NEXT GENERATION SEQUENCING(NGS) BASED REVERSE VACCINOLOGY APPROACHES FOR THE DEVELOPMENT OF VACCINES AGAINST PATHOGENS
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Next generation sequencing and reverse vaccinology approaches allow for vaccine development directly from pathogen genomic data. Reverse vaccinology was first used in the 1990s to develop a vaccine for Neisseria meningitidis B by sequencing its genome, predicting potential antigens, and testing them experimentally. The workflow involves sequencing pathogen genomes, predicting proteins and epitopes, filtering for antigenic proteins, predicting T-cell and B-cell epitopes, evaluating epitopes, predicting structures, and designing vaccine candidates for testing.
NEXT GENERATION SEQUENCING(NGS) BASED REVERSE VACCINOLOGY APPROACHES FOR THE DEVELOPMENT OF VACCINES AGAINST PATHOGENS
- 1. NEXT GENERATION SEQUENCING(NGS) BASED REVERSE VACCINOLOGY APPROACHES FOR THE DEVELOPMENT OF VACCINES AGAINST PATHOGENS Present by Gowdham M
- 2. Reverse Vaccinology: Reverse Vaccinology is a new approach to design vaccines from genomic information with the help of bioinformatics. History Reverse vaccinology was first used to predict potential antigens for a vaccine against the B strains of Neisseria meningitidis (meningococci) in the 1990’s. Rappuoli team at the J. Craig Venter Institute first sequenced the N. meningitidis B genome. They scanned the sequenced genome for potential antigens. They found over 600 possible antigens, which were tested by expression in Escherichia coli. Several proved to function successfully in mice, however, not alone for a good immune response. Later, the vaccine was proven to be safe and effective in adult humans
- 3. Overview of workflow Drug Designing Potential antigens Epitopes Prediction Protein Filtrations Proteomes Analysis Genome Analysis NGS data
- 4. Various NGS Platform illumina Ion Torrent Pacific Biosciences Roche 454 SOLiD Sequencing Nanopore
- 5. Sequenced Genomes Assembled Genomes Unassembled Genome Data’s Transcriptomic Data’s Existing data from Primary Biological databases
- 6. Assembled Genomes Gene predictions AUGUSTUS FGENESH GeneMark GLIMMER GLIMMERHMM GENSCAN ORF FINDER mGENE Unassembled Genome Data’s Quality Control (QC) Analysis Remove Low Quality Sequences Trimming Adapters from sequences Remove Short Read Sequences (<10 bp) Homopolymer Sequences Trimming(Repeat) Ambiguity Filtering(Optional) Genome Assembly ABySS,ALE-Assembly,Allpaths-LG, ALLPATHS,*CANU,*FALCON,*MaSuRCA *Velvet,*SPAdes Gene predictions Transcriptomic Data’s Genome Assembly Trinity SPAdes Velvet/Oases Protein Predictions TransDecoder
- 8. Proteomes (NGS Proteomes/ Primary Protein Databases) Blast against host proteomes and remove homologues proteins (Blastp program) Prioritizing of proteins based on subcellular Localization (DeepLoc,Targetp,SignaP, Phobius,WoLF PSORT,PSORTb) Detection of Virulent proteins( VFDB,Victors,etc..)rs Functional Domain Annotations (Pfam, Interproscan) Pathways (KEGG) Membrane Protein Predictions (TMHMM)
- 9. Proteomes (NGS Proteomes/ Primary Protein Databases) Antigenic Protein Determination ( Vaxijen server) Physical and Chemical Properties (Molecular Weight, Theoretical Pi, Amino Acid Composition, Atomic Composition, Extinction Coefficient, Estimated Half-Life, Instability Index, Aliphatic Index and Grand Average of Hydropathicity (GRAVY)) Protein-Protein Interaction Analysis (STRING,BioGRID) Ortholog Analysis (Orthovenn Webserver)
- 10. Filtered Antigenic Proteins T-cell Epitope Predictions MHC-I Prediction IEDB MHC –I predictor NETMHCPAN-I server MHC-II Prediction IEDB MHC -II predictor NETMHCPAN-II server B-cell Epitope Predictions Linear B-cell Epitope Predictions Bepipred Linear Epitope Chou & Fasman Beta-Turn Emini Surface Accessibility Karplus & Schulz Flexibility Kolaskar & Tongaonkar Antigenicity Parker Hydrophilicity Discontinuous B-cell Epitope Predictions DiscoTope Ellipro Epitopia
- 11. Predicted Epitopes Antigenecity (Vaxijen server) Allergenicity (AllergenPro,AllerBase, AllergenOnline, AllerTOP,Allerdictor) Physical and Chemical Properties Population Coverage and Epitope Conservancy Analysis Epitope Clustering EpitopeEvaluations
- 12. Epitopes Structures Predictions Secondary Structure Analysis (PSIPRED) Tertiary structure prediction, Refinement and Validations. (PEPstrMOD, PEP-FOLD) Protein-Peptide Docking CABS-dock server, HPEPDOCK Server, Galaxy PepDock, Rosetta FlexPepDock, Gold, Schrodinger
- 13. Vaccines Designing Clustering Epitopes Sequences 3D Structures of Epitopes HLA allele 3D structures of Allele From PDB Docking of Allele and Epitopes Toxicity prediction of docked peptides MD Simulation of Selected peptides from Toxicity prediction Clinical Trail studies