Biological Control of Fusarium culmorum, Fusarium graminearum and Fusarium poae by Antagonistic Yeasts

The genus Fusarium is considered to be one of the most pathogenic, phytotoxic and toxin-producing group of microorganisms in the world. Plants infected by these fungi are characterized by a reduced consumer and commercial value, mainly due to the contamination of crops with mycotoxins. Therefore, effective methods of reducing fungi of the genus Fusarium must be implemented already in the field before harvesting, especially with alternative methods to pesticides such as biocontrol. In this study we identified yeasts that inhibit the growth of the pathogenic fungi Fusarium culmorum, F. graminearum and F. poae. Tested yeasts came from different culture collections, or were obtained from organic and conventional cereals. The greater number of yeast isolates from organic cereals showed antagonistic activity against fungi of the genus Fusarium compared to isolates from the conventional cultivation system. Cryptococcus carnescens (E22) isolated from organic wheat was the only isolate that limited the mycelial growth of all three tested fungi and was the best antagonist against F. poae. Selected yeasts showed various mechanisms of action against fungi, including competition for nutrients and space, production of volatile metabolites, reduction of spore germination, production of siderophores or production of extracellular lytic enzymes: chitinase and β-1,3-glucanase. Of all the investigated mechanisms of yeast antagonism against Fusarium, competition for nutrients and the ability to inhibit spore germination prevailed.

Volatile Organic Compound Profiles From Wheat Diseases Are Pathogen-Specific and Can Be Exploited for Disease Classification

Plants and fungi emit volatile organic compounds (VOCs) that are either constitutively produced or are produced in response to changes in their physico-chemical status. We hypothesized that these chemical signals could be utilized as diagnostic tools for plant diseases. VOCs from several common wheat pathogens in pure culture (Fusarium graminearum, Fusarium culmorum, Fusarium avenaceum, Fusarium poae, and Parastagonospora nodorum) were collected and compared among isolates of the same fungus, between pathogens from different species, and between pathogens causing different disease groups [Fusarium head blight (FHB) and Septoria nodorum blotch (SNB)]. In addition, we inoculated two wheat varieties with either F. graminearum or P. nodorum, while one variety was also inoculated with Blumeria graminis f.sp. tritici (powdery mildew, PM). VOCs were collected 7, 14, and 21 days after inoculation. Each fungal species in pure culture emitted a different VOC blend, and each isolate could be classified into its respective disease group based on VOCs with an accuracy of 71.4 and 84.2% for FHB and SNB, respectively. When all collection times were combined, the classification of the tested diseases was correct in 84 and 86% of all cases evaluated. Germacrene D and sativene, which were associated with FHB infection, and mellein and heptadecanone, which were associated with SNB infection, were consistently emitted by both wheat varieties. Wheat plants infected with PM emitted significant amounts of 1-octen-3-ol and 3,5,5-trimethyl-2-hexene. Our study suggests that VOC blends could be used to classify wheat diseases. This is the first step toward a real-time disease detection in the field based on chemical signatures of wheat diseases.

Effect of Thiosemicarbazone Derivatives and Fusarium culmorum (Wm.G. Sm.) Sacc. Infection of Winter Wheat Seedlings on Their Health Status and Soil Biological Activity

Climate change has an impact on agricultural production, including the greater persistence of plant pathogens in the environment. Therefore, the question arises as to how to effectively and safely protect plants by using chemicals, the number of which is decreasing each year. The aim of this study was to evaluate the fungistatic effect of 2 acetylthiophenethiosemicarbazone (2actpTS) and Cis-jasmonethiosemicarbazone (JTS) against Fusarium culmorum (Fc) on winter wheat seedlings. The influence of thiosemicarbazones (TSs) on the health status and phytotoxicity of seedlings and soil biological activity was investigated. Before TSs application (watering or spraying), soil was infected with F. culmorum (strain No. 37). The substance type and method of its application significantly influenced only the fresh weight of the seedlings. The varying phytotoxicity of the molecules depended primarily on their application method to the plants. The highest seedling phytotoxicity was recorded when compounds were applied during watering and the lowest when they were sprayed. The results showed that the application of substances 2actpTS and JTS, both in the form of watering and spraying, had a positive effect on plant health status, as expressed by the disease index (DI) value. The presence of the infectious agent and the type of chemical compound increased soil enzyme activity. The highest total number of bacteria was found in the soil that was infected with Fc and in soil samples where JTS was applied by watering and spraying. The highest utilization of amines, amides and amino acids by microorganisms was found in the samples where JTS was applied. The obtained results will be used to create intermediate products for the protection of cereals against Fusarium diseases, not only at the stage of germination and tillage of plants, but also at subsequent developmental stages.

Superabsorbent Polymer Seed Coating Reduces Leaching of Fungicide but Does Not Alter Their Effectiveness in Suppressing Pathogen Infestation

Superabsorbent polymers (SAPs) applied to soil have been recognized as water reservoirs that allow plants to cope with periods of drought. Their application as a seed coat makes water available directly to the seeds during their germination and early growth phase, but on the other hand, it can affect the efficiency of plant protection substances used in seed dressing. In our experiments, we evaluated the effect of seed coating with SAP on fungicide leaching and changes in their effectiveness in suppressing Fusarium culmorum infestation. Leaching of fungicide from wheat seeds coated with SAP after fungicide dressing, as measured by the inhibition test of mycelium growth under in vitro conditions, was reduced by 14.2–15.8% compared to seeds without SAP coating. Germination of maize seeds and growth of juvenile plants in artificially infected soil did not differ significantly between seeds dressed with fungicide alone and seeds treated with SAP and fungicide. In addition, plants from the seeds coated with SAP alone grew significantly better compared to untreated seeds. Real-time PCR also confirmed this trend by measuring the amount of pathogen DNA in plant tissue. Winter wheat was less tolerant to F. culmorum infection and without fungicide dressing, the seeds were unable to germinate under strong pathogen attack. In the case of milder infection, similar results were observed as in the case of maize seeds.

The Effect of Nitrogen Fertilization and Fusarium culmorum Inoculation on the Biomarkers of Oxidative Stress in Wheat Flag Leaves

During cultivation, wheat is exposed to several abiotic and/or biotic stress conditions that may adversely impact the wheat yield and quality. The impact of abiotic stress caused by nitrogen deficiency and biotic stress caused by phytopathogenic fungus Fusarium culmorum on biomarkers of oxidative stress in the flag leaf of nine winter wheat varieties (Ficko, U-1, Galloper, BC Mandica, BC Opsesija, Ingenio, Isengrain, Felix, and Bezostaya-1) was analyzed in this study. Hydrogen peroxide concentration and lipid peroxidation level were measured as indicators of oxidative stress, while the antioxidant response was determined by measuring the concentration of phenolic compounds and activities of antioxidant enzymes. Wheat variety and nitrogen treatment had a significant effect on all examined biomarkers of oxidative stress in the flag leaf, while the impact of Fusarium treatment was less pronounced. The most significant impact on the measured stress biomarkers had a low nitrogen level, which mainly increased hydrogen peroxide concentration and lipid peroxidation level and decreased activities of antioxidant enzymes in most varieties. The obtained results were discussed and compared with the previous study in which biochemical analyzes were performed on the wheat spike. There was no significant strong correlation between flag leaf and spike response in the measured parameters, which, in addition to the variety-specific response, also indicates a tissue-specific antioxidant response.

Transformation of Selected Fusarium Toxins and Their Masked Forms during Malting of Various Cultivars of Wheat

This study investigated the impact of malting of six wheat cultivars inoculated with Fusarium culmorum on the dynamics of content changes of selected Fusarium toxins. The grains of all the tested cultivars showed a high content of deoxynivalenol (DON), zearalenone (ZEN), and their derivatives, whereas nivalenol (NIV) and its glucoside were found only in the Legenda cultivar. Our experiments confirmed that the malting process of wheat grain enables the secondary growth of Fusarium, and mycotoxin biosynthesis. The levels of toxins in malt were few-fold higher than those in grain; an especially high increase was noted in the case of ZEN and its sulfate as the optimal temperature and pH conditions for the biosynthesis of these toxins by the pathogen are similar to those used in the grain malting process. This is the first paper reporting that during the malting process, biosynthesis of ZEN sulfate occurs, instead of glycosylation, which is a typical modification of mycotoxins by plant detoxication enzymes.

Potential of Trichoderma harzianum and Its Metabolites to Protect Wheat Seedlings against Fusarium culmorum and 2,4-D

Wheat is a critically important crop. The application of fungi, such as Trichoderma harzianum, to protect and improve crop yields could become an alternative solution to synthetic chemicals. However, the interaction between the fungus and wheat in the presence of stress factors at the molecular level has not been fully elucidated. In the present work, we exposed germinating seeds of wheat (Triticum aestivum) to the plant pathogen Fusarium culmorum and the popular herbicide 2,4-dichlorophenoxyacetic acid (2,4-D) in the presence of T. harzianum or its extracellular metabolites. Then, the harvested roots and shoots were analyzed using spectrometry, 2D-PAGE, and MALDI-TOF/MS techniques. Although F. culmorum and 2,4-D were found to disturb seed germination and the chlorophyll content, T. harzianum partly alleviated these negative effects and reduced the synthesis of zearalenone by F. culmorum. Moreover, T. harzianum decreased the activity of oxidoreduction enzymes (CAT and SOD) and the contents of the oxylipins 9-Hode, 13-Hode, and 13-Hotre induced by stress factors. Under the influence of various growth conditions, changes were observed in over 40 proteins from the wheat roots. Higher volumes of proteins and enzymes performing oxidoreductive functions, such as catalase, ascorbate peroxidase, cytochrome C peroxidase, and Cu/Zn superoxide dismutase, were found in the Fusarium-inoculated and 2,4-D-treated wheat roots. Additionally, observation of the level of 12-oxo-phytodienoic acid reductase involved in the oxylipin signaling pathway in wheat showed an increase. Trichoderma and its metabolites present in the system leveled out the mentioned proteins to the control volumes. Among the 30 proteins examined in the shoots, the expression of the proteins involved in photosynthesis and oxidative stress response was found to be induced in the presence of the herbicide and the pathogen. In summary, these proteomic and metabolomic studies confirmed that the presence of T. harzianum results in the alleviation of oxidative stress in wheat induced by 2,4-D or F. culmorum.

In vitro variation in pathogenicity of Fusarium species causing head blight in relation to their isolation from barley head

Till now, no published study is available on the variation in pathogenicity of Fusarium head blight (FHB) pathogens in relation to their isolation origin in barley head. To end this, two barley cultivars of contrasting quantitative resistance were artificially infected by four FHB species under field conditions over two consecutive growing seasons. Then, pathogenicity tests were conducted under in vitro conditions on single-spore cultures originated from both kernels and glumes in the heads. Different pathogenicity was detected among Fusarium species originated from both kernels and glumes, indicating that the same isolate from glumes and kernels differs in pathogenicity on leaves/seedlings. Isolates of Fusarium culmorum and Fusarium verticillioides originated from infected kernels had shorter latent periods and higher area under disease progress curves compared to isolates originated from glumes, and the reverse was observed for the Fusarium equiseti isolate. In the case of Fusarium solani, isolates originated from kernels or from glumes were equally pathogenic. Primarily findings in this first in-depth study have implications for breeding programs relied principally on actual quantification of pathogenicity in Fusarium species present in a given environment. The sampling of fungi should take into account the presence of Fusarium species of interest on kernels or glumes.

Expression Patterns of miR398, miR167, and miR159 in the Interaction between Bread Wheat (Triticum aestivum L.) and Pathogenic Fusarium culmorum and Beneficial Trichoderma Fungi

Bread wheat (Triticum aestivum L.) is an agronomically significant cereal cultivated worldwide. Wheat breeding is limited by numerous abiotic and biotic stresses. One of the most deleterious factors is biotic stress provoked by the Fusarium culmorum fungus. This pathogen is a causative agent of Fusarium root rot and Fusarium head blight. Beneficial fungi Trichoderma atroviride and T. cremeum are strong antagonists of mycotoxigenic Fusarium spp. These fungi promote plant growth and enhance their tolerance of negative environmental conditions. The aim of the study was to determine and compare the spatial (in above- and underground organs) and temporal (early: 6 and 22 hpi; and late: 5 and 7 dpi reactions) expression profiles of three mature miRNAs (miR398, miR167, and miR159) in wheat plants inoculated with two strains of F. culmorum (KF846 and EW49). Moreover, the spatial expression patterns in wheat response between plants inoculated with beneficial T. atroviride (AN35) and T. cremeum (AN392) were assessed. Understanding the sophisticated role of miRNAs in wheat–fungal interactions may initiate a discussion concerning the use of this knowledge to protect wheat plants from the harmful effects of fungal pathogens. With the use of droplet digital PCR (ddPCR), the absolute quantification of the selected miRNAs in the tested material was carried out. The differential accumulation of miR398, miR167, and miR159 in the studied groups was observed. The abundance of all analyzed miRNAs in the roots demonstrated an increase in the early and reduction in late wheat response to F. culmorum inoculation, suggesting the role of these particles in the initial wheat reaction to the studied fungal pathogen. The diverse expression patterns of the studied miRNAs between Trichoderma–inoculated or F. culmorum–inoculated plants and control wheat, as well as between Trichoderma–inoculated and F. culmorum–inoculated plants, were noticed, indicating the need for further analysis.