Flow cytometry as a tool to investigate nuclear senescence in symbiotic and pathogenic systems

Arbuscular mycorrhizae have been reported to moderate negative effects (growth inhibition, necrosis, death) caused by soil-borne pathogenic fungi. In this paper, we assessed root viability with two DNA fluorescent flow cytometry stains that can be usefully combined to evaluate chromatin changes. Nuclei were extracted from arbuscular mycorrhizal (Glomus mosseae) and non-mycorrhizal tomato roots, infected or uninfected with Phytophthora nicotianae var parasitica and watered with a low phosphate nutrient solution. Nuclei extracted from non-mycorrhizal roots infected with the pathogenic fungus showed lower fluorescence values after staining, compared to arbuscular mycorrhizal roots, suggesting that the presence of the pathogen may result in DNA loss and condensation. Infection by either fungus (symbiotic and pathogen) reduced the ratio of 4c to 2c nuclei in the differentiated root.La cytométrie de flux comme moyen pour étudier la sénescence nucléaire dans des systèmes symbiotiques et pathogènes. Les mycorhizes arbusculaires sont connues pour réduire les effets négatifs (croissance, nécrose, mort) causés par les pathogènes telluriques. Dans ce travail, nous avons évalué par cytométrie de flux, la viabilité des racines, en utilisant deux colorants d'ADN qui, en combinaison, permettent d'estimer les changements au niveau de la chromatine. Les noyaux ont été extraits des racines de tomate mycorhizées ou non avec Glomus mosseae ou infectées ou non avec Phytophthora nicotiana var parasitica et ayant reçu une solution nutritive pauvre en phosphore. Les noyaux extraits des racines non mycorhizées, infectées avec le champignon pathogène, ont donné des valeurs plus faibles de florescence par rapport à ceux obtenus avec des noyaux extraits des racines mycorhizées. Cela suggère que la présence de champignons pathogènes se traduit au niveau de l'ADN par une perte de condensation. L'infection par les deux champignons (symbiotique et pathogène) réduit le rapport du nombre de noyaux 4c par rapport aux 2c dans les racines différenciées

The Golden touch – Fungal bioconversion of agroindustrial waste in high value compounds

Reuse what you can, recycle what cannot be reused, repair what is broken, remanufacture what cannot be repaired1. These are the main pillars of the circular economy system. In spite of a linear economy system, these sustainable processes would turn goods that are at the end of their service life into resources for others. This is the way to close loops in industrial ecosystems and minimize waste, a concept that should soon disappear. Is it feasible? Microbial biodiversity could help to change the fate of agroindustrial wastes, transforming them in new products. Thanks to their natural capability to grow on cellulose and lignin, fungi could be good candidates. Fungal fermentation is independent from land exploitation, being ethical sustainable by not competing with food and farm industry. In the optimal conditions, fungal growth could be economic and energetic sustainable, as required by the principle of the green economy. Within the Fungal Kingdom, a great variety of fungal features and metabolic pathways could be found. Unfortunately, this metabolic richness is still poorly known. Moreover, several fungi are edible, and some have a good nutritional quality, being rich in protein, polysaccharides, with low rate of fatty acids, high PUFA levels, and other nutritional compounds as vitamins and essential amino acids. Basidiomycetes and Ascomycetes have been used for centuries as folk medicines, especially in Asian countries, where their medicinal properties are well known. The immunomodulatory and antitumoral properties of substances extracted from various species of edible mushrooms are now also extensively recognized at a scientific level. This project was aimed to investigate the methods to produce such a high-value biomass with low economic and environmental impact. In particular, this work was aimed to find fungal strains which could grow on agroindustrial waste, as for example food industry or insect breeding wastes. These wastes could then find a new exploitation to have good biomass yields, with the desired nutritional features. In this work, medicinal mushrooms have been selected, including Pleurotus ostreatus and Ganoderma lucidum, both known to metabolise many kinds of substrates and to contain a wide variety of bioactive components. To investigate their ability to metabolise agroindustrial wastes and transform them in high value products, media composition was designed using not-easily accessible C and N sources. Twenty cultural lines were set up in submerged fermentation using different wastes. The first goal was to evaluate how biomass production yields were affected by different C and N sources and their ratio. Preliminary results about the biomass recovery and the fungal ability to metabolise agroindustrial wastes look very promising. More in details, fungal strains were able to grow in the presence of most of the media and, in particular, in the presence of insect breeding wastes where the biomass recovery was comparable to the control. Additional efforts are now focused to study the fungal broths and the biomass extracts. They will be analysed from a nutraceutical point of view, evaluating also their antioxidant and radical scavenging properties using spettrophotometric assays. For instance, Folin-Ciocalteu test, ABTS and DPPH assays will be used to assess the presence of bioactive molecules that could a positive outcome on animal and human health

Solution-Phase Parallel Synthesis of Aryloxyimino Amides via a Novel Multicomponent Reaction among Aromatic (Z)-Chlorooximes, lsocyanides, and Electron-Deficient Phenols

A library of 41 aryloxyimino amides was prepared via solution phase parallel synthesis by extending the multicomponent reaction of (Z)-chlorooximes and isocyanides to the use of electron-deficient phenols. The resulting aryloxyiminoamide derivatives can be used as intermediates for the synthesis of benzo[d]isoxazole-3-carboxamides, dramatically reducing the number of synthetic steps required by other methods reported in literature

Mycoprotein: a way for a more sustainable food system

The seventeen Sustainable Development Goals (SDG) are the blueprint to achieve a better and more sustainable future for all. One way to reach this goal is to look and be inspired by nature biodiversity. Microbial biodiversity could be indeed a key factor to achieve several SDG, as environmental degradation, climate change, and sustainable food system. The transition to more sustainable food systems will require many changes, including the search for a new source of food or feed with a lower impact of intensive agriculture and farming. Fungal biomass, known as mycoprotein, could be an alternative source of protein. Why mycoprotein could be a sustainable novel food? First, fungal fermentation is independent of land exploitation, avoiding ethical issues by not competing with food and farm industries. Moreover, fungal growth could be handled in an economic and energetic sustainable way, as required by the principle of the green economy. Indeed, fungal fermentation may be carried out using agro-industrial by-products (AIBPs), valorizing them as valuable feedstocks for a new production system. This project was aimed to investigate the production of high-value fungal biomass with low economic and environmental impact. Medicinal mushrooms were chosen because they are known to metabolize many kinds of substrates and to produce high-value compounds. Pleurotus ostreatus, Ganoderma lucidum, Cordyceps militaris and Pleurotus eryngii were grown in different cultural conditions. Twenty-five cultural lines were set up in submerged fermentation using different by-products including AIBPs. The first goal was to assess the effects of different AIBPs, C, and N sources and ratios on the biomass production yields: results indicated the optimal growth medium was often strain-dependent. More in detail, the best yields were obtained in the presence of insect breeding and AIBPs, where the biomass recovery of some strains was comparable to the control (rich synthetic medium). Once the optimal growth condition was set for each fungus, the nutritional quality of the biomass was evaluated. It should be indeed noted that great variability in the nutritional composition may be found among fungi, due to species, strains, type of substrate used, fermentation time, type of storage, and conservation process. In this project, the chemical composition of the fungal biomasses was investigated using centesimal composition analysis. Fungal biomass was extracted using different methodologies and the extracts were analyzed through spectrophotometric analysis (Folin-Ciocalteu assay, FRAP, and DPPH assays) to assess the presence of bioactive molecules that could have a positive outcome on animal and human health. Data confirmed the major role of culture media on the nutritional value of the produced biomass. Moreover, different strains shown different biomass quality as well. G. lucidum grown on insect breeding by-product had a very high protein content, while higher lipid content was measured in C. militaris grown on cereal molasses by-products. Results are indeed very promising but, in the future, additional effort should be done to optimize fermentation conditions, in order to achieve good nutritional value and to set up the optimal conditions for biomass production scale-up