| Summary: | Cryptosporidium parvum, a primary cause of cryptosporidiosis in humans and livestock
worldwide, has a complex life cycle that includes an environmental oocyst stage, and stages of
merogony, gametogony, and sporogony that are completed in a single host. Development
within the host takes place in a protected intracellular but extracytoplasmic niche at the apical
surface of epithelial cells. The life cycle can be described as having alternating extracellular
invasive and intracellular replicative stages. With no effective chemotherapeutics,
understanding the mechanism of host cell invasion by this pathogen is paramount.
The first aim dissertation was to identify functions of sporozoite surface proteins and
rhomboid proteases (CpROMs) during motility and invasion of host cells. We demonstrate that
two CpROMs distinctively and collectively cleaved five thrombospondin-family proteins
(TSPs) and a mucin-like glycoprotein in a heterologous assay. Further, there was differential
co-expression and co-localization of the CpROMs and their substartes during in vitro life cycle
development; CpROM4 and CpTSP10 proteins colocalized to the anterior, middle and posterior
of sporozoites and in developing intracellular stages while CpROM5 and TRAP-C1 colocalized
to intact and non-intact oocyst walls, the anterior of sporozoites, and intracellular stages as
early as 2 h post infection. CpTSP7, also localized to the oocyst wall, the anterior and posterior
of sporozoites and intracellular stages from 6 h post infection. Similar to CpTSP10, CpTSP9
was not present in the oocyst wall; however, it was expressed in sporozoites and intracellular
stages from 6 h post infection. Short synthetic peptides derived from adhesive ectodomains in
thrombospondins including a TRAP-C1 apple domain (TAAP), thrombospondin type I
domains in CpTSP7 (7TS) and CpTSP9 (9TS), and a kringle domain in CpTSP10 (10K1) as
well as their corresponding antibodies demonstrated competitive and neutralization inhibition effect of C. parvum infection of host cells. Polyclonal antibodies against TAAP caused
sporozoites to agglutinate in a concentrationdependent manner, suggesting a contribution to
reduced infectivity. In conclusion, the specificity and expression profiles of CpROM4 and
CpROM5 indicate that they have distinct functions in shedding surface adhesins during
excystation, motility, invasion, and intracellular development.
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