Differential gene expression and metabolomic analyses of Brachypodium distachyon infected by deoxynivalenol producing and non-producing strains of Fusarium graminearum

Abstract

Background: Fusarium Head Blight (FHB) caused primarily by Fusarium graminearum (Fg) is one of the major diseases of small-grain cereals including bread wheat. This disease both reduces yields and causes quality losses due to the production of deoxynivalenol (DON), the major type B trichothecene mycotoxin. DON has been described as a virulence factor enabling efficient colonization of spikes by the fungus in wheat, but its precise role during the infection process is still elusive. Brachypodium distachyon (Bd) is a model cereal species which has been shown to be susceptible to FHB. Here, a functional genomics approach was performed in order to characterize the responses of Bd to Fg infection using a global transcriptional and metabolomic profiling of B. distachyon plants infected by two strains of F. graminearum: a wild-type strain producing DON (Fgdon+) and a mutant strain impaired in the production of the mycotoxin (Fgdon-).

Results: Histological analysis of the interaction of the Bd21 ecotype with both Fg strains showed extensive fungal tissue colonization with the Fgdon+ strain while the florets infected with the Fgdon- strain exhibited a reduced hyphal extension and cell death on palea and lemma tissues. Fungal biomass was reduced in spikes inoculated with the Fgdon- strain as compared with the wild-type strain. The transcriptional analysis showed that jasmonate and ethylene-signalling pathways are induced upon infection, together with genes encoding putative detoxification and transport proteins, antioxidant functions as well as secondary metabolite pathways. In particular, our metabolite profiling analysis showed that tryptophan-derived metabolites, tryptamine, serotonin, coumaroyl-serotonin and feruloyl-serotonin, are more induced upon infection by the Fgdon+ strain than by the Fgdon- strain. Serotonin was shown to exhibit a slight direct antimicrobial effect against Fg.

Conclusion: Our results show that Bd exhibits defense hallmarks similar to those already identified in cereal crops. While the fungus uses DON as a virulence factor, the host plant preferentially induces detoxification and the phenylpropanoid and phenolamide pathways as resistance mechanisms. Together with its amenability in laboratory conditions, this makes Bd a very good model to study cereal resistance mechanisms towards the major disease FHB.

Figure 1

Fusarium head blight symptoms on Brachypodium distachyon spikes. A: Fg don ⁺ strain-induced symptoms, 96 h after point inoculation. B: Fg don ^- strain-induced symptoms, 96 h after point inoculation. Bars: 1 cm.

Figure 2

Histology of early infection of B. distachyon floral cavities by F. graminearum Fg don ⁺ and Fg don ^- strains. A, B: Penetration of feathery stigmata 24 h after point inoculation by Fg don ⁺ and Fg don ^- strains, respectively. C, D: Symptoms and fungal colonization on palea 72 h after point inoculation by Fg don ⁺ and Fg don ^- strains, respectively. Bars: 100 μm. Arrows indicate fungal hyphae.

Figure 3

Estimation of B. distachyon spike (let)s colonization of F. graminearum Fg don ⁺ and Fg don ^- strains . A: Quantification of fungal DNA in infected spikelets (n.q. = not quantifiable, different letters indicate significant differences between conditions; t-test, p-value ≤ 0.01). B: Production of macroconidia on B. distachyon spikes infected by the Fg don ⁺ or the Fg don ^- strain; different letters indicate the significance of differences between conditions (Duncan test, p-value ≤ 0.01). C: Evaluation of B. distachyon rachis colonization by the Fg don ⁺ or the Fg don ^- strain on infected spikes collected 7 dai; left panel: localization of the different rachis sections collected, right panel: quantification of sections presenting out of which fungal growth was observed.

Figure 4

Venn diagrams showing differentially accumulating transcripts in B. distachyon spikelets inoculated with F. graminearum wild-type ( Fg don ⁺ ) or tri5 mutant ( Fg don ^- ) strains or mock-inoculated (Tween). A: Up-regulated transcripts. B: Down-regulated transcripts.

Figure 5

Induction of the tryptophan pathway during the interaction between B. distachyon and F. graminearum . A: Reconstructed tryptophan pathway in B. distachyon. The accession numbers of B. distachyon genes encoding enzymes potentially involved in the metabolic pathway and exhibiting a differential expression between the conditions used for transcriptome analyses are indicated over or next to each arrow (for more information see Additional file 4). B: Metabolic profiling of soluble secondary metabolites from B. distachyon spikelets 96 h after infection by the F. graminearum Fg don ⁺ strain. HPLC profiles were obtained from aqueous methanolic extracts. Chromatograms were analysed by UV detection at 280 nm. For a correspondence of peak numbers, see Additional file 5. C: Relative quantification of metabolites derived from the tryptophan pathway (different letters indicate significance of the difference between conditions; t-test p-value ≤ 0.02).