Background Studies show higher rates of sexually transmitted infections (STIs) among transgender (trans) and non-binary (TNB) persons compared with the general population. Scant studies have examined non-HIV STI testing (henceforth referred to as STI testing); fewer inclusive of trans men and non-binary persons. We characterised the prevalence of STI testing and time since last STI test and gender-based differences in these outcomes among TNB persons. Methods Data were analysed from a 2018 community-based participatory cross-sectional survey (n = 528). Prevalence of lifetime STI testing history and time since last STI test were reported overall and compared across genders (trans men, trans women, non-binary assigned female at-birth, non-binary assigned male at-birth) using Chi-squared, then bivariable and multivariable logistic regression analyses to compare lifetime STI testing history (ever vs never) across sociodemographic and health care characteristics. Results Most (n = 425; 80.5%) participants reported having ever had an STI test; over half (59.8%) ever tested had tested within the past year. Bivariate analyses showed no significant gender differences in lifetime STI testing history (P = 0.298) or time since last STI test (P = 0.118). In a multivariable model, higher age, reporting multiple committed partners (vs single/divorced), known HIV status, and ever receiving information about pre-exposure prophylaxis (PrEP) were positively associated with ever having had an STI test, whereas Latinx race/ethnicity (vs white) was negatively associated. Conclusions Findings showed high rates of lifetime STI testing and recent testing, with no gender-based differences. Never testing rates were concerning considering screening recommendations. Broad based (non-gender specific) TNB-focused interventions may be warranted to increase uptake.