aspergillus section Usti.

    PubMed

    Samson, R A; Varga, J; Meijer, M; Frisvad, J C

    2011-06-30

    Based on phylogenetic analysis of sequence data, Aspergillus section Usti includes 21 species, inclucing two teleomorphic species Aspergillus heterothallicus (= Emericella heterothallica) and Fennellia monodii. Aspergillus germanicus sp. nov. was isolated from indoor air in Germany. This species has identical ITS sequences with A. insuetusCBS 119.27, but is clearly distinct from that species based on β-tubulin and calmodulin sequence data. This species is unable to grow at 37 °C, similarly to A. keveii and A. insuetus. Aspergillus carlsbadensis sp. nov. was isolated from the Carlsbad Caverns National Park in New Mexico. This taxon is related to, but distinct from a clade including A. calidoustus, A. pseudodeflectus, A. insuetus and A. keveii on all trees. This species is also unable to grow at 37 °C, and acid production was not observed on CREA. Aspergillus californicus sp. nov. is proposed for an isolate from chamise chaparral (Adenostoma fasciculatum) in California. It is related to a clade including A. subsessilis and A. kassunensis on all trees. This species grew well at 37 °C, and acid production was not observed on CREA. The strain CBS 504.65 from soil in Turkey showed to be clearly distinct from the A. deflectus ex-type strain, indicating that this isolate represents a distinct species in this section. We propose the name A. turkensis sp. nov. for this taxon. This species grew, although rather restrictedly at 37 °C, and acid production was not observed on CREA. Isolates from stored maize, South Africa, as a culture contaminant of Bipolaris sorokiniana from indoor air in Finland proved to be related to, but different from A. ustus and A. puniceus. The taxon is proposed as the new species A. pseudoustus. Although supported only by low bootstrap values, F. monodii was found to belong to section Usti based on phylogenetic analysis of either loci BLAST searches to the GenBank database also resulted in closest hits from section Usti. This species obviously

  9. 21 CFR 173.120 - Carbohydrase and cellulase derived from Aspergillus niger.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... cellulase derived from Aspergillus niger. Carbohydrase and cellulase enzyme preparation derived from Aspergillus niger may be safely used in food in accordance with the following prescribed conditions: (a) Aspergillus niger is classified as follows: Class, Deuteromycetes; order, Moniliales; family,...

  10. In vitro interactions of antifungal agents and tacrolimus against Aspergillus biofilms.

    PubMed

    Gao, Lujuan; Sun, Yi

    2015-11-01

    Aspergillus biofilms were prepared from Aspergillus fumigatus, Aspergillus flavus, and Aspergillus terreus via a 96-well plate-based method, and the combined antifungal activity of tacrolimus with azoles or amphotericin B against Aspergillus biofilms was investigated via a broth microdilution checkerboard technique system. Our results suggest that combinations of tacrolimus with voriconazole or amphotericin B have synergistic inhibitory activity against Aspergillus biofilms. However, combinations of tacrolimus with itraconazole or posaconazole exhibit no synergistic or antagonistic effects.

  11. Deletion of the Ustilago maydis ortholog of the Aspergillus sporulation regulator medA affects mating and virulence through pheromone response.

    PubMed

    Chacko, Nadia; Gold, Scott

    2012-06-01

    Mating of compatible haploid cells of Ustilago maydis is essential for infection and disease development in the host. For mating and subsequent filamentous growth and pathogenicity, the transcription factor, prf1 is necessary. Prf1 is in turn regulated by the cAMP and MAPK pathways and other regulators like rop1 and hap1. Here we describe the identification of another putative Prf1 regulator, med1, the ortholog of the Aspergillus nidulans medusa (medA) transcription factor and show that it is required for mating and full virulence in U. maydis. med1 deletion mutants show both pre- and post-mating defects and are unresponsive to external pheromone. The expression of prf1 is down-regulated in Δmed1 compared to the wild type, suggesting that med1 is upstream of prf1. Additionally, indicative of a role in secondary metabolism regulation, deletion of the med1 gene de-represses the production of glycolipids in U. maydis.

  12. Pathogenesis of Aspergillus fumigatus in Invasive Aspergillosis

    PubMed Central

    Dagenais, Taylor R. T.; Keller, Nancy P.

    2009-01-01

    Summary: Aspergillus species are globally ubiquitous saprophytes found in a variety of ecological niches. Almost 200 species of aspergilli have been identified, less than 20 of which are known to cause human disease. Among them, Aspergillus fumigatus is the most prevalent and is largely responsible for the increased incidence of invasive aspergillosis (IA) in the immunocompromised patient population. IA is a devastating illness, with mortality rates in some patient groups reaching as high as 90%. Studies identifying and assessing the roles of specific factors of A. fumigatus that contribute to the pathogenesis of IA have traditionally focused on single-gene deletion and mutant characterization. In combination with recent large-scale approaches analyzing global fungal responses to distinct environmental or host conditions, these studies have identified many factors that contribute to the overall pathogenic potential of A. fumigatus. Here, we provide an overview of the significant findings regarding A. fumigatus pathogenesis as it pertains to invasive disease. PMID:19597008

  13. Comparative Reannotation of 21 Aspergillus Genomes

    SciTech Connect

    Salamov, Asaf; Riley, Robert; Kuo, Alan; Grigoriev, Igor

    2013-03-08

    We used comparative gene modeling to reannotate 21 Aspergillus genomes. Initial automatic annotation of individual genomes may contain some errors of different nature, e.g. missing genes, incorrect exon-intron structures, 'chimeras', which fuse 2 or more real genes or alternatively splitting some real genes into 2 or more models. The main premise behind the comparative modeling approach is that for closely related genomes most orthologous families have the same conserved gene structure. The algorithm maps all gene models predicted in each individual Aspergillus genome to the other genomes and, for each locus, selects from potentially many competing models, the one which most closely resembles the orthologous genes from other genomes. This procedure is iterated until no further change in gene models is observed. For Aspergillus genomes we predicted in total 4503 new gene models ( ~;;2percent per genome), supported by comparative analysis, additionally correcting ~;;18percent of old gene models. This resulted in a total of 4065 more genes with annotated PFAM domains (~;;3percent increase per genome). Analysis of a few genomes with EST/transcriptomics data shows that the new annotation sets also have a higher number of EST-supported splice sites at exon-intron boundaries.

  14. Aspergillus asexual reproduction and sexual reproduction are differentially affected by transcriptional and translational mechanisms regulating stunted gene expression.

    PubMed Central

    Wu, J; Miller, B L

    1997-01-01

    The Stunted protein (StuAp) is a member of a family of transcription factors that regulate fungal development and cell cycle progression. Regulated stuA gene expression is required for correct cell pattern formation during asexual reproduction (conidiation) and for initiation of the sexual reproductive cycle in Aspergillus nidulans. Transcriptional initiation from two different promoters yields overlapping mRNAs (stuA alpha and stuAbeta) that upon translation yield the same protein. Here we show that multiple regulatory mechanisms interact to control (i) developmental competence-dependent expression of both transcripts and (ii) induction-dependent expression of stuA alpha, but not stuAbeta, by the conidiation-specific Bristle (BrlAp) transcriptional activator. Quantitative levels of both mRNAs are further modulated by (i) an activator(s) located at a far-upstream upstream activation sequence, (ii) feedback regulation by StuAp, and (iii) positive translational regulation that requires the peptide product of a micro-open reading frame unique to the stuA alpha mRNA 5' untranslated region. Gradients in stuA alpha expression were most important for correct cell and tissue type development. Threshold requirements were as follows: metula-phialide differentiation < ascosporogenesis < cleistothecial shell-Hülle cell differentiation. Altered stuA expression affected conidiophore morphology and conidial yields quantitatively but did not alter the temporal development of cell types or conidiophore density. By contrast, the sexual cycle showed both temporal delay and quantitative reduction in the number of cleistothecial initials but normal morphogenesis of tissue types. PMID:9315680

  15. Antibiotic Extraction as a Recent Biocontrol Method for Aspergillus Niger andAspergillus Flavus Fungi in Ancient Egyptian mural paintings

    NASA Astrophysics Data System (ADS)

    Hemdan, R. Elmitwalli; Fatma, Helmi M.; Rizk, Mohammed A.; Hagrassy, Abeer F.

    Biodeterioration of mural paintings by Aspergillus niger and Aspergillus flavus Fungi has been proved in different mural paintings in Egypt nowadays. Several researches have studied the effect of fungi on mural paintings, the mechanism of interaction and methods of control. But none of these researches gives us the solution without causing a side effect. In this paper, for the first time, a recent treatment by antibiotic "6 penthyl α pyrone phenol" was applied as a successful technique for elimination of Aspergillus niger and Aspergillus flavus. On the other hand, it is favorable for cleaning Surfaces of Murals executed by tembera technique from the fungi metabolism which caused a black pigments on surfaces.

  16. Fractionation of Aspergillus niger cellulases by combined ion exchange affinity chromatography

    SciTech Connect

    Boyer, R.F.; Allen, T.L.; Dykema, P.A.

    1987-02-05

    Eight chemically modified cellulose supports were tested for their ability to adsorb components of the Aspergillus niger cellulase system. At least two of the most effective adsorbents, aminoethyl cellulose and carboxymethyl cellulose, were shown to be useful for the fractionation of cellulases. These supports apparently owe their resolving capacity to both ion exchange and biospecific binding effects; however, the relative importance of each effect is unknown. These observations form the basis for a new cellulase fractionation technique, combined ion exchange-affinity chromatography. 22 references.

  17. Phylogeny, identification and nomenclature of the genus Aspergillus.

    PubMed

    Samson, R A; Visagie, C M; Houbraken, J; Hong, S-B; Hubka, V; Klaassen, C H W; Perrone, G; Seifert, K A; Susca, A; Tanney, J B; Varga, J; Kocsubé, S; Szigeti, G; Yaguchi, T; Frisvad, J C

    2014-06-01

    Aspergillus comprises a diverse group of species based on morphological, physiological and phylogenetic characters, which significantly impact biotechnology, food production, indoor environments and human health. Aspergillus was traditionally associated with nine teleomorph genera, but phylogenetic data suggest that together with genera such as Polypaecilum, Phialosimplex, Dichotomomyces and Cristaspora, Aspergillus forms a monophyletic clade closely related to Penicillium. Changes in the International Code of Nomenclature for algae, fungi and plants resulted in the move to one name per species, meaning that a decision had to be made whether to keep Aspergillus as one big genus or to split it into several smaller genera. The International Commission of Penicillium and Aspergillus decided to keep Aspergillus instead of using smaller genera. In this paper, we present the arguments for this decision. We introduce new combinations for accepted species presently lacking an Aspergillus name and provide an updated accepted species list for the genus, now containing 339 species. To add to the scientific value of the list, we include information about living ex-type culture collection numbers and GenBank accession numbers for available representative ITS, calmodulin, β-tubulin and RPB2 sequences. In addition, we recommend a standard working technique for Aspergillus and propose calmodulin as a secondary identification marker. PMID:25492982

  18. Fatal coinfection with Legionella pneumophila serogroup 8 and Aspergillus fumigatus.

    PubMed

    Guillouzouic, Aurélie; Bemer, Pascale; Gay-Andrieu, Françoise; Bretonnière, Cédric; Lepelletier, Didier; Mahé, Pierre-Joachim; Villers, Daniel; Jarraud, Sophie; Reynaud, Alain; Corvec, Stéphane

    2008-02-01

    Legionella pneumophila is an important cause of community-acquired and nosocomial pneumonia. We report on a patient who simultaneously developed L. pneumophila serogroup 8 pneumonia and Aspergillus fumigatus lung abscesses. Despite appropriate treatments, Aspergillus disease progressed with metastasis. Coinfections caused by L. pneumophila and A. fumigatus remain exceptional. In apparently immunocompetent patients, corticosteroid therapy is a key risk factor for aspergillosis.

  19. Biodiversity of Aspergillus Species in Some Important Agricultural Products

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The genus Aspergillus is one of the most important filamentous fungal genera. Aspergillus species are used in the fermentation industry, but they are also responsible of various plant and food secondary rot, with the consequence of possible accumulation of mycotoxins. The aflatoxin-producing A. fl...

  20. Genomic Islands in Pathogenic Filamentous Fungus Aspergillus fumigatus

    Technology Transfer Automated Retrieval System (TEKTRAN)

    We present the genome sequences of a new clinical isolate, CEA10, of an important human pathogen, Aspergillus fumigatus, and two closely related, but rarely pathogenic species, Neosartorya fischeri NRRL181 and Aspergillus clavatus NRRL1. Comparative genomic analysis of CEA10 with the recently sequen...

  1. Phylogeny, identification and nomenclature of the genus Aspergillus

    PubMed Central

    Samson, R.A.; Visagie, C.M.; Houbraken, J.; Hong, S.-B.; Hubka, V.; Klaassen, C.H.W.; Perrone, G.; Seifert, K.A.; Susca, A.; Tanney, J.B.; Varga, J.; Kocsubé, S.; Szigeti, G.; Yaguchi, T.; Frisvad, J.C.

    2014-01-01

    Aspergillus comprises a diverse group of species based on morphological, physiological and phylogenetic characters, which significantly impact biotechnology, food production, indoor environments and human health. Aspergillus was traditionally associated with nine teleomorph genera, but phylogenetic data suggest that together with genera such as Polypaecilum, Phialosimplex, Dichotomomyces and Cristaspora, Aspergillus forms a monophyletic clade closely related to Penicillium. Changes in the International Code of Nomenclature for algae, fungi and plants resulted in the move to one name per species, meaning that a decision had to be made whether to keep Aspergillus as one big genus or to split it into several smaller genera. The International Commission of Penicillium and Aspergillus decided to keep Aspergillus instead of using smaller genera. In this paper, we present the arguments for this decision. We introduce new combinations for accepted species presently lacking an Aspergillus name and provide an updated accepted species list for the genus, now containing 339 species. To add to the scientific value of the list, we include information about living ex-type culture collection numbers and GenBank accession numbers for available representative ITS, calmodulin, β-tubulin and RPB2 sequences. In addition, we recommend a standard working technique for Aspergillus and propose calmodulin as a secondary identification marker. PMID:25492982

  2. Nationwide Surveillance of Azole Resistance in Aspergillus Diseases.

    PubMed

    Vermeulen, Edith; Maertens, Johan; De Bel, Annelies; Nulens, Eric; Boelens, Jerina; Surmont, Ignace; Mertens, Anna; Boel, An; Lagrou, Katrien

    2015-08-01

    Aspergillus disease affects a broad patient population, from patients with asthma to immunocompromised patients. Azole resistance has been increasingly reported in both clinical and environmental Aspergillus strains. The prevalence and clinical impact of azole resistance in different patient populations are currently unclear. This 1-year prospective multicenter cohort study aimed to provide detailed epidemiological data on Aspergillus resistance among patients with Aspergillus disease in Belgium. Isolates were prospectively collected in 18 hospitals (April 2011 to April 2012) for susceptibility testing. Clinical and treatment data were collected with a questionnaire. The outcome was evaluated to 1 year after a patient's inclusion. A total of 220 Aspergillus isolates from 182 patients were included. The underlying conditions included invasive aspergillosis (n = 122 patients), allergic bronchopulmonary aspergillosis (APBA) (n = 39 patients), chronic pulmonary aspergillosis (n = 10 patients), Aspergillus bronchitis (n = 7 patients), and aspergilloma (n = 5 patients). The overall azole resistance prevalence was 5.5% (95% confidence interval [CI] 2.8 to 10.2%) and was 7.0% (4/57; 95% CI, 2.3 to 17.2%) in patients with APBA, bronchitis, aspergilloma, or chronic aspergillosis and 4.6% in patients with invasive aspergillosis (5/108; 95% CI, 1.7 to 10.7%). The 6-week survival in invasive aspergillosis was 52.5%, while susceptibility testing revealed azole resistance in only 2/58 of the deceased patients. The clinical impact of Aspergillus fumigatus resistance was limited in our patient population with Aspergillus diseases.

  3. Fatal coinfection with Legionella pneumophila serogroup 8 and Aspergillus fumigatus.

    PubMed

    Guillouzouic, Aurélie; Bemer, Pascale; Gay-Andrieu, Françoise; Bretonnière, Cédric; Lepelletier, Didier; Mahé, Pierre-Joachim; Villers, Daniel; Jarraud, Sophie; Reynaud, Alain; Corvec, Stéphane

    2008-02-01

    Legionella pneumophila is an important cause of community-acquired and nosocomial pneumonia. We report on a patient who simultaneously developed L. pneumophila serogroup 8 pneumonia and Aspergillus fumigatus lung abscesses. Despite appropriate treatments, Aspergillus disease progressed with metastasis. Coinfections caused by L. pneumophila and A. fumigatus remain exceptional. In apparently immunocompetent patients, corticosteroid therapy is a key risk factor for aspergillosis. PMID:17945454

  4. Aspergillus pragensis sp. nov. discovered during molecular reidentification of clinical isolates belonging to Aspergillus section Candidi

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The identity of nine clinical isolates from Czech patients presumably belonging to Aspergillus section Candidi based on morphology of colonies was revised using sequences of ß-tubulin, calmodulin, and internal transcribed spacer (ITS) rDNA. The set of isolates included six isolates from suspected (n...

  5. Reduction of aflatoxin production by Aspergillus flavus and Aspergillus parasiticus in interaction with Streptomyces.

    PubMed

    Verheecke, C; Liboz, T; Anson, P; Diaz, R; Mathieu, F

    2015-05-01

    The aim of this study is to investigate aflatoxin gene expression during Streptomyces-Aspergillus interaction. Aflatoxins are carcinogenic compounds produced mainly by Aspergillus flavus and Aspergillus parasiticus. A previous study has shown that Streptomyces-A. flavus interaction can reduce aflatoxin content in vitro. Here, we first validated this same effect in the interaction with A. parasiticus. Moreover, we showed that growth reduction and aflatoxin content were correlated in A. parasiticus but not in A. flavus. Secondly, we investigated the mechanisms of action by reverse-transcriptase quantitative PCR. As microbial interaction can lead to variations in expression of household genes, the most stable [act1, βtub (and cox5 for A. parasiticus)] were chosen using geNorm software. To shed light on the mechanisms involved, we studied during the interaction the expression of five genes (aflD, aflM, aflP, aflR and aflS). Overall, the results of aflatoxin gene expression showed that Streptomyces repressed gene expression to a greater level in A. parasiticus than in A. flavus. Expression of aflR and aflS was generally repressed in both Aspergillus species. Expression of aflM was repressed and was correlated with aflatoxin B1 content. The results suggest that aflM expression could be a potential aflatoxin indicator in Streptomyces species interactions. Therefore, we demonstrate that Streptomyces can reduce aflatoxin production by both Aspergillus species and that this effect can be correlated with the repression of aflM expression.

  6. Constitutive expression of fluorescent protein by Aspergillus var. niger and Aspergillus carbonarius to monitor fungal colonization in maize plants

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Aspergillus niger and A. carbonarius are two species in the Aspergillus section Nigri (black-spored aspergilli) frequently associated with peanut (Arachis hypogea), maize (Zea mays), and other plants as pathogens. These infections are symptomless and as such are major concerns since some black aspe...

  7. Empyema necessitatis due to Aspergillus fumigatus.

    PubMed

    Lee, Hyun Woo; Kim, Yeon Wook; Cho, Jaeyoung; Lee, Chang-Hoon

    2014-01-01

    We present an extremely rare case of empyema necessitatis secondary to Aspergillus fumigatus infection. A 58-year-old woman presented to our hospital with a painful skin rash on the right thorax. Three fistulas communicating with the pleural space were found. Since she did not show a clinical improvement despite antituberculous and antibacterial treatment, we looked for other causes. Pleural fungus culture showed A. fumigatus and chest wall biopsy revealed numerous fungal hyphae. Treatment with necrotic tissue debridement and antifungal agents was successful. PMID:25452298

  8. Testing the efficacy of RNA interference constructs in Aspergillus fumigatus.

    PubMed

    Henry, Christine; Mouyna, Isabelle; Latgé, Jean-Paul

    2007-04-01

    We recently developed a silencing vector in Aspergillus fumigatus which carries a hygromycin resistance marker and a transcriptional unit for hairpin RNA expression under the control of the inducible glucoamylase promoter (pGla) (Mouyna et al. in FEMS Microbiol Lett 237:317-324, 2004). We showed previously that this vector can be used for the RNA interference application of two genes ALB1 and FKS1 of which reduced mRNA levels occurred for both, with phenotypic consequences resembling disruptions of genes involved in melanin (ALB1) and beta(1-3)glucan biosynthesis (FKS1). We reported here the silencing of KRE6 and CRH1, two other genes putatively involved in cell wall biosynthesis using a similar construction under the control of the constitutive promoter glyceraldehyde-3-phosphate dehydrogenase (pgpdA). Silencing of the expression of these two genes was obtained. Further analysis of the transformants showed however that (1) a 100% loss of expression was never achieved for all genes tested (2) the vector used for RNAi is lost or modified over successive transfers resulting in an inhibition of the silencing. These disadvantages of RNAi indicate that classical gene disruption by gene replacement remains the most efficient method for a molecular analysis of gene function in A. fumigatus. PMID:17273823

  9. Aspergillus terreus recovered from a corneal scraping.

    PubMed

    Campbell, Suzanne

    2014-01-01

    A 52 year old, healthy male presented to his optometrist complaining of redness and irritation in the right eye. A foreign body was removed from the eye. The patient was started on ophthalmic solutions of vigamox and systane. At 48 hours, the patient reported increased redness, limited vision, and yellow discharge from the eye. The patient was referred to an ophthalmologist for further evaluation. Physical assessment revealed a superlative central infiltrate (extreme, centrally located injury that had permeated the cornea), diffuse corneal haze, and edema with a 3- to 4+ conjunctival injection and a 1 millimeter hypopyon (an effusion of pus into the anterior chamber of the eye). Corneal scrapings were collected for aerobic and anaerobic bacterial and fungal cultures. The patient was then prescribed. vancomycin, tobramycin, and natamycin ophthalmic eyedrops. On day three, fungal culture results indicated possible fungal forms seen. On day 12, results from the fungal culture of the corneal scraping revealed the causative agent to be Aspergillus terreus. Voriconazole eyedrops were added to the treatment regimen and continued for 10 weeks. The physician order for a fungal culture as well as laboratory data providing the final identification of Aspergillus terreus and laboratory comments indicating an elevated minimum inhibitory concentration (MIC) (> 2 microg/mL) to amphotericin B is associated with treatment failure positively impacted the patient outcome. After completion of the treatment regimen, a photo-therapeutic keratectomy (PTK) was performed in an attempt to remove the dense corneal scarring caused by the fungal infection.

  10. Chronological aging in conidia of pathogenic Aspergillus: Comparison between species.

    PubMed

    Oliveira, Manuela; Pereira, Clara; Bessa, Cláudia; Araujo, Ricardo; Saraiva, Lucília

    2015-11-01

    Aspergillus fumigatus, Aspergillus flavus, Aspergillus terreus and Aspergillus niger are common airborne fungi, and the most frequent causative agents of human fungal infections. However, the resistance and lifetime persistence of these fungi in the atmosphere, and the mechanism of aging of Aspergillus conidia are unknown.With this work, we intended to study the processes underlying conidial aging of these four relevant and pathogenic Aspergillus species. Chronological aging was therefore evaluated in A. fumigatus, A. flavus, A. terreus and A. niger conidia exposed to environmental and human body temperatures. The results showed that the aging process in Aspergillus conidia involves apoptosis,with metacaspase activation, DNA fragmentation, and reactive oxygen species production, associated with secondary necrosis. Distinct results were observed for the selected pathogenic species. At environmental conditions, A. niger was the species with the highest resistance to aging, indicating a higher adaption to environmental conditions, whereas A. flavus followed by A. terreus were the most sensitive species. At higher temperatures (37 °C), A. fumigatus presented the longest lifespan, in accordance with its good adaptation to the human body temperature. Altogether,with this work new insights regarding conidia aging are provided, which may be useful when designing treatments for aspergillosis.

  11. 2-hydroxyisocaproic acid is fungicidal for Candida and Aspergillus species.

    PubMed

    Sakko, M; Moore, C; Novak-Frazer, L; Rautemaa, V; Sorsa, T; Hietala, P; Järvinen, A; Bowyer, P; Tjäderhane, L; Rautemaa, R

    2014-04-01

    The amino acid derivative 2-hydroxyisocaproic acid (HICA) is a nutritional additive used to increase muscle mass. Low levels can be detected in human plasma as a result of leucine metabolism. It has broad antibacterial activity but its efficacy against pathogenic fungi is not known. The aim was to test the efficacy of HICA against Candida and Aspergillus species. Efficacy of HICA against 19 clinical and reference isolates representing five Candida and three Aspergillus species with variable azole antifungal sensitivity profiles was tested using a microdilution method. The concentrations were 18, 36 and 72 mg ml(-1) . Growth was determined spectrophotometrically for Candida isolates and by visual inspection for Aspergillus isolates, viability was tested by culture and impact on morphology by microscopy. HICA of 72 mg ml(-1) was fungicidal against all Candida and Aspergillus fumigatus and Aspergillus terreus isolates. Lower concentrations were fungistatic. Aspergillus flavus was not inhibited by HICA. HICA inhibited hyphal formation in susceptible Candida albicans and A. fumigatus isolates and affected cell wall integrity. In conclusion, HICA has broad antifungal activity against Candida and Aspergillus at concentrations relevant for topical therapy. As a fungicidal agent with broad-spectrum bactericidal activity, it may be useful in the topical treatment of multispecies superficial infections.

  12. GH11 xylanase from Emericella nidulans with low sensitivity to inhibition by ethanol and lignocellulose-derived phenolic compounds.

    PubMed

    Silva, Caio de Oliveira Gorgulho; Aquino, Elaine Nascimento; Ricart, Carlos André Ornelas; Midorikawa, Gláucia Emy Okida; Miller, Robert Neil Gerard; Filho, Edivaldo Ximenes Ferreira

    2015-07-01

    An endo-β-1,4-xylanase (X22) was purified from crude extract of Emericella nidulans when cultivated on submerged fermentation using sugarcane bagasse as the carbon source. The purified protein was identified by mass spectrometry and was most active at pH and temperature intervals of 5.0-6.5 and 50-60°C, respectively. The enzyme showed half-lives of 40, 10 and 7 min at 28, 50 and 55°C, respectively, and pH 5.0. Apparent Km and Vmax values on soluble oat spelt xylan were 3.39 mg/mL and 230.8 IU/mg, respectively, while Kcat and Kcat/Km were 84.6 s(-1) and 25.0 s(-1) mg(-1) mL. Incubation with phenolic compounds showed that tannic acid and cinnamic acid had an inhibitory effect on X22 but no time-dependent deactivation. On the other hand, ferulic acid, 4-hydroxybenzoic acid, vanillin and p-coumaric acid did not show any inhibitory effect on X22 activity, although they changed X22 apparent kinetic parameters. Ethanol remarkably increased enzyme thermostability and apparent Vmax and Kcat values, even though the affinity and catalytic efficiency for xylan were lowered.

  13. Changes in intensity and spectral distribution of fluorescence. Effect of light treatment on normal and DCMU-poisoned Anacystis nidulans.

    PubMed

    Papageorgiou, G; Govindjee

    1967-07-01

    The intensity of the "steady-state" fluorescence of "aerobic" Anacystis nidulans is variable under prolonged illumination with orange (590 mmu) or blue (440 mmu) light for both normally photosynthesizing and DCMU-poisoned cells. In general, orange light illumination causes an increase of the fluorescence intensity followed by a decrease, while blue light causes an increase until a steady level is reached. Poisoned Anacystis cells show four to eight times larger changes in fluorescence intensity than the normal cells; the detailed time course of fluorescence changes is also different in poisoned and normal cells. When algae are cooled to -196 degrees C in light, the light-induced changes in the "steady-state" fluorescence disappear in both types of cells. Difference fluorescence spectra, constructed by subtracting the fluorescence spectra taken after 5-15 min of illumination from those after 60-90 min of illumination, show a doublet structure of the difference band with a major peak coinciding with the Anacystis emission maximum (685 mmu) and a minor peak located at about 693 mmu.

  14. Aspergillus flavus contamination in two Portuguese wastewater treatment plants.

    PubMed

    Viegas, C; Dias, R; Gomes, A Quintal; Meneses, M; Sabino, R; Viegas, S

    2014-01-01

    Filamentous fungi from genus Aspergillus were previously detected in wastewater treatment plants (WWTP) as being Aspergillus flavus (A. flavus), an important toxigenic fungus producing aflatoxins. This study aimed to determine occupational exposure adverse effects due to fungal contamination produced by A. flavus complex in two Portuguese WWTP using conventional and molecular methodologies. Air samples from two WWTP were collected at 1 m height through impaction method. Surface samples were collected by swabbing surfaces of the same indoor sites. After counting A. flavus and identification, detection of aflatoxin production was ensured through inoculation of seven inoculates in coconut-milk agar. Plates were examined under long-wave ultraviolet (UV; 365 nm) illumination to search for the presence of fluorescence in the growing colonies. To apply molecular methods, air samples were also collected using the impinger method. Samples were collected and collection liquid was subsequently used for DNA extraction. Molecular identification of A. flavus was achieved by real-time polymerase chain reaction (RT-PCR) using the Rotor-Gene 6000 qPCR detection system (Corbett). Among the Aspergillus genus, the species that were more abundant in air samples from both WWTP were Aspergillus versicolor (38%), Aspergillus candidus (29.1%), and Aspergillus sydowii (12.7%). However, the most commonly species found on surfaces were A. flavus (47.3%), Aspergillus fumigatus (34.4%), and Aspergillus sydowii (10.8%). Aspergillus flavus isolates that were inoculated in coconut agar medium were not identified as toxigenic strains and were not detected by RT-PCR in any of the analyzed samples from both plants. Data in this study indicate the need for monitoring fungal contamination in this setting. Although toxigenic strains were not detected from A. flavus complex, one cannot disregard the eventual presence and potential toxicity of aflatoxins. PMID:25072712

  15. Aspergillus surveillance project at a large tertiary-care hospital.

    PubMed

    Curtis, L; Cali, S; Conroy, L; Baker, K; Ou, C-H; Hershow, R; Norlock-Cruz, F; Scheff, P

    2005-03-01

    A one-year surveillance project was conducted at a large tertiary hospital, which had extensive indoor renovation and extensive demolition/building at several nearby sites. This study collected viable fungi samples in the hospital every six days and analysed 74 duct dust samples for Aspergillus fumigatus mycelial asp f1 protein. Mean total fungi were 257.8 cfu/m3 outdoors, 53.2 cfu/m3 in all indoor samples and 83.5 cfu/m3 in the bone marrow transplant patient rooms. Mean total aspergillus was 6.8 cfu/m3 outdoors, 12.1 cfu/m3 in all indoor samples and 7.3 cfu/m3 in the bone marrow transplant patient rooms. The five most prevalent Aspergillus species collected inside the hospital (mean cfu/m3) were Aspergillus niger 7.57 cfu/m3, Aspergillus candidus 1.72 cfu/m3, Aspergillus flavus 0.97 cfu/m3, A. fumigatus 0.88 cfu/m3 and Aspergillus glaucus 0.45 cfu/m3. In rooms undergoing duct cleaning, mean A. fumigatus concentrations were 11.0 cfu/m3. Forty-eight of 74 (65%) duct dust samples had measurable levels of asp f1 protein, with a mean level of 0.41 ppm and maximum level of 1.94 ppm. Three major incidents involved increased hospital aspergillus concentrations. A. niger levels reached 680 cfu/m3 in an organ transplant room after a water leak from a ceiling pipe. Total aspergillus concentrations rose to 77 cfu/m3 in a bone marrow transplant patient room after improper sealing and water infiltration of the unit's dedicated high-efficiency particulate air filter system. Total aspergillus levels of 160 cfu/m3 were recorded in a renovation area during wood cutting. The higher concentrations of aspergillus seen inside the hospital compared with outdoors and the various moisture/HEPA filter/renovation incidents suggest that numerous small to moderate sources of aspergillus exist in the hospital. PMID:15694975

  16. Antifungal susceptibility profile of cryptic species of Aspergillus.

    PubMed

    Alastruey-Izquierdo, Ana; Alcazar-Fuoli, Laura; Cuenca-Estrella, Manuel

    2014-12-01

    The use of molecular tools has led to the description of new cryptic species among different Aspergillus species complexes. Their frequency in the clinical setting has been reported to be between 10 and 15%. The susceptibility to azoles and amphotericin B of many of these species is low, and some of them, such as Aspergillus calidoustus or Aspergillus lentulus, are considered multi-resistant. The changing epidemiology, the frequency of cryptic species, and the different susceptibility profiles make antifungal susceptibility testing an important tool to identify the optimal antifungal agent to treat the infections caused by these species.

  17. Role of growth media and chemical enhancers in secondary metabolites production from Aspergillus carbonarius (NRL-369) and their pharmaceutical potentials.

    PubMed

    Khan, Abid Ali; Bacha, Nafess; Ahmad, Bashir; Cox, R J; Bakht, Jehan

    2016-07-01

    The present study investigates the effect of different growth media and chemical enhancer on silent genes in Aspergillus carbonarius (NRL-369) for secondary metabolites production and its in vitro biological activities. Results revealed that Aspergillus carbonarius (NRL-369) grown in Czapeak yeast extract broth medium produced more metabolites compared with other media. Chemical epigenetic modifiers (suberoyl-anilide hydroxamic acid (SAHA) and 5-azacytidine (5-AZA) at concentration of 15mM were effective for the expression of silent genes resulting in increased secondary metabolites production. Secondary metabolites extracted in ethyl acetate and fractionized in n-Hexane showed variable degree of growth inhibitions of the tested microorganisms. Similarly, these samples were also active against brine shrimps and Lemna. PMID:27393435

  18. Effect of gamma radiation on Aspergillus flavus and Aspergillus ochraceus ultrastructure and mycotoxin production

    NASA Astrophysics Data System (ADS)

    Ribeiro, J.; Cavaglieri, L.; Vital, H.; Cristofolini, A.; Merkis, C.; Astoreca, A.; Orlando, J.; Carú, M.; Dalcero, A.; Rosa, C. A. R.

    2011-05-01

    The aim of this work was to study the effect of gamma radiation (2 kGy) on Aspergillus flavus and Aspergillus ochraceus ultrastructure. Moreover, the influence on aflatoxin B 1 and ochratoxin A production was also observed. Irradiated A. flavus strain showed a dull orangish colony while control strain showed the typical green color. Minor differences were observed on stipes, metulae and conidia size between control and irradiated A. flavus and A. ochraceus strains. Irradiated fungi showed ultrastructural changes on cell wall, plasmalema and cytoplasm levels. The levels of mycotoxins produced by irradiated strains were two times greater than those produced by control strains. Successive transferences of irradiated strains on malt extract agar allowed the fungus to recuperate morphological characteristics. Although minor changes in the fungal morphology were observed, ultrastructural changes at cell wall level and the increase of mycotoxin production ability were observed. Inappropriate storage of irradiated food and feed would allow the development of potentially more toxicogenic fungal propagules.

  19. Ochratoxigenic Aspergillus species on grapes from Chilean vineyards and Aspergillus threshold levels on grapes.

    PubMed

    Díaz, Gonzalo A; Torres, René; Vega, Mario; Latorre, Bernardo A

    2009-07-31

    This study reports the incidence of ochratoxigenic strains of Aspergillus on Chilean grapes (Vitis vinifera) and wineries, and production of OTA levels in wines with grapes having different levels of contamination with OTA-producing Aspergillus carbonarius was studied. A. carbonarius, A. niger, A. niveus, A. paradoxus, A. versicolor, A. wentii, and A. westerdijkiae were identified on apparently healthy clusters of red and white grape cultivars. However, A. carbonarius and A. niger were the most frequently identified species, more abundant on red than white grape cultivars. Aspergillus spp. populations increased between veraison and harvest, but the isolation frequencies were relatively low over the entire growing season. At the winery, A. carbonarius, A. niger and A. westerdijkiae were occasionally found in the air, exclusively during winemaking. OTA-producing strains were only found among isolates of A. carbonarius, A. niger, A. wenti, and A. westerdijkiae, producing 2 to 17 microg/L of OTA in liquid medium; however, A. westerdijkiae produced the highest OTA concentration in vitro. Red wines elaborated with 0.5% of grapes infected with an OTA-producing strain of A. carbonarius (Aspuc-SB36) exceeded the 2 microg/L of OTA tolerance established for wines by the European Community. Therefore, a threshold below 0.5% infected berries is proposed for red wines. ELISA tests proved to be useful for detecting OTA in broth culture as in wine samples.

  20. Aspergillus pragensis sp. nov. discovered during molecular reidentification of clinical isolates belonging to Aspergillus section Candidi.

    PubMed

    Hubka, Vit; Lyskova, Pavlina; Frisvad, Jens C; Peterson, Stephen W; Skorepova, Magdalena; Kolarik, Miroslav

    2014-08-01

    The identity of nine clinical isolates recovered from Czech patients and presumptively identified as Aspergillus sp. section Candidi based on colony morphology was revised using sequences of β-tubulin, calmodulin gene sequence, and internal transcribed spacer rDNA. Six isolates were from suspected and proven onychomycosis, one from otitis externa, and two associated with probable invasive aspergillosis. The results showed that one Aspergillus candidus isolate was the cause of otitis externa, and both isolates obtained from sputa of patients with probable invasive aspergillosis were reidentified as A. carneus (sect. Terrei) and A. flavus (sect. Flavi). Three isolates from nail scrapings were identified as A. tritici, a verified agent of nondermatophyte onychomycosis. One isolate from toenail was determined to be A. candidus and the two isolates belonged to a hitherto undescribed species, Aspergillus pragensis sp. nov. This species is well supported by phylogenetic analysis based on β-tubulin and calmodulin gene and is distinguishable from other members of sect. Candidi by red-brown reverse on malt extract agar, slow growth on Czapek-Dox agar and inability to grow at 37°C. A secondary metabolite analysis was also provided with comparison of metabolite spectrum to other species. Section Candidi now encompasses five species for which a dichotomous key based on colony characteristics is provided. All clinical isolates were tested for susceptibilities to selected antifungal agents using the Etest and disc diffusion method. Overall sect. Candidi members are highly susceptible to common antifungals.

  1. A novel fungal fruiting structure formed by Aspergillus niger and Aspergillus carbonarius in grape berries.

    PubMed

    Pisani, Cristina; Nguyen, Trang Thoaivan; Gubler, Walter Douglas

    2015-09-01

    Sour rot, is a pre-harvest disease that affects many grape varieties. Sour rot symptoms include initial berry cracking and breakdown of berry tissue. This is a disease complex with many filamentous fungi and bacteria involved, but is usually initiated by Aspergillus niger or Aspergillus carbonarius. Usually, by the time one sees the rot there are many other organisms involved and it is difficult to attribute the disease to one species. In this study two species of Aspergillus were shown to produce a previously unknown fruiting structure in infected berries. The nodulous morphology, bearing conidia, suggests them to be an 'everted polymorphic stroma'. This structure forms freely inside the berry pulp and assumes multiple shapes and sizes, sometimes sclerotium-like in form. It is composed of a mass of vegetative hyphae with or without tissue of the host containing spores or fruiting bodies bearing spores. Artificially inoculated berries placed in soil in winter showed the possible overwintering function of the fruiting body. Inoculated berry clusters on standing vines produced fruiting structures within 21 d post inoculation when wounds were made at veraison or after (July-September). Histological studies confirmed that the fruiting structure was indeed fungal tissue.

  2. A novel fungal fruiting structure formed by Aspergillus niger and Aspergillus carbonarius in grape berries.

    PubMed

    Pisani, Cristina; Nguyen, Trang Thoaivan; Gubler, Walter Douglas

    2015-09-01

    Sour rot, is a pre-harvest disease that affects many grape varieties. Sour rot symptoms include initial berry cracking and breakdown of berry tissue. This is a disease complex with many filamentous fungi and bacteria involved, but is usually initiated by Aspergillus niger or Aspergillus carbonarius. Usually, by the time one sees the rot there are many other organisms involved and it is difficult to attribute the disease to one species. In this study two species of Aspergillus were shown to produce a previously unknown fruiting structure in infected berries. The nodulous morphology, bearing conidia, suggests them to be an 'everted polymorphic stroma'. This structure forms freely inside the berry pulp and assumes multiple shapes and sizes, sometimes sclerotium-like in form. It is composed of a mass of vegetative hyphae with or without tissue of the host containing spores or fruiting bodies bearing spores. Artificially inoculated berries placed in soil in winter showed the possible overwintering function of the fruiting body. Inoculated berry clusters on standing vines produced fruiting structures within 21 d post inoculation when wounds were made at veraison or after (July-September). Histological studies confirmed that the fruiting structure was indeed fungal tissue. PMID:26321727

  3. Extrolites of Aspergillus fumigatus and Other Pathogenic Species in Aspergillus Section Fumigati

    PubMed Central

    Frisvad, Jens C.; Larsen, Thomas O.

    2016-01-01

    Aspergillus fumigatus is an important opportunistic human pathogen known for its production of a large array of extrolites. Up to 63 species have been described in Aspergillus section Fumigati, some of which have also been reliably reported to be pathogenic, including A. felis, A. fischeri, A. fumigatiaffinis, A. fumisynnematus, A. hiratsukae, A. laciniosus, A. lentulus, A. novofumigatus, A. parafelis, A. pseudofelis, A. pseudoviridinutans, A. spinosus, A. thermomutatus, and A. udagawae. These species share the production of hydrophobins, melanins, and siderophores and ability to grow well at 37°C, but they only share some small molecule extrolites, that could be important factors in pathogenicity. According to the literature gliotoxin and other exometabolites can be contributing factors to pathogenicity, but these exometabolites are apparently not produced by all pathogenic species. It is our hypothesis that species unable to produce some of these metabolites can produce proxy-exometabolites that may serve the same function. We tabulate all exometabolites reported from species in Aspergillus section Fumigati and by comparing the profile of those extrolites, suggest that those producing many different kinds of exometabolites are potential opportunistic pathogens. The exometabolite data also suggest that the profile of exometabolites are highly specific and can be used for identification of these closely related species. PMID:26779142

  4. The Aspergillus Genome Database, a curated comparative genomics resource for gene, protein and sequence information for the Aspergillus research community.

    PubMed

    Arnaud, Martha B; Chibucos, Marcus C; Costanzo, Maria C; Crabtree, Jonathan; Inglis, Diane O; Lotia, Adil; Orvis, Joshua; Shah, Prachi; Skrzypek, Marek S; Binkley, Gail; Miyasato, Stuart R; Wortman, Jennifer R; Sherlock, Gavin

    2010-01-01

    The Aspergillus Genome Database (AspGD) is an online genomics resource for researchers studying the genetics and molecular biology of the Aspergilli. AspGD combines high-quality manual curation of the experimental scientific literature examining the genetics and molecular biology of Aspergilli, cutting-edge comparative genomics approaches to iteratively refine and improve structural gene annotations across multiple Aspergillus species, and web-based research tools for accessing and exploring the data. All of these data are freely available at http://www.aspgd.org. We welcome feedback from users and the research community at aspergillus-curator@genome.stanford.edu.

  5. Primary role of the cytoplasmic membrane in thermal acclimation evidenced in nitrate-starved cells of the blue-green alga, Anacystis nidulans

    SciTech Connect

    Gombos, Z.; Vigh, L.

    1986-02-01

    The lipid phase transition of the cytoplasmic membrane and the chilling susceptibility were studied in nitrate-starved Anacystis nidulans cells. Nitrate starvation resulted in the disappearance of the thylakoid membrane system, without any effect on chilling susceptibility. The chilling susceptibility of the algal cells depended on the growth temperature. Temperatures of lipid phase transitions of the cytoplasmic membranes were detected by chilling-induced spectral changes in the carotenoid region, in vivo. These values were identical to those of cultures containing intact thylakoid systems. Our results suggest that cytoplasmic membrane plays a determinative role in the thermal acclimation of the alga cell.

  6. A tyrosinase inhibitor from Aspergillus niger.

    PubMed

    Vasantha, K Y; Murugesh, C S; Sattur, A P

    2014-10-01

    Tyrosinase, in the presence of oxygen, is the main culprit in post harvest browning of food products, resulting in the drop in its commercial value. In an effort to seek natural tyrosinase inhibitors for food applications, a screening programme was undertaken. Of the 26 fungal cultures isolated from soil samples of Agumbe forest, India, one isolate S16, identified as Aspergillus niger, gave an inhibition of 84 % against the enzyme. The inhibitor was isolated by following an enzyme inhibition assay guided purification protocol. The structure of the inhibitor was elucidated and found to be kojic acid. The IC50 of the Competitive inhibitor was found to be 8.8 μg with a Ki of 0.085 mM.

  7. Toxic Metabolite Produced by Aspergillus wentii

    PubMed Central

    Wu, M. T.; Ayres, J. C.; Koehler, P. E.; Chassis, G.

    1974-01-01

    Mycelial extracts of an Aspergillus wentii strain grown on yeast-extract sucrose medium and initially isolated from country-cured ham were highly toxic when inoculated into chicken embryos or fed to mice. Moldy corn and rice were less toxic when fed to mice. Water extracts of moldy corn or rice or culture filtrates from yeast-extract sucrose medium were not toxic. Purification by thin-layer chromatography followed by crystallization yielded orange-red crystals that showed high toxicity and had a melting point of 285 to 286 C. Chloroform solutions of the crystals had absorption maxima at 270, 295, and 452 nm. The smallest amount of this component necessary to have zero hatchability of fertile eggs was 50 μg/egg. PMID:4823420

  8. Environmental fungicides and triazole resistance in Aspergillus.

    PubMed

    Bowyer, Paul; Denning, David W

    2014-02-01

    Fungal diseases are problematic in both human health and agriculture. Treatment options are limited and resistance may emerge. The relatively recent recognition of triazole resistance in Aspergillus fumigatus has prompted questioning of the origin of resistance. While multiple mechanisms are described in clinical isolates from triazole-treated patients, some de novo resistance is also recognised, especially attributable to TR34 /L98H. Such strains probably arose in the environment, and, indeed, multiple studies have now demonstrated TR(34) /L98H triazole resistance strains of A. fumigatus from soil. Docking and other in vitro studies are consistent with environmental resistance induction through exposure to certain triazole fungicides, notably difenoconazole, propiconazole, epoxiconazole, bromuconazole and tebuconazole. This article addresses the potential implications of this issue for both human health and food security.

  9. Production of cyclopiazonic acid by Aspergillus tamarii Kita.

    PubMed Central

    Dorner, J W

    1983-01-01

    Production of the mycotoxin cyclopiazonic acid by Aspergillus tamarii Kita is reported for the first time. Examination of 23 isolates of the fungus showed that 22 produced the toxin under the culture conditions utilized. PMID:6660879

  10. Nutrient environment influences competition among Aspergillus flavus genotypes

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Structures of Aspergillus flavus populations, shaped by intraspecific competition, influence the incidences and severities of crop aflatoxin contamination. Competition for nutrients may be one factor modulating intraspecific interactions, but influences of specific types and concentrations of nutrie...

  11. Septic arthritis due to tubercular and Aspergillus co-infection.

    PubMed

    Kumar, Mukesh; Thilak, Jai; Zahoor, Adnan; Jyothi, Arun

    2016-01-01

    Aspergillus septic arthritis is a rare and serious medical and surgical problem. It occurs mainly in immunocompromised patients. Aspergillus fumigatus is the most common causative organism followed by Aspergillus flavus. The most common site affected is knee followed by shoulder, ankle, wrist, hip and sacroiliac joint. Debridement and voriconazole are primary treatment of articular aspergilosis. To the best of our knowledge, there are no reported cases of co-infection of tuberculosis (TB) and Aspergillus infecting joints. We report a case of co-infection of TB and A. flavus of hip and knee of a 60-year-old male, with type 2 diabetes mellitus. He was treated with debridement, intravenous voriconazole, and antitubercular drugs.

  12. Emerging infections caused by non-Aspergillus filamentous fungi.

    PubMed

    Douglas, A P; Chen, S C-A; Slavin, M A

    2016-08-01

    There are three broad groups of non-Aspergillus moulds: the mucormycetes, the hyalohyphomycetes and the phaeohyphomycetes. Infections with these pathogens are increasingly reported, particularly in the context of increasing use of immunosuppressant agents and improved diagnostics. The epidemiology of non-Aspergillus mould infections varies with geography, climate and level of immunosuppression. Skin and soft-tissue infections are the predominant presentation in the immunocompetent host and pulmonary and other invasive infections in the immunocompromised host. The more common non-Aspergillus moulds include Rhizopus, Mucor, Fusarium and Scedosporium species; however, other emerging pathogens are Rasamsonia and Verruconis species, which are discussed in this article. Outbreaks of non-Aspergillus mould infections have been increasingly reported, with contaminated medical supplies and natural disasters as common sources. Currently culture and other conventional diagnostic methods are the cornerstone of diagnosis. Molecular methods to directly detect and identify mould pathogens in tissue and body fluids are increasingly used. PMID:26812445

  13. Aspergillus Thyroiditis after Allogeneic Hematopoietic Stem Cell Transplantation

    PubMed Central

    Ataca, Pinar; Atilla, Erden; Saracoglu, Pelin; Yilmaz, Gulden; Civriz Bozdag, Sinem; Toprak, Selami Kocak; Yuksel, Meltem Kurt; Ceyhan, Koray; Topcuoglu, Pervin

    2015-01-01

    Aspergillus thyroiditis is a rare disorder detected in immunocompromised patients during disseminated infections. Early management is essential to prevent high mortality. A 61-year-old allogeneic stem cell male recipient presented with painful thyroid nodular enlargement. He had low TSH and low free T4 levels. The thyroid ultrasound showed a hypoechoic nodule; biopsy indicated suppurative Aspergillus thyroiditis. He was successfully treated by amphotericin B. PMID:26640727

  14. Functional analysis of FarA transcription factor in the regulation of the genes encoding lipolytic enzymes and hydrophobic surface binding protein for the degradation of biodegradable plastics in Aspergillus oryzae.

    PubMed

    Garrido, Sharon Marie; Kitamoto, Noriyuki; Watanabe, Akira; Shintani, Takahiro; Gomi, Katsuya

    2012-05-01

    FarA is a Zn(II)(2)Cys(6) transcription factor which upregulates genes required for growth on fatty acids in filamentous fungi like Aspergillus nidulans. FarA is also highly similar to the cutinase transcription factor CTF1α of Fusarium solani which binds to the cutinase gene promoter in this plant pathogen. This study determines whether FarA transcriptional factor also works in the regulation of genes responsible for the production of cutinase for the degradation of a biodegradable plastic, poly-(butylene succinate-co-adipate) (PBSA), in Aspergillus oryzae. The wild-type and the farA gene disruption strains were grown in minimal agar medium with emulsified PBSA, and the wild-type showed clear zone around the colonies while the disruptants did not. Western blot analysis revealed that the cutinase protein CutL1 and a hydrophobic surface binding protein such as HsbA were produced by the wild-type but not by the disruptants. In addition, the expressions of cutL1, triacylglycerol lipase (tglA), and mono- and di-acylglycerol lipase (mdlB) genes as well as the hsbA gene were significantly lower in the disruptants compared to the wild-type. These results indicated that the FarA transcriptional factor would be implicated in the expression of cutL1 and hsbA genes that are required for the degradation of PBSA as well as lipolytic genes such as mdlB and tglA for lipid hydrolysis.

  15. Modified cyanobacteria

    DOEpatents

    Vermaas, Willem F J.

    2014-06-17

    Disclosed is a modified photoautotrophic bacterium comprising genes of interest that are modified in terms of their expression and/or coding region sequence, wherein modification of the genes of interest increases production of a desired product in the bacterium relative to the amount of the desired product production in a photoautotrophic bacterium that is not modified with respect to the genes of interest.

  16. [Identification of Target Extracellular Proteases--Activators of Proteins of Haemostasis System Produced by Micromycetes Aspergillus ochraceus and Aspergillus terreus].

    PubMed

    Zvonareva, E S; Osmolovskiy, A A; Kreyer, V G; Baranova, N A; Kotova, I B; Egorov, N S

    2015-01-01

    Effects of extracellular proteases of Aspergillus ochraceus and Aspergillus terreus on plasma hemostasis proteins, consist of initiating the activation of prothrombin complex proteins, was detected. Was discovered, that A. ochraceus proteases have a direct influence on protein C and coagulation factor X, and A. terreus proteases causes their activation indirectly through kallikrein system stimulation. The ability of extracellular proteases of micromycetes activate prekallikrein in human blood plasma on the example of A. terreus was first demonstrated.

  17. Validation of a new Aspergillus real-time PCR assay for direct detection of Aspergillus and azole resistance of Aspergillus fumigatus on bronchoalveolar lavage fluid.

    PubMed

    Chong, Ga-Lai M; van de Sande, Wendy W J; Dingemans, Gijs J H; Gaajetaan, Giel R; Vonk, Alieke G; Hayette, Marie-Pierre; van Tegelen, Dennis W E; Simons, Guus F M; Rijnders, Bart J A

    2015-03-01

    Azole resistance in Aspergillus fumigatus is increasingly reported. Here, we describe the validation of the AsperGenius, a new multiplex real-time PCR assay consisting of two multiplex real-time PCRs, one that identifies the clinically relevant Aspergillus species, and one that detects the TR34, L98H, T289A, and Y121F mutations in CYP51A and differentiates susceptible from resistant A. fumigatus strains. The diagnostic performance of the AsperGenius assay was tested on 37 bronchoalveolar lavage (BAL) fluid samples from hematology patients and 40 BAL fluid samples from intensive care unit (ICU) patients using a BAL fluid galactomannan level of ≥1.0 or positive culture as the gold standard for detecting the presence of Aspergillus. In the hematology and ICU groups combined, there were 22 BAL fluid samples from patients with invasive aspergillosis (IA) (2 proven, 9 probable, and 11 nonclassifiable). Nineteen of the 22 BAL fluid samples were positive, according to the gold standard. The optimal cycle threshold value for the presence of Aspergillus was <36. Sixteen of the 19 BAL fluid samples had a positive PCR (2 Aspergillus species and 14 A. fumigatus samples). This resulted in a sensitivity, specificity, and positive and negative predictive values of 88.9%, 89.3%, 72.7%, and 96.2%, respectively, for the hematology group and 80.0%, 93.3%, 80.0%, and 93.3%, respectively, in the ICU group. The CYP51A real-time PCR confirmed 12 wild-type and 2 resistant strains (1 TR34-L98H and 1 TR46-Y121F-T289A mutant). Voriconazole therapy failed for both patients. The AsperGenius multiplex real-time PCR assay allows for sensitive and fast detection of Aspergillus species directly from BAL fluid samples. More importantly, this assay detects and differentiates wild-type from resistant strains, even if BAL fluid cultures remain negative.

  18. New and revisited species in Aspergillus section Nigri

    PubMed Central

    Varga, J.; Frisvad, J.C.; Kocsubé, S.; Brankovics, B.; Tóth, B.; Szigeti, G.; Samson, R.A.

    2011-01-01

    Four new species, Aspergillus eucalypticola, A. neoniger, A. fijiensis and A. indologenus are described and illustrated. Aspergillus eucalypticola was isolated from Eucalyptus leaf from Australia, and is related to A. tubingensis and A. costaricaensis, but could clearly be distinguished from them based on either β-tubulin or calmodulin sequence data. Aspergillus eucalypticola produced pyranonigrin A, funalenone, aurasperone B and other naphtho-γ-pyrones. Aspergillus neoniger is also a biseriate species isolated from desert sand in Namibia, and mangrove water in Venezuela, which produces aurasperone B and pyranonigrin A. Aspergillus fijiensis is a uniseriate species related to A. aculeatinus, and was isolated from soil in Fiji, and from Lactuca sativa in Indonesia. This species is able to grow at 37 °C, and produces asperparalines and okaramins. Aspergillus indologenus was isolated from soil, India. This species also belongs to the uniseriate group of black aspergilli, and was found to be related to, but clearly distinguishable from A. uvarum based on β-tubulin, calmodulin and ITS sequence data. Aspergillus indologenus produced the insecticidal compounds okaramins A, B, H, and two types of indol-alkaloids which have not been structure elucidated. Two other species, A. violaceofuscus and A. acidus, are revalidated based on molecular and extrolite data. Aspergillus violaceofuscus was found to be related to A. japonicus, and produced some of the same interesting indol-alkaloids as A. indologenus, and also produced several families of partially characterised extrolites that were also found in A. heteromorphus. Aspergillus acidus (previously known as A. foetidus var. pallidus and A. foetidus var. acidus) is also a valid species, while A. foetidus is a synonym of A. niger based on molecular and physiological data. Two other species described previously, A. coreanus and A. lacticoffeatus, were found to be colour mutants of A. acidus and A. niger, respectively. Methods

  19. Does fungicide application in vineyards induce resistance to medical azoles in Aspergillus species?

    PubMed

    Lago, Magali; Aguiar, Ana; Natário, André; Fernandes, Carla; Faria, Miguel; Pinto, Eugénia

    2014-09-01

    This study assessed if the use of sterol demethylase inhibitor fungicides in vineyard production can induce resistance to azoles in Aspergillus strains and if it can induce selection of resistant species. We also tried to identify the Aspergillus species most prevalent in the vineyards. Two vineyards from northern Portugal were selected from "Vinhos Verdes" and "Douro" regions. The vineyards were divided into plots that were treated or not with penconazole (PEN). In each vineyard, air, soil, and plant samples were collected at three different times. The strains of Aspergillus spp. were isolated and identified by morphological and molecular techniques. We identified 46 Aspergillus section Nigri, eight Aspergillus fumigatus, seven Aspergillus lentulus, four Aspergillus wentii, two Aspergillus flavus, two Aspergillus terreus, one Aspergillus calidoustus, one Aspergillus westerdijkiae, one Aspergillus tamarii, and one Eurotium amstelodami. Aspergillus strains were evaluated for their susceptibility to medical azoles used in human therapy (itraconazole, posaconazole, and voriconazole) and to agricultural azoles (PEN) used in the prevention and treatment of plant diseases. The isolates showed moderate susceptibility to voriconazole. We did not observe any decrease of susceptibility to the medical azoles tested throughout the testing period in any of the treated plots, although some of the resistant species were isolated from there.

  20. KdmA, a histone H3 demethylase with bipartite function, differentially regulates primary and secondary metabolism in A spergillus nidulans

    PubMed Central

    Gacek‐Matthews, Agnieszka; Noble, Luke M.; Gruber, Clemens; Berger, Harald; Sulyok, Michael; Marcos, Ana T.

    2015-01-01

    Summary A spergillus nidulans kdmA encodes a member of the KDM4 family of jumonji histone demethylase proteins, highly similar to metazoan orthologues both within functional domains and in domain architecture. This family of proteins exhibits demethylase activity towards lysines 9 and 36 of histone H3 and plays a prominent role in gene expression and chromosome structure in many species. Mass spectrometry mapping of A . nidulans histones revealed that around 3% of bulk histone H3 carried trimethylated H3K9 (H3K9me3) but more than 90% of histones carried either H3K36me2 or H3K36me3. KdmA functions as H3K36me3 demethylase and has roles in transcriptional regulation. Genetic manipulation of KdmA levels is tolerated without obvious effect in most conditions, but strong phenotypes are evident under various conditions of stress. Transcriptome analysis revealed that – in submerged early and late cultures – between 25% and 30% of the genome is under KdmA influence respectively. Transcriptional imbalance in the kdm A deletion mutant may contribute to the lethal phenotype observed upon exposure of mutant cells to low‐density visible light on solid medium. Although KdmA acts as transcriptional co‐repressor of primary metabolism genes, it is required for full expression of several genes involved in biosynthesis of secondary metabolites. PMID:25712266

  1. Mutational analysis of the Aspergillus ambient pH receptor PalH underscores its potential as a target for antifungal compounds

    PubMed Central

    Lucena‐Agell, Daniel; Hervás‐Aguilar, América; Múnera‐Huertas, Tatiana; Pougovkina, Olga; Rudnicka, Joanna; Galindo, Antonio; Tilburn, Joan; Arst, Herbert N.

    2016-01-01

    Summary The pal/RIM ambient pH signalling pathway is crucial for the ability of pathogenic fungi to infect hosts. The Aspergillus nidulans 7‐TMD receptor PalH senses alkaline pH, subsequently facilitating ubiquitination of the arrestin PalF. Ubiquitinated PalF triggers downstream signalling events. The mechanism(s) by which PalH transduces the alkaline pH signal to PalF is poorly understood. We show that PalH is phosphorylated in a signal dependent manner, resembling mammalian GPCRs, although PalH phosphorylation, in contrast to mammalian GPCRs, is arrestin dependent. A genetic screen revealed that an ambient‐exposed region comprising the extracellular loop connecting TM4‐TM5 and ambient‐proximal residues within TM5 is required for signalling. In contrast, substitution by alanines of four aromatic residues within TM6 and TM7 results in a weak ‘constitutive’ activation of the pathway. Our data support the hypothesis that PalH mechanistically resembles mammalian GPCRs that signal via arrestins, such that the relative positions of individual helices within the heptahelical bundle determines the Pro316‐dependent transition between inactive and active PalH conformations, governed by an ambient‐exposed region including critical Tyr259 that potentially represents an agonist binding site. These findings open the possibility of screening for agonist compounds stabilizing the inactive conformation of PalH, which might act as antifungal drugs against ascomycetes. PMID:27279148

  2. Enhanced diversity and aflatoxigenicity in interspecific hybrids of Aspergillus flavus and Aspergillus parasiticus.

    PubMed

    Olarte, Rodrigo A; Worthington, Carolyn J; Horn, Bruce W; Moore, Geromy G; Singh, Rakhi; Monacell, James T; Dorner, Joe W; Stone, Eric A; Xie, De-Yu; Carbone, Ignazio

    2015-04-01

    Aspergillus flavus and A. parasiticus are the two most important aflatoxin-producing fungi responsible for the contamination of agricultural commodities worldwide. Both species are heterothallic and undergo sexual reproduction in laboratory crosses. Here we examine the possibility of interspecific matings between A. flavus and A. parasiticus. These species can be distinguished morphologically and genetically, as well as by their mycotoxin profiles. Aspergillus flavus produces both B aflatoxins and cyclopiazonic acid (CPA), B aflatoxins or CPA alone, or neither mycotoxin; Aspergillus parasiticus produces B and G aflatoxins or the aflatoxin precursor O-methylsterigmatocystin, but not CPA. Only four of forty-five attempted interspecific crosses between opposite mating types of A. flavus and A. parasiticus were fertile and produced viable ascospores. Single ascospore strains from each cross were shown to be recombinant hybrids using multilocus genotyping and array comparative genome hybridization. Conidia of parents and their hybrid progeny were haploid and predominantly monokaryons and dikaryons based on flow cytometry. Multilocus phylogenetic inference showed that experimental hybrid progeny were grouped with naturally occurring A. flavus L strain and A. parasiticus. Higher total aflatoxin concentrations in some F1 progeny strains compared to midpoint parent aflatoxin levels indicate synergism in aflatoxin production; moreover, three progeny strains synthesized G aflatoxins that were not produced by the parents, and there was evidence of allopolyploidization in one strain. These results suggest that hybridization is an important diversifying force resulting in the genesis of novel toxin profiles in these agriculturally important fungi.

  3. Biotransformation of the mycotoxin zearalenone by fungi of the genera Rhizopus and Aspergillus.

    PubMed

    Brodehl, Antje; Möller, Anne; Kunte, Hans-Jörg; Koch, Matthias; Maul, Ronald

    2014-10-01

    Zearalenone (ZEN) is a nonsteroidal estrogenic mycotoxin biosynthesized by various Fusarium fungi. These fungal species frequently infest grains; therefore, ZEN represents a common contaminant in cereal products. The biotransformation of ZEN differs significantly from species to species, and several metabolites are known to be formed by animals, plants, and microorganisms. The aim of the present study was to investigate the microbial conversion of ZEN by species of the genera Rhizopus and Aspergillus representing relevant fungi for food processing (e.g. fermentation). To monitor the ZEN metabolism, ZEN was added to liquid cultures of the different fungal species. After a period of 3 days, the media were analyzed by HPLC-MS/MS for metabolite formation. Two Aspergillus oryzae strains and all seven Rhizopus species were able to convert ZEN into various metabolites, including ZEN-14-sulfate as well as ZEN-O-14- and ZEN-O-16-glucoside. Microbial transformation of ZEN into the significantly more estrogenic α-zearalenol (α-ZEL) was also observed. Additionally, a novel fungal metabolite, α-ZEL-sulfate, was detected. Semi-quantification of the main metabolites indicates that more than 50% of initial ZEN may be modified. The results show that fungal strains have the potential to convert ZEN into various metabolites leading to a masking of the toxin, for example in fermented food. PMID:25145804

  4. Biotransformation of the mycotoxin zearalenone by fungi of the genera Rhizopus and Aspergillus.

    PubMed

    Brodehl, Antje; Möller, Anne; Kunte, Hans-Jörg; Koch, Matthias; Maul, Ronald

    2014-10-01

    Zearalenone (ZEN) is a nonsteroidal estrogenic mycotoxin biosynthesized by various Fusarium fungi. These fungal species frequently infest grains; therefore, ZEN represents a common contaminant in cereal products. The biotransformation of ZEN differs significantly from species to species, and several metabolites are known to be formed by animals, plants, and microorganisms. The aim of the present study was to investigate the microbial conversion of ZEN by species of the genera Rhizopus and Aspergillus representing relevant fungi for food processing (e.g. fermentation). To monitor the ZEN metabolism, ZEN was added to liquid cultures of the different fungal species. After a period of 3 days, the media were analyzed by HPLC-MS/MS for metabolite formation. Two Aspergillus oryzae strains and all seven Rhizopus species were able to convert ZEN into various metabolites, including ZEN-14-sulfate as well as ZEN-O-14- and ZEN-O-16-glucoside. Microbial transformation of ZEN into the significantly more estrogenic α-zearalenol (α-ZEL) was also observed. Additionally, a novel fungal metabolite, α-ZEL-sulfate, was detected. Semi-quantification of the main metabolites indicates that more than 50% of initial ZEN may be modified. The results show that fungal strains have the potential to convert ZEN into various metabolites leading to a masking of the toxin, for example in fermented food.

  5. Isolation of pathogenic Aspergillus species from commercially-prepared potting media.

    PubMed

    Kenyon, E M; Russell, L H; McMurray, D N

    1984-09-30

    Twelve commercially-prepared potting soils were screened for the presence of pathogenic Aspergillus species. Pathogenic Aspergillus species were isolated from 67% of the soils. A fumigatus was isolated from 42% and A. flavus and A. niger from 33%.

  6. Detection and discrimination of four Aspergillus section Nigri species by PCR.

    PubMed

    Palumbo, J D; O'Keeffe, T L

    2015-02-01

    Species of Aspergillus section Nigri are not easily distinguished by traditional morphological techniques, and typically are identified by DNA sequencing methods. We developed four PCR primers to distinguish between Aspergillus niger, Aspergillus welwitschiae, Aspergillus carbonarius and Aspergillus tubingensis, based on species-conserved differences in the calmodulin gene sequence. PCR amplification from total DNA using these primers was species specific; no amplification occurred from nontarget species DNA for each primer pair. Species-specific PCR could distinguish between species in mixed DNA templates, indicating a utility in determining culture uniformity of isolated Aspergillus strains. In addition, with these primer sets, each species could be detected in soil following mixed-species inoculation with Aspergillus spores. This indicates that PCR with these species-specific primers may be useful in determining the distribution of Aspergillus species in environmental samples without the need for species identification from isolated strains, as well as detecting species that may be infrequently isolated by culture-based methods.

  7. What is the importance of classifying Aspergillus disease in cystic fibrosis patients?

    PubMed

    Jones, Andrew M; Horsley, Alex; Denning, David W

    2014-08-01

    Aspergillus species are commonly isolated from lower respiratory tract samples of patients with cystic fibrosis (CF) and markers of immunological sensation to Aspergillus are frequently encountered in this group of patients; however, the contribution of Aspergillus to CF lung disease outside of the typical complications of ABPA and aspergilloma formation remains largely unclear. Patients with CF show discretely different responses to Aspergillus, though the underlying reasons for this variation are unknown. Recent work has begun to allow us to categorize patient responses to Aspergillus based upon molecular markers of infection and immune sensitization. Aspergillus sensitization and/or airway infection is associated with worse FEV1, in CF and other patients (asthma, chronic obstructive pulmonary disease, bronchiectasis). Classification of different clinical phenotypes of Aspergillus will enable future studies to determine the natural history of different manifestations of Aspergillus disease and evaluate the effects of intervention with antifungal therapy.

  8. Shedding light on Aspergillus niger volatile exometabolome

    PubMed Central

    Costa, Carina Pedrosa; Gonçalves Silva, Diogo; Rudnitskaya, Alisa; Almeida, Adelaide; Rocha, Sílvia M.

    2016-01-01

    An in-depth exploration of the headspace content of Aspergillus niger cultures was performed upon different growth conditions, using a methodology based on advanced multidimensional gas chromatography. This volatile fraction comprises 428 putatively identified compounds distributed over several chemical families, being the major ones hydrocarbons, alcohols, esters, ketones and aldehydes. These metabolites may be related with different metabolic pathways, such as amino acid metabolism, biosynthesis and metabolism of fatty acids, degradation of aromatic compounds, mono and sesquiterpenoid synthesis and carotenoid cleavage. The A. niger molecular biomarkers pattern was established, comprising the 44 metabolites present in all studied conditions. This pattern was successfully used to distinguish A. niger from other fungi (Candida albicans and Penicillium chrysogenum) with 3 days of growth by using Partial Least Squares-Discriminant Analysis (PLS-DA). In addition, PLS-DA-Variable Importance in Projection was applied to highlight the metabolites playing major roles in fungi distinction; decreasing the initial dataset to only 16 metabolites. The data pre-processing time was substantially reduced, and an improvement of quality-of-fit value was achieved. This study goes a step further on A. niger metabolome construction and A. niger future detection may be proposed based on this molecular biomarkers pattern. PMID:27264696

  9. Characterization of an exopolygalacturonase from Aspergillus niger.

    PubMed

    Fahmy, Afaf S; El-Beih, Fawkia M; Mohamed, Saleh A; Abdel-Gany, Somia S; Abd-Elbaky, Engy A

    2008-06-01

    Polygalacturonase (PGI) from Aspergillus niger NRRL3 was purified about 12.0-fold from the cell-free broth using diethylaminoethyl-Sepharose and Sephacryl S-200 columns. The molecular weight of the PGI was 32,000 Da as estimated by gel filtration and sodium dodecyl sulfate-polyacrylamide gel electrophoresis. PGI had an isoelectric point of 7.6 and an optimum pH of 5.0. PGI was active on polygalacturonic acid and esterified pectins, but the activity on pectin decreased with an increase in degree of esterification. PGI had higher affinity (low Km) and turnover number (Vmax/Km and Kcat/Km) toward polygalacturonic acid. PGI was found to have a temperature optimum at 40 degrees C and was approximately stable up to 30 degrees C. All the examined metal cations had partial inhibitory effects on PGI, while Mn+2 at 5 mM caused a complete inhibition for the enzyme. Comparison of viscosity reduction rates with release of reducing sugars indicated that the enzyme from A. niger is exoacting. The storage stability study of PGI showed that the enzyme in powder form retained 56% of its activity after 9 months of storage at 4 degrees C. The above properties of PGI may be suitable for food processing. PMID:18500582

  10. Aspergillus Biofilm In Vitro and In Vivo.

    PubMed

    Beauvais, Anne; Latgé, Jean-Paul

    2015-08-01

    In vivo, Aspergillus fumigatus grows as a typical biofilm with hyphae covered by an extracellular matrix (ECM) composed of polysaccharides, galactomannan, and galactosaminogalactan. α1,3 glucans and melanin are also constitutive of the ECM in aspergilloma but not in invasive aspergillosis. In vitro, two biofilm models were established to mimic the in vivo situation. The first model (model 1) uses submerged liquid conditions and is characterized by slow growth, while the second model (model 2) uses agar medium and aerial conditions and is characterized by rapid growth. The composition of the ECM was studied only in the second model and has been shown to be composed of galactomannan, galactosaminogalactan (GAG), and α1,3 glucans, melanin, antigens, and hydrophobins. The presence of extracellular DNA was detected in model 1 biofilm but not in model 2. Transcriptomic analysis employing both biofilm models showed upregulation of genes coding for proteins involved in the biosynthesis of secondary metabolites, adhesion, and drug resistance. However, most data on A. fumigatus biofilms have been obtained in vitro and should be confirmed using in vivo animal models. There is a need for new therapeutic antibiofilm strategies that focus on the use of combination therapy, since biofilm formation poses an important clinical problem due to their resistance to antifungal agents. Furthermore, in vivo investigations of A. fumigatus biofilms that incorporate the associated microbiota are needed. Such studies will add another layer of complexity to our understanding of the role of A. fumigatus biofilm during lung invasion. PMID:26350307

  11. Emergence of Azole Resistance in Aspergillus.

    PubMed

    Wiederhold, Nathan P; Patterson, Thomas F

    2015-10-01

    Resistance to the azole antifungals itraconazole, voriconazole, and posaconazole in Aspergillus species is a growing concern. This is especially alarming for A. fumigatus, where acquired resistance has been documented in patients with invasive disease caused by this species that were exposed to these agents, as well as in azole-naive individuals. The primary mechanisms of resistance that have been described in clinical strains include different point mutations in the CYP51A gene, which encodes the enzyme responsible for converting lanosterol to ergosterol via demethylation. Some resistant isolates also contain a tandem repeat in the promoter region of this gene that causes increased expression. These mutations, including TR34/L98H and TR46/Y121F/T289A have also been found in the environment in several areas of the world and have been demonstrated to cause resistance to azole fungicides used in agriculture, thus raising the concern of environmental spread of resistance. Treatment options are limited in patients with infections caused by azole-resistant isolates and include amphotericin B formulations or combination therapy involving an echinocandin. However, there are few clinical data available to help guide therapy, and infections caused by resistant A. fumigatus isolates have been reported to have high mortality rates. PMID:26398534

  12. Bioconversion of Capsaicin by Aspergillus oryzae.

    PubMed

    Lee, Minji; Cho, Jeong-Yong; Lee, Yu Geon; Lee, Hyoung Jae; Lim, Seong-Il; Park, So-Lim; Moon, Jae-Hak

    2015-07-01

    This study identified metabolites of capsaicin bioconverted by Aspergillus oryzae, which is generally used for mass production of gochujang prepared by fermenting red pepper powder in Korea. A. oryzae was incubated with capsaicin in potato dextrose broth. Capsaicin decreased depending on the incubation period, but new metabolites increased. Five capsaicin metabolites purified from the ethyl acetate fraction of the capsaicin culture were identified as N-vanillylcarbamoylbutyric acid, N-vanillyl-9-hydroxy-8-methyloctanamide, ω-hydroxycapsaicin, 8-methyl-N-vanillylcarbamoyl-6(E)-octenoic acid, and 2-methyl-N-vanillylcarbamoyl-6(Z)-octenoic acid by nuclear magnetic resonance (NMR) and mass spectrometry (MS). The capsaicin metabolites in gochujang were confirmed and quantitated by selective multiple reaction monitoring detection after liquid chromatography electrospray ionization MS using the isolated compounds as external standards. On the basis of the structures of the capsaicin metabolites, it is proposed that capsaicin metabolites were converted by A. oryzae by ω-hydroxylation, alcohol oxidation, hydrogenation, isomerization, and α- and/or β-oxidation.

  13. Fumonisin B2 production by Aspergillus niger.

    PubMed

    Frisvad, Jens C; Smedsgaard, Jørn; Samson, Robert A; Larsen, Thomas O; Thrane, Ulf

    2007-11-14

    The carcinogenic mycotoxin fumonisin B2 was detected for the first time in the industrially important Aspergillus niger. Fumonisin B2, known from Fusarium verticillioides and other Fusaria, was detected in cultures of three full genome sequenced strains of A. niger, in the ex type culture and in a culture of F. verticillioides by electrospray LC-MS analysis of methanolic extracts from agar plugs of cultures grown on several substrates. Whereas F. verticillioides produced fumonisins B1, B2, and B3 on agar media based on plant extracts, such as barley malt, oat, rice, potatoes, and carrots, A. niger produced fumonisin B2 best on agar media with a low water activity, including Czapek yeast autolysate agar with 5% NaCl. Of the media tested, only rice corn steep agar supported fumonisin production by both F. verticillioides and A. niger. However, A. niger had a different regulation of fumonisin production and a different quantitative profile of fumonisins, producing only B2 as compared to F. verticillioides. Fumonisin production by A. niger, which is a widely occurring species and an extremely important industrial organism, will have very important implications for biotechnology and especially food safety. A. niger is used for the production of citric acid and as producer of extracellular enzymes, and also as a transformation host for the expression of heterologous proteins. Certain strains of A. niger produce both ochratoxin A and fumonisins, so some foods and feeds may potentially contain two types of carcinogenic mycotoxins from this species.

  14. Infection of the pleura by Aspergillus fumigatus

    PubMed Central

    Krakówka, Paweł; Rowińska, Ewa; Halweg, Halina

    1970-01-01

    Pleural aspergillosis occurs mostly in established cases of pleural empyema with a broncho-pleural fistula. Ten such patients are reported here: in all, Aspergillus fumigatus infection was related to tuberculosis. In three cases with an active, sputum-positive tuberculous process the pleural empyema was a complication of spontaneous pneumothorax in two, and of lung resection in one. In two cases the empyema occurred as a complication of tuberculous pleuritis, but A. fumigatus infection was noted only after the sputum had become negative for tubercle bacilli. In five patients with inactive tuberculosis, the empyema was a late complication of pneumothorax therapy. The diagnosis of pleural aspergillosis is made on the basis of microscopical examination and culture of A. fumigatus in the pleural pus. The cultures were positive in seven of the 10 cases presented. In two cases in which the culture was negative microscopical examination of the pus revealed the presence of numerous fungal hyphae which was evidence of fungal necrosis. In one case the diagnosis was not made until necropsy. Serum precipitin tests with filtrates of A. fumigatus are further valuable evidence of aspergillous infection. Of 10 presented patients, this test was positive in all seven cases in which it was done. The treatment of pleural aspergillosis by local instillation of nystatin or amphotericin B was effective in six out of seven cases in which it was used. Images PMID:5441996

  15. Degradation of melanin by Aspergillus fumigatus.

    PubMed Central

    Luther, J P; Lipke, H

    1980-01-01

    A strain of Aspergillus fumigatus from composted coffee and garden wastes utilized natural deproteinized insect, banana, hair, octopus, and synthetic tyrosine and dopa melanins as sole sources of carbon. With a sucrose supplement, degradation was essentially complete after 50 days in Czapek medium pH 6.5 at 30 degrees C. The catabolic rate differed for each substrate pigment, as did the molecular weight distribution of products accumulating in the medium. After incubation with L-[U-14C]melanin, over 50% was recovered in a dark fungal pigment, the remainder appearing as cell protein, chitin, lipid, CO2, and polar metabolites. When grown on melanin, the normally pale mycelia darkened with the production of a fungal allomelanin, with infrared spectrum and alkali fusion products differing from those of the substrate pigment. Isotope distribution in amino acids for A. fumigatus grown on labeled melanin supplemented with sucrose suggested separate pools for synthesis of cell proteins and melanoproteins. Deposition of allomelanin increased resistance of conidia, sterigma, and conidiophores to lytic carbohydrases as judged by scanning electron microscopy. Images PMID:6996615

  16. A new diketopiperazine alkaloid from Aspergillus oryzae.

    PubMed

    Shaaban, Mohamed; El-Metwally, Mohammad Magdy; Nasr, Hamdi

    2014-01-01

    Investigation of bioactive secondary metabolites from terrestrial Aspergillus oryzae sp. MMAO1 using M2 medium afforded a new diketopiperazine alkaloid, 7,9-dihydroxy-3-(1H-indol-3-ylmethyl)-8-methoxy-2,3,11,11a-tetrahydro-6H-pyrazino[1,2-b]isoquinoline-1,4-dione (1a), containing the unusual amino acid L-6,8-dihydroxy-7-methoxyphenylalanine. This was co-isolated with ditryptophenaline (2), cyclo-(Tryp,Tyr) (4), cyclo-(Pro,Val), α-cyclopiazonic acid (3), kojic acid and uridine. Re-cultivation of the fungal strain on Dox medium led to the production of bisdethio(bismethylthio)gliotoxin (5), pseurotin A (6) along with linoleic acid, α-cyclopiazonic acid (3) and kojic acid. The chemical structure of the new diketopiperazine alkaloid including the relative configuration was determined by 1D and 2D NMR spectroscopy and HR-ESI-MS spectrometry, and by comparison with the related literature. The new alkaloid (1a) showed no antimicrobial activity or cytotoxicity against brine shrimps.

  17. Comparison of glucose oxidases from Penicillium adametzii, Penicillium Funiculosum and Aspergillus Niger in the design of amperometric glucose biosensors.

    PubMed

    Ramanavicius, Arunas; Voronovic, Jaroslav; Semashko, Tatiana; Mikhailova, Raisa; Kausaite-Minkstimiene, Asta; Ramanaviciene, Almira

    2014-01-01

    The properties of amperometric glucose biosensors based on three different glucose oxidases and various redox mediators were evaluated. Glucose oxidases (GOx) from Penicillium adametzii, Penicillium funiculosum and Aspergillus niger and artificial redox mediators, such as ferrocene, ferrocenecarboxaldehyde, α-methylferrocene methanol and ferrocenecarboxylic acid, were used for modifying the graphite rod electrode and amperometrical reagent-less glucose detection. The obtained results were compared using N-methylphenazonium methyl sulphate in the solution. Taking into account the experimental kinetic parameters and the stability of the tested enzymatic electrodes, GOx from Penicillium funiculosum proved to be more suitable for glucose biosensor design in comparison with other evaluated enzymes. PMID:25492463

  18. 40 CFR 180.1206 - Aspergillus flavus AF36; exemption from the requirement of a tolerance.

    Code of Federal Regulations, 2010 CFR

    2010-07-01

    ... 40 Protection of Environment 23 2010-07-01 2010-07-01 false Aspergillus flavus AF36; exemption... FOOD Exemptions From Tolerances § 180.1206 Aspergillus flavus AF36; exemption from the requirement of a... pesticide Aspergillus flavus AF36 in or on cotton, gin byproducts; cotton, hulls; cotton, meal;...

  19. Flash kinetics and light intensity dependence of oxygen evolution in the blue-green alga Anacystis nidulans.

    PubMed

    Ley, A C; Babcock, G T; Sauer, K

    1975-05-15

    Patterns of oxygen evolution in flashing light for the glue-green alga Anacystis nidulans are compared with those for broken spinach chloroplasts and whole cells of the green alga Chlorella pyrenoidosa. The oscillations of oxygen yield with flash number that occur in both Anacystis and Chlorella, display a greater degree of damping than do those of isolated spinach chloroplasts. The increase in damping results from a two- to threefold increase in the fraction (alpha) of reaction centers "missed" by a flash. The increase in alpha cannot be explained by non-saturing flash intensities or by the dark reduction of the oxidized intermediates formed by the flash. Anaerobic conditions markedly increase alpha in Anacystis and Chlorella but have no effect on alpha in broken spinach chloroplasts. The results signify that the mechanism of charge separation and water oxidation involved in all three orgainsms is the same, but that the pool of secondary electron acceptors between Photosystem II and Photosystem I is more reduced in the dark, in the algal cells, than in the isolated spinach chloroplasts. Oxygen evolution in flashing light for Anacystis and Chlorella show light saturation curves for the oxygen yield of the third flash (Y3) that differ markedly from those of the steady-state flashes(YS). In experiments in which all flashes are uniformly attenuated, Y3 requires nearly twice as much light as YS to reach half-saturation. Under these conditions Y3 has a sigmoidal dependence on intensity, while that of YS is hyperbolic. These differences depend on the number of flashes attenuated. When any one of the first three flashes is attenuated, the variation of Y3 with intensity resembles that of YS. When two of the first three flashes are attenuated, Y3 is intermediate in shape between the two extremes. A quantitative interpretation of these results based on the model of Kok et al. (Kik, B., Forbush, B.and McGloin, M. (1970) Photochem. Photobiol. 14, 307-321) fits the experimental

  20. Effect of Microgravity on Fungistatic Activity of an α-Aminophosphonate Chitosan Derivative against Aspergillus niger

    PubMed Central

    Lee, Yang Soo; Kim, Byoung-Suhk

    2015-01-01

    Biocontamination within the international space station is ever increasing mainly due to human activity. Control of microorganisms such as fungi and bacteria are important to maintain the well-being of the astronauts during long-term stay in space since the immune functions of astronauts are compromised under microgravity. For the first time control of the growth of an opportunistic pathogen, Aspergillus niger, under microgravity is studied in the presence of α-aminophosphonate chitosan. A low-shear modelled microgravity was used to mimic the conditions similar to space. The results indicated that the α-aminophosphonate chitosan inhibited the fungal growth significantly under microgravity. In addition, the inhibition mechanism of the modified chitosan was studied by UV-Visible spectroscopy and cyclic voltammetry. This work highlighted the role of a bio-based chitosan derivative to act as a disinfectant in space stations to remove fungal contaminants. PMID:26468641

  1. Effect of Microgravity on Fungistatic Activity of an α-Aminophosphonate Chitosan Derivative against Aspergillus niger.

    PubMed

    Devarayan, Kesavan; Sathishkumar, Yesupatham; Lee, Yang Soo; Kim, Byoung-Suhk

    2015-01-01

    Biocontamination within the international space station is ever increasing mainly due to human activity. Control of microorganisms such as fungi and bacteria are important to maintain the well-being of the astronauts during long-term stay in space since the immune functions of astronauts are compromised under microgravity. For the first time control of the growth of an opportunistic pathogen, Aspergillus niger, under microgravity is studied in the presence of α-aminophosphonate chitosan. A low-shear modelled microgravity was used to mimic the conditions similar to space. The results indicated that the α-aminophosphonate chitosan inhibited the fungal growth significantly under microgravity. In addition, the inhibition mechanism of the modified chitosan was studied by UV-Visible spectroscopy and cyclic voltammetry. This work highlighted the role of a bio-based chitosan derivative to act as a disinfectant in space stations to remove fungal contaminants. PMID:26468641

  2. Effect of Microgravity on Fungistatic Activity of an α-Aminophosphonate Chitosan Derivative against Aspergillus niger.

    PubMed

    Devarayan, Kesavan; Sathishkumar, Yesupatham; Lee, Yang Soo; Kim, Byoung-Suhk

    2015-01-01

    Biocontamination within the international space station is ever increasing mainly due to human activity. Control of microorganisms such as fungi and bacteria are important to maintain the well-being of the astronauts during long-term stay in space since the immune functions of astronauts are compromised under microgravity. For the first time control of the growth of an opportunistic pathogen, Aspergillus niger, under microgravity is studied in the presence of α-aminophosphonate chitosan. A low-shear modelled microgravity was used to mimic the conditions similar to space. The results indicated that the α-aminophosphonate chitosan inhibited the fungal growth significantly under microgravity. In addition, the inhibition mechanism of the modified chitosan was studied by UV-Visible spectroscopy and cyclic voltammetry. This work highlighted the role of a bio-based chitosan derivative to act as a disinfectant in space stations to remove fungal contaminants.

  3. Expression and export: recombinant protein production systems for Aspergillus.

    PubMed

    Fleissner, André; Dersch, Petra

    2010-07-01

    Several Aspergillus species, in particular Aspergillus niger and Aspergillus oryzae, are widely used as protein production hosts in various biotechnological applications. In order to improve the expression and secretion of recombinant proteins in these filamentous fungi, several novel genetic engineering strategies have been developed in recent years. This review describes state-of-the-art genetic manipulation technologies used for strain improvement, as well as recent advances in designing the most appropriate engineering strategy for a particular protein production process. Furthermore, current developments in identifying bottlenecks in the protein production and secretion pathways are described and novel approaches to overcome these limitations are introduced. An appropriate combination of expression vectors and optimized host strains will provide cell factories customized for each production process and expand the great potential of Aspergilli as biotechnology workhorses to more complex multi-step industrial applications.

  4. Heterologous expression of Aspergillus terreus fructosyltransferase in Kluyveromyces lactis.

    PubMed

    Spohner, Sebastian C; Czermak, Peter

    2016-06-25

    Fructo-oligosaccharides are prebiotic and hypocaloric sweeteners that are usually extracted from chicory. They can also be produced from sucrose using fructosyltransferases, but the only commercial enzyme suitable for this purpose is Pectinex Ultra, which is produced with Aspergillus aculeatus. Here we used the yeast Kluyveromyces lactis to express a secreted recombinant fructosyltransferase from the inulin-producing fungus Aspergillus terreus. A synthetic codon-optimised version of the putative β-fructofuranosidase ATEG 04996 (XP 001214174.1) from A. terreus NIH2624 was secreted as a functional protein into the extracellular medium. At 60°C, the purified A. terreus enzyme generated the same pattern of oligosaccharides as Pectinex Ultra, but at lower temperatures it also produced oligomers with up to seven units. We achieved activities of up to 986.4U/mL in high-level expression experiments, which is better than previous reports of optimised Aspergillus spp. fermentations. PMID:27084521

  5. Toxigenic Aspergillus and Penicillium Isolates from Weevil-Damaged Chestnuts

    PubMed Central

    Wells, John M.; Payne, Jerry A.

    1975-01-01

    Aspergillus and Penicillium were among the most common genera of fungi isolated on malt-salt agar from weevil-damaged Chinese chestnut kernels (16.8 and 40.7% occurrence, respectively). Chloroform extracts of 21 of 50 Aspergillus isolates and 18 of 50 representative Penicillium isolates, grown for 4 weeks at 21.1 C on artificial medium, were toxic to day-old cockerels. Twelve of the toxic Aspergillus isolates were identified as A. wentii, eight as A. flavus, and one as A. flavus var. columnaris. Nine of the toxic Penicillium isolates were identified as P. terrestre, three as P. steckii, two each as P. citrinum and P. funiculosum, and one each as P. herquei (Series) and P. roqueforti (Series). Acute diarrhea was associated with the toxicity of A. wentii and muscular tremors with the toxicity of P. terrestre, one isolate of P. steckii, and one of P. funiculosum. PMID:1190758

  6. Biosorption potency of Aspergillus niger for removal of chromium (VI).

    PubMed

    Srivastava, Shaili; Thakur, Indu Shekhar

    2006-09-01

    Aspergillus niger isolated from soil and effluent of leather tanning mills had higher activity to remove chromium. The potency of Aspergillus niger was evaluated in shake flask culture by absorption of chromium at pH 6 and temperature 30 degrees C. The results of the study indicated removal of more than 75% chromium by Aspergillus niger determined by diphenylcarbazide colorimetric assay and atomic absorption spectrophotometry after 7 days. Study of microbial Cr(VI) reduction and identification of reduction intermediates has been hindered by the lack of analytical techniques that can identify the oxidation state with subcellular spatial resolution. Therefore, removal of chromium was further substantiated by transmission electron microscopy (TEM), scanning electron microscopy (SEM), and energy-dispersive X-ray spectroscopy (EDX), which indicated an accumulation of chromium in the fungal mycelium. PMID:16874547

  7. Aspergillus Mediastinitis after Orthotopic Heart Transplantation: A Case Report.

    PubMed

    El-Sayed Ahmed, Magdy M; Almanfi, Abdelkader; Aftab, Muhammad; Singh, Steve K; Mallidi, Hari R; Frazier, O H

    2015-10-01

    A 55-year-old woman was admitted for orthotopic heart transplantation. Her medical history was notable for multiple cardiovascular problems, including ischemic cardiomyopathy that necessitated circulatory support with a left ventricular assist device. Five weeks after undergoing orthotopic heart transplantation, she developed Aspergillus calidoustus mediastinitis, for which she underwent a prolonged course of antifungal treatment that comprised (in sequence) posaconazole for 11 days, voriconazole for 10 days, and amphotericin B for 42 days. During this period, she also underwent repeated mediastinal drainage and sternal débridement, followed by sternal wiring and coverage with bilateral pectoralis advancement flaps. Four months postoperatively, she was discharged from the hospital with a successfully controlled infection and a healed sternum. To our knowledge, only 3 previous cases of Aspergillus mediastinitis after orthotopic heart transplantation have been reported in the literature, none of which was Aspergillus calidoustus.

  8. Biodiversity of Aspergillus species in some important agricultural products

    PubMed Central

    Perrone, G.; Susca, A.; Cozzi, G.; Ehrlich, K.; Varga, J.; Frisvad, J.C.; Meijer, M.; Noonim, P.; Mahakarnchanakul, W.; Samson, R.A.

    2007-01-01

    The genus Aspergillus is one of the most important filamentous fungal genera. Aspergillus species are used in the fermentation industry, but they are also responsible of various plant and food secondary rot, with the consequence of possible accumulation of mycotoxins. The aflatoxin producing A. flavus and A. parasiticus, and ochratoxinogenic A. niger, A. ochraceus and A. carbonarius species are frequently encountered in agricultural products. Studies on the biodiversity of toxigenic Aspergillus species is useful to clarify molecular, ecological and biochemical characteristics of the different species in relation to their different adaptation to environmental and geographical conditions, and to their potential toxigenicity. Here we analyzed the biodiversity of ochratoxin producing species occurring on two important crops: grapes and coffee, and the genetic diversity of A. flavus populations occurring in agricultural fields. Altogether nine different black Aspergillus species can be found on grapes which are often difficult to identify with classical methods. The polyphasic approach used in our studies led to the identification of three new species occurring on grapes: A. brasiliensis, A. ibericus, and A. uvarum. Similar studies on the Aspergillus species occurring on coffee beans have evidenced in the last five years that A. carbonarius is an important source of ochratoxin A in coffee. Four new species within the black aspergilli were also identified in coffee beans: A. sclerotioniger, A. lacticoffeatus, A. sclerotiicarbonarius, and A. aculeatinus. The genetic diversity within A. flavus populations has been widely studied in relation to their potential aflatoxigenicity and morphological variants L- and S-strains. Within A. flavus and other Aspergillus species capable of aflatoxin production, considerable diversity is found. We summarise the main recent achievements in the diversity of the aflatoxin gene cluster in A. flavus populations, A. parasiticus and the non

  9. Electrochemical monitoring of citric acid production by Aspergillus niger.

    PubMed

    Kutyła-Olesiuk, Anna; Wawrzyniak, Urszula E; Ciosek, Patrycja; Wróblewski, Wojciech

    2014-05-01

    Hybrid electronic tongue was developed for the monitoring of citric acid production by Aspergillus niger. The system based on various potentiometric/voltammetric sensors and appropriate chemometric techniques provided correct qualitative and quantitative classification of the samples collected during standard Aspergillus niger culture and culture infected with yeast. The performance of the proposed approach was compared with the monitoring of the fermentation process carried out using classical methods. The results obtained proved, that the designed hybrid electronic tongue was able to evaluate the progress and correctness of the fermentation process.

  10. Modern taxonomy of biotechnologically important Aspergillus and Penicillium species.

    PubMed

    Houbraken, Jos; de Vries, Ronald P; Samson, Robert A

    2014-01-01

    Taxonomy is a dynamic discipline and name changes of fungi with biotechnological, industrial, or medical importance are often difficult to understand for researchers in the applied field. Species belonging to the genera Aspergillus and Penicillium are commonly used or isolated, and inadequate taxonomy or uncertain nomenclature of these genera can therefore lead to tremendous confusion. Misidentification of strains used in biotechnology can be traced back to (1) recent changes in nomenclature, (2) new taxonomic insights, including description of new species, and/or (3) incorrect identifications. Changes in the recent published International Code of Nomenclature for Algae, Fungi and Plants will lead to numerous name changes of existing Aspergillus and Penicillium species and an overview of the current names of biotechnological important species is given. Furthermore, in (biotechnological) literature old and invalid names are still used, such as Aspergillus awamori, A. foetidus, A. kawachii, Talaromyces emersonii, Acremonium cellulolyticus, and Penicillium funiculosum. An overview of these and other species with their correct names is presented. Furthermore, the biotechnologically important species Talaromyces thermophilus is here combined in Thermomyces as Th. dupontii. The importance of Aspergillus, Penicillium, and related genera is also illustrated by the high number of undertaken genome sequencing projects. A number of these strains are incorrectly identified or atypical strains are selected for these projects. Recommendations for correct strain selection are given here. Phylogenetic analysis shows a close relationship between the genome-sequenced strains of Aspergillus, Penicillium, and Monascus. Talaromyces stipitatus and T. marneffei (syn. Penicillium marneffei) are closely related to Thermomyces lanuginosus and Th. dupontii (syn. Talaromyces thermophilus), and these species appear to be distantly related to Aspergillus and Penicillium. In the last part of

  11. Polyphasic taxonomy of Aspergillus section Fumigati and its teleomorph Neosartorya

    PubMed Central

    Samson, R.A.; Hong, S.; Peterson, S.W.; Frisvad, J.C.; Varga, J.

    2007-01-01

    The taxonomy of Aspergillus section Fumigati with its teleomorph genus Neosartorya is revised. The species concept is based on phenotypic (morphology and extrolite profiles) and molecular (β-tubulin and calmodulin gene sequences) characters in a polyphasic approach. Four new taxa are proposed: N. australensis N. ferenczii, N. papuaensis and N. warcupii. All newly described and accepted species are illustrated. The section consists of 33 taxa: 10 strictly anamorphic Aspergillus species and 23 Neosartorya species. Four other Neosartorya species described previously were not available for this monograph, and consequently are relegated to the category of doubtful species. PMID:18490953

  12. Malic acid production from thin stillage by Aspergillus species.

    PubMed

    West, Thomas P

    2011-12-01

    The ability of Aspergillus strains to utilize thin stillage to produce malic acid was compared. The highest malic acid was produced by Aspergillus niger ATCC 9142 at 17 g l(-1). Biomass production from thin stillage was similar with all strains but ATCC 10577 was the highest at 19 g l(-1). The highest malic acid yield (0.8 g g(-1)) was with A. niger ATCC 9142 and ATCC 10577 on the stillage. Thus, thin stillage has the potential to act as a substrate for the commercial production of food-grade malic acid by the A. niger strains.

  13. A study on Aspergillus species in houses of asthmatic patients from Sari City, Iran and a brief review of the health effects of exposure to indoor Aspergillus.

    PubMed

    Hedayati, Mohammad T; Mayahi, Sabah; Denning, David W

    2010-09-01

    To study the distribution of Aspergillus spp. in outdoor and indoor air of asthmatic patients' houses, as well as a review on the health effects of exposure to indoor Aspergillus. Open plates containing malt extract agar media were used to isolate fungi from the indoor (n = 360) and outdoor (n = 180) air of 90 asthmatic patients' houses living in Sari City, Iran. Plates were incubated at room temperature for 7-14 days. Cultured Aspergillus spp. were identified by standard mycological techniques. All culture plates grew fungi, a testament to the ubiquitous nature of fungal exposure. Cladosporium spp. (29.2%), Aspergillus spp. (19.0%), and Penicillium spp. (18.3%) were most common inside the houses while Cladosporium spp. (44.5%), Aspergillus spp. (12.4%), and Alternaria spp. (11.1%) were most common outside the houses. Aspergillus flavus (30.1%) and A. fumigatus (23.1%) are the most commonly isolated species in indoor air. Aspergillus flavus (44.5%) and A. fumigatus (42.6%) were the most prevalent Aspergillus spp. outside. The most colony numbers of Aspergillus were isolated from kitchens (30.4%) and the least from bedrooms (21.1%). Aspergillus flavus was the most prevalent species in all sampled rooms except in the kitchen where A. fumigatus was the most common. Aspergillus flavus is the most prevalent species among the Aspergillus spp. in the indoor and outdoor of a warm climate area. In these areas, A. flavus can be a major source of allergen in the air. Therefore, minimizing indoor fungal exposure could play an important role in reducing allergic symptoms in susceptible persons. PMID:19697147

  14. Aqueous extracts of Tulbaghia violacea inhibit germination of Aspergillus flavus and Aspergillus parasiticus conidia.

    PubMed

    Somai, Benesh Munilal; Belewa, Vuyokazi

    2011-06-01

    Aspergillus flavus and Aspergillus parasiticus are important plant pathogens and causal agents of pre- and postharvest rots of corn, peanuts, and tree nuts. These fungal pathogens cause significant crop losses and produce aflatoxins, which contaminate many food products and contribute to liver cancer worldwide. Aqueous preparations of Tulbaghia violacea (wild garlic) were antifungal and at 10 mg/ml resulted in sustained growth inhibition of greater than 50% for both A. flavus and A. parasiticus. Light microscopy revealed that the plant extract inhibited conidial germination in a dose-dependent manner. When exposed to T. violacea extract concentrations of 10 mg/ml and above, A. parasiticus conidia began germinating earlier and germination was completed before that of A. flavus, indicating that A. parasiticus conidia were more resistant to the antifungal effects of T. violacea than were A. flavus conidia. At a subinhibitory extract dose of 15 mg/ml, hyphae of both fungal species exhibited increased granulation and vesicle formation, possibly due to increased reactivity between hyphal cellular components and T. violacea extract. These hyphal changes were not seen when hyphae were formed in the absence of the extract. Transmission electron microscopy revealed thickening of conidial cell walls in both fungal species when grown in the presence of the plant extract. Cell walls of A. flavus also became considerably thicker than those of A. parasiticus, indicating differential response to the extract. Aqueous preparations of T. violacea can be used as antifungal treatments for the control of A. flavus and A. parasiticus. Because the extract exhibited a more pronounced effect on A. flavus than on A. parasiticus, higher doses may be needed for control of A. parasiticus infections. PMID:21669082

  15. Refining the pH response in A spergillus nidulans: a modulatory triad involving PacX, a novel zinc binuclear cluster protein

    PubMed Central

    Bussink, Henk‐Jan; Bignell, Elaine M.; Múnera‐Huertas, Tatiana; Lucena‐Agell, Daniel; Scazzocchio, Claudio; Espeso, Eduardo A.; Bertuzzi, Margherita; Rudnicka, Joanna; Negrete‐Urtasun, Susana; Peñas‐Parilla, Maria M.; Rainbow, Lynne; Peñalva, Miguel Á.; Arst, Herbert N.

    2015-01-01

    Summary The A spergillus nidulans PacC transcription factor mediates gene regulation in response to alkaline ambient pH which, signalled by the Pal pathway, results in the processing of PacC72 to PacC27 via PacC53. Here we investigate two levels at which the pH regulatory system is transcriptionally moderated by pH and identify and characterise a new component of the pH regulatory machinery, PacX. Transcript level analysis and overexpression studies demonstrate that repression of acid‐expressed pal F, specifying the Pal pathway arrestin, probably by PacC27 and/or PacC53, prevents an escalating alkaline pH response. Transcript analyses using a reporter and constitutively expressed pac C  trans‐alleles show that pac C preferential alkaline‐expression results from derepression by depletion of the acid‐prevalent PacC72 form. We additionally show that pac C repression requires PacX. pac X mutations suppress PacC processing recalcitrant mutations, in part, through derepressed PacC levels resulting in traces of PacC27 formed by pH‐independent proteolysis. pac X was cloned by impala transposon mutagenesis. PacX, with homologues within the Leotiomyceta, has an unusual structure with an amino‐terminal coiled‐coil and a carboxy‐terminal zinc binuclear cluster. pacX mutations indicate the importance of these regions. One mutation, an unprecedented finding in A . nidulans genetics, resulted from an insertion of an endogenous Fot1‐like transposon. PMID:26303777

  16. Internalization of Aspergillus fumigatus conidia by epithelial and endothelial cells.

    PubMed Central

    Paris, S; Boisvieux-Ulrich, E; Crestani, B; Houcine, O; Taramelli, D; Lombardi, L; Latgé, J P

    1997-01-01

    The internalization of conidia of the opportunistic fungus Aspergillus fumigatus by primary cell cultures of nonprofessional phagocytes was investigated. This study is the first to show that A. fumigatus conidia were able to be engulfed by tracheal epithelial, alveolar type II, and endothelial cells. PMID:9119494

  17. Field ecology, fungal sex and food contamination involving Aspergillus species

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Several species within the genus Aspergillus are capable of producing a myriad of toxic secondary metabolites, with aflatoxin being of most concern. These fungi happen to colonize important agricultural commodities, thereby having the potential to contaminate our food with carcinogenic aflatoxins. P...

  18. Genetic Response to Seed Colonizatin by Aspergillus flavus in Peanut

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Previous studies to evaluate peanut genotypes for in vitro resistance to seed colonization by Aspergillus flavus have not resulted in the development of cultivars with resistance to aflatoxin contamination in the field. New breeding lines showing pre-harvest field resistance to aflatoxin contaminat...

  19. Polyphasic taxonomy of Aspergillus section Fumigati and its teleomorph Neosartorya

    Technology Transfer Automated Retrieval System (TEKTRAN)

    We revised the taxonomy of Aspergillus section Fumigati along with its teleomorph genus Neosartorya. Our species concept is based phenotype (morphology and extrolite profiles) and molecular (beta-tubulin and calmodulin gene sequences) characters in a polyphasic approach. Four new taxa are proposed:...

  20. Potential of Aspergillus flavus Genomics for Applications in Biotechnology

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Aspergillus flavus is a common saprophyte and opportunistic pathogen that survives in the natural environment by extracting nutrition from plant debris, insect carcasses and a variety of other carbon sources. A. flavus produces numerous secondary metabolites and hydrolytic enzymes. The primary obj...

  1. Nuclear heterogeneity in conidial populations of Aspergillus flavus

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Aspergillus flavus is a major producer of aflatoxin and an opportunistic pathogen for a wide range of hosts. Understanding genotypic and phenotypic variations within strains of A. flavus is important for controlling disease and reducing aflatoxin contamination. A. flavus is multinucleate and predomi...

  2. 21 CFR 866.3040 - Aspergillus spp. serological reagents.

    Code of Federal Regulations, 2012 CFR

    2012-04-01

    ... 21 Food and Drugs 8 2012-04-01 2012-04-01 false Aspergillus spp. serological reagents. 866.3040 Section 866.3040 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) MEDICAL DEVICES IMMUNOLOGY AND MICROBIOLOGY DEVICES Serological Reagents § 866.3040...

  3. 21 CFR 866.3040 - Aspergillus spp. serological reagents.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... 21 Food and Drugs 8 2010-04-01 2010-04-01 false Aspergillus spp. serological reagents. 866.3040 Section 866.3040 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) MEDICAL DEVICES IMMUNOLOGY AND MICROBIOLOGY DEVICES Serological Reagents § 866.3040...

  4. 21 CFR 866.3040 - Aspergillus spp. serological reagents.

    Code of Federal Regulations, 2014 CFR

    2014-04-01

    ... 21 Food and Drugs 8 2014-04-01 2014-04-01 false Aspergillus spp. serological reagents. 866.3040 Section 866.3040 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) MEDICAL DEVICES IMMUNOLOGY AND MICROBIOLOGY DEVICES Serological Reagents § 866.3040...

  5. 21 CFR 866.3040 - Aspergillus spp. serological reagents.

    Code of Federal Regulations, 2011 CFR

    2011-04-01

    ... 21 Food and Drugs 8 2011-04-01 2011-04-01 false Aspergillus spp. serological reagents. 866.3040 Section 866.3040 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) MEDICAL DEVICES IMMUNOLOGY AND MICROBIOLOGY DEVICES Serological Reagents § 866.3040...

  6. 21 CFR 866.3040 - Aspergillus spp. serological reagents.

    Code of Federal Regulations, 2013 CFR

    2013-04-01

    ... 21 Food and Drugs 8 2013-04-01 2013-04-01 false Aspergillus spp. serological reagents. 866.3040 Section 866.3040 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) MEDICAL DEVICES IMMUNOLOGY AND MICROBIOLOGY DEVICES Serological Reagents § 866.3040...

  7. QUANTITATIVE PCR OF SELECTED ASPERGILLUS, PENICILLIUM AND PAECILOMYCES SPECIES

    EPA Science Inventory

    A total of 65 quantitative PCR (QPCR) assays, incorporating fluorigenic 5' nuclease (TaqMan®) chemistry and directed at the nuclear ribosomal RNA operon, internal transcribed spacer regions (ITS1 or ITS2) was developed and tested for the detection of Aspergillus, Penicillium and ...

  8. Heritability study of eGFP-transformed Aspergillus flavus strains

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Pre-harvest prevention of aflatoxin contamination of corn, cottonseed, and peanut through field inoculation with non-aflatoxigenic Aspergillus flavus appears to be the only method for biocontrol currently being used. Until recently, evidence for out-crossing in A. flavus was observed in agar slants...

  9. The maize rachis affects Aspergillus flavus movement during ear development

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Aspergillus flavus expressing green fluorescent protein (GFP) was used to follow infection in ears of maize hybrids resistant and susceptible to the fungus. Developing ears were needle-inoculated with GFP-transformed A. flavus 20 days after silk emergence, and GFP fluorescence in the pith was evalu...

  10. Aflatoxin production and oxidative stress in Aspergillus flavus

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The colonization of crops by Aspergillus flavus results in the production of aflatoxins. Aflatoxin production is also exacerbated by abiotic stresses in the field. Here, we investigated the role of reactive oxygen species (ROS), which accumulate in plant tissues in response to drought and heat stres...

  11. Taxonomic revision of Eurotium and transfer of species to Aspergillus

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Section Aspergillus contains economically important, xerophilic species widely distributed in nature and the human environment that are known for their ability to grow on substrates with low water activity. The high level of phenoplasticity and frequent occurrence of mutants with atypical morphology...

  12. Population dynamics of Aspergillus flavus following biocontrol treatment of corn

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Aspergillus flavus is a fungal pathogen of many agronomically important crops worldwide. We sampled A. flavus strains from a cornfield in Rocky Mount, North Carolina, over a period of two years. The field was planted in 2010 and plots were inoculated at tasselling with either AF36 or NRRL 21882 (=Af...

  13. Population structure of Aspergillus flavus before and after biocontrol treatment

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Aspergillus flavus is a fungal pathogen of many important crops worldwide. We sampled A. flavus strains from a cornfield in Rocky Mount, North Carolina, over a period of two years. Plots were inoculated at tasselling with either AF36 or NRRL 21882 (=Afla-Guard) biocontrol strains, both of which are ...

  14. Biotransformation of quinazoline and phthalazine by Aspergillus niger.

    PubMed

    Sutherland, John B; Heinze, Thomas M; Schnackenberg, Laura K; Freeman, James P; Williams, Anna J

    2011-03-01

    Cultures of Aspergillus niger NRRL-599 in fluid Sabouraud medium were grown with quinazoline and phthalazine for 7 days. Metabolites were purified by high-performance liquid chromatography and identified by mass spectrometry and proton nuclear magnetic resonance spectroscopy. Quinazoline was oxidized to 4-quinazolinone and 2,4-quinazolinedione, and phthalazine was oxidized to 1-phthalazinone.

  15. Cryptic Sexuality Influences Aflatoxigenicity in Aspergillus parasiticus and A. flavus

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Ascomycetous fungi of the genus Aspergillus comprise a wide variety of species of biotechnological importance as well as pathogens and toxin producers. Recent studies report A. fumigatus to be heterothallic and possibly undergoing sexual reproduction. We therefore investigated whether compatible mat...

  16. Genomic sequence for the aflatoxigenic filamentous fungus Aspergillus nomius

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The genome of the A. nomius type strain was sequenced using a personal genome machine. Annotation of the genes was undertaken, followed by gene ontology and an investigation into the number of secondary metabolite clusters. Comparative studies with other Aspergillus species involved shared/unique ge...

  17. Kinesin-3 in the basidiomycete Ustilago maydis transports organelles along the entire microtubule array.

    PubMed

    Steinberg, Gero

    2015-01-01

    The molecular motor kinesin-3 transports early endosomes along microtubules in filamentous fungi. It was reported that kinesin-3 from the ascomycete fungi Aspergillus nidulans and Neurospora crassa use a subset of post-translationally modified and more stable microtubules. Here, I show that kinesin-3 from the basidiomycete Ustilago maydis moves along all hyphal microtubules. This difference is likely due to variation in cell cycle control and associated organization of the microtubule array.

  18. Metabolomics Analysis Reveals Specific Novel Tetrapeptide and Potential Anti-Inflammatory Metabolites in Pathogenic Aspergillus species.

    PubMed

    Lee, Kim-Chung; Tam, Emily W T; Lo, Ka-Ching; Tsang, Alan K L; Lau, Candy C Y; To, Kelvin K W; Chan, Jasper F W; Lam, Ching-Wan; Yuen, Kwok-Yung; Lau, Susanna K P; Woo, Patrick C Y

    2015-01-01

    Infections related to Aspergillus species have emerged to become an important focus in infectious diseases, as a result of the increasing use of immunosuppressive agents and high fatality associated with invasive aspergillosis. However, laboratory diagnosis of Aspergillus infections remains difficult. In this study, by comparing the metabolomic profiles of the culture supernatants of 30 strains of six pathogenic Aspergillus species (A. fumigatus, A. flavus, A. niger, A. terreus, A. nomius and A. tamarii) and 31 strains of 10 non-Aspergillus fungi, eight compounds present in all strains of the six Aspergillus species but not in any strain of the non-Aspergillus fungi were observed. One of the eight compounds, Leu-Glu-Leu-Glu, is a novel tetrapeptide and represents the first linear tetrapeptide observed in Aspergillus species, which we propose to be named aspergitide. Two other closely related Aspergillus-specific compounds, hydroxy-(sulfooxy)benzoic acid and (sulfooxy)benzoic acid, may possess anti-inflammatory properties, as 2-(sulfooxy)benzoic acid possesses a structure similar to those of aspirin [2-(acetoxy)benzoic acid] and salicylic acid (2-hydroxybenzoic acid). Further studies to examine the potentials of these Aspergillus-specific compounds for laboratory diagnosis of aspergillosis are warranted and further experiments will reveal whether Leu-Glu-Leu-Glu, hydroxy-(sulfooxy)benzoic acid and (sulfooxy)benzoic acid are virulent factors of the pathogenic Aspergillus species. PMID:26090713

  19. Metabolomics Analysis Reveals Specific Novel Tetrapeptide and Potential Anti-Inflammatory Metabolites in Pathogenic Aspergillus species.

    PubMed

    Lee, Kim-Chung; Tam, Emily W T; Lo, Ka-Ching; Tsang, Alan K L; Lau, Candy C Y; To, Kelvin K W; Chan, Jasper F W; Lam, Ching-Wan; Yuen, Kwok-Yung; Lau, Susanna K P; Woo, Patrick C Y

    2015-06-17

    Infections related to Aspergillus species have emerged to become an important focus in infectious diseases, as a result of the increasing use of immunosuppressive agents and high fatality associated with invasive aspergillosis. However, laboratory diagnosis of Aspergillus infections remains difficult. In this study, by comparing the metabolomic profiles of the culture supernatants of 30 strains of six pathogenic Aspergillus species (A. fumigatus, A. flavus, A. niger, A. terreus, A. nomius and A. tamarii) and 31 strains of 10 non-Aspergillus fungi, eight compounds present in all strains of the six Aspergillus species but not in any strain of the non-Aspergillus fungi were observed. One of the eight compounds, Leu-Glu-Leu-Glu, is a novel tetrapeptide and represents the first linear tetrapeptide observed in Aspergillus species, which we propose to be named aspergitide. Two other closely related Aspergillus-specific compounds, hydroxy-(sulfooxy)benzoic acid and (sulfooxy)benzoic acid, may possess anti-inflammatory properties, as 2-(sulfooxy)benzoic acid possesses a structure similar to those of aspirin [2-(acetoxy)benzoic acid] and salicylic acid (2-hydroxybenzoic acid). Further studies to examine the potentials of these Aspergillus-specific compounds for laboratory diagnosis of aspergillosis are warranted and further experiments will reveal whether Leu-Glu-Leu-Glu, hydroxy-(sulfooxy)benzoic acid and (sulfooxy)benzoic acid are virulent factors of the pathogenic Aspergillus species.

  20. [Indoor fungal exposure: What impact on clinical and biological status regarding Aspergillus during cystic fibrosis].

    PubMed

    Pricope, D; Deneuville, E; Frain, S; Chevrier, S; Belaz, S; Roussey, M; Gangneux, J-P

    2015-06-01

    The sources of exposure during diseases due to Aspergillus fungi in cystic fibrosis patients are still poorly explored. We assessed home fungal exposure in patients suffering from cystic fibrosis and analysed its impact on the presence of Aspergillus biological markers, the colonisation of airways, as well as the sensitization and Aspergillus serology. Between March 2012 and August 2012, 34 patients benefited from a visit performed by a home environment medical adviser including sampling for mycological analysis. The number of colonies of Aspergillus was not significantly different in the various sampling sites (P=0.251), but the number of non-Aspergillus colonies was much higher in the kitchen (P=0.0045). Subsequently, home fungal exposure was compared between the groups "absence of Aspergillus-related markers" and "presence of Aspergillus-related markers". Home exposure to Aspergillus (P=0.453) and non-Aspergillus (P=0.972) flora was not significant between the 2 groups. Within this series of 34 patients that should be expanded, we note an absence of clear relationship between home exposure and the Aspergillus-linked markers in patients suffering from cystic fibrosis. This result should be taken into account regarding too restrictive hygiene advices provided to families, given the fact that fungal exposure can also results from activities performed away from home.

  1. Clinical Performance of Aspergillus PCR for Testing Serum and Plasma: a Study by the European Aspergillus PCR Initiative.

    PubMed

    White, P Lewis; Barnes, Rosemary A; Springer, Jan; Klingspor, Lena; Cuenca-Estrella, Manuel; Morton, C Oliver; Lagrou, Katrien; Bretagne, Stéphane; Melchers, Willem J G; Mengoli, Carlo; Donnelly, J Peter; Heinz, Werner J; Loeffler, Juergen

    2015-09-01

    Aspergillus PCR testing of serum provides technical simplicity but with potentially reduced sensitivity compared to whole-blood testing. With diseases for which screening to exclude disease represents an optimal strategy, sensitivity is paramount. The associated analytical study confirmed that DNA concentrations were greater in plasma than those in serum. The aim of the current investigation was to confirm analytical findings by comparing the performance of Aspergillus PCR testing of plasma and serum in the clinical setting. Standardized Aspergillus PCR was performed on plasma and serum samples concurrently obtained from hematology patients in a multicenter retrospective anonymous case-control study, with cases diagnosed according to European Organization for Research and Treatment of Cancer/Invasive Fungal Infections Cooperative Group and the National Institute of Allergy and Infectious Diseases Mycoses Study Group (EORTC/MSG) consensus definitions (19 proven/probable cases and 42 controls). Clinical performance and clinical utility (time to positivity) were calculated for both kinds of samples. The sensitivity and specificity for Aspergillus PCR when testing serum were 68.4% and 76.2%, respectively, and for plasma, they were 94.7% and 83.3%, respectively. Eighty-five percent of serum and plasma PCR results were concordant. On average, plasma PCR was positive 16.8 days before diagnosis and was the earliest indicator of infection in 13 cases, combined with other biomarkers in five cases. On average, serum PCR was positive 10.8 days before diagnosis and was the earliest indicator of infection in six cases, combined with other biomarkers in three cases. These results confirm the analytical finding that the sensitivity of Aspergillus PCR using plasma is superior to that using serum. PCR positivity occurs earlier when testing plasma and provides sufficient sensitivity for the screening of invasive aspergillosis while maintaining methodological simplicity.

  2. Loss of msnA, a putative stress regulatory gene, in Aspergillus parasiticus and Aspergillus flavus increased production of conidia, aflatoxins and kojic acid

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Production of the harmful carcinogenic aflatoxins by Aspergillus parasiticus and Aspergillus flavus has been postulated to be a mechanism to relieve oxidative stress. The msnA gene, the ortholog of Saccharomyces cerevisiae MSN2 associated with multi-stress response, of the two species was disrupted....

  3. Variants of Aspergillus alutaceus var. alutaceus (formerly Aspergillus ochraceus) with altered ochratoxin A production.

    PubMed Central

    Chelack, W S; Borsa, J; Szekely, J G; Marquardt, R R; Frohlich, A A

    1991-01-01

    The present studies, using Aspergillus alutaceus var. alutaceus Berkeley et Curtis (formerly A. ochraceus Wilhelm) NRRL 3174 along with three other wild-type strains, were undertaken in an attempt to understand the effects of irradiation and other treatments on mycotoxin production in grain. Bedford barley was inoculated with spores of NRRL 3174, gamma irradiated, and incubated at 28 degrees C and 25% moisture. After 10 days of incubation, two colony types, ochre (parental) and yellow (variant), were isolated from the grain. Further culturing of the yellow variant resulted in the spontaneous appearance of a white variant that exhibited greatly enhanced fluorescence under UV light. In subsequent work, we have also isolated variants producing a soluble red pigment. In addition, in model experiments involving irradiation (1 kGy) of pure cultures, induction frequencies ranging between 2 and 4% (survival basis) were observed for the yellow and red variants. Inoculation of these variants into wheat and incubation for 14 days at 28 degrees C and 32% moisture resulted in ochratoxin A production in the relative amounts of 0.09:1:4.6:9.3 for the red, ochre (parental), yellow, and white variants, respectively. Additional characteristics of these isolates are described. Confirmation that the white high-ochratoxin-A-producing variants were derived from the parental strain was demonstrated by obtaining revertant sectors in monoclonal cultures of the variants. Images PMID:1768122

  4. Voriconazole Exposure and Risk of Cutaneous Squamous Cell Carcinoma, Aspergillus Colonization, Invasive Aspergillosis and Death in Lung Transplant Recipients.

    PubMed

    Mansh, M; Binstock, M; Williams, K; Hafeez, F; Kim, J; Glidden, D; Boettger, R; Hays, S; Kukreja, J; Golden, J; Asgari, M M; Chin-Hong, P; Singer, J P; Arron, S T

    2016-01-01

    Voriconazole is a triazole antifungal used to prevent and treat invasive fungal infections after lung transplantation, but it has been associated with an increased risk of developing cutaneous squamous cell carcinoma (SCC). Despite widespread use, there are no clear guidelines for optimal prophylactic regimens that balance the competing risks and benefits. We conducted a retrospective cohort study of all lung transplant recipients at the University of California, San Francisco, who were transplanted between October 1991 and December 2012 (n = 455) to investigate whether voriconazole exposure affected development of SCC, Aspergillus colonization, invasive aspergillosis and all-cause mortality. Voriconazole exposure was associated with a 73% increased risk of developing SCC (hazard ratio [HR] 1.73; 95% confidence interval [CI]: 1.04-2.88; p = 0.03), with each additional 30-day exposure at the standard dose increasing the risk by 3.0% (HR 1.03; 95% CI: 1.02-1.04; p < 0.001). Voriconazole exposure reduced risk of Aspergillus colonization by 50% (HR 0.50; 95% CI: 0.34-0.72; p < 0.001), but we were underpowered to detect risk reduction for invasive aspergillosis. Voriconazole exposure significantly reduced all-cause mortality among subjects who developed Aspergillus colonization (HR 0.34; 95% CI: 0.13-0.91; p = 0.03) but had no significant impact on those without colonization. Physicians should consider patient-specific factors that modify the potential risks and benefits of voriconazole for the care of lung transplant recipients. PMID:26372838

  5. Rhamnogalacturonan I modifying enzymes: an update.

    PubMed

    Silva, Inês R; Jers, Carsten; Meyer, Anne S; Mikkelsen, Jørn Dalgaard

    2016-01-25

    Rhamnogalacturonan I (RGI) modifying enzymes catalyse the degradation of the RGI backbone and encompass enzymes specific for either the α1,2-bond linking galacturonic acid to rhamnose or the α1,4-bond linking rhamnose to galacturonic acid in the RGI backbone. The first microbial enzyme found to be able to catalyse the degradation of the RGI backbone, an endo-hydrolase (EC 3.2.1.171) derived from Aspergillus aculeatus, was discovered 25 years ago. Today the group of RGI modifying enzymes encompasses endo- and exo-hydrolases as well as lyases. The RGI hydrolases, EC 3.2.1.171-EC 3.2.1.174, have been described to be produced by Aspergillus spp. and Bacillus subtilis and are categorized in glycosyl hydrolase families 28 and 105. The RGI lyases, EC 4.2.2.23-EC 4.2.2.24, have been isolated from different fungi and bacterial species and are categorized in polysaccharide lyase families 4 and 11. This review brings together the available knowledge of the RGI modifying enzymes and provides a detailed overview of biocatalytic reaction characteristics, classification, structure-function traits, and analyses the protein properties of these enzymes by multiple sequence alignments in neighbour-joining phylogenetic trees. Some recently detected unique structural features and dependence of calcium for activity of some of these enzymes (notably the lyases) are discussed and newly published results regarding improvement of their thermostability by protein engineering are highlighted. Knowledge of these enzymes is important for understanding microbial plant cell wall degradation and for advancing enzymatic processing and biorefining of pectinaceous plant biomass.

  6. Vaccination approaches against opportunistic fungal infections caused by Aspergillus fumigatus.

    PubMed

    Reichard, Utz; Herrmann, Sahra; Asif, Abdul R

    2014-01-01

    Although innate immunity primarily combats systemic infections of opportunistic fungi such as Aspergillus and Candida spp., acquired and protective immunoreactions were observed long ago in animal trials following sublethal systemic infections caused by viable fungi or after challenging animals with inactivated fungal cells. Based on these observations, fungal antigens should exist which mediate such protective immunoreactions and have in part already been identified. In this context, this review focuses primarily on the various approaches that have been used to identify protection-mediating Aspergillus-antigens and their rationale. Emphasis is placed on screening methods that have exploited genetic or proteomic approaches on the basis of the corresponding fungal genome projects. Thereby, a survey and description is given of the antigens so far known to be capable of inducing immune responses that protect animals against acquiring lethal systemic aspergillosis.

  7. Purification of soyasaponin -β-galactosidase from Aspergillus sp.39

    NASA Astrophysics Data System (ADS)

    Tian, Jing; Zhao, Ping; Xu, Longquan; Fei, Xu; Wang, Yi

    In order to increase physiological activity of soyasaponin, enzyme hydrolysis of soyasaponin was studied. The enzyme which hydrolyzes soyasaponin to lower sugar soyasaponin was obtained from Aspergillus sp.39s. And it was purified by the method of biologic chromatography system. The method of SDS-polyacrylamide gel electrophoresis was used to determine the molecular weight of the enzyme produced by Aspergillus sp.39s. The molecular weight was about 50 kDa. The optimum pH and temperature of soyasaponin-β-galactosidase produced from sp.39s was 5.0 and 40°C respectively. Soyasaponin-β-galactosidase was comparatively stable in the pH range from 3.0 to 7.0 and in the temperature range from 20°C to 60°C.

  8. Rapid and Sensitive Plate Method for Detection of Aspergillus fumigatus

    PubMed Central

    Bauters, T. G. M.; Nelis, H. J.

    2000-01-01

    The routine identification of Aspergillus fumigatus in clinical samples involves, apart from direct examination, the isolation of the organism on a plate followed by its microscopic characterization. This approach lacks sensitivity, specificity, and speed. A new procedure has been developed combining microcolony formation on a nylon membrane filter at 45°C with the detection of a specific 4-methylumbelliferyl-α-l-arabinopyranoside cleaving enzyme activity in digitonin permeabilized cells. The test takes approximately 14 h and has an efficiency of 98.2% and false-positive and -negative rates of 0 and 3.1%, respectively. When applied to 188 clinical samples taken from patients with proven or nonproven presence of Aspergillus species, a good agreement with the conventional plate-microscopy method was obtained. PMID:11015405

  9. Fitness Studies of Azole-Resistant Strains of Aspergillus fumigatus

    PubMed Central

    Valsecchi, Isabel; Mellado, Emilia; Beau, Rémi; Raj, Shriya

    2015-01-01

    Isogenic bar-coded strains of Aspergillus fumigatus carrying the G54W or M220K mutation in Cyp51A were constructed. In vitro, the growth and conidiation capacities of the mutants were similar to those of the parental strain. Competition studies in the absence of azoles showed that there was no adverse fitness cost for the azole-resistant A. fumigatus strains in vitro or in vivo compared to the parental strain. PMID:26416854

  10. Single cell transcriptomics of neighboring hyphae of Aspergillus niger

    PubMed Central

    2011-01-01

    Single cell profiling was performed to assess differences in RNA accumulation in neighboring hyphae of the fungus Aspergillus niger. A protocol was developed to isolate and amplify RNA from single hyphae or parts thereof. Microarray analysis resulted in a present call for 4 to 7% of the A. niger genes, of which 12% showed heterogeneous RNA levels. These genes belonged to a wide range of gene categories. PMID:21816052

  11. Mycotoxins produced by Aspergillus fumigatus isolated from silage.

    PubMed

    Cole, R J; Kirksey, J W; Dorner, J W; Wilson, D M; Johnson, J; Bedell, D; Springer, J P; Chexal, K K; Clardy, J; Cox, R H

    1977-01-01

    Results are presented which show that Aspergillus fumigatus was one of the predominant fungi contaminating moldy silage. Growth of A. fumigatus on silage appeared to depend on a preliminary aerobic fermentation by other natural microflora in silage. The clavine alkaloid, fumigaclavine A, and a new clavine alkaloid designated fumigaclavine C were produced by A. fumigatus. The LD50 of fumigaclavine C was approximately 150 mg/kg oral dose in day-old cockerels. PMID:350117

  12. Monitoring environmental Aspergillus spp. contamination and meteorological factors in a haematological unit.

    PubMed

    Cavallo, M; Andreoni, S; Martinotti, M G; Rinaldi, M; Fracchia, L

    2013-12-01

    The opportunistic pathogens belonging to the Aspergillus genus are present in almost all seasons of the year, and their concentration is related to meteorological conditions. The high density of Aspergillus spp. conidia in a haematological hospital ward may be a significant risk factor for developing invasive fungal diseases in immunocompromised patients. Aim of the present study was to evaluate the variability of airborne Aspergillus spp. conidia contamination in a Haematological Unit (HU) within a period of 16 months in relation with some meteorological parameters. An environmental Aspergillus surveillance was conducted in the HU in four rooms and their bathrooms, in the corridor and in three external sites using an agar impact sampler. During each sampling, temperature and relative humidity at each site were recorded and current wind speed and rainfall events were taken from the official weather service. Aspergillus spp. conidia concentration differed significantly across the sampling sites. Internal Aspergillus spp. loads were significantly dependent on temperature, internal relative humidity and rain. External conidia concentrations were significantly influenced by outdoor temperature and relative humidity. A suitable indicator was introduced to evaluate the seasonal distribution of Aspergillus spp. conidia in the sampling sites, and a significant dependence on this indicator was observed inside the HU. Seventeen different fungal species belonging to the Aspergillus genus were detected during the sampling period. Aspergillus fumigatus was the most frequently isolated species and its distribution depended significantly on the seasonal indicator both inside and outside the hospital ward.

  13. The distribution of Aspergillus spp. opportunistic parasites in hives and their pathogenicity to honey bees.

    PubMed

    Foley, Kirsten; Fazio, Géraldine; Jensen, Annette B; Hughes, William O H

    2014-03-14

    Stonebrood is a disease of honey bee larvae caused by fungi from the genus Aspergillus. As very few studies have focused on the epidemiological aspects of stonebrood and diseased brood may be rapidly discarded by worker bees, it is possible that a high number of cases go undetected. Aspergillus spp. fungi are ubiquitous and associated with disease in many insects, plants, animals and man. They are regarded as opportunistic pathogens that require immunocompromised hosts to establish infection. Microbiological studies have shown high prevalences of Aspergillus spp. in apiaries which occur saprophytically on hive substrates. However, the specific conditions required for pathogenicity to develop remain unknown. In this study, an apiary was screened to determine the prevalence and diversity of Aspergillus spp. fungi. A series of dose-response tests were then conducted using laboratory reared larvae to determine the pathogenicity and virulence of frequently occurring isolates. The susceptibility of adult worker bees to Aspergillus flavus was also tested. Three isolates (A. flavus, Aspergillus nomius and Aspergillus phoenicis) of the ten species identified were pathogenic to honey bee larvae. Moreover, adult honey bees were also confirmed to be highly susceptible to A. flavus infection when they ingested conidia. Neither of the two Aspergillus fumigatus strains used in dose-response tests induced mortality in larvae and were the least pathogenic of the isolates tested. These results confirm the ubiquity of Aspergillus spp. in the apiary environment and highlight their potential to infect both larvae and adult bees.

  14. Aspergillus as a multi-purpose cell factory: current status and perspectives.

    PubMed

    Meyer, Vera; Wu, Bo; Ram, Arthur F J

    2011-03-01

    Aspergilli have a long history in biotechnology as expression platforms for the production of food ingredients, pharmaceuticals and enzymes. The achievements made during the last years, however, have the potential to revolutionize Aspergillus biotechnology and to assure Aspergillus a dominant place among microbial cell factories. This mini-review will highlight most recent breakthroughs in fundamental and applied Aspergillus research with a focus on new molecular tools, techniques and products. New trends and concepts related to Aspergillus genomics and systems biology will be discussed as well as the challenges that have to be met to integrate omics data with metabolic engineering attempts.

  15. Biodegradation of phenol by Antarctic strains of Aspergillus fumigatus.

    PubMed

    Gerginova, Maria; Manasiev, Jordan; Yemendzhiev, Husein; Terziyska, Anna; Peneva, Nadejda; Alexieva, Zlatka

    2013-01-01

    Taxonomic identification of three newly isolated Antarctic fungal strains by their 18S rDNA sequences revealed their affiliation with Aspergillus fumigatus. Phenol (0.5 g/l) as the sole carbon source was completely degraded by all strains within less than two weeks. Intracellular activities of three key enzymes involved in the phenol catabolism were determined. Activities of phenol hydroxylase (EC 1.14.13.7), hydroquinone hydroxylase (EC 1.14.13.x), and catechol 1,2-dioxygenase (EC 1.13.11.1) varied significantly between strains. The rates of phenol degradation in the three strains correlated best with the activity of catechol 1,2-dioxygenase. Six pairs of oligonucleotide primers were designed on the basis of the Aspergillus fumigatus Af293 genome sequence (NCBI Acc. No. XM_743491.1) and used to amplify phenol hydroxylase-related gene sequences. DNA sequences of about 1200 bp were amplified from all three strains and found to have a high degree of sequence identity with the corresponding gene of Aspergillus fumigatus Af293.

  16. Molecular analysis of Aspergillus section Flavi isolated from Brazil nuts.

    PubMed

    Gonçalves, Juliana Soares; Ferracin, Lara Munique; Carneiro Vieira, Maria Lucia; Iamanaka, Beatriz Thie; Taniwaki, Marta Hiromi; Pelegrinelli Fungaro, Maria Helena

    2012-04-01

    Brazil nuts are an important export market in its main producing countries, including Brazil, Bolivia, and Peru. Approximately 30,000 tons of Brazil nuts are harvested each year. However, substantial nut contamination by Aspergillus section Flavi occurs with subsequent production of aflatoxins. In our study, Aspergillus section Flavi were isolated from Brazil nuts (Bertholletia excelsa), and identified by morphological and molecular means. We obtained 241 isolates from nut samples, 41% positive for aflatoxin production. Eighty-one isolates were selected for molecular investigation. Pairwise genetic distances among isolates and phylogenetic relationships were assessed. The following Aspergillus species were identified: A. flavus, A. caelatus, A. nomius, A. tamarii, A. bombycis, and A. arachidicola. Additionally, molecular profiles indicated a high level of nucleotide variation within β-tubulin and calmodulin gene sequences associated with high genetic divergence from RAPD data. Among the 81 isolates analyzed by molecular means, three of them were phylogenetically distinct from all other isolates representing the six species of section Flavi. A putative novel species was identified based on molecular profiles.

  17. Allergic Aspergillus sinusitis and its association with allergic bronchopulmonary aspergillosis

    PubMed Central

    Panjabi, Chandramani

    2011-01-01

    Allergic Aspergillus sinusitis (AAS) is a three decade old clinicopathologic entity in which mucoid impaction akin to that of allergic bronchopulmonary aspergillosis (ABPA) occurs in the paranasal sinuses. Features such as radiographic evidence of pansinusitis, passage of nasal plugs and recurrent nasal polyposis in patients with an atopic background is suggestive of AAS. Histopathlogic confirmation from the inspissated mucus is a sine qua non for the diagnosis. Heterogeneous densities on computed tomography of the paranasal sinuses are caused by the 'allergic mucin' in the sinuses. Many patients give a history of having undergone multiple surgical procedures for symptomatic relief. The current approach to treatment appears to include an initial surgical debridement followed by postoperative oral corticosteroids for long durations. Although both ABPA and AAS are classified as Aspergillus-related hypersensitivity respiratory disorders, their co-occurrence appears to be an infrequently recognised phenomenon. This could perhaps be attributed to the fact that these two diseases are often treated by two different specialties. A high index of suspicion is required to establish the diagnoses of ABPA and AAS. All patients with asthma and/or rhinosinusitis along with sensitisation to Aspergillus antigens are at an increased risk of developing ABPA and/or AAS. ABPA must be excluded in all patients with AAS and vice versa. Early diagnosis and initiation of appropriate therapy could plausibly alter the course of the disease processes and prevent the possible development of long term sequelae. PMID:22053309

  18. Aspergillus fumigatus-Related Species in Clinical Practice

    PubMed Central

    Lamoth, Frédéric

    2016-01-01

    Aspergillus fumigatus is the main etiologic agent of invasive aspergillosis (IA). Other Aspergillus species belonging to the section Fumigati (A. fumigatus complex) may occasionally be the cause of IA. These strains are often misidentified, as they cannot be distinguished from A. fumigatus by conventional morphological analysis and sequencing methods. This lack of recognition may have important consequences as these A. fumigatus-related species often display some level of intrinsic resistance to azoles and other antifungal drugs. A. lentulus, A. udagawae, A. viridinutans, and A. thermomutatus (Neosartorya pseudofischeri) have been associated with refractory cases of IA. Microbiologists should be able to suspect the presence of these cryptic species behind a putative A. fumigatus isolate on the basis of some simple characteristics, such as defect in sporulation and/or unusual antifungal susceptibility profile. However, definitive species identification requires specific sequencing analyses of the beta-tubulin or calmodulin genes, which are not available in most laboratories. Multiplex PCR assays or matrix-assisted laser desorption ionization – time-of-flight mass spectrometry (MALDI-TOF MS) gave promising results for rapid and accurate distinction between A. fumigatus and other Aspergillus spp. of the section Fumigati in clinical practice. Improved diagnostic procedures and antifungal susceptibility testing may be helpful for the early detection and management of these particular IA cases. PMID:27242710

  19. In vitro activity of disinfectants against Aspergillus spp.

    PubMed

    Mattei, A S; Madrid, I M; Santin, R; Schuch, L F D; Meireles, M C A

    2013-01-01

    Fungi of the Aspergillus genus are widespread and contaminate the environment. Thousands of conidia are released from each phialide and dispersed in the air every day. These fungi are considered important mycose-causing agents in hospitals. Due to this, research to determine prevalent fungi from the Aspergillus genus in hospital environments, and an adequate disinfection program in these areas is are needed. This study evaluated the susceptibility of Aspergillus spp. isolated from a veterinary environment against four disinfectants. Successive dilutions of disinfectants (log2) were used according to CLSI M38-A2 microdilution technique adapted to chemical agents against 18 isolates of this genus. After 72 hours of incubation, the Minimum Inhibiting Concentration and Minimum Fungicidal Concentration capable of inhibiting 50% and 90% of the isolates were determined. Chlorexidine-cetrimine, benzalconium chloride and a chlorophenol derivative proved to be effective against all isolates with a lower MIC than that suggested by the manufacturer, except for the A. flavus strain. Sodium hypochlorite was ineffective against three A. fumigatus, three A. flavus and one A. niger isolate. These results demonstrated that all studied disinfectants were effective against environmental isolates, with the exception of sodium hypochlorite, which showed lower effectiveness.

  20. Multi-triazole-resistant Aspergillus fumigatus infections in Australia.

    PubMed

    Kidd, Sarah E; Goeman, Emma; Meis, Jacques F; Slavin, Monica A; Verweij, Paul E

    2015-06-01

    The emergence of triazole resistance, including multi-triazole-resistant Aspergillus fumigatus is being reported around the world, but there has been little evidence of this problem to date in Australia. Here we describe a retrospective search of antifungal susceptibility results of all Australian clinical A. fumigatus isolates referred to the National Mycology Reference Centre, Adelaide, Australia between 2000 and 2013, yielding 13 isolates with elevated minimum inhibitory concentrations to itraconazole, posaconazole and/or voriconazole. Four isolates were found to be Aspergillus lentulus, a closely related, morphologically similar species known to have reduced susceptibility to triazoles. Analysis of the cyp51A gene of nine confirmed A. fumigatus isolates revealed two carrying the TR34 /L98H mutation, one apparently locally acquired in 2004, and the other probably acquired abroad in 2012. Four isolates possessed the G54R, F46Y, Y431S and G448S mutations, respectively, whereas three isolates did not possess known cyp51A resistance mutations, raising the possibility of other, undetected resistance mechanisms. Routine antifungal susceptibility testing is definitively recommended in patients on long term and sub-therapeutic triazole therapy with breakthrough Aspergillus infection and recommended for all clinically relevant A. fumigatus isolates. PMID:25885568

  1. Biosynthesis and Characterization of Silver Nanoparticles by Aspergillus Species.

    PubMed

    Zomorodian, Kamiar; Pourshahid, Seyedmohammad; Sadatsharifi, Arman; Mehryar, Pouyan; Pakshir, Keyvan; Rahimi, Mohammad Javad; Arabi Monfared, Ali

    2016-01-01

    Currently, researchers turn to natural processes such as using biological microorganisms in order to develop reliable and ecofriendly methods for the synthesis of metallic nanoparticles. In this study, we have investigated extracellular biosynthesis of silver nanoparticles using four Aspergillus species including A. fumigatus, A. clavatus, A. niger, and A. flavus. We have also analyzed nitrate reductase activity in the studied species in order to determine the probable role of this enzyme in the biosynthesis of silver nanoparticles. The formation of silver nanoparticles in the cell filtrates was confirmed by the passage of laser light, change in the color of cell filtrates, absorption peak at 430 nm in UV-Vis spectra, and atomic force microscopy (AFM). There was a logical relationship between the efficiencies of studied Aspergillus species in the production of silver nanoparticles and their nitrate reductase activity. A. fumigatus as the most efficient species showed the highest nitrate reductase activity among the studied species while A. flavus exhibited the lowest capacity in the biosynthesis of silver nanoparticles which was in accord with its low nitrate reductase activity. The present study showed that Aspergillus species had potential for the biosynthesis of silver nanoparticles depending on their nitrate reductase activity. PMID:27652264

  2. Aspergillus fumigatus Endophthalmitis with Necrotizing Scleritis following Pars Plana Vitrectomy.

    PubMed

    Gruener, Anna M; Allen, Felicity; Stanford, Miles R; Graham, Elizabeth M

    2016-01-01

    We present a case of Aspergillus fumigatus endophthalmitis complicated by necrotizing scleritis in a 68-year-old man with diet-controlled diabetes, after retinal detachment repair. He was initially treated with systemic steroids for surgically induced necrotizing scleritis following routine pars plana vitrectomy. An additional diagnosis of endophthalmitis was made when the patient developed a hypopyon. Repeat vitreous culture isolated Aspergillus fumigatus. Symptoms improved following antifungal treatment leaving the patient with scleromalacia and an advanced postoperative cataract. Fungal scleritis and endophthalmitis are rare complications of intraocular surgery with sight-threatening consequences, and, as this case demonstrates, may even occur concomitantly. The overlapping features of both conditions can make differentiating one from the other difficult. A fungal aetiology should be considered in cases of postoperative scleritis and endophthalmitis that are protracted and refractory to standard therapy. Even in cases of early diagnosis and treatment, visual outcomes in Aspergillus endophthalmitis and scleritis are variable and often disappointing, not infrequently necessitating enucleation of a painful blind eye.

  3. Aspergillus fumigatus Endophthalmitis with Necrotizing Scleritis following Pars Plana Vitrectomy.

    PubMed

    Gruener, Anna M; Allen, Felicity; Stanford, Miles R; Graham, Elizabeth M

    2016-01-01

    We present a case of Aspergillus fumigatus endophthalmitis complicated by necrotizing scleritis in a 68-year-old man with diet-controlled diabetes, after retinal detachment repair. He was initially treated with systemic steroids for surgically induced necrotizing scleritis following routine pars plana vitrectomy. An additional diagnosis of endophthalmitis was made when the patient developed a hypopyon. Repeat vitreous culture isolated Aspergillus fumigatus. Symptoms improved following antifungal treatment leaving the patient with scleromalacia and an advanced postoperative cataract. Fungal scleritis and endophthalmitis are rare complications of intraocular surgery with sight-threatening consequences, and, as this case demonstrates, may even occur concomitantly. The overlapping features of both conditions can make differentiating one from the other difficult. A fungal aetiology should be considered in cases of postoperative scleritis and endophthalmitis that are protracted and refractory to standard therapy. Even in cases of early diagnosis and treatment, visual outcomes in Aspergillus endophthalmitis and scleritis are variable and often disappointing, not infrequently necessitating enucleation of a painful blind eye. PMID:27379189

  4. Biosynthesis and Characterization of Silver Nanoparticles by Aspergillus Species

    PubMed Central

    Pourshahid, Seyedmohammad; Mehryar, Pouyan; Pakshir, Keyvan; Rahimi, Mohammad Javad; Arabi Monfared, Ali

    2016-01-01

    Currently, researchers turn to natural processes such as using biological microorganisms in order to develop reliable and ecofriendly methods for the synthesis of metallic nanoparticles. In this study, we have investigated extracellular biosynthesis of silver nanoparticles using four Aspergillus species including A. fumigatus, A. clavatus, A. niger, and A. flavus. We have also analyzed nitrate reductase activity in the studied species in order to determine the probable role of this enzyme in the biosynthesis of silver nanoparticles. The formation of silver nanoparticles in the cell filtrates was confirmed by the passage of laser light, change in the color of cell filtrates, absorption peak at 430 nm in UV-Vis spectra, and atomic force microscopy (AFM). There was a logical relationship between the efficiencies of studied Aspergillus species in the production of silver nanoparticles and their nitrate reductase activity. A. fumigatus as the most efficient species showed the highest nitrate reductase activity among the studied species while A. flavus exhibited the lowest capacity in the biosynthesis of silver nanoparticles which was in accord with its low nitrate reductase activity. The present study showed that Aspergillus species had potential for the biosynthesis of silver nanoparticles depending on their nitrate reductase activity. PMID:27652264

  5. In vitro activity of disinfectants against Aspergillus spp

    PubMed Central

    Mattei, A.S.; Madrid, I.M.; Santin, R.; Schuch, L.F.D.; Meireles, M.C.A.

    2013-01-01

    Fungi of the Aspergillus genus are widespread and contaminate the environment. Thousands of conidia are released from each phialide and dispersed in the air every day. These fungi are considered important mycose-causing agents in hospitals. Due to this, research to determine prevalent fungi from the Aspergillus genus in hospital environments, and an adequate disinfection program in these areas is are needed. This study evaluated the susceptibility of Aspergillus spp. isolated from a veterinary environment against four disinfectants. Successive dilutions of disinfectants (log2) were used according to CLSI M38-A2 microdilution technique adapted to chemical agents against 18 isolates of this genus. After 72 hours of incubation, the Minimum Inhibiting Concentration and Minimum Fungicidal Concentration capable of inhibiting 50% and 90% of the isolates were determined. Chlorexidine-cetrimine, benzalconium chloride and a chlorophenol derivative proved to be effective against all isolates with a lower MIC than that suggested by the manufacturer, except for the A. flavus strain. Sodium hypochlorite was ineffective against three A. fumigatus, three A. flavus and one A. niger isolate. These results demonstrated that all studied disinfectants were effective against environmental isolates, with the exception of sodium hypochlorite, which showed lower effectiveness. PMID:24294243

  6. Association of airborne Aspergillus with asthma exacerbation in Southern Pakistan

    PubMed Central

    Zubairi, Ali Bin Sarwar; Azam, Iqbal; Awan, Safia; Zafar, Afia

    2014-01-01

    Background Exposure to airborne fungi has been related with exacerbation of asthma in adults and children leading to increased outpatient, emergency room visits, and hospitalizations. Hypersensitivity to these airborne fungi may be an important initial predisposing factor in the development and exacerbation of asthma. Objective This study was conducted to determine an association between fungal types and spore concentrations with the risk of asthma exacerbation in adults. Methods This cross-sectional study was conducted from May 2008 to August 2009 at the Aga Khan University Hospital Karachi, Pakistan. All adult (age≥16 years) patients presenting to the hospital with acute asthma exacerbation were enrolled after informed consent. A home survey was conducted for each patient to assess their environmental characteristics. Indoor air samples were also obtained from the patient's home to determine the type and spore concentration of fungi within the week of their enrollment in the study. Results Three hundred and ninety-one patients with an acute asthma exacerbation were enrolled during the study period. The mean age of participants was 46 years (standard deviation, ±18 years) and 247 (63.2%) were females. A trend of higher asthma enrollment associated with higher Aspergillus concentrations was found in two consecutive summers. A total of nineteen types of fungi were found in air samples. Aspergillus spp. was the most frequently isolated fungus with acute asthma exacerbation. Conclusion An association of higher concentration of indoor Aspergillus spp. with asthma exacerbation in adults was observed in this study. PMID:24809014

  7. Biosynthesis and Characterization of Silver Nanoparticles by Aspergillus Species

    PubMed Central

    Pourshahid, Seyedmohammad; Mehryar, Pouyan; Pakshir, Keyvan; Rahimi, Mohammad Javad; Arabi Monfared, Ali

    2016-01-01

    Currently, researchers turn to natural processes such as using biological microorganisms in order to develop reliable and ecofriendly methods for the synthesis of metallic nanoparticles. In this study, we have investigated extracellular biosynthesis of silver nanoparticles using four Aspergillus species including A. fumigatus, A. clavatus, A. niger, and A. flavus. We have also analyzed nitrate reductase activity in the studied species in order to determine the probable role of this enzyme in the biosynthesis of silver nanoparticles. The formation of silver nanoparticles in the cell filtrates was confirmed by the passage of laser light, change in the color of cell filtrates, absorption peak at 430 nm in UV-Vis spectra, and atomic force microscopy (AFM). There was a logical relationship between the efficiencies of studied Aspergillus species in the production of silver nanoparticles and their nitrate reductase activity. A. fumigatus as the most efficient species showed the highest nitrate reductase activity among the studied species while A. flavus exhibited the lowest capacity in the biosynthesis of silver nanoparticles which was in accord with its low nitrate reductase activity. The present study showed that Aspergillus species had potential for the biosynthesis of silver nanoparticles depending on their nitrate reductase activity.

  8. Pseudallescheria boydii with Aspergillus fumigatus and Aspergillus terreus in a Critically Ill Hematopoietic Stem Cell Recipient with ARDS.

    PubMed

    Lahmer, Tobias; Messer, Marlena; Ehmer, Ursula; Eser, Stefan; Beitz, Analena; Fekecs, Lisa; Schmid, Roland M; Huber, Wolfgang

    2016-04-01

    Pseudallescheria boydii is a fungal organism known to affect immunocompromised patients. This organism is known to cause, in severe cases, invasive infection of various organs such as the central nervous, cardiovascular, and respiratory systems. We report an unusual case of pulmonary P. boydii pneumonia in an immunocompromised critically ill patient with a co-infection of Aspergillus fumigatus and Aspergillus terreus with ARDS. This case highlights the importance of a high index of suspicion for superimposed fungal infections in patients who are critically ill and immunocompromised. Uncommon fungal pathogens should be considered in the differential diagnosis of respiratory failure, especially if diagnostic markers such as galactomannan (from BAL and serum) or 1,3-beta-D-glucan are elevated. Further diagnostic interventions are warranted when insufficient clinical improvement is observed to prevent treatment failure and adverse outcomes. PMID:26455910

  9. Proteomic analysis of temperature dependent extracellular proteins from Aspergillus fumigatus grown under solid-state culture condition.

    PubMed

    Adav, Sunil S; Ravindran, Anita; Sze, Siu Kwan

    2013-06-01

    Fungal species of the genus Aspergillus are filamentous ubiquitous saprophytes that play a major role in lignocellulosic biomass recycling and also are considered as cell factories for the production of organic acids, pharmaceuticals, and industrially important enzymes. Analysis of extracellular secreted biomass degrading enzymes using complex lignocellulosic biomass as a substrate by solid-state fermentation could be a more practical approach to evaluate application of the enzymes for lignocellulosic biorefinery. This study isolated a fungal strain from compost, identified as Aspergillus fumigatus, and further analyzed it for lignocellulolytic enzymes at different temperatures using label free quantitative proteomics. The profile of secretome composition discovered cellulases, hemicellulases, lignin degrading proteins, peptidases and proteases, and transport and hypothetical proteins; while protein abundances and further their hierarchical clustering analysis revealed temperature dependent expression of these enzymes during solid-state fermentation of sawdust. The enzyme activities and protein abundances as determined by exponentially modified protein abundance index (emPAI) indicated the maximum activities at the range of 40-50 °C, demonstrating the thermophilic nature of the isolate A. fumigatus LF9. Characterization of the thermostability of secretome suggested the potential of the isolated fungal strain in the production of thermophilic biomass degrading enzymes for industrial application. PMID:23647126

  10. Determination of Ochratoxin A in apples contaminated with Aspergillus ochraceus by using a microfluidic competitive immunosensor with magnetic nanoparticles.

    PubMed

    Fernández-Baldo, Martín A; Bertolino, Franco A; Fernández, Gastón; Messina, Germán A; Sanz, María I; Raba, Julio

    2011-07-01

    Ochratoxin A (OTA) is a mycotoxin produced by filamentous fungi of the genus Aspergillus and Penicillium that presents carcinogenic, teratogenic and nephrotoxic properties. In this work, we have developed, characterized and applied an immunoassay methodology comprised of magnetic nanoparticles (MNPs) as platform for immobilizing bioactive materials incorporated into a microfluidic system for rapid and sensitive quantification of Ochratoxin A (OTA) in apples (Red Delicious) contaminated with Aspergillus ochraceus. The sensor has the potential for automation and the detection of OTA was carried out using a competitive indirect immunoassay method based on the use of anti-OTA monoclonal antibodies immobilized on 3-aminopropyl-modified MNPs. The total assay time into the microfluidic competitive immunosensor was 16 min, and the calculated detection limit was 0.05 µg kg(-1). Moreover, the intra- and inter-assay coefficients of variation were below 6.5%. The proposed method can be a very promising analytical tool for the determination of OTA in apparently healthy fruits post-harvest and for its application in the agricultural industry.

  11. Comparative mapping of aflatoxin pathway gene clusters in Aspergillus parasiticus and Aspergillus flavus.

    PubMed Central

    Yu, J; Chang, P K; Cary, J W; Wright, M; Bhatnagar, D; Cleveland, T E; Payne, G A; Linz, J E

    1995-01-01

    Aflatoxins are toxic and carcinogenic secondary metabolites produced by the fungi Aspergillus flavus and A. parasiticus. Aflatoxins are synthesized by condensation of acetate units; their synthesis is estimated to involve at least 16 different enzymes. In this study we have shown that at least nine genes involved in the aflatoxin biosynthetic pathway are located within a 60-kb DNA fragment. Four of these genes, nor-1, aflR, ver-1, and omtA (previously named omt-1), have been cloned in A. flavus and A. parasiticus. In addition, five other genes, pksA, uvm8, aad, ord-1, and ord-2 have been recently cloned in A. parasiticus. The pksA, aad, and uvm8 genes exhibit sequence homologies to polyketide synthase, aryl-alcohol dehydrogenase, and fatty acid synthase genes, respectively. The cDNA sequences of ord-1 and ord-2 genes, which may be involved in later steps of aflatoxin biosynthesis, have been determined; the ord-1 gene product exhibits homology to cytochrome P-450-type enzymes. By characterizing the overlapping regions of the DNA inserts in different cosmid and lambda DNA clones, we have determined the order of these aflatoxin pathway genes within this 60-kb DNA region to be pksA, nor-1, uvm8, aflR, aad, ver-1, ord-1, ord-2, and omtA in A. parasiticus and nor-1, aflR, ver-1, ord-1, ord-2, and omtA in A. flavus. The order is related to the order in enzymatic steps required for aflatoxin biosynthesis. The physical distances (in kilobase pairs) and the directions of transcription of these genes have been determined for both aflatoxigenic species. PMID:7793957

  12. Constitutive expression of fluorescent protein by Aspergillus var. niger and Aspergillus carbonarius to monitor fungal colonization in maize plants.

    PubMed

    Palencia, Edwin Rene; Glenn, Anthony Elbie; Hinton, Dorothy Mae; Bacon, Charles Wilson

    2013-09-01

    Aspergillus niger and Aspergillus carbonarius are two species in the Aspergillus section Nigri (black-spored aspergilli) frequently associated with peanut (Arachis hypogea), maize (Zea mays), and other plants as pathogens. These infections are symptomless and as such are major concerns since some black aspergilli produce important mycotoxins, ochratoxins A, and the fumonisins. To facilitate the study of the black aspergilli-maize interactions with maize during the early stages of infections, we developed a method that used the enhanced yellow fluorescent protein (eYFP) and the monomeric red fluorescent protein (mRFP1) to transform A. niger and A. carbonarius, respectively. The results were constitutive expressions of the fluorescent genes that were stable in the cytoplasms of hyphae and conidia under natural environmental conditions. The hyphal in planta distribution in 21-day-old seedlings of maize were similar wild type and transformants of A. niger and A. carbonarius. The in planta studies indicated that both wild type and transformants internally colonized leaf, stem and root tissues of maize seedlings, without any visible disease symptoms. Yellow and red fluorescent strains were capable of invading epidermal cells of maize roots intercellularly within the first 3 days after inoculation, but intracellular hyphal growth was more evident after 7 days of inoculation. We also tested the capacity of fluorescent transformants to produce ochratoxin A and the results with A. carbonarius showed that this transgenic strain produced similar concentrations of this secondary metabolite. This is the first report on the in planta expression of fluorescent proteins that should be useful to study the internal plant colonization patterns of two ochratoxigenic species in the Aspergillus section Nigri.

  13. Aspergillus steynii and Aspergillus westerdijkiae as potential risk of OTA contamination in food products in warm climates.

    PubMed

    Gil-Serna, Jessica; Patiño, Belén; Cortes, Laura; Gonzalez-Jaen, Maria Teresa; Vazquez, Covadonga

    2015-04-01

    Aspergillus steynii and Aspergillus westerdijkiae are the main ochratoxin A (OTA) producing species of Aspergillus section Circumdati. Due to its recent description, few data are available about the influence of ecophysiological factors on their growth and OTA production profiles. In this work, the effect of temperature (20, 24 and 28 °C) and water activity (aw) (0.928, 0.964 and 0.995) on growth, sporulation and OTA production by these fungi was examined in CYA and media prepared from paprika, green coffee, anise, grapes, maize and barley. Growth was positively affected by the highest temperature and aw values indicating that both species might be expected in warm climates or storage conditions. However, optimal growth conditions showed differences depending on the medium. OTA production was markedly affected by substrate and showed qualitative and quantitative differences. Both species, especially A. steynii, represent a great potential risk of OTA contamination due to their high production in a variety of conditions and substrates, in particular in barley and paprika-based media. Additionally, neither growth nor sporulation did result good indicators of OTA production by A. steynii or A. westerdijkiae; therefore, specific and highly-sensitive detection methods become essential tools for control strategies to reduce OTA risk by these species. PMID:25475281

  14. Decontamination of Aspergillus flavus and Aspergillus parasiticus spores on hazelnuts via atmospheric pressure fluidized bed plasma reactor.

    PubMed

    Dasan, Beyhan Gunaydin; Mutlu, Mehmet; Boyaci, Ismail Hakki

    2016-01-01

    In this study, an atmospheric pressure fluidized bed plasma (APFBP) system was designed and its decontamination effect on aflatoxigenic fungi (Aspergillus flavus and Aspergillus parasiticus) on the surface of hazelnuts was investigated. Hazelnuts were artificially contaminated with A. flavus and A. parasiticus and then were treated with dry air plasma for up to 5min in the APFBP system at various plasma parameters. Significant reductions of 4.50 log (cfu/g) in A. flavus and 4.19 log (cfu/g) in A. parasiticus were achieved after 5min treatments at 100% V - 25kHz (655W) by using dry air as the plasma forming gas. The decontamination effect of APFBP on A. flavus and A. parasiticus spores inoculated on hazelnuts was increased with the applied reference voltage and the frequency. No change or slight reductions were observed in A. flavus and A. parasiticus load during the storage of plasma treated hazelnuts whereas on the control samples fungi continued to grow under storage conditions (30days at 25°C). Temperature change on hazelnut surfaces in the range between 35 and 90°C was monitored with a thermal camera, and it was demonstrated that the temperature increase taking place during plasma treatment did not have a lethal effect on A. flavus and A. parasiticus spores. The damage caused by APFBP treatment on Aspergillus spp. spores was also observed by scanning electron microscopy.

  15. Overexpression of Aspergillus tubingensis faeA in protease-deficient Aspergillus niger enables ferulic acid production from plant material.

    PubMed

    Zwane, Eunice N; Rose, Shaunita H; van Zyl, Willem H; Rumbold, Karl; Viljoen-Bloom, Marinda

    2014-06-01

    The production of ferulic acid esterase involved in the release of ferulic acid side groups from xylan was investigated in strains of Aspergillus tubingensis, Aspergillus carneus, Aspergillus niger and Rhizopus oryzae. The highest activity on triticale bran as sole carbon source was observed with the A. tubingensis T8.4 strain, which produced a type A ferulic acid esterase active against methyl p-coumarate, methyl ferulate and methyl sinapate. The activity of the A. tubingensis ferulic acid esterase (AtFAEA) was inhibited twofold by glucose and induced twofold in the presence of maize bran. An initial accumulation of endoglucanase was followed by the production of endoxylanase, suggesting a combined action with ferulic acid esterase on maize bran. A genomic copy of the A. tubingensis faeA gene was cloned and expressed in A. niger D15#26 under the control of the A. niger gpd promoter. The recombinant strain has reduced protease activity and does not acidify the media, therefore promoting high-level expression of recombinant enzymes. It produced 13.5 U/ml FAEA after 5 days on autoclaved maize bran as sole carbon source, which was threefold higher than for the A. tubingensis donor strain. The recombinant AtFAEA was able to extract 50 % of the available ferulic acid from non-pretreated maize bran, making this enzyme suitable for the biological production of ferulic acid from lignocellulosic plant material. PMID:24664515

  16. Aspergillus steynii and Aspergillus westerdijkiae as potential risk of OTA contamination in food products in warm climates.

    PubMed

    Gil-Serna, Jessica; Patiño, Belén; Cortes, Laura; Gonzalez-Jaen, Maria Teresa; Vazquez, Covadonga

    2015-04-01

    Aspergillus steynii and Aspergillus westerdijkiae are the main ochratoxin A (OTA) producing species of Aspergillus section Circumdati. Due to its recent description, few data are available about the influence of ecophysiological factors on their growth and OTA production profiles. In this work, the effect of temperature (20, 24 and 28 °C) and water activity (aw) (0.928, 0.964 and 0.995) on growth, sporulation and OTA production by these fungi was examined in CYA and media prepared from paprika, green coffee, anise, grapes, maize and barley. Growth was positively affected by the highest temperature and aw values indicating that both species might be expected in warm climates or storage conditions. However, optimal growth conditions showed differences depending on the medium. OTA production was markedly affected by substrate and showed qualitative and quantitative differences. Both species, especially A. steynii, represent a great potential risk of OTA contamination due to their high production in a variety of conditions and substrates, in particular in barley and paprika-based media. Additionally, neither growth nor sporulation did result good indicators of OTA production by A. steynii or A. westerdijkiae; therefore, specific and highly-sensitive detection methods become essential tools for control strategies to reduce OTA risk by these species.

  17. Overexpression of Aspergillus tubingensis faeA in protease-deficient Aspergillus niger enables ferulic acid production from plant material.

    PubMed

    Zwane, Eunice N; Rose, Shaunita H; van Zyl, Willem H; Rumbold, Karl; Viljoen-Bloom, Marinda

    2014-06-01

    The production of ferulic acid esterase involved in the release of ferulic acid side groups from xylan was investigated in strains of Aspergillus tubingensis, Aspergillus carneus, Aspergillus niger and Rhizopus oryzae. The highest activity on triticale bran as sole carbon source was observed with the A. tubingensis T8.4 strain, which produced a type A ferulic acid esterase active against methyl p-coumarate, methyl ferulate and methyl sinapate. The activity of the A. tubingensis ferulic acid esterase (AtFAEA) was inhibited twofold by glucose and induced twofold in the presence of maize bran. An initial accumulation of endoglucanase was followed by the production of endoxylanase, suggesting a combined action with ferulic acid esterase on maize bran. A genomic copy of the A. tubingensis faeA gene was cloned and expressed in A. niger D15#26 under the control of the A. niger gpd promoter. The recombinant strain has reduced protease activity and does not acidify the media, therefore promoting high-level expression of recombinant enzymes. It produced 13.5 U/ml FAEA after 5 days on autoclaved maize bran as sole carbon source, which was threefold higher than for the A. tubingensis donor strain. The recombinant AtFAEA was able to extract 50 % of the available ferulic acid from non-pretreated maize bran, making this enzyme suitable for the biological production of ferulic acid from lignocellulosic plant material.

  18. Morphological and molecular identification of filamentous Aspergillus flavus and Aspergillus parasiticus isolated from compound feeds in South Africa.

    PubMed

    Iheanacho, Henry E; Njobeh, Patrick B; Dutton, Francis M; Steenkamp, Paul A; Steenkamp, Lucia; Mthombeni, Julian Q; Daru, Barnabas H; Makun, Anthony H

    2014-12-01

    Isolation of filamentous species of two Aspergillum genera from compound feeds produced in South Africa, and subsequent extraction of their individual DNA in this study, presents a simple but rapid molecular procedure for high through-put analysis of the individual morphological forms. DNA was successfully isolated from the Aspergillus spp. from agar cultures by use of a commercial kit. Agarose gel electrophoresis fractionation of the fungi DNA, showed distinct bands. The DNA extracted by this procedure appears to be relatively pure with a ratio absorbance at 260 and 280 nm. However, the overall morphological and molecular data indicated that 67.5 and 51.1% of feed samples were found to be contaminated with Aspergillus flavus and Aspergillus parasiticus, respectively, with poultry feed having the highest contamination mean level of 5.7 × 105 CFU/g when compared to cattle (mean: 4.0 × 106 CFU/g), pig (mean: 2.7 × 104 CFU/g) and horse (1.0 × 102 CFU) feed. This technique presents a readily achievable, easy to use method in the extraction of filamentous fungal DNA and it's identification. Hence serves as an important tool towards molecular study of these organisms for routine analysis check in monitoring and improving compound feed quality against fungal contamination.

  19. Decontamination of Aspergillus flavus and Aspergillus parasiticus spores on hazelnuts via atmospheric pressure fluidized bed plasma reactor.

    PubMed

    Dasan, Beyhan Gunaydin; Mutlu, Mehmet; Boyaci, Ismail Hakki

    2016-01-01

    In this study, an atmospheric pressure fluidized bed plasma (APFBP) system was designed and its decontamination effect on aflatoxigenic fungi (Aspergillus flavus and Aspergillus parasiticus) on the surface of hazelnuts was investigated. Hazelnuts were artificially contaminated with A. flavus and A. parasiticus and then were treated with dry air plasma for up to 5min in the APFBP system at various plasma parameters. Significant reductions of 4.50 log (cfu/g) in A. flavus and 4.19 log (cfu/g) in A. parasiticus were achieved after 5min treatments at 100% V - 25kHz (655W) by using dry air as the plasma forming gas. The decontamination effect of APFBP on A. flavus and A. parasiticus spores inoculated on hazelnuts was increased with the applied reference voltage and the frequency. No change or slight reductions were observed in A. flavus and A. parasiticus load during the storage of plasma treated hazelnuts whereas on the control samples fungi continued to grow under storage conditions (30days at 25°C). Temperature change on hazelnut surfaces in the range between 35 and 90°C was monitored with a thermal camera, and it was demonstrated that the temperature increase taking place during plasma treatment did not have a lethal effect on A. flavus and A. parasiticus spores. The damage caused by APFBP treatment on Aspergillus spp. spores was also observed by scanning electron microscopy. PMID:26398284

  20. Nonfunctionality of Aspergillus sojae aflR in a strain of Aspergillus parasiticus with a disrupted aflR gene.

    PubMed

    Takahashi, Tadashi; Chang, Perng-Kuang; Matsushima, Kenichiro; Yu, Jiujiang; Abe, Keietsu; Bhatnagar, Deepak; Cleveland, Thomas E; Koyama, Yasuji

    2002-08-01

    Aspergillus sojae belongs to the Aspergillus section Flavi but does not produce aflatoxins. The functionality of the A. sojae aflR gene (aflRs) was examined by transforming it into an DeltaaflR strain of A. parasiticus, derived from a nitrate-nonutilizing, versicolorin A (VERA)-accumulating strain. The A. parasiticus aflR gene (aflRp) transformants produced VERA, but the aflRs transformants did not. Even when aflRs was placed under the control of the amylase gene (amyB) promoter of Aspergillus oryzae, the amy(p)::aflRs transformants did not produce VERA. A chimeric construct containing the aflRs promoter plus the aflRs N- and aflRp C-terminal coding regions could restore VERA production, but a construct containing the aflRp promoter plus the aflRp N- and aflRs C-terminal coding regions could not. These results show that the A. sojae aflR promoter is functional in A. parasiticus and that the HAHA motif does not affect the function of the resulting hybrid AflR. We conclude that the lack of aflatoxin production by A. sojae can be attributed, at least partially, to the premature termination defect in aflRs, which deletes the C-terminal transcription activation domain that is critical for the expression of aflatoxin biosynthetic genes.