Cross-sectional studies have suggested that infection with human immunodeficiency virus (HIV) type 1 could reduce the mitochondrial DNA (mtDNA) content of blood cells. We investigated mtDNA content in peripheral blood mononuclear cells (PBMCs) obtained from 36 antiretroviral therapy-naive documented HIV-1 seroconverters, before and after seroconversion. mtDNA content statistically significantly decreased 1 year after seroconversion and showed a nonsignificant decrease during the subsequent 4 years. These findings confirm that infection with HIV-1 may, itself, reduce mtDNA content, at least within PBMCs. This could have implications for the subsequent development of mitochondrial toxicities associated with the use of nucleoside analogue reverse-transcriptase inhibitors.