BACKGROUND: Substance use frequently goes undetected in primary care. Though barriers to implementing systematic screening for alcohol and drug use have been examined in urban settings, less is known about screening in rural primary care. OBJECTIVE: To identify current screening practices, barriers, facilitators, and recommendations for the implementation of substance use screening in rural federally qualified health centers (FQHCs). DESIGN: As part of a multi-phase study implementing electronic health record-integrated screening, focus groups (n = 60: all stakeholder groups) and individual interviews (n = 10 primary care providers (PCPs)) were conducted. PARTICIPANTS: Three stakeholder groups (PCPs, medical assistants (MAs), and patients) at three rural FQHCs in Maine. APPROACH: Focus groups and interviews were recorded, transcribed, and content analyzed. Themes surrounding current substance use screening practices, barriers to screening, and recommendations for implementation were identified and organized by the Knowledge to Action (KTA) Framework. KEY RESULTS: Identifying the problem: Stakeholders unanimously agreed that screening is important, and that universal screening is preferred to targeted approaches. Adapting to the local context: PCPs and MAs agreed that screening should be done annually. Views were mixed regarding the delivery of screening; patients preferred self-administered, tablet-based screening, while MAs and PCPs were divided between self-administered and face-to-face approaches. Assessing barriers: For patients, barriers to screening centered around a perceived lack of rapport with providers, which contributed to concerns about trust, judgment, and privacy. For PCPs and MAs, barriers included lack of comfort, training, and preparedness to address screening results and offer treatment. CONCLUSIONS: Though stakeholders agree on the importance of implementing universal screening, concerns about the patient-provider relationship, the consequences of disclosure, and privacy appear heightened by the rural context. Findings highlight that strong relationships with providers are critical for patients, while in-clinic resources and training are needed to increase provider comfort and preparedness to address substance use.