Troubling disparities in viral suppression persist among transgender (trans) women living with HIV in the US. We utilized baseline data from a randomized controlled trial of a behavioral intervention among trans women living with HIV in San Francisco and Los Angeles, to identify the socio-ecological correlates of biologically confirmed viral suppression (< 200 HIV-1 RNA copies/mL). Among 253 participants, the mean age was 43 (SD = 11), 46% identified as Black or African American and 35% were virally non-suppressed. In adjusted Poisson regression models, the following barriers to viral suppression were identified: injection drug use [adjusted risk ratio (aRR) 0.78, 95% CI 0.65-0.93, Z = - 2.64, p = 0.008], methamphetamine use (aRR 0.65, 95% CI 0.51-0.83, Z = - 3.45, p = 0.001), amphetamine use (aRR 0.62, 95% CI 0.44-0.87, Z = - 2.75, p = 0.006), homelessness (aRR 0.79, 95% CI 0.63-0.98, Z = - 2.06, p = 0.039), and sex work (aRR 0.60, 95% CI 0.41-0.86, Z = - 2.77, p = 0.009). These findings underscore the importance of interventions that address the socio-ecological barriers to viral suppression among trans women in urban settings.