title: Stable expression of Gal/GalNAc lectin of Entamoeba histolytica in transgenic chloroplasts and immunogenicity in mice towards vaccine development for amoebiasis.
abstract: Chloroplast genetic engineering offers several advantages, including high levels of transgene expression, transgene containment via maternal inheritance and multigene engineering in a single transformation event. Entamoeba histolytica infects 50 million people, causing about 100,000 deaths annually, but there is no approved vaccine against this pathogen. LecA, a potential target for blocking amoebiasis, was expressed for the first time in transgenic plants. Stable transgene integration into chloroplast genomes and homoplasmy were confirmed by polymerase chain reaction and Southern blot analyses. LecA expression was evaluated by Western blots and quantified by enzyme-linked immunosorbent assay (up to 6.3% of total soluble protein or 2.3 mg LecA/g leaf tissue). Subcutaneous immunization of mice with crude extract of transgenic leaves resulted in higher immunoglobulin G titres (up to 1:10,000) than in previous reports. An average yield of 24 mg of LecA per plant should produce 29 million doses of vaccine antigen per acre of transgenic plants. Such high levels of expression and immunogenicity should facilitate the development of a less expensive amoebiasis vaccine.
authors.affiliation: Department of Molecular Biology and Microbiology, University of Central Florida, Biomolecular Science, Bldg. #20, Room 336, Orlando, FL 32816-2364, USA.
authors.forename: Seethamahalakshmi
authors.initials: S
authors.lastname: Chebolu
authors.forename: Henry
authors.initials: H
authors.lastname: Daniell
mesh_terms: D000562:Amebiasis / Q000276:immunology*; D000818:Animals; D000953:Antigens, Protozoan / Q000235:genetics* / Q000276:immunology; D015139:Blotting, Southern; D015153:Blotting, Western; D002736:Chloroplasts / Q000235:genetics* / Q000378:metabolism; D004748:Entamoeba histolytica / Q000276:immunology*; D004797:Enzyme-Linked Immunosorbent Assay; D005260:Female; D005818:Genetic Engineering; D037102:Lectins / Q000235:genetics / Q000276:immunology; D051379:Mice; D008807:Mice, Inbred BALB C; D030821:Plants, Genetically Modified; D016133:Polymerase Chain Reaction; D017434:Protein Structure, Tertiary; D015800:Protozoan Proteins / Q000235:genetics* / Q000276:immunology; D016052:Protozoan Vaccines / Q000235:genetics* / Q000276:immunology; D014026:Nicotiana / Q000166:cytology / Q000235:genetics; D014614:Vaccines, Synthetic / Q000276:immunology
chemical_list: D000953:Antigens, Protozoan; D037102:Lectins; D015800:Protozoan Proteins; D016052:Protozoan Vaccines; D014614:Vaccines, Synthetic