Long-term effects of combining anaerobic digestate with other organic waste products on soil microbial communities.

Introduction: Agriculture is undergoing an agroecological transition characterized by adopting new practices to reduce chemical fertilizer inputs. In this context, digestates are emerging as sustainable substitutes for mineral fertilizers. However, large-scale application of digestates in agricultural fields requires rigorous studies to evaluate their long-term effects on soil microbial communities, which are crucial for ecosystem functioning and resilience. Material and methods: This study presents provides a comparative analysis in long-term field conditions of fertilization strategies combining annual applications of raw digestate with biennial applications of different organic waste products (OWPs)-biowaste compost (BIO), farmyard manure (FYM), and urban sewage sludge (SLU)-and compares them to combinations of the same OWPs with mineral fertilizers. The cumulative effects of repeated OWP applications, paired with two nitrogen sources-organic (digestate) and chemical (mineral fertilizer)-were assessed through soil physicochemical and microbial analyses. We hypothesized that the combined effect varied according to the N-supply sources and that this effect also depended on the type of OWP applied. Soil microbial communities were characterized using high-throughput sequencing targeting 16S and 18S ribosomal RNA genes, following DNA extraction from soil samples collected in 2022, six years after the initial digestate application. Results: The results indicated that combining OWPs rich in stable and recalcitrant organic matter, such as BIO and FYM, with raw digestate, offers an improved fertilization practice. This approach maintains soil organic carbon (SOC) levels, increases soil phosphorus and potassium content, and stimulates microbial communities differently than nitrogen supplied via mineral fertilizers. While microbial biomass showed no significant variation across treatments, microbial diversity indices exhibited differences based on the type of OWP and nitrogen source. The fertilization strategies moderately influenced prokaryotic and fungal community structures, with distinct patterns depending on the OWP and nitrogen source. Notably, fungal communities responded more strongly to treatment variations than prokaryotic communities. Discussion: This study provides new insights into the cumulative effects of substituting mineral fertilizers with digestates on soil microbial communities and soil physicochemical parameters. The sustainable development of agroecosystems significantly depends on a better understanding of the complex responses of soil microbial communities to different fertilization regimes. Future research should continue to assess the long-term impact of digestate application on soil microbiota in real agronomic field conditions, considering associated agricultural practices.