Reduced susceptibility to Fusarium head blight in Brachypodium distachyon through priming with the Fusarium mycotoxin deoxynivalenol.

The fungal cereal pathogen Fusarium graminearum produces deoxynivalenol (DON) during infection. The mycotoxin DON is associated with Fusarium head blight (FHB), a disease that can cause vast grain losses. Whilst investigating the suitability of Brachypodium distachyon as a model for spreading resistance to F. graminearum, we unexpectedly discovered that DON pretreatment of spikelets could reduce susceptibility to FHB in this model grass. We started to analyse the cell wall changes in spikelets after infection with F. graminearum wild-type and defined mutants: the DON-deficient Δtri5 mutant and the DON-producing lipase disruption mutant Δfgl1, both infecting only directly inoculated florets, and the mitogen-activated protein (MAP) kinase disruption mutant Δgpmk1, with strongly decreased virulence but intact DON production. At 14 days post-inoculation, the glucose amounts in the non-cellulosic cell wall fraction were only increased in spikelets infected with the DON-producing strains wild-type, Δfgl1 and Δgpmk1. Hence, we tested for DON-induced cell wall changes in B. distachyon, which were most prominent at DON concentrations ranging from 1 to 100 ppb. To test the involvement of DON in defence priming, we pretreated spikelets with DON at a concentration of 1 ppm prior to F. graminearum wild-type infection, which significantly reduced FHB disease symptoms. The analysis of cell wall composition and plant defence-related gene expression after DON pretreatment and fungal infection suggested that DON-induced priming of the spikelet tissue contributed to the reduced susceptibility to FHB.

Secreted fungal effector lipase releases free fatty acids to inhibit innate immunity-related callose formation during wheat head infection.

The deposition of the (1,3)-ß-glucan cell wall polymer callose at sites of attempted penetration is a common plant defense response to intruding pathogens and part of the plant's innate immunity. Infection of the Fusarium graminearum disruption mutant Δfgl1, which lacks the effector lipase FGL1, is restricted to inoculated wheat (Triticum aestivum) spikelets, whereas the wild-type strain colonized the whole wheat spike. Our studies here were aimed at analyzing the role of FGL1 in establishing full F. graminearum virulence. Confocal laser-scanning microscopy revealed that the Δfgl1 mutant strongly induced the deposition of spot-like callose patches in vascular bundles of directly inoculated spikelets, while these callose deposits were not observed in infections by the wild type. Elevated concentrations of the polyunsaturated free fatty acids (FFAs) linoleic and α-linolenic acid, which we detected in F. graminearum wild type-infected wheat spike tissue compared with Δfgl1-infected tissue, provided clear evidence for a suggested function of FGL1 in suppressing callose biosynthesis. These FFAs not only inhibited plant callose biosynthesis in vitro and in planta but also partially restored virulence to the Δfgl1 mutant when applied during infection of wheat spikelets. Additional FFA analysis confirmed that the purified effector lipase FGL1 was sufficient to release linoleic and α-linolenic acids from wheat spike tissue. We concluded that these two FFAs have a major function in the suppression of the innate immunity-related callose biosynthesis and, hence, the progress of F. graminearum wheat infection.

Resistance of callose synthase activity to free fatty acid inhibition as an indicator of Fusarium head blight resistance in wheat.

The fungal pathogen Fusarium graminearum is the causal agent of Fusarium head blight (FHB); a devastating crop disease resulting in heavy yield losses and grain contamination with mycotoxins. We recently showed that the secreted lipase FGL1, a virulence factor of F. graminearum, targets plant defense-related callose biosynthesis during wheat head infection. This effector-like function is based on a FGL1-mediated release of polyunsaturated free fatty acids (FFA) that can inhibit callose synthase activity. The importance of FGL1 in successful wheat head colonization was demonstrated in FGL1 disruption mutants (Δfgl1), where infection was restricted to directly inoculated spikelets and accompanied by strong callose deposition in the spikelet's phloem. The application of polyunsaturated FFA to Δfgl1-infected spikelets prevented callose deposition in the phloem and partially restored wheat head colonization.   The comparative analysis of 3 wheat cultivars revealed that the level of resistance to FHB correlated with resistance to FFA-dependent inhibition of callose biosynthesis. Therefore, resistance of callose biosynthesis to FFA inhibition might be used as marker and/or direct target in the breeding of FHB-resistant wheat cultivars.

Functional characterization of a eukaryotic melibiose transporter.

Pathogenic fungi drastically affect plant health and cause significant losses in crop yield and quality. In spite of their impact, little is known about the carbon sources used by these fungi in planta and about the fungal transporters importing sugars from the plant-fungus interface. Here, we report on the identification and characterization of MELIBIOSE TRANSPORTER1 (MBT1) from the hemibiotrophic fungus Colletotrichum graminicola (teleomorph Glomerella graminicola), the causal agent of leaf anthracnose and stalk rot disease in maize (Zea mays). Functional characterization of the MBT1 protein in baker's yeast (Saccharomyces cerevisiae) expressing the MBT1 cDNA revealed that α-D-galactopyranosyl compounds such as melibiose, galactinol, and raffinose are substrates of MBT1, with melibiose most likely being the preferred substrate. α-D-glucopyranosyl disaccharides like trehalose, isomaltose, or maltose are also accepted by MBT1, although with lower affinities. The MBT1 gene shows low and comparable expression levels in axenically grown C. graminicola and upon infection of maize leaves both during the initial biotrophic development of the fungus and during the subsequent necrotrophic phase. Despite these low levels of MBT1 expression, the MBT1 protein allows efficient growth of C. graminicola on melibiose as sole carbon source in axenic cultures. Although Δmbt1 mutants are unable to grow on melibiose, they do not show virulence defects on maize.