PIP: HIV-1 vaccine candidates being tested in the US and Europe are mainly based upon subtype B strains. Worldwide, however, there are multiple strains of HIV against which vaccines must also be developed. The authors present methodology and findings from an investigation into which subtypes of HIV-1 are present in an urban center in southern India. Lymphocyte and serum samples were collected from HIV-1 infected patients attending an outpatient screening clinic at Apollo Hospital, a private urban hospital in Hyderabad, Andhra Pradesh. Sera were tested and confirmed with ELISA and Western blot for the presence of antibody to HIV-1, with peripheral blood leukocytes obtained from five of the patients through density gradient centrifugation of fresh blood and subsequently shipped at -70 degrees Celsius to Johns Hopkins University where they were subjected to polymerase chain reaction and neutralization assays. No information was available about when the individuals became infected with HIV-1 and only limited clinical and risk factor data were available. In contrast to previous reports of exclusively subtype C strains in India, the study found and presents the first published evidence of the presence of subtype B HIV-1 in the country. The sequences identified in the study appear to be more similar to subtype B isolates from North America and Europe than those reported from Thailand, and are distinct from a subtype B sequence collected in 1992 from Vellore in southeast India. The presence of neutralizing activity against subtype B strains from sera matched with the phylogenetic analysis provides strong evidence for the presence of HIV-1 subtype B infection in India. It would not be surprising if additional HIV-1 subtypes were detected in India given the frequent travel which occurs between India and Europe, North America, Africa, and other areas of Asia including Thailand. These results underscore the necessity for a comprehensive and nationwide analysis of HIV-1 strain variation throughout India in the interest of developing and disseminating an effective vaccine against HIV-1 in India.