@article{antiga_la starza_miccoli_d'angeli_scala_zaccaria_shu_obrian_beccaccioli_payne_et al._2020, title={Aspergillus flavus Exploits Maize Kernels Using an "Orphan" Secondary Metabolite Cluster}, volume={21}, ISSN={["1422-0067"]}, DOI={10.3390/ijms21218213}, abstractNote={Aspergillus flavus is a saprophytic cosmopolitan fungus, capable of infecting crops both pre- and post-harvest and exploiting different secondary metabolites, including aflatoxins. Aflatoxins are known carcinogens to animals and humans, but display no clear effect in host plants such as maize. In a previous study, we mined the genome of A. flavus to identify secondary metabolite clusters putatively involving the pathogenesis process in maize. We now focus on cluster 32, encoding for fungal effectors such as salicylate hydroxylase (SalOH), and necrosis- and ethylene-inducing proteins (npp1 domain protein) whose expression is triggered upon kernel contact. In order to understand the role of this genetic cluster in maize kernel infection, mutants of A. flavus, impaired or enhanced in specific functions (e.g., cluster 32 overexpression), were studied for their ability to cause disease. Within this frame, we conducted histological and histochemical experiments to verify the expression of specific genes within the cluster (e.g., SalOH, npp1), the production of salicylate, and the presence of its dehydroxylated form. Results suggest that the initial phase of fungal infection (2 days) of the living tissues of maize kernels (e.g., aleuron) coincides with a significant increase of fungal effectors such as SalOH and Npp1 that appear to be instrumental in eluding host defences and colonising the starch-enriched tissues, and therefore suggest a role of cluster 32 to the onset of infection.}, number={21}, journal={INTERNATIONAL JOURNAL OF MOLECULAR SCIENCES}, author={Antiga, Ludovica and La Starza, Sonia Roberta and Miccoli, Cecilia and D'Angeli, Simone and Scala, Valeria and Zaccaria, Marco and Shu, Xiaomei and Obrian, Gregory and Beccaccioli, Marzia and Payne, Gary A. and et al.}, year={2020}, month={Nov} }
 @article{gilbert_medina_mack_lebar_rodriguez_bhatnagar_magan_obrian_payne_2018, title={Carbon Dioxide Mediates the Response to Temperature and Water Activity Levels in Aspergillus flavus during Infection of Maize Kernels}, volume={10}, ISSN={["2072-6651"]}, DOI={10.3390/toxins10010005}, abstractNote={Aspergillus flavus is a saprophytic fungus that may colonize several important crops, including cotton, maize, peanuts and tree nuts. Concomitant with A. flavus colonization is its potential to secrete mycotoxins, of which the most prominent is aflatoxin. Temperature, water activity (aw) and carbon dioxide (CO2) are three environmental factors shown to influence the fungus-plant interaction, which are predicted to undergo significant changes in the next century. In this study, we used RNA sequencing to better understand the transcriptomic response of the fungus to aw, temperature, and elevated CO2 levels. We demonstrate that aflatoxin (AFB1) production on maize grain was altered by water availability, temperature and CO2. RNA-Sequencing data indicated that several genes, and in particular those involved in the biosynthesis of secondary metabolites, exhibit different responses to water availability or temperature stress depending on the atmospheric CO2 content. Other gene categories affected by CO2 levels alone (350 ppm vs. 1000 ppm at 30 °C/0.99 aw), included amino acid metabolism and folate biosynthesis. Finally, we identified two gene networks significantly influenced by changes in CO2 levels that contain several genes related to cellular replication and transcription. These results demonstrate that changes in atmospheric CO2 under climate change scenarios greatly influences the response of A. flavus to water and temperature when colonizing maize grain.}, number={1}, journal={TOXINS}, author={Gilbert, Matthew K. and Medina, Angel and Mack, Brian M. and Lebar, Matthew D. and Rodriguez, Alicia and Bhatnagar, Deepak and Magan, Naresh and Obrian, Gregory and Payne, Gary}, year={2018}, month={Jan} }
 @article{dolezal_obrian_nielsen_woloshuk_boston_payne_2013, title={Localization, morphology and transcriptional profile of Aspergillus flavus during seed colonization}, volume={14}, ISSN={["1364-3703"]}, DOI={10.1111/mpp.12056}, abstractNote={Summary}, number={9}, journal={MOLECULAR PLANT PATHOLOGY}, author={Dolezal, Andrea L. and Obrian, Gregory R. and Nielsen, Dahlia M. and Woloshuk, Charles P. and Boston, Rebecca S. and Payne, Gary A.}, year={2013}, month={Dec}, pages={898–909} }
 @article{johnson_egner_obrian_glassbrook_roebuck_sutter_payne_kensler_groopman_2008, title={Quantification of urinary aflatoxin B-1 dialdehyde metabolites formed by aflatoxin aldehyde reductase using isotope dilution tandem mass spectrometry}, volume={21}, ISSN={["0893-228X"]}, DOI={10.1021/tx700397n}, abstractNote={The aflatoxin B 1 aldehyde reductases (AFARs), inducible members of the aldo-keto reductase superfamily, convert aflatoxin B 1 dialdehyde derived from the exo- and endo-8,9-epoxides into a number of reduced alcohol products that might be less capable of forming covalent adducts with proteins. An isotope dilution tandem mass spectrometry method for quantification of the metabolites, C-8 monoalcohol, dialcohol, and C-6a monoalcohol, was developed to ascertain their possible role as urinary biomarkers for application to chemoprevention investigations. This method uses a novel (13)C 17-aflatoxin B 1 dialcohol internal standard, synthesized from (13)C 17-aflatoxin B 1 biologically produced by Aspergillus flavus. Chromatographic standards of the alcohols were generated through sodium borohydride reduction of the aflatoxin B 1 dialdehyde. This method was then explored for sensitivity and specificity in urine samples of aflatoxin B 1-dosed rats that were pretreated with 3 H-1,2-dithiole-3-thione to induce the expression of AKR7A1, a rat isoform of AFAR. One of the two known monoalcohols and the dialcohol metabolite were detected in all urine samples. The concentrations were 203.5 +/- 39.0 ng of monoalcohol C-6a/mg of urinary creatinine and 10.0 +/- 1.0 ng of dialcohol/mg of creatinine (mean +/- standard error). These levels represented about 8.0 and 0.4% of the administered aflatoxin B 1 dose that was found in the urine at 24 h, respectively. Thus, this highly sensitive and specific isotope dilution method is applicable to in vivo quantification of urinary alcohol products produced by AFAR. Heretofore, the metabolic fate of the 8,9-epoxides that are critical for aflatoxin toxicities has been measured by biomarkers of lysine-albumin adducts, hepatic and urinary DNA adducts, and urinary mercapturic acids. This urinary detection of the alcohol products directly contributes to the goal of mass balancing the fate of the bioreactive 8,9-epoxides of AFB 1 in vivo.}, number={3}, journal={CHEMICAL RESEARCH IN TOXICOLOGY}, author={Johnson, Denise N. and Egner, Patricia A. and OBrian, Greg and Glassbrook, Norman and Roebuck, Bill D. and Sutter, Thomas R. and Payne, Gary A. and Kensler, Thomas W. and Groopman, John D.}, year={2008}, month={Mar}, pages={752–760} }
 @article{cary_obrian_nielsen_nierman_harris-coward_yu_bhatnagar_cleveland_payne_calvo_2007, title={Elucidation of veA-dependent genes associated with aflatoxin and sclerotial production in Aspergillus flavus by functional genomics}, volume={76}, ISSN={0175-7598 1432-0614}, url={http://dx.doi.org/10.1007/s00253-007-1081-y}, DOI={10.1007/s00253-007-1081-y}, abstractNote={The aflatoxin-producing fungi, Aspergillus flavus and A. parasiticus, form structures called sclerotia that allow for survival under adverse conditions. Deletion of the veA gene in A. flavus and A. parasiticus blocks production of aflatoxin as well as sclerotial formation. We used microarray technology to identify genes differentially expressed in wild-type veA and veA mutant strains that could be involved in aflatoxin production and sclerotial development in A. flavus. The DNA microarray analysis revealed 684 genes whose expression changed significantly over time; 136 of these were differentially expressed between the two strains including 27 genes that demonstrated a significant difference in expression both between strains and over time. A group of 115 genes showed greater expression in the wild-type than in the veA mutant strain. We identified a subgroup of veA-dependent genes that exhibited time-dependent expression profiles similar to those of known aflatoxin biosynthetic genes or that were candidates for involvement in sclerotial production in the wild type.}, number={5}, journal={Applied Microbiology and Biotechnology}, publisher={Springer Science and Business Media LLC}, author={Cary, J. W. and OBrian, G. R. and Nielsen, D. M. and Nierman, W. and Harris-Coward, P. and Yu, J. and Bhatnagar, D. and Cleveland, T. E. and Payne, G. A. and Calvo, A. M.}, year={2007}, month={Jul}, pages={1107–1118} }
 @article{du_obrian_payne_2007, title={Function and regulation of aflJ in the accumulation of aflatoxin early pathway intermediate in Aspergillus flavus}, volume={24}, ISSN={["0265-203X"]}, DOI={10.1080/02652030701513826}, abstractNote={aflJ resides within the aflatoxin biosynthetic gene cluster adjacent to the pathway regulatory gene aflR and is involved in aflatoxin production, but its function is unknown. Over-expression of aflJ in the aflatoxin-producing strain 86-10 resulted in increased aflatoxin. In an effort to study the function and regulation of aflJ, strain 649-1 lacking the entire biosynthetic cluster was transformed with either reporter constructs, expression constructs, or cosmid clones and analysed for gene expression or metabolite accumulation. Over-expression of aflJ did not result in elevated transcription of ver-1, omtA or aflR. To determine if over-expression of aflJ leads to an increase in early pathway intermediates, strain 649-1 was transformed with cosmid 5E6 and either gpdA::aflJ alone, gpdA::aflR alone, or aflJ and aflR together. Cosmid 5E6 contains the genes pksA, nor-1, fas-1, and fas-2, which are required for the biosynthesis of the early pathway intermediate averantin. 649-1 transformants containing 5E6 alone produced no detectable averantin. In contrast, 5E6 transformants with gpdA::aflR produced averantin, but only half as much as those transformants containing both aflR and aflJ. Northern blot analysis showed that 5E6 transformants containing both aflR and aflJ had five times more pksA transcripts and four times more nor-1 transcripts than 5E6 transformants containing gpdA::aflR alone. Further, aflJ transcription was regulated by aflR. Over-expression of aflR resulted in elevated aflJ transcription. aflJ appears to modulate the regulation of early genes in aflatoxin biosynthesis.}, number={10}, journal={FOOD ADDITIVES AND CONTAMINANTS}, author={Du, W. and Obrian, G. R. and Payne, G. A.}, year={2007}, pages={1043–1050} }
 @article{obrian_georgianna_wilkinson_yu_abbas_bhatnagar_cleveland_nierman_payne_2007, title={The effect of elevated temperature on gene transcription and aflatoxin biosynthesis}, volume={99}, ISSN={["0027-5514"]}, DOI={10.3852/mycologia.99.2.232}, abstractNote={The molecular regulation of aflatoxin biosynthesis is complex and influenced by several environmental conditions; one of these is temperature. Aflatoxins are produced optimally at 28-30 C, and production decreases as temperatures approach 37 C, the optimum temperature for fungal growth. To better characterize the influence of temperature on aflatoxin biosynthesis, we monitored the accumulation of aflatoxin and the expression of more than 5000 genes in Aspergillus flavus at 28 C and 37 C. A total of 144 genes were expressed differentially (P < 0.001) between the two temperatures. Among the 103 genes more highly expressed at 28 C, approximately 25% were involved in secondary metabolism and about 30% were classified as hypothetical. Genes encoding a catalase and superoxide dismutase were among those more highly expressed at 37 C. As anticipated we also found that all the aflatoxin biosynthetic genes were much more highly expressed at 28 C relative to 37 C. To our surprise expression of the pathway regulatory genes aflR and aflS, as well as aflR antisense, did not differ between the two temperatures. These data indicate that the failure of A. flavus to produce aflatoxin at 37 C is not due to lack of transcription of aflR or aflS. One explanation is that AFLR is nonfunctional at high temperatures. Regardless, the factor(s) sensing the elevated temperatures must be acute. When aflatoxin-producing cultures are transferred to 37 C they immediately stop producing aflatoxin.}, number={2}, journal={MYCOLOGIA}, author={OBrian, G. R. and Georgianna, D. R. and Wilkinson, J. R. and Yu, J. and Abbas, H. K. and Bhatnagar, D. and Cleveland, T. E. and Nierman, W. and Payne, G. A.}, year={2007}, pages={232–239} }
 @article{xue_isleib_payne_novitzky_obrian_2005, title={Aflatoxin production in peanut lines selected to represent a range of linoleic acid concentrations}, volume={68}, ISSN={["1944-9097"]}, DOI={10.4315/0362-028X-68.1.126}, abstractNote={To determine whether concentrations of linoleate in peanut (Arachis hypogaea L.) seed oil could be used to predict an ability to support aflatoxin production, seeds of genotypes representing a range of linoleate content were inoculated with Aspergillus flavus Link ex Fries and assayed for aflatoxin content. Seeds were blanched and quartered, inoculated with conidia of A. flavus, placed on moistened filter paper in petri dishes, and incubated for 8 days at 28 degrees C. Multiple regression analysis was used to account for the variation among lines with the use of fatty acid concentrations as independent variables. In test 1, linoleate accounted for 39 to 44% of the variation among lines for aflatoxin B1 and B2 and total aflatoxin (26 to 27% after log transformation). Oleate accounted for substantial additional variation (27 to 29%) among lines (20 to 23% after log transformation). Other fatty acids accounted for small fractions of among-line variation. In test 2, linoleate accounted for about 35 to 44% of the variation among entries across traits (29 to 37% for log-transformed data); arachidate accounted for 19 to 29% (27 to 33% after log transformation). Eicosenoate accounted for a small part of the total entry variation. In both experiments, residual variation among entries was significant. Low-linoleate lines consistently contained more aflatoxin, whereas normal- to high-linoleate lines contained variable amounts. Although fatty acid concentrations accounted for significant portions of genetic variation, it is not practical to use them as predictors for susceptibility to aflatoxin contamination, especially for lines in the normal range for oleate and linoleate.}, number={1}, journal={JOURNAL OF FOOD PROTECTION}, author={Xue, HQ and Isleib, TG and Payne, GA and Novitzky, WF and Obrian, G}, year={2005}, month={Jan}, pages={126–132} }
 @article{xue_isleib_payne_wilson_novitzky_g o'brian_2003, title={Comparison of aflatoxin production in normal- and high-oleic backeross-derived peanut lines}, volume={87}, ISSN={["1943-7692"]}, DOI={10.1094/PDIS.2003.87.11.1360}, abstractNote={ The effect of the high-oleate trait of peanut on aflatoxin production was tested by comparing normal oleic lines with high-oleic backcross-derived lines. Seeds were blanched, quartered, and inoculated with Aspergillus flavus conidia, placed on moistened filter paper in petri dishes, and incubated for 8 days. In one experiment, dishes were stacked in plastic bags in a Latin square design with bags and positions in stacks as blocking variables. High-oleic lines averaged nearly twice as much aflatoxin as normal lines. Background genotype had no significant effect on aflatoxin content, and interaction between background genotype and oleate level was not detected. In a second experiment, dishes were arranged on plastic trays enclosed in plastic bags and stacked with PVC spacers between trays. Fungal growth and aflatoxin production were greater than in the first experiment. Background genotype, oleate level, and their interaction were significant. The mean of high-oleic lines was almost twice that of normal lines, but the magnitude of the difference varied with background genotype. Special care should be taken with high-oleic lines to prevent growth of Aspergillus spp. and concomitant development of aflatoxin contamination. }, number={11}, journal={PLANT DISEASE}, author={Xue, HQ and Isleib, TG and Payne, GA and Wilson, RF and Novitzky, WP and G O'Brian}, year={2003}, month={Nov}, pages={1360–1365} }
 @article{obrian_fakhoury_payne_2003, title={Identification of genes differentially expressed during aflatoxin biosynthesis in Aspergillus flavus and Aspergillus parasiticus}, volume={39}, ISSN={["1087-1845"]}, DOI={10.1016/S1087-1845(03)00014-8}, abstractNote={A complex regulatory network governs the biosynthesis of aflatoxin. While several genes involved in aflatoxin production are known, their action alone cannot account for its regulation. Arrays of clones from an Aspergillus flavus cDNA library and glass slide microarrays of ESTs were screened to identify additional genes. An initial screen of the cDNA clone arrays lead to the identification of 753 unique ESTs. Many showed sequence similarity to known metabolic and regulatory genes; however, no function could be ascribed to over 50% of the ESTs. Gene expression analysis of Aspergillus parasiticus grown under conditions conducive and non-conductive for aflatoxin production was evaluated using glass slide microarrays containing the 753 ESTs. Twenty-four genes were more highly expressed during aflatoxin biosynthesis and 18 genes were more highly expressed prior to aflatoxin biosynthesis. No predicted function could be ascribed to 18 of the 24 genes whose elevated expression was associated with aflatoxin biosynthesis.}, number={2}, journal={FUNGAL GENETICS AND BIOLOGY}, author={OBrian, GR and Fakhoury, AM and Payne, GA}, year={2003}, month={Jul}, pages={118–127} }