The Solvent Dimethyl Sulfoxide Affects Physiology, Transcriptome and Secondary Metabolism of Aspergillus flavus

Dimethyl sulfoxide (DSMO) is a simple molecule widely used because of its great solvating ability, but this solvent also has little-known biological effects, especially on fungi. Aspergillus flavus is a notorious pathogenic fungus which may contaminate a large variety of crops worldwide by producing aflatoxins, endangering at the same time food safety and international trade. The aim of this study was to characterize the effect of DMSO on A. flavus including developmental parameters such as germination and sporulation, as well as its transcriptome profile using high-throughput RNA-sequencing assay and its impact on secondary metabolism (SM). After DMSO exposure, A. flavus displayed depigmented conidia in a dose-dependent manner. The four-day exposition of cultures to two doses of DMSO, chosen on the basis of depigmentation intensity (35 mM “low” and 282 mM “high”), led to no significant impact on fungal growth, germination or sporulation. However, transcriptomic data analysis showed that 4891 genes were differentially regulated in response to DMSO (46% of studied transcripts). A total of 4650 genes were specifically regulated in response to the highest dose of DMSO, while only 19 genes were modulated upon exposure to the lowest dose. Secondary metabolites clusters genes were widely affected by the DMSO, with 91% of clusters impacted at the highest dose. Among these, aflatoxins, cyclopiazonic acid and ustiloxin B clusters were totally under-expressed. The genes belonging to the AFB1 cluster were the most negatively modulated ones, the two doses leading to 63% and 100% inhibition of the AFB1 production, respectively. The SM analysis also showed the disappearance of ustiloxin B and a 10-fold reduction of cyclopiazonic acid level when A. flavus was treated by the higher DMSO dose. In conclusion, the present study showed that DMSO impacted widely A. flavus’ transcriptome, including secondary metabolism gene clusters with the aflatoxins at the head of down-regulated ones. The solvent also inhibits conidial pigmentation, which could illustrate common regulatory mechanisms between aflatoxins and fungal pigment pathways. Because of its effect on major metabolites synthesis, DMSO should not be used as solvent especially in studies testing anti-aflatoxinogenic compounds.

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The 14-3-3 Protein Homolog ArtA Regulates Development and Secondary Metabolism in the Opportunistic Plant PathogenAspergillus flavus

Applied and Environmental Microbiology ◽

10.1128/aem.02241-17 ◽

2017 ◽

Vol 84 (5) ◽

Cited By ~ 4

Author(s):

Beatriz A. Ibarra ◽

Jessica M. Lohmar ◽

Timothy Satterlee ◽

Taylor McDonald ◽

Jeffrey W. Cary ◽

...

Keyword(s):

Secondary Metabolism ◽

Aspergillus Flavus ◽

Pathogenic Fungus ◽

Cyclopiazonic Acid ◽

Gene Clusters ◽

Opportunistic Pathogen ◽

Aflatoxin Contamination ◽

Morphological Development ◽

Dependent Manner ◽

Content Type

ABSTRACTThe opportunistic plant-pathogenic fungusAspergillus flavusproduces carcinogenic mycotoxins termed aflatoxins (AF). Aflatoxin contamination of agriculturally important crops, such as maize, peanut, sorghum, and tree nuts, is responsible for serious adverse health and economic impacts worldwide. In order to identify possible genetic targets to reduce AF contamination, we have characterized theartAgene, encoding a putative 14-3-3 homolog inA. flavus. TheartAdeletion mutant presents a slight decrease in vegetative growth and alterations in morphological development and secondary metabolism. Specifically,artAaffects conidiation, and this effect is influenced by the type of substrate and culture condition. In addition, normal levels ofartAare required for sclerotial development. Importantly,artAnegatively regulates AF production as well as the concomitant expression of genes in the AF gene cluster. An increase in AF is also observed in seeds infected with theA. flavusstrain lackingartA. Furthermore, the expression of other secondary metabolite genes is alsoartAdependent, including genes in the cyclopiazonic acid (CPA) and ustiloxin gene clusters, in this agriculturally important fungus.IMPORTANCEIn the current study,artA, which encodes a 14-3-3 homolog, was characterized in the agriculturally and medically important fungusAspergillus flavus, specifically, its possible role governing sporulation, formation of resistant structures, and secondary metabolism. The highly conservedartAis necessary for normal fungal morphogenesis in an environment-dependent manner, affecting the balance between production of conidiophores and the formation of resistant structures that are necessary for the dissemination and survival of this opportunistic pathogen. This study reports a 14-3-3 protein affecting secondary metabolism in filamentous fungi. Importantly,artAregulates the biosynthesis of the potent carcinogenic compound aflatoxin B1 (AFB1) as well as the production of other secondary metabolites.

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The kinetochore protein Spc105, a novel interaction partner of LaeA, regulates development and secondary metabolism in Aspergillus flavus

10.1101/487280 ◽

2018 ◽

Author(s):

QingQing Zhi ◽

Lei He ◽

JieYing Li ◽

Jing Li ◽

ZhenLong Wang ◽

...

Keyword(s):

Secondary Metabolism ◽

Aspergillus Flavus ◽

Leucine Zipper ◽

Cyclopiazonic Acid ◽

Gene Clusters ◽

Interaction Partner ◽

Kinetochore Protein ◽

Metabolism Regulation ◽

Fungal Secondary Metabolism

Nuclear protein LaeA is known as the global regulator of secondary metabolism in Aspergillus. LaeA connects with VeA and VelB to form a heterotrimeric complex, which coordinates fungal development and secondary metabolism. Here, we describe a new interaction partner of LaeA, the kinetochore protein Spc105, from the aflatoxin-producing fungus Aspergillus flavus. We showed that in addition to involvement in nuclear division, Spc105 is required for normal conidiophore development and sclerotia production of A. ﬂavus. Moreover, Spc105 positively regulates the production of secondary metabolites such as aflatoxin and kojic acid, and negatively regulates the production of cyclopiazonic acid. Transcriptome analysis of the ?spc105 strain revealed that 23 backbone genes were differentially expressed, corresponding to 19 of the predicted 56 secondary metabolite gene clusters, suggesting a broad regulatory role of Spc105 in secondary metabolism. Notably, the reduced expression of laeA in our transcriptome data led to the discovery of the correlation between Spc105 and LaeA, and double mutant analysis indicated a functional interdependence between Spc105 and LaeA. Further, yeast two-hybrid (Y2H) and GST pull-down assays revealed that Spc105 interacts directly with the S-adenosylmethionine (SAM)-binding domain of LaeA, and that the leucine zipper motif in Spc105 is required for this interaction. The Spc105-LaeA interaction identified in our study indicates a cooperative interplay of distinct regulators in A. flavus, providing new insights into fungal secondary metabolism regulation networks.

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Transcriptome Analysis of Aspergillus flavus RevealsveA-Dependent Regulation of Secondary Metabolite Gene Clusters, Including the Novel Aflavarin Cluster

Eukaryotic Cell ◽

10.1128/ec.00092-15 ◽

2015 ◽

Vol 14 (10) ◽

pp. 983-997 ◽

Cited By ~ 44

Author(s):

J. W. Cary ◽

Z. Han ◽

Y. Yin ◽

J. M. Lohmar ◽

S. Shantappa ◽

...

Keyword(s):

Secondary Metabolism ◽

Secondary Metabolite ◽

Aspergillus Flavus ◽

Gene Clusters ◽

Fungal Species ◽

The Novel ◽

Content Type ◽

Expression Of Genes ◽

Number Of Genes ◽

Fungal Secondary Metabolism

ABSTRACTThe global regulatoryveAgene governs development and secondary metabolism in numerous fungal species, includingAspergillus flavus. This is especially relevant sinceA. flavusinfects crops of agricultural importance worldwide, contaminating them with potent mycotoxins. The most well-known are aflatoxins, which are cytotoxic and carcinogenic polyketide compounds. The production of aflatoxins and the expression of genes implicated in the production of these mycotoxins areveAdependent. The genes responsible for the synthesis of aflatoxins are clustered, a signature common for genes involved in fungal secondary metabolism. Studies of theA. flavusgenome revealed many gene clusters possibly connected to the synthesis of secondary metabolites. Many of these metabolites are still unknown, or the association between a known metabolite and a particular gene cluster has not yet been established. In the present transcriptome study, we show thatveAis necessary for the expression of a large number of genes. Twenty-eight out of the predicted 56 secondary metabolite gene clusters include at least one gene that is differentially expressed depending on presence or absence ofveA. One of the clusters under the influence ofveAis cluster 39. The absence ofveAresults in a downregulation of the five genes found within this cluster. Interestingly, our results indicate that the cluster is expressed mainly in sclerotia. Chemical analysis of sclerotial extracts revealed that cluster 39 is responsible for the production of aflavarin.

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Microevolution in the pansecondary metabolome of Aspergillus flavus and its potential macroevolutionary implications for filamentous fungi

Proceedings of the National Academy of Sciences ◽

10.1073/pnas.2021683118 ◽

2021 ◽

Vol 118 (21) ◽

pp. e2021683118

Author(s):

Milton T. Drott ◽

Tomás A. Rush ◽

Tatum R. Satterlee ◽

Richard J. Giannone ◽

Paul E. Abraham ◽

...

Keyword(s):

Secondary Metabolism ◽

Aspergillus Flavus ◽

High Performance ◽

High Resolution Mass Spectrometry ◽

Population Level ◽

Gene Clusters ◽

The United States ◽

Ecological Interactions ◽

Different Populations ◽

Fungal Secondary Metabolism

Fungi produce a wealth of pharmacologically bioactive secondary metabolites (SMs) from biosynthetic gene clusters (BGCs). It is common practice for drug discovery efforts to treat species’ secondary metabolomes as being well represented by a single or a small number of representative genomes. However, this approach misses the possibility that intraspecific population dynamics, such as adaptation to environmental conditions or local microbiomes, may harbor novel BGCs that contribute to the overall niche breadth of species. Using 94 isolates of Aspergillus flavus, a cosmopolitan model fungus, sampled from seven states in the United States, we dereplicate 7,821 BGCs into 92 unique BGCs. We find that more than 25% of pangenomic BGCs show population-specific patterns of presence/absence or protein divergence. Population-specific BGCs make up most of the accessory-genome BGCs, suggesting that different ecological forces that maintain accessory genomes may be partially mediated by population-specific differences in secondary metabolism. We use ultra-high-performance high-resolution mass spectrometry to confirm that these genetic differences in BGCs also result in chemotypic differences in SM production in different populations, which could mediate ecological interactions and be acted on by selection. Thus, our results suggest a paradigm shift that previously unrealized population-level reservoirs of SM diversity may be of significant evolutionary, ecological, and pharmacological importance. Last, we find that several population-specific BGCs from A. flavus are present in Aspergillus parasiticus and Aspergillus minisclerotigenes and discuss how the microevolutionary patterns we uncover inform macroevolutionary inferences and help to align fungal secondary metabolism with existing evolutionary theory.

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Inhibition of Aspergillus flavus growth and aflatoxin B1 production by natamycin

World Mycotoxin Journal ◽

10.3920/wmj2020.2620 ◽

2021 ◽

pp. 1-16

Author(s):

P. Chang ◽

B. Tai ◽

M. Zheng ◽

Q. Yang ◽

F. Xing

Keyword(s):

Aspergillus Flavus ◽

Animal Health ◽

Fungal Growth ◽

Outer Wall ◽

Aflatoxin Production ◽

Ergosterol Biosynthesis ◽

Dependent Manner ◽

Conidia Germination ◽

Streptomyces Spp ◽

Inhibitory Mechanisms

Aspergillus flavus causes huge crop losses, reduces crop quality and has adverse effects on human and animal health. A large amount of food contaminated with aflatoxin can greatly increase the risk of liver cancer. Therefore, prevention and control of aflatoxin production have aroused attention of research in various countries. Natamycin extracted from Streptomyces spp. has been widely used in production practice due to its good specificity and safety. Here, we found that natamycin could significantly inhibit fungal growth, conidia germination, ergosterol and AFB1 production by A. flavus in a dose-dependent manner. Scanning electron microscope analysis indicated that the number of conidia was decreased, the outer wall of conidia was destroyed, and the mycelia were shrivelled and tangled by natamycin. RNA-Seq data indicated that natamycin inhibited fungal growth and conidia development of A. flavus by significantly down-regulating some genes involved in ergosterol biosynthesis, such as Erg13, HMG1 and HMG2. It inhibited conidia germination by significantly down-regulating some genes related to conidia development, such as FluG and VosA. After natamycin exposure, the decreased ratio of aflS/aflR caused by the down-regulation of all the structural genes, which subsequently resulted in the suppression of AFB1 production. In conclusion, this study served to reveal the inhibitory mechanisms of natamycin on fungal growth and AFB1 biosynthesis in A. flavus and to provide solid evidence for its application in controlling AFB1 contamination.

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Caryolan-1-ol, an antifungal volatile produced by Streptomyces spp., inhibits the endomembrane system of fungi

Open Biology ◽

10.1098/rsob.170075 ◽

2017 ◽

Vol 7 (7) ◽

pp. 170075 ◽

Cited By ~ 12

Author(s):

Gyeongjun Cho ◽

Junheon Kim ◽

Chung Gyoo Park ◽

Corey Nislow ◽

David M. Weller ◽

...

Keyword(s):

Fungal Growth ◽

Gene Clusters ◽

Terpene Synthase ◽

Dependent Manner ◽

Endomembrane System ◽

Chemical Genomics ◽

Streptomyces Spp ◽

Antifungal Volatiles ◽

Dose Dependent ◽

Volatile Terpene

Streptomyces spp. have the ability to produce a wide variety of secondary metabolites that interact with the environment. This study aimed to discover antifungal volatiles from the genus Streptomyces and to determine the mechanisms of inhibition. Volatiles identified from Streptomyces spp. included three major terpenes, geosmin, caryolan-1-ol and an unknown sesquiterpene. antiSMASH and KEGG predicted that the volatile terpene synthase gene clusters occur in the Streptomyces genome. Growth inhibition was observed when fungi were exposed to the volatiles. Biological activity of caryolan-1-ol has previously not been investigated. Fungal growth was inhibited in a dose-dependent manner by a mixture of the main volatiles, caryolan-1-ol and the unknown sesquiterpene, from Streptomyces sp. S4–7. Furthermore, synthesized caryolan-1-ol showed similar antifungal activity. Results of chemical-genomics profiling assays showed that caryolan-1-ol affected the endomembrane system by disrupting sphingolipid synthesis and normal vesicle trafficking in the fungi.

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Antifungal activity and inhibition of aflatoxins production by Zingiber officinale Roscoe essential oil against Aspergillus flavus in stored maize grains

Ciência Rural ◽

10.1590/0103-8478cr20190779 ◽

2020 ◽

Vol 50 (6) ◽

Author(s):

Samuel Botião Nerilo ◽

Jéssica Cristina Zoratto Romoli ◽

Lydiana Polis Nakasugi ◽

Natana Souza Zampieri ◽

Simone Aparecida Galerani Mossini ◽

...

Keyword(s):

Essential Oil ◽

Aspergillus Flavus ◽

Fungal Growth ◽

Zingiber Officinale ◽

Aflatoxin Production ◽

Gas Chromatography Mass Spectrometry ◽

Dependent Manner ◽

Zingiber Officinale Roscoe ◽

Dose Dependent

ABSTRACT: Essential oils are possible alternatives to the use of synthetic pesticides for control of fungal contamination. Ginger (Zingiber officinale) essential oil (GEO) is known for having antifungal and antiaflatoxigenic properties, but its use as a fumigant in situ has not been studied yet. The aim of this study was to evaluate GEO’s effects upon Aspergillus flavus as a fumigant agent in stored maize grains. The main compounds reported in GEO were α-zingiberene (23.85%) and geranial (14.16%), characterized by gas chromatography-mass spectrometry and nuclear magnetic resonance. The GEO was used as a fumigant in irradiated maize grains in concentrations ranging from 5 to 50 µg/g and the resulting effects were compared to a synthetic antifungal agent (carbendazim and thiram), an antifungal traditionally used for seed treatment. The antifungal efficacy of GEO against A. flavus has been proven in a dose-dependent manner through in situ (maize grains) test. The GEO inhibited aflatoxin production at concentrations 25 and 50 µg/g and controlled fungal growth. Therefore, GEO can be used as an effective and non-toxic alternative to conventional treatments in stored maize grains for the natural control of A. flavus.

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The genome of the butternut canker pathogen, Ophiognomonia clavigignenti-juglandacearum shows an elevated number of genes associated with secondary metabolism and protection from host resistance responses

PeerJ ◽

10.7717/peerj.9265 ◽

2020 ◽

Vol 8 ◽

pp. e9265

Author(s):

Guangxi Wu ◽

Taruna A. Schuelke ◽

Gloria Iriarte ◽

Kirk Broders

Keyword(s):

Secondary Metabolism ◽

Pathogenic Fungus ◽

Gene Families ◽

Gene Clusters ◽

Large Numbers ◽

Genome Wide ◽

A Genome ◽

Number Of Genes ◽

Hardwood Tree ◽

Branch Dieback

Ophiognomonia clavigignenti-juglandacearum (Oc-j) is a plant pathogenic fungus that causes canker and branch dieback diseases in the hardwood tree butternut, Juglans cinerea. Oc-j is a member of the order of Diaporthales, which includes many other plant pathogenic species, several of which also infect hardwood tree species. In this study, we sequenced the genome of Oc-j and achieved a high-quality assembly and delineated its phylogeny within the Diaporthales order using a genome-wide multi-gene approach. We also further examined multiple gene families that might be involved in plant pathogenicity and degradation of complex biomass, which are relevant to a pathogenic life-style in a tree host. We found that the Oc-j genome contains a greater number of genes in these gene families compared to other species in the Diaporthales. These gene families include secreted CAZymes, kinases, cytochrome P450, efflux pumps, and secondary metabolism gene clusters. The large numbers of these genes provide Oc-j with an arsenal to cope with the specific ecological niche as a pathogen of the butternut tree.

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The Inhibitory Effects ofCurcuma longaL. Essential Oil and Curcumin onAspergillus flavusLink Growth and Morphology

The Scientific World JOURNAL ◽

10.1155/2013/343804 ◽

2013 ◽

Vol 2013 ◽

pp. 1-6 ◽

Cited By ~ 20

Author(s):

Flávio Dias Ferreira ◽

Simone Aparecida Galerani Mossini ◽

Francine Maery Dias Ferreira ◽

Carla Cristina Arrotéia ◽

Christiane Luciana da Costa ◽

...

Keyword(s):

Essential Oil ◽

Aspergillus Flavus ◽

Culture Media ◽

Curcuma Longa ◽

Fungal Growth ◽

Aflatoxin Production ◽

Dependent Manner ◽

Inhibitory Effects ◽

Concentration Dependent Manner ◽

Spore Viability

The essential oil fromCurcuma longaL. was analysed by GC/MS. The major components of the oil were ar-turmerone (33.2%),α-turmerone (23.5%) andβ-turmerone (22.7%). The antifungal activities of the oil were studied with regard toAspergillus flavusgrowth inhibition and altered morphology, as preliminary studies indicated that the essential oil fromC. longainhibitedAspergillus flavusLink aflatoxin production. The concentration of essential oil in the culture media ranged from 0.01% to 5.0% v/v, and the concentration of curcumin was 0.01–0.5% v/v. The effects on sporulation, spore viability, and fungal morphology were determined. The essential oil exhibited stronger antifungal activity than curcumin onA. flavus. The essential oil reduced the fungal growth in a concentration-dependent manner.A. flavusgrowth rate was reduced byC. longaessential oil at 0.10%, and this inhibition effect was more efficient in concentrations above 0.50%. Germination and sporulation were 100% inhibited in 0.5% oil. Scanning electron microscopy (SEM) ofA. flavusexposed to oil showed damage to hyphae membranes and conidiophores. Because the fungus is a plant pathogen and aflatoxin producer,C. longaessential oil may be used in the management of host plants.

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The genome of the butternut canker pathogen, Ophiognomonia clavigignenti-juglandacearum shows an elevated number of genes associated with secondary metabolism and protection from host resistance responses in comparison with other members of the Diaporthales

10.1101/820977 ◽

2019 ◽

Author(s):

Guangxi Wu ◽

Taruna A. Schuelke ◽

Kirk Broders

Keyword(s):

Secondary Metabolism ◽

Pathogenic Fungus ◽

Gene Families ◽

Gene Clusters ◽

Large Numbers ◽

Genome Wide ◽

A Genome ◽

Number Of Genes ◽

Hardwood Tree ◽

Branch Dieback

AbstractOphiognomonia clavigignentijuglandacearum (Oc-j) is a plant pathogenic fungus that causes canker and branch dieback diseases in the hardwood tree butternut, Juglans cinerea. Oc-j is a member of the order of Diaporthales, which includes many other plant pathogenic species, several of which also infect hardwood tree species. In this study, we sequenced the genome of Oc-j and achieved a high-quality assembly and delineated the phylogeny of Oc-j within the Diaporthales order using a genome-wide multi-gene approach. We also further examined multiple gene families that might be involved in plant pathogenicity and degradation of complex biomass, which are relevant to a pathogenic life-style in a tree host. We found that the Oc-j genome contains a greater number of genes in these gene families compared to other species in Diaporthales. These gene families include secreted CAZymes, kinases, cytochrome P450, efflux pumps, and secondary metabolism gene clusters. The large numbers of these genes provide Oc-j with an arsenal to cope with the specific ecological niche as a pathogen of the butternut tree.

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