Development Of Effective Vaccine Research Articles

An immunoinformatic investigation on Rift Valley fever virus protein reveals possible epitopes for vaccines.

This immunoinformatic study identified potential epitopes from the envelopment polyprotein (Gn/Gc) of Rift Valley fever virus (RVFV), a pathogenic virus causing severe fever in humans and livestock. Effective vaccination is crucial for controlling RVFV outbreaks. The identification of suitable epitopes is crucial for the development of safe and effective vaccines. Protein sequences were obtained from the UniProt database, and evaluated through VaxiJen v2.0 to predict the B and T-cell epitopes within the RVFV glycoprotein. Gn/Gc protein sequences were analyzed with bioinformatics tools and algorithms. The predicted T-cell and B-cell epitopes were evaluated for antigenicity, allergenicity, and toxicity by the VaxiJen v2.0 system, AllerTop v2.0, and ToxinPred server, respectively. We employed computational methods to screen the RVFV envelopment polyprotein encompassing N-terminal and C-terminal glycoprotein segments, to discover antigenic T- and B-cell epitopes. Our analysis unveiled multiple potential epitopes within the RVFV glycoprotein, specifically within the Gn/Gc protein sequences. Subsequently, we selected eleven cytotoxic T-lymphocytes (CTL) and four helper T-lymphocytes (HTL) for population coverage analysis, which collectively extended to cover 97.04% of the world's population, representing diverse ethnicities and regions. Notably, the CTL epitope VQADLTLMF exhibited binding affinity to numerous human leukocyte antigen (HLA) alleles. The identification of glycoprotein (Gn/Gc) epitopes through this immunoinformatic study bears significant implications for advancing the development of an effective RVFV vaccine. These findings provide valuable insights into the immunological aspects of the disease and may contribute towards the development of broad-spectrum antiviral therapies targeting other RNA viruses with similar polymerase enzymes.

From pathogenesis to antigens: the key to shaping the future of TB vaccines

Tuberculosis (TB) remains one of the gravest global health challenges. Mycobacterium tuberculosis (M. tuberculosis), the causative agent, employs sophisticated immune evasion and pathogenesis strategies. Its capability to thrive within immune cells and incite robust inflammatory responses prolongs infection and dissemination. Mycobacterial advanced adaptations facilitate navigation through the human immune system and present a variable antigenic profile throughout different infection stages. Investigating these strategies unfolds targeted approaches to effective vaccine development against TB. This review delves into the most advanced and exhaustive insights into the immune evasion tactics and pathogenic processes of M. tuberculosis across various infection stages. The knowledge distilled from this analysis holds the promise of guiding the creation of innovative TB vaccines and translating theoretical groundwork into practical immunological defenses.

Licensed H5N1 vaccines generate cross-neutralizing antibodies against highly pathogenic H5N1 clade 2.3.4.4b influenza virus.

Global emergence of highly pathogenic avian influenza (HPAI) H5N1 clade 2.3.4.4b viruses, and their transmission to dairy cattle and animals, including humans, pose a significant global public health threat. Therefore, development of effective vaccines and therapeutics against H5N1 clade 2.3.4.4b virus is considered a public health priority. In the U.S., three H5N1 vaccines derived from earlier strains of HPAI H5N1 (A/Vietnam; clade 1 and A/Indonesia; clade 2.1) virus, with (MF59 or AS03) or without adjuvants, are licensed and stockpiled for pre-pandemic preparedness, but whether they can elicit neutralizing antibodies against circulating H5N1 clade 2.3.4.4b viruses is unknown. In this study, we evaluated the binding, hemagglutination inhibition and neutralizing antibody response generated following vaccination of adults with the three licensed vaccines. Individuals vaccinated with the two adjuvanted licensed H5N1 vaccines generate cross-reactive binding and cross-neutralizing antibodies against the HPAI clade 2.3.4.4b A/Astrakhan/3212/2020 virus. Seroconversion rates of 60% to 95% against H5 clade 2.3.4.4b were observed following two doses of AS03-adjuvanted-A/Indonesia or three doses of MF59-adjuvanted-A/Vietnam vaccine. These findings suggest that the stockpiled U.S. licensed adjuvanted H5N1 vaccines generate cross-neutralizing antibodies against circulating HPAI H5N1 clade 2.3.4.4b in humans and may be useful as bridging vaccines until updated H5N1 vaccines become available.

Genetic diversity and natural selection analysis of VAR2CSA and vir genes: implication for vaccine development

Variable surface antigens (VSAs) encoded by var and vir genes in Plasmodium falciparum and Plasmodium vivax, respectively, are known to be involved in malaria pathogenesis and host immune escape through antigenic variations. Knowledge of the genetic diversity of these antigens is essential for malaria control and effective vaccine development. In this study, we analysed the genetic diversity and evolutionary patterns of two fragments (DBL2X and DBL3X) of VAR2CSA gene and four vir genes (vir 4, vir 12, vir 21 and vir 27) from different endemic regions, including Southeast Asia and sub-Saharan Africa. High levels of segregating sites (S) and haplotype diversity (Hd) were observed in both var and vir genes. Among vir genes, vir 12 (S = 131, Hd = 0.996) and vir 21 (S = 171, Hd = 892) were found to be more diverse as compared to vir 4 (S = 11, Hd = 0.748) and vir 27 (S = 23, Hd = 0.814). DBL2X (S = 99, Hd = 0.996) and DBL3X (S = 307, Hd = 0.999) fragments showed higher genetic diversity. Our analysis indicates that var and vir genes are highly diverse and follow the similar evolutionary pattern globally. Some codons showed signatures of positive or negative selection pressure, but vir and var genes are likely to be under balancing selection. This study highlights the high variability of var and vir genes and underlines the need of functional experimental studies to determine the most relevant allelic forms for effective progress towards vaccine formulation and testing.

Strep Easy Kit; a bio-enrichment dual ICG-strip test for simultaneous detection of Streptococcus agalactiae serotypes Ia and III in fish samples.

The Strep Easy Kit, a bio-enrichment dual ICG-strip test, is a diagnostic tool designed for the detection of Streptococcus agalactiae, an important pathogenic bacterium in tilapia farming. The kit can simultaneously identify two different serotypes of S. agalactiae, serotype Ia and serotype III. This capability is crucial for the collection of valuable epidemiological data and facilitates strategic planning for effective vaccine development and deployment. The Strep Easy Kit consists of two main steps: pathogen enrichment and pathogen detection. The enrichment step increases the concentration of bacteria so that the bacterial load is raised to the level reliably detectable by the subsequent ICG strip test. This is achieved by incubating the fish samples in a suitable liquid medium under specified temperature and time conditions. The second step involves the use of the dual-ICG strip test. This strip test consists of two monoclonal antibodies and one polyclonal antibody that are specific to S. agalactiae and can distinguish between S. agalactiae serotype Ia and S. agalactiae serotype III. This dual capability enables the ICG strip test to simultaneously detect both serotypes of S. agalactiae in a single test kit. The detection limit of the test kit, which consists of a dual ICG-Strip test combined with an enrichment step, is 100 CFU/mL. The kit can be used to detect S. agalactiae in both live and dead fish samples, making it versatile for various testing scenarios. The test results obtained using the Strep Easy Kit have shown a 94.4% correlation with the standard method (Thai Agricultural Standard; TAS 10453-2010), with 90.2% sensitivity and 100% specificity. Significant advantages of the Strep Easy Kit lie in its simplicity and portability, allowing farmers to perform the test by themselves and on-site. This makes it a practical and accessible tool for the tilapia farming industry.

GIHP: Graph convolutional neural network based interpretable pan-specific HLA-peptide binding affinity prediction.

Accurately predicting the binding affinities between Human Leukocyte Antigen (HLA) molecules and peptides is a crucial step in understanding the adaptive immune response. This knowledge can have important implications for the development of effective vaccines and the design of targeted immunotherapies. Existing sequence-based methods are insufficient to capture the structure information. Besides, the current methods lack model interpretability, which hinder revealing the key binding amino acids between the two molecules. To address these limitations, we proposed an interpretable graph convolutional neural network (GCNN) based prediction method named GIHP. Considering the size differences between HLA and short peptides, GIHP represent HLA structure as amino acid-level graph while represent peptide SMILE string as atom-level graph. For interpretation, we design a novel visual explanation method, gradient weighted activation mapping (Grad-WAM), for identifying key binding residues. GIHP achieved better prediction accuracy than state-of-the-art methods across various datasets. According to current research findings, key HLA-peptide binding residues mutations directly impact immunotherapy efficacy. Therefore, we verified those highlighted key residues to see whether they can significantly distinguish immunotherapy patient groups. We have verified that the identified functional residues can successfully separate patient survival groups across breast, bladder, and pan-cancer datasets. Results demonstrate that GIHP improves the accuracy and interpretation capabilities of HLA-peptide prediction, and the findings of this study can be used to guide personalized cancer immunotherapy treatment. Codes and datasets are publicly accessible at: https://github.com/sdustSu/GIHP.

Reverse vaccinology approach for identification of epitopes from E1 protein as peptide vaccine against HCV: A proof of concept

The development of effective vaccines against Hepatitis C Virus (HCV) remains a global health priority and challenge. In this study, we employed an integrative approach combining computational epitope prediction with experimental validation to identify immunogenic peptides targeting the E1 glycoprotein of HCV. In the present report, computational data from various epitope prediction algorithms such as IEDB and SYFPEITHI, followed by molecular dynamics (MD) simulations and immuno-informatics analysis is presented. Through computational screening, we identified potential epitope candidates, with QVRNSSGLY (P3) and QLFTFSPRH (P7) emerging as promising candidates. MD simulations revealed stable interactions between these epitopes and MHC molecule, further validated by free energy estimations using MMPBSA method. Immuno-informatics analysis supported these findings, showing high binding potential and immunogenicity scores for the selected peptides. Subsequent synthesis and characterization of epitope peptides confirmed their structural integrity and purity required for conducting immune activation assays. Experimental immunological assays carried out in this study involved epitope peptide induced activation of CD8 + and CD4 + T cells from healthy human subjects and HCV- recovered patients. Data from experimental validation revealed significant cytokine release upon exposure to epitope peptides, particularly TNF-a, IL-6, and GM-CSF, indicative of robust immune responses. Notably, peptides P3 and P7 exhibited the most pronounced cytokine induction profiles, underscoring their potential as vaccine candidates. Further investigations addressing the mechanism of action of these epitope peptides under preclinical and clinical settings may help in developing effective vaccine against HCV.

Single-cell sequencing: Current applications in various tuberculosis specimen types.

Tuberculosis (TB) is a chronic disease caused by Mycobacterium tuberculosis (M.tb) and responsible for millions of deaths worldwide each year. It has a complex pathogenesis that primarily affects the lungs but can also impact systemic organs. In recent years, single-cell sequencing technology has been utilized to characterize the composition and proportion of immune cell subpopulations associated with the pathogenesis of TB disease since it has a high resolution that surpasses conventional techniques. This paper reviews the current use of single-cell sequencing technologies in TB research and their application in analysing specimens from various sources of TB, primarily peripheral blood and lung specimens. The focus is on how these technologies can reveal dynamic changes in immune cell subpopulations, genes and proteins during disease progression after M.tb infection. Based on the current findings, single-cell sequencing has significant potential clinical value in the field of TB research. Next, we will focus on the real-world applications of the potential targets identified through single-cell sequencing for diagnostics, therapeutics and the development of effective vaccines.

Six adenoviral vectored African swine fever virus genes protect against fatal disease caused by genotype I challenge.

African swine fever virus causes a lethal hemorrhagic disease in domestic pigs and has killed millions of animals across Europe and Asia since 2007. Development of safe and effective subunit vaccines against African swine fever has been problematic due to the complexity of the virus and a poor understanding of protective immunity. In a previous study, we demonstrated that a complex combination of eight different virus genes delivered using two different viral vector vaccine platforms protected domestic pigs from fatal disease. In this study, we show that three of the eight genes are required for protection and that one viral vector is sufficient, significantly reducing the complexity of the vaccine. Unfortunately, this combination did not protect against the current outbreak strain of African swine fever virus, suggesting that more work to identify immunogenic and protective viral proteins is required to develop a truly effective African swine fever vaccine.

Determining genetic diversity of prevalent G and P genotype of Bovine Rotavirus A from neonatal calves of Gujarat, India.

Neonatal calf diarrhea is a major cause of mortality in newborn calves worldwide, posing a significant challenge in bovine herds. Group A Bovine Rotaviruses (BRVA) are the primary contributors to severe gastroenteritis in calves under two months old. This study examined the prevalence and molecular characterization of BRVA in neonatal calves in Gujarat, India. Sixty-nine diarrheic fecal samples were collected and subjected to various molecular methods of BRVA detection, isolation, and characterization. The latex agglutination test (LAT), electropherotyping (RNA-PAGE), and reverse transcription polymerase chain reaction revealed positivity rates of 39.13%, 20.30%, and 37.70%, respectively. RNA-PAGE identified 11 bands with a 4:2:3:2 migration pattern, indicative of the segmented genome of BRVA. BRVA was successfully isolated from LAT-positive samples, with 26 samples exhibiting clear cytopathic effects upon passage in MA-104 cell lines. Genotyping identified G10 as the predominant G genotype, with P[11] genotypes comprising 76.92% of the isolates. The most common G/P combination was G10P[11], highlighting its zoonotic potential. These findings underscore the importance of molecular detection and genotyping for effective vaccine development. This study provides crucial insights into the prevalent G and P genotypes of BRVA in Gujarat, India, aiding in the development of targeted control measures.

Identification of Two Linear Epitopes on MGF_110-13L Protein of African Swine Fever Virus with Monoclonal Antibodies.

African swine fever caused by African swine fever virus (ASFV) is an acute, highly contagious swine disease with high mortality. To facilitate effective vaccine development and find more serodiagnostic targets, fully exploring the ASFV antigenic proteins is urgently needed. In this study, the MGF_110-13L was identified as an immunodominant antigen among the seven transmembrane proteins. The main outer-membrane domain of MGF_110-13L was expressed and purified. Two monoclonal antibodies (mAbs; 8C3, and 10E4) against MGF_110-13L were generated. The epitopes of two mAbs were preliminary mapped with the peptide fusion proteins after probing with mAbs by enzyme-linked immunosorbent assay (ELISA) and Western blot. And the two target epitopes were fine-mapped using further truncated peptide fusion protein strategy. Finally, the core sequences of mAbs 8C3 and 10E4 were identified as 48WDCQDGICKNKITESRFIDS67, and 122GDHQQLSIKQ131, respectively. The peptides of epitopes were synthesized and probed with ASFV antibody positive pig sera by a dot blot assay, and the results showed that epitope 10E4 was an antigenic epitope. The epitope 10E4 peptide was further evaluated as a potential antigen for detecting ASFV antibodies. To our knowledge, this is the first report of antigenic epitope information on the antigenic MGF_110-13L protein of ASFV.

A Novel Multi-epitope Vaccine Based on Virulence Stimulating Two-component Protein EsrA Against Fish Pathogenic Edwardsiella tarda – An Immunoinformatic Approach

Edwardsiellosis, caused by the gram-negative pathogen Edwardsiella tarda, is a major global aquaculture threat. Despite vaccination being a potential strategy, development of effective vaccines remains a challenge. The study aimed to develop a multi-epitope vaccine (EsrAmev) against virulent EsrA antigen in fish pathogenic E. tarda using an immunoinformatic approach. The study identified antigenic, non-allergenic, and nontoxic epitopes for cytotoxic T lymphocytes, helper T-lymphocytes, and linear Blymphocytes. They were combined to create the EsrAmev construct, with TLR4 agonist added for the immunogenicity. The physiochemical analysis of EsrAmev by ProtParam server showed a molecular weight of 29 kDa, an antigenicity score of 0.5966, and a solubility of 0.823563 with immunogenic, non-allergenic and non-toxic properties. Robetta, Mod, and Galaxy refiner servers refined the EsrAmev 3D structure, achieving 97.4% RAMA-favoured region, stable interactions, binding stability, and structural stiffness with TLR4 receptor. The EsrAmev construct, optimized for high immune responses in Escherichia coli K12 model, was successfully cloned into the pET28a (+) vector in silico for future wet lab applications. These findings suggests that the EsrAmev multi-epitope vaccine might have potential for the prophylaxis of E.tardacausing Fish Edwardsiellosis.

A Systematic Review on Artificial Intelligence (AI) Assisted Drug & Vaccine Development against SARS-CoV-2

Background: Adaptation and application of Artificial Intelligence (AI) technology for the development of drugs against the deadly and continuously mutating Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) virus has been extremely beneficial, cost-effective, and time saving for the scientific community. A systematic review is necessary for complete picturization of the overall AI assistance in developing drugs and vaccines against SARS-CoV-2. Materials and Methods: A systematic analysis and review of the research literature available on the application of AI in the development of drugs and vaccines against SARS-CoV-2 from various online platforms has been performed, and relevant full papers have been selected on certain selection criteria and have been used for this review. Results: Utilization of AI tools has enabled the selection, modification, evaluation, and prediction of the effectiveness of drug formulations against coronavirus disease (COVID-19) in a very rapid and efficient manner. Vaccine development against the deadly SARS-CoV-2 virus has also been aided and benefited immensely by using AI tools and techniques. Discussion: Thousands of studies regarding the development of effective drugs and vaccines against the constantly evolving, mutating, and prevailing SARS-CoV-2 virus have been conducted, and several thousands are still being conducted around the world. Conclusion: AI is a powerful tool, and its application has been highly beneficial in developing effective drugs and vaccines against the deadly SARS-CoV-2 in a cost-effective and time-effective frame. This systematic review briefs the findings and achievements till the date of writing this article in the field of AI-assisted drug and vaccine development against COVID-19.

Dengue virus: pathogenesis and potential small moleculeantivirals.

Dengue, caused by dengue virus (DENV), is now endemic in nearly 100 countries and infection incidence is reported in another 30 countries. Yearly an estimated 400 million cases and 2200 deaths are reported. Effective vaccinesagainst DENV are limited and there has been significant focus on the development of effective antiviral against the disease.The World Health Organization has initiated research programs to prioritize the development and optimization ofantiviral agents against several viruses including Flaviviridae. A significant effort has been taken by the researchers to develop effective antivirals against DENV. Several potential small-molecule inhibitors like efavirenz, tipranavir and dasabuvir have been tested against envelope and non-structural proteins of DENV, and are in clinical trials around the world. We recently developed one small molecule, namely 7D, targeting the host PF4-CXCR3 axis. 7D inhibited all 4 serotypes of DENV in vitro and specifically DENV2 infection in two different mice models. Although the development of dengue vaccines remains a high priority, antibody cross reactivity among the serotypes and resulting antibody-dependent enhancement (ADE) of infection are major concerns that have limited the development of effective vaccine against DENV. Therefore, there has been a significant emphasis on the development of antiviral drugs against dengue.This review article describes the rescue effects of some of the small molecule inhibitors to viral/host factors associated with DENV pathogenesis.

Generation of single-round infectious rotavirus with a mutation in the intermediate capsid protein VP6.

Rotavirus causes severe diarrhea in infants. Although live attenuated rotavirus vaccines are available, vaccine-derived infections have been reported, which warrants development of next-generation rotavirus vaccines. A single-round infectious virus is a promising vaccine platform; however, this platform has not been studied extensively in the context of rotavirus. Here, we aimed to develop a single-round infectious rotavirus by impairing the function of the viral intermediate capsid protein VP6. Recombinant rotaviruses harboring mutations in VP6 were rescued using a reverse genetics system. Mutations were targeted at VP6 residues involved in virion assembly. Although the VP6-mutated rotavirus expressed viral proteins, it did not produce progeny virions in wild-type cells; however, the virus did produce progeny virions in VP6-expressing cells. This indicates that the VP6-mutated rotavirus is a single-round infectious rotavirus. Insertion of a foreign gene, and replacement of the VP7 gene segment with that of human rotavirus clinical isolates, was successful. No infectious virions were detected in mice infected with the single-round infectious rotavirus. Immunizing mice with the single-round infectious rotavirus induced neutralizing antibody titers as high as those induced by wild-type rotavirus. Taken together, the data suggest that this single-round infectious rotavirus has potential as a safe and effective rotavirus vaccine. This system is also applicable for generation of safe and orally administrable viral vectors.IMPORTANCERotavirus, a leading cause of acute gastroenteritis in infants, causes an annual estimated 128,500 infant deaths worldwide. Although live attenuated rotavirus vaccines are available, they are replicable and may cause vaccine-derived infections. Thus, development of safe and effective rotavirus vaccine is important. In this study, we report the development of a single-round infectious rotavirus that can replicate only in cells expressing viral VP6 protein. We demonstrated that (1) the single-round infectious rotavirus did not replicate in wild-type cells or in mice; (2) insertion of foreign genes and replacement of the outer capsid gene were possible; and (3) it was as immunogenic as the wild-type virus. Thus, the mutated virus shows promise as a next-generation rotavirus vaccine. The system is also applicable to orally administrable viral vectors, facilitating development of vaccines against other enteric pathogens.

Characteristics of Norovirus capsid protein-specific CD8+ T-Cell responses in previously infected individuals

ABSTRACT Norovirus (NV) infection causes acute gastroenteritis in children and adults. Upon infection with NV, specific CD8+ T cells, which play an important role in anti-infective immunity, are activated in the host. Owing to the NV’s wide genotypic variability, it is challenging to develop vaccines with cross-protective abilities against infection. To aid effective vaccine development, we examined specific CD8+ T-cell responses towards viral-structural protein (VP) epitopes, which enable binding to host susceptibility receptors. We isolated peripheral blood mononuclear cells from 196 participants to screen and identify predominant core peptides towards NV main and small envelope proteins using ex vivo and in vitro intracellular cytokine staining assays. Human leukocyte antigen (HLA) restriction characteristics were detected using next-generation sequencing. Three conservative immunodominant VP-derived CD8+ T-cell epitopes, VP294–102 (TDAARGAIN), VP2153–161 (RGPSNKSSN), and VP1141–148 (FPHIIVDV), were identified and restrictively presented by HLA-Cw * 0102, HLA-Cw * 0702, and HLA-A *1101 alleles, separately. Our findings provide useful insights into the development of future vaccines and treatments for NV infection.

Global elimination of HBV: Is it really achievable?

Hepatitis B virus (HBV) infection is a major cause of premature death worldwide. In 2016, the World Health Organization (WHO) called for HBV elimination and set up very ambitious elimination targets. The development of effective vaccines, accurate diagnostic tools and safe antiviral drugs make HBV elimination a realistic goal. However, the most constrained-resource regions, which bear the highest burden of HBV, are facing major challenges in implementing strategies to reduce HBV incidence and mortality. Developing simplified approaches adapted to resource-limited settings and scaling up interventions for the prevention and control of HBV globally are urgently needed. Whether HBV elimination will be achieved in an equitable manner and in a reasonable timeframe remains highly uncertain.

Engineering W/O/W pickering emulsion for the enhanced antigen delivery and immune responses

Tumor immunotherapy, particularly cancer vaccines, holds promise for combating cancer by harnessing tailored immune responses against malignant cells. However, conventional approaches face challenges in efficiently delivering antigens for optimal immune activation. Emulsion adjuvants, like MF59, enhance cellular uptake but struggle to induce robust CD8+ T cell responses. Here, we introduce a novel strategy employing a water-in-oil-in-water (W/O/W) multiple Pickering emulsion (mPE) for antigen delivery. The mPE, utilizing biocompatible, pH-sensitive particles, encapsulates antigens within the inner water phase, ensuring enhanced intracellular processing and dictating the intracellular fate of antigens for improved cross-presentation. In vitro and in vivo studies demonstrated that mPEs induced robust dendritic cells activation and antigen cross-presentation, leading to a significantly enhanced immune response. Notably, calcium phosphate-stabilized mPE (CaP-mPE) illustrated the more robust IFN-γ+ T cell responses. In comparison with traditional surfactant-stabilized multiple emulsions, CaP-mPE significantly inhibit tumor growth and effectively prolong the survival of tumor-bearing mice. This innovative approach offers a promising avenue for the development of effective cancer vaccines with potent cellular immune responses.

A novel subunit vaccine based on Fiber1/2 knob domain provides full protection against fowl adenovirus serotype 4 and induces stronger immune responses than a Fiber2 subunit vaccine

Outbreaks of hepatitis-hydropericardium syndrome (HHS) caused by fowl adenovirus serotype 4 (FAdV-4) have resulted in huge economic losses to the poultry industry in China since 2015. However, commercially available vaccines against the FAdV-4 infection remain scarce. In our study, subunit vaccine candidates derived from the bacterially expressed recombinant Fiber1 knob domain and Fiber2 knob domain fusion protein (termed as Fiber1/2 knob subunit vaccine) and Fiber2 protein (termed as Fiber2 subunit vaccine) of the FAdV-4 SDSX strain were developed. Immunogenicity evaluation showed that the Fiber1/2 knob subunit vaccine induced the production of antibodies at 7 d postvaccination (dpv), earlier than the Fiber2 subunit vaccine. Moreover, the neutralizing antibody level of the Fiber1/2 subunit vaccine group was higher than the Fiber2 subunit vaccine group, showing significant differences at 14, 21, and 28 dpv. Immune protection test results revealed that both Fiber1/2 knob subunit and Fiber2 subunit vaccines could protect chickens from death against FAdV-4 challenge, although the weight of chickens in the Fiber1/2 knob subunit vaccine group decreased less. Furthermore, analysis of plasma Glutamic oxaloacetic transaminase (AST) and blood glutamic pyruvic transaminase (ALT) levels suggested that the Fiber1/2 subunit vaccine can significantly inhibit liver damage caused by FAdV-4 infection and is more effective in blocking the pathogenicity of FAdV-4 in target organs. In addition, the Fiber1/2 knob subunit vaccine further reduced the viral load in different tissues and virus shedding in chickens than the Fiber2 subunit vaccine. Overall, the Fiber1/2 knob subunit vaccine was more effective than the Fiber2 subunit vaccine. These findings lay the foundation for the development of more effective FAdV-4 subunit vaccines.

Identification and characterization of a nonbiological small-molecular mimic of a Zika virus conformational neutralizing epitope

Antigenic similarities between Zika virus (ZIKV) and other flaviviruses pose challenges to the development of virus-specific diagnostic tools and effective vaccines. Starting with a DNA-encoded one-bead-one-compound combinatorial library of 508,032 synthetic, non-natural oligomers, we selected and characterized small molecules that mimic ZIKV epitopes. High-throughput fluorescence-activated cell sorter-based bead screening was used to select molecules that bound IgG from ZIKV-immune but not from dengue-immune sera. Deep sequencing of the DNA from the "Zika-only" beads identified 40 candidate molecular structures. A lead candidate small molecule "CZV1-1" was selected that correctly identifies serum specimens from Zika-experienced patients with good sensitivity and specificity (85.3% and 98.4%, respectively). Binding competition studies of purified anti-CZV1-1 IgG against known ZIKV-specific monoclonal antibodies (mAbs) showed that CZV1-1 mimics a nonlinear, neutralizing conformational epitope in the domain III of the ZIKV envelope. Purified anti-CZV1-1 IgG neutralized infection of ZIKV in cell cultures with potencies comparable to highly specific ZIKV-neutralizing mAbs. This study demonstrates an innovative approach for identification of synthetic non-natural molecular mimics of conformational virus epitopes. Such molecular mimics may have value in the development of accurate diagnostic assays for Zika, as well as for other viruses.