The molecular diversity and demographic characteristics among 976 anti-HIV-1-positive heterosexuals attending 15 sexually transmitted infection (STI) clinics participating in an unlinked anonymous HIV prevalence serosurvey in England and Wales during 1997-2000 were investigated. Subtypes were assigned by heteroduplex mobility assay or sequencing of the p17/p24 region of gag and the V3/V4 region of env and by sequencing of the protease gene. Overall, there was no significant change in the subtype distribution, with subtype C accounting for the majority (32%) of subtyped infections. Subtypes B (29%), A (12%), circulating recombinant forms (CRFs, 9%), unique recombinant forms (URFs, 8%), and subtypes D-H (8%) were also detected. Thirty-nine percent of infections in men were with subtype B, whereas subtype C was most common (38%) in women. Logistic regression analyses showed the relative risk (RR) of infection with a non-B subtype, compared with subtype B, to be greater in African-born individuals (RR = 28.9, P < 0.01), among newly diagnosed infections (RR = 3.4, P < 0.01), and in women (RR = 2.4, P < 0.01). These findings indicate a high level of genetic diversity among HIV-infected heterosexual STI clinic attendees in England and Wales. Recently, subtype C has become most prevalent, particularly in younger age groups, suggesting recent acquisition of this viral strain. The high proportion of non-B, CRF, and URF infections among UK-born individuals is consistent with mixing between migrants and UK-born individuals in England and Wales. As migration patterns change, continued monitoring of HIV genetic diversity will aid understanding of transmission patterns.