title: Chloroplast-derived vaccine antigens confer dual immunity against cholera and malaria by oral or injectable delivery.
abstract: Cholera and malaria are major diseases causing high mortality. The only licensed cholera vaccine is expensive; immunity is lost in children within 3 years and adults are not fully protected. No vaccine is yet available for malaria. Therefore, in this study, the cholera toxin-B subunit (CTB) of Vibrio cholerae fused to malarial vaccine antigens apical membrane antigen-1 (AMA1) and merozoite surface protein-1 (MSP1) was expressed in lettuce and tobacco chloroplasts. Southern blot analysis confirmed homoplasmy and stable integration of transgenes. CTB-AMA1 and CTB-MSP1 fusion proteins accumulated up to 13.17% and 10.11% (total soluble protein, TSP) in tobacco and up to 7.3% and 6.1% (TSP) in lettuce, respectively. Nine groups of mice (n = 10/group) were immunized subcutaneously (SQV) or orally (ORV) with purified antigens or transplastomic tobacco leaves. Significant levels of antigen-specific antibody titres of immunized mice completely inhibited proliferation of the malarial parasite and cross-reacted with the native parasite proteins in immunoblots and immunofluorescence studies. Protection against cholera toxin challenge in both ORV (100%) and SQV (89%) mice correlated with CTB-specific titres of intestinal, serum IgA and IgG1 in ORV and only IgG1 in SQV mice, but no other immunoglobulin. Increasing numbers of interleukin-10(+) T cell but not Foxp3(+) regulatory T cells, suppression of interferon-gamma and absence of interleukin-17 were observed in protected mice, suggesting that immunity is conferred via the Tr1/Th2 immune response. Dual immunity against two major infectious diseases provided by chloroplast-derived vaccine antigens for long-term (>300 days, 50% of mouse life span) offers a realistic platform for low cost vaccines and insight into mucosal and systemic immunity.
authors.affiliation: Department of Molecular Biology and Microbiology, College of Medicine, University of Central Florida, Orlando, FL 32816-2364, USA.
authors.forename: Abdoreza
authors.initials: A
authors.lastname: Davoodi-Semiromi
authors.forename: Melissa
authors.initials: M
authors.lastname: Schreiber
authors.forename: Samson
authors.initials: S
authors.lastname: Nalapalli
authors.forename: Dheeraj
authors.initials: D
authors.lastname: Verma
authors.forename: Nameirakpam D
authors.initials: ND
authors.lastname: Singh
authors.forename: Robert K
authors.initials: RK
authors.lastname: Banks
authors.forename: Debopam
authors.initials: D
authors.lastname: Chakrabarti
authors.forename: Henry
authors.initials: H
authors.lastname: Daniell
mesh_terms: D000284:Administration, Oral; D000818:Animals; D000907:Antibodies, Bacterial / Q000097:blood; D000913:Antibodies, Protozoan / Q000097:blood; D000953:Antigens, Protozoan / Q000235:genetics / Q000276:immunology; D015496:CD4-Positive T-Lymphocytes / Q000276:immunology; D002736:Chloroplasts / Q000276:immunology* / Q000378:metabolism; D002771:Cholera / Q000276:immunology / Q000517:prevention & control*; D002772:Cholera Toxin / Q000235:genetics / Q000276:immunology; D022121:Cholera Vaccines / Q000096:biosynthesis* / Q000235:genetics / Q000276:immunology; D003429:Cross Reactions; D005260:Female; D056724:Immunity, Humoral; D007070:Immunoglobulin A / Q000097:blood; D007074:Immunoglobulin G / Q000097:blood; D007279:Injections, Subcutaneous; D018545:Lactuca / Q000235:genetics / Q000276:immunology; D008288:Malaria / Q000276:immunology / Q000517:prevention & control*; D017780:Malaria Vaccines / Q000096:biosynthesis* / Q000235:genetics / Q000276:immunology; D020066:Merozoite Surface Protein 1 / Q000235:genetics / Q000276:immunology; D051379:Mice; D008807:Mice, Inbred BALB C; D030821:Plants, Genetically Modified / Q000235:genetics / Q000276:immunology; D011993:Recombinant Fusion Proteins / Q000276:immunology; D014026:Nicotiana / Q000235:genetics / Q000276:immunology
chemical_list: D000907:Antibodies, Bacterial; D000913:Antibodies, Protozoan; D000953:Antigens, Protozoan; D022121:Cholera Vaccines; D007070:Immunoglobulin A; D007074:Immunoglobulin G; D017780:Malaria Vaccines; D020066:Merozoite Surface Protein 1; D011993:Recombinant Fusion Proteins; D002772:Cholera Toxin