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Background: The Democratic Republic of Congo (DRC) has faced emerging infectious diseases such as Ebola, Mpox and Yellow fever, and antimicrobial resistance is a growing concern. To address these issues, in 2011 the country embarked on implementing the One Health (OH) approach at the national and provincial levels. This study investigates OH institutionalization and implementation in the DRC, describes the process of OH decentralization, and identifies the opportunities and challenges of sustaining these efforts. Methods: We conducted a qualitative study based on literature, document review and key informant interviews. The literature search targeted PubMed, Google Scholar and the document depository of the national One Health platform (NOHP). Key informant identified at the national level included ministry representatives, OH platform members and donors supporting OH implementation. These interviews were conducted in-person and online, recorded, transcribed, and imported into Dedoose software (version 9.2.006) for coding. Content analysis was performed to identify activities, processes, and achievements during the implementation of OH in DRC. Findings: Results of the literature and document review (n = 72) and analysis of stakeholder interviews (n = 24) indicate that a national OH platform, initiated in 2011, is hosted at the Ministry of Higher Education and coordinates other sectors. It comprises governmental departments, academic institutions, and civil society organizations working at the human, animal, and environment sectors. The governance structure includes a national coordinator, a permanent secretariat, technical working groups, and subnational entities at provincial and territorial levels. Following the establishment of the national OH platform, a structured process foresees to facilitate OH implementation at the provincial and territorial levels. Achievements up to today include the development of training programs, establishment of OH committees in some provinces, assessments of workforce needs, formulation of a national strategy, development of governance manuals, and support to the Mpox response coordination.Nevertheless, OH implementation in the DRC faces challenges, including leadership tensions at the national level, inadequate domestic funding, limited training and capacity building for professionals, and insufficient infrastructure for data collection and sharing. Strengthening leadership and coordination, advocating for domestic resource mobilization, and strengthening infrastructure for data collection and sharing while ensuring equity across sectors is essential for advancing the OH agenda and ensuring its efficacy.