Participant Protection The study protocol and informed consent documents were reviewed by the local ethics committees of the University of KwaZulu-Natal, the University of Cape Town, the University of the Witwatersrand in Johannesburg, and by the Prevention Sciences Review Committee (PSRC) of the Division of AIDS (DAIDS, National Institutes of Health, U.S.A.). The consent forms were translated into isiZulu and written informed consent is obtained at each stage of the study (screening, enrolment into HIV-negative cohort, enrolment into acute infection phase, and for sample storage). All women screened for participation, whether ultimately enrolled into the study cohort or not, receive HIV pre- and post-test counselling, risk reduction counselling, male and female condoms, access to clinical care, and treatment for sexually transmitted infections (STIs). HIV/STI risk reduction counselling, condom provision, and prevention education supplies are administered at each subsequent study visit. One-on-one counsel- ling was provided by a non-governmental organization which offered the same service in the public sector in the region, to be consistent with the services in the region. Overcoming one of the key challenges to doing HIV-related work in this context [11], participants who were HIV infected during the study were referred to the CAPRISA Antiretroviral Treatment (CAT) programme where they were offered ongoing care, and treatment for HIV when clinically eligible. Study Visits and Procedures Once enrolled into the HIV-negative observational cohort, the participants underwent a baseline evaluation that included: a HIV behaviour risk assessment, a clinical evaluation, blood collection for routine laboratory assessment and HIV status, and a specimen collection for sexually transmitted infection (STI) diagnosis. Following the baseline evaluation, participants in the HIV- negative cohort attend monthly follow-up evaluations for a maximum of 24 months. The monthly evaluations consist of a clinical evaluation, two HIV antibody rapid tests, HIV EIA, and pooled HIV-1 RNA testing (Amplicor v1.5: Roche Diagnostics, Rotkreuz, Switzerland). A urine dipstick is done on clinical suspicion of pregnancy, or if requested by the participant. In addition, a routine laboratory assessment, urinalysis, and STI screening are performed at 6-monthly intervals. Women from the HIV-negative cohort are diagnosed with acute HIV infection by (1) the detection of HIV-1 antibodies within 5 months of a previously negative HIV-1 test; or (2) evidence of HIV-1 viral replication in the absence of HIV-1 antibodies. While women in this cohort were screened monthly allowing for rapid identification of acute infection, a 5 months window period was allowed to enable to recruit from other prevention cohorts where follow-up was only quarterly. Time of infection is defined as the mid-point between the last HIV antibody negative test and the first HIV antibody positive test; or if a positive RNA PCR assay is available on the same date as a negative HIV EIA, the date of HIV infection is estimated at 14 days prior to the first positive RNA PCR assay. Staff Training and Infrastructure Development Given the extended scope and longitudinal nature of this study, CAPRISA was required to expand the size of the staff and the ability of the organization to manage a large cohort. After hiring additional clinicians, nurses, counsellors, and laboratory person- nel, all team members underwent extensive training in both good clinical practice (GCP) and in the particulars of this study with respect to the case report forms (CRFs) for documentation, the quality control plan, and Human Subjects Protections (HSP). CAPRISA 002 Study Furthermore, the capacity of the laboratory had to be increased to handle the large quantity of specimens related to the study. Data Management and Statistical Analysis All data is entered onto case report forms (CRFs) at the study sites and faxed into the CAPRISA Data Management Centre, using the DataFax system (Clinical DataFax Systems Inc., Ontario, Canada). This capability did not previously exist at site, and had to be implemented in order to collect the data for this study. Source documents and original CRFs are maintained on site while an electronic version of the CRF is maintained by the Data Management Centre. Quality assurance and quality control are maintained and checked at specified intervals. All analyses are conducted using the SAS statistical package version 9.1 (SAS Institute, Cary, NC, U.S.A.). Results Establishing the Cohort Between August 2004 and May 2005, 775 women who were classified as high-risk either by self-identification as sex workers or self-report of more than three sexual partners in the previous 3 months were screened for participation in the observational cohort. Recruitment was done at known FSW sites in the city, therefore the majority of these women (n = 193, 78.8%) were self- identified as sex workers. The cumulative number of women screened and enrolled over the entire 10-month period can be seen in Figure 1. Of the 775 women screened, 59.6% (95% CI 55.9%–62.8%) were found to be HIV-positive. The mean ages of the women who were screened and found to be HIV-positive and HIV negative were 29.3 years (range 16–58) and 34.2 years (range 18–58), respectively (p,0.001). The HIV prevalence among the screening participants changed dramatically over time from 83% in the first month of screening to 17% in the last month of screening (Figure 2). Of the 313 women who were HIV-negative, 245 HIV-negative women were eligible and agreed to participate in the HIV- negative cohort. Exclusion criteria for women who were uninfected at screening were as follows: less than three sexual partners in the previous 3 months (n = 22), women who were pregnant at the time of screening (n = 16), women who planned to relocate (n = 4), women younger than 18 years (n = 3), and women Figure 1. Cumulative HIV serostatus of screening participants over time. doi:10.1371/journal.pone.0001954.g001 PLoS ONE | www.plosone.org 3 April 2008 | Volume 3 | Issue 4 | e1954