A protocol for the handling of small intestinal biopsies from HIV-infected patients is presented. This protocol includes the Warthin-Starry stain for the detection of microsporidia. This stain has proved a reliable and sensitive diagnostic technique for microsporidial infections as it stains both Enterocytozoon bieneusi and Septata intestinalis in duodenal enterocytes. Because the stain demonstrates Septata intestinalis in lamina propria macrophages as well as enterocytes, it allows for the practical differentiation of these two microsporidial infections. The Warthin-Starry stain has also demonstrated Septata intestinalis in nasal and colonic biopsies in some of these patients. Since the completion of an earlier study, a further 40 cases of Enterocytozoon bieneusi and three cases of Septata intestinalis have been diagnosed in just over 240 consecutive duodenal biopsies from HIV positive patients presenting with diarrhoea and other gastrointestinal complaints. Other opportunistic infections include cytomegalovirus in four cases, mycobacteria in eight cases, cryptosporidia in nine cases, giardia in four cases and Isospora belli in one case. Since the ratio of these opportunistic infections has remained much the same as in the previous study of 180 consecutive duodenal biopsies, we suggest that these rates may reflect the actual prevalence of microsporidial infections in AIDS patients in Sydney, Australia.