To date, the highly adapted cave microbial communities are challenged by the expanding anthropization of these subterranean habitats. Although recent advances in characterizing show-caves microbiome composition and functionality, the anthropic effect on promoting the establishment, or reducing the presence of specific microbial guilds has never been studied in detail. This work aims to investigate the whole microbiome (Fungi, Algae, Bacteria and Archaea) of four Italian show-caves, displaying different environmental and geo-morphological conditions and one recently discovered natural cave to highlight potential human-induced microbial traits alterations. Results indicate how show-caves share common microbial traits in contrast to the natural one; the first are characterized by microorganisms related to outdoor environment and/or capable of exploiting extra inputs of organic matter eventually supplied by tourist flows (i.e. Chaetomium and Phoma for fungi and Pseudomonas for bacteria). Yet, variation in microalgae assemblage composition was reported in show-caves, probably related to the effect of the artificial lighting. This study provides insights into the potential microbiome cave contamination by human-related bacteria (e.g. Lactobacillus and Staphylococcus) and commensal/opportunistic human associated fungi (e.g. Candida) and dermatophytes. This work is critical to untangle caves microbiome towards management and conservation of these fragile ecosystems.

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Fungi can run feedstock pretreatment to improve the hydrolysis and utilization of recalcitrant lignocellulose-rich biomass during anaerobic digestion (AD). In this study, three fungal strains (Coprinopsis cinerea MUT 6385, Cyclocybe aegerita MUT 5639, Cephalotrichum stemonitis MUT 6326) were inoculated in the non-sterile solid fraction of digestate, with the aim to further (re)use it as a feedstock for AD. The application of fungal pretreatments induced changes in the plant cell wall polymers, and different profiles were observed among strains. Significant increases (p < 0.05) in the cumulative biogas and methane yields with respect to the untreated control were observed. The most effective pretreatment was carried out for 20 days with C. stemonitis, causing the highest hemicellulose, lignin, and cellulose reduction (59.3%, 9.6%, and 8.2%, respectively); the cumulative biogas and methane production showed a 182% and 214% increase, respectively, compared to the untreated control. The increase in AD yields was ascribable both to the addition of fungal biomass, which acted as an organic feedstock, and to the lignocellulose transformation due to fungal activity during pretreatments. The developed technologies have the potential to enhance the anaerobic degradability of solid digestate and untap its biogas potential for a further digestion step, thus allowing an improvement in the environmental and economic sustainability of the AD process and the better management of its by-products.

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