Microsporidia in the small intestine of HIV-infected patients. A new diagnostic technique and a new species.

OBJECTIVES: To determine whether microsporidian infections occur in Australian patients infected with human immunodeficiency virus (HIV), to assess the incidence, and to discuss microscopic detection methods. DESIGN AND PATIENTS: 180 consecutive HIV-infected patients (109 with chronic diarrhoea and 71 with other indications) underwent upper gastrointestinal tract endoscopy and pinch biopsies of the second part of the duodenum. The biopsies were handled by a protocol: four levels, with haematoxylin and eosin stain (H&E) at each level, periodic acid Schiff reagent after diastase (DiPAS) and auramine stain at the second level, and Warthin-Starry (WS) stain and cytomegalovirus early antigen immunoperoxidase study at the third level. Electron microscopy was carried out on samples from the first 95 patients, and thereafter from selected patients.
SETTING: The patients came from the HIV Medicine Unit of a teaching hospital and from the practice of a gastroenterologist. MAIN OUTCOME MEASURES: Diagnosis of microsporidia was to based on the H&E stain, with electron microscopy as the definitive test because the microsporidia are often difficult to see with H&E. Empirically, the WS stain was found to stain the microorganisms and it replaced electron microscopy during the study as the screening diagnostic test.
RESULTS: Microsporidia were present in 36 of the 109 patients with diarrhoea (33%) and one of 71 patients without diarrhoea. The WS stain in all cases showed developing spores in the enterocytes and in four cases in macrophages as well. The H&E stain showed non-specific duodenitis and was not diagnostic in some cases. Electron microscopy on samples from the first 95 consecutive patients showed 100% concordance with the WS stain. In 33 cases, electron microscopy showed the multinucleated plasmodia and the spores of Enterocytozoon bieneusi and in the four cases confirmed the spores in macrophages and showed a new Encephalitozoon-like species with a septate parasitophorous vacuole. Other causes of duodenal infection were cytomegalovirus (11 cases), mycobacteria (8), cryptosporidia (8) and Giardia lamblia (5).
CONCLUSION: E. bieneusi was the commonest microorganism found in our series of 180 consecutive patients. The actual prevalence of the two microsporidia species within the HIV-positive population and general community awaits further study. The WS stain provides a sensitive diagnostic test for the presence of E. bieneusi and the new Encephalitozoon-like species, avoiding the cost and potential sampling error of electron microscopy. The detailed ultrastructure and taxonomy of the new species requires further study.