-Yanshi
-B.V.Sc.&A.H
Introduction:
From the past century, vaccination has widely been accepted as a tool to combat diseases both in case of human and animals. Livestock vaccination has gained socio-economic importance in the recent years, but the rate of adoption and diffusion of vaccination technology has been very low at field level. The use of edible vaccines has come to the rescue in recent times which offers a new strategy for the development of safe, inexpensive vaccines against diseases. Edible vaccines are the transgenic plant and animal – based production of or those agents produced in an edible format (i.e. a plant part, its fruit, or sub products derived from the plant) that upon oral ingestion will stimulate the immune system. In simple words, it can be said that plant or animal-made pharmaceuticals are edible vaccines. It does not necessarily mean nutritious, tasty, or organoleptically pleasing.
Concept of Edible vaccines
The concept of edible vaccines was developed by Arntzen in 1990s which proved that such vaccines can annihilate the restrictions in production of traditional vaccines. The earliest demonstration of an edible vaccine was in tobacco plant in which a surface antigen from the bacterium Streptococcus mutans was expressed. The bacterium was responsible for causing dental carries. It was assumed that mucosal immune response stimulation would prevent the bacteria from colonizing the teeth thereby preventing tooth decay. This demonstration was the milestone in edible vaccine production.
Production of Edible vaccines:
The edible vaccines are subunit vaccines which consist of antigenic proteins and are devoid of pathogenic genes. The gene encoding antigens from pathogenic organisms are produced in the edible parts of the plants in two ways:
• Epitope within the antigen are identified and the DNA fragments encoding these are used to construct genes by fusion with coat protein gene of plant virus. The recombinant virus is then used to infect stabilized plants to produce a huge number of new plants.
• In another method, the entire structural gene is incorporated with plant vector by transformation which allows transcription and accumulation of coding sequence in plants.
The other approaches that can be used are:
Agrobacterium mediated gene transfer
The most common transfer vector for DNA is Agrobacterium tumefaciens which is co-cultured with plant cells or tissues which needs to be transformed. The first step includes identification, isolation and characterization of a pathogenic antigen. The antigen needs to elicit a strong and specific immune response in order to be effective. The gene is cloned in the transfer vector into the transfer DNA (T-DNA) to produce antigenic protein. It is then inserted into genome, expressed and inherited into mendalian fashion which results in the antigen being expressed in the fruit or plant. This approach is slow with lower yield, however shows satisfactory results in dicotyledonous plants.
Biolistic method:
It is a sophisticated method that involves the use of gene gun that fires the gene containing DNA coated metal such as gold, tungsten particles at the plant cells. The plants take up the DNA, grow into new plants and then are clone to produce large numbers of genetically identical crops. This approach is highly attractive as the gene transfer is independent of the regeneration ability of the species, but the requirement of expensive device particle gun is a major drawback to this method. It can express antigens through nuclear and chloroplast transformation.
Electroporation:
The DNA is inserted into the cells and is exposed to high voltage electrical pulse which produces transient pores within the plasma lemma. It requires mild enzymatic treatment to weaken the cell wall as it acts as an effective barrier against entry of DNA into cell cytoplasm.
Mechanism of Action:
When an edible vaccine is consumed by an individual, the outer plant’s cell wall protects the antigens from degradation by gastric secretion, which enables the antigens to be delivered to the intestinal mucosal surfaces, where they are absorbed to stimulate a strong and specific immune response.
The main route of antigen capture at the intestinal level is through Microfold (M) cells which represent a small number of specialized follicular-associated epithelium (FAE) enterocytes found primarily in the gastrointestinal tract. The M cells capture a wide variety of macromolecules and microorganisms from the lumen of the small intestine to submucosal antigen-presenting cells (APCs) on Peyer’s patches. Dendritic cells (DCs) are the most potent antigen-presenting cells in priming naive T cells to initiate an adaptive immune response. Dendritic cells are found in an immature stage in a steady state and are characterized by high endocytic activity and a low capability to prime naive T cells.
But in inflammatory conditions, these dendritic cells mature resulting in an increase in the expression of co-stimulatory molecules and migration to T-cell-rich zones in lymph nodes, where they present antigens along with the release of cytokines which facilitates the differentiation of naive antigen-specific T cells into effector cells and their migration to the specific inflammation site.
Intestinal DCs can promote the activation of naive T cells and the differentiation to follicular T helper cells (Tfh) either by directly promoting Tfh differentiation or indirectly by promoting Th17 cells that will later become Tfh. These T helper cells specifically promote the activation of follicular B cells and the generation of IgG and IgA-secreting plasma cells. These activated B cells then leave the lymphoid follicles and migrate to the mucosa associated lymphoid tissue (MALT), where plasma cells secreting immunoglobulin A (IgA) antibodies can be found. These IgA antibodies are transported across epithelial cells in secretions to the lumen, where they interact with antigens. DCs are critically important in IgA class switching and secretion in B cells. Also, DCs can directly capture luminal antigens by projecting dendrites through the epithelial cell layer and into the lumen.
The other mechanism of antigen capture in the small intestine involves goblet cells which are involved in the production of mucins and can directly capture and deliver antigens to intestinal DCs. An efficient, edible vaccine stimulates specific T and B cell responses, which promote long-lasting memory cells for subsequent encounters wherein the antigen is presented in the course of an actual infection.
Advantages of Edible vaccines:
Edible vaccines possess a lot of advantages over traditional vaccines which includes:
• Edible vaccines are comparatively cost effective as they do not require expensive manufacturing equipments but rich soil.
• These do not require sterilized production facility or biosafety standards to cultivate certain pathogenic agents.
• Edible vaccines do not require strict refrigeration storage and have good genetic and heat stability.
• The seeds from an edible vaccine plant can be easily dehydrated and preserved for cheap and quick distribution, making them easily accessible in times of need. Also, plants with oil or their aqueous extracts possess more storage opportunities.
• It requires simpler means of administration, making it economical and reducing the need for medical personnel and sterile injection conditions, which are not always achievable.
• Traditional vaccines developed from cultured mammalian cells can lead to contamination with other animal viruses, but it is not the case with edible vaccines as plant viruses can’t impact animals.
• Moreover, due to numerous antigens being integrated, M cells are randomly stimulated leading to possibility of second generation vaccines.
• Edible vaccines do not require subsidiary elements to stimulate immune response like traditional vaccines as edible vaccines have efficient mode of action for immunization. They specifically stimulate both mucosal and humoral immune system.
• Edible vaccines lack certain toxic compounds and only contain therapeutic proteins which are free of pathogens and toxins due to which risk of potential side effects and allergic reactions are greatly reduced. Moreover, the antigen remains protected through bio encapsulation.
• The production process can be scaled up rapidly by breeding.
Disadvantage of Edible vaccines
Some limitations of edible vaccines are:
• The animal may develop immune tolerance to a particular vaccine protein or peptide if the dose is not given properly.
• The dosage requirement varies from generation to generation, plant to plant, protein content, quantity of food eaten and patient’s age and weight.
• The administration requires methods for standardization of plant material and its products as low doses may result in lesser number of antibodies production and high doses may cause immune tolerance.
• Plant stability is one of the major limitations in case of humans as certain foods can’t be eaten raw and needs cooking that can cause denaturation or weaken the protein present in it. This may not be a problem in animals as animals are mostly fed raw feed.
• The vaccines are prone to get microbial infestation if not properly stored.
• A proper demarcation line may not be there to differentiate between ‘vaccine feed’ and ‘normal feed’ to avoid misadministration of the vaccine.
• The function of the edible vaccine can be hampered due to vast differences in the glycosylation pattern of plants and animals.
• The continuity of the vaccine production cannot be guaranteed in plants as they are living and can change themselves.
• The gastric enzymes and the acidic environment of the stomach can cause the breakdown of the vaccine before it can activate an immune response.
• The animal may develop an allergy to the feed expressing the foreign antigen.
• The effects and risk of using pesticides on the plants can be negative towards both the plant vaccine and the animal.
• There is also a possibility of transgenic escape into the surrounding environment, but this can be prevented or reduced by regulating growing practices and locations.
Examples of Edible vaccines:
Lettuce
The plant is an effective model system against enteric diseases caused by E.coli in both animals and humans. The glycoprotein E2 expressed lettuce for classical swine fear hog pest virus has been developed. The plant is used in raw form and has utmost efficacy to be used as edible vaccine.
Alfalfa:
This is the plant used to develop edible vaccines mainly for veterinary use. Transgenic alfalfa containing hog pest virus glycoprotein E2 has also been developed.
Corn:
Edible vaccines expressing antigenic glycoproteins against rabies virus in humans and animals and transmissible gastroenteritis coronavirus (TGEV) in pigs have been prepared and tested. Transgenic corn expressing the fusion protein of the Newcastle disease virus (NDV) has shown to produce immunogenic effects and confer protective immunity in poultry.
Quinoa:
An edible vaccine based on quinoa has been developed by expressing the VP2 antigen from infectious bursitis virus which mainly affects poultry.
Tobacco:
Transgenic tobacco expressing VP1 protein against chicken infectious anemia has been prepared.
Papaya:
Transgenic papaya expressing antigenic glycoproteins for Taenia solium has been made to combat cysticercosis both in humans and pigs.
Algae:
Chlamydomonas reinhardtii as host algae has been used to prepare edible vaccines for foot and mouth disease and classical swine flu virus.
Applications of Edible vaccines:
Edible vaccines have immense applications in diverse fields such as:
Cancer therapy:
Several plants have been engineered to generate monoclonal antibodies which have been verified as effective cancer therapy agents. An example is of monoclonal antibody in soyabean for BR-96 which attacks doxorubicin responsible for causing breast cancer, ovarian cancer, colon cancer and lung tumors.
Birth control:
Antibodies formed due to administration of TMV produced protein found on zona pellucida (ZB3 protein) are capable of preventing fertilization of eggs.
Autoimmune diseases:
It has been found that the proteins expressed in genetically modified potatoes are successful in suppressing immune attack and delaying the onset of high sugar level.
Recombinant drugs/proteins:
The plants can be engineered to produce enzymes and drugs for treating various diseases in animals.
Chloroplast transformation:
Transgenic protein allow transmission and accumulation of chloroplast genome in ample quantities which otherwise is not possible via usual cross pollination.
Current status and future aspect of Edible vaccines
Many recombinant proteins as vaccine and other pharmaceutical compounds have been developed to combat human and animals diseases and have reached various phases of clinical trials. There has been growth in the research areas of edible vaccines in the past few years, but still many aspects have to unfurl. But, the resistance towards genetically modified crops and transgenic contamination may present a threat to the rising of edible vaccines.
Conclusion:
Edible vaccines can represent a valuable solution for treating various diseases whose control and prevention is restricted due to limitations of traditional vaccines such as production cost, storage requirements, their harmful side effects, etc. However there are some challenges which need to be overcome to make the saying ‘Let thy food be thy medicine’ turned into a reality.
******
