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. 2018 Nov 9;17(1):417.
doi: 10.1186/s12936-018-2566-0.

Specificity of the IgG antibody response to Plasmodium falciparum, Plasmodium vivax, Plasmodium malariae, and Plasmodium ovale MSP119 subunit proteins in multiplexed serologic assays

Jeffrey W Priest  1 , Mateusz M Plucinski  2   3 , Curtis S Huber  2 , Eric Rogier  2 , Bunsoth Mao  4 , Christopher J Gregory  5 , Baltazar Candrinho  6 , James Colborn  7 , John W Barnwell  2
Affiliations

Specificity of the IgG antibody response to Plasmodium falciparum, Plasmodium vivax, Plasmodium malariae, and Plasmodium ovale MSP119 subunit proteins in multiplexed serologic assays

Jeffrey W Priest et al. Malar J. .

Abstract

Background: Multiplex bead assays (MBA) that measure IgG antibodies to the carboxy-terminal 19-kDa sub-unit of the merozoite surface protein 1 (MSP119) are currently used to determine malaria seroprevalence in human populations living in areas with both stable and unstable transmission. However, the species specificities of the IgG antibody responses to the malaria MSP119 antigens have not been extensively characterized using MBA.

Methods: Recombinant Plasmodium falciparum (3D7), Plasmodium malariae (China I), Plasmodium ovale (Nigeria I), and Plasmodium vivax (Belem) MSP119 proteins were covalently coupled to beads for MBA. Threshold cut-off values for the assays were estimated using sera from US citizens with no history of foreign travel and by receiver operator characteristic curve analysis using diagnostic samples. Banked sera from experimentally infected chimpanzees, sera from humans from low transmission regions of Haiti and Cambodia (N = 12), and elutions from blood spots from humans selected from a high transmission region of Mozambique (N = 20) were used to develop an antigen competition MBA for antibody cross-reactivity studies. A sub-set of samples was further characterized using antibody capture/elution MBA, IgG subclass determination, and antibody avidity measurement.

Results: Total IgG antibody responses in experimentally infected chimpanzees were species specific and could be completely suppressed by homologous competitor protein at a concentration of 10 μg/ml. Eleven of 12 samples from the low transmission regions and 12 of 20 samples from the high transmission area had antibody responses that were completely species specific. For 7 additional samples, the P. falciparum MSP119 responses were species specific, but various levels of incomplete heterologous competition were observed for the non-P. falciparum assays. A pan-malaria MSP119 cross-reactive antibody response was observed in elutions of blood spots from two 20-30 years old Mozambique donors. The antibody response from one of these two donors had low avidity and skewed almost entirely to the IgG3 subclass.

Conclusions: Even when P. falciparum, P. malariae, P. ovale, and P. vivax are co-endemic in a high transmission setting, most antibody responses to MSP119 antigens are species-specific and are likely indicative of previous infection history. True pan-malaria cross-reactive responses were found to occur rarely.

Keywords: MSP119; Malaria; Multiplex; Serology; Specificity.

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Figures

Fig. 1
Fig. 1
Alignment of predicted Plasmodium spp. MSP119 protein sequences using COBALT [61]. Residues in the P. malariae sequence that differ from the Cameroon sequence of Birkenmeyer et al. [38] are shaded. Predicted protein sequences resulting from the oligonucleotides used in PCR amplification are underlined. The positions of residues conserved among all the presented MSP119 protein sequences are indicated in the consensus with divergent residues indicated by a dot. GenBank accession numbers are MH577181, P. ovale Nigeria I strain; MH577182, P. malariae China I strain; MH577183, P. malariae Greece I strain; MH577184, P. malariae Uganda I strain; and MH577185, P. malariae Guyana strain
Fig. 2
Fig. 2
Assessment of coupling efficiency using a dilution series of goat anti-GST IgG antibody. Goat anti-GST IgG antibody was diluted 1:1 × 103, 1:1 × 104, 1:5 × 104, 1:1 × 105, 1:5 × 105, 1:1 × 106, 1:5 × 106, and 1:1 × 107 in modified Buffer A lacking the E. coli extract. Following a 1-h incubation (50 μl/well) at room temperature, bound anti-GST antibody was detected with a biotinylated rabbit anti-goat IgG secondary antibody (1:500 dilution in Buffer B, 50 μl/well, 1 h at room temperature). Wells were developed with R-phycoerythrin-labelled streptavidin and read on a BioPlex 200 instrument (BioRad) as described in “Methods”
Fig. 3
Fig. 3
Antibody competition titration assays using homologous MSP119 proteins. Dilutions (1:400) of P. falciparum Lot 6 defined human serum or of sera from chimpanzees experimentally infected with either P. malariae (Klimatis), P. ovale (Alpert) or P. vivax (Duff) were incubated with the indicated concentrations of the homologous MSP119 competitor protein for 1 h at room temperature. Multiplex bead assays were performed as described in “Methods”, and the multiplex responses in MFI-bg units are plotted versus the competitor concentration
Fig. 4
Fig. 4
Antibody competition titration assays using MSP119 proteins from four Plasmodium species. A combined dilution (1:400 of each serum) containing sera from chimpanzees experimentally infected with either P. malariae (Klimatis), P. ovale (Alpert) or P. vivax (Duff) was incubated with the indicated concentrations of the MSP119 competitor protein for 1 h at room temperature. Competitor proteins used were: a P. falciparum MSP119; b P. malariae MSP119; c P. ovale MSP119; d P. vivax MSP119. Multiplex bead assays were performed as described in “Methods” and the multiplex response in MFI-bg units are plotted versus the competitor concentration. Multiplex responses are presented as a percentage of the assay results for the PBS control
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