This work was aimed to study the HIV-1 subtype B epidemics in the Basque Country, Spain. 1727 HIV-1 subtype B sequences comprising protease and reverse transcriptase (PR/RT) coding regions, sampled between 2001 and 2008, were analyzed. 156 transmission clusters were detected by means of phylogenetic analyses. Most of them comprised less than 4 individuals and, in total, they included 441 patients. Six clusters comprised 10 or more patients and were further analyzed in order to study their origin and diversification. Four clusters included men who had unprotected homosexual sex (MSM), one group was formed by intravenous drug users (IDUs), and another included both IDUs and people infected through unprotected heterosexual sex (HTs). Most of these clusters originated from the mid-1980s to the mid-1990s. Only one cluster, formed by MSM, originated after 2000. The time between infections was significantly lower in MSM groups than in those containing IDUs (P-value < 0.0001). Nucleoside RT and non-nucleoside RT inhibitor (NRTI and NNRTI)-resistance mutations to antiretroviral treatment were found in these six clusters except the most recent MSM group, but only the IDU clusters presented protease inhibitor (PI)-resistance mutations. The most prevalent mutations for each inhibitor class were PI L90M, NRTI T215D/Y/F, and NNRTI K103N, which were also among the most prevalent resistant variants in the whole dataset. In conclusion, while most infections occur as isolated introductions into the population, the number of infections found to be epidemiologically related within the Basque Country is significant. Public health control measures should be reinforced to prevent the further expansion of transmission clusters and resistant mutations occurring within them.