Fusarium Head Blight (FHB) is a re-emerging disease of wheat that causes extensive agricultural damage through direct losses in yield and quality due to the presence of Fusarium damaged kernels and their associated mycotoxins such as the trichothecene deoxynivalenol (DON). Biological control, including the treatment of crop residues with antagonists such as mycoparasitic Trichoderma species to reduce pathogen inoculum of FHB, holds considerable promise. The main purpose of this PhD thesis is to select fungal antagonists able to compete with FHB causal agents, particularly Fusarium graminearum and Fusarium culmorum, in order to prevent the presence of DON on wheat.
Potential antagonists were screened on the basis of their growth in presence of DON in liquid cultures in multi-wells microplates. For each isolate, growth curves in presence/absence of DON were submitted to linear regression and statistically compared. 10 out of 100 Trichoderma spp., 56 out of 93 F. oxysporum and 2 out of 40 Pythium spp. isolates showed a statistically comparable growth rate in presence/absence of DON. At the end of growth period, cultural filtrates were submitted to HPLC analysis in order to investigate the fate of the mycotoxin. No one of cultural substrates showed a DON content lower than the initial one, enabling us to reject the hypothesis of mycotoxin degradation.
The ten Trichoderma isolates and two Pythium spp., selected for their ability to grow in presence of DON, were investigated as potential antagonists against Fusarium culmorum and F. graminearum mycotoxigenic isolates in plate confrontation assays. The results showed that three Trichoderma isolates were successful antagonists and exhibited antibiosis and mycoparasitism.
In addition, we assessed the transcription of some chitinase-encoding genes, known for their involvement during mycoparasitic growth conditions, in various stages of plate confrontation assays with F. culmorum and F. graminearum by RT-PCR. The results showed that almost all investigated genes encoding chitinases from subgroups A, B and C responded to mycoparasitic conditions and were up-regulated before contact or/and at contact with the host.
In order to evaluate the ability to prevent DON production by F. graminearum, competition tests on natural substrates of these three Trichoderma isolates against mycotoxigenic F. graminearum confirmed the positive results achieved on agar plates. Among these 3 isolates, one showed the best capacity of inhibiting DON production and was chosen for the following analysis.
An in vitro competition test on natural substrates (wheat straw and rice) was performed using qPCR as a tool to estimate the effect of the selected T. gamsii isolate on pathogen’s growth and DON accumulation. Results achieved on rice, confirmed the ability of T. gamsii 6085 to antagonize F. graminearum and F. culmorum isolates whose biomass, in presence of Trichoderma, was lower then the control. On wheat straw, a substrate very poor in nutrients, T. gamsii 6085 seemed to develop very badly and growth of both Fusarium was unaffected by the presence of the antagonist.
In the last part of the work we investigated a possible mechanism possibly involved in DON resistance in Trichoderma. We evaluated the gene expression of some PDR-ABC transporters in confrontation plate assay and in the above mentioned competition test on rice
