Vaccine Design: Methods and Protocols, Volume 2. Vaccines for Veterinary Diseases (Methods in Molecular Biology, 2411)

Dedication
Preface
Contents
Contributors
Part I: Veterinary Vaccines: Introduction
	Chapter 1: Challenges in Veterinary Vaccine Development
		1 Introduction
		2 Severe Acute Respiratory Syndrome Coronavirus-2 (SARS-COV-2)
		3 Challenges in Development of Vaccines for Poultry
		4 Challenges in the Development oF Vaccines for Companion Animals
		5 Challenges in the Development of Vaccines for Pack Animals
			5.1 Llamas and Alpacas
			5.2 Yak
			5.3 Buffalo
		6 Challenges in the Development of Vaccines for Cattle
			6.1 Timing of Vaccination
			6.2 Vaccine Safety and Delivery Considerations
			6.3 Economic Considerations
		7 Challenges in Development of Vaccines for Sheep and Goat
		8 Challenges in the Development of Vaccines Against Ectoparasites
		9 Challenges in the Development of Fish Vaccines
		10 Conclusion
		References
Part II: Trends in Veterinary Vaccines
	Chapter 2: Mammalian Cell Culture as a Platform for Veterinary Vaccines
		1 Introduction
		2 Materials
			2.1 PEI Preparation
			2.2 Glucose Preparation
			2.3 Dulbecco´s Modified Eagle´s Medium (DMEM)
			2.4 Fetal Bovine Serum (FBS)
			2.5 CO2 Incubator
			2.6 Laminar Flow Cabinet
			2.7 Cell Culture Flasks, Dishes, and Plates
			2.8 Inverted Microscopes
		3 Methods
			3.1 Transfection
				3.1.1 Transient Transfection of HEK293 Cell Line by Using PEI (See Note 9)
				3.1.2 Lentivirus-Mediated Transduction of HEK293 Cell Line
					Lentivirus Production in HEK293-FT Cells
					Lentivirus Titration by Using an Assay Based on the HIV p24 Capsid Protein (See Note 21)
					Transduction of HEK293 Cell Line and Generation of Stable Recombinant Protein-Expressing Cell Pools in 24-Well Plates
			3.2 Obtain Protein-Expressing Recombinant Clones by Limiting Dilution (See Note 30)
			3.3 Cell Adaptation to Chemically Defined Protein-Free Media and Suspension Culture (See Note 36)
			3.4 Cryopreservation Protocol
			3.5 Growing Cell Clones in Suspension Culture in CDPFM
		4 Notes
		References
	Chapter 3: Avian Paramyxoviruses as Vectors for Vaccine Development
		1 Introduction
		2 Materials
			2.1 Preparation of a Foreign Gene (Insert) for Cloning
			2.2 Transfection
		3 Methods
			3.1 Construction of Full-Length APMV Genome Containing the Gene of a Protective Antigen
			3.2 Recovery of APMV Vectored Vaccine
		4 Notes
		References
Part III: Vaccines for Poultry
	Chapter 4: Reverse Genetics and Its Usage in the Development of Vaccine Against Poultry Diseases
		1 Introduction
			1.1 Reverse Genetics in the Context of Virology
			1.2 Reverse Genetics Platform for the Development of Poultry Vaccines
				1.2.1 Avian Influenza Virus
					Inactivated and Live Influenza Vaccines
				1.2.2 Newcastle Disease Virus
					A better Attenuated and Genotype-Matched Vaccine Against  NDV
					Bivalent NDV-Vectored Vaccines Against Poultry Viruses
					DIVA Strategy
				1.2.3 Avian Coronavirus (Infectious Bronchitis Virus)
		2 Materials
			2.1 Cell and Viruses
			2.2 Reverse Genetics Construction
			2.3 Transfection and Recovery
			2.4 Characterization of the Recombinant Virus
			2.5 Immunization and Challenge
		3 Methods
			3.1 Cells and Viruses
			3.2 Reverse Genetics Construction and Sequencing
				3.2.1 Avian Influenza
				3.2.2 Newcastle Disease Virus
				3.2.3 Infectious Bronchitis Virus
			3.3 Transfection and Recovery of the Recombinant Virus
				3.3.1 Avian Influenza
				3.3.2 Newcastle Disease Virus
				3.3.3 Infectious Bronchitis Virus
			3.4 Characterization of the Recombinant Virus
			3.5 Immunization and Challenge Study
		4 Notes
		References
Part IV: Vaccines for Farm Animals
	Chapter 5: Use of a Ferret Model to Test Efficacy and Immunogenicity of Live Attenuated Mycobacterium avium Subspecies paratub...
		1 Introduction
		2 Materials
		3 Methods
			3.1 Overall Experimental Timeline
			3.2 Monitoring and Data Collection
			3.3 Animals and Procedures
				3.3.1 Animal Type and Handling
				3.3.2 Blood Collection
			3.4 Infection of Ferrets
				3.4.1 Culture and Inoculum Preparation
				3.4.2 Infection
				3.4.3 IFN-γ Assay
				3.4.4 Serum ELISA Assay for Antibody Production
				3.4.5 Skin Test Assay
				3.4.6 Fecal Culture
				3.4.7 PCR Assay
			3.5 Experimental Endpoint
				3.5.1 Histopathology on Ferret Tissues
		4 Notes
		References
	Chapter 6: Recombinant Escherichia coli Cell Lysates as a Low-Cost Alternative for Vaccines Against Veterinary Clostridial Dis...
		1 Introduction
		2 Materials
			2.1 Strain and Plasmids
			2.2 Transformation, Expression, and Processing of E. coli Recombinant Proteins
			2.3 Polyacrylamide Gel Electrophoresis and Western Blotting
			2.4 Antigen Quantification
			2.5 Vaccine Formulation and Immunogenicity Evaluation
		3 Methods
			3.1 Transformation and Storage E. coli BL21 (DE3) with Plasmid/Gene-of-Interest Vector
			3.2 Expression of Target Protein in E. coli Cell Culture
			3.3 Preparation of Soluble Cell Fraction
			3.4 Preparation of Insoluble Cell Fraction
			3.5 SDS-PAGE and Western Blotting Analyses
			3.6 Antigen Quantification
			3.7 Vaccine Formulation
			3.8 Seroneutralization
		4 Notes
		References
	Chapter 7: Clostridium spp. Toxins: A Practical Guide for Expression and Characterization in Escherichia coli
		1 Introduction
		2 Materials
		3 Methods
			3.1 Cloning
			3.2 Protein Expression
			3.3 Characterization of the Recombinant Protein
			3.4 Antigenicity Evaluation
		4 Notes
		References
Part V: Vaccines for Veterinary Parasites
	Chapter 8: Macrophage Stimulation as a Useful Approach for Immunoscreening of Potential Vaccine Candidates Against Toxoplasma ...
		1 Introduction
		2 Materials
			2.1 Recombinant TgPrx1 Vaccine
			2.2 NcGRA6 Vaccine
			2.3 Macrophages Preparation and Stimulation
		3 Methods
			3.1 Preparation of Recombinant TgPrx1-GST Vaccine Antigen
				3.1.1 Gene Amplification and Cloning
				3.1.2 Protein Expression and Purification Procedures of TgPrx1-GST
			3.2 Preparation of rNcGRA6-GST Vaccine Antigen
				3.2.1 Gene Amplification and Cloning
				3.2.2 Protein Expression and Purification of NcGRA6-GST
			3.3 Isolation and Propagation of Thioglycolate-Elicited Peritoneal Macrophages
		4 Notes
		References
Part VI: Vaccines for Fish
	Chapter 9: Fundamentals of Fish Vaccination
		1 Introduction
		2 Definition and Properties of the Vaccine
		3 Immunological Basis of Fish Vaccine
		4 Type of Vaccine
		5 Modalities of Administration of the Fish Vaccine
		6 Challenges in Fish Vaccine Development
		7 Opportunities and Advancement for Fish Vaccine Development
		8 Correlates of Vaccine Protection Efficacy
		9 Conclusion
		References
	Chapter 10: Teleost Fish as an Experimental Model for Vaccine Development
		1 Introduction
		2 Fish Immune System
			2.1 Innate Immunity
			2.2 Adaptative Immunity
			2.3 Maternal Immunity
			2.4 Mucosal Immunity
		3 Fish as a Model for Vaccine Development
			3.1 Vaccine Delivery System
		4 Zebrafish (Danio rerio)
		References
	Chapter 11: Development of Nano-Conjugated DNA Vaccine Against Edwardsiellosis Disease in Fish
		1 Introduction
		2 Construction of Bicistronic DNA Vaccine (pGPD + IFN)
			2.1 Designing of Primers for GAPDH Gene of E. tarda and IFN-γ Gene from L. rohita
			2.2 Isolation of Genomic DNA from E. tarda
			2.3 Extraction of GAPDH Gene from Genomic DNA of E. tarda
			2.4 Extraction of IFN-γ Gene from L. rohita
			2.5 Cloning of GAPDH Gene and IFN-γ Gene in pTZ57R/T Cloning Vector
			2.6 Cloning of GAPDH Gene and IFN-γ Gene in pIRES Expression Vector
		3 Extraction of Plasmid DNA Construct (pGPD + IFN)
		4 Conjugation of pGPD + IFN with Chitosan  NPs
			4.1 Preparation of Chitosan Nanoparticles (NPs)
			4.2 Determination of Zeta Potential and Size of Nanoparticles
			4.3 Conjugation of pGPD + IFN with Chitosan  NPs
		References
	Chapter 12: Development and Evaluation of DNA Vaccine Against Salmonid Alphavirus
		1 Introduction
		2 Materials
			2.1 DNA Plasmid Preparation
			2.2 Transfection and Immunostaining of Antigen Protein
			2.3 Immunization and Virus Infection
			2.4 Neutralization Activity of Antibody Against Salmonid Alphavirus
			2.5 Tissue Sampling for Quantitative Reverse Transcription PCR (qRT-PCR)
			2.6 ELISA for Antigen-Specific Antibody Response
			2.7 Histopathology
		3 Methods
			3.1 Design the Constructs
			3.2 Production and Confirmation of the DNA Vaccines
				3.2.1 Plasmid Production and Qualification
				3.2.2 Transfection and Surface Protein Staining
			3.3 Salmonid Alphavirus Propagation and Immunofluorescence Staining
			3.4 Schedule for Vaccination, Challenge and Sampling
			3.5 Evaluation of the Vaccine Efficiency
		4 Notes
		References
	Chapter 13: Novel Vaccine Development for Fish Culture Based on the Multiepitope Concept
		1 Introduction
			1.1 Subunit Vaccine
			1.2 Multiepitope Vaccine
			1.3 Concept of Chimeric Multiepitope Vaccine
		2 Materials
			2.1 Experimental Fish, Bacterial Strain, Plasmid, Antibody, Antibiotics, and Media
			2.2 Immunoproteomics Analysis
			2.3 Chimeric Multiepitope Vaccine Design
			2.4 Codon Optimization
			2.5 Chimeric Multiepitope Vaccine Characterization
			2.6 Chimeric Multiepitope Vaccine Preparation
			2.7 Vaccine Efficacy Analysis
		3 Methods
			3.1 Immunoproteomics Analysis
				3.1.1 Immunogenic Protein Precipitation
				3.1.2 Gel-Free Digestion for Immune-Proteomics Approach
				3.1.3 Immunogenic Proteins Identification by LC-MS/MS
			3.2 Chimeric Multiepitope Vaccine Design
			3.3 Codon Optimization
			3.4 Chimeric Multiepitope Vaccine Characterization
			3.5 Chimeric Multiepitope Vaccine Preparation
				3.5.1 Construction of Plasmid Harboring Chimeric Multiepitope Vaccine
				3.5.2 Chimeric Multiepitope Protein Vaccine Expression in the Prokaryotic System
				3.5.3 Chimeric Multiepitope DNA Vaccine Expression in the Eukaryotic System
				3.5.4 Chimeric Multiepitope Recombinant Protein Vaccine Purification
				3.5.5 Chimeric Multiepitope DNA Vaccine Preparation
			3.6 Vaccine Efficacy Analysis
				3.6.1 Formalin-Killed Vaccine Preparation
				3.6.2 Fish Vaccination
				3.6.3 Challenge  Test
				3.6.4 Immunoblotting Assay
		4 Notes
		References
	Chapter 14: Testing Novel Inactivation Methods and Adjuvants for Vaccines Against Streptococcus agalactiae in Nile Tilapia Ore...
		1 Introduction
		2 Materials
			2.1 Bacterial Strain
			2.2 Growth Media
			2.3 Reagents for Vaccine Elaboration
			2.4 ELISA Reagents
			2.5 Equipment
		3 Methods
			3.1 Vaccine Preparation
				3.1.1 Formaldehyde-Inactivated Bacteria
				3.1.2 H2O2-Inactivated Bacteria
				3.1.3 Inactivation by pH Manipulation
			3.2 Vaccination and Challenge
			3.3 Inclusion de Adjuvants in a H2O2-Inactivated Vaccine
			3.4 Determination of Anti-S. agalactiae IgM Antibodies
		4 Notes
		References
Part VII: Vaccines Against Ticks
	Chapter 15: Anti-Tick Vaccines: Current Advances and Future Prospects
		1 Introduction
			1.1 Overview of Recent Advances in Anti-Tick Development
			1.2 Current Promising Anti-Tick Antigens and Commercial Vaccines
			1.3 Overview of Anti-Tick  RV
			1.4 Overview of Anti-Tick Effector Mechanisms: Gaps and Future Prospects
		2 Computational Methods in Anti-Tick Vaccine Development
			2.1 General Considerations
			2.2 Retrieving Protein Sequence  Data
			2.3 Amino Acid Sequence Analysis
			2.4 Identification of Peptide Motifs that Are Unique to Tick Protein Sequences
			2.5 Motif Mapping on the Protein 3D Structure
			2.6 Antigenicity Prediction of Tick Peptide Motifs
				2.6.1 T-Cell Receptor Binding Affinity Prediction
				2.6.2 B-Cell Receptor Binding Affinity Prediction
		3 Notes
		References
	Chapter 16: Vaccines Against Vector-Borne Diseases
		1 Introduction
		2 Materials
			2.1 Selection of Vaccine Candidates
			2.2 Cloning and Expression of Recombinant Proteins from E. coli
			2.3 Purification of Recombinant Proteins
			2.4 Immunization of Mice
			2.5 Vaccination and Challenge Studies
		3 Methods
			3.1 Selection of Vaccine Candidate
			3.2 Cloning and Expression of Recombinant Proteins from E. coli
			3.3 Expression and Purification of Recombinant Proteins from E. coli
			3.4 Purification of Recombinant Proteins (See Note 30)
			3.5 Immunization of Mice (see Note 35)
			3.6 Vaccination and Challenge Studies
				3.6.1 Assessment of the Vaccine Candidate Protein for the Prevention of Pathogen Acquisition in the Vector (see Note 38)
				3.6.2 Assessment of the Vaccine Candidate Protein for the Prevention of Pathogen Transmission to Hosts
				3.6.3 Assessment of the Vaccine Candidate Protein for the Prevention of Tick Infestation
		4 Notes
		References
	Chapter 17: A Quantum Vaccinomics Approach Based on Protein-Protein Interactions
		1 Introduction
			1.1 Vaccines and Vector-Borne Diseases
			1.2 From Quantum Biology to Quantum Immunology
			1.3 Quantum Vaccinomics
		2 Materials
			2.1 Reagents, Consumables, Kits, Equipment, and Software
			2.2 Buffers
			2.3 Cultured Tick Cells
			2.4 Antibodies from Vaccinated Cattle
		3 Methods
			3.1 General Considerations
			3.2 Yeast Two-Hybrid (Y2H)
				3.2.1 cDNA Library Construction
				3.2.2 Yeast Two-Hybrid Screen
			3.3 In Vitro Protein-Protein Interactions (Fig. 2)
				3.3.1 Protein-Protein Pull-Down
				3.3.2 Corroboration of Protein-Protein Interactions by Western Blot
			3.4 Musical Scores and Protein Interacting Domains (Fig. 3)
				3.4.1 Epistemological Bases for a Musical Characterization
				3.4.2 Data Sounding and Musical Analysis
				3.4.3 Determination of Pitches for each Amino Acid
				3.4.4 Rhythmic Characterization
				3.4.5 Formal Analysis: Melodic, Harmonic, Formal and Counterpoint Study
			3.5 Identification of Protective Epitopes or Immunological Quantum (Fig. 4)
				3.5.1 Staining Protocol
				3.5.2 Data Analysis
		4 Notes
		References
	Chapter 18: Screening for the ``Achilles Heel´´ of Hyalomma anatolicum Ticks by RNA Interference Technology and an Update on A...
		1 Introduction
		2 Materials
			2.1 Chemicals and Reagents
			2.2 Buffers
			2.3 Reference Biological Material
			2.4 Experimental Animals
		3 Methods
			3.1 Isolation of Total RNA and cDNA Synthesis
			3.2 Primer Design and Synthesis
			3.3 Amplification of FER2, TPM, and SUB Genes of H. anatolicum
			3.4 Cloning of FER2, TPM & SUB Genes of H. anatolicum in pTZ57R/T Vector System
			3.5 Plasmid Isolation from Positive Clones for Double-Stranded RNA Synthesis
			3.6 Primer Designing for RNAi
			3.7 Generation of dsRNA
			3.8 Injection of dsRNA into the Unfed Adult of H. anatolicum
			3.9 Evaluation of RNAi Effects
				3.9.1 Evaluation of RNAi Effects on Biological Activity
				3.9.2 Evaluation by Quantification of Targeted Genes
				3.9.3 Evaluation of Impact of RNAi on Cellular Architecture of Oocyte of Injected Ticks
					Tick Dissection and Fixation of Sample
					Dehydration
					Infiltration and Embedding in Glycol Methacrylate Resin
					Microtomy
					H and E Staining
		4 Notes
		5 Concluding Notes
			5.1 Future Approaches and Technologies for Novel Anti-Tick Vaccine Design
				5.1.1 Target Identification and Screening of Anti-Tick Vaccine Candidate Antigens
				5.1.2 Anti-Tick Vaccine Target Screening
			5.2 Next-Generation Anti-Tick Vaccines Designs
		References
	Chapter 19: Host Immunization with Recombinant Tick Antigen and Evaluation of Host Immune Response
		1 Introduction
		2 Materials
			2.1 For Recombinant Protein Preparation
			2.2 For Recombinant Protein Purification by Fast Protein Liquid Chromatography (FPLC)
			2.3 Mouse Immunization
			2.4 Measurement of Antibody Titers by ELISA
			2.5 Tick-Infestation Challenge
		3 Methods
			3.1 Large-Scale Recombinant Protein Synthesis and Extraction
			3.2 Purification of Recombinant Protein
			3.3 Mouse Immunization
			3.4 Blood Collection and Measurement of Antibody Titer
			3.5 Tick-Infestation Challenge
		4 Notes
		References
	Chapter 20: Fundamental Tick Vaccinomic Approach to Evade Host Autoimmune Reaction
		1 Introduction
		2 Materials
			2.1 In Silico Data Acquisition for Identifying a Unique Tick-Specific Peptide (Fig. 2)
				2.1.1 In Silico Analysis to Identify the Homology Relationship Between the Tick Peptide(s) and its Host(s)
				2.1.2 In Silico B-Cell Epitopes Prediction
			2.2 Recombinant Protein Production of Tick and Host Antigens
				2.2.1 RNA Extraction and RT-PCR
				2.2.2 Cloning and Expression of Both Animal Host and Tick sp. Recombinant Proteins
				2.2.3 Purification of Both Expressed Recombinant Proteins
			2.3 Synthetic Peptide Production
			2.4 Host Immunization Using the Validated Animal Host and Tick Recombinant Proteins and the Tick Synthetic Peptide
			2.5 Detection of Autoimmune Reaction
				2.5.1 In Vitro Autoimmune Analysis
				2.5.2 In Vivo Autoimmune Analysis
			2.6 Vaccine Efficacy by Tick Challenge
		3 Methods
			3.1 In Silico Data Acquisition to Identify a Unique Tick-Specific Peptide Sequence
				3.1.1 In Silico Analysis to Identify the Homology Relationship Between the Tick Peptide(s) and its Host(s) Sequences
				3.1.2 In Silico B-Cell Epitope Prediction
			3.2 Recombinant Protein Production of Tick and Host Antigens
				3.2.1 RNA Extraction and RT-PCR
				3.2.2 Cloning and Expression of Both Animal Host and Tick sp. Recombinant Proteins
				3.2.3 Purification of Both Expressed Recombinant Proteins
			3.3 Synthetic Peptide Production
			3.4 Host Immunization Using the Validated Animal Host, Tick Recombinant Proteins, and the Unique Tick Synthetic Peptide
			3.5 Analysis of Autoimmune Reaction
				3.5.1 In Vitro Autoimmune Analysis
				3.5.2 In Vivo Autoimmune Analysis
			3.6 Vaccine Efficacy Evaluation
		4 Notes
		References
Index