Diarrhea occurs commonly in African human immunodeficiency virus (HIV) infections. A case-control (HIV-positive vs. -negative) study of adults with diarrhea was done in Lusaka, Zambia, to determine the prevalence of intestinal infection by HEp-2 cell-adherent Escherichia coli. Adherent E. coli were more common in HIV-positive patients with acute diarrhea than among HIV-negative controls (60% vs. 33%) and were found significantly more often in HIV-positive patients with chronic diarrhea than among HIV-negative controls with chronic diarrhea (79% vs. 17%, P < .002). Adherent strains were found significantly more often among HIV-positive patients (69%) than in 22 asymptomatic subjects (36%, P < .02). The HEp-2 cell adherence of the E. coli strains did not show a common pattern. Adherent bacteria were also observed in colonic biopsies from 32% of Zambians with chronic diarrhea who underwent endoscopy. Adherent E. coli may be an important cause of HIV-associated diarrhea in Zambia.