The possible presence of Pneumocystis among healthy adults was examined by detecting Pneumocystis jirovecii-specific DNA in prospectively obtained oropharyngeal wash samples from 50 persons without underlying lung disease or immunosuppression. Pneumocystis carriage, defined by detecting Pneumocystis DNA by nested polymerase chain reaction in 2 independent analyses plus successful mitochondrial large subunit ribosomal RNA typing by direct sequencing, was found in 20% of cases. All carriers were asymptomatic, anti-HIV negative, and had normal total lymphocyte and CD4+ cell counts. A second sample obtained in the 6-month follow-up was positive in 2 of 9 available carriers. Genotype analysis showed different polymorphisms; 85A/248C (40%) and 85C/248C (30%) were most frequently observed. This study provides the first evidence that P. jirovecii DNA can be frequently detected in the respiratory tract of immunocompetent adults, which agrees with the hypothesis that the general population could be a reservoir and source of this infection.