Sequence analysis of HIV-1 from 440 therapy-naive individuals included within the CASCADE study, who seroconverted within 18 months of the last negative test, identified 65 persons infected with a strain carrying resistance-associated mutations. Population-based sequencing was performed for 20 of these individuals during the therapy-free follow-up period. The median time of follow-up was 15 months (interquartile range from 10 to 23 months). Of these individuals, 12 showed subsequent evolution at the resistance positions, whereas the virus of 8 people was stable during this period. In the reverse transcriptase (RT) gene, the drug-resistant 215Y or 215F codons evolved to alternative codons in all six cases, 70R reverted to the wild-type 70K in 3 of the 4 individuals, 67N evolved only in 1 of 4 patients to a wild-type 67D, 215S evolved to wild-type 215T in 1 of 3 patients, 219N evolved to 219K in 1 of 2 patients, and one patient with 184V reversed to the wild-type 184M. The 181C variant evolved to the wild-type 181Y in 1 of 2 individuals. These codon changes were caused by single nucleotide mutations. No evolution was observed for other RT mutations: 41L, 69D, 69N, 190S, 210W, 215L, 215C, 215E and 219Q. In the protease gene, resistance mutations 84V and 90M were stable in 2 individuals. Comparing the CD4+ T-cell count of the 12 evolving versus the 8 stable cases revealed no statistically significant difference at the date of the first sequence following seroconversion. Interestingly, a lower CD4+ T-cell count was observed in the group without evolution at the second sequence time point (P = 0.043). No difference in HIV-1 RNA load was observed. These results, together with the apparent pressure to mutate at the resistance-associated positions exemplify the decreased fitness of viruses carrying 21 5Y/F, 70R or 184V.