Hydrogel capsules as new delivery system for Trichoderma koningiopsis Th003 to control Rhizoctonia solani in rice (Oryza sativa).

The incorporation of biological control agents (BCAs) such as Trichoderma spp. in agricultural systems favors the transition towards sustainable practices of plant nutrition and diseases control. Novel bioproducts for crop management are called to guarantee sustainable antagonism activity of BCAs and increase the acceptance of the farmers. The encapsulation in polymeric matrices play a prominent role for providing an effective carrier/protector and long-lasting bioproduct. This research aimed to study the influence of biopolymer in hydrogel capsules on survival and shelf-life of T. koningiopsis. Thus, two hydrogel capsules prototypes based on alginate (P1) and amidated pectin (P2), containing conidia of T. koningiopsis Th003 were formulated. Capsules were prepared by the ionic gelation method and calcium gluconate as crosslinker. Conidia releasing under different pH values of the medium, survival of conidia in drying capsules, storage stability, and biocontrol activity against rice sheath blight (Rhizoctonia solani) were studied. P2 prototype provided up to 98% survival to Th003 in fluid bed drying, faster conidia releasing at pH 5.8, storage stability greater than 6 months at 18 °C, and up to 67% of disease reduction. However, both biopolymers facilitate the antagonistic activity against R. solani, and therefore can be incorporated in novel hydrogel capsules-based biopreparations. This work incites to develop novel biopesticides-based formulations with potential to improve the delivery process in the target site and the protection of the active ingredient from the environmental factors.

Biological Efficacy of Plant Growth-Promoting Bacteria and Arbuscular Mycorrhizae Fungi: Assessments in Laboratory and Greenhouse Conditions.

Utilizing the interactions of microorganisms with plants offers a favorable path to increase crop production and replace the use of synthetic fertilizers. Different bacteria and fungi have been used as biofertilizers to improve agricultural production, yield, and sustainability. Beneficial microorganisms can act as free-living organisms, symbiotes, and endophytes. Soil bacteria called plant growth-promoting bacteria (PGPB) and fungi called arbuscular mycorrhizae fungi (AMF) stimulate the growth and health of plants by direct and indirect mechanisms including nitrogen fixation, phosphorus solubilization, phytohormone production, enzyme production, antibiotic synthesis, and induced systemic resistance. To use these microorganisms as a biofertilizer, it is necessary to assess their efficacy under laboratory and greenhouse conditions. Few reports detail the methods used to develop a test under different environmental conditions, and without these details it is difficult to develop suitable methodologies to evaluate microorganism-plant relationships. We describe four protocols that go from sample preparation to testing in vitro the efficacy of different biofertilizers. Each protocol can be used to test a different biofertilizer microorganism, focusing on bacteria such as Rhizobium sp., Azotobacter sp., Azospirillum sp., Bacillus sp. as well as AMF such as Glomus sp. These protocols can be used in several stages of biofertilizer development, including microorganism selection, microorganism characterization, and in vitro evaluation of efficacy for the registration process. © 2023 Wiley Periodicals LLC. Basic Protocol 1: Evaluating the biological effect of biofertilizer based on PGPB under laboratory conditions Basic Protocol 2: Evaluating the biological effect of biofertilizer based on PGPB under greenhouse conditions Basic Protocol 3: Evaluating the biological effect of biofertilizer based on symbiotic nitrogen-fixing bacteria Basic Protocol 4: Evaluating the biological effect of biofertilizer based on AMF.

Trichoderma koningiopsis Survival on Coated Seeds and Effect on Plant Growth Promotion in Rice (Oryza sativa).

Seed coating is an alternative delivery system for beneficial plant microorganisms into the soil. Although seed coats are widely used for the application of agrochemicals, the incorporation of beneficial microorganisms has not been explored deeply and their survival on seeds while in storage is unknown. The study aimed to evaluate the effect of the coating process on microbial survival and on plant growth promotion. Two coating formulations were designed, and assessed by two coating processes: rotating drum and fluidized bed. The rotating drum process resulted in more uniform coatings than in the fluidized bed process. In addition, with this coating technique, lower viability losses over time were observed. The rotatory drum prototype containing a biopolymer and a clay mineral derivate (P90) showed the best behavior at the three temperatures evaluated, with superior viabilities compared to the other prototypes and the lowest loss of viability after 12 months. The formulation of this coating prototype may preserve the viability of Trichoderma koningiopsis Th003 up to 15 months at 8 °C, 9 months at 18 °C, and 3 months at 28 °C, which are very promising shelf-life results. Regarding the effect of seed coating on plant growth, prototypes showed higher yields > 16% than the control, comparable to the conventional use of Tricotec® WG, which may reduce the number of applications and water consumption for dissolution of the inoculant. The results demonstrated that the formulation composition, as well as the coating process may impact the microbial survival on seeds.

High-Throughput Assessments for Storage Stability, In Vitro Release, and Particle Size of Encapsulated Biocontrol Fungi in Hydrogel Beads.

Use of biocontrol fungi (BF) such as Trichoderma spp. minimizes fungicide input and increases both plant nutrition and protection from disease. Thus, the introduction of BF by novel inoculants in crop management is an excellent strategy to promote sustainable antagonism activity. Within these strategies, encapsulation in polymeric matrices such as hydrogel beads will play a prominent role in providing an effective carrier/protector and long-lasting bioproduct. These studies have used biomaterials with tunable physicochemical features, providing differential morphologies, compaction, and disintegration, among other parameters. Aiming at developing bioproducts within polymeric hydrogel beads, viability of encapsulated conidia, storage stability, release of active ingredient, and particle size are essential. However, there are no reports that detail standardized and comprehensive methods to evaluate the characterization of these bioproducts. We describe step-by-step protocols that go from sample preparation to testing the viability and storage stability using vacuum-sealed aluminum foil bags. We also describe a high-throughput in vitro method for quantifying released fungal conidia of BF at different pH values. Finally, the particle size of beads is established by bright-field microscopy. These protocols could be transferable to other biological actives, accessible to researchers in the microbiology and bioengineering communities. © 2022 Wiley Periodicals LLC. Basic Protocol 1: Viability of encapsulated conidia Basic Protocol 2: Study of stability of prototypes under storage Basic Protocol 3: Release profile of encapsulated conidia at different pH conditions Basic Protocol 4: Determination of capsule size.

Efecto del estrés térmico y la radiación ultravioleta durante la producción masiva de Nomuraea rileyi / Effect of thermal stress and ultraviolet radiation during massive production of Nomuraea rileyi

Resumen El hongo entomopatógeno Nomuraea rileyi (aislamiento Nm006) ha demostrado un alto potencial para ser utilizado en el control biológico de gusano cogollero del maíz Spodoptera frugiperda. Sin embargo, este microorganismo es altamente susceptible a condiciones abióticas de estrés, lo que dificulta el desarrollo tecnológico de un bioplaguicida. Teniendo en cuenta lo anterior, el objetivo del trabajo fue evaluar el efecto individual de temperatura y luz ultravioleta mediante choques de estrés implementados en el proceso de fermentación sólida. Los conidios obtenidos de los diferentes tratamientos se caracterizaron microbiológicamente (rendimiento y germinación), enzimática (β-esterasa, N-acetilglucosaminidasa y quimoelastasa proteasa Pr1) y biológicamente mediante un bioensayo. Los choques de temperatura no afectaron el rendimiento, germinación y actividad biológica del aislamiento, pero si potenciaron la actividad β-esterasa y la actividad N-acetilglucosaminidasa en comparación con los conidios no expuestos. Con respecto a los choques con luz UV, éstos no mejoraron las características de los conidios de N. rileyi. Con base en los resultados, los choques térmicos a 5 °C y 45 °C, se seleccionaron para la fermentación del hongo, porque no afectaron negativamente ninguna característica y aumentaron las actividades enzimáticas β-esterasa y N-acetilglucosaminidasa de los conidios, lo que podría mejorar la actividad insecticida sobre S. frugiperda.

Abstract The entomopathogenic fungus Nomuraea rileyi (isolate Nm006) is a possible microbial control agent against "fall armyworm" Spodoptera frugiperda. However, this fungus is affected by abiotic stress conditions that cause a slow development of the biopesticide. The objective of this study was to assess the individual effect of abiotic stress (temperature and UV light) during solid fermentation of Nomuraea rileyi Nm006. Conidia were produced under different stress conditions and then characterized microbiologically (yield and germination), enzymatically (β-esterese, N-acetylglucosaminidase and subtilin-like protease Pr1) and biologically against Spodoptera frugiperda. All the treatments were compare with conidia produced without stress. Yield and N-acetylglucosaminidase activity did not improve in conidia produced under temperature stress. Germination, efficacy and β-esterase activity were significantly higher in conidia exposed to thermal shock compared to unexposed conidia. UV light shocks did not improve any characteristics of Nm006. Thermal stress at 5 °C and 45 °C were select for fungus fermentation because conidia had higher enzymatic activity (β-esterase y N-acetilglucosaminidase) and any characteristic was negatively affect. This thermal stress could be a strategy to improve the insecticidal activity against S. frugiperda.