Microsporidia spore surface proteins are an important, under investigated aspect of spore/host cell attachment and infection. For comparison analysis of surface proteins, we required an antibody control specific for an intracellular protein. An endoplasmic reticulum-associated heat shock protein 70 family member (Hsp70; ECU02_0100; “C1”) was chosen for further analysis. DNA encoding the C1 hsp70 was amplified, cloned and used to heterologously express the C1 Hsp70 protein, and specific antiserum was generated. Two-dimensional Western blotting analysis showed that the purified antibodies were monospecific. Immunoelectron microscopy of developing and mature E. cuniculi spores revealed that the protein localized to internal structures and not to the spore surface. In spore adherence inhibition assays, the anti-C1 antibodies did not inhibit spore adherence to host cell surfaces, whereas antibodies to a known surface adhesin (EnP1) did so. In future studies, the antibodies to the ‘C1’ Hsp70 will be used to delineate spore surface protein expression. 1. Introduction Microsporidia are spore-forming, obligate intracellular divergent fungi with an extensive host range that includes most vertebrates and invertebrates. Although the first species of microsporidia was described over 150 years ago, microsporidiosis was rarely diagnosed in humans prior to the AIDS pandemic. Today, microsporidia are recognized as opportunistic pathogens of humans [1]. Most microsporidia infections in humans are thought to arise via the fecal-oral route. Ingestion of the environmentally stable spores leads to primary infection in the small intestine where replication of the organisms results in destruction of the epithelium. Therefore, the most common clinical manifestations of microsporidiosis are self-limiting diarrhea in immunocompetent individuals and persistent diarrhea perhaps leading to a wasting syndrome in the immunocompromised [2]. All microsporidia possess a unique invasion apparatus known as the polar tube or polar filament, which must be discharged in order to infect the host cell. Upon extrusion, the polar tube penetrates the host cell plasma membrane and allows the passage of infectious sporoplasm from the spore through the hollow polar tube into the host cell cytoplasm where replication occurs. We hypothesize that infection of the host cell is facilitated by adherence of the microsporidia spore to the host cell surface prior to or during the activation process. Our previous studies have demonstrated that microsporidia spores of the genus Encephalitozoon adhere to the
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