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Biomimetic calcium phosphate nanoparticles (CaP) have been actively used in biomedicine, due to their high biodegradability and biocompatibility. However, much less progress has been made regarding their application in precision agriculture, i.e., for the controlled delivery of active species to plants. Herein, we report a straightforward and green synthetic method to dope CaP with potassium (K) and nitrogen (N, as nitrate and urea). By modulating the synthetic conditions in terms of maturation time (at 37 °C) and doping, we prepared K- and N-doped nanoparticles in the form of either amorphous calcium phosphate (ACP) or nanocrystalline apatite (Ap) and studied the impact of the dopants on the ACP-to-Ap transformation pathways. Importantly, we found out that ACP, isolated at low maturation times, incorporates nitrogen (in the form of nitrate and urea) to a larger extent than Ap (2.6 vs 1.1 wt %, respectively). Multinutrient nanofertilizers (so-called nanoU-NPK) with the following composition (wt %) were obtained: Ca (23.3), P (10.1), K (1.5), NO

Phytophthora root rot caused by the pathogen Phytophthora cinnamomi is one of the main causes of oak mortality in Mediterranean open woodlands, the so-called dehesas. Disease control is challenging, therefore, new alternative measures are needed. This study focused on searching for natural biocontrol agents with the aim of developing integrated pest management (IPM) strategies in dehesas as a part of adaptive forest management (AFM) strategies. Native Trichoderma spp. were selectively isolated from healthy trees growing in damaged areas by P. cinnamomi root rot, using Rose Bengal selective medium. All Trichoderma (n = 95) isolates were evaluated against P. cinnamomi by mycelial growth inhibition (MGI). Forty-three isolates presented an MGI higher than 60%. Twenty-one isolates belonging to the highest categories of MGI were molecularly identified as T. gamsii, T. viridarium, T. hamatum, T. olivascens, T. virens, T. paraviridescens, T. linzhiense, T. hirsutum, T. samuelsii, and T. harzianum. Amongst the identified strains, 10 outstanding Trichoderma isolates were tested for mycoparasitism, showing values on a scale ranging from 3 to 4. As far as we know, this is the first report referring to the antagonistic activity of native Trichoderma spp. over P. cinnamomi strains cohabiting in the same infected dehesas. The analysis of the tree health status and MGI suggest that the presence of Trichoderma spp. might diminish or even avoid the development of P. cinnamomi, protecting trees from the worst effects of P. cinnamomi root rot.

Natural compounds and living organisms continue to play a limited role in crop protection, and few of them have reached the market, despite their attractiveness and the efforts made in research. Very often these products have negative characteristics compared to synthetic compounds, e.g., higher costs of production, lower effectiveness, lack of persistence, and inability to reach and penetrate the target plant. Conversely, nanotechnologies are having an enormous impact on all human activities, including agriculture, even if the production of some nanomaterials is not environmentally friendly or could have adverse effects on agriculture and the environment. Thus, certain nanomaterials could facilitate the development of formulated natural pesticides, making them more effective and more environmentally friendly. Nanoformulations can improve efficacy, reduce effective doses, and increase shelf-life and persistence. Such controlled-release products can improve delivery to the target pest. This review considers certain available nanomaterials and nanotechnologies for use in agriculture, discussing their properties and the feasibility of their use in sustainable crop protection, in particular, in improving the effectiveness of natural bio-based agrochemicals. © 2019 Society of Chemical Industry.

Nanotechnology is emerging as a very promising tool towards more efficient and sustainable practices in agriculture. In this work, we propose the use of non-toxic calcium phosphate nanoparticles doped with urea (U-ACP) for the fertilization of Triticum durum plants. U-ACP nanoparticles present very similar morphology, structure, and composition than the amorphous precursor of bone mineral, but contain a considerable amount of nitrogen as adsorbed urea (up to ca. 6 wt % urea). Tests on Triticum durum plants indicated that yields and quality of the crops treated with the nanoparticles at reduced nitrogen dosages (by 40%) were unaltered in comparison to positive control plants, which were given the minimum N dosages to obtain the highest values of yield and quality in fields. In addition, optical microscopy inspections showed that Alizarin Red S stained nanoparticles were able to penetrate through the epidermis of the roots or the stomata of the leaves. We observed that the uptake through the roots occurs much faster than through the leaves (1 h vs. 2 days, respectively). Our results highlight the potential of engineering nanoparticles to provide a considerable efficiency of nitrogen uptake by durum wheat and open the door to design more sustainable practices for the fertilization of wheat in fields., This research was funded by Fondazione CARIPLO (project no. 2016-0648: Romancing the stone: size-controlled HYdroxyaPATItes for sustainable Agriculture–HYPATIA) and the Spanish Ministerio de Ciencia, Innovación y Universidades (MCIU/AEI/FEDER) with the Projects NanoSmart (RYC-2016-21042) and NanoVIT (RTI-2018-095794-A-C22). GBRR also acknowledges the Spanish MICINN for her postdoctoral contract within the Juan de la Cierva Program (JdC-2017).

Fungal spores germinate and undergo vegetative growth, leading to either asexual or sexual reproductive dispersal. Previous research has indicated that among developmental regulatory genes, expression is conserved across nutritional environments, whereas pathways for carbon and nitrogen metabolism appear highly responsive—perhaps to accommodate differential nutritive processing. To comprehensively investigate conidial germination and the adaptive life history decision-making underlying these two modes of reproduction, we profiled transcription of Neurospora crassa germinating on two media: synthetic Bird medium, designed to promote asexual reproduction; and a natural maple sap medium, on which both asexual reproduction and sexual reproduction manifest. A later start to germination but faster development was observed on synthetic medium. Metabolic genes exhibited altered expression in response to nutrients—at least 34% of the genes in the genome were significantly downregulated during the first two stages of conidial germination on synthetic medium. Knockouts of genes exhibiting differential expression across development altered germination and growth rates, as well as in one case causing abnormal germination. A consensus Bayesian network of these genes indicated especially tight integration of environmental sensing, asexual and sexual development, and nitrogen metabolism on a natural medium, suggesting that in natural environments, a more dynamic and tentative balance of asexual and sexual development may be typical of N. crassa colonies. IMPORTANCE One of the most remarkable successes of life is its ability to flourish in response to temporally and spatially varying environments. Fungi occupy diverse ecosystems, and their sensitivity to these environmental changes often drives major fungal life history decisions, including the major switch from vegetative growth to asexual or sexual reproduction. Spore germination comprises the first and simplest stage of vegetative growth. We examined the dependence of this early life history on the nutritional environment using genome-wide transcriptomics. We demonstrated that for developmental regulatory genes, expression was generally conserved across nutritional environments, whereas metabolic gene expression was highly labile. The level of activation of developmental genes did depend on current nutrient conditions, as did the modularity of metabolic and developmental response network interactions. This knowledge is critical to the development of future technologies that could manipulate fungal growth for medical, agricultural, or industrial purposes.

Natural compounds and living organisms still have a limited use in crop protection, and only a few of them have reached the market, despite their attractiveness and the efforts made in research. Very often those products have negative characteristics compared to the synthetic compounds, e.g. higher costs of production, lower effectiveness, lack of persistence, and inability to reach and penetrate the target plant. Conversely, nanotechnologies are having an enormous impact in all human activities, including agriculture, even if some nanomaterials are not environmentally friendly to produce, or could have adverse effects in the agriculture and the environment. Thus, some nanomaterials could facilitate the development of formulated natural pesticides, making them more effective and more environmentally friendly. Nanoformulations can improve efficacy, reduce effective doses, and increase shelf-life and persistence. Such controlled release products can improve delivery to the target pest. This review considers some available nanomaterials and nanotechnologies to be used in agriculture, discussing their properties and feasibility in the perspective of their use in sustainable crop protection, in particular to improve the effectiveness of natural bio-based agrochemicals.

Natural compounds and living organisms continue to play a limited role in crop protection, and few of them have reached the market, despite their attractiveness and the efforts made in research. Very often these products have negative characteristics compared to synthetic compounds, e.g., higher costs of production, lower effectiveness, lack of persistence, and inability to reach and penetrate the target plant. Conversely, nanotechnologies are having an enormous impact on all human activities, including agriculture, even if the production of some nanomaterials is not environmentally friendly or could have adverse effects on agriculture and the environment. Thus, certain nanomaterials could facilitate the development of formulated natural pesticides, making them more effective and more environmentally friendly. Nanoformulations can improve efficacy, reduce effective doses, and increase shelf-life and persistence. Such controlled-release products can improve delivery to the target pest. This review considers certain available nanomaterials and nanotechnologies for use in agriculture, discussing their properties and the feasibility of their use in sustainable crop protection, in particular, in improving the effectiveness of natural bio-based agrochemicals. © 2019 Society of Chemical Industry.

Long noncoding RNA (lncRNA) plays important roles in sexual development in eukaryotes. In filamentous fungi, however, little is known about the expression and roles of lncRNAs during fruiting body formation. By profiling developmental transcriptomes during the life cycle of the plant-pathogenic fungus Fusarium graminearum, we identified 547 lncRNAs whose expression was highly dynamic, with about 40% peaking at the meiotic stage. Many lncRNAs were found to be antisense to mRNAs, forming 300 sense-antisense pairs. Although small RNAs were produced from these overlapping loci, antisense lncRNAs appeared not to be involved in gene silencing pathways. Genome-wide analysis of small RNA clusters identified many silenced loci at the meiotic stage. However, we found transcriptionally active small RNA clusters, many of which were associated with lncRNAs. Also, we observed that many antisense lncRNAs and their respective sense transcripts were induced in parallel as the fruiting bodies matured. The nonsense-mediated decay (NMD) pathway is known to determine the fates of lncRNAs as well as mRNAs. Thus, we analyzed mutants defective in NMD and identified a subset of lncRNAs that were induced during sexual development but suppressed by NMD during vegetative growth. These results highlight the developmental stage-specific nature and functional potential of lncRNA expression in shaping the fungal fruiting bodies and provide fundamental resources for studying sexual stage-induced lncRNAs., IMPORTANCE Fusarium graminearum is the causal agent of the head blight on our major staple crops, wheat and corn. The fruiting body formation on the host plants is indispensable for the disease cycle and epidemics. Long noncoding RNA (lncRNA) molecules are emerging as key regulatory components for sexual development in animals and plants. To date, however, there is a paucity of information on the roles of lncRNAs in fungal fruiting body formation. Here we characterized hundreds of lncRNAs that exhibited developmental stage-specific expression patterns during fruiting body formation. Also, we discovered that many lncRNAs were induced in parallel with their overlapping transcripts on the opposite DNA strand during sexual development. Finally, we found a subset of lncRNAs that were regulated by an RNA surveillance system during vegetative growth. This research provides fundamental genomic resources that will spur further investigations on lncRNAs that may play important roles in shaping fungal fruiting bodies.

Long noncoding RNA (lncRNA) plays important roles in morphological differentiation and development in eukaryotes. In filamentous fungi, however, little is known about lncRNAs and their roles in sexual development. Here we describe sexual stage-induced lncRNAs during the formation of perithecia, the sexual fruiting bodies of Fusarium graminearum. We have identified 547 lncRNAs whose expression was developmental stage-specific, with about 40% of which peaked during the development of asci, the sac-like structures containing meiospores. A large fraction of the lncRNAs were found to be antisense to mRNAs, forming 300 sense–antisense pairs. Although small RNAs (sRNAs) were produced from these overlapping loci, most of the antisense lncRNAs appeared not to be involved in gene silencing pathways. Genome-wide analysis of sRNA clusters identified many silenced loci at the meiotic stage. However, we found transcriptionally-active sRNA clusters, many of which were associated with lncRNAs. Also, we observed that many antisense lncRNAs and their respective sense transcripts were induced in parallel as the perithecia matured. To identify regulatory components for lncRNA expression, we analyzed mutants defective in the nonsense-mediated decay (NMD) pathway. A subset of the lncRNAs appeared to be targeted by the NMD before the perithecia formation, suggesting a suppressive role of the NMD in lncRNA expression during vegetative stage. This research provides fundamental genomic resources that will spur further investigations on developmental lncRNAs that may play important roles in shaping the fungal fruiting bodies.

Summary F-box proteins determine substrate specificity of the ubiquitin–proteasome system. Previous work has demonstrated that the F-box protein Fbp1, a component of the SCFFbp1 E3 ligase complex, is essential for invasive growth and virulence of the fungal plant pathogen Fusarium oxysporum. Here, we show that, in addition to invasive growth, Fbp1 also contributes to vegetative hyphal fusion and fungal adhesion to tomato roots. All of these functions have been shown previously to require the mitogen-activated protein kinase (MAPK) Fmk1. We found that Fbp1 is required for full phosphorylation of Fmk1, indicating that Fbp1 regulates virulence and invasive growth via the Fmk1 pathway. Moreover, the Δfbp1 mutant is hypersensitive to sodium dodecylsulfate (SDS) and calcofluor white (CFW) and shows reduced phosphorylation levels of the cell wall integrity MAPK Mpk1 after SDS treatment. Collectively, these results suggest that Fbp1 contributes to both the invasive growth and cell wall integrity MAPK pathways of F. oxysporum.

Summary F-box proteins function in the recruitment of proteins for SCF ubiquitination and proteasome degradation. Here, we studied the role of Fbp1, a nonessential F-box protein of the tomato pathogen Fusarium oxysporum f. sp. lycopersici. The Δfbp1 mutant showed a significant delay in the production of wilt symptoms on tomato plants and was impaired in invasive growth on cellophane membranes and on living plant tissue. To search for target proteins recruited by Fbp1, a combination of sodium dodecylsulphate-polyacrylamide gel electrophoresis (SDS-PAGE) and matrix-assisted laser desorption/ionization time-of-flight/time-of-flight (MALDI-TOF/TOF) was used to compare proteins in mycelia of the wild-type and Δfbp1 mutant. The proteomic approach identified 41 proteins differing significantly in abundance between the two strains, 17 of which were more abundant in the Δfbp1 mutant, suggesting a possible regulation by proteasome degradation. Interestingly, several of the identified proteins were related to vesicle trafficking. Microscopic analysis revealed an impairment of the Δfbp1 strain in directional growth and in the structure of the Spitzenkorper, suggesting a role of Fbp1 in hyphal orientation. Our results indicate that Fbp1 regulates protein turnover and pathogenicity in F. oxysporum.

F‐box proteins determine substrate specificity of the ubiquitin–proteasome system. Previous work has demonstrated that the F‐box protein Fbp1, a component of the SCF(Fbp1) E3 ligase complex, is essential for invasive growth and virulence of the fungal plant pathogen Fusarium oxysporum. Here, we show that, in addition to invasive growth, Fbp1 also contributes to vegetative hyphal fusion and fungal adhesion to tomato roots. All of these functions have been shown previously to require the mitogen‐activated protein kinase (MAPK) Fmk1. We found that Fbp1 is required for full phosphorylation of Fmk1, indicating that Fbp1 regulates virulence and invasive growth via the Fmk1 pathway. Moreover, the Δfbp1 mutant is hypersensitive to sodium dodecylsulfate (SDS) and calcofluor white (CFW) and shows reduced phosphorylation levels of the cell wall integrity MAPK Mpk1 after SDS treatment. Collectively, these results suggest that Fbp1 contributes to both the invasive growth and cell wall integrity MAPK pathways of F. oxysporum.

F‐box proteins function in the recruitment of proteins for SCF ubiquitination and proteasome degradation. Here, we studied the role of Fbp1, a nonessential F‐box protein of the tomato pathogen Fusarium oxysporum f. sp. lycopersici. The Δfbp1 mutant showed a significant delay in the production of wilt symptoms on tomato plants and was impaired in invasive growth on cellophane membranes and on living plant tissue. To search for target proteins recruited by Fbp1, a combination of sodium dodecylsulphate‐polyacrylamide gel electrophoresis (SDS‐PAGE) and matrix‐assisted laser desorption/ionization time‐of‐flight/time‐of‐flight (MALDI‐TOF/TOF) was used to compare proteins in mycelia of the wild‐type and Δfbp1 mutant. The proteomic approach identified 41 proteins differing significantly in abundance between the two strains, 17 of which were more abundant in the Δfbp1 mutant, suggesting a possible regulation by proteasome degradation. Interestingly, several of the identified proteins were related to vesicle trafficking. Microscopic analysis revealed an impairment of the Δfbp1 strain in directional growth and in the structure of the Spitzenkörper, suggesting a role of Fbp1 in hyphal orientation. Our results indicate that Fbp1 regulates protein turnover and pathogenicity in F. oxysporum.