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We retrospectively screened 1836 men who have sex with men (MSM) participating in the Amsterdam Cohort Studies (1984–2003) for hepatitis C virus (HCV) antibodies. HCV incidence was 0.18/100 person-years (PY) in HIV-positive MSM (8/4408 PY [95% confidence interval [CI], 0.08–0.36]) but was 0/100 PY in MSM without HIV (0/7807 PY [95% CI, 0.00–0.05]). After 2000, HCV incidence among HIV-positive men increased 10-fold to 0.87/100 PY (5/572 PY [95% CI, 0.28–2.03]). Additional hospital cases (n=34) showed that MSM in Amsterdam who acquired HCV infection after 2000 reported high rates of ulcerative sexually transmitted infections (59%) and rough sexual techniques (55%), denied injection drug use, and were mainly infected with the difficult-to-treat HCV genotypes 1 (56%) and 4 (36%). Phylogenetic analysis showed 3 monophyletic clusters of MSM-specific HCV strains. The emergence of an …