Background: Commercial sex plays an increasingly important role in China’s growing HIV and sexually transmitted infection (STI) epidemics. In China, street-based sex workers (SSWs) are a subgroup of female sex workers with a particularly high risk of HIV/STI infections but are neglected in responses to HIV. This study assesses changes in HIV voluntary counseling and testing (VCT) utilization and high-risk sexual behaviors following a three-month HIV preventive intervention among SSWs in Chongqing, China. Methods: A three-month intervention was conducted by a team of peer educators, outreach workers from community-based organizations and health professionals. It mainly included distribution of free pamphlets and condoms and delivery of onsite and clinic-based VCT. Cross-sectional surveys were conducted prior to (n = 100) and immediately following (n = 112) the intervention to assess its impact. In-depth interviews were conducted among 12 SSWs after the intervention to further explore potential barriers to HIV prevention. Results: The intervention significantly increased SSWs’ participation in VCT (from 2.0%–15.2%, P < 0.001). Despite participants’ improved HIV-related knowledge level (from 24.0%–73.2%, P < 0.001), there were minimal changes in the levels of condom use with clients. Qualitative research revealed that fear of police arrest and stigma were the main barriers to VCT utilization. Low condom use was associated with family financial constraints, inadequate power in condom negotiation, low awareness and misconceptions of HIV infection risks. Conclusion: HIV intervention improved VCT utilization and knowledge but we did not observe an increase in condom use after this short intervention. SSWs faced substantial economic, social and environmental barriers to VCT utilization and condom use. Full article