It has been reported that the COVID-19 pandemic has predisposed adolescents to risky behaviors such as substance use and subsequent substance use disorder (SUD). However, it is unknown how the pandemic has changed the prevalence of SUD among adolescents in Uganda. We aimed to determine the prevalence of SUD and associated factors among adolescents in southwestern Uganda. Retrospectively, psychiatry ward records from November 2018 to July 2021 were collected from the largest tertiary hospital in southwestern Uganda. A total of 441 adolescent records were included in the analysis, with a mean age was 17±1.88 years, and the majority were males (50.34%). The overall prevalence of SUD was 7.26% (5.90% and 9.80% before and during the pandemic). Despite a little rise in SUD (3.9% increment) during the COVID-19 pandemic, there was no statistical difference compared to before the pandemic. The likelihood of being diagnosed with SUD was more among older adolescents at any period. In addition, having a diagnosis of bipolar mood disorder reduced the likelihood of SUD during the pandemic. This study indicated no statistical change in the diagnosis of SUD among adolescents before and during the COVID-19 pandemic. As older-male adolescents (17 to 19 years) were at higher risk of SUD, there is a need for early intervention for this group.