Antigenic Structure Research Articles

Interaction of Specific Monoclonal Antibodies with Leukocyte Antigens in Camels

The dromedary camel as a livestock species significantly impacts the economy of arid and semi-arid regions worldwide. The identification of cross-reactive antibodies against pivotal immune cell markers acts as a valuable method to investigate the immune system of camels. The aim of the present study was to identify new monoclonal antibodies that react with camel leukocyte subsets using flow cytometry and multicolor immunofluorescence. The expression patterns of the tested antibodies indicated cross-reactivity of the anti-bovine CD9 monoclonal antibody clones LT86A and Hl9a with different binding potential. Although all leukocyte subpopulations stained positively with the CD9 antibodies, monocytes showed the highest CD9 abundance, compared to lymphocytes and granulocytes. No cross-reactivity was identified for the tested monoclonal antibodies against equine CD8a (clone: ETC142BA1), mouse CD3 (clone: CD3-12), human CD3 (clone: T3/2/16A9), human CD206 (clone: MMR), and bovine granulocytes (clone: CH138A). The present study revealed that only camel monocytes showed positive staining with the anti-ovine CD5 mAb (clone ST1), which is in contrast to the human and murine systems. The present findings indicated low homogeneity between camels and other species in the antigenic structure of leukocyte antigens, highlighting the need to develop camel-specific mAbs against the main immune cell markers.

Identification of Linear Epitopes in the C-Terminal Region of ASFV p72 Protein.

African swine fever, which is induced by the African swine fever virus (ASFV), poses a significant threat to the global pig industry due to its high lethality in domestic pigs and wild boars. Despite the severity of the disease, there is a lack of effective vaccines and drugs against the ASFV. The p72 protein, constituting 31 to 33% of the total virus particle mass, serves as the primary capsid protein of ASFV. It is a crucial antigen for the development of ASF subunit vaccines and serological diagnostic methods. In this investigation, 27 monoclonal antibodies (mAbs) were generated through mouse immunization with the truncated C-terminal p72 protein expressed by Escherichia coli. Among these, six mAbs exhibited binding to the p72 trimer, with their respective recognized epitopes identified as 542VTAHGINLIDKF553, 568GNAIKTP574, and 584FALKPREEY592. All three epitopes were situated within the interval sequences of functional units of the C-terminal jelly-roll barrel of p72. Notably, two epitopes, 568GNAIKTP574 and 584FALKPREEY592, were internal to the p72 trimer, while the epitope 542VTAHGINLIDKF553 was exposed on the surface of the trimer and consistently conserved across all ASFV genotypes. These findings enhance our comprehension of the antigenic function and structure of the p72 protein, facilitating the utilization of p72 in the development of diagnostic techniques for ASFV.

GGNpTCR: A Generative Graph Structure Neural Network for Predicting Immunogenic Peptides for T-cell Immune Response.

Identifying the interactions between T-cell receptor (TCRs) and human antigens is a crucial step in developing new vaccines, diagnostics, and immunotherapy. Current methods primarily focus on learning binding patterns from known TCR binding repertoires by using sequence information alone without considering the binding specificity of new antigens or exogenous peptides that have not appeared in the training set. Furthermore, the spatial structure of antigens plays a critical role in immune studies and immunotherapy, which should be addressed properly in the identification of interacting TCR-antigen pairs. In this study, we introduced a novel deep learning framework based on generative graph structures, GGNpTCR, for predicting interactions between TCR and peptides from sequence information. Results of real data analysis indicate that our model achieved excellent prediction for new antigens unseen in the training data set, making significant improvements compared to existing methods. We also applied the model to a large COVID-19 data set with no antigens in the training data set, and the improvement was also significant. Furthermore, through incorporation of additional supervised mechanisms, GGNpTCR demonstrated the ability to precisely forecast the locations of peptide-TCR interactions within 3D configurations. This enhancement substantially improved the model's interpretability. In summary, based on the performance on multiple data sets, GGNpTCR has made significant progress in terms of performance, universality, and interpretability.

Conserved antigen structures and antibody-driven variations on foot-and-mouth disease virus serotype A revealed by bovine neutralizing monoclonal antibodies.

Foot-and-mouth disease virus (FMDV) serotype A is antigenically most variable within serotypes. The structures of conserved and variable antigenic sites were not well resolved. Here, a historical A/AF72 strain from A22 lineage and a latest A/GDMM/2013 strain from G2 genotype of Sea97 lineage were respectively used as bait antigen to screen single B cell antibodies from bovine sequentially vaccinated with A/WH/CHA/09 (G1 genotype of Sea97 lineage), A/GDMM/2013 and A/AF72 antigens. Total of 39 strain-specific and 5 broad neutralizing antibodies (bnAbs) were isolated and characterized. Two conserved antigenic sites were revealed by the Cryo-EM structures of FMDV serotype A with two bnAbs W2 and W125. The contact sites with both VH and VL of W125 were closely around icosahedral threefold axis and covered the B-C, E-F, and H-I loops on VP2 and the B-B knob and H-I loop on VP3; while contact sites with only VH of W2 concentrated on B-B knob, B-C and E-F loops on VP3 scattering around the three-fold axis of viral particle. Additional highly conserved epitopes also involved key residues of VP158, VP1147 and both VP272 / VP1147 as determined respectively by bnAb W153, W145 and W151-resistant mutants. Furthermore, the epitopes recognized by 20 strain-specific neutralization antibodies involved the key residues located on VP3 68 for A/AF72 (11/20) and VP3 175 position for A/GDMM/2013 (9/19), respectively, which revealed antigenic variation between different strains of serotype A. Analysis of antibody-driven variations on capsid of two virus strains showed a relatively stable VP2 and more variable VP3 and VP1. This study provided important information on conserve and variable antigen structures to design broad-spectrum molecular vaccine against FMDV serotype A.

The OL101 O antigen locus specifies a novel Klebsiella pneumoniae serotype O13 structure

Klebsiella pneumoniae is one of the priority objects for the development of new therapies against infections. The species has been perceived as of limited variety of O antigens (11 O serotypes identified to date). That trait makes lipopolysaccharide an attractive target for protective antibodies. Nowadays, K. pneumoniae O antigens encoding genes are often analysed by bioinformatic tools, such as Kaptive, indicating higher actual diversity of the O antigen loci. One of the novel K. pneumoniae O loci for which the antigen structure has not been elucidated so far is OL101. In this study, four clinical isolates predicted as OL101 were characterized and found to have the O antigen structure composed of β-Kdop-[→3)-α-l-Rhap-(1 → 4)-α-d-Glcp]n, representing a novel serotype O13. Identification of the β-Kdop terminus was based on the analysis of the complete LPS molecule by the HR-MAS NMR spectroscopy. The bioinformatic analysis of 71,377 K. pneumoniae genomes from public databases (July 2023) revealed a notable OL101 prevalence of 6.55 %.

Comparison of Neutrophil-Lymphocyte Ratio in Patients with COVID-19 and Dengue Hemorrhagic Fever

Coronavirus Disease-19 (COVID-19) and Dengue Hemorrhagic Fever (DHF) can be found together in patients and are difficult to distinguish because they have the same antigenic viral structure, symptoms, and laboratory findings. Neutrophil-Lymphocyte Ratio (NLR) is one of the inflammatory markers that are both easy and have fast results, so it is considered adequate for managing both diseases. This study aimed to compare NLR values and cut-off based on degrees in COVID-19 and DHF patients. The study used data from medical records of 459 COVID-19 patients and 95 DHF patients treated at UNHAS RSPTN and Sayang Rakyat Hospital from July 2020 to August 2021. They had routine hematological examination results when they were first admitted. COVID-19 samples were grouped into severe and moderate degrees, and DHF samples were grouped into DHF without shock and DHF in shock. Mann-Whitney test, independent T-test, and ROC curve were used to compare the two groups (significant if the p-value was <0.05). There was a difference in NLR median between moderate COVID-19 (2.57) and severe COVID-19 (6.39) (p<0.001); between DHF without-shock (0.49) and DHF in-shock (0.43) (p<0.001). Neutrophil lymphocyte ratio cut-off between moderate and severe COVID-19 based on ROC curve was 5.66 (sensitivity 79.6%, specificity 73.5%), and between DHF without-shock and DHF in-shock was 0.39 (sensitivity 93.2%, specificity 83.3%). Neutrophil lymphocyte ratio values in COVID-19 are different from DHF. Neutrophil-lymphocyte ratio values are higher in severe COVID-19 than moderate, while NLR values are lower in DHF in shock rather than without shock.

Alloimmune hemolytic disease of the fetus and newborn: genetics, structure, and function of the commonly involved erythrocyte antigens.

Hemolytic disease of the fetus and newborn (HDFN) can occur when a pregnant woman has antibody directed against an erythrocyte surface antigen expressed by her fetus. This alloimmune disorder is restricted to situations where transplacental transfer of maternal antibody to the fetus occurs, and binds to fetal erythrocytes, and significantly shortens the red cell lifespan. The pathogenesis of HDFN involves maternal sensitization to erythrocyte "non-self" antigens (those she does not express). Exposure of a woman to a non-self-erythrocyte antigen principally occurs through either a blood transfusion or a pregnancy where paternally derived erythrocyte antigens, expressed by her fetus, enter her circulation, and are immunologically recognized as foreign. This review focuses on the genetics, structure, and function of the erythrocyte antigens that are most frequently involved in the pathogenesis of alloimmune HDFN. By providing this information we aim to convey useful insights to clinicians caring for patients with this condition.

Design, structure and plasma binding of ancestral β-CoV scaffold antigens

We report the application of ancestral sequence reconstruction on coronavirus spike protein, resulting in stable and highly soluble ancestral scaffold antigens (AnSAs). The AnSAs interact with plasma of patients recovered from COVID-19 but do not bind to the human angiotensin-converting enzyme 2 (ACE2) receptor. Cryo-EM analysis of the AnSAs yield high resolution structures (2.6–2.8 Å) indicating a closed pre-fusion conformation in which all three receptor-binding domains (RBDs) are facing downwards. The structures reveal an intricate hydrogen-bonding network mediated by well-resolved loops, both within and across monomers, tethering the N-terminal domain and RBD together. We show that AnSA-5 can induce and boost a broad-spectrum immune response against the wild-type RBD as well as circulating variants of concern in an immune organoid model derived from tonsils. Finally, we highlight how AnSAs are potent scaffolds by replacing the ancestral RBD with the wild-type sequence, which restores ACE2 binding and increases the interaction with convalescent plasma.

Epidemiological effectiveness of domestic influenza vaccines in cases with vaccination against a new coronavirus infection in 2022–2023

The study presents the results of an epidemiological prospective field cohort study on a comparative assessment of the epidemiological effectiveness of domestic inactivated influenza vaccines and vaccines against new coronavirus infection during immunization of adults aged 18 years. Statistically significant differences were found in the incidence of influenza, acute upper respiratory infections, and new coronavirus infection between the vaccinated and unvaccinated groups. The etiologies of cases of acute upper respiratory infections registered in the study participants during the epidemic season of influenza in 2022–2023 were verified. The clinical symptoms of influenza in the vaccinated group were significantly less severe than those in the unvaccinated group. No significant differences were found in the clinical picture of acute upper respiratory infections between the compared groups. Influenza immunoprophylaxis is an urgent problem, which consists of the ability of viruses to change their antigenic structure and avoid immunity, remaining in constant circulation. Thus, the antigenic composition of influenza vaccines must be systematically updated to ensure the formation of effective population immunity. More studies on the etiological spectrum of respiratory pathogens and assessment of the epidemiological effectiveness of vaccination against vaccine-controlled infections in the analyzed groups are needed. Therefore, to assess immunological protection, including the assessment of the epidemiological effectiveness of seasonal vaccines used against influenza and new coronavirus infection, the priority age group includes individuals aged 18–25 years (especially those belonging to organized collectives) and adults with an unfavorable premorbid background. The use of domestic vaccines against influenza and the new coronavirus infection for specific immunoprophylaxis is generally recognized as the most effective sanitary and anti-epidemic (preventive) measure in the morbidity management of several infectious diseases.

Synthesis, molecular modeling investigation, molecular dynamic and ADME prediction of some novel Mannich bases derived from 1,2,4-triazole, and assessment of their anticancer activity

A series of biologically active novel Mannich bases containing with a 1H-1,2,4-triazole-5-one ring were developed to evaluate the cytotoxic activity. For this purpose, the synthesized Schiff Bases (S1-5) were reacted with formaldehyde and morpholine, which is a secondary amine to yield novel N-Mannich bases (M1-5) via the Mannich reaction. The structures of the compounds (M1-5) were determined structurally employing 1H/13C-NMR, IR and elemental analysis. In this study, we evaluated the cytotoxic potential of the compounds (M1-5) on the human hypopharyngeal carcinoma FaDu cells. We found that the compound (M3) possesses a significant anticancer feature against FaDu cells that might be evaluated with further in vitro and in vivo studies to understand its anticancer potential better. Lastly, comparisons were made using molecular docking calculations to find the theoretical activities of the compounds (M1-5). The docking score parameter of the compound (M3) against the 2DO4 protein is −5.67, the docking score parameter against the 5JPZ protein is −5.72, and finally, the docking score parameter against the 2H80 protein is −5.50. Molecular dynamic calculations are made for 0–100 ns. The ADME/T calculations were performed to find the drug potential of the compounds (M1-5). The results suggest that our drug candidate compound exhibits strong potential for co-administration with the antigen structures, owing to the low rate of interactions that decreased over time. Communicated by Ramaswamy H. Sarma

Synthesis and Immunological Evaluation of Escherichia coli O1- Derived Oligosaccharide–Protein Conjugates toward Avian Pathogenic Escherichia coli O1 Vaccine Development

AbstractAvian pathogenic Escherichia coli O1 (APEC O1) is a pathogenic bacterium that causes significant economic losses in the poultry industry and raises concerns about zoonotic infections. The development of effective vaccines against APEC O1 is essential due to antibiotic resistance and the potential for severe symptoms in both chickens and humans. In this context, we have been focusing on the O1A, O1B, and O1C antigen structures derived from E. coli O1 lipopolysaccharide (LPS). In this study, the first synthesis of the pentasaccharide repeating units of the O1B and O1C antigens was successfully achieved. The synthesis and immunological evaluation of their conjugates with bovine serum albumin (BSA) were conducted. Only the O1A-pentasaccharide structure is a glycotope candidate for APEC O1. Keyhole limpet hemocyanin (KLH)–O1A-pentasaccharide conjugate was also synthesized, and its immunogenicity was evaluated by the ELISA assay. The efficient production of antibodies capable of binding to both APEC O1 LPS and the O1A-pentasaccharide structure was observed, indicating that O1A-pentasaccharide is a promising vaccine candidate against APEC O1.

The Influenza B Virus Victoria and Yamagata Lineages Display Distinct Cell Tropism and Infection-Induced Host Gene Expression in Human Nasal Epithelial Cell Cultures.

Understanding Influenza B virus infections is of critical importance in our efforts to control severe influenza and influenza-related diseases. Until 2020, two genetic lineages of influenza B virus-Yamagata and Victoria-circulated in the population. These lineages are antigenically distinct, but the differences in virus replication or the induction of host cell responses after infection have not been carefully studied. Recent IBV clinical isolates of both lineages were obtained from influenza surveillance efforts of the Johns Hopkins Center of Excellence in Influenza Research and Response and characterized in vitro. B/Victoria and B/Yamagata clinical isolates were recognized less efficiently by serum from influenza-vaccinated individuals in comparison to the vaccine strains. B/Victoria lineages formed smaller plaques on MDCK cells compared to B/Yamagata, but infectious virus production in primary human nasal epithelial cell (hNEC) cultures showed no differences. While ciliated epithelial cells were the dominant cell type infected by both lineages, B/Victoria lineages had a slight preference for MUC5AC-positive cells, and B/Yamagata lineages infected more basal cells. Finally, while both lineages induced a strong interferon response 48 h after infection of hNEC cultures, the B/Victoria lineages showed a much stronger induction of interferon-related signaling pathways compared to B/Yamagata. This demonstrates that the two influenza B virus lineages differ not only in their antigenic structure but also in their ability to induce host innate immune responses.

A bioinformatic analysis: Previous allergen exposure may support anti- SARS-CoV-2 immune response

COVID-19, caused by infection with the SARS-CoV-2 has become a global health problem due to significant mortality rates; the exact pathophysiological mechanism remains uncertain. Articles reporting patient data are quite heterogeneous and have several limitations. Surviving patients develop a CD4 and CD8 T-cell response to the virus SARS-CoV-2 during COVID-19. Interestingly, pre-existing virus-reactive T-cells have been found in patients that were not infected before, suggesting some form of cross-reactivity or immunological mimicry. To better understand this phenomenon, we performed a bioinformatic study, which was aimed to identify antigenic structures that may explain the presence of such “reactive” T-cells, which may support or modulate the immune response to SARS-CoV-2 infections. Seven different common environmental allergen epitopes identical to the SARS-CoV-2 S-protein were identified that share affinity to 8 MHCI-specific epitope regions. Pollen showed the greatest similarity with the S protein epitope. In the epitope similarity analysis between the S protein and MHC-II / T helper epitopes, the highest similarity was determined for mites. When S-protein that stimulates B cells and identical epitope antigens are examined, the most common allergens were hornbeam and wheat. The high epitope similarity observed for the allergens examined and S protein epitopes suggest that these allergens may be a reason for pre-existing SARS-CoV-2 – reactive T-cells in previously non-infected subjects and such a previous exposure may affect the course of the disease in COVID-19 infection. It remains to be determined whether such a previous existence of SARS-CoV-2 reactive cells can support the clearance of the virus or if they, in contrast, may even aggravate the disease course. (Table 4, Ref 54)

Heterologous DNA-prime/protein-boost immunization with a monomeric SARS-CoV-2 spike antigen redundantizes the trimeric receptor-binding domain structure to induce neutralizing antibodies in old mice.

A multitude of alterations in the old immune system impair its functional integrity. Closely related, older individuals show, for example, a reduced responsiveness to severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) vaccines. However, systematic strategies to specifically improve the efficacy of vaccines in the old are missing or limited to simple approaches like increasing the antigen concentration or injection frequencies. We here asked whether the intrinsic, trimeric structure of the SARS-CoV-2 spike (S) antigen and/or a DNA- or protein-based antigen delivery platform affects priming of functional antibody responses particularly in old mice. The used S-antigens were primarily defined by the presence/absence of the membrane-anchoring TM domain and the closely interlinked formation/non-formation of a trimeric structure of the receptor binding domain (S-RBD). Among others, we generated vectors expressing prefusion-stabilized, cell-associated (TM+) trimeric "S2-P" or secreted (TM-) monomeric "S6-PΔTM" antigens. These proteins were produced from vector-transfected HEK-293T cells under mild conditions by Strep-tag purification, revealing that cell-associated but not secreted S proteins tightly bound Hsp73 and Grp78 chaperones. We showed that both, TM-deficient S6-PΔTM and full-length S2-P antigens elicited very similar S-RBD-specific antibody titers and pseudovirus neutralization activities in young (2-3 months) mice through homologous DNA-prime/DNA-boost or protein-prime/protein-boost vaccination. The trimeric S2-P antigen induced high S-RBD-specific antibody responses in old (23-24 months) mice through DNA-prime/DNA-boost vaccination. Unexpectedly, the monomeric S6-PΔTM antigen induced very low S-RBD-specific antibody titers in old mice through homologous DNA-prime/DNA-boost or protein-prime/protein-boost vaccination. However, old mice efficiently elicited an S-RBD-specific antibody response after heterologous DNA-prime/protein-boost immunization with the S6-PΔTM antigen, and antibody titers even reached similar levels and neutralizing activities as in young mice and also cross-reacted with different S-variants of concern. The old immune system thus distinguished between trimeric and monomeric S protein conformations: it remained antigen responsive to the trimeric S2-P antigen, and a simple change in the vaccine delivery regimen was sufficient to unleash its reactivity to the monomeric S6-PΔTM antigen. This clearly shows that both the antigen structure and the delivery platform are crucial to efficiently prime humoral immune responses in old mice and might be relevant for designing "age-adapted" vaccine strategies.

Transport Functions of Serum Immunoglobulins Among the Residents of the European Arctic of the Russian Federation

The paper presents data on the level of serum immunoglobulins in residents of the North and Arctic of the European territory of the Russian Federation. In people living in the Arctic, the average concentration of IgM, IgA and IgE is 1.4–2.6 times higher, the frequency of elevated concentrations is 2.4–8.8 times higher. A high frequency of IgG deficiency of 72.3% was established, which indicates inhibition of switching of antibody synthesis with a predominant predominance of IgM and IgA. In an unfavorable climate, the spectrum of antigenic structures increases and expands significantly. Activation of antibody production is due to an increased level of antigenic effects on the body and an increase in the content, diversity in the intravascular environment of tissue metabolism products with the properties of autoantigens. It was revealed that the concentration of immunoglobulins is significant in the formation of circulating immune complexes (CIC). In residents of the Arctic of the European territory of the Russian Federation, increased concentrations of IgM and IgA are associated with an increase in the content of neutrophil granulocytes and interferon-gamma cytokine (IFN-γ), which in turn is aimed at ensuring the effectiveness of the clearance of waste products in hypoxia.

Dynamic structural analysis-based epitope prediction of Exendin-4 in aqueous solution.

The study of epitopes has a broad range of applications in drug discovery, vaccine design, and immunotherapy. In this study, an epitope prediction method was developed based on the dynamic structure of protein antigens. Solvent accessible surface area, charge, and root mean square fluctuation were introduced as the key residue property parameters. The epitope prediction algorithm was established by constructing a three-parameter complex metrics of seven-peptide groups. The method was applied to predict the epitopes of Exendin-4, an effective antidiabetic drug. The epitopes of both the natural and C-terminal amidated forms of Exendin-4 were predicted and compared in their folded and intermediate states. In the folded state, the epitopes of natural Exendin-4 (His1-Phe6 and Asp9-Val19) were found to be nearly identical to the epitopes of C-terminal aminated Exendin-4 (His1-Thr7 and Asp9-Val19). In the intermediate state, however, the epitopes of natural Exendin-4 (His1-Gly4, Phe6 and Lys12-Arg20) covered fewer amino acids than the epitopes of C-terminal aminated Exendin-4 (His1-Gly4, Phe6, Asp9-Val19 and Trp25-Lys27). The comparison with the results from other prediction tools demonstrates the reliability of our predicted epitopes of Exendin-4.

БИЗНЕС-ЛОГИСТИЧЕСКИЙ ПОДХОД К ИССЛЕДОВАНИЮ ФАРМАЦЕВТИЧЕСКОГО РЫНКА ПРЕПАРАТОВ ХАБАРОВСКОГО КРАЯ ДЛЯ ЛЕЧЕНИЯ ГРИППА, ОРВИ

Analyzing the infectious morbidity of the population, it should be noted that the predominant places in the general structure belong to such nosologies as influenza and SARS, which more than 35 million citizens get sick in the country every year. In this regard, the generated economic damage reaches, according to various sources, 50-100 billion rubles /year, while the total damage from the entire group of infectious diseases ranges from 80-125 billion rubles/year. Due to the fact that the peak incidence occurs in the autumn-spring period, it is at this time that the greatest demand for antiviral drugs (LP) is noted. Due to the high variability of the antigenic structure of RNA-containing viruses (more than two thousand variants of strains) and the airborne transmission pathway, which increases the distribution area, data infections are widespread among the population and are registered annually not by single outbreaks, but cause a wave of epidemics with a lethality of 250-650 thousand people/year. The greatest danger of these viruses is for people with not yet formed and (or) weakened immunity: children, pregnant women, the elderly, especially if it occurs not in mild or moderate, but in severe or toxic forms, causing a number of complications. The article presents an overview of the research of the Khabarovsk Territory pharmaceutical market of drugs for the treatment of influenza, acute respiratory viral infections for the period of 2016-2020. The aim of the research was to study the consumption range of drugs for the treatment of influenza and ARVI in the Khabarovsk Territory of the Russian Federation. Based on the data obtained, we can draw conclusions on the quality of this region’s pharmaceutical care to the population with influenza and ARVI in the period under review.

Identification of Potential Novel B-Cell Epitopes of Capsid Protein VP2 in Senecavirus A.

Senecavirus A (SVA) is a type of nonenveloped single-stranded, positive-sense RNA virus. The VP2 protein is a structural protein that plays an important role in inducing early and late immune responses of the host. However, its antigenic epitopes have not been fully elucidated. Therefore, defining the B epitopes of the VP2 protein is of great importance to revealing its antigenic characterization. In this study, we analyzed B-cell immunodominant epitopes (IDEs) of the VP2 protein from the SVA strain CH/FJ/2017 using the Pepscan approach and a bioinformatics-based computational prediction method. The following four novel IDEs of VP2 were identified: IDE1, 41TKSDPPSSSTDQPTTT56; IDE2, 145PDGKAKSLQELNEEQW160; IDE3, 161VEMSDDYRTGKNMPF175; and IDE4, 267PYFNGLRNRFTTGT280. Most of the IDEs were highly conserved among the different strains. To our knowledge, the VP2 protein is a major protective antigen of SVA that can induce neutralizing antibodies in animals. Here, we analyzed the immunogenicity and neutralization activity of four IDEs of VP2. Consequently, all four IDEs showed good immunogenicity that could elicit specific antibodies in guinea pigs. A neutralization test in vitro showed that the peptide-specific guinea pig antisera of IDE2 could neutralize SVA strain CH/FJ/2017, and IDE2 was identified as a novel potential neutralizing linear epitope. This is the first time VP2 IDEs have been identified by using the Pepscan method and a bioinformatics-based computational prediction method. These results will help elucidate the antigenic epitopes of VP2 and clarify the basis for immune responses against SVA. IMPORTANCE The clinical symptoms and lesions caused by SVA are indistinguishable from those of other vesicular diseases in pigs. SVA has been associated with recent outbreaks of vesicular disease and epidemic transient neonatal losses in several swine-producing countries. Due to the continuing spread of SVA and the lack of commercial vaccines, the development of improved control strategies is urgently needed. The VP2 protein is a crucial antigen on the capsids of SVA particles. Furthermore, the latest research showed that VP2 could be a promising candidate for the development of novel vaccines and diagnostic tools. Hence, a detailed exploration of epitopes in the VP2 protein is necessary. In this study, four novel B-cell IDEs were identified using two different antisera with two different methods. IDE2 was identified as a new neutralizing linear epitope. Our findings will help in the rational design of epitope vaccines and further understanding of the antigenic structure of VP2.

2019–2020 H1N1 clade A5a.1 viruses have better in vitro fitness compared with the co-circulating A5a.2 clade

Surveillance for emerging human influenza virus clades is important for identifying changes in viral fitness and assessing antigenic similarity to vaccine strains. While fitness and antigenic structure are both important aspects of virus success, they are distinct characteristics and do not always change in a complementary manner. The 2019–2020 Northern Hemisphere influenza season saw the emergence of two H1N1 clades: A5a.1 and A5a.2. While several studies indicated that A5a.2 showed similar or even increased antigenic drift compared with A5a.1, the A5a.1 clade was still the predominant circulating clade that season. Clinical isolates of representative viruses from these clades were collected in Baltimore, Maryland during the 2019–2020 season and multiple assays were performed to compare both antigenic drift and viral fitness between clades. Neutralization assays performed on serum from healthcare workers pre- and post-vaccination during the 2019–2020 season show a comparable drop in neutralizing titers against both A5a.1 and A5a.2 viruses compared with the vaccine strain, indicating that A5a.1 did not have antigenic advantages over A5a.2 that would explain its predominance in this population. Plaque assays were performed to investigate fitness differences, and the A5a.2 virus produced significantly smaller plaques compared with viruses from A5a.1 or the parental A5a clade. To assess viral replication, low MOI growth curves were performed on both MDCK-SIAT and primary differentiated human nasal epithelial cell cultures. In both cell cultures, A5a.2 yielded significantly reduced viral titers at multiple timepoints post-infection compared with A5a.1 or A5a. Receptor binding was then investigated through glycan array experiments which showed a reduction in receptor binding diversity for A5a.2, with fewer glycans bound and a higher percentage of total binding attributable to the top three highest bound glycans. Together these data indicate that the A5a.2 clade had a reduction in viral fitness, including reductions in receptor binding, that may have contributed to the limited prevalence observed after emergence.

Chapter 2b: The molecular and antigenic structure of the TBEV

Chapter 2b: The molecular and antigenic structure of the TBEV