Giovanna Cristina Varese

Cultivating fungi on agro-industrial by-products (BP) in submerged fermentation is a growing trend used in different research fields, among which are those producing alternative (non-animal) proteins, including mycoproteins. Six BP-based media were screened using four fungal strains, identifying Ganoderma resinaceum MUT3345 grown on stale bread as the most productive combination (18.6 g/L). Using two full factorial designs, this submerged fermentation was optimised via response surface methodology, increasing the protein concentration (+56.27 %). Among downstream processes tested, freeze-drying without homogenisation was selected to produce a stable, gluten-free, protein and fibre-rich mycoflour (20.1 % and 19.6 %, respectively). Amino acid analysis of the freeze-dried fungal biomass showed a similar profile to soy flour and high essential amino acid density (EAA/AA = 41.4%). This analysis led to a tailored nitrogen-to-protein conversion factor (kP = 5.16), much lower and more aligned with other fungal studies than the conventional Jones’ (i.e. 6.25). In conclusion, this study demonstrates the potential of stale bread to sustain alone the growth of a nutritionally valuable basidiomycete, which was processed into a valuable and versatile product that can be used as food ingredient.

Preserving ex situ soil microbial communities is essential for bacterial biodiversity conservation in microbiome biobanking, accurate microbiome research, and future biotechnological applications in sustainable agriculture. The effect of three soil preservation methods—cryopreservation at −80 °C, refrigeration at 4 °C, and lyophilization—on the viability, diversity and metabolic functionality of the soil microbiome was evaluated using soil from untreated and solarized strawberry fields stored at 0, 6 and 12 months. Microbial viability, ecological strategies, functionality, and diversity were assessed using plate counts, r/K strategist profiling, BIOLOG EcoPlates, and 16S rRNA/ITS gene amplicon-based sequencing, respectively. Cryopreservation outperformed the other preservation strategies by generally maintaining total bacterial counts, especially in control soil, preserving the balance between oligotrophic and copiotrophic communities, and sustaining high alpha diversity indices over 12 months. Refrigeration provided moderate preservation, though microbial viability and community stability declined over time. In contrast, lyophilization significantly reduced microbial abundance, altered community composition, and diminished metabolic diversity and substrate utilization. Beta diversity analyses confirmed that cryopreserved samples remained similar in microbial structure to the initial timepoint, while lyophilized samples diverged substantially. Functional analyses also revealed reduced metabolic activity in lyophilized soils, underscoring the negative impact on the microbiome’s ecological roles. Overall, the results of this study identify cryopreservation as the most effective storage method for maintaining soil microbiome integrity, providing critical guidance for preserving soil samples in microbiome research and sustainable agricultural biotechnology.

Polycyclic aromatic hydrocarbons (PAHs) are persistent and toxic pollutants that accumulate in urban soils, reducing microbial diversity and compromising ecosystem functioning. Developing effective bioremediation strategies requires identifying native degraders and understanding their ecological dynamics under pollutant pressure. Here, we investigated fungal and bacterial communities from PAH-contaminated soil subjected to three consecutive enrichment steps using phenanthrene, fluoranthene, benzo(a)pyrene, benzo(g,h,i)perylene, and their mixture as the sole carbon sources. High-throughput sequencing of ITS2 and V3-V4 amplicons revealed a decline in alpha diversity and a strong restructuring of both communities during the enrichment. Distance-based redundancy analysis showed that contaminant type and enrichment progression jointly shaped community composition, selecting for stress-tolerant taxa. Culturomics yielded 102 fungal isolates, representing 19 taxa, predominantly within Ascomycota. The most represented taxa were Galactomyces pseudocandidus (19 strains), Fusarium oxysporum (five), Stilbella aciculosa and Exophiala attenuata (four each) and Fusarium solani (three). Approximately one-third of isolates harbored associated bacteria, mainly Stenotrophomonas, Bosea and Chitinophaga species. Functional assays identified biosurfactant-producing strains, while microplate screening highlighted Fusarium solani, Galactomyces pseudocandidus and Trametes versicolor as capable of growing under PAH-selective conditions. Overall, our results demonstrate that PAH-contaminated soils host fungal taxa able to persist under pollutant pressure together with recurrent fungi-associated bacteria of potential ecological relevance for bioremediation.

The persistence of PAHs in soil poses a significant challenge for common strategies of soil recovery, affecting, for instance, the general perception of urban agriculture, being highly exposed to different sources of pollution. Sustainable strategies for their mitigation are needed, such as mycoremediation, since it relies on fungi to transform pollutants into products with lower environmental burden. The current study investigates fungi of polluted soil, aimed at urban agriculture, with the intent of creating an active microbial consortium capable of transforming the pollutants of interest and decreasing the toxicity of the site. The analysis of key community members revealed a diverse mycobiota, mainly dominated by Ascomycota. Among the 28 isolated taxa and 16 genera, the most abundant ones were Aspergillus , Scedosporium , Trichoderma , and Fusarium . Fungal isolates were evaluated for their potential to grow in the presence of LMW and HMW PAHs, showing a significant difference due to inter- and intra-specific variabilities. Data showed that some fungi can also produce biosurfactants. Coupling this information with the capability of fungi to grow on carriers, six fungi were selected for in situ soil remediation. Biostimulation and bioaugmentation (with strains in co-cultures) were evaluated for pollutant mitigation. The best results were obtained by a microbial consortium that also revealed high tolerance to HMW PAHs. The chronic ecotoxicity of the soil did not show any effect on Eisenia fetida , with the only exception of biostimulation, which led to a 44% toxicity decrease. This study provides a foundation for fungal-based approaches for soil bioremediation and quality recovery, unveiling for the first time the capability of many species to inhabit PAHs-contaminated soil and be actively involved in their degradation. Key points: • The microbiota of polluted soil was mainly dominated by Ascomycota. • The isolated 28 taxa mostly belong to Aspergillus, Scedosporium, Trichoderma, and Fusarium. • Many fungi degrade PAHs and/or produce biosurfactants. • A mix of millet and canary seeds worked as an efficient fungal carrier. • Biostimulation and bioaugmentation were tested against a PAHs-polluted soil.

Corals maintain complex symbiotic relationships with diverse microorganisms, including fungi, which are often overlooked but represent a critical component of the coral holobiont. This study explores the fungal diversity associated with the tissue and skeleton of the red coral Corallium rubrum , a key species in Mediterranean Marine Animal Forests (MAFs). Using a culture‐based approach, we recovered a broad spectrum of fungal diversity, dominated by Ascomycota such as Penicillium , Cladosporium and Aspergillus . The discovery of numerous taxa with known bioactive properties underscores the potential ecological and biotechnological relevance of coral‐associated fungi. At the same time, the presence of species such as Aspergillus sydowii , which is considered pathogenic under elevated temperatures, raises concerns about coral vulnerability during increasingly frequent Mediterranean marine heatwaves. These taxa should be further investigated to evaluate their pathogenic potential. Overall, our results expand current knowledge of coral–fungal associations, providing a foundation for future work on their ecological significance, role in coral resilience and potential applications in biotechnology.

Background: Colistin (Col) resistance and heteroresistance in extensively drug-resistant (XDR) Acinetobacter baumannii severely limit therapeutic options. We investigated the activity and mechanism of human albumin nanoparticles (haNPs) as colistin potentiators against genetically characterized clinical isolates. Methods: Sixteen clinical isolates were analyzed. Col MICs were determined by broth microdilution, and heteroresistance by population analysis profiling. Potentiation of Col activity was assessed using both Col-loaded haNPs (Col/haNPs) and free Col co-administered with empty haNPs, alongside the proton motive force (PMF) uncoupler carbonyl cyanide 3-chlorophenylhydrazone (CCCP). Assays included checkerboard synergy (FICI), membrane potential analysis (DiOC₂(3)), intracellular Col quantification (UPLC–MS/MS), zeta potential measurements, transmission electron microscopy (TEM), protein leakage, and ROS detection. Results: Heteroresistance was detected in 9/16 isolates. Col/haNPs reduced Col MICs by 4–64-fold in resistant strains and shifted MICs to ≤2 mg/L in most heteroresistant isolates. Empty haNPs displayed no intrinsic antibacterial activity yet selectively potentiated Col, with strong synergy (FICI down to 0.035). Membrane depolarization and increased intracellular Col accumulation under haNP-treated conditions paralleled the effects of CCCP, indicating that haNPs elicit a CCCP-like functional response. These findings are compatible with perturbation of membrane energetics and possible downstream effects on PMF-dependent transport processes. TEM and surface charge analyses supported direct nanoparticle–envelope interaction and progressive membrane disruption. Conclusions: haNPs enhance Col activity across genetically diverse A. baumannii isolates, with particularly strong effects in heteroresistant strains. The combined effects of PMF modulation, increased intracellular drug availability, and envelope interaction provide a mechanistic rationale for the use of albumin-based nanoparticles, either as Col carriers or in combination with free drug, to overcome Col resistance and heteroresistance.

The Nagoya Protocol establishes an international framework for access and benefit-sharing including for microbial research. Yet many microbiologists have only a vague understanding of what the Nagoya Protocol requires and are unsure how to navigate its complexities, despite the fact that non-compliance can have significant legal consequences and far-reaching reputational and legal impacts. This paper discusses common misconceptions and practical challenges that microbiologists may encounter when complying with the Nagoya Protocol and a step-by-step guide on how to “do” the Nagoya Protocol. We present three case studies to showcase real-life experiences and provide best practice principles for access and benefit-sharing while fostering biodiversity conservation, equitable collaboration, and sustainable innovation.

The circular bioeconomy, centred on the principles of “Reduce, Reuse, and Recycle”, is central to achieving the goals of the European Green Deal and the “Farm to Fork” strategy. The key approach, within this framework, is the valorisation of food processing side-streams through various processing technologies to produce fertilisers that, when applied to the soil, act as potential “soil improvers”. When applied to soil, these materials affect not only the physical, chemical, and biological properties of soil but also impact greenhouse gas emissions, nutrient leaching, and the levels of contaminants or pathogens. A systematic review of field studies conducted in European soils between 2014 and 2024 was performed with stringent inclusion and exclusion criteria, which allowed the identification of suitable articles in three independent steps. The main soil improvers derived from various agri-food side-streams were biochar, compost, and digestate. Their effects, along with the potential risks associated with their use in agricultural fields, were carefully evaluated and discussed. This review forms part of the DeliSoil project “Delivering safe, sustainable, tailored & societally accepted soil improvers from circular food production for boosting soil health” funded under the European Union “A Soil Deal for Europe”. This systematic review showed that soil improvers enhance soil health by improving microbial diversity, organic matter, and water retention, and by boosting plant growth. However, potential risks require careful management, and long-term studies are needed to fully assess their environmental and agronomic impacts. Graphical Abstract

The wide geographic distribution of microorganisms, combined with their vast taxonomic and functional diversity, make them indispensable reservoirs of genetic variation that sustain ecosystem resilience and fuel biotechnological innovation. However, to use this diversity, microbiologists must navigate a complex legal and regulatory landscape governed by multiple United Nations treaties and their respective access and benefit-sharing frameworks as well as regulatory frameworks specific to particular ecosystems, biosecurity, pathogens, and intellectual property. This complex regulatory web is also actively growing and changing, which makes it immensely challenging for a “regular” microbiologist to navigate. For policymakers and negotiators, it is also difficult to appreciate the full complexity that practitioners experience. This policy briefing provides a concise regulatory guide for practitioners and policymakers alike, summarized in a graphical overview, to provide more clarity and understanding for those at the edge of decision-making and practice.

The white-rot fungus Coriolopsis trogii MUT3379 produces Lac3379-1 laccase at high yields due to the previous development of a robust fermentation process. Throughout the extended use of this strain, we observed the occurrence of substrate-specific guaiacol and ABTS (2,2’-azino-bis(3-ethylbenzothiazoline-6-sulphonic acid)) oxidizing enzymes other than Lac3379-1 Since we did not succeed in producing these enzymes in significant amounts by conventional strain selection and fermentation tools, we developed an approach based on protoplast preparation and regeneration to isolate stable producers of these alternative oxidative enzymes from the complex multinucleate mycelium of C. trogii MUT3379. A cost-effective and efficient protocol for protoplast preparation was developed by using the enzymatic cocktail VinoTaste Pro by Novozymes. A total of 100 protoplast-derived clones were selected and screened to produce laccases and other oxidative enzymes. A variable spectrum of oxidative activity levels, including both high and low producers, was revealed. Notably, a subset of clones exhibited diverse guaiacol/ABTS positive enzymatic patterns. These findings suggest that it is possible to isolate different lineages from the mycelium of C. trogii MUT337, each producing a distinct pattern of oxidative enzymes. This highlights the potential of protoplast-mediated genome separation to uncover novel metabolic traits that would otherwise remain cryptic. These data hold outstanding significance for accessing and producing novel oxidative enzymes from native fungal populations.