Constructing and transient expression of a gene cassette containing edible vaccine elements and shigellosis, anthrax and cholera recombinant antigens in tomato

Abstract

Shigella dysenteriae causing shigellosis is one of the diseases that threaten the health of human society in the developing countries. In Shigella, IpaD gene is one of the key pathogenic genes causing strong mucosal immune system reactions. Anthrax disease is caused by Bacillus anthracis. PA protective antigen is one of the subunits in anthrax toxin complex responsible for the transfer of other subunits into the cytosol of host cells. The 20 kDa subunit of PA (PA20) has the property of immunogenicity. CTxB or B subunit of Vibrio cholerae toxin (CT) is a non-toxic protein and has the function to transfer toxic subunit into cytosol of the host cells by binding to GM1 receptor. The aim of this study was to fuse PA20, ipaD and CTxB and transform tomato plants by this cassette in order to produce an oral vaccine against shigellosis, anthrax and cholera. CTxB was used for these two antigens as an immune adjuvant. IpaD and PA20 genes were cloned in pBI121 containing the CTxB gene and Extensin signal peptide. In order to evaluate the transient expression of Shigellosis, Anthrax and Cholera antigens, agro-infiltrated tomato tissues were inoculated with Agrobacterium tumefaciens containing the gene cassette. Cloning was confirmed by PCR, enzymatic digestion and sequencing techniques. Expression of the antigens was examined by SDS-PAGE, dot blot and ELISA. Maturate green fruits demonstrated the highest expression of the recombinant proteins. The first phase of this study was carried out for cloning and expressing of CtxB, ipaD and PA20 antigens in tomato. In the next phase, we aim to analyze the immunogenicity of this vaccine candidate in laboratory animals.

Designer Exosomes: A New Platform for Biotechnology Therapeutics

“…Studies on vaccine development showed that host cells infected with parasites, viruses, or bacterial pathogens secrete exosomes filled with several antigens of those pathogens [ 128 ]. Experiments in vaccine development showed that cocktail and multi-epitopic vaccines (MEVs) could overcome antigenic escape of pathogens from the immune system [ 129 , 130 ]. Hopefully, using exosome engineering methods, exosomes could be recruited to present multiple antigens as designer vaccine exosomes.…”

A cross talk between the immunization and edible vaccine: Current challenges and future prospects

“…Cholera is a diarrhoeal disease caused by bacteria Vibrio cholerae , a bacterium of comma shape. It colonizes the intestine and produces enterotoxin cholera toxin B (CTB) and causes acute watery diarrhoea [ 78 , 79 ]. When taken orally CTB acts as a mucosal immunogen.…”

Designing a chimeric subunit vaccine for influenza virus, based on HA2, M2e and CTxB: a bioinformatics study

“…Another type of vaccine that is receiving a great deal of attention is subunit vaccines. Today, these vaccines are produced by biotechnology methods, including recombinant DNA technology [12]. These vaccines are produced in various expression systems, such as the E. coli (prokaryotic expression system) or eukaryotic expression systems such as yeast [13], HEK-293 and CHO cell lines [14] as well as plant cells [12].…”

“…Today, these vaccines are produced by biotechnology methods, including recombinant DNA technology [12]. These vaccines are produced in various expression systems, such as the E. coli (prokaryotic expression system) or eukaryotic expression systems such as yeast [13], HEK-293 and CHO cell lines [14] as well as plant cells [12]. Due to the antigenic property of HA, NA, and M2e proteins, various studies have been done on the development of a subunit vaccine against influenza virus based on these proteins [15].…”

“…Various bioinformatics tools can be used in the development of a vaccine [18,19]. In our previous studies, bioinformatics analyses were performed before in vitro development of the chimeric vaccine candidate for the three anthrax, shigellosis, and cholera diseases [12]. Before the expression of the vaccine in a host by biotechnology techniques, bioinformatics analyses were conducted to optimize the gene construct in terms of its stability and half-life as well as to retain the third structure and epitopes intact [20].…”