title: A plant based protective antigen [PA(dIV)] vaccine expressed in chloroplasts demonstrates protective immunity in mice against anthrax.
abstract: The currently available anthrax vaccines are limited by being incompletely characterized, potentially reactogenic and have an expanded dosage schedule. Plant based vaccines offer safe alternative for vaccine production. In the present study, we expressed domain IV of Bacillus anthracis protective antigen gene [PA(dIV)] in planta (by nuclear agrobacterium and chloroplast transformation) and E. coli [rPA(dIV)]. The presence of transgene and the expression of PA(dIV) in planta was confirmed by molecular analysis. Expression levels up to 5.3% of total soluble protein (TSP) were obtained with AT rich (71.8% AT content) PA(dIV) gene in transplastomic plants while 0.8% of TSP was obtained in nuclear transformants. Further, we investigated the protective response of plant and E. coli derived PA(dIV) in mice by intraperitoneal (i.p.) and oral immunizations with or without adjuvant. Antibody titers of >10(4) were induced upon i.p. and oral immunizations with plant derived PA(dIV) and oral immunization with E. coli derived PA(dIV). Intraperitoneal injections with adjuvanted E. coli derived PA(dIV), generated highest antibody titers of >10(5). All the immunized groups demonstrated predominant IgG1 titers over IgG2a indicating a polarized Th2 type response. We also evaluated the mucosal antibody response in orally immunized groups. When fecal extracts were analyzed, low sIgA titer was demonstrated in adjuvanted plant and E. coli derived PA(dIV) groups. Further, PA(dIV) antisera enhanced B. anthracis spore uptake by macrophages in vitro and also demonstrated an anti-germinating effect suggesting a potent role at mucosal surfaces. The antibodies from various groups were efficient in neutralizing the lethal toxin in vitro. When mice were challenged with B. anthracis, mice immunized with adjuvanted plant PA(dIV) imparted 60% and 40% protection while E. coli derived PA(dIV) conferred 100% and 80% protection upon i.p. and oral immunizations. Thus, our study is the first attempt in highlighting the efficacy of plant expressed PA(dIV) by oral immunization in murine model.
authors.affiliation: Laboratory of Molecular Biology and Genetic Engineering, School of Biotechnology, Jawaharlal Nehru University, New Delhi, India.
authors.forename: Jyotsna
authors.initials: J
authors.lastname: Gorantala
authors.forename: Sonam
authors.initials: S
authors.lastname: Grover
authors.forename: Divya
authors.initials: D
authors.lastname: Goel
authors.forename: Amit
authors.initials: A
authors.lastname: Rahi
authors.forename: Sri Krishna
authors.initials: SK
authors.lastname: Jayadev Magani
authors.forename: Subhash
authors.initials: S
authors.lastname: Chandra
authors.forename: Rakesh
authors.initials: R
authors.lastname: Bhatnagar
mesh_terms: D000818:Animals; D000881:Anthrax / Q000276:immunology / Q000517:prevention & control*; D022122:Anthrax Vaccines / Q000008:administration & dosage / Q000235:genetics / Q000276:immunology* / Q000378:metabolism; D000907:Antibodies, Bacterial / Q000097:blood / Q000276:immunology; D057134:Antibodies, Neutralizing / Q000097:blood / Q000276:immunology; D000942:Antigens, Bacterial / Q000235:genetics / Q000276:immunology* / Q000378:metabolism; D001408:Bacillus anthracis / Q000276:immunology; D001427:Bacterial Toxins / Q000235:genetics / Q000276:immunology* / Q000378:metabolism; D002736:Chloroplasts / Q000235:genetics / Q000378:metabolism*; D018928:Immunity, Mucosal; D007070:Immunoglobulin A / Q000097:blood / Q000276:immunology; D007071:Immunoglobulin A, Secretory / Q000276:immunology; D007074:Immunoglobulin G / Q000097:blood / Q000276:immunology; D051379:Mice; D008807:Mice, Inbred BALB C; D008969:Molecular Sequence Data; D030821:Plants, Genetically Modified; D012231:Rhizobium / Q000235:genetics / Q000378:metabolism; D014026:Nicotiana / Q000235:genetics / Q000378:metabolism / Q000382:microbiology; D014170:Transformation, Genetic; D014611:Vaccination; D019444:Vaccines, DNA / Q000008:administration & dosage / Q000235:genetics / Q000276:immunology* / Q000378:metabolism
chemical_list: D022122:Anthrax Vaccines; D000907:Antibodies, Bacterial; D057134:Antibodies, Neutralizing; D000942:Antigens, Bacterial; D001427:Bacterial Toxins; D007070:Immunoglobulin A; D007071:Immunoglobulin A, Secretory; D007074:Immunoglobulin G; D019444:Vaccines, DNA; C030325:anthrax toxin