Emanuela Gobbi

The increasing generation of agro-industrial residues requires the development of sustainable valorization strategies for converting low-value biomasses into high-added-value products within a circular bioeconomy framework. In this study, tomato peels (TP) derived from the tomato-processing industry and green waste (GW) from urban pruning activities were investigated as substrates for solid-state fermentation (SSF) mediated by Trichoderma harzianum, with the aim of evaluating fungal growth, metabolomic remodeling, and the production of antioxidant bioactive compounds. Different substrate formulations containing TP and GW were subjected to SSF for 7 days, and fungal colonization was monitored. The highest fungal colonization was observed in substrates containing high proportions of GW, whereas pure tomato peels showed negligible colonization, indicating a strong substrate-dependent effect on fungal development. UPLC-QToF-MS metabolomic profiling was subsequently performed on the fermented substrate formulations using three independent biological replicates per treatment, whereas the 100% TP formulation, which showed negligible fungal colonization, was excluded from metabolomic analysis. Results revealed marked differences among fermented substrates, and 22 discriminant metabolites significantly enriched in those containing higher proportions of TP were identified. These metabolites mainly included hydroxycinnamic acid derivatives, flavonoids, lignans, phenolic glycosides, and organic acids, such as coumaric acid, hydroxycaffeic acid, cinnamoylglucose, citric acid, and cyanidin glycosides, suggesting fermentation-associated transformation and release of phenolic compounds. Fermented extracts obtained from mixed substrates enriched in TP exhibited the highest antioxidant activity, with DPPH and ABTS radical scavenging capacities reaching up to 63.48 µmol TE/g dw and 119.67 µmol TE/g dry weight, respectively, together with increased total phenolic content. The integration of microbiological, metabolomic, and antioxidant analyses demonstrated that co-fermentation of tomato-processing byproducts with green waste can modulate fermentation outcomes and enhance antioxidant potential. Overall, this study provides new insights into substrate-driven metabolic transformations during Trichoderma-mediated SSF and highlights the potential of mixed agro-industrial residues as sustainable feedstocks for the production of antioxidant-rich extracts with possible nutraceutical and biotechnological applications.

The increasing demand for food production requires innovative approaches to protect crops from pathogens that significantly reduce yield and quality. Phytoplasmas, persistent bacterial pathogens transmitted by phloem-feeding insects, cause severe damage to economically important crops, including tomato plants. Early detection of these pathogens can be crucial considering that traditional molecular diagnostic methods, such as polymerase chain reaction (PCR), often fail during early infection stages due to low pathogen concentrations. In this study, we explore the use of Raman spectroscopy as a rapid, non-invasive tool for monitoring alterations in plant metabolites caused by Candidatus Phytoplasma solani infection in tomato plants. Grafting experiments were performed, and Raman spectra were collected at different time intervals post-infection. Changes in the spectral intensities of chlorophyll, carotenoids, and polyphenols were identified as early as two weeks post-infection, prior to the pathogen's detectability by molecular methods. These findings highlight the potential of Raman spectroscopy to fill the diagnostic gap in the early stages of phytoplasma infections, offering a window for timely intervention and a further tool in precision agriculture.

Agro-industrial residues have transitions from being an environmental problem to being a cost-effective source of biopolymers and value-added chemicals. However, the efficient extraction of the desired products from these residues requires pretreatments. Fungal biorefinery is a fascinating approach for the biotransformation of raw materials into multiple products in a single batch. In this study, the ability of Trichoderma asperellum R to convert fruit scrap and green waste into value-added chemicals was tested in solid-state and in nonsterile conditions. A solid-state fermentation protocol for a tray bioreactor was developed using spawn as the inoculum for nonsterile substrates. T. asperellum R drove the fermentation of both substrates, shaping the metabolites that were enriched in the secondary plant metabolites. Strain R showed cellulase activity only when inoculated on fruit scraps, resulting in increased amounts of polysaccharides in the crude extract. This extract was also enriched in vanillic acid and limonoid, which are intriguing compounds due to the increasing interest in their potential as biological nitrification inhibitors or food additives. Finally, trimethoxybenzaldehyde, an interesting chemical building block, was identified in the extracts of the Trichoderma-guided fermentation. The overall results showed that the application of T. asperellum R has potential as a driver to facilitate the extraction of bioactive substances from nonsterile recalcitrant substrates.

Several agri-food byproducts represent valuable sources of compounds to be reutilized for nutritional, nutraceutical, and cosmetic purposes. Examples especially comprise byproducts from the processing of fruits such as pomace, because of their richness in nutrients (e.g., fibers) and bioactive compounds (e.g., polyphenols) that can be destined for animal and human use. However, in agreement with the principles of circular economy that are being promoted during the most recent years, other understudied agri-food byproducts of both plant and animal origin are being evaluated to assess their possible reutilization and valorization. In this review, we aim at summarizing the most recent research dealing with the extraction of nutrients and bioactive compounds from agri-food byproducts using innovative and sustainable approaches. Specifically, the review is focused on byproducts generated in large amounts (tons/year) by the food industry of Northeast Italy, namely, honey, red fruits (grapes and berries), and chicory, which are especially of interest for their content in phenolic acids, flavonoids, anthocyanins, and dietary fiber. The potential applications of these byproducts and extracts in cosmetic, nutraceutical, and nutritional fields are also discussed, referring to the published literature, as well as their potential utilization as sources of novel bioactive compounds with pharmacological applications.

The continuous increase in biogas production poses the need for innovative applications for its by-products. Solid-state fermentation (SSF) has regained attention in the development of several products because of the possibility to use low-cost and easily available substrates, such as organic wastes. SSF represents a valuable process for agricultural digestate valorization in terms of enzyme production. In the present study, cellulase and esterase were produced by Trichoderma asperellum R on a digestate-based substrate in SSF, with esterase as the highest obtained activity. After assessing the effect of light on it, the esterase production in SSF was optimized using response surface methodology. The influence of substrate composition, temperature and humidity on the enzyme production was evaluated on two sets of data generated based on digestate concentration (50% and 70% w/w). The statistical analyses revealed that these parameters affected esterase production only when Trichoderma asperellum grew on substrate containing 50% w/w of digestate. The best esterase activity (264.6 mU/mg total protein) was achieved with the following optimized SSF parameters: 50% digestate, 50% fruits, 10% sawdust, 30 °C. The current finding of esterase production on digestate-based substrates makes the SSF method presented here a sustainable and completely circular technology.

Earth’s life may have originated as self-replicating RNA, and it has been argued that RNA viruses and viroid-like elements are remnants of such pre-cellular RNA world. RNA viruses are defined by linear RNA genomes encoding an RNA-dependent RNA polymerase (RdRp), whereas viroid-like elements consist of small, single-stranded, circular RNA genomes that, in some cases, encode paired self-cleaving ribozymes. Here we show that the number of candidate viroid-like elements occurring in geographically and ecologically diverse niches is much higher than previously thought. We report that, amongst these circular genomes, fungal ambiviruses are viroid-like elements that undergo rolling circle replication and encode their own viral RdRp. Thus, ambiviruses are distinct infectious RNAs showing hybrid features of viroid-like RNAs and viruses. We also detected similar circular RNAs, containing active ribozymes and encoding RdRps, related to mitochondrial-like fungal viruses, highlighting fungi as an evolutionary hub for RNA viruses and viroid-like elements. Our findings point to a deep co-evolutionary history between RNA viruses and subviral elements and offer new perspectives in the origin and evolution of primordial infectious agents, and RNA life.

In this study, the agricultural digestate from anaerobic biogas production mixed with food wastes was used as a substrate to grow Trichoderma reesei RUT-C30 and Trichoderma atroviride Ta13 in solid-state fermentation (SSF) and produce high-value bioproducts, such as bioactive molecules to be used as ingredients for biostimulants. The Trichoderma spp. reached their maximum growth after 6 and 3 SSF days, respectively. Both Trichoderma species were able to produce cellulase, esterase, and citric and malic acids, while T. atroviride also produced gibberellins and oxylipins as shown by ultraperformance liquid chromatography with quadrupole time-of-flight mass spectrometry (UPLC-QTOF-MS) profiling. Experimental evaluation of germination parameters highlighted a significant promotion of tomato seed germination and root elongation induced by T. atroviride crude extracts from SSF. This study suggests an innovative sustainable use of the whole digestate mixed with agro-food waste as a valuable substrate in fungal biorefineries. Here, it has been applied to produce plant growth-promoting fungi and bioactive molecules for sustainable agriculture.

Earth’s life may have originated as self-replicating RNA. Some of the simplest current RNA replicators are RNA viruses, defined by linear RNA genomes encoding an RNA-dependent RNA polymerase (RdRP), and subviral agents with single-stranded, circular RNA genomes, such as viroids encoding paired self-cleaving ribozymes. Amongst a massive expansion of candidate viroid and viroid-like elements, we report that fungal pathogens, ambiviruses, are viroid-like elements which undergo rolling circle replication and encode their own viral RdRP, thus they are a distinct hybrid infectious agent. These findings point to a deep evolutionary history between modern RNA viruses and sub-viral elements and offer new perspectives on the evolution of primordial infectious agents, and RNA life. One-Sentence Summary Novel infectious agents resembling self-cleaving viroid-like RNAs whilst encoding a viral RNA-dependent RNA polymerase.