Evaluation of DNA encoding acidic ribosomal protein P2 of Cryptosporidium parvum as a potential vaccine candidate for cryptosporidiosis

Abstract

The Cryptosporidium parvum acidic ribosomal protein P2 (CpP2) is an important immunodominant marker in C. parvum infection. In this study, the CpP2 antigen was evaluated as a vaccine candidate using a DNA vaccine model in adult C57BL/6 IL-12 knockout (KO) mice, which are susceptible to C. parvum infection. Our data show that subcutaneous immunization in the ear with DNA encoding CpP2 (CpP2-DNA) cloned into the pUMVC4b vector induced a significant anti-CpP2 IgG antibody response that was predominantly of the IgG1 isotype. Compared to control KO mice immunized with plasmid alone, CpP2-immunized mice demonstrated specific in vitro spleen cell proliferation as well as enhanced IFN-γ production to recombinant CpP2. Further, parasite loads in CpP2 DNA-immunized mice were compared to control mice challenged with C. parvum oocysts. Although a trend in reduction of infection was observed in the CpP2 DNA-immunized mice, differences between groups were not statistically significant. These results suggest that a DNA vaccine encoding the C. parvum P2 antigen is able to provide an effective means of eliciting humoral and cellular responses and has the potential to generate protective immunity against C. parvum infection but may require using alternative vectors or adjuvant to generate a more potent and balanced response.

Cryptosporidium parvum antigens induce mouse and human dendritic cells to generate Th1‐enhancing cytokines

“…The rCp23 (22), rCp40 (22), rCp17 (18) and rCpP2 (19,24) proteins were fused to a Schistosoma japonicum glutathione‐ S ‐transferase (GST) tag expressed from plasmid pGex4T‐2 in Escherichia coli BL21 cells following the manufacturer’s instructions. The GST fusion tag was cleaved with thrombin (GE Healthcare, Piscataway, NJ, USA), and then, thrombin was removed using pAmino Benzamidine‐Agarose (SIGMA # A7155).…”

“…Endotoxin was removed using Detoxi‐Gel Endotoxin Removing Columns (Thermo Fisher Scientific). rCpP2 was also expressed as a 6 × His fusion protein in pQE81 vector (Qiagen, Valencia, CA, USA) using E. coli DH5α cells (Invitrogen, Carlsbad, CA, USA) and purified as previously described (19,24). Protein concentrations were determined using the Micro BCA Protein assay (Thermo Fisher Scientific).…”

A review of the global burden, novel diagnostics, therapeutics, and vaccine targets for cryptosporidium

“…146 Other antigens being explored for vaccine use include P2 antigen, profilin, Cryptosporidium apyrase, Muc4, and Muc5. 143,147,148 …”

Treatment of Cryptosporidiosis

“…Evaluation of a DNA vaccine comprised of P2 (CpP2), an important immunodominant marker in C. parvum infection (Priest et al 2010), showed that CpP2-DNA followed with P2 protein (prime-boost), significantly increase antibody production over immunization with just the protein or CpP2-DNA alone. When challenged, reduction in oocysts production was not statistically significant, although a trend in reduction of infection was observed in the CpP2-DNA-immunized mice (Benitez et al 2011).…”