Chloroplast-derived vaccine antigens confer dual immunity against cholera and malaria by oral or injectable delivery

Plant Biotechnol J. 2010 Feb;8(2):223-42. doi: 10.1111/j.1467-7652.2009.00479.x. Epub 2009 Dec 28.

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.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, U.S. Gov't, Non-P.H.S.

MeSH terms

  • Administration, Oral
  • Animals
  • Antibodies, Bacterial / blood
  • Antibodies, Protozoan / blood
  • Antigens, Protozoan / genetics
  • Antigens, Protozoan / immunology
  • CD4-Positive T-Lymphocytes / immunology
  • Chloroplasts / immunology*
  • Chloroplasts / metabolism
  • Cholera / immunology
  • Cholera / prevention & control*
  • Cholera Toxin / genetics
  • Cholera Toxin / immunology
  • Cholera Vaccines / biosynthesis*
  • Cholera Vaccines / genetics
  • Cholera Vaccines / immunology
  • Cross Reactions
  • Female
  • Immunity, Humoral
  • Immunoglobulin A / blood
  • Immunoglobulin G / blood
  • Injections, Subcutaneous
  • Lettuce / genetics
  • Lettuce / immunology
  • Malaria / immunology
  • Malaria / prevention & control*
  • Malaria Vaccines / biosynthesis*
  • Malaria Vaccines / genetics
  • Malaria Vaccines / immunology
  • Merozoite Surface Protein 1 / genetics
  • Merozoite Surface Protein 1 / immunology
  • Mice
  • Mice, Inbred BALB C
  • Plants, Genetically Modified / genetics
  • Plants, Genetically Modified / immunology
  • Recombinant Fusion Proteins / immunology
  • Tobacco / genetics
  • Tobacco / immunology

Substances

  • Antibodies, Bacterial
  • Antibodies, Protozoan
  • Antigens, Protozoan
  • Cholera Vaccines
  • Immunoglobulin A
  • Immunoglobulin G
  • Malaria Vaccines
  • Merozoite Surface Protein 1
  • Recombinant Fusion Proteins
  • Cholera Toxin