Background The coronavirus disease 2019 (COVID-19) misinformation and inadequate access to hygiene and sanitation amenities could hamper efforts to contain COVID-19 spread in resource-limited settings. In this study, we describe knowledge of COVID-19 symptoms and preventive measures, sources of information, and access to adequate handwashing among patients with chronic diseases in three Rwandan rural districts during the onset of COVID-19 in Rwanda. Methods This was a cross-sectional survey conducted among patients who were enrolled in the HIV/AIDS, non-communicable diseases, mental health, oncology, and pediatric development programs at health facilities in Kayonza, Kirehe and Burera districts. The study sample was randomly selected and stratified by district and clinical program. Telephone-based data collection occurred between 23 April and 11 May 2020. Primary caregivers responded to the survey when the selected patient was a child under age 18 or severely ill. We defined good knowledge of COVID-19 symptoms and preventive measures as knowing that a dry cough and fever were common symptoms and social distancing or staying home and regular handwashing could prevent COVID-19 infection. Access to adequate handwashing was defined as living in a household with a handwashing station and regular access to clean water and soap. We used Fisher’s exact tests and logistic regression to measure associations between the source of information and good knowledge about COVID-19 and between socio-economic characteristics and access to adequate handwashing. Results In total, 150 patients and 70 caregivers responded to the survey. Forty-eight (22.3%) respondents had no formal education. Sources of COVID-19 information included mass media (86.8%), local government leaders (27.3%), healthcare workers (15.9%) and social media (6.8%). Twenty-seven percent (n=59) of respondents had good knowledge of COVID-19 symptoms and preventive measures. In the adjusted analysis, getting information from news media was associated with having good knowledge about COVID-19 (adjusted odds ratio, aOR: 5.46; 95% CI: 1.43-20.75]. Seventy-nine (35.9%) respondents reported access to adequate handwashing at home, with access varying significantly by the district in favour of Kayonza (61.3%). Conclusions COVID-19-related knowledge and access to adequate handwashing were low among patients with chronic diseases at the beginning of the pandemic in Rwanda. Efforts to mitigate COVID-19 spread among chronic care populations may include investment in targeted COVID-19-related education and access to adequate handwashing.