Organic agriculture is perceived as environmentally sustainable, but, under its umbrella, different production systems exist ranging from simplified organic productions to well diversified systems with a full implementation of agro-ecological approaches. Among several developed tools for agriculture sustainability assessment, multi-criteria models are increasingly gaining importance. In this study, we evaluated the use of the multi-criteria DEXi-BIOrt tool, coupled with data from long-term experiments, for the sustainability assessment of different organic vegetable production scenarios. These scenarios were applied in two Italian areas: the Adriatic coast of Marche Region and the Metaponto plan of Basilicata Region. Despite the presence of some critical issues, DEXI-BIOrt proved to be a valid tool for the sustainability evaluation of organic vegetable productions. In both areas, the most sustainable scenarios resulted the well diversified organic systems characterized by proper rotations, inclusion of agro-ecological service crops, cultivation of local and different cultivars, and presence of short supply chain mechanisms. Conversely, the implementation of the simplified organic substitution scenarios does not guarantee a suitable level of sustainability. The findings of this study could support decision makers in the implementation of appropriate measures for enhancing organic production sustainability in the framework of post-2020 Common Agricultural Policy.