We obtained specimens from 128 HIV-1 seroconverters identified from 1995 through 1998 in a prospective cohort study of 1,209 HIV-negative injecting drug users (IDUs) in Bangkok, Thailand. Epidemiologic data indicated that parenteral transmission accounted for nearly all infections. HIV-1 DNA from the C2-V4 env region was sequenced, and phylogenetic analyses determined that 102 (79.7%) of the specimens were subtype E and 26 (20.3%) subtype B strains. All subtype B strains clustered with strains often referred to in previous studies as Thai B or B'. The interstrain nucleotide distance (C2-V4) within subtype E strains was low (mean, 6.8%), and pairwise comparisons with a prototype subtype E strain, CM244, showed limited divergence (mean, 5.6%). The subtype B stains showed greater interstrain divergence (mean, 9.2%) and were significantly divergent from the prototype B strain HIV-MN (mean, 13.0%; p < 0.0001). The subtype E strains had significantly lower mean V3 loop charge than did subtype B strains (p = 0.017) and, on the basis of analysis of amino acid sequences, were predicted to be predominantly (91%) non-syncytium-inducing (NSI), chemokine coreceptor CCR5-using (CCR5+) viruses. The subtype B strains had a higher mean V3 loop charge, and a smaller proportion (23%) were predicted to be NSI/CCR5+ viruses. This study demonstrates that most incident HIV1 infections among Bangkok IDUs are due to subtype E viruses, with a narrow spectrum of genetic diversity. The characterization of incident HIV-1 strains from 1995 to 1998 will provide important baseline information for comparison with any breakthrough infections that occur among IDUs in Bangkok who are participating in an HIV-1 vaccine efficacy trial initiated in 1999.