In December 2004, infection with a strain of multidrug-resistant (MDR), dual-tropic human immunodeficiency virus (HIV)-1 was newly diagnosed in a man aged 46 years in New York City (NYC). The man (i.e., the index patient) had no history of antiretroviral treatment and reported having sex with multiple named and anonymous male partners, using crystal methamphetamine, and engaging in unprotected insertive and receptive anal intercourse. He had rapid progression to acquired immunodeficiency syndrome (AIDS) after experiencing signs and symptoms of acute HIV infection. The case was reported to the New York City Department of Health and Mental Hygiene (NYCDOH) in late January 2005 and has been described previously. This report describes the public health investigation of the index patient's reported contacts and a review of viral genetic sequencing (genotype) results from other HIV-infected patients in the NYC region to estimate the prevalence of this strain of HIV. The investigation, conducted by NYCDOH, Connecticut Department of Public Health, Aaron Diamond AIDS Research Center, New York State Department of Health, and CDC, identified three other patients with similar risk factors who engaged in high-risk sexual activity at the same time and in the same venues as the index patient and who were infected with a genotypically homologous strain of HIV. The findings demonstrate the usefulness of population-based reporting of HIV genotyping data to identify exact matches of new HIV mutations associated with drug resistance and to determine their characteristics and public health importance. The findings also demonstrate the continued risk for HIV transmission among men who have sex with men (MSM) through high-risk behaviors and the need to find effective methods to prevent HIV transmission in this population.