The document discusses different types of vaccines and how plant-based vaccines are produced. It describes the process of producing plant vaccines which involves selecting an antigen-encoding gene and plant host, inserting the gene into the plant, and harvesting and processing the plant tissue to purify or consume the vaccine. Plant vaccines stimulate both mucosal and systemic immunity. Examples of plant vaccines discussed include those against bacteria, viruses, parasites, and SARS-CoV-2.

1. MADRAS CHRISTIAN COLLEGE ( AUTONOMOUS) , CHENNAI
3rd ICA Component Presentation On
Presented By:
Alok Kumar
3rd BSc Botany

2. What is a vaccine ?
According to World Health Organization (WHO), a vaccine is
any preparation intended to produce immunity to a disease by
stimulating the production of antibodies (WHO 2018).
A vaccine could possess purified microbial toxins, biomolecules, killed
or attenuated microorganisms, and recombinant vectors. Any vaccine
possesses two basic immunological characteristics: specificity & memory.
First vaccine is reported in 1796, when Edward Jenner attempted
vaccination for smallpox on an 8-year-old boy, James Phipps.
Louis Pasteur named these attenuated strains as vaccine (Latin vacca, meaning cow).

3. Types Of Vaccines:-
Live Attenuated Vaccines
A live attenuated vaccine is a live microbial preparation in which microbes have been weakened by growing
them on nonspecific host. Attenuated vaccine produces similar immune response as of wild strain, however
doesn’t cause disease. It produces a strong immune response that often lasts lifelong.
Ex- Influenza, MMR.
Inactivated Vaccine
Inactivated vaccine is a killed microbial preparation in which microbes have been killed by chemical or physical
treatment. These vaccines are safer and more stable than attenuated vaccine. An inactivated vaccine provides
low immunity therefore is needed to be administered in multiple doses.
Ex- Polio vaccine, Rabies virus.
Subunit Vaccines
Subunit vaccines do not use the entire microorganism; in its place thbese vaccines use only the antigens that
can induce the immune system effectively. These vaccines can be prepared by purifying the desired antigen
from the pathogenic organism or by genetic engineering. A gene coding the vaccine protein of interest is
inserted into a vector and subsequently expressed in a host system. These vaccines are safe and possess
minimum side effects due to the use of a specific gene.
Ex- Hepatitis B, Human Papilloma Virus.

4. Toxoid
A disease caused by microbial toxins instead of microorganism itself can be cured by toxoid. A toxoid vaccine is
an inactivated form of microbial toxins; the inactivation can be achieved by the treatment of toxin by chemical
such as formaldehyde.
Ex- Tetanus , Diphtheria
Conjugate Vaccines
Conjugate vaccines are a variation of subunit vaccines. This vaccine was developed to provide immunization
against certain microorganisms that possess a polysaccharide layer around them. This layer protects the
microorganism from the host’s immune system. In conjugate vaccine, antigenic proteins are linked with
microbial polysaccharide so that immune system can produce immunity against it. This polysaccharide
attachment helps the immune system to defend the body by reacting with the polysaccharide coating of the
pathogen.
Ex- Hib vaccine, Meningococcal vaccine
DNA Vaccines
This is a very recent technology. In this technique, gene encoding for microbial antigens is directly introduced
into host body. Then the microbial gene controls the host cell to produce the antigen protein of interest.
Expression of microbial protein in host cells result in the specific immune response in host cell.
Ex - Zydus Cadila ( SARS Cov-2)

5. Recombinant Vector Vaccines
Recombinant vector vaccines are functionally similar to DNA vaccines, but attenuated virus or bacteria are used
to introduce the microbial DNA into the host cells. Vector term is used for the attenuated virus or bacteria that
used to carry the microbial DNA.
Ex- VSV (Vesicular Stomatitis Virus), CMV( Cytomegalovirus)
Plant Vaccines
In the early 1990s, a novel approach to produce edible vaccine in plants emerged. Plant vaccines are a type of
subunit vaccines and commonly called edible vaccines. Plant-derived vaccines are less toxic and cheaper than
the conventional vaccines. The oral activity and heat stability of these vaccines make them an obvious choice for
poor countries.
Currently many plant-derived vaccines are available, and few of them are in clinical trials.

6. Production of Plant Vaccines :
Mason and Arntzen described detailed procedure for
the production of plant vaccines. Plant vaccines are
produced by molecular farming (Pharming).
Plant vaccine is produced by inserting a microbial gene
(encodes the microbial protein) into a plant.
Expression of this microbial gene in plant cells will
produce the protein that can be purified or consumed
along with the plant tissue.
Hence, It is an advanced biological technique used for
the production of pharmaceutically beneficial proteins
in plants.

7. Production of Plant Based Vaccines:
This process involves following crucial steps-
 Selection of the Gene
The gene encoding the antigenic proteins that stimulate the immune system should be selected for the process.
The requisitions of the gene include its ability to assemble in virus or plasmid, to survive in GIT (Gastro Intestinal
Tract), and also to trigger the oral and mucosal immune responses.
 Selection of Plant
A plant selected for the plant vaccine production should be easily available and must be the one that can grow
in the local region. The plant should be easy to transform and needs to express high quantity of protein. In
addition, the plant must not contain any toxic compound in it. An edible plant is most preferred for both human
and animal application. Currently, many plants have been used for the production of the plant vaccines such as
tobacco, potato, tomato, banana, cereals, alfalfa, etc.

8.  Insertion of the Gene in the Plant
The microbial gene can be transferred in plants by two methods-
Stable Genomic Interaction
The structural gene (antigenic gene) is loaded on a plant transformation vector. The vector is introduced into
plant cells using Agrobacterium tumefaciens chromosomal integration or by microprojectile bombardment
techniques.
Transient Expression Using Viral Vectors
The microbial DNA encoding for the epitope is inserted in a plant pathogenic virus. This genetically engineered
virus infects the plant cells and starts replication there. Along with viral genes the microbial genes get expressed
and produce protein that gets accumulated inside these cells.
 Harvesting
Expression of microbial protein occurs in various parts of the plant. The plant tissue can be consumed directly or
after cooking. Alternately, the protein can be purified by downstream processing and subsequently used in
developing tablets, capsules, and other pharmaceutical products. Generally, plant vaccines are considered safe;
however, it is important to check the potential and toxicity of the product in animal models before being used
commercially.

9. Action OF Plant Made Vaccines:
Here,
(A) plant vaccine;
(B) plant vaccine inside the GIT;
(C) release of the antigen in the intestine;
(D) absorbed of antigen by M cells;
(E) M cells pass the antigens to antigen-presenting
cells(APC);
(F) activation of T and B lymphocytes;
(G) differentiated of B cells into plasma cells;
(H)production of IgA antibody;
(I) neutralization of antigen

10. A plant vaccine administered orally stimulates the mucosal and humoral immune system of the host body.
Mucosal immune system is the primary defense system and an appropriate site for vaccination. After the
administration of plant vaccine, it stimulates the immune system in a systemic manner.
 Entry of the Antigen
Antigen present in the plant cell can be consumed in the form of raw plant products such as fruits and
vegetables. The tough outer cell wall protects the antigen from degradation by gastric enzymes and breaks to
release of the antigen in the intestine.
 Absorption of Antigen
Free antigen present in the intestine are absorbed by M cells (or microfold cells are found in the follicle-
associated epithelium of the Peyer’s patch) in the intestinal lining that are present over the Peyer’s patches (in
the ileum) and the gut-associated lymphoid tissue (GALT).
 Immune Response
M cells pass the antigens to the macrophages and other antigen-presenting cells which further present the
antigens to local T and B lymphocytes. These activated B cells migrate toward mucosal-associated lymphoid
tissue (MALT). B lymphocytes differentiate into plasma cells and synthesize IgA antibodies at mucosal surfaces
that neutralize the antigens in the intestine. In addition, few antigens taken up by the intestinal dendritic cells
(DCs) that further induces the production of IgG antibodies, imparting systemic immunity.

11. Types Of Plant Vaccines:
 Plant Vaccines Against Bacterial Infections
Pathogenic bacteria cause a variety of infections such as tetanus, typhoid fever, diphtheria, syphilis, cholera,
food-borne illness, leprosy, and tuberculosis. Tuberculosis ranks second the world’s leading causes of death
worldwide.
In year 2012, 8.6 million cases and 1.3 million deaths were reported due to tuberculosis (WHO 2013).
Streptococcus (group B Streptococcus) is another frequent cause of life-threatening infection during the first 2
months of life. Food- and water-borne bacteria such as Salmonella and Campylobacter are responsible for a
recent dramatic increase in diarrheal disease.
The discovery of new organisms and new strains of many familiar bacteria presents a sturdy challenge to the
researchers toward development of microbial control measures. There is a large and fast-growing list of target
protein/peptide from microbial pathogens that have been expressed by plants. Some of them are listed in
adapt to new environment by making changes in the structure.
Application of conventional vaccines has controlled the death rate of viral diseases; however, development of
plant vaccines could possibly provide a support to the conventional vaccines. Various viral proteins are
expressed in plants for the preparation of edible vaccine.

13. Vaccines Against Viral Infections:
Viruses cause various dreadful diseases in human and animals.
Most of the epidemic diseases are caused by viruses because
of their ability to spread fast.
Viral infections are difficult to treat, because they grow inside
the host cell and quickly adapt to new environment by making
changes in the structure. Human immunodeficiency virus,
hepatitis virus, rabies virus, variola virus, polio virus, and
Japanese encephalitis virus are some of the most common
viral pathogens. These viral pathogens cause severe infections
in humans and resulted in several death worldwide.
Application of conventional vaccines has controlled the death
rate of viral diseases; however, development of plant vaccines
could possibly provide a support to the conventional vaccines.
Various viral proteins are expressed in plants for the
preparation of edible vaccine;

14. Plant Vaccines Against Parasites:
In tropical and subtropical regions of the world, parasitic infections are leading causes of diseases and
subsequent deaths. Malaria, lymphatic filariasis, and toxoplasmosis are the most common and severe parasitic
diseases.
Malaria is an endemic disease occurring in almost 108 countries of the world and kills a large number of
people every year (WHO 2016). Nearly 30 million people in the United States are affected with toxoplasmosis
according to the report of Centers for Disease Control and Prevention (CDC) (CDC 2017).
About 120 million people are infected with lymphatic filariasis worldwide, 65% of them are present in
Southeast Asia region (CDC 2013). Development of edible vaccines to prevent the parasitic infection would
provide an alternative to the existing prophylaxis and treatment methods. At present, some parasitic proteins
are expressed in plants for the preparation of ediblevaccine,

15. Plant Vaccines Against SARS CoV -2
Plant-based vaccines for COVID-19 can be developed either by expressing the antigenic (plantigens) component
of SARS-CoV-2 for inducing active immunity or expressing the antibody (plant anti bodies) against the virus to
provide passive protection.
Plant-based vaccines are considered as third-generation vaccines. The production approach of a plant-based
vaccine involves cloning the vaccine candidate into a plant expression system, which is capable of promoting the
expression of the candidate gene in the plant, which then produces the antigenic or protective protein.
Coronaviruses (CoVs), enveloped positive-sense RNA viruses, are characterized by club-like spikes that project
from their surface, an unusually large RNA genome, and a unique replication strategy.
Coronaviruses cause a variety of diseases in mammals and birds ranging from enteritis in cows and pigs and upper
respiratory disease in chickens to potentially lethal human respiratory infections
Coronaviruses (CoVs) are the largest group of viruses belonging to the Nidovirales order, which
includes Coronaviridae, Arteriviridae, Mesoniviridae, and Roniviridae families.

16. Novel CORONA Virus :
SARS Co-2 is novel family of viruses which causes respiratory
illness. SARS stands for ‘SEVERE ACUTE RESPIRATORY SYNDROME’.
New disease which got emerged out from Wuhan, China was
named by Covid-19 which abbreviates for ‘CORONA VIRUS
DISEASE 2019.
Plant Vaccines Against Covid-19 :
Serum Institute of India (SII) has announced it has partnered
with UK biotech company, Spy biotech, a spin-off of the
University of Oxford, to develop a COVID-19
virus-like-particle (VLP)-based vaccine.
The vaccine has entered a Phase 1/2 trial in Australia.

17. VLP resembles the SARS-CoV-2 virus. The VLP is constructed incorporating all key structural proteins, mirroring
the molecular structure of coronavirus. However, it doesn't cause infection as it doesn't have the genetic
material that helps it replicate like a natural virus. VLPs can be produced using mammalian cells, insect cells,
bacteria, yeast and plant cells.
Of the five COVID-19 VLP vaccines in clinical trials listed by the World Health Organization (WHO), one plant-
based VLP vaccine is in a phase 3 clinical trial. Medicago developed the CoVLP vaccine within 20 days after
receiving the SARS-CoV-2 genetic sequence.
To design a VLP, the company selected the spike protein (S protein) of SARS-CoV-2, the most widely used target
antigen for COVID-19 vaccines among the structural envelope protein (E), membrane protein (M), and
nucleocapsid protein (N) because of its high immunogenicity and crucial role in viral cell entry.
Medicago’s first product, a quadrivalent VLP vaccine for seasonal influenza, completed the phase 3 clinical trial
in 2020 and was the first plant-derived human vaccine that was well tolerated and provided substantial
protection from influenza viruses in adults.
In the trial, 3.76, 7.5, and 15 μg of CoVLP with or without ASO3 and CpG 1018 were intramuscularly injected
into 18–55-year-olds at an interval of 21 days in a randomized, partially blinded clinical trial . The
immunogenicity results demonstrated that adjuvant formulations have a greater potential to improve humoral
and cellular immune responses to the CoVLP vaccine compared with non-adjuvant formulations. All subjects
receiving the adjuvanted vaccine developed neutralizing antibodies as the immune response after the second
dose in all dose groups.