The impact of bio-compost on the diversity, composition, and structure of soil microbial communities remains a topic requiring further exploration. Globally, many agricultural soils face degradation, prompting the development of diverse strategies for their restoration. Beneficial soil microorganisms are garnering significant attention due to their crucial role in promoting sustainable agricultural practices, particularly the application of bio-compost. This study presents preliminary findings on the potential effects of a bio-compost on the microbial communities of treated soils, assessing its sustainability in consideration of possible impact on the spread of antibiotic-resistant bacteria (ARBs) in the view of a One Health approach. Experiments were conducted using an ex-situ pilot plant setup, where agricultural soil was treated with two types of bio-compost derived from organic waste. Biomolecular analyses were applied to both untreated and bio-compost-treated soils, enabling an assessment of microbial evolution over time. Early results from a one-year trial revealed that bio-compost treatment fosters bacterial proliferation in the soil, showing consistent growth across the study period. However, species analysis indicated that certain microorganisms introduced to the bio-compost gained dominance, leading to the disappearance of some native species during the experiment. ARBs were searched by cultivation on selective media. Preliminary data indicate the presence of ARBs in the bio-compost highlighting the importance in performing such analyses when considering an agricultural practice in a One Health view. Data obtained from experiments on the ex-situ plant, suggest that the treatment with the bio-composts does not alter the autochthonous bacteria community in terms of ARBs.