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Deoxynivalenol (DON) is a potent mycotoxin and virulence factor produced by Fusarium graminearum. We examined the expression of the core DON biosynthetic gene Tri5 during wheat head infection of susceptible and resistant cultivars and susceptible cultivars treated with strobilurin fungicides (e.g., azoxystrobin). DON was quantified to correlate expression with toxin accumulation. The highest Tri5 expression relative to housekeeping genes occurred at the infection front. As infection progressed, earliest-infected kernels showed diminished relative Tri5 expression but Tri5 expression never ceased during the 21 days observed. Azoxystrobin treatment showed no significant effect on either relative Tri5 expression or DON quantity. The resistant cultivar 'Alsen' showed minimal spread of the fungus, with no fungus detected by day 21. DON was not detected in significant quantities in Alsen in the later stages sampled. In Wheaten, DON levels were negligible at 8 days postinoculation (dpi), with detectable DON at later-sampled time points. Tri5 was detected even in fully senesced kernels 21 dpi. Our data demonstrate the presence of Tri5 transcripts in a susceptible cultivar over a much longer time period than has been previously documented. This suggests the ability of the fungus to rapidly resume toxin biosynthesis in dried infected grain should conducive environmental conditions be present, and provides a possible mechanism for high DON levels in asymptomatic grain.

Fusarium graminearum, the causal agent of head blight of wheat, was the third filamentous fungus to have a completed genome sequence. Since the release of the genome sequence in 2003, F. graminearum has become a model for studies of genomics and transcriptomics, mycotoxins, fungal population genetics, gene function, and sexual development. Herein we present the techniques we have used in our laboratory to perform expression analyses on life cycle stages of F. graminearum and techniques to functionally characterize those genes identified as potentially interesting.

Fusarium graminearum (teleomorph: Gibberella zeae) is an Ascomycete fungal plant pathogen which infects a range of agriculturally important crops, including wheat, barley, and maize. A random plasmid insertion mutagenesis approach was used to analyze the pathogenicity of the PH-1 strain, for which full genomic information is available. Fungal transformants were initially screened for their ability to infect wheat ears. From a total of 1,170 transformants screened, eight were confirmed to be highly reduced in pathogenicity toward wheat ears and roots. These were designated disease-attenuated F. graminearum (daf) mutants. The in vitro growth rate and appearance of each daf mutant was equivalent to the parental strain. Deoxynivalenol (DON) was not detected in threshed grain recovered from ears inoculated with the daf10 mutant. Plasmid rescue and sequencing of the mutant daf10 revealed a deletion of approximately 350 kb from one end of chromosome 1. This chromosome segment is predicted to contain 146 genes. Microarray analysis of daf10 gene expression during growth in DON-inducing conditions confirmed the large deletion. The identities of the genes deleted and their potential role in DON production, pathogenesis, and other life processes are discussed.

Fusarium graminearum, a devastating pathogen of small grains, overwinters on crop residues and produces ephemeral perithecia. Accumulation of lipids in overwintering hyphae would provide reserves for overwinter survival and perithecium development. Fatty acid composition of cultures during perithecium development indicated a drop in neutral lipid levels during development but little change in fatty acid composition across stages. Microscopic examination of cultures early in sexual development revealed hyphal cells engorged with lipid bodies. In comparison, vegetative hyphae contained few lipid bodies. Microarray analysis was performed on wheat stems at stages of colonization through perithecium development. Gene expression analysis during stages of perithecium development both in planta and in vitro (previously published) supports the view that lipid biosynthesis occurs during early stages of wheat colonization leading to sexual development and that lipid oxidation occurs as perithecia are developing. Analysis of gene expression during the stages of wheat stem colonization also revealed sets of genes unique to these stages. These results support the view that lipids accumulate in hyphae colonizing wheat stalks and are subsequently used in perithecium formation on stalk tissue. These results indicate that extensive colonization of plant tissue prior to harvest is essential for subsequent sporulation on crop residues and, thus, has important implications for inoculum reduction.

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Maize is a staple crop in Guatemala, especially in the rural regions where it is consumed in high amounts. Given that traditional pre- and post-harvest practices lead to exposure to the environmental surroundings where pests and microorganisms may be present, maize quality and safety can be compromised severely. In order to assess the potential degree of risk, an exploratory study involving maize mycobiota from six farms from Huehuetenango, Guatemala was conducted. DNA was extracted from the maize samples, and the ITS1 region was subjected to Illumina sequencing. This survey identified 52 fungal taxa in the 90-day maize storage period. For the samples where the maize moisture content exceeded 20%, a high yeast content was observed which can reflect spoilage during storage. A significant amount of Fusarium and Aspergillus – mycotoxin-producing molds – was found, representing a potential for mycotoxin contamination. This indicates a plausible health risk in a region where maize represents a significant portion of the diet. Potential maize pathogens in the genera Acremonium and Cladosproium, and Stenocarpella maydis, were also common. Results from this study can help better understand the potential health-risk scenario in the Highlands of Guatemala if poor grain handling practices are adopted. [ABSTRACT FROM AUTHOR]