A highly conserved WDYPKCDRA epitope in the RNA directed RNA polymerase of human coronaviruses can be used as epitope-based universal vaccine design

BACKGROUND: Coronaviruses are the diverse group of RNA virus. From 1960, six strains of human coronaviruses have emerged that includes SARS-CoV and the recent infection by deadly MERS-CoV which is now going to cause another outbreak. Prevention of these viruses is urgent and a universal vaccine for all strain could be a promising solution in this circumstance. In this study we aimed to design an epitope based vaccine against all strain of human coronavirus. RESULTS: Multiple sequence alignment (MSA) approach was employed among spike (S), membrane (M), enveloped (E) and nucleocapsid (N) protein and replicase polyprotein 1ab to identify which one is highly conserve in all coronaviruses strains. Next, we use various in silico tools to predict consensus immunogenic and conserved peptide. We found that conserved region is present only in the RNA directed RNA polymerase protein. In this protein we identified one epitope WDYPKCDRA is highly immunogenic and 100% conserved among all available human coronavirus strains. CONCLUSIONS: Here we suggest in vivo study of our identified novel peptide antigen in RNA directed RNA polymerase protein for universal vaccine – which may be the way to prevent all human coronavirus disease

Immunoinformatic analysis of the SARS-CoV-2 envelope protein as a strategy to assess cross-protection against COVID-19

Envelope protein of coronaviruses is a structural protein existing in both monomeric and homo-pentameric form. It has been related to a multitude of roles including virus infection, replication, dissemination and immune response stimulation. In the present study, we employed an immunoinformatic approach to investigate the major immunogenic domains of the SARS-CoV-2 envelope protein and map them among the homologue proteins of coronaviruses with tropism for animal species that are closely inter-related with the human beings population all over the world. Also, when not available, we predicted the envelope protein structural folding and mapped SARS-CoV-2 epitopes. Envelope sequences alignment provides evidence of high sequence homology for some of the investigated virus specimens; while the structural mapping of epitopes resulted in the interesting maintenance of the structural folding and epitope sequence localization also in the envelope proteins scoring a lower alignment score. In line with the One-Health approach, our evidences provide a molecular structural rationale for a potential role of taxonomically related coronaviruses in conferring protection from SARS-CoV-2 infection and identifying potential candidates for the development of diagnostic tools and prophylactic-oriented strategies

Immunobioinformatics analysis and phylogenetic tree construction of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) in Indonesia: spike glycoprotein gene

The outbreak of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), which causes coronavirus disease 2019 (COVID-19), has spread worldwide and as a result, the World Health Organization (WHO) declared it a pandemic. At present, there are no approved vaccines against SARS-CoV-2. Therefore, the aim of this study was to predict epitope-based vaccines using bioinformatics approaches and phylogenetic tree construction of SARS-CoV-2 against the backdrop of the COVID-19 pandemic. In this study, we employed 27 isolates of SARS-CoV-2 spike glycoprotein genes retrieved from GenBank® (National Center for Biotechnology Information, USA) and the GISAID EpiCoV™ Database (Germany). We analyzed the candidate epitopes using the Immune Epitope Database and Analysis Resource. Furthermore, we performed a protective antigen prediction with VaxiJen 2.0. Data for B-cell epitope prediction, protective antigen prediction, and the underlying phylogenetic tree of SARS-CoV-2 were obtained in this research. Therefore, these data could be used to design an epitope-based vaccine against SARS-CoV-2. However, the advanced study is recommended for confirmation (in vitro and in vivo)

Immunoinformatics Patterns and Characteristic of Epitope-Based Peptide Vaccine candidates against COVID-19

Vaccination as defined by the WHO is “the administration of agent-specific, but safe, antigenic components that in vaccinated individuals can induce protective immunity against the corresponding infectious agent”. Regardless of their debated history, the standard vaccine approaches have been unsuccessful in providing vaccines for numerous infectious organisms. In the recent three decades, an enormous amount of immunological data was retrieved from clinical studies  due to the advancement in human genome sequencing. These data are being deposited in databases and numerous scientific literature. The development of several bioinformatics tools to analyze this rapidly increasing immunological databank has given rise to the field of immunoinformatics. This approach allows the selection of immunogenic residues from the pathogen genomes. The ideal residues could be industrialized as vaccine candidates to provide protective immune responses in the hosts. This methodology will significantly decrease the time and cost needed for the vaccine development.  This review focus on  published articles that proposed as vaccine candidates through immunoinformatics analysis. The reviewed  Published immunoinformatics studies provided vaccine peptide candidates against SARS-COV-2, which is based on functional and non functional immunogenic proteins like open reading frame , spike protein, envelope protein and membranous protein .All of which  are designed by unique strategies like reverse vaccinology . Spike protein was the most common used target with different suggeststed B and T cell peptides  due to the difference in methodology between the findings

Betacoronavirus genomes: How genomic information has been used to deal with past outbreaks and the COVID-19 pandemic

In the 21st century, three highly pathogenic betacoronaviruses have emerged, with an alarming rate of human morbidity and case fatality. Genomic information has been widely used to understand the pathogenesis, animal origin and mode of transmission of betacoronaviruses in the aftermath of the 2002-03 severe acute respiratory syndrome (SARS) and 2012 Middle East respiratory syndrome (MERS) outbreaks. Furthermore, genome sequencing and bioinformatic analysis have had an unprecedented relevance in the battle against the 2019-20 coronavirus disease 2019 (COVID-19) pandemic, the newest and most devastating outbreak caused by a coronavirus in the history of mankind, allowing the follow up of disease spread and transmission dynamics in near real time. Here, we review how genomic information has been used to tackle outbreaks caused by emerging, highly pathogenic, betacoronavirus strains, emphasizing on SARS-CoV, MERS-CoV and SARS-CoV-2.In the 21st century, three highly pathogenic betacoronaviruses have emerged, with an alarming rate of human morbidity and case fatality. Genomic information has been widely used to understand the pathogenesis, animal origin and mode of transmission of betacoronaviruses in the aftermath of the 2002-03 severe acute respiratory syndrome (SARS) and 2012 Middle East respiratory syndrome (MERS) outbreaks. Furthermore, genome sequencing and bioinformatic analysis have had an unprecedented relevance in the battle against the 2019-20 coronavirus disease 2019 (COVID-19) pandemic, the newest and most devastating outbreak caused by a coronavirus in the history of mankind, allowing the follow up of disease spread and transmission dynamics in near real time. Here, we review how genomic information has been used to tackle outbreaks caused by emerging, highly pathogenic, betacoronavirus strains, emphasizing on SARS-CoV, MERS-CoV and SARS-CoV-2

Specificity Determining Features at the Interface of Biomolecular Complexes as Regulators of Biological Functions

Amino acid residues at the biomolecular interface play essential roles in many biological and cellular processes; relevant to this thesis, protein-protein interactions regulate signaling pathways and enzymatic activity, whereas protein-DNA interactions control gene expression, and protein-peptide interactions are central to the immune system. Biomolecular recognition and binding stability are largely determined by residues at the molecular interface. In this thesis, we focused on three biological datasets that are related to humans and human health: 1) dysregulated citrullination in the inflamed joints of rheumatoid arthritis patients, 2) a novel family of PRD-like transcription factors critical to the first few cell divisions in human life, and 3) epitopes that likely activate a cytotoxic T cell-mediated immune response against SARS-CoV-2 infection. For each dataset, in order to study the structural and functional consequences of molecular interactions, we applied a wide range of bioinformatics techniques to analyze sequences, structures and biological data retrieved from various databases, as well as taking into account experimental results from collaborators and from the literature. In rheumatoid arthritis, normally cytoplasmic peptidylarginine deiminase (PAD) enzymes citrullinate arginine residues in extracellular matrix (ECM) proteins. To examine specificity determining features that regulate the citrullination activity, we analyzed the sequence and structure data of the ECM proteins that were found citrullinated in chronically inflamed human joints. For citrullination, we found that an arginine side chain needs to be exposed to solvent but can arise from β-strands, α-helices, loops and β-turns. Moreover, there is no sequence motif linked to enzymatic activity. In addition, we studied the effect of citrullination on proteins important for a normal ECM, focusing on integrin binding to fibronectin and transforming growth factor-β (TGF-β). Citrullination of these proteins was found to inhibit cell attachment and spreading since PAD-treatment of the isoDGR motif in fibronectin and the RGD motif in TGF-β significantly reduced their binding with integrin αVβ3 and αVβ6, respectively. The expression of the human paired (PRD)-like transcription factors (TFs) are limited to the period of embryonic genome activation up to the 8-cell stage. We identified that one of these PRD-like TFs, LEUTX, binds to a TAATCC sequence motif. Sequence comparisons revealed that LEUTX protein is comprised of two domains: the DNA-binding homeodomain and a Leutx domain containing a transactivation domain. We identified specificity determining residues in the LEUTX homeodomain that are important for recognition of the TAATCC-containing 36 bp DNA motif enriched in genes involved in embryonic genome activation. We demonstrated using molecular models why a heterozygotic missense mutation A54V at the DNA-specificity determining position of LEUTX has significantly reduced overall transcriptional activity, as well as why the double mutant – I47T and A54V – form of LEUTX restores binding to the DNA motif similarly to that seen in the I47T mutation alone. At the onset of the COVID-19 pandemic we sought to understand the molecular factors that trigger the cytotoxic T cell-mediated immune response against the SARS-CoV-2 virus, taking advantage of binding data and 3D structures for related viruses and other pathogenic organisms. We first predicted the MHC class I (MHC-I)-specific immunogenic epitopes of length 8- to 11 amino acids from the SARS-CoV-2 proteins. Next, we predicted that the 9-mer epitopes would have the highest potential to elicit a strong immune response. For experimental validation, the predicted 9-mer epitopes were matched with the SARS-CoV-derived epitopes that are known to elicit an effective T cell response in vitro. Furthermore, our observations provide a structural explanation for the binding of SARS-CoV-2 epitopes to MHC-I molecules, identifying conserved immunogenic epitopes essential for understanding the pathogenesis of COVID-19. The three investigated datasets were made in concert with collaborative experimental studies and/or considering publicly available experimental data. The experimental studies generally provided the starting point for the in silico studies, which in turn had the objective of providing a detailed explanation of the experimental results. Furthermore, the in silico results could be used to devise novel and focused experiments, suggesting that bioinformatics predictions and wet-laboratory experimental investigations optimally take place with multiple advantages. Overall, this thesis demonstrates the synergy that is possible by applying this interdisciplinary approach to understanding the consequences of molecular interactions.Aminosyror i kontaktytan mellan olika biomolekyler spelar en viktig roll i många biologiska och cellulära processer; relevanta interaktioner för den här avhandlingen är protein-protein interaktioner som reglerar signaleringsrutter och enzymatisk aktivitet, protein-DNA interaktioner som kontrollerar genexpression, samt protein-peptid interaktioner som har en central roll i immunförsvaret. Biomolekylär igenkänning och bindningsstabilitet beror till stor del på de aminosyror som finns i den molekylära kontaktytan. I den här avhandlingen fokuserade vi på tre biologiska dataset som är relaterade till människor och människors hälsa: 1) felreglerad citrullinering i inflammerade leder hos patienter med reumatoid artrit, 2) en nyupptäckt familj av PRD (human paired)-lika transkriptionsfaktorer som är nödvändiga för de första celldelningarna i människolivet, och 3) epitoper som troligen aktiverar en cytotoxisk T-cell-förmedlad immunrespons mot SARS-CoV-2 infektioner. För att studera de strukturella och funktionella konsekvenserna av de molekylära interaktionerna i varje dataset, användes en mängd olika bioinformatiska tekniker för att analysera sekvenser, strukturer och biologiska data från olika databaser och dessutom beaktades experimentella resultat från samarbetspartners och från litteraturen. I reumatoid artrit citrullinerar vanligen PAD (cytoplasmatiska peptidyl arginin deiminas)-enzymer arginin-aminosyror i proteiner i det extracellulära matrixet (ECM). För att undersöka egenskaper som avgör specificiteten hos citrullineringsaktiviteten analyserade vi sekvens- och strukturdata för ECM-proteiner som blir citrullinerade i kroniskt inflammerade leder hos människor. Vi upptäckte att en argininsidokedja måste vara i kontakt med det omgivande lösningsmedlet för att kunna citrullineras, att de kan finnas i beta-strängar, alfa-helixar och beta-svängar, samt att det inte finns några sekvensmotiv som är kopplade till enzymatisk aktivitet. Utöver detta studerade vi effekten av citrullinering på proteiner som är viktiga för normal extracellulär matrix, med fokus på integrinbinding till fibronektin och TGF-β (transforming growth factor-β). Citrullinering av dessa proteiner upptäcktes inhibera cellvidhäftning och spridning eftersom PAD-behandling av isoDGR-motivet i fibronektin och RGD-motivet i TGF-β ordentligt reducerar deras bindning till integrin αVβ3 och αVβ6, respektive. Expressionsnivåerna av PRD-lika transkriptionsfaktorer (TF) är begränsade till perioden av zygotens genomaktivering upp till 8-cells stadiet. Vi identifierade att en av dessa PRD-lika transkriptionsfaktorer, LEUTX, binder till ett TAATCC sekvensmotiv. Sekvensjämförelser avslöjade att LEUTX proteinet består av två domäner, det DNA-bindande homeodomänet och en leutx-domän som innehåller en transaktiveringsdomän. Vi identifierade specificitetsbestämmande aminosyror i LEUTX homeodomänen som är viktiga för igenkänning av TAATCC-innehållande 36 baspars DNA-motivet som är berikad med gener involverade i zygotens genomaktivering. Vi använde molekylära modeller för att visa varför en heterozygotisk missense-mutation, A54V, i DNA-specificitetsbestämmande positionen i LEUTX har ordentligt minskad generell transkriptionsaktivitet, och varför dubbelmutanten I47T och A54V återställer bindning till DNA-motivet på samma sätt som observerats i enbart I47T mutationen. När COVID-19 pandemin inleddes försökte vi förstå de molekylära faktorer som startar den cytotoxiska T-cell-förmedlade immunresponsen mot SARS-CoV-2 viruset, genom att utnyttja bindningsdata och 3D strukturer för relaterade virus och andra patogena organismer. Vi förutspådde först MHC klass I (MHC-I)-specifika immunogena epitoper av längden 8 till 11 aminosyror från SARS-CoV-2 proteiner. Därefter förutspådde vi att epitoper bestående av 9 aminosyror hade den högsta potentialen att orsaka en stark immunrespons. För experimentell validering matchades de 9 aminosyror långa epitoperna med epitoper från SARS-CoV som man vet att orsakar en effektiv T-cell respons in vitro. Våra observationer bidrar också med en strukturell förklaring för bindningen av SARS-CoV-2 epitoper till MHC-I molekyler, vilket identifierar konserverade immunogena epitoper som är nödvändiga för att förstår patogenesen hos COVID-19. De tre undersökta dataseten gjordes i samarbete med experimentella studier och/eller genom att ta allmänt tillgängliga experimentella data i beaktande. De experimentella studierna gav en startpunkt för in silico-studierna, vilka i sin tur hade som mål att ge en detaljerad förklaring till de experimentella resultaten. In silico-resultaten kan också användas för att utveckla nya och fokuserade experiment, vilket indikerar att bioinformatiska förutspåelser och experimentella studier optimalt sker med många fördelar. Över lag visar denna avhandling synergin som är möjlig genom att använda detta interdisciplinära arbetssätt för att förstå konsekvenserna av molekylära interaktioner

사람 코로나바이러스 (SARS-CoV-2)와 돼지 코로나바이러스 (PEDV)의 유전학적 분석과 유전적 변이가 바이러스 항원성과 진단에 영향을 미칠 가능성

학위논문 (박사) -- 서울대학교 대학원 : 수의과대학 수의학과, 2021. 2. 박용호.바이러스는 면역 회피를 유발할 수 있는 유전적 돌연변이를 통해 숙주 면역과 계속해서 싸우고 있으며, 이러한 바이러스의 생존 전략은 앞으로도 계속 될 것이다. 특히, 유전자 변이에 따른 아미노산 서열의 비상동성 (non-synonymous) 변화는 바이러스 에피톱의 항원성을 변화시킬 수 있으며, 이러한 변화는 기존에 개발된 백신의 방어능을 저하시킬 수 있다. 또한, 바이러스의 염기서열의 변이는 현재 일상적으로 사용되는 진단 기술인 중합효소연쇄반응 (PCR)과 효소결합면역흡착분석법 (ELISA)의 진단 정확도를 저해 할 수 있다. 따라서 바이러스가 진화함에 따른 중요한 그들의 유전학적 변화를 조사하고 추적하는 것은 바이러스에 대한 적절한 예방 및 진단 전략을 수립하는 데 매우 큰 도움이 된다. 이 연구에서는 현재 사람과 돼지에서 심각한 문제가 되고 있는 코로나바이러스의 유전적 변이와 그 변이들이 바이러스의 항원성과 진단에 영향을 미칠 가능성에 대해 조사하였다. 첫 번째 장에서는 최근 사람에서 문제가 되고 있는 severe acute respiratory syndrome coronavirus type 2 (SARS-CoV-2)를 분석하였다. 코로나바이러스의 스파이크 (S) 단백질은 바이러스의 세포 내 유입에 결정적인 역할을 하는 표면 단백질이다. 따라서, 이 연구에서는 항원성과 면역학적 특징을 확인하기 위해 SARS-CoV-2의 S 유전자를 분석하였다. S 유전자를 기반으로 한 계통학적 분석에서 SARS-CoV-2 분리주들 사이에 두 개의 유전자 그룹이 존재하는 것을 확인하였다. 이 두 개의 유전자 그룹은 하나의 특이적인 염기서열 변이인 D614G에 의해 나뉘었다. 이 변이는 SARS-CoV-2가 숙주의 면역체계를 회피하는데 결정적인 역할을 할 것으로 생각되었다. D614G 염기서열 변이를 포함하는 S1 domain의 에피톱 부위에 대해 항원 지수 분석을 시행한 결과, SARS-CoV-2b 유전자 그룹이 SARS-CoV-2a 유전자 그룹에 비해 유의적으로 감소한 항원 지수를 보이는 것으로 확인하였다. 따라서, 이 유전적 변이에 의해 두 유전자 그룹간의 항원성 차이가 발생하였을 것으로 생각되었다. 두 유전자 그룹간 항원성 차이가 발생하였다면 두 유전자 그룹을 백신에 포함시키는 것이 COVID-19을 방어하는데 보다 효율적일 것이다. 그러므로, 실제로 두 유전자 그룹간 항원성 차이가 발생하였는지 확인하는 것이 시급하다. 두 번째 장에서는 전 세계 돼지 산업에 지속적이고 심각한 피해를 입히고 있는 돼지유행성설사바이러스 (PEDV)를 분석하였다. 최근 양돈장의 PEDV 유병률은 약 9.92 %로 지속적으로 문제가 되고 있음이 확인되었다. 뉴클레오캡시드 (N) 유전자를 기반으로 베이지안 계통 분석을 진행한 결과, 세 개의 주요 N 유전자 기반 유전자 그룹 (N1, N2 및 N3)과 두 개의 하위 유전자 그룹 (N3a과 N3b) 을 확인하였다. N 단백질에 포함된 에피톱 부분의 항원 지수를 분석한 결과, 유전자 그룹간 항원성에 차이가 있을 것으로 강하게 의심되었다. 에피톱 부위에서 N3 유전자 그룹의 항원 지수는 N1 및 N2 유전자 그룹의 항원 지수에 비해 유의하게 낮았다. 이러한 변화는 N1 단백질을 항원으로 사용하는 ELISA 키트의 진단 결과에 영향을 미칠 것으로 판단되었다. 또한, 최근 확인된 한국 PED 바이러스들의 S 유전자를 분석한 결과, 스파이크 단백질 (COE, S1D 및 2C10)의 B 세포 에피토프 서열의 일부에서 유의적으로 낮은 항원 지수가 확인되었다. 이러한 S 및 N 유전자의 면역학적 주요 부위에 유전적 변이가 발생한 PED 바이러스들은 기존에 확립된 숙주 면역을 회피하여 돼지 농장에 심각한 손상을 줄 수 있기 때문에 지속적인 감시가 필요하다. 이 연구에서 바이러스의 면역 회피나 진단 오류를 유발할 수 있는 중요한 유전적 변이를 현재 심각한 문제가 되고 있는 사람과 돼지 코로나바이러스에서 확인하였다. 이러한 발견은 바이러스 감염 예방에 대한 더 나은 이해와 보다 정확한 진단법을 개발하는데 도움을 줄 것으로 기대한다. 나아가 향후 코로나바이러스 진화에 적절히 대응할 수 있도록 유전자 분석을 통한 지속적인 감시가 유지되어야 한다.Aspect of virus evolution, viruses have continued to fight with host immune through genetic mutation facilitating immune evasion and this strategy of viruses for their survival will continue in the future. Genetic mutation, which may change structural form of viral proteins by non-synonymous changes, may alter antigenicity of a viral epitope and in turn can lead to decreased efficacy of previously developed vaccine for the virus. Furthermore, such genetic mutation can hamper diagnostic accuracy of polymerase chain reaction (PCR) and enzyme-linked immunosorbent assay (ELISA), which are routinely used diagnostic techniques. Thus, it is very important work to investigate and track critical genetic events along with virus evolution. These efforts can give worthy information and insight to establish appropriate prevent and diagnostic strategy for viruses. Herein, human and animal coronaviruses causing sever disease were investigated by genetic and phylogenetic analysis. As stated in chapter I, The S glycoprotein of coronaviruses is important for viral entry and pathogenesis with most variable sequences. Therefore, we analyzed the S gene sequences of SARS-CoV-2 to better understand the antigenicity and immunogenicity of this virus in this study. In phylogenetic analysis, two subtypes (SARS-CoV-2a and -b) were confirmed within SARS-CoV-2 strains. These two subtypes were divided by a novel non-synonymous mutation of D614G. This may play a crucial role in the evolution of SARS-CoV-2 to evade the host immune system. The region containing this mutation point was confirmed as a B-cell epitope located in the S1 domain, and SARS-CoV-2b strains exhibited severe reduced antigenic indexes compared to SARS-CoV-2a in this area. This may allow these two subtypes to have different antigenicity. If the two subtypes have different serological characteristics, a vaccine for both subtypes will be more effective to prevent COVID-19. Thus, further study is urgently required to confirm the antigenicity of these two subtypes. As stated in chapter I, Porcine epidemic diarrhea virus (PEDV) causes continuous, significant damage to the swine industry worldwide. By RT-PCR-based methods, this study demonstrated the ongoing presence of PEDV in pigs of all ages in Korea at the average detection rate of 9.92%. By the application of Bayesian phylogenetic analysis, it was found that the nucleocapsid (N) gene of PEDV could evolve at similar rates to the spike (S) gene at the order of 10−4 substitutions/site/year. Based on branching patterns of PEDV strains, three main N gene-based genogroups (N1, N2, and N3) and two sub-genogroups (N3a, N3b) were proposed in this study. By analyzing the antigenic index, possible antigenic differences also emerged in both the spike and nucleocapsid proteins between the three genogroups. The antigenic indexes of genogroup N3 strains were significantly lower compared with those of genogroups N1 and N2 strains in the B-cell epitope of the nucleocapsid protein. Indeed, there is different antigenicity between the genogroups based on the N gene, it may affect diagnostic results using commercial ELISA kits based on N1 protein. Similarly, significantly lower antigenic indexes in some parts of the B-cell epitope sequences of the spike protein (COE, S1D, and 2C10) were also identified. PEDV mutants derived from genetic mutations of the S and N genes may cause severe damage to swine farms by evading established host immunities. In conclusion, the crucial genetic variations, which may induce immune evasion or diagnostic error, were revealed in these coronavirus. It is expected that these results provide better understanding for preventing viral infection and more precise diagnosis. Also, constant surveillances through genetic analysis should maintain to appropriately respond to coronavirus evolution in the future.Abstract iv List of Figures ix List of Tables x General introduction 11 Literature review 14 1. Coronavirus 15 1.1. General overveiw 17 1.2. SARS-CoV-2: Human betacoronavirus 17 1.3. Porcine epidemic diarrhea virus: Animal alphacoronavirus 19 2. Virus evolution and genetic analysis on viruses 23 2.1. Antigenic drift and genetic shift 23 2.2. Genetic mutation and recombination 23 2.3. Mutation rate of DNA and RNA viruses 25 2.4. Phenotypic Variation by Mutations 26 2.5. Phylogenetic analysis on viruses 26 3. Impact of genetic mutation on viral antigenicity and diagnosis 28 Chapter I 31 Abstract 32 1. Introduction 33 2. Material and Methods 34 2.1. Sample collection and Phylogenetic analysis 34 2.2. Epitope prediction and antigenic index analysis on S gene sequences 34 3. Results 35 3.1. Phylogenetic analysis on S gene of SARS-CoV-2 35 3.2. Epitope prediction of S protein 35 3.3. Antigenic index analysis on the epitope of S1 subunit 35 4. Discussion 36  Chapter II 43 Abstract 44 1. Introduction 45 2. Materials and Methods 47 2.1. Sample collection, PEDV detection by PCR, and complete sequencing 47 2.2. Genetic analysis of recombination 48 2.3. Bayesian phylogenetic analysis 50 2.4. Pairwise genetic distance (p-Distance) analysis 51 2.5. Inferring ancestral amino acid changes 51 2.6. Amino acids and antigenic index analysis of N gene 51 2.7. Antigenic index analysis of B-cell epitopes in Korean PEDV strains 52 3. RESULTS 53 3.1. The detection of PEDV in Korea from 2017 to 2018 53 3.2. Phylogenetic analysis of global PEDV strains 56 3.3. Evolutionary rates of PEDV genes 60 3.4. Amino acids and antigenic index analysis of N gene sequences 60 3.5. Antigenic index analysis of S protein B-cell epitopes in Korean PEDV strains 61 4. Discussion 66 General conclusions 69 References 71 국문 초록 88Docto

Novel epitope based peptides for vaccine against SARS-CoV-2 virus: immunoinformatics with docking approach

Background: Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is the causative viral strain for the contagious pandemic respiratory illness in humans which is a public health emergency of international concern. There is a desperate need for vaccines and antiviral strategies to combat the rapid spread of SARS-CoV-2 infection.Methods: The present study based on computational methods has identified novel conserved cytotoxic T-lymphocyte epitopes as well as linear and discontinuous B-cell epitopes on the SARS-CoV-2 spike (S) protein. The predicted MHC class I and class II binding peptides were further checked for their antigenic scores, allergenicity, toxicity, digesting enzymes and mutation.Results: A total of fourteen linear B-cell epitopes where GQSKRVDFC displayed the highest antigenicity-score and sixteen highly antigenic 100% conserved T-cell epitopes including the most potential vaccine candidates MHC class-I peptide KIADYNYKL and MHC class-II peptide VVFLHVTYV were identified. Furthermore, the potential peptide QGFSALEPL with high antigenicity score attached to larger number of human leukocyte antigen alleles. Docking analyses of the allele HLA-B*5201 predicted to be immunogenic to several of the selected epitopes revealed that the peptides engaged in strong binding with the HLA-B*5201 allele.Conclusions: Collectively, this research provides novel candidates for epitope-based peptide vaccine design against SARS-CoV-2 infection

The global emergency of novel coronavirus (SARS-CoV-2). An update of the current status and forecasting

Over the past two decades, there have been two major outbreaks where the crossover of animal Betacoronaviruses to humans has resulted in severe acute respiratory syndrome coronavirus (SARS-CoV) and Middle East respiratory syndrome coronavirus (MERS-CoV). In December 2019, a global public health concern started with the emergence of a new strain of coronavirus (SARS-CoV-2 or 2019 novel coronavirus, 2019-nCoV) which has rapidly spread all over the world from its origin in Wuhan, China. SARS-CoV-2 belongs to the Betacoronavirus genus, which includes human SARS-CoV, MERS and two other human coronaviruses (HCoVs), HCoV-OC43 and HCoV-HKU1. The fatality rate of SARS-CoV-2 is lower than the two previous coronavirus epidemics, but it is faster spreading and the large number of infected people with severe viral pneumonia and respiratory illness, showed SARS-CoV-2 to be highly contagious. Based on the current published evidence, herein we summarize the origin, genetics, epidemiology, clinical manifestations, preventions, diagnosis and up to date treatments of SARS-CoV-2 infections in comparison with those caused by SARS-CoV and MERS-CoV. Moreover, the possible impact of weather conditions on the transmission of SARS-CoV-2 is also discussed. Therefore, the aim of the present review is to reconsider the two previous pandemics and provide a reference for future studies as well as therapeutic approaches