This study aimed to assess the knowledge and practices of the general public in the Middle Eastern countries during the COVID-19 pandemic. A cross-sectional study using an online survey was conducted between the 19th of March and the 6th of April 2020 in three Middle Eastern countries (Jordan, Saudi Arabia, and Kuwait) to explore the knowledge and practices of the Middle Eastern population regarding COVID-19. A previously developed questionnaire was adapted and used for this study. Multiple linear regression analysis was used to identify predictors of COVID-19 knowledge. A total of 1208 participants (members of the public) participated from the three countries (Jordan = 389, Saudi Arabia = 433, and Kuwait = 386). The majority of participants ( = 810, 67.2%) were females aged 30 to 49 years ( = 501, 41.5%). Participants had moderate overall COVID-19 knowledge, with a mean (SD) score of 7.93 (±1.72) out of 12 (66.1%). Participants had better knowledge about disease prevention and control (83.0%), whereas the lowest sub-scale scores were for questions about disease transmission routes (43.3%). High education level was an important predictor of greater COVID-19 knowledge scores ( < 0.01). Further public education is needed to address the relatively low level of education regarding the transmission of COVID-19 in the Middle Eastern countries. Policymakers are recommended to develop informative COVID-19 related campaigns that specifically target young people (university students), unemployed individuals, and those with lower levels of education.