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The red pigmentation of Fusarium graminearum and related species that cause stem and head blight of cereals is due to the deposition of aurofusarin in the cell walls. To determine the importance of this polyketide for fungal physiology and pathogenicity, aurofusarin deficient mutants were produced by random and targeted mutagenesis of F. pseudograminearum and F. graminearum. We show that a gene cluster, including the F. graminearum PKS12 gene, is responsible for the biosynthesis of aurofusarin. Three F. pseudograminearum aurofusarin deficient mutants were disrupted in a region upstream from a gene with sequence homology to the aflatoxin regulatory gene aflR. Comparative PCR analyses of the aurofusarin gene cluster in F. graminearum, F. culmorum, and F. pseudograminearum show conserved organization and expression analyses detected no PKS12 transcripts in any of the mutants. To confirm that PKS12 encodes the precursor for aurofusarin, targeted mutagenesis was carried out in F. graminearum. All disruptants showed an albino phenotype. The DeltaPKS12 mutants have higher growth rate and a 10-fold increase in conidia production compared to the wild type. Aurofusarin does not appear to aid in radiation protection and all the mutants are fully pathogenic on wheat and barley. HPLC analyses of aurofusarin deficient mutants confirm the absence of aurofusarin and show an increase in the level of the mycotoxin zearalenone.

Fusarium graminearum is the causal agent of the Fusarium head blight (FHB) and a destructive pathogen of cereals accounting for high grain yield losses especially on wheat and maize. Like other fungal pathogens, F. graminearum secretes various extracellular enzymes, which are hypothesized to be involved in host infection. Extracellular lipolytic activity of F. graminearum was strongly induced in culture by wheat germ oil; this allowed us to isolate, clone, and characterize a gene (FGL1) encoding a secreted lipase. Expression analysis indicated that FGL1 is induced by lipid-containing substrates and repressed by glucose. In planta, FGL1 transcription was detected 1 day post-infection of wheat spikes. The function of the FGL1 gene product was verified by specifically demonstrating lipase activity after expression in a heterologous host. Ebelactone B, a known lipase inhibitor, repressed the lipolytic activity of the enzyme. Disease severity was strongly reduced when wild-type conidia were supplemented with ebelactone B. Transformation-mediated disruption of FGL1 led to reduced extracellular lipolytic activity in culture and to reduced virulence to both wheat and maize.

Fusarium graminearum is the main causative agent of Fusarium head blight on small grain cereals and of ear rot on maize. The disease leads to dramatic yield losses and to an accumulation of mycotoxins. The most dominant F. graminearum mycotoxins are the trichothecenes, with deoxynivalenol and nivalenol being the most prevalent derivatives. To investigate the involvement of trichothecenes in the virulence of the pathogen, the gene coding for the initial enzyme of the trichothecene pathway was disrupted in three field isolates, differing in chemotype and in virulence. From each isolate three individual disruption mutants were tested for their virulence on wheat, barley and maize. Despite the different initial virulence of the three wild-type progenitor strains on wheat, all disruption mutants caused disease symptoms on the inoculated spikelet, but the symptoms did not spread into other spikelets. On barley, the trichothecene deficient mutants showed no significant difference compared to the wild-type strains: all were equally aggressive. On maize, mutants derived from the NIV-producing strain caused less disease than their wild-type progenitor strain, while mutants derived from DON-producing strains caused the same level of disease as their progenitor strains. These data demonstrate that trichothecenes influence the virulence of F. graminearum in a highly complex manner, which is strongly host as well as moderately chemotype specific. [ABSTRACT FROM AUTHOR]

Fusarium graminearumis the causal agent of the Fusarium head blight (FHB) and a destructive pathogen of cereals accounting for high grain yield losses especially on wheat and maize. Like other fungal pathogens,F. graminearumsecretes various extracellular enzymes, which are hypothesized to be involved in host infection. Extracellular lipolytic activity ofF. graminearumwas strongly induced in culture by wheat germ oil; this allowed us to isolate, clone, and characterize a gene (FGL1) encoding a secreted lipase. Expression analysis indicated thatFGL1is induced by lipid-containing substrates and repressed by glucose.In planta,FGL1transcription was detected 1 day post-infection of wheat spikes. The function of theFGL1gene product was verified by specifically demonstrating lipase activity after expression in a heterologous host. Ebelactone B, a known lipase inhibitor, repressed the lipolytic activity of the enzyme. Disease severity was strongly reduced when wild-type conidia were supplemented with ebelactone B. Transformation-mediated disruption ofFGL1led to reduced extracellular lipolytic activity in culture and to reduced virulence to both wheat and maize. [ABSTRACT FROM AUTHOR]

Abstract: Fusarium graminearum is a filamentous fungus that causes devastating diseases on plants of economic importance including maize, wheat, and barley. F. graminearum is able to utilize triglycerides as a carbon source during growth. Extracellular lipases are the preferred enzymes to catalyze the hydrolysis of fats and oils. Lipases are ubiquitous enzymes of considerable physiological significance and industrial potential. Previously, FGL1 was the first described F. graminearum extracellular lipase associated with virulence. We report the biochemical characterization of FGL1 and four new secreted lipases of F. graminearum. The lipase genes of F. graminearum wild-type strain 8/1 were sequenced, cloned and over-expressed in Pichia pastoris. We show that the lipases have their temperature optimum between 30 and 40°C and a pH optimum of ∼7. A broad range of lipase substrates, from C4 to C18 p-nitrophenyl esters, were hydrolyzed efficiently by the lipases, indicating the true lipolytic activity of the enzymes. Expression patterns of these lipases were also analyzed by semiquantitative RT-PCR in F. graminearum cultured in water supplemented with 2% (v/v) wheat germ oil at 28°C. Transcripts of all examined lipases are detectable and the genes are regulated differently under these culture conditions. Our data indicated that F. graminearum possesses a ubiquitous source of secreted lipases, which could be used for industrial intentions. We also provided the foundation of lipase expression in vitro, which is necessary for further characterization. [Copyright &y& Elsevier]