Evaluation of Immunogenicity of an Oral Salmonella Vaccine Expressing Recombinant Plasmodium berghei Merozoite Surface Protein-1

Abstract

A repetitive region of Plasmodium berghei merozoite surface protein-1 (PbMSP-1) was expressed as a fusion protein with either maltose binding protein or the B subunit of heat-labile enterotoxin from Escherichia coli. Vaccination of mice with the fusion proteins indicates that this region of PbMSP-1 is antigenic as evidenced by an antibody response. The fusion proteins were also expressed in Salmonella and mice were orally immunized with the recombinant Salmonella. Some of the vaccinated mice survived a challenge with P. berghei blood-stage parasites without developing parasitemia. All control mice became patent and succumbed to the challenge infection. This partial protection was also observed with purified recombinant protein and was independent of the adjuvant used. Mice immunized with recombinant Salmonella showed either extremely low or no antibody response to PbMSP-1, suggesting that cell-mediated immunity is important for the observed protection. These studies show that it is feasible to develop a cost effective oral vaccine against the blood stage of the malarial parasite.

The complete sequence of Plasmodium berghei merozoite surface protein-1 and its inter- and intra-species variability

“…The size-fractionated DNA was ligated into HindIII of pBluescriptII KS + (Stratagene). Following transformation, colony lifts were screened with a radioactive probe derived from previously cloned fragment of PbMSP-1 [20]. A positive recombinant clone with an insert of 4.4 kb, designated as pPb230-4.4, was obtained.…”

“…A portion of MSP-1 from P. berghei was previously cloned and sequenced [20] and that clone was used as a probe in cloning the complete PbMSP-1 gene reported herein. PbMSP-1 exhibits similar conserved and variable blocks as MSP-1 from other species, but differs in the amount of repetitiveness and homogeneity of the tandem repeats found within the variable blocks.…”

Alternative approaches to vaccine delivery

“…Since then, various approaches have been taken to improve the immunogenicity of CSP delivered by Salmonella, [52][53][54][55] and to examine how well it may deliver other Plasmodium antigens. [56][57][58][59][60] There are also concomitant improvements in the general technology of bacterial vectors, with new strategies for the expression and presentation of recombinant proteins together with safer bacterial strains. 61 In recent years, food grade lactic acid bacteria have emerged as live vaccine vehicles for oral delivery.…”

Plasmodial serine repeat antigen homologues with properties of schizont cysteine proteases

“…Parasite proteins were separated by SDS-PAGE under reducing conditions and blotted onto an Immobilon-P membrane (Millipore), and immunoblotting was performed with control antisera generated against acrylamide alone (1:500 dilution; lane 1), and with antisera generated against SERA vinckei -1 diluted at 1:2000 (lane 2), 1:1000 (lane 3), and 1:500 (lane 4). Membranes were washed, incubated with alkaline phosphatase-conjugated goat anti-mouse serum, washed, and evaluated by colorimetric detection as previously described [20]. For both immunoblots, molecular weight standards (kDa) are labeled, and the arrow identifies the 105 kDa protein recognized by the anti-SERA vinckei -1 serum.…”

“…The protein was expressed in BL21(DE3)pLysS strain E. coli and affinity purified as previously described [15]. Purified SERA vinckei -1 was then electrophoresed on an SDS-PAGE gel, and CD-1 mice were immunized intraperitoneally with 50 -100 vg of the protein in gel slices as previously described [20]. Control mice were immunized in the same fashion with gel slices not containing protein.…”