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.
Project description: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 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 out of forty-five attempted interspecific crosses between compatible mating types of A. flavus and A. parasiticus were fertile and produced viable ascospores. Single ascospore strains from each cross were isolated and 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 putative 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 species.
Project description:To better understand the effect of temperature on mycotoxin biosynthesis, RNA-Seq technology was used to profile the Aspergillus flavus transcriptome under different temperature conditions. This approachallowed us to quantify transcript abundance for over 80% of fungal genes including 1,153 genes that were differentially expressed at 30M-BM-0C and 37M-BM-0C. Wleven of the 55 secondary metabolite clusters were up-regulated at the lower temperature, including aflatoxin biosynthesis genes, which were among the most highly up-expressed genes. On average, transcript abundance for the 30 aflatoxin biosynthesis genes was 3,300 times greater at 30M-BM-0C as compared to 37M-BM-0C. The results are consistent with the view that high temperature negatively affects aflatoxin production by turning down transcription of the two key transcriptional regulators, aflR and aflS. Subtle changes in the expression levels of aflS to aflR appear to control transcription activation of the aflatoxin cluster. 2 samples examined: from the fungus grown at 30M-BM-0C and 37M-BM-0C
Project description:We examined the antifungal activity of artemisinin against Aspergillus fumigatus (A. fumigatus), a pathogenic filamentous fungus responsible for allergic and invasive aspergillosis in humans and analyzed transcript profiles of the fungus on exposure to Artemisinin. A. fumigatus spores were cultured for 48 h and then treated with artemisinin (at MIC50 concentration) or solvent control (DMSO) for 3 h to study its transcriptomic profiles.
Project description:Protein lysine acetylation is a prevalent post-translational modification that plays pivotal roles in various biological processes in both prokaryotes and eukaryotes. Aspergillus flavus, as a fungus of aflatoxin producer, has attracted tremendous attention due to its health impact on agricultural commodities. Here, we performed the first lysine-acetylome mapping in this filamentous fungus using immune-affinity-based purification integrated with high-resolution mass spectrometry. Overall, we identified 1383 lysine-acetylation sites in 652 acetylated proteins, which account for 5.18% of the total proteins in A. flavus. The acetylated proteins are involved in various cellular processes involving the ribosome, carbon metabolism, antibiotic biosynthesis, secondary metabolites, and the citrate cycle and are distributed in diverse subcellular locations according to bioinformatics analysis. We demonstrated for the first time the acetylation of fatty acid synthase α and β encoded by aflA and aflB involved in the aflatoxin-biosynthesis pathway (cluster 54), as well as backbone enzymes from secondary metabolite clusters 20 and 21 encoded by AFLA_062860 and AFLA_064240, suggesting important roles for acetylation associated with these processes. Our findings illustrating abundant lysine acetylation in A. flavus expanded the understanding of the fungal acetylome and provided insight into the regulatory roles of acetylation in secondary metabolism.
Project description:The filamentous fungus Aspergillus fumigatus is the cause of a variety of pulmonary infections (chronic pulmonary aspergillosis) and life-threatening systemic infection that can infect a variety of different organs (invasive aspergillosis). A. fumigatus is widely distributed in the environment and produces large numbers of small conidia that are inhaled daily. A rapid immune response eliminates these in the immunocompetent host but in cases of pulmonary disease or immunosuppression conidia can quickly germinate and grow in the body. Statins interfere with biosynthesis of cholesterol and are therefore used in treatment of hypercholesterolemia. There is increasing evidence for the potential use of statins in preventing and treating fungal infections. The aim of this study was to assess the effect of statins on A fumigatus and to characterize the proteomic alterations occurring in A. fumigatus in response to statin. Pre-growth of A fumigatus in the presence of statin resulted in lower levels of ergosterol. Cells also showed increased permeability as measured by elevated amino acid and protein leakage. Gliotoxin release increased about nine fold in atorvastatin treated cells. Proteomic analysis revealed differential abundance of proteins involved in oxidative stress response such as glutathione S-transferase family protein (8.43 fold increase) and heat shock protein Hsp30/Hsp42 (2.02 fold increase). Protein related to secondary metabolite biosynthesis like nonribosomal peptide synthetase fmpE (3.06 increase) and 3- Aflatoxin B1-aldehyde reductase GliO-like (-2.86 fold decrease) These results indicate that statins have the ability to reduce the growth of A. fumigatus.
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.
Project description:The molecular mechanisms underlying aflatoxin production have been well-studied in strains of the fungus Aspergillus flavus (A. flavus) under artificial conditions. However, aflatoxin biosynthesis has rarely been studied in natural isolates of A. flavus strains. In the present study, tandem mass tag (TMT) labeling and high-performance liquid chromatography (HPLC) coupled with tandem-mass spectrometry analysiswere used for proteomic quantification in natural isolates of high- and low-aflatoxin-yield A. flavus strains.
Project description:Dimethyl sulfoxide (DSMO) is a simple molecule widely used because of its great solvating ability. Beyond its physico-chemical properties, it is also biologically active, including on fungal species. Aspergillus flavus is a saprophytic and famous pathogenic fungus able to produce Aflatoxin B1 (AFB1), a potent carcinogenic mycotoxin which may contaminate many food crops. The aim of this study was to characterize the effect of DMSO on A. flavus transcriptome profile using high-throughput RNA-sequencing assay.
Project description:To better understand the effect of temperature on mycotoxin biosynthesis, RNA-Seq technology was used to profile the Aspergillus flavus transcriptome under different temperature conditions. This approachallowed us to quantify transcript abundance for over 80% of fungal genes including 1,153 genes that were differentially expressed at 30°C and 37°C. Wleven of the 55 secondary metabolite clusters were up-regulated at the lower temperature, including aflatoxin biosynthesis genes, which were among the most highly up-expressed genes. On average, transcript abundance for the 30 aflatoxin biosynthesis genes was 3,300 times greater at 30°C as compared to 37°C. The results are consistent with the view that high temperature negatively affects aflatoxin production by turning down transcription of the two key transcriptional regulators, aflR and aflS. Subtle changes in the expression levels of aflS to aflR appear to control transcription activation of the aflatoxin cluster.