Background: Herpes Simplex Virus Type 2 (HSV2) causes genital herpes, an incurable, lifelong sexually transmitted infection (STI). This study was conducted to describe the seroprevalence and associated factors of HSV2 infection in a cohort of Female Sex Workers (FSWs). Methods: A descriptive cross-sectional study was conducted on 136 FSWs who attended the central STD Clinic, in Colombo. They were assessed using an interviewer-administered questionnaire followed by serum sampling. Serum samples were tested for HSV2 IgG using an enzyme immunoassay specific for glycoprotein gG2 of HSV2. Results: HSV2 seroprevalence of FSW was 66.9% (91/136) of which only 14.3% (13/91) reported a history of genital infection. Age ≥35 years (OR 3.35, 95%CI 1.54-7.31, p=0.002), education ≥ grade 5 (OR 8.35, 95%CI 13.1- 64.7, p<0.001), lifetime sexual partners ≥ 200 (OR 5.33, 95% CI 2.3-12.36), p<0.001), duration of sex work ≥1 year (OR 4.5, 95% CI 2.09-9.65, p<0.001) and history of STI (OR 4.52, 95% CI 1.9-10.78, p<0.001) were statistically significant risk factors for high HSV2 prevalence. Consistent condom use by commercial partners in the last 3 months was not a significant protective factor in preventing HSV2 infection (OR 0.44, 95% CI 0.15-1.26, p = 0.12) in this cohort. Conclusions: The seroprevalence of HSV2 is high in this population and most of them did not report any genital infection. Multiple risk factors for higher prevalence were observed. Findings support the need for regular surveillance, monitoring of HSV-2 infection in high-risk populations, and expanding awareness of HSV-2 infection in the country.