Plasmodium vivax gametocyte proteins, Pvs48/45 and Pvs47, induce transmission-reducing antibodies by DNA immunization.

Malaria transmission-blocking vaccines (TBV) aim to interfere with the development of the malaria parasite in the mosquito vector, and thus prevent spread of transmission in a community. To date three TBV candidates have been identified in Plasmodium vivax; namely, the gametocyte/gamete protein Pvs230, and the ookinete surface proteins Pvs25 and Pvs28. The Plasmodium falciparum gametocyte/gamete stage proteins Pfs48/45 and Pfs47 have been studied as TBV candidates, and Pfs48/45 shown to induce transmission-blocking antibodies, but the candidacy of their orthologs in P. vivax, Pvs48/45 (PVX_083235) and Pvs47 (PVX_083240), for vivax TBV have not been tested. Herein we investigated whether targeting Pvs48/45 and Pvs47 can inhibit parasite transmission to mosquitoes, using P. vivax isolates obtained in Thailand. Mouse antisera directed against the products from plasmids expressing Pvs48/45 and Pvs47 detected proteins of approximately 45- and 40-kDa, respectively, in the P. vivax gametocyte lysate, by Western blot analysis under non-reducing conditions. In immunofluorescence assays Pvs48/45 was detected predominantly on the surface and Pvs47 was detected in the cytoplasm of gametocytes. Membrane feeding transmission assays demonstrated that anti-Pvs48/45 and -Pvs47 mouse sera significantly reduced the number of P. vivax oocysts developing in the mosquito midgut. Limited amino acid polymorphism of these proteins was observed among 27 P. vivax isolates obtained from Thailand, Vanuatu, and Colombia; suggesting that polymorphism may not be an impediment for the utilization of Pvs48/45 and Pvs47 as TBV antigens. In one Thai isolate we found that the fourth cysteine residue in the Pvs47 cysteine-rich domain (CRD) III (amino acid position 337) is substituted to phenylalanine. However, antibodies targeting Pvs47 CRDI-III showed a significant transmission-reducing activity against this isolate, suggesting that this substitution in Pvs47 was not critical for recognition by the generated antibodies. In conclusion, our results indicate that Pvs48/45 and Pvs47 are potential transmission-blocking vaccine candidates of P. vivax.

Blocking of transmission to mosquitoes by antibody to Plasmodium vivax malaria vaccine candidates Pvs25 and Pvs28 despite antigenic polymorphism in field isolates.

We have previously demonstrated that mouse antisera against yeast-produced recombinant forms of the ookinete surface proteins of Plasmodium vivax (Pvs25 and Pvs28) blocks transmission of the homologous P. vivax (Sal I strain). In this study, we developed mouse and rabbit antisera against Pvs25 and Pvs28 and evaluated the efficacy of these vaccine candidates against natural isolates of P. vivax in Thailand. Although both Pvs25 and Pvs28 genes are polymorphic, sera from mice immunized using alum adjuvant completely inhibited oocyst development for most human isolates, whereas sera from rabbits immunized with either alum or Freund's adjuvant were partially inhibitory. All inhibition occurred in an antibody dose dependent fashion. Data from this study clearly demonstrates that antibodies raised against Sal I-based vaccines overcome the genetic polymorphism of Pvs25 and Pvs28 present in natural isolates of P. vivax, suggesting the wide range applicability of Sal I based vaccines.

Serum antibodies induced by intranasal immunization of mice with Plasmodium vivax Pvs25 co-administered with cholera toxin completely block parasite transmission to mosquitoes.

Transmission-blocking vaccines (TBVs) targeting ookinete surface proteins expressed on sexual-stage malaria parasites are considered one promising strategy for malaria control. To evaluate the prospect of developing non-invasive and easy-to-administer mucosal malaria transmission-blocking vaccines, mice were immunized intranasally with a Plasmodium vivax ookinete surface protein, Pvs25 with a mucosal adjuvant cholera toxin (CT). Immunization induced significant serum IgG with high IgG1/IgG2a ratio (indicative of Th-2 type immune response). Feeding Anopheles dirus mosquitoes with mixtures of immune sera and gametocytemic blood derived from vivax-infected volunteer patients in Thailand significantly reduced both the number of midgut oocysts as well as the percentage of infected mosquitoes. The observed transmission-blocking effect was dependent on immune sera dilution. This study demonstrates for the first time that the mucosally induced mouse immune sera against a human malaria ookinete surface protein can completely block parasite transmission to vector mosquitoes, suggesting the possibility of non-invasive mucosal vaccines against mucosa-unrelated important pathogens like malaria.

Comparative evaluation of selected diagnostic assays for the detection of IgG and IgM antibody to Orientia tsutsugamushi in Thailand.

We compared the performance of 2 commercially available dipstick assays, 2 enzyme-linked immunosorbent assays (ELISAs), and an indirect immunofluorescent antibody (IFA) assay for the diagnosis of scrub typhus, using the indirect immunoperoxidase (IIP) test as the reference standard. The dipstick assays were the Integrated Diagnostics (Baltimore, MD) Dip-S-Ticks Scrub Recombinant (r56) dipstick test (INDX assay) and the PanBio (Brisbane, Australia) Scrub Typhus IgM and IgG Rapid Immunochromatographic test (PanBio assay). One of the ELISAs used pooled cell lysates of Karp, Kato, and Gilliam strain Orientia tsutsugamushi as antigen (pooled-antigen ELISA), and the other used a recombinant r56 protein as the antigen (recombinant ELISA). With a panel of 123 positive and 227 negative sera, sensitivity and specificity of the assays were as follows: INDX assay, IgG, 60% and 95%, IgM, 60% and 97%; PanBio assay, IgG, 94% and 96%, IgM, 83% and 93%; IFA (1:400 cutoff), IgG, 91% and 96%, IgM, 85% and 98%; pooled-antigen ELISA, IgG (1:1600 cutoff), 97% and 89%, IgM (1:400 cutoff), 94% and 91%; recombinant ELISA, IgG (1:1600 cutoff), 97% and 92%, IgM (1:400 cutoff), 93% and 94%. Because of its excellent performance and use of a standardized, commercially available antigen, the recombinant ELISA is suitable for use in a diagnostic laboratory, where it may be able to replace the IFA and IIP assays. In contrast, the PanBio dipstick assay was easy to perform and did not require sophisticated equipment, making it suitable for use in rural areas where more sophisticated diagnostic tests such as the ELISA and IFA may not be available.