Browsing by Author "Geneshan, H."
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Item Comparison of the impact of allelic polymorphisms in PfAMA1 on the induction of T Cell responses in high and low malaria endemic communities in Ghana(Malaria Journal, 2021) Ofori, E.A.; Tetteh, J.K.A.; Frimpong, A.; Geneshan, H.; Belmonte, M.; Peters, B.; Villasante, E.; Sedegah, M.; Ofori, M.F.; Kusi, K.A.Background: Malaria eradication requires a combined efort involving all available control tools, and these eforts would be complemented by an efective vaccine. The antigen targets of immune responses may show polymor phisms that can undermine their recognition by immune efectors and hence render vaccines based on antigens from a single parasite variant inefective against other variants. This study compared the infuence of allelic polymor phisms in Plasmodium falciparum apical membrane antigen 1 (PfAMA1) peptide sequences from three strains of P. fal ciparum (3D7, 7G8 and FVO) on their function as immunodominant targets of T cell responses in high and low malaria transmission communities in Ghana. Methods: Peripheral blood mononuclear cells (PBMCs) from 10 subjects from a high transmission area (Obom) and 10 subjects from a low transmission area (Legon) were tested against 15 predicted CD8+T cell minimal epitopes within the PfAMA1 antigen of multiple parasite strains using IFN-γ ELISpot assay. The peptides were also tested in simi lar assays against CD8+enriched PBMC fractions from the same subjects in an efort to characterize the responding T cell subsets. Results: In assays using unfractionated PBMCs, two subjects from the high transmission area, Obom, responded pos itively to four (26.7%) of the 15 tested peptides. None of the Legon subject PBMCs yielded positive peptide responses using unfractionated PBMCs. In assays with CD8+enriched PBMCs, three subjects from Obom made positive recall responses to six (40%) of the 15 tested peptides, while only one subject from Legon made a positive recall response to a single peptide. Overall, 5 of the 20 study subjects who had positive peptide-specifc IFN-γ recall responses were from the high transmission area, Obom. Furthermore, while subjects from Obom responded to peptides in PfAMA1 from multiple parasite strains, one subject from Legon responded to a peptide from 3D7 strain only. Conclusions: The current data demonstrate the possibility of a real efect of PfAMA1 polymorphisms on the induc tion of T cell responses in malaria exposed subjects, and this efect may be more pronounced in communities with higher parasite exposure.Item Measuring naturally acquired immune responses to candidate malaria vaccine antigens in Ghanaian adults(2011-06-20) Dodoo, D.; Hollingdale, M.R.; Anum, D.; Koram, K.A.; Gyan, B.; Akanmori, B.D.; Ocran, J.; Adu-Amankwah, S.; Geneshan, H.; Abot, E.; Legano, J.; Banania, G.; Sayo, R.; Brambilla, D.; Kumar, S.; Doolan, D.L.; Rogers, W.O.; Epstein, J.; Richie, T.L.; Sedegah, M.Abstract Background To prepare field sites for malaria vaccine trials, it is important to determine baseline antibody and T cell responses to candidate malaria vaccine antigens. Assessing T cell responses is especially challenging, given genetic restriction, low responses observed in endemic areas, their variability over time, potential suppression by parasitaemia and the intrinsic variability of the assays. Methods In Part A of this study, antibody titres were measured in adults from urban and rural communities in Ghana to recombinant Plasmodium falciparum CSP, SSP2/TRAP, LSA1, EXP1, MSP1, MSP3 and EBA175 by ELISA, and to sporozoites and infected erythrocytes by IFA. Positive ELISA responses were determined using two methods. T cell responses to defined CD8 or CD4 T cell epitopes from CSP, SSP2/TRAP, LSA1 and EXP1 were measured by ex vivo IFN-γ ELISpot assays using HLA-matched Class I- and DR-restricted synthetic peptides. In Part B, the reproducibility of the ELISpot assay to CSP and AMA1 was measured by repeating assays of individual samples using peptide pools and low, medium or high stringency criteria for defining positive responses, and by comparing samples collected two weeks apart. Results In Part A, positive antibody responses varied widely from 17%-100%, according to the antigen and statistical method, with blood stage antigens showing more frequent and higher magnitude responses. ELISA titres were higher in rural subjects, while IFA titres and the frequencies and magnitudes of ex vivo ELISpot activities were similar in both communities. DR-restricted peptides showed stronger responses than Class I-restricted peptides. In Part B, the most stringent statistical criteria gave the fewest, and the least stringent the most positive responses, with reproducibility slightly higher using the least stringent method when assays were repeated. Results varied significantly between the two-week time-points for many participants. Conclusions All participants were positive for at least one malaria protein by ELISA, with results dependent on the criteria for positivity. Likewise, ELISpot responses varied among participants, but were relatively reproducible by the three methods tested, especially the least stringent, when assays were repeated. However, results often differed between samples taken two weeks apart, indicating significant biological variability over short intervals.Item Measuring naturally acquired immune responses to candidate malaria vaccine antigens in Ghanaian adults.(2011) Dodoo, D.; Hollingdale, M. R; Anum, D.; Koram, K. A; Gyan, B.; Akanmori, B. D.; Ocran, J.; Adu-Amankwah, S.; Geneshan, H.; Abot, E.; Legano, J.; Banania, G.; Sayo, R.; Brambilla, D.; Kumar, S.; Doolan, D.L.; Roger, W.O.; Epstein, J.; Richie, T.L.; Sedegah, M.Background: To prepare field sites for malaria vaccine trials, it is important to determine baseline antibody and T cell responses to candidate malaria vaccine antigens. Assessing T cell responses is especially challenging, given genetic restriction, low responses observed in endemic areas, their variability over time, potential suppression by parasitaemia and the intrinsic variability of the assays. Methods. In Part A of this study, antibody titres were measured in adults from urban and rural communities in Ghana to recombinant Plasmodium falciparum CSP, SSP2/TRAP, LSA1, EXP1, MSP1, MSP3 and EBA175 by ELISA, and to sporozoites and infected erythrocytes by IFA. Positive ELISA responses were determined using two methods. T cell responses to defined CD8 or CD4 T cell epitopes from CSP, SSP2/TRAP, LSA1 and EXP1 were measured by ex vivo IFN- ELISpot assays using HLA-matched Class I- and DR-restricted synthetic peptides. In Part B, the reproducibility of the ELISpot assay to CSP and AMA1 was measured by repeating assays of individual samples using peptide pools and low, medium or high stringency criteria for defining positive responses, and by comparing samples collected two weeks apart. Results: In Part A, positive antibody responses varied widely from 17%-100%, according to the antigen and statistical method, with blood stage antigens showing more frequent and higher magnitude responses. ELISA titres were higher in rural subjects, while IFA titres and the frequencies and magnitudes of ex vivo ELISpot activities were similar in both communities. DR-restricted peptides showed stronger responses than Class I-restricted peptides. In Part B, the most stringent statistical criteria gave the fewest, and the least stringent the most positive responses, with reproducibility slightly higher using the least stringent method when assays were repeated. Results varied significantly between the two-week time-points for many participants. Conclusions: All participants were positive for at least one malaria protein by ELISA, with results dependent on the criteria for positivity. Likewise, ELISpot responses varied among participants, but were relatively reproducible by the three methods tested, especially the least stringent, when assays were repeated. However, results often differed between samples taken two weeks apart, indicating significant biological variability over short intervals. © 2011 Dodoo et al; licensee BioMed Central Ltd.