Unknown,Transcriptomics,Genomics,Proteomics

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Genotypic and Phenotypic Versatility of Aspergillus flavus during Maize Exploitation

ABSTRACT: Several are the inputs which are able to modulate mycotoxin synthesis. In particular, when a fungus receives an external stimulus reacts by activating, through a quite well-defined signal cascade, an evident switch in its lifestyle. This profound change is also due to the activation of global gene regulators and, in particular, of transcription factors able to switch on the mycotoxin gene clusters expression. Aflatoxins (AF) are harmful carcinogenic compounds produced mainly by Aspergillus flavus and A. parasiticus. AF are produced during the contamination of maize kernels into the field, even if their role in phyto-toxicity is not yet assessed. Nevertheless, AF biosynthesis is tightly regulated by host-derived signals. Recently, the nature of some of these signals have been elucidated. In particular, a role in susceptibility and resistance of maize to A. flavus contamination has been assigned to some plant oxylipins. These findings draw a scenario in which a complex interplay is under way between A. flavus and maize. For uncovering all the implications of this cross-talk we decide to follow a holistic approach. In particular, we designed experimental conditions aimed to mimic the different phases of A. flavus infection cycle on maize and then by performing a microarray analysis on the harvested mycelia. The microarray data set has been processed for performing the differential expression analysis of almost 14000 gene probes, the pathway analysis based on the gene ontology and inter pro annotations, the secondary metabolites cluster co-expression analysis and an identification of groups of co-expressed neighbor genes, possibly associated with production of secondary metabolites. Analysis of 12 microarrays monitoring gene expression of Aspergillus flavus over various growth conditions

ORGANISM(S): Aspergillus flavus

SUBMITTER: Paolo Uva

PROVIDER: E-GEOD-28163 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Genotypic and phenotypic versatility of Aspergillus flavus during maize exploitation.

Reverberi Massimo M Punelli Marta M Scala Valeria V Scarpari Marzia M Uva Paolo P Mentzen Wieslawa I WI Dolezal Andrea L AL Woloshuk Charles C Pinzari Flavia F Fabbri Anna A AA Fanelli Corrado C Payne Gary A GA

PloS one 20130719 7

Aspergillus flavus is a cosmopolitan fungus able to respond to external stimuli and to shift both its trophic behaviour and the production of secondary metabolites, including that of the carcinogen aflatoxin (AF). To better understand the adaptability of this fungus, we examined genetic and phenotypic responses within the fungus when grown under four conditions that mimic different ecological niches ranging from saprophytic growth to parasitism. Global transcription changes were observed in both ...[more]

PMID: 23894339

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Project description:Aspergillus flavus is a common saprophyte and opportunistic pathogen producing aflatoxin (AF) and many other secondary metabolites. 5-Azacytidine (5-AC), a derivative of nucleoside cytidine, is widely used for studies in epigenetics and cancer biology as an inactivator of DNA methyltransferase and is also used for studying secondary metabolism in fungi. Our previous studies showed that 5-AC affects development and inhibits AF production in A. flavus, and that A. flavus lacks DNA methylation. How this common DNA methyltransferase inhibitor affects development and AF production is not clear. In this study, we applied an RNA-Seq approach to elucidate the mechanism of 5-ACM-bM-^@M-^Ys effect on A. flavus. In our current study, we identified 240 significantly differently expressed (Q-value<0.05) genes after 5-AC treatment, including two backbone genes in secondary metabolite clusters #27 and #35, which are involved in development or survival of sclerotia. With 5-AC treatment, about three quarters of the genes in the AF biosynthetic gene cluster in A. flavus were down-regulated to a certain degree. Strikingly, at least two genes aflI and aflLa, were completely inhibited. Interestingly, several genes involved in fungal development were down-regulated, especially veA, which is a gene that encodes protein bridges VelB and LaeA. This result supports the hypothesis that 5-AC affects development and AF production through weakening or even interrupting the connection between VelB and LaeA and then causing dysregulation of the expression pattern of genes involved in development and secondary metabolism. Our results improved the A. flavus genome annotation, provided a comprehensive view of the transcriptome of A. flavus responding to 5-AC and confirmed that fungal development and secondary metabolism are co-regulated. In additon, the RNA-Seq data of another sample treated with gallic acid was used to improve A. flavus genome annotation. mRNA of Aspergillus flavus cultured in three different culture media PDB, PDB+5-AC(5-Azacytidine),and PDB+GA(gallic acid) was subjected to sequence independently.

Project description:Aspergillus flavus is one of the major fungal molds that colonize peanut in the field and during storage. The impact to human and animal health and to economy in agriculture and commerce are significant since this mold produces the most potent natural toxins, aflatoxins, which are carcinogenic, mutagenic, immunosuppressive, and teratogenic. A strain of marine Bacillus megaterium isolated from the Yellow Sea of East China was evaluated for its effect to inhibit aflatoxin formation through down-regulating aflatoxin pathway gene expression in A. flavus as demonstrated by genechip analysis in liquid medium and peanuts. The results showed that aflatoxin accumulation in potato dextrose broth liquid medium and liquid minimal medium was almost totally (more than 98%) inhibited by B. megaterium. The expression of many of the aflatoxin biosynthetic genes in the fungus was confirmed to be turned down. Some of the target genes down-regulated by B. megaterium within the whole genome and within the aflatoxin pathway gene cluster (aflF, aflT, aflS, aflJ, aflL, aflX) were identified. These target genes could be used for controlling aflatoxin contamination in crops such as corn, cotton, and peanut. Importantly, the expression of the regulatory gene aflS was found to be significantly down-regulated. The effect of B. megaterium on aflatoxin biosynthesis and genes expression of pathogen was firstly tested in potato dextrose broth (PDB) and glucose minimal salts medium (MM). The cell suspension of B. megaterium (concentration in PDB and MM was finally adjusted to 108 CFU/ml) or sterile distilled water as a control was added into the 100 ml beaker flask containing 15 ml PDB or MM, respectively. Then 100 M-NM-<l of spore suspension (5 M-CM-^W 106 spores/ml) of A. flavus were added into each beaker flask. After 48 h of incubation at 28M-BM-0C at 200 rpm, mycelia were collected, fresh frozen with liquid nitrogen, ground to a fine powder in liquid nitrogen, and stored at -80M-BM-0C for further analysis. The effect of B. megaterium on aflatoxin biosynthesis and genes expression in the A. flavus fungal pathogen was also tested in two types of peanut kernels, UF 715133-1 and Jinhua 1012, respectively. Peanut kernels were wounded (6 mm diameter and approximately 3 mm deep) using a sterile borer and then 20 M-NM-<l of 1 M-CM-^W 108 CFU/ml cell suspension of B. megaterium was inoculated on wounded peanut kernels respectively. Sterile distilled water was also used for inoculation as control. Two hours after bacterial inoculation, 10 M-NM-<l A. flavus spore suspension was inoculated into each wound at a concentration of 106 spores/ml. The kernels were placed in artificial weather chamber to maintain high humidity (85%) and incubated at 28M-BM-0C for 7 days. Each treatment was replicated three times with 20 peanut kernels in each test. The mycelia on kernels were harvested at day 7 and fresh frozen immediately in liquid nitrogen, ground into powder, and stored at -80M-BM-0C for further analysis.

Dataset Information

Genotypic and Phenotypic Versatility of Aspergillus flavus during Maize Exploitation

Publications

Genotypic and phenotypic versatility of Aspergillus flavus during maize exploitation.

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