Grapevine trunk diseases (GTDs) are a major threat to the wine and grape industry. The aim of the study was to investigate the antifungal activity against Neofusicoccum parvum, Diplodia seriata, and Botryosphaeria dothidea of ε-polylysine, chitosan oligomers, their conjugates, Streptomyces rochei and S. lavendofoliae culture filtrates, and their binary mixtures with chitosan oligomers. In vitro mycelial growth inhibition tests suggest that the efficacy of these treatments, in particular those based on ε-polylysine and ε-polylysine:chitosan oligomers 1:1 w/w conjugate, against the three Botryosphaeriaceae species would be comparable to or higher than that of conventional synthetic fungicides. In the case of ε-polylysine, EC₉₀ values as low as 227, 26.9, and 22.5 µg·mL⁻¹ were obtained for N. parvum, D. seriata, and B. dothidea, respectively. Although the efficacy of the conjugate was slightly lower, with EC₉₀ values of 507.5, 580.2, and 497.4 µg·mL⁻¹, respectively, it may represent a more cost-effective option to the utilization of pure ε-polylysine. The proposed treatments may offer a viable and sustainable alternative for controlling GTDs.

Phytophthora cinnamomi, responsible for “root rot” or “dieback” plant disease, causes a significant amount of economic and environmental impact. In this work, the fungicide action of nanocomposites based on silver nanoparticles and polyphenol inclusion compounds, which feature enhanced bioavailability and water solubility, was assayed for the control of this soil-borne water mold. Inclusion compounds were prepared by an aqueous two-phase system separation method through extraction, either in an hydroalcoholic solution with chitosan oligomers (COS) or in a choline chloride:urea:glycerol deep eutectic solvent (DES). The new inclusion compounds were synthesized from stevioside and various polyphenols (gallic acid, silymarin, ferulic acid and curcumin), in a [6:1] ratio in the COS medium and in a [3:1] ratio in the DES medium, respectively. Their in vitro response against Phytophthora cinnamomi isolate MYC43 (at concentrations of 125, 250 and 500 µg·mL⁻¹) was tested, which found a significant mycelial growth inhibition, particularly high for the composites prepared using DES. Therefore, these nanocomposites hold promise as an alternative to fosetyl-Al and metalaxyl conventional systemic fungicides.

Fusarium head blight (FHB) is a complex disease of cereals caused by Fusarium species, which causes severe damages in terms of yield quality and quantity worldwide, and which produces mycotoxin contamination, posing a serious threat to public health. In the study presented herein, the antifungal activity against Fusarium culmorum of chitosan oligomers (COS)–amino acid conjugate complexes was investigated both in vitro and in vivo. The amino acids assayed were cysteine, glycine, proline and tyrosine. In vitro tests showed an enhancement of mycelial growth inhibition, with EC₅₀ and EC₉₀ effective concentration values ranging from 320 to 948 µg·mL⁻¹ and from 1107 to 1407 µg·mL⁻¹ respectively, for the conjugate complexes, as a result of the synergistic behavior between COS and the amino acids, tentatively ascribed to enhanced cell membrane damage originating from lipid peroxidation. Tests on colonies showed a maximum percentage reduction in the number of colonies at 1500 µg·mL⁻¹ concentration, while grain tests were found to inhibit fungal growth, reducing deoxynivalenol content by 89%. The formulation that showed the best performance, i.e., the conjugate complex based on COS and tyrosine, was further investigated in a small-scale field trial with artificially inoculated spelt (Triticum spelta L.), and as a seed treatment to inhibit fungal growth in spelt seedlings. The field experiment showed that the chosen formulation induced a decrease in disease severity, with a control efficacy of 83.5%, while the seed tests showed that the treatment did not affect the percentage of germination and resulted in a lower incidence of root rot caused by the pathogen, albeit with a lower control efficacy (50%). Consequently, the reported conjugate complexes hold enough promise for crop protection applications to deserve further examination in larger field trials, with other Fusarium spp. pathogens and/or Triticum species.

In a context in which the incidence and severity of grapevine fungal diseases is increasing as a result of both climate change and modern management culture practices, reducing the excessive use of phytosanitary products in viticulture represents a major challenge. Specifically, grapevine trunk diseases (GTDs), caused by several complexes of wood decay or xylem-inhabiting fungi, pose a major challenge to vineyard sustainability. In this study, the efficacy of chitosan oligomers (COS)–amino acid conjugate complexes against three fungal species belonging to the Botryosphaeriaceae family (Neofusicoccum parvum, Diplodia seriata, and Botryosphaeria dothidea) was investigated both in vitro and in planta. In vitro tests led to EC₅₀ and EC₉₀ effective concentrations in the 254.6−448.5 and 672.1−1498.5 µg·mL⁻¹ range, respectively, depending on the amino acid involved in the conjugate complex (viz. cysteine, glycine, proline or tyrosine) and on the pathogen assayed. A synergistic effect between COS and the amino acids was observed against D. seriata and B. dothidea (synergy factors of up to 2.5 and 2.8, respectively, according to Wadley’s method). The formulations based on COS and on the conjugate complex that showed the best inhibition rates, COS−tyrosine, were further investigated in a greenhouse trial on grafted vines of two varieties (”Tempranillo” on 775P and “Garnacha” on 110R rootstock), artificially inoculated with the mentioned three Botryosphaeriaceae species. The in planta bioassay revealed that the chosen formulations induced a significant decrease in disease severity against N. parvum and B. dothidea. In summary, the reported conjugate complexes may be promising enough to be worthy of additional examination in larger field trials.

Fusarium head blight (FHB) is a disease that poses a major challenge in cereal production that has important food and feed safety implications due to trichothecene contamination. In this study, the effect of stevioside—a glycoside found in the leaves of candyleaf (Stevia rebaudiana Bertoni)—was evaluated in vitro against Fusarium culmorum (W.G. Smith) Sacc., alone and in combination (in a 1:1 molar ratio) with polyphenols obtained from milk thistle seeds (Silybum marianum (L.) Gaertn). Different concentrations, ranging from 32 to 512 µg·mL⁻¹, were assayed, finding EC₅₀ and EC₉₀ inhibitory concentrations of 156 and 221 µg·mL⁻¹, respectively, for the treatment based only on stevioside, and EC₅₀ and EC₉₀ values of 123 and 160 µg·mL⁻¹, respectively, for the treatment based on the stevioside–polyphenol conjugate complexes. Colony formation inhibition results were consistent, reaching full inhibition at 256 µg·mL⁻¹. Given that synergistic behavior was observed for this latter formulation (SF = 1.43, according to Wadley’s method), it was further assessed for grain protection at storage, mostly directed against mycotoxin contamination caused by the aforementioned phytopathogen, confirming that it could inhibit fungal growth and avoid trichothecene contamination. Moreover, seed tests showed that the treatment did not affect the percentage of germination, and it resulted in a lower incidence of root rot caused by the pathogen in Kamut and winter wheat seedlings. Hence, the application of these stevioside–S. marianum seed extract conjugate complexes may be put forward as a promising and environmentally friendly treatment for the protection of cereal crops and stored grain against FHB.

Fusarium culmorum is a soil-borne fungus able to cause Fusarium head blight, one of the most important cereal diseases worldwide, which can result in significant yield losses of up to 50% and which jeopardizes food and feed safety due to the mycotoxins produced. In the study presented herein, the enhancement of the antifungal activity against this pathogen, resulting from the addition of silver nanoparticles (AgNPs) to different polyphenol-stevioside inclusion compounds, dispersed either in a chitosan oligomers hydroalcoholic solution or in a choline chloride:urea:glycerol deep eutectic solvent, was investigated in vitro. The polyphenols assayed were curcumin, ferulic acid, gallic acid and silymarin. Four composite concentrations (62.5, 125, 250 and 500 µg·mL⁻¹), with and without AgNPs, were assessed, finding noticeable differences in mycelial growth inhibition, with EC₅₀ and EC₉₀ values ranging from 118 to 579 µg·mL⁻¹ and from 333 to 2604 µg·mL⁻¹, respectively. The obtained results evidenced the improved efficacy of the composites with AgNPs, a superior performance of the composites based on curcumin and ferulic acid, and the advantages of the deep eutectic solvent-based dispersion medium over the chitosan oligomers-based one. The reported composites hold promise for crop protection applications.