BACKGROUND Highly active antiretroviral therapy (HAART) in HIV patients is considered successful when plasma viral load (VL) reaches < 50 copies/ml. However, many patients have a persistent VL of 50 to 1000 copies/ml, and treatment guidelines do not recommend genotypic resistance testing at these levels because of poor performance. The aim of this study was to evaluate the usefulness of a concentration technique for HIV-1 sequencing in samples with < 1000 copies/ml, and determine the virological consequences of HAART treatment changes guided by resistance testing in this scenario. METHODS Observational study performed in 51 patients with plasma VL between 50 and 1000 copies/m; 27 patients had these levels for at least 12 consecutive months. Prior to RNA extraction, virions were concentrated from 3-ml plasma samples and then genotyped following standard procedures. RESULTS Forty-seven of the 51 samples were successfully sequenced, resulting in a sensitivity of 92%. Among these 47 patients, 27 showed a persistent viral load of 50-1000 copies/ml for 12 months, and 20 patients achieved undetectable viral load following the genotype-guided HAART change (intention-to-treat analysis: NC = F; 20 of 27 [74.1%]; on-treatment analysis: 20 of 23 [86.9%]). CONCLUSIONS We report a simple method for genotype sequencing in patients with persistent low-level viremia that allowed a modification of the HAART regimen leading to undetectable plasma viremia.