Background: Female sex workers (FSW) bear a high burden of sexually transmitted infections (STI). In this paper, we estimate the prevalence of human immunodeficiency virus (HIV), HBV = hepatitis B virus (HBV), HCV = hepatitis C virus (HCV), syphilis and coinfections in the second Biological and Behavioral Surveillance Survey among FSW in Brazil. Method: The survey was conducted in 12 Brazilian cities from July to November 2016. We used respondent-driven sampling (RDS) to recruit 350 FSW in each city. Rapid tests were used for screening HIV, syphilis, HCV, and HBV. Confirmatory tests were performed on all samples with reactive rapid test result. All testing algorithms and interpretations were done according to the recommendations of the Department of STI/AIDS and viral hepatitis, Ministry of Health. The STI diagnoses were given by: confirmed HIV infection by a positive result on Western blot; active syphilis infection, defined by a RPR titer equal or greater than 1/8; viremia period of HBV and HCV infections, characterized by a detectable (or quantifiable) viral load. Prevalence estimates and standard errors were calculated using statistical procedures suitable for data collected by RDS. Results:Excluding the seeds, 4245 FSW were enrolled. Prevalence estimates were: HIV 5.3% (95% CI: 4.4%–6.2%); active syphilis 8.5% (95% CI: 7.3%–9.7%); HBV 0.4% (95% CI: 0.2%–0.7%); and, HCV 0.9% (95% CI: 0.6%–1.3%). Among the 4154 FSW tested for the 4 infections, 13.3%; (95% CI 12.0%–14.8%) were diagnosed with at least one of the infections, of which 87.6% (95% CI: 83.3%–90.9%) had single infections. The prevalence of HIV/syphilis co-infection was 1.09% (95% CI: 0.7%– 1.6%) and of HIV/HCV or HBV infections was 0.4% (95% CI: 0.2%–0.7%). Conclusions:Our results reveal the need to conduct more studies to estimate the prevalence of STI and co-infections among FSW in Brazil. Longitudinal trends in the prevalence estimates of HIV and other STI provide information to monitor changes in this high-risk population. Additionally, the study highlights the importance of measuring the hepatitis burden among FSW living with HIV, and the need of including FSW in all aspects of STI prevention, care, and treatment programs.