Respiratory and fecal specimens from 111 patients with cystic fibrosis (CF) were cultured on 394 occasions to determine which site Pseudomonas aeruginosa colonizes first. By an enrichment and selection culture technique, P. aeruginosa was recovered from fecal cultures in 21 (42%) of 50 respiratory tract-positive patients but from only 3 (6.3%) of 48 respiratory tract-negative patients and from 1 (2.2%) of 45 control subjects. In 4 (22%) of the 18 patients who became culture-positive during the study, P. aeruginosa was recovered from a fecal specimen before respiratory colonization. Only 1 of these 4 became persistently colonized with P. aeruginosa in the respiratory tract; however, the serotype of the respiratory isolate was different from that of the fecal isolate. P. aeruginosa is recovered rarely from the feces of patients with CF who are culture-negative in the respiratory tract. In patients with CF, primary colonization with P. aeruginosa probably occurs in the respiratory tract.