Determination Of Protein Research Articles

Designing a multi-epitope influenza vaccine: an immunoinformatics approach

Influenza continues to be one of the top public health problems since it creates annual epidemics and can start a worldwide pandemic. The virus’s rapid evolution allows the virus to evade the host defense, and then seasonal vaccines need to be reformulated nearly annually. However, it takes almost half a year for the influenza vaccine to become accessible. This delay is especially concerning in the event of a pandemic breakout. By producing the vaccine through reverse vaccinology and phage display vaccines, this time can be reduced. In this study, epitopes of B lymphocytes, cytotoxic T lymphocytes, and helper T lymphocytes of HA, NA, NP, and M2 proteins from two strains of Influenza A were anticipated. We found two proper epitopes (ASFIYNGRL and LHLILWITDRLFFKC) in Influenza virus proteins for CTL and HTL cells, respectively. Optimal epitopes and linkers in silico were cloned into the N-terminal end of M13 protein III (pIII) to create a multi-epitope-pIII construct, i.e., phage display vaccine. Also, prediction of tertiary structure, molecular docking, molecular dynamics simulation, and immune simulation were performed and showed that the designed multi-epitope vaccine can bind to the receptors and stimulate the immune system response.

Thermal Denaturation of Fresh Frozen Tissue Enhances Mass Spectrometry Detection of Peptides.

Thermal denaturation (TD), known as antigen retrieval, heats tissue samples in a buffered solution to expose protein epitopes. Thermal denaturation of formalin-fixed paraffin-embedded samples enhances on-tissue tryptic digestion, increasing peptide detection using matrix-assisted laser desorption ionization imaging mass spectrometry (MALDI IMS). We investigated the tissue-dependent effects of TD on peptide MALDI IMS and liquid chromatography-tandem mass spectrometry signal in unfixed, frozen human colon, ovary, and pancreas tissue. In a triplicate experiment using time-of-flight, orbitrap, and Fourier-transform ion cyclotron resonance mass spectrometry platforms, we found that TD had a tissue-dependent effect on peptide signal, resulting in a (22.5%) improvement in peptide detection from the colon, a (73.3%) improvement in ovary tissue, and a (96.6%) improvement in pancreas tissue. Biochemical analysis of identified peptides shows that TD facilitates identification of hydrophobic peptides.

Multiplexed chromatin immunoprecipitation sequencing for quantitative study of histone modifications and chromatin factors.

ChIP-seq is a widely used technique for studying histone post-translational modifications and DNA-binding proteins. DNA fragments associated with a specific protein or histone modification epitope are captured by using antibodies, sequenced and mapped to a reference genome. Albeit versatile and popular, performing many parallel ChIP-seq experiments to compare different conditions, replicates and epitopes is laborious, is prone to experimental variation and does not allow quantitative comparisons unless adequate spike-in chromatin is included. We present a detailed protocol for performing and analyzing a multiplexed quantitative chromatin immunoprecipitation-sequencing experiment (MINUTE-ChIP), in which multiple samples are profiled against multiple epitopes in a single workflow. Multiplexing not only dramatically increases the throughput of ChIP-seq experiments (e.g., profiling 12 samples against multiple histone modifications or DNA-binding proteins in a single experiment), but also enables accurate quantitative comparisons. The protocol consists of four parts: sample preparation (i.e., lysis, chromatin fragmentation and barcoding of native or formaldehyde-fixed material), pooling and splitting of the barcoded chromatin into parallel immunoprecipitation reactions, preparation of next-generation sequencing libraries from input and immunoprecipitated DNA and data analysis using our dedicated analysis pipeline. This pipeline autonomously generates quantitatively scaled ChIP-seq tracks for downstream analysis and visualization, alongside necessary quality control indicators. The entire workflow requires basic knowledge in molecular biology and bioinformatics and can be completed in 1 week. MINUTE-ChIP empowers biologists to perform every ChIP-seq experiment with an appropriate number of replicates and control conditions, delivering more statistically robust, exquisitely quantitative and biologically meaningful results.

A public, cross-reactive glycoprotein epitope confounds Ebola virus serology.

Ebola disease (EBOD) in humans is a severe disease caused by at least four related viruses in the genus Orthoebolavirus, most often by the eponymous Ebola virus. Due to human-to-human transmission and incomplete success in treating cases despite promising therapeutic development, EBOD is a high priority in public health research. Yet despite almost 50 years since EBOD was first described, the sources of these viruses remain undefined and much remains to be understood about the disease epidemiology and virus emergence and spread. One important approach to improve our understanding is detection of antibodies that can reveal past human infections. However, serosurveys routinely describe seroprevalences that imply infection rates much higher than those clinically observed. Proposed hypotheses to explain this difference include existence of common but less pathogenic strains or relatives of these viruses, misidentification of EBOD as something else, and a higher proportion of subclinical infections than currently appreciated. The work presented here maps B-cell epitopes in the spike protein of Ebola virus and describes a single epitope that is cross-reactive with an antigen seemingly unrelated to orthoebolaviruses. Antibodies against this epitope appear to explain most of the unexpected reactivity towards the spike, arguing against common but unidentified infections in the population. Importantly, antibodies of cross-reactive donors from within and outside the known EBOD geographic range bound the same epitope. In light of this finding, it is plausible that epitope mapping enables broadly applicable specificity improvements in the field of serology.

Differential patterns of antibody response against SARS-CoV-2 nucleocapsid epitopes detected in sera from patients in acute phase of COVID-19, convalescents and pre-pandemic individuals.

The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) have already infected more than 0.7 billion people and caused over 7 million deaths worldwide. At the same time, our knowledge about this virus is still incipient. In some cases, there is a pre-pandemic immunity, however its source is unknown. The analysis of patients' humoral responses might shed a light on this puzzle. In this paper, we evaluated the antibody recognition of nucleocapsid protein, one of the structural proteins of SARS-CoV-2. For this purpose, we used pre-pandemic, acute COVID-19 and convalescent patients' sera to identify and map nucleocapsid protein epitopes. We identified a common epitope KKSAAEASKKPRQKRTATKA recognized by sera antibodies from all three groups. Some motifs of this sequence are widespread among various coronaviruses, plant or human proteins indicating that there might be more sources of nucleocapsid-reactive antibodies than previous infection with seasonal coronavirus. The two sequences MSDNGPQNQRNAPRITFGGP and KADETQALPQRQKKQQTVTL were detected as specific for sera from patients in acute phase of infection and convalescents making them suitable for future development of vaccine against SARS-CoV-2. Knowledge of the humoral response to SARS-CoV-2 infection is essential for the design of appropriate diagnostic tools and vaccine antigens.

HLA-C Peptide Repertoires as Predictors of Clinical Response during Early SARS-CoV-2 Infection.

The human leukocyte antigen (HLA) system plays a pivotal role in the immune response to viral infections, mediating the presentation of viral peptides to T cells and influencing both the strength and specificity of the host immune response. Variations in HLA genotypes across individuals lead to differences in susceptibility to viral infection and severity of illness. This study uses observations from the early phase of the COVID-19 pandemic to explore how specific HLA class I molecules affect clinical responses to SARS-CoV-2 infection. By analyzing paired high-resolution HLA types and viral genomic sequences from 60 patients, we assess the relationship between predicted HLA class I peptide binding repertoires and infection severity as measured by the sequential organ failure assessment score. This approach leverages functional convergence across HLA-C alleles to identify relationships that may otherwise be inaccessible due to allelic diversity and limitations in sample size. Surprisingly, our findings show that severely symptomatic infection in this cohort is associated with disproportionately abundant binding of SARS-CoV-2 structural and non-structural protein epitopes by patient HLA-C molecules. In addition, the extent of overlap between a given patient's predicted HLA-C and HLA-A peptide binding repertoires correlates with worse prognoses in this cohort. The findings highlight immunologic mechanisms linking HLA-C molecules with the human response to viral pathogens that warrant further investigation.

Potential contribution of gut microbiota in the development of autoantibodies in T1D children carrying HLA-DRB1/DQB1 risk alleles: an experimental and in silico analysis.

This study aimed to investigate the prevalence of insulin autoantibody (IAA), glutamic acid decarboxylase antibody (GADA), and insulinoma-associated antigen-2 antibody (IA-2A) in type 1 diabetes (T1D) children based on the presence of predisposing HLA-II alleles. Additionally, to assess the sequence homology between autoantigens of islet cells and selected proteins derived from gut bacteria in terms of their binding capacities to HLA risk alleles, HLA-DRB1/DQB1 alleles were determined by PCR-SSOP in 111 T1D children (probands) along with 222 parents and 133 siblings. Autoantibodies were measured by ELISA, and in silico analysis was run as follows: protein extraction, homology and epitope prediction, peptide alignment, and HLA-peptide docking. Higher significant frequencies of DRB1*03:01, DQB1*02:01, and DQB1*03:02 alleles and DRB1*03:01 ~ DQB1*02:01 haplotype and lower frequencies of DRB1*11:01, DRB1*14:01, and DQB1*03:01 alleles were found in probands compared to parents and siblings. DRB1*11:01 ~ DQB1*03:01, DRB1*14:01 ~ DQB1*05:03, and DRB1*15:01-DQB1*06:02 haplotypes were significantly less frequent in the probands compared to parents. Out of 111 probands, 21 were seronegative, 90 tested positive for one autoantibody, and 15 showed the concurrent presence of three autoantibodies. Logistic regression analysis revealed that DRB1*04 ~ DQB1*03:02 haplotype was associated with the induction of GADA and IA-2A, while DRB1*11:01 ~ DQB1*03:01 was associated with seronegativity. Epitopes derived from GAD and gut bacteria showed strong binding capacities to HLA risk alleles. Due to the sequence similarities between gut bacteria-derived proteins and islet cell autoantigens and their potential for binding to HLA risk alleles, dysbiosis of gut microbiota can be considered another risk factor for the development of T1D, especially in genetically susceptible individuals.

Salmonella pathogenesis-based In-silico design and immunoinformatic analysis of multi-epitope vaccine constructs in broiler veterinary medicine

Salmonellosis, a zoonotic gastrointestinal disease, presents a significant global health burden with a high incidence rate. Transmission primarily occurs through the consumption of contaminated poultry products, although water and contact with asymptomatic animals are also vectors. The disease’s pervasiveness has prompted international health organizations to advocate for robust prevention and control strategies. This study focuses on the in-silico design of a multi-epitope vaccine targeting Salmonella enterica serovar Typhimurium’s fimH protein, a fimbriae component crucial for bacterial adhesion and pathogenicity. The vaccine construct was developed by identifying and synthesizing non-allergenic, antigenic, and non-toxic epitopes for both Cytotoxic T Lymphocytes and Helper T Lymphocytes. Adjuvants were incorporated to enhance immunogenicity, and the vaccine’s structure was modeled using advanced bioinformatics tools. The proposed vaccine demonstrated promising antigenicity and immunogenicity profiles, with a favorable physical-chemical property analysis. The vaccine’s structures, designed by computational analysis, suggests high likelihood to native protein configurations. Antigenicity and allergenicity assessments validate the vaccine’s immunogenic potential and hypoallergenic nature. Physicochemical evaluations indicate favorable stability and solubility profiles, essential for vaccine efficacy. This comprehensive approach to vaccine design expressed in Chlorella vulgaris holds promises for effective salmonellosis control. The multi-epitope vaccine, designed through meticulous in-silico methods, emerges as a promising candidate for controlling salmonellosis. Its strategic construction based on the fimH protein epitopes offers a targeted approach to elicit a robust immune response, potentially curbing the spread of this disease in poultry.

SUB-immunogold-SEM reveals nanoscale distribution of submembranous epitopes

Electron microscopy paired with immunogold labeling is the most precise tool for protein localization. However, these methods are either cumbersome, resulting in small sample numbers and restricted quantification, or limited to identifying protein epitopes external to the membrane. Here, we introduce SUB-immunogold-SEM, a scanning electron microscopy technique that detects intracellular protein epitopes proximal to the membrane. We identify four critical sample preparation factors contributing to the method’s sensitivity. We validate its efficacy through precise localization and high-powered quantification of cytoskeletal and transmembrane protein distribution. We evaluate the capabilities of SUB-immunogold-SEM on cells with highly differentiated apical surfaces: (i) auditory hair cells, revealing the presence of nanoscale MYO15A-L rings at the tip of stereocilia; and (ii) respiratory multiciliate cells, mapping the distribution of the SARS-CoV-2 receptor ACE2 along the motile cilia. SUB-immunogold-SEM extends the application of SEM-based nanoscale protein localization to the detection of intracellular epitopes on the exposed surfaces of any cell.

Identification and evaluation of multi-antigenic epitopes of immunodominant protein from the selected Crimean–Congo hemorrhagic fever virus genome towards the development of diagnostic and vaccine candidates by reverse vaccinology approach

Identification and evaluation of multi-antigenic epitopes of immunodominant protein from the selected Crimean–Congo hemorrhagic fever virus genome towards the development of diagnostic and vaccine candidates by reverse vaccinology approach

Improving Specificity for Ovarian Cancer Screening Using a Novel Extracellular Vesicle–Based Blood Test: Performance in a Training and Verification Cohort

The low incidence of ovarian cancer (OC) dictates that any screening strategy needs to be both highly sensitive and highly specific. This study explored the utility of detecting multiple colocalized proteins or glycosylation epitopes on single tumor-associated extracellular vesicles (EVs) from blood. The novel OC Test employs immunoaffinity capture of tumor-associated EVs followed by proximity-ligation qPCR to detect combinations of up to three biomarkers to maximize specificity and measures multiple combinations to maximize sensitivity. A high-grade serous carcinoma (HGSC) case-control training set of EDTA plasma samples from 397 women was used to lock down the test design, the data interpretation algorithm, and the cut-off between cancer and non-cancer. Performance was verified and compared to CA125 in an independent blinded case-control set of serum samples from 390 women (132 controls, 66 HGSC, 83 non-HGSC OC, 109 benign). In the verification study, the OC Test showed a specificity of 97.0% (128/132; 95% CI: 92.4%-99.6%), a HGSC sensitivity of 97.0% (64/66; 95% CI: 87.8%-99.2%), and an AUC of 0.97 (95% CI 0.93-0.99) and also detected 73.5% (61/83; 95% CI: 62.7%-82.6%) of the non-HGSC OC cases. This test exhibited fewer false positives in subjects with benign ovarian tumors, non-ovarian cancers, and inflammatory conditions when compared to CA125. The combined sensitivity and specificity of this new test suggests it may have potential in OC screening.

Molecular similarities between the genes for Trypanosoma cruzi microtubule-associated proteins, mammalian interferons, and TRIMs

Initial studies using bioinformatics analysis revealed DNA sequence similarities between Trypanosoma cruzi GenBank® M21331, coding for Antigen 36 (Ag 36), and tripartite motif (TRIM) genes. TRIM40 showed 9.7% identity to GenBank M21331, and four additional TRIM genes had identities greater than 5.0%. TRIM37 showed a continuous stretch of identity of 12 nucleotides, that is, at least 25% longer than any of the other TRIMs. When we extended our analysis on the relationships of GenBank M21331 to further innate immune genes, using the Needleman-Wunsch (NW) algorithm for alignment, identities to human IFN-α, IFN-β, and IFN-γ genes of 13.6%, 12.6%, and 17.9%, respectively, were found. To determine the minimum number of genes coding for proteins closely related to Ag 36, a BLAST-p search was conducted with it versus the T. cruzi genome. The BLAST-p search revealed that T. cruzi GenBank M21331 had 14 gene sequences homologous to microtubule-associated protein (MAP) genes with 100% amino acid sequence identity. To verify the similarities in non-human genes, a study comparing TRIM21 region sequences among mammalian species to the comparable human TRIM21 region showed that related sequences were also present in 11 mammalian species. The MAP genes homologous to Ag 36 form a family of at least 14 genes which mimic human immune genes in the IFN and TRIM families. This mimicry is of gene sequences and not their protein products or epitopes. These results appear to be the first description of molecular mimicry of immune genes in humans by a protozoan parasite.

B-cell epitope prediction of MPB83 protein as a candidate for serodiagnostic antigen of bovine tuberculosis in human: In silico study

Background: Bovine tuberculosis (bTB) can be transmitted to humans by inhalation or consumption of incomplete pasteurized milk and dairy products derived from infected cows. Most cases of Mycobacterium tuberculosis (M. bovis) infection are resistant to tuberculosis (TB) drugs. The risk of death during treatment for bTB has been reported to be 2.55 times higher than for TB. However, the quality of diagnostic methods for bTB remains relatively low.Objective: We aim to evaluate the potential of the B-cell epitope of the MPB83 protein as a candidate bTB serodiagnostic antigen using an in silico approach. Methods: This study was a computer-based descriptive study using secondary data from the National Center for Biotechnology Information (NCBI) protein database. The MPB83 protein sequence was obtained from M. tuberculosis variant bovis AF2122/97 from the United Kingdom. We described the characteristics of the linear epitope of the M. bovis B-cell protein MPB83 by measuring antigenicity, molecular weight, instability index, and Grand Average of Hydropathy (GRAVY) score. The tools used in this study were IBIVU PRALINE, VaxiJen v2.0, IEDB, ExPASy ProtParam, Cluspro, and the PyMOL application.Results: We found an epitope that could be used for bTB serodiagnostic antigen with low conservation, the 106KLNPDVNLVDTLN118 epitope. It has the molecular weight, instability index, and GRAVY score of 1638.76 Da, -28.44, and -0.300, respectively. Epitopes with the best criteria were simulated by docking to human major histocompatibility complex (MHC) class II. Docking results showed that the lowest binding energy was -644.8 kcal/mol. Further analysis using the PyMOL application obtained 14 hydrogen bonds with bond distances ranging from 1.7 Å to 2.2 Å, all of which showed strong hydrogen bonds.Conclusion: The B-cell epitope of MPB83 protein sequence 106KLNPDVNLVDTLN118 has a potential serodiagnostic antigen candidate for human bTB.

Advancement in the Antigenic Epitopes and Vaccine Adjuvants of African Swine Fever Virus.

African swine fever virus (ASFV), a highly virulent double-stranded DNA virus, poses a significant threat to global pig farming, with mortality rates in domestic pigs reaching up to 100%. Originating in Kenya in 1921, ASFV has since proliferated to Western Europe, Latin America, Eastern Europe, and most recently China in 2018, resulting in substantial global agricultural losses. Antigenic epitopes, recognized by the immune system's T cells and B cells, are pivotal in antiviral immune responses. The identification and characterization of these antigenic epitopes can offer invaluable insights into the immune response against ASFV and aid in the development of innovative immunotherapeutic strategies. Vaccine adjuvants, substances that amplify the body's specific immune response to antigens, also play a crucial role. This review provides an overview of the progress in studying T/B-cell epitopes in ASFV proteins and ASFV vaccine adjuvants, highlighting their role in the immune response and potential use in new vaccine development.

Phylodynamic and Epistatic Analysis of Coxsackievirus A24 and Its Variant.

Coxsackievirus A24 (CV-A24) is a human enterovirus that causes acute flaccid paralysis. However, a Coxsackievirus A24 variant (CV-A24v) is the most common cause of eye infections. The causes of these variable pathogenicity and tissue tropism remain unclear. To elucidate the phylodynamics of CV-A24 and CV-A24v, we analyzed a dataset of 66 strains using Bayesian phylodynamic approach, along with detailed sequence variation and epistatic analyses. Six CV-A24 strains available in GenBank and 60 CV-A24v strains, including 11 Taiwanese strains, were included in this study. The results revealed striking differences between CV-A24 and CV-A24v exhibiting long terminal branches in the phylogenetic tree, respectively. CV-A24v presented distinct ladder-like clustering, indicating immune escape mechanisms. Notably, 10 genetic recombination events in the 3D regions were identified. Furthermore, 11 missense mutation signatures were detected to differentiate CV-A24 and CV-A24v; among these mutations, the F810Y substitution may significantly affect the secondary structure of the GH loop of VP1 and subsequently affect the epitopes of the capsid proteins. In conclusion, this study provides critical insights into the evolutionary dynamics and epidemiological characteristics of CV-A24 and CV-A24v, and highlights the differences in viral evolution and tissue tropism.

Molecular mimicry in multisystem inflammatory syndrome in children

Multisystem inflammatory syndrome in children (MIS-C) is a severe, post-infectious sequela of SARS-CoV-2 infection1,2, yet the pathophysiological mechanism connecting the infection to the broad inflammatory syndrome remains unknown. Here we leveraged a large set of samples from patients with MIS-C to identify a distinct set of host proteins targeted by patient autoantibodies including a particular autoreactive epitope within SNX8, a protein involved in regulating an antiviral pathway associated with MIS-C pathogenesis. In parallel, we also probed antibody responses from patients with MIS-C to the complete SARS-CoV-2 proteome and found enriched reactivity against a distinct domain of the SARS-CoV-2 nucleocapsid protein. The immunogenic regions of the viral nucleocapsid and host SNX8 proteins bear remarkable sequence similarity. Consequently, we found that many children with anti-SNX8 autoantibodies also have cross-reactive T cells engaging both the SNX8 and the SARS-CoV-2 nucleocapsid protein epitopes. Together, these findings suggest that patients with MIS-C develop a characteristic immune response to the SARS-CoV-2 nucleocapsid protein that is associated with cross-reactivity to the self-protein SNX8, demonstrating a mechanistic link between the infection and the inflammatory syndrome, with implications for better understanding a range of post-infectious autoinflammatory diseases.

Predicting immune response targets in orthoflaviviruses through sequence homology and computational analysis.

Flaviviruses cause severe encephalitic or hemorrhagic diseases in humans. Its members, Kyasanur forest disease virus (KFDV) and Alkhumra hemorrhagic fever virus (ALKV), cause hemorrhagic fever and are prevalent in India and Saudi Arabia, respectively, while the tick-borne encephalitis virus (TBEV) causes a dangerous encephalitic infection in Europe and Asia. However, little information is available about the targets of immune responses for these deadly viruses. Here, we predict potential antigenic peptide epitopes of viral envelope protein for inducing a cell-mediated and humoral immune response. Using the Immune Epitope Database and Analysis Resource (IEDB-AR), we identified 13 MHC-I and two MHC-II dominant conserved epitopes in KFDV and ALKV and six MHC-I and three MHC-II epitopes in TBEV envelope proteins. Parallelly, we also predicted B-cell linear and discontinuous envelope protein epitopes for these viruses. Interestingly, the epitopes are conserved in all three viral envelope proteins. Further, the discontinuous epitopes are structurally compared with the available DENV, ZIKV, WNV, TBEV, and LIV envelope protein antibody structures. Overall structural comparison analyses highlight (i) lateral ridge epitope in the ED-III domain of E protein, and (ii) envelope dimer epitope (EDE) could be targeted for developing potent vaccine candidates as well as therapeutic antibody production. Moreover, existing structural and biochemical functions of the same epitopes in homologous viruses are predicted to have a reduced antibody-dependent enhancement (ADE) effect on flaviviral infection.

Signal amplification by cyclic extension enables high-sensitivity single-cell mass cytometry.

Mass cytometry uses metal-isotope-tagged antibodies to label targets of interest, which enables simultaneous measurements of ~50 proteins or protein modifications in millions of single cells, but its sensitivity is limited. Here, we present a signal amplification technology, termed Amplification by Cyclic Extension (ACE), implementing thermal-cycling-based DNA in situ concatenation in combination with 3-cyanovinylcarbazole phosphoramidite-based DNA crosslinking to enable signal amplification simultaneously on >30 protein epitopes. We demonstrate the utility of ACE in low-abundance protein quantification with suspension mass cytometry to characterize molecular reprogramming during the epithelial-to-mesenchymal transition as well as the mesenchymal-to-epithelial transition. We show the capability of ACE to quantify the dynamics of signaling network responses in human T lymphocytes. We further present the application of ACE in imaging mass cytometry-based multiparametric tissue imaging to identify tissue compartments and profile spatial aspects related to pathological states in polycystic kidney tissues.

Enhanced Assessment of Cross-Reactive Antigenic Determinants within the Spike Protein.

Despite successful vaccination efforts, the emergence of new SARS-CoV-2 variants poses ongoing challenges to control COVID-19. Understanding humoral responses regarding SARS-CoV-2 infections and their impact is crucial for developing future vaccines that are effective worldwide. Here, we identified 41 immunodominant linear B-cell epitopes in its spike glycoprotein with an SPOT synthesis peptide array probed with a pool of serum from hospitalized COVID-19 patients. The bioinformatics showed a restricted set of epitopes unique to SARS-CoV-2 compared to other coronavirus family members. Potential crosstalk was also detected with Dengue virus (DENV), which was confirmed by screening individuals infected with DENV before the COVID-19 pandemic in a commercial ELISA for anti-SARS-CoV-2 antibodies. A high-resolution evaluation of antibody reactivity against peptides representing epitopes in the spike protein identified ten sequences in the NTD, RBD, and S2 domains. Functionally, antibody-dependent enhancement (ADE) in SARS-CoV-2 infections of monocytes was observed in vitro with pre-pandemic Dengue-positive sera. A significant increase in viral load was measured compared to that of the controls, with no detectable neutralization or considerable cell death, suggesting its role in viral entry. Cross-reactivity against peptides from spike proteins was observed for the pre-pandemic sera. This study highlights the importance of identifying specific epitopes generated during the humoral response to a pathogenic infection to understand the potential interplay of previous and future infections on diseases and their impact on vaccinations and immunodiagnostics.

Comprehensive Analysis of Conserved B-Cell Epitopes in DENV NS1 Protein for Enhanced Rapid Diagnostic Tests Development in Indonesia

Dengue virus (DENV) constitutes a formidable public health threat within tropical and subtropical regions. The escalation of cases in Southeast Asia, notably Indonesia, is a matter of considerable concern. Timely identification of DENV infection remains imperative for efficient disease management. The non-structural protein-1 (NS1), distinguished by its heightened immunogenicity and early detectability during infection, stands as a pivotal target for diagnostic modalities. The current investigation delves into the exploration of B-cell epitopes within Indonesian DENV NS1 protein isolates, with the overarching objective of enhancing the sensitivity and specificity of early detection systems, such as the Rapid Diagnostic Test (RDT). This study has successfully delineated five B-cell epitopes that exhibit conservation across DENV serotypes within Indonesian isolates (accessions number: QBB90021.1, QBE90252.1, QBB90023.1, and UDW38833.1). These epitopes were discerned through comprehensive screening leveraging the IEDB platforms. Noteworthy variations in antigenicity, allergenicity, and toxicity profiles were observed among these identified epitopes. Molecular docking analysis substantiated a robust binding affinity between the predicted epitope and the B-cell receptor. The ensuing in silico protein-peptide docking analyses offer valuable insights into potential B-cell epitopes on the Indonesian DENV NS1 protein. The identified epitopes, particularly the SQHNYRPGY epitope characterized by its antigenic potency and non-allergenic attributes, hold promise for advancing the development of sensitive and specific RDTs tailored for DENV detection in the Indonesian context. However, it is imperative to underscore the requisite for subsequent experimental validation to affirm the efficacy of these epitopes in diagnostic applications.