Pneumocystis pneumonia has long been recognized as a cause of morbidity and mortality in immunocompromised populations, particularly those with HIV infection. Pneumocystis colonization-that is, detection of the organism or its DNA, without signs or symptoms of pneumonia-has recently been described, and accumulating evidence suggests that it may be an important clinical phenomenon. Sensitive molecular techniques such as polymerase chain reaction are frequently used to identify Pneumocystis colonization. Low levels of Pneumocystis in the lungs may stimulate pulmonary inflammation and may play a role in the development of lung diseases such as chronic obstructive pulmonary disease. In this review, we discuss evidence for the occurrence of Pneumocystis colonization in animals as well as the epidemiology and risk factors for Pneumocystis colonization in various human populations. We also evaluate the clinical significance of Pneumocystis colonization and its relationship to lung disease.