The discovery of a 44% (44 out of 100) prevalence rate of HIV infection among female prostitutes working in brothels in Chiangmai in Thailand in June 1989, prompted this follow-up study in August to confirm the high prevalence rate and to look for risk factors for infection. We studied 238 female prostitutes working in 14 brothels and confirmed this high prevalence rate. Eighty-seven (36.5%) out of 238 prostitutes were found to be HIV-positive by enzyme-linked immunosorbent assay with IFA or Western blot confirmation. Logistic regressions found a significant association between HIV infection and frequency of sexual intercourse > 3 times per day [odds ratio (OR) = 2.82, 95% confidence interval (Cl) = 1.47–5.41], sexual service charge < 150 Baht (OR = 9.1, 95% Cl = 2.9–33.3), and post sexual cleansing with water alone (OR = 3.85, 95% Cl = 1.90–7.80). Of 56 women found seronegative in the June survey, 35 were re-tested in the August study. Seven (20%) of them were seropositive, giving an HIV seroconversion incidence rate of 10% per month. The findings of this study prompted intensive health education programmes among prostitutes, their customers, and owners of brothels.