Herpes Simplex Virus-2 (HSV-2) as an ulcerative mucocutaneous disease has been shown to facilitate the transmission of HIV infection. Therefore early identification and treatment of HSV-2 is fast becoming a strategy for preventing HIV transmission. This study aimed to determine the seroprevalence of HSV-2 among attendees of an HIV counselling and testing Centre (HCT), at a large HIV treatment centre in Lagos, Nigeria. It was a cross sectional study among clients presenting for voluntary HCT, who were counselled and screened for HIV and HSV-2. SPSS for window Version 17 was used for data analysis to carry out socio-demographic distribution of participants, χ 2 for association of variables with HSV-2 and the strength of the association determined by Pearson correlation. Two hundred and fifty eight participants enrolled for the study with 60% of participants being female. The most prominent age group was 24-29 years (22.6%) for female and 30-35 years for male (25.2%). The prevalence of HIV and HSV-2 among the participants was 29.8% and 9.7% respectively. In addition, 7.8% tested positive to both HSV and HIV, with HSV-2 prevalence among the participants that tested positive to HIV significantly higher than participants that tested negative to HIV (P < 0.001). The strength of the association by partial correlation demonstrated a medium correlation for HIV and HSV-2 and a weak correlation for age group and HSV-2 (0.359 and 0.187 respectively). The association between HSV-2 and HIV infection is attested by this study showing a high prevalence of HSV infection among HIV positive participants.