Early intracellular development in vitro of the cyst-forming protozoon Sarcocystis singaporensis and the influence of a monoclonal antibody on invasion, intracellular localization, and development of sporozoites were studied. As revealed by immunofluorescence using parasite-specific antibodies which labeled the parasitophorous vacuole membrane (PVM) and by ultrastructural analysis, sporozoites invaded pneumonocytes of the rat via formation of a parasitophorous vacuole (PV). About half of the sporozoites left this compartment within the first 8 h postinfection to enter the host cell cytosol. By semiquantitative analysis of acetyl-histone H4 expression of sporozoites, a marker linked to early gene expression of eukaryotic cells, we show (supported by ultrastructural analysis) that escape from the PV appears to be necessary for early intracellular development. More than 90% of sporozoites located in the cytosol expressed high levels of acetylated histone H4 in the nucleus, whereas only a quarter of the intravacuolar sporozoites exhibited a similar signal. As revealed by ultrastructural analysis, young schizonts all resided in the cytosol. Specific binding of a monoclonal antibody (11D5/H3) to sporozoites before invasion significantly enhanced their escape from the PV, whereas cell invasion itself remained unaffected. The antibody actually increased proliferation of the parasites in vitro, providing a further link between residence in the cytosol and successful intracellular development. Monoclonal antibody 11D5/H3 precipitated a major 58-kDa antigen from oocyst-sporocyst extracts and reacted with the cytoplasm and the surface of sporozoites in immunofluorescence assays. Collectively, the observed antibody-parasite interaction suggests the existence of a signaling event that influences intracellular development of Sarcocystis.Cyst-forming coccidia of the genus Sarcocystis are among the most prevalent parasites of livestock and are responsible for considerable economic losses (10, 15). Furthermore, recent outbreaks of Sarcocystis-induced disease among humans in tropical countries (3) underscore the increasing importance of these parasites for public health (31).A peculiar aspect of Sarcocystis infection has been known for a long time but has received little attention: the possible escape of sporozoites from the parasitophorous vacuole (PV) after invasion of host cells. Early observations in vivo and ultrastructural studies of infected cell cultures of bovine pulmonary artery endothelial cells and bovine monocytes revealed that Sarcocystis cruzi sporozoites as well as the resulting schizonts were located free in the host cell cytoplasm, i.e., not surrounded by a PV (11, 34). We have extended this observation to Sarcocystis singaporensis, a species that infects rats as intermediate hosts and specifically develops inside endothelial cells and pneumonocytes, a characteristic that renders it a suitable model for Sarcocystis infections in the laboratory (21). To date, however, nothing is known about a possible escape from ...
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