Chronic diarrhea accompanied by weight loss is a common and often debilitating problem associated with human immunodeficiency virus (HIV) infection. Enterocytozoon bieneusi, a newly identified species of the phylum of protozoa, Microspora, has been reported associated with chronic diarrhea and wasting in 11 acquired immunodeficiency syndrome (AIDS) patients in the United States, Europe, and Africa. Diagnosis has been based solely on the ultrastructural identification of this small, intracellular parasite in bowel biopsies. Seventy-one small bowel biopsies from 67 homosexual AIDS and AIDS-related complex patients with chronic diarrhea and with no pathogens identified by light microscopy on paraffin sections, were embedded in plastic and studied by light and transmission electron microscopy. Enterocytozoon bieneusi microsporidiosis was diagnosed by electron microscopy in 20 (22 biopsies) of the patients. More jejunal biopsies (16 of 36) were positive than duodenal biopsies (six of 35). Parasites and spores were clearly visible at the light microscopic level in the semi-thin plastic sections from 17 and 21 of the biopsies, respectively. In retrospect, parasites could be identified by light microscopy in standard hematoxylin and eosin-stained paraffin sections. Infection was confined to enterocytes covering the villi, especially the tips, and was associated with villous atrophy and cell degeneration, necrosis, and sloughing. Release of spores into the bowel lumen was evident. Colorectal biopsies from two of the patients with small bowel microsporidiosis were negative for microsporidia. Enterocytozoon bieneusi infection of the small bowel may be an important cause of diarrhea in HIV-infected persons. © 1990.