One of the most challenging worldwide obstacles limiting the efficacy of current antiretroviral therapy is the emergence of drug-resistant mutants of HIV-1. Drug resistance is reported to occur in 30-40% of treated patients and has important clinical and epidemiologic consequences. For individuals, drug resistance restricts subsequent antiretroviral treatment choices and can exhaust therapeutic options, culminating in HIV-1 disease progression and death. Drug-resistant variants are also transmitted when new infections occur, effectively multiplying the individual drug failures and creating a growing public health concern. Surveillance studies report that the prevalence of drug-resistant mutations among recently infected drug-nave individuals is 5-10% and is increasing. As such, transmission of drug resistance threatens to reverse the reductions in morbidity and mortality accomplished by antiretroviral therapy. New strategies are needed to prevent the emergence and spread of HIV-1 drug resistance. These strategies should be based on in-depth knowledge of the in vivo biology of drug resistance. Studies being conducted by the In Vivo Biology Group of the HIV Drug Resistance Program (DRP) have been specifically designed to provide such knowledge.Several groups have reported that viremia can still be detected in patients on currently recommended antiretroviral treatment regimens despite suppression of plasma HIV-1 RNA to <50 copies/ml, as determined by commercial assays (limit of quantification 50 copies/ml). Controversy exists about whether this persistent, low-level viremia is the consequence of activation of HIV-1 from latently infected cellular reservoirs or represents HIV-1 produced from new cycles of viral replication. This issue has important implications for strategies designed to eradicate HIV-1 infection or to further reduce HIV-1 replication to prevent drug resistance. HIV-1 RNA assays of greater sensitivity, accuracy, and precision are needed to address this issue. The DRP Virology Core has developed such an assay that is now being applied to characterize the prevalence, level, and change over time of persistent viremia in patients on antiretroviral regimens of differing potency. In addition, a prospective, randomized trial of lopinavir/ritonavir intensification of antiretroviral therapy in patients with viremia of 10-50 copies/ml is being enrolled to address whether increasing the potency of a regimen affects the level of persistent viremia.