From 1994 to 1995 four presumably autochthonous malaria cases were diagnosed by blood smear microscopy in Evros Province, northern Greece. Alarmed by these unexpected infections a serological survey was performed from 1997 to 1999 in ten rural villages, including those where the malaria cases had occurred. Among the 1,102 blood samples examined, nine turned out to contain specific antibodies against plasmodial parasites as detected by indirect fluorescent antibody test, including two of the former patients. The remaining seven samples were taken from healthy individuals with no history of recent infection or of having travelled to endemic areas. A further 21 sera showed borderline reactivity with Plasmodium falciparum antigen. Although no retrospective examination of the blood specimens could be performed to confirm the serological results by direct parasite detection, we can conclude that at least the seropositive persons have actually undergone infection with malaria parasites but developed no or only mild clinical symptoms which went unnoticed. It is becoming obvious that even in European countries where climatic and vector conditions are favourable for the development of the parasite there is a potential risk of incidental malaria transmission by indigenous Anopheles mosquitoes.

By using different staining techniques, 479 stool specimens from 212 diarrheic patients with AIDS were examined for microsporidian spores. Calcofluor fluorescence staining of 119 specimens revealed fluorescent ovoid structures of microsporidian size. Staining of these samples according to the method of Weber et al. (R. Weber, R. T. Bryan, R. L. Owen, C. M. Wilcox, L. Gorelkin, and G. S. Visvesvara, N. Engl. J. Med. 326:161–166, 1992) with trichrome produced six specimens with pinkish spores containing the characteristic microsporidian belt-like structure. The 6 specimens were processed for transmission electron microscopy, as were another 21 specimens which did not present the belt-like structure after trichrome staining but which looked highly suspicious after fluorescence staining. In these 21 samples, only fungal spores and, particularly, bacterial Clostridium spores were demonstrated, whereas in the 6 samples diagnosed positive after trichrome staining, the existence of microsporidia could be verified by electron microscopy. Based on our observations, we propose that the belt-like structure seen with the Weber stains in microsporidian spores corresponds to structures existing in priming-stage spores. The results suggest that routine microscopical fecal diagnosis for microsporidian infection should include a screening by fluorescence staining and, subsequently, a confirmatory viewing of fluorescence-positive samples after trichrome staining.