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Background: Plasmodium ookinete surface proteins as post-fertilization target antigens are potential malaria transmission-blocking vaccine (TBV) candidates. Putative secreted ookinete protein 25 (PSOP25) is a highly conserved ookinete surface protein, and has been shown to be a promising novel TBV target. Here, we further investigated the TBV activities of the full-length recombinant PSOP25 (rPSOP25) protein in Plasmodium berghei, and characterized the potential functions of PSOP25 during the P. berghei life-cycle. Methods: We expressed the full-length P. berghei PSOP25 protein in a prokaryotic expression system, and developed polyclonal mouse antisera and a monoclonal antibody (mAb) against the recombinant protein. Indirect immunofluorescence assay (IFA) and Western blot were used to test the specificity of antibodies. The transmission-blocking (TB) activities of antibodies were evaluated by the in vitro ookinete conversion assay and by direct mosquito feeding assay (DFA). Finally, the function of PSOP25 during Plasmodium development was studied by deleting the psop25 gene. Results: Both polyclonal mouse antisera and anti-rPSOP25 mAb recognized the PSOP25 proteins in the parasites, and IFA showed the preferential expression of PSOP25 on the surface of zygotes, retorts and mature ookinetes. In vitro, these antibodies significantly inhibited ookinetes formation in an antibody concentration-dependent manner. In DFA, mice immunized with the rPSOP25 and those receiving passive transfer of the anti-rPSOP25 mAb reduced the prevalence of mosquito infection by 31.2 and 26.1%, and oocyst density by 66.3 and 63.3%, respectively. Genetic knockout of the psop25 gene did not have a detectable impact on the asexual growth of P. berghei, but significantly affected the maturation of ookinetes and the formation of midgut oocysts. Conclusions: The full-length rPSOP25 could elicit strong antibody response in mice. Polyclonal and monoclonal antibodies against PSOP25 could effectively block the formation of ookinetes in vitro and transmission of the parasites to mosquitoes. Genetic manipulation study indicated that PSOP25 is required for ookinete maturation in P. berghei. These results support further testing of the PSOP25 orthologs in human malaria parasites as promising TBV candidates. [ABSTRACT FROM AUTHOR]

Background: Transmission-blocking vaccines (TBVs) are a promising strategy for malaria control and elimination. However, candidate TBV antigens are currently limited, highlighting the urgency of identifying new antigens for TBV development. Methods: Using a combination of bioinformatic analysis and functional studies in the rodent malaria model Plasmodium berghei, we identified a conserved Plasmodium protein PbPH (PBANKA_041720) containing a pleckstrin homology (PH) domain. The expression of PbPH was detected by Western blot and indirect immunofluorescence assay (IFA). The function of PbPH was tested by genetic knockout. The TB activity was confirmed by in vitro ookinete conversion assay and mosquito feeding. Results: PbPH was detected in Western blot as highly expressed in sexual stages (gametocytes and ookinetes). IFA revealed localizations of PbPH on the surface of gametes, zygotes, and ookinetes. Deletion of the pbph gene did not affect asexual growth, but significantly reduced the formation of gametocytes, ookinetes, and oocysts, indicating that PbPH protein is required for parasite sexual development. Recombinant PbPH expressed and purified from bacteria elicited strong antibody responses in mice and the antibodies significantly inhibited exflagellation of male gametocytes and formation of ookinetes in a concentration-dependent manner. Mosquito feeding experiments confirmed that mosquitoes fed on mice immunized with PbPH had 13 % reduction in the prevalence of infection and almost 48 % reduction in oocyst density. Conclusions: Pbph is a highly conserved Plasmodium gene and is required for parasite sexual development. PbPH protein is expressed on the surface of gametes and ookinetes. Immunization of mice against the recombinant PbPH protein induced strong antibody responses that effectively reduced the formation of male gametes and ookinetes in vitro and blocked transmission of the parasites to mosquitoes. These results highlight PbPH as a potential TBV candidate that is worth future investigations in human malaria parasites. [ABSTRACT FROM AUTHOR]