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edible vaccine is based on oral vaccine in which certain protein is inserted which act as a antigen and body start to produce antibody against foreign particle .

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Edible vaccine

BY :-Rewat pd kafle SANN int l college Kathmandu

IntroductionTypes of Vaccines Preparing of Edible Vaccines Advantages of Edible Vaccines Limitations of Edible Vaccines Conclusion

Immunization HistoryEdward Jenner, noticed that milkmaids who developed cowpox, did not develop the deadly smallpox. In 1796, Jenner took the fluid from a cowpox pustule on a dairymaid's hand and inoculated an 8-year-old boy. Six weeks later, he exposed the boy to smallpox, and the boy did not develop any symptoms. Jenner coined the term "vaccine" from the word "vaca" which means "cow" in Latin.

yBackgroundy 80% of worlds infants immunized y 20% remaining account for 2,000,00 death annually y Mostly remote, impoverished nations y Foods could produce vaccines

y Vaccine y A vaccine is a biological preparation that

improves immunity to a particular disease y typically contains an agent that resembles a disease-causing microorganism, and is often made from weakened or killed forms of the microbe or its toxins y agent stimulates the body's immune system to recognize the agent as foreign, destroy it, and recognize it.

y Vaccinationy Vaccination is the administration of antigenic material to

produce immunity to a disease. material administered can either be live but weakened forms of pathogen (Bacteria or virus ) killed or inactivated forms of these pathogens, or purified material such as proteins.

(Child receiving an oral polio vaccine).

y Types of vaccinesy Killed(virulent, micro-organisms that have been destroyed with chemicals or heat) y Attenuated(live viruses that have been cultivated under conditions that disabletheir virulent properties)

y Toxoid (toxoid vaccines are made from inactivated toxic compounds ) y Subunit(introducing an inactivated or attenuated micro-organism to an immunesystem)

y Conjugate, recombinant Vector vaccine etc.

Do we think this boy is happy?

y Introduction of edible vacciney Edible vaccines are sub-unit vaccines where the selected

genes are introduced into the plants and the transgenic plant is then induced to manufacture the encoded protein. y Edible vaccines are mucosal targeted vaccines where stimulation of both systematic and mucosal immune network takes place.y Edible vaccines against cholera toxin B subunit, hepatitis B

surface antigen, E. coli heat labile enterotoxin protein have been developed and tested for efficacy in animal and human trials.

y oral subunit vaccine for the respiratory syncytial virus a

y y y y

serious pathogen that causes bronchiolitis and pneumoniatype diseases in human RSV is a leading cause of viral lower respiratory tract illness in infants and children. The virus enters the human body mainly through the nose and eyes, but also through the mouth. Tomato was selected for the production and delivery of vaccine . The tomato is one of the most important vegetable crops of commercial importance.

y constructs carrying the RSV-F

antigenic protein gene. y constructs consisted of the universal constitutive cauliflower mosaic virus (CaMV) 35S promoter along with enhancer elements or leader sequences added to the RSVF protein gene y CaMV 35S-driven gene constructs carrying the antigenic F-protein were transformed into tomato leaf protoplasts to determine whether the F-protein could be correctly processed in a plant cell. Using SDS-PAGE and Western analyses, the correct size protein (68 kd) was found in transformed in tomato protoplasts .

Fertile transgenic tomato plants carrying the RSV-F antigen growing in the greenhouse

y immunoblotting with a monoclonal antibody against the F-

protein, it was confirmed that the F protein was expressedy the level of expression of the F-protein increased in

constructs carrying the leader sequence AMV RNA4 and the P268 enhancey the antigenic F protein transferred into tomato using Agro

bacterium-mediated transformation, and transgenic tomato plant.

y The presence of the transgene was confirmed in 71 out of 74

tomato plants growing in the greenhouse using Southern blotting. y harvested fruit was analyzed using ELISA With the Epromoter. y the F-protein was expressed only in the fruit of all transgenic plants carrying the F-antigenic protein gene expression level of the F-protein among plants was ranging from 9.0 to 32.5 g/g of fruit fresh weight.y Fruits from the highest F-protein expressing plants were fed

to mice, and among 25 orally immunized mice, 22 showed significant immune responses and produced anti-RSV-F antibodies.

y Upon analysis of the tomato fruit for the localization of the

antigenic RSV-F protein, it was found that the majority of the antigenic protein is localized in the seed, while the pulp contained only marginal levels of the antigen . This suggested that in order to deliver a high amount level of the RSV-F antigen vaccine, the whole tomato fruit (seed and pulp) must be homogenized and used for the delivery of the vaccine to insure presence of high enough levels of the antigen.

Southern blot analysis of tomato lines. Lanes 1 5 correspond to transformed tomato lines carrying the RSV-F transgene and showing a single copy of the transgene; lane 6 corresponds to an untransformed tomato plant (control), and lane 7 corresponds to the plasmid DNA showing the RSV-F antigen

Mechanism of Action

y The goal of oral vaccine is to stimulate both mucosal and

humoral immunity against pathogen . y Edible vaccine when taken orally undergoes mastication process and majority of plant cell degradation occurs in intestine as a result of action on digestive or bacterial enzyme on edible vaccine . y Peyers patches are rich sources of IgA producing plasma cells y The break down of edible vaccine occurs near pp.y IgA antibodies are transported across epithelial cell into

secretion of lumen where they intact with antigen present in lumen .

Engineering Edible Vaccines

y Introduction of foreign DNA y plasmid needs to be disarmed by deleting the genes for auxin y and cytokinin synthesis y Genes for antibiotic resistance are used to select out the

transformed cells and whole plants, which contain the foreign gene; and for expressing the desired product, which can then be regenerated from them so that it does not produce tumor. y Each single antigen expressed in plants must be tested for its proper assembly and can be verified by animal studies, Western blot; and quantified by enzyme-linked immunosorbent assay (ELISA)

y Choice of plant species for vaccine deliveryBest food plant Ability to be eaten raw and unprocessed Suitable for infants Widely and easily grown Easily stored Resistant to spoiling

yAdvantages of edible vaccineLow-cost preserving system (numerous seeds, stored for long time) Easy purification of antibodies stored in the seeds, at any location Large harvests, number of times/year Commonly used in baby food because of low allergenic potential High expression of proteins Easy storage/transportation Expressed protein is heat-stable Direct consumption Stored for long periods without refrigeration Inexpensive Grown widely in developing countries produce large amounts of recombinant proteins High content of vitamin A may boost immune response

y Limitation of edible vacciney though edible vaccines are stable and easily accessible there are some limitations which restricts its development

Eg. Develop immunotolerance to the vaccine peptide or proteiny whole fruit or vegetable vaccines is the consistency of dosage from fruit to fruit, plant to plant y Fruit vaccines should be easily identifiable to avoid the misadministration of the vaccine, which may lead to complications such as immunotolerance

Conclusionsafer and more effective immunization. overcome the difficulties associated with traditional vaccines, like production, distribution and delivery . cost-effective, safe and efficacious delivery system Essential component in our disease-prevention.

ySafety issuey Allergic and toxic (e.g., glycans, nicotine, etc.) y Risk of a typical measles

(in plants with cloned measles virus genes)y Health and environmental risks of genetically modified organisms y Prevention of misuse/overuse

yEthical issuey Dangerous/harmful gene products or potent immunogens allergens may also be

incorporatedy Terminator crops with suicide genes responsible for male sterility. y GM food CaMV- 35 S promoter may responsible in the stomach and small intestine

effects y Antibiotic resistance marker genes can spread from transgenic food to pathogenic bacteria, making infections very difficult to treat.y risk of creating altogether new strains of infectious agents, like super viruses.