• Edible vaccines
• Phytopharmacology and molecular pharming
• Plant vaccines
• Plant-based therapeutics
• Viral hepatitis

• COVID-19 vaccine
• oral vaccines
• transgenic plant-based oral vaccines

• bioprocess development
• chimeric vaccines
• heterologous expression
• high-throughput technology
• virus-like particles

• Chlamydomonas reinhardtii
• Chloroplast expression
• Interleukin 29 (IL29)
• TN72 strain
• Therapeutic protein

Among novel strategies proposed in pest management, synergistic agents are used to improve insecticide efficacy through an elevation of intracellular calcium concentration that activates the calcium-dependent intracellular pathway. This leads to a changed target site conformation and to increased sensitivity to insecticides while reducing their concentrations. Because virus-like particles (VLPs) increase the intracellular calcium concentration, they can be used as a synergistic agent to synergize the effect of insecticides. VLPs are self-assembled viral protein complexes, and by contrast to entomopathogen viruses, they are devoid of genetic material, which makes them non-infectious and safer than viruses. Although VLPs are well-known to be used in human health, we propose in this study the development of a promising strategy based on the use of VLPs as synergistic agents in pest management. This will lead to increased insecticides efficacy while reducing their concentrations.

• insecticide
• intracellular calcium
• pest management
• synergistic agent
• virus-like particles

• Nicotiana benthamiana
• SAG1
• adjuvant
• fusion protein
• heat shock protein
• toxoplasmosis
• vaccine

• R01 AI129807 NIAID NIH HHS
• Agencia Nacional de Promoción Científica y Tecnológica
• NIAID

• HBV
• HCV
• edible plants
• mucosal immunity
• oral vaccines

The past 30 years have seen the growth of plant molecular farming as an approach to the production of recombinant proteins for pharmaceutical and biotechnological uses. Much of this effort has focused on producing vaccine candidates against viral diseases, including those caused by enveloped viruses. These represent a particular challenge given the difficulties associated with expressing and purifying membrane-bound proteins and achieving correct assembly. Despite this, there have been notable successes both from a biochemical and a clinical perspective, with a number of clinical trials showing great promise. This review will explore the history and current status of plant-produced vaccine candidates against enveloped viruses to date, with a particular focus on virus-like particles (VLPs), which mimic authentic virus structures but do not contain infectious genetic material.

• Bunyavirales
• Flaviviridae
• Influenza virus
• alphavirus
• coronavirus
• hepatitis B virus
• human immunodeficiency virus
• newcastle disease virus
• plant molecular farming
• plant-produced vaccines
• rhabdovirus
• virus-like particles

• Cancer
• Plant-based expression system
• Recombinant proteins
• Therapeutic vaccine

• Enterotoxigenic Escherichia coli
• Nicotiana benthamiana
• immunotherapy
• monoclonal antibody
• passive immunization
• secretory IgA

• Animals
• Antibodies, Bacterial/genetics
• Antibodies, Bacterial/immunology
• Antibodies, Bacterial/metabolism
• Antibodies, Bacterial/therapeutic use
• Antibodies, Monoclonal/genetics
• Antibodies, Monoclonal/immunology
• Antibodies, Monoclonal/metabolism
• Antibodies, Monoclonal/therapeutic use
• Antibody Affinity
• Bacterial Adhesion/drug effects
• Caco-2 Cells
• Enterotoxigenic Escherichia coli/immunology
• Escherichia coli Infections/microbiology
• Escherichia coli Infections/therapy
• Gastric Acid/metabolism
• Glycosylation
• Humans
• Immunoglobulin A, Secretory/genetics
• Immunoglobulin A, Secretory/immunology
• Immunoglobulin A, Secretory/metabolism
• Immunoglobulin A, Secretory/therapeutic use
• Immunotherapy
• Mice
• Plants, Genetically Modified
• Nicotiana/genetics
• Nicotiana/metabolism

• biopharmaceuticals
• clinical trials
• magnifection
• plantibodies
• protein
• therapeutics
• vaccines

Newcastle disease is a major avian disease that causes enormous economic loss in poultry industry. There have been a number of reports on the suitability of plant-based recombinant vaccine against this disease. Fusion (F) and hemagglutinin-neuraminidase (HN) epitopes of the Newcastle disease virus (NDV) represent the major immunogenic sites for development of recombinant anti-ND vaccines in plant hosts.

The main objective of this research was to evaluate the ability of a recombinant anti-ND vaccine in induction of immune responses in animal model.

In this study, immunogenicity of recombinant fusion (F) and hemagglutinin-neuraminidase (HN) epitopes of the Newcastle disease virus (NDV) is investigated in an animal model. The corresponding genes encoding amino acids 65-81 of the F protein and 346-353 amino acids of HN were expressed in tobacco seedling using agrobacterium-mediated transformation. Expression of the foreign gene in the tobacco seedlings was investigated by a number of molecular assays including Real-Time PCR and ELISA. Transgenic plant extract was used to induce immunogenic response in animal model.

Integration of the foreign gene in plant host genome was confirmed by polymerase chain reaction (PCR). Expression of the foreign recombinant protein was confirmed by Real-Time PCR and ELISA assays. Immunogenicity of the recombinant protein was investigated in rabbit by subcutaneous injection. Results indicated that the transgenic plant extract can induce immune responses in the host as confirmed by presence of specific antibodies in the sera in ELISA assay. Western blot assays showed that the foreign gene was actually expressed in transgenic seedlings.

The results obtained in this research provide further evidence on applicability of plant-based recombinant vaccines for protection of poultry against Newcastle disease.

• ELISA
• Immunogenic Response
• Mucosal Immunity
• Newcastle Disease
• Recombinant Vaccine
• Western Blot

• Agrobacterium-mediated transformation
• Flammulina velutipes
• hepatitis B
• mating
• oral vaccine

American cockroaches (Periplaneta americana) are an important indoor allergen source and a major risk factor for exacerbations and poor control of asthma. We previously reported that allergen components from American cockroaches exhibit varying levels of pathogenicity. Sensitization to major American cockroach allergen, Per a 2, correlated with more severe clinical phenotypes among patients with allergic airway diseases.

In this study, we examined whether oral plant vaccine-encoding full-length Per a 2 clone-996 or its hypoallergenic clone-372 could exert a prophylactic role in Per a 2-sensitized mice. The cDNAs coding Per a 2–996 and Per a 2–372 were inserted into TuMV vector and expressed in Chinese cabbage. Adult female BALB/c mice were fed with the cabbage extracts for 21 days and subsequently underwent two-step sensitization with recombinant Per a 2.

Per a 2-specific IgE measured by in-house ELISA in the sera of Per a 2-372-treated groups were significantly lower than in the control groups after allergen challenge but not the Per a 2-996-treated group. Moreover, Per a 2–372 vaccine markedly decreased airway hyper-responsiveness and infiltration of inflammatory cells into the lungs, as well as reduced mRNA expression of IL-4 and IL-13 in comparison with the control mice.

Our data suggest that oral administration of edible plant vaccine encoding Per a 2 hypo-allergen may be used as a prophylactic strategy against the development of cockroach allergy.

• Administration, Oral
• Allergens/genetics
• Allergens/immunology
• Allergens/pharmacology
• Animals
• Asthma/genetics
• Asthma/immunology
• Asthma/pathology
• Asthma/therapy
• Brassica/genetics
• Brassica/immunology
• Chenopodium quinoa/genetics
• Chenopodium quinoa/immunology
• Female
• Insect Proteins/genetics
• Insect Proteins/immunology
• Insect Proteins/pharmacology
• Mice
• Mice, Inbred BALB C
• Vaccines/genetics
• Vaccines/immunology
• Vaccines/pharmacology

• Taichung Veterans General Hospital

• E1E2 heterodimer
• Hepatitis C virus
• N-glycosylation
• edible vaccine
• lettuce
• lettuce-produced proteins
• molecular farming
• transient expression

• Administration, Oral
• Animals
• Female
• HEK293 Cells
• Humans
• Immunity, Humoral
• Lactuca/genetics
• Mice, Inbred BALB C
• Plants, Genetically Modified
• Protein Engineering/methods
• Protein Multimerization
• Recombinant Proteins/genetics
• Recombinant Proteins/immunology
• Recombinant Proteins/isolation & purification
• Viral Envelope Proteins/genetics
• Viral Envelope Proteins/immunology
• Viral Hepatitis Vaccines/administration & dosage
• Viral Hepatitis Vaccines/genetics
• Viral Hepatitis Vaccines/immunology

• NSP4
• VP6
• oral vaccine
• plant-based vaccine
• rotavirus

• Chlamydomonas reinhardtii
• Microalga
• chloroplast
• recombinant product

Although viruses are simple biological systems, they are capable of evolving highly efficient techniques for infecting cells, expressing their genomes, and generating new copies of themselves. It is possible to genetically manipulate most of the different classes of known viruses in order to produce recombinant viruses that express foreign proteins. Recombinant viruses have been used in gene therapy to deliver selected genes into higher organisms, in vaccinology and immunotherapy, and as important research tools to study the structure and function of these proteins.

Virus-like particles (VLPs) are multiprotein structures that mimic the organization and conformation of authentic native viruses but lack the viral genome. They have been applied not only as prophylactic and therapeutic vaccines but also as vehicles in drug and gene delivery and, more recently, as tools in nanobiotechnology.

In this chapter, basic and advanced features of viruses and VLPs are presented and their major applications are discussed. The different production platforms based on animal cell technology are explained, and their main challenges and future perspectives are explored. The implications of large-scale production of viruses and VLPs are discussed in the context of process control, monitoring, and optimization. The main upstream and downstream technical challenges are identified and discussed accordingly.

• bioencapsulation
• chloroplast transformation
• human infectious diseases
• molecular farming
• mucosal immunity
• oral delivery

• Chloroplasts/immunology
• Communicable Diseases
• Humans
• Molecular Farming
• Poliovirus Vaccine, Oral/immunology
• Vaccination

• R01 GM063879 NIGMS NIH HHS
• R01 HL107904 NHLBI NIH HHS
• R01 HL109442 NHLBI NIH HHS

• antigen
• hepatitis B virus
• molecular farming
• plant edible vaccine
• recombinant protein
• virus-like particles

• Animals
• Biotechnology/methods
• Gene Expression
• Hepatitis B/immunology
• Hepatitis B/prevention & control
• Hepatitis B Surface Antigens/genetics
• Hepatitis B Surface Antigens/immunology
• Hepatitis B Vaccines/genetics
• Hepatitis B Vaccines/immunology
• Hepatitis B Vaccines/therapeutic use
• Hepatitis B virus/genetics
• Hepatitis B virus/immunology
• Humans
• Plants, Genetically Modified/genetics
• Vaccines, Edible/genetics
• Vaccines, Edible/immunology
• Vaccines, Edible/therapeutic use
• Vaccines, Synthetic/genetics
• Vaccines, Synthetic/immunology
• Vaccines, Synthetic/therapeutic use

• Antigen production
• Arabidopsis thaliana
• Plant vaccine
• Rift valley fever virus
• Transformation

• Enterocolitis, Necrotizing/genetics
• Enterocolitis, Necrotizing/pathology
• Epidermal Growth Factor/administration & dosage
• Epidermal Growth Factor/biosynthesis
• ErbB Receptors/genetics
• ErbB Receptors/metabolism
• Gastrointestinal Microbiome/genetics
• HeLa Cells
• Humans
• Phosphorylation
• Plants, Genetically Modified/genetics
• Seeds/genetics
• Glycine max/genetics

• R21 DK094065 NIDDK NIH HHS
• National Institutes of Health

Oral vaccines delivered as tablets offer a number of advantages over traditional parenteral-based vaccines including the ease of delivery, lack of needles, no need for trained medical personnel, and the ability to formulate into temperature-stable tablets. We have been evaluating an oral vaccine platform based on recombinant adenoviral vectors for the purpose of creating a prophylactic vaccine to prevent influenza, and have demonstrated vaccine efficacy in animal models and substantial immunogenicity in humans. These studies have evaluated monovalent vaccines to date. To protect against the major circulating A and B influenza strains, a multivalent influenza vaccine will be required.

In this study, the immunogenicity of orally delivered monovalent, bivalent, trivalent, and quadrivalent vaccines was tested in ferrets and mice. The various vaccine combinations were tested by blending monovalent recombinant adenovirus vaccines, each expressing hemagglutinin from a single strain. Human tablet delivery was modeled in animals by oral gavage in mice and by endoscopic delivery in ferrets.

We demonstrated minimal interference between the various vaccine vectors when used in combination and that the oral quadrivalent vaccine compared favorably to an approved trivalent inactivated vaccine.

The quadrivalent vaccine presented here produced immune responses that we predict should be capable of providing protection against multiple influenza strains, and the platform should have applications to other multivalent vaccines.

Vaxart, Inc.

• Ferret
• Influenza
• Mouse
• Oral
• Vaccination

Plant molecular farming (PMF), defined as the practice of using plants to produce human therapeutic proteins, has received worldwide interest. PMF has grown and advanced considerably over the past two decades. A number of therapeutic proteins have been produced in plants, some of which have been through pre-clinical or clinical trials and are close to commercialization. Plants have the potential to mass-produce pharmaceutical products with less cost than traditional methods. Tobacco-derived antibodies have been tested and used to combat the Ebola outbreak in Africa. Genetically engineered immunoadhesin (DPP4-Fc) produced in green plants has been shown to be able to bind to MERS-CoV (Middle East Respiratory Syndrome), preventing the virus from infecting lung cells. Biosafety concerns (such as pollen contamination and immunogenicity of plant-specific glycans) and costly downstream extraction and purification requirements, however, have hampered PMF production from moving from the laboratory to industrial application. In this review, the challenges and opportunities of PMF are discussed. Topics addressed include; transformation and expression systems, plant bioreactors, safety concerns, and various opportunities to produce topical applications and health supplements.

• edible vaccine
• humanized glycan
• plant molecular farming
• seed platform
• transient expression

• Hepatitis B surface antigen
• Mucosal vaccine
• Supercritical fluid extraction
• TEM
• Virus-like particle
• Zea mays

• ELISA
• Hepatitis B surface antigen
• Immunogenicity
• Pre-clinical trials
• Transgenic marker-free potato plants

The use of plants for production of recombinant proteins has evolved over the past 25 years. The first plant-based vaccines were expressed in stably transgenic plants, with the idea to conveniently deliver “edible vaccines” by ingestion of the antigen-containing plant material. These systems provided a proof of concept that oral delivery of vaccines in crude plant material could stimulate antigen-specific serum and mucosal antibodies. Transgenic grains like rice in particular provide a stable and robust vehicle for antigen delivery. However, some issues exist with stably transgenic plants, including relatively low expression levels and regulatory issues. Thus, many recent studies use transient expression with plant viral vectors to achieve rapid high expression in Nicotiana benthamiana, followed by purification of antigen and intranasal delivery for effective stimulation of mucosal immune responses.

• Administration, Oral
• Animals
• Antibody Formation/immunology
• Blotting, Western
• Freeze Drying
• Hepatitis B/immunology
• Hepatitis B Surface Antigens/immunology
• Hepatitis B Vaccines/administration & dosage
• Hepatitis B Vaccines/immunology
• Lactuca/genetics
• Lactuca/physiology
• Mice, Inbred BALB C
• Plants, Genetically Modified
• Reproducibility of Results
• Sucrose/analysis
• Temperature
• Virion/metabolism

• Arabidopsis thaliana
• Carrot
• Edible vaccine
• HIV-1 p24
• Oral immunization
• Transgenic plants

• Administration, Oral
• Animals
• Antigens/biosynthesis
• Antigens/immunology
• Bioreactors
• Humans
• Plants, Edible/metabolism
• Technology, Pharmaceutical
• Vaccination/methods
• Vaccines, Edible/administration & dosage
• Vaccines, Edible/immunology
• Vaccines, Subunit/administration & dosage
• Vaccines, Subunit/immunology

• Antigens, Plant/administration & dosage
• Antigens, Plant/genetics
• Communicable Disease Control/methods
• Communicable Disease Control/trends
• Humans
• Phytotherapy/methods
• Phytotherapy/trends
• Plants/genetics
• Plants/metabolism
• Plants, Genetically Modified/genetics
• Vaccination/methods
• Vaccination/trends
• Vaccines, Synthetic/administration & dosage
• Vaccines, Synthetic/biosynthesis
• Vaccines, Synthetic/genetics

Transmissible gastroenteritis virus (TGEV) is a commercially important pathogen of hog farms and causes contagious, lethal diarrhea in piglets. While orally and parenterally administered vaccines made from inactivated or attenuated TGEV are commercially available, they require individual administration to piglets, which is time and labor intensive, and run the risk of reversion to pathogenicity. Also, parenteral vaccines produce neutralizing serum antibodies which may be less effective against an orally transmitted pathogen, compared to an oral vaccine that would induce the production of mucosal antibodies. There has been an effort to produce subunit vaccines in an edible form in plants for convenient administration through feed. These efforts towards the expression of the S-antigen of TGEV in maize seed, its effectiveness at inducing neutralizing antibody production in the colostrum of gilts, and its efficacy in protecting piglets against challenge by virulent TGEV are summarized here.

• Allergens/genetics
• Allergens/immunology
• Allergens/metabolism
• Animals
• Antibodies/blood
• Antibodies/immunology
• Antigens, Dermatophagoides/genetics
• Antigens, Dermatophagoides/immunology
• Antigens, Dermatophagoides/metabolism
• Arthropod Proteins/genetics
• Arthropod Proteins/immunology
• Arthropod Proteins/metabolism
• Asthma/blood
• Asthma/immunology
• Asthma/therapy
• Blotting, Western
• Bronchoalveolar Lavage Fluid/chemistry
• Bronchoalveolar Lavage Fluid/immunology
• Cytokines/immunology
• Cytokines/metabolism
• Dermatophagoides farinae/genetics
• Dermatophagoides farinae/immunology
• Dermatophagoides pteronyssinus/genetics
• Dermatophagoides pteronyssinus/immunology
• Desensitization, Immunologic/methods
• Electrophoresis, Polyacrylamide Gel
• Enzyme-Linked Immunosorbent Assay
• Female
• Genetic Vectors/genetics
• Immunoglobulin E/genetics
• Immunoglobulin E/immunology
• Mice
• Mice, Inbred BALB C
• Protein Binding/immunology
• Recombinant Fusion Proteins/genetics
• Recombinant Fusion Proteins/immunology
• Recombinant Fusion Proteins/metabolism
• Nicotiana/genetics
• Nicotiana/metabolism
• Tobacco Mosaic Virus/genetics

Hepatitis B persists as a common human disease despite effective vaccines having been employed for almost 30 years. Plants were considered as alternative sources of vaccines, to be mainly orally administered. Despite 20-year attempts, no real anti-HBV plant-based vaccine has been developed. Immunization trials, based on ingestion of raw plant tissue and conjugated with injection or exclusively oral administration of lyophilized tissue, were either impractical or insufficient due to oral tolerance acquisition. Plant-produced purified HBV antigens were highly immunogenic when injected, but their yields were initially insufficient for practical purposes. However, knowledge and technology have progressed, hence new plant-derived anti-HBV vaccines can be proposed today. All HBV antigens can be efficiently produced in stable or transient expression systems. Processing of injection vaccines has been developed and needs only to be successfully completed. Purified antigens can be used for injection in an equivalent manner to the present commercial vaccines. Although oral vaccines require improvement, plant tissue, lyophilized or extracted and converted into tablets, etc., may serve as a boosting vaccine. Preliminary data indicate also that both vaccines can be combined in an effective parenteral-oral immunization procedure. A partial substitution of injection vaccines with oral formulations still offers good prospects for economically viable and efficacious anti-HBV plant-based vaccines.