White mold is an agricultural disease caused by the fungus Sclerotinia sclerotiorum, which affects important crops. There are different ways of controlling this organism, but none provides inhibition of its resistance structures (sclerotia). Nanotechnology offers promising applications in agricultural area. Here, silver nanoparticles were biogenically synthesized using the fungus Trichoderma harzianum and characterized. Cytotoxicity and genotoxicity were evaluated, and the nanoparticles were initially tested against white mold sclerotia. Their effects on soybean were also investigated with no effects observed. The nanoparticles showed potential against S. sclerotiorum, inhibiting sclerotia germination and mycelial growth. Nanoparticle characterization data indicated spherical morphology, satisfactory polydispersity and size distribution. Cytotoxicity and genotoxicity assays showed that the nanoparticles caused both the effects, although, the most toxic concentrations were above those applied for white mold control. Given the potential of the nanoparticles against S. sclerotiorum, we conclude that this study presents a first step for a new alternative in white mold control.

Background Biogenic nanoparticles possess a capping of biomolecules derived from the organism employed in the synthesis, which contributes to their stability and biological activity. These nanoparticles have been highlighted for the control of phytopathogens, so there is a need to understand their composition, mechanisms of action, and toxicity. This study aimed to investigate the importance of the capping and compare the effects of capped and uncapped biogenic silver nanoparticles synthesized using the filtrate of Trichoderma harzianum against the phytopathogenic fungus Sclerotinia sclerotiorum. Capping removal, investigation of the composition of the capping and physico-chemical characterization of the capped and uncapped nanoparticles were performed. The effects of the nanoparticles on S. sclerotiorum were evaluated in vitro. Cytotoxicity and genotoxicity of the nanoparticles on different cell lines and its effects on nontarget microorganisms were also investigated. Results The capped and uncapped nanoparticles showed spherical morphology, with greater diameter of the uncapped ones. Functional groups of biomolecules, protein bands and the hydrolytic enzymes NAGase, β-1,3-glucanase, chitinase and acid protease from T. harzianum were detected in the capping. The capped nanoparticles showed great inhibitory potential against S. sclerotiorum, while the uncapped nanoparticles were ineffective. There was no difference in cytotoxicity comparing capped and uncapped nanoparticles, however higher genotoxicity of the uncapped nanoparticles was observed towards the cell lines. Regarding the effects on nontarget microorganisms, in the minimal inhibitory concentration assay only the capped nanoparticles inhibited microorganisms of agricultural importance, while in the molecular analysis of the soil microbiota there were major changes in the soils exposed to the uncapped nanoparticles. Conclusions The results suggest that the capping played an important role in controlling nanoparticle size and contributed to the biological activity of the nanoparticles against S. sclerotiorum. This study opens perspectives for investigations concerning the application of these nanoparticles for the control of phytopathogens.

Trichoderma harzianum is a biological control agent used against phytopathogens and biostimulation in agriculture. However, its efficacy can be affected by biotic and abiotic factors, and microencapsulation has been used to maximize the efficacy. The objective was to develop polymeric microparticles to encapsulate T. harzianum, to perform physicochemical characterization to evaluate its stability, to evaluate effects on the soil microbiota, antifungal activity in vitro and enzymatic activity. Size distribution of wet and dry microparticles was 2000 and 800 µm, respectively. Scanning electron microscopy showed spherical morphology and encapsulation of T. harzianum. Photostability assays showed that encapsulation protected the fungus against ultraviolet radiation. The evaluation of the microbiota showed that the proportion of denitrifying bacteria increased when compared to the control. The T. harzianum encapsulation showed an improvement in the chitinolytic and cellulosic activity. In vitro tests showed that encapsulated fungus were able to provide a greater control of S. sclerotiorum.

In this study, we prepared, characterized, and performed toxicity analyses of poly(ε-caprolactone) nanocapsules loaded with neem oil. Three formulations were prepared by the emulsion/solvent evaporation method. The nanocapsules showed a mean size distribution around 400 nm, with polydispersity below 0.2 and were stable for 120 days. Cytotoxicity and genotoxicity results showed an increase in toxicity of the oleic acid + neem formulations according to the amount of oleic acid used. The minimum inhibitory concentrations demonstrated that all the formulations containing neem oil were active. The nanocapsules containing neem oil did not affect the soil microbiota during 300 days of exposure compared to the control. Phytotoxicity studies indicated that NC_20 (200 mg of neem oil) did not affect the net photosynthesis and stomatal conductance of maize plants, whereas use of NC_10 (100:100 of neem:oleic acid) and NC_15 (150:50 of neem:oleic acid) led to negative effects on these physiological parameters. Hence, the use of oleic acid as a complement in the nanocapsules was not a good strategy, since the nanocapsules that only contained neem oil showed lower toxicity. These results demonstrate that evaluation of the toxicity of nanopesticides is essential for the development of environmentally friendly formulations intended for applications in agriculture.

The biogenic synthesis of metallic nanoparticles can contribute to resolving problems related to pests and soil fertilization. Among the different types of metallic nanoparticles, iron nanoparticles have shown good results, especially concerning toxicity because this metal is an essential micronutrient for all plants and can assist their growth, increasing the levels of carbohydrates, proteins, and chlorophyll. This work performed the green synthesis of biogenic iron oxide nanoparticles using the biological control agent Trichoderma harzianum as a stabilizing agent. The physicochemical properties of the nanoparticles were evaluated using the following techniques: dynamic light scattering, nanoparticle tracking analysis, scanning electron microscopy, X-ray diffraction, and Fourier transform infrared spectroscopy. Cytotoxicity was evaluated using different cell lines, while comet and Allium cepa assays were used to assess genotoxicity. In addition, as a proof of concept, the biological activity of the nanoparticles against the pathogen Sclerotinia sclerotiorum (white mold) was evaluated using an in vitro antifungal activity test. The effect of the nanoparticles on seed germination was also evaluated. The results indicated that the nanoparticles consisted of hematite (α-Fe2O3) and had a mean size diameter of 207 ± 2 nm, polydispersity index of 0.45 ± 0.07, and zeta potential of 13 ± 2 mV. The biogenic iron oxide nanoparticles did not alter cell viability, compared to the controls, and did not lead to changes in the mitotic index, at the concentrations used. Furthermore, they were able to increase the proliferation of Trichoderma, which led to the inhibition of emergence of the pathogen S. sclerotiorum and did not affect the germination of the seeds. Therefore, the green synthesis of biogenic iron oxide nanoparticles based on T. harzianum is an attractive option for pest control, aiming at sustainable agricultural practices.