BACKGROUND: Female sex workers (FSWs) are at risk for sexually transmitted infections (STIs), including HIV. We implemented an HIV/STI preventive intervention among FSWs in Shanghai that aimed to increase condom use, improve HIV knowledge, and reduce STI and HIV incidence. METHODS: From six districts in Shanghai, 750 randomly selected venue-based FSWs were allocated to either a behavioural intervention or control group. In the intervention and control groups, 221 and 278 participants, respectively, had at least one follow-up at three or six months. In analysis, we randomly selected 57 lost to follow-up cases in the intervention group and imputed baseline values to equalize the arms at n = 278 (74.1% follow-up rate in each group). The impacts of the intervention on condom use, HIV/STI risk perception and knowledge, and STI incidence were assessed using either a logistic or linear model, adjusting for the baseline measure of the outcome and venue type. RESULTS: The intervention improved consistent condom use with any partner type in the previous month (AOR = 2.09, 95% CI, 1.43-3.04, p = 0.0001). Consistent condom use with clients in the three most recent sex acts increased in both arms, and with primary partners in the intervention arm, but there was no difference between groups after adjusting for baseline condom use and venue type. There were no differences in cumulative incidence of any STI (i.e., chlamydia, gonorrhoea, syphilis) between groups. HIV transmission knowledge (p = 0.0001), condom use skill (p = 0.0421), and self-efficacy for using condoms (p = 0.0071) were improved by the intervention. HIV-related stigma declined (p = 0.0119) and HIV and STI risk perception were improved (4.6 to 13.9%, and 9.4 to 20.0%, respectively). The intervention was associated with these improvements after adjusting for the baseline measure and venue type. CONCLUSION: Following a preventive intervention among Shanghai FSWs, our findings demonstrate that a simple, community-based educational intervention improved overall condom use, HIV and STI knowledge, and attitudes in relation to HIV/AIDS. The intervention should be implemented widely after tailoring educational materials regarding condom negotiation with different partner types (i.e., commercial sex clients and primary partners).