Characterization of Conserved Evolution in H7N9 Avian Influenza Virus Prior Mass Vaccination.

Vaccination is crucial for the prevention and mitigation of avian influenza infections in China. The inactivated H7N9 vaccine, when administered to poultry, significantly lowers the risk of infection among both poultry and humans, while also markedly decreasing the prevalence of H7N9 detections. Highly pathogenic (HP) H7N9 viruses occasionally appear, whereas their low pathogenicity (LP) counterparts have been scarcely detected since 2018. However, these contributing factors remain poorly understood. We conducted an exploratory investigation of the mechanics via the application of comprehensive bioinformatic approaches. We delineated the Yangtze River Delta (YRD) H7N9 lineage into 5 clades (YRD-A to E). Our findings highlight the emergence and peak occurrence of the LP H7N9-containing YRD-E clade during the 5th epidemic wave in China's primary poultry farming areas. A more effective control of LP H7N9 through vaccination was observed compared to that of its HP H7N9 counterpart. YRD-E exhibited a tardy evolutionary trajectory, denoted by the conservation of its genetic and antigenic variation. Our analysis of YRD-E revealed only minimal amino acid substitutions along its phylogenetic tree and a few selective sweep mutations since 2016. In terms of epidemic fitness, the YRD-E was measured to be lower than that of the HP variants. Collectively, these findings underscore the conserved evolutionary patterns distinguishing the YRD-E. Given the conservation presented in its evolutionary patterns, the YRD-E LP H7N9 is hypothesized to be associated with a reduction following the mass vaccination in a relatively short period owing to its lower probability of antigenic variation that might affect vaccine efficiency.

Neutralization potency of the 2023-24 seasonal influenza vaccine against circulating influenza H3N2 strains.

Seasonal influenza is a severe disease that significantly impacts public health, causing millions of infections and hundreds of thousands of deaths each year. Seasonal influenza viruses, particularly the H3N2 subtype, exhibit high antigenic variability, often leading to mismatch between vaccine strains and circulating strains. Therefore, rapidly assessing the alignment between existing seasonal influenza vaccine and circulating strains is crucial for enhancing vaccine efficacy. This study, based on a pseudovirus platform, evaluated the match between current influenza H3N2 vaccine strains and circulating strains through cross-neutralization assays using clinical human immune sera against globally circulating influenza virus strains. The research results show that although mutations are present in the circulating strains, the current H3N2 vaccine strain still imparting effective protection, providing a scientific basis for encouraging influenza vaccination. This research methodology can be sustainably applied for the neutralization potency assessment of subsequent circulating strains, establishing a persistent methodological framework.

Hyper-diverse antigenic variation and resilience to transmission-reducing intervention in falciparum malaria.

Intervention efforts against falciparum malaria in high-transmission regions remain challenging, with rapid resurgence typically following their relaxation. Such resilience co-occurs with incomplete immunity and a large transmission reservoir from high asymptomatic prevalence. Incomplete immunity relates to the large antigenic variation of the parasite, with the major surface antigen of the blood stage of infection encoded by the multigene and recombinant family known as var. With a stochastic agent-based model, we investigate the existence of a sharp transition in resurgence ability with intervention intensity and identify molecular indicators informative of its proximity. Their application to survey data with deep sampling of var sequences from individual isolates in northern Ghana suggests that the transmission system was brought close to transition by intervention with indoor residual spraying. These results indicate that sustaining and intensifying intervention would have pushed malaria dynamics to a slow-rebound regime with an increased probability of local parasite extinction.

Original antigenic sin: A potential double-edged effect for vaccine improvement.

Original antigenic sin (OAS) influences the immune response to subsequent infections with related variants following initial pathogen exposure. This phenomenon is characterized by cross-reactivity, which, although it may worsen infections, also provides a degree of protection against immune evasion caused by variations. This paradox complicates the development of creating universal vaccinations, as they frequently show diminished effectiveness against these emerging variants. This review aims to elucidate the diverse impacts of OAS on the immune response to various infections, emphasizing the complicated balance between beneficial and harmful outcomes. Moreover, we evaluate the influence of adjuvants and other variables on the extent of OAS, hence affecting the effectiveness of vaccines. Understanding the mechanisms of OAS that cause persistent infections and evasion of the immune system is crucial for the developing innovative vaccines. And it has significant potential for clinical applications.

Antigenic cartography using variant-specific hamster sera reveals substantial antigenic variation among Omicron subvariants.

Severe acute respiratory syndrome Coronavirus 2 (SARS-CoV-2) has developed substantial antigenic variability. As the majority of the population now has pre-existing immunity due to infection or vaccination, the use of experimentally generated animal immune sera can be valuable for measuring antigenic differences between virus variants. Here, we immunized Syrian hamsters by two successive infections with one of nine SARS-CoV-2 variants. Their sera were titrated against 16 SARS-CoV-2 variants, and the resulting titers were visualized using antigenic cartography. The antigenic map shows a condensed cluster containing all pre-Omicron variants (D614G, Alpha, Delta, Beta, Mu, and an engineered B.1+E484K variant) and considerably more diversity among a selected panel of Omicron subvariants (BA.1, BA.2, BA.4/BA.5, the BA.5 descendants BF.7 and BQ.1.18, the BA.2.75 descendant BN.1.3.1, the BA.2-derived recombinants XBB.2 and EG.5.1, and the BA.2.86 descendant JN.1). Some Omicron subvariants were as antigenically distinct from each other as the wildtype is from the Omicron BA.1 variant. Compared to titers measured in human sera, titers in hamster sera are of higher magnitude, show less fold change, and result in a more compact antigenic map topology. The results highlight the potential of sera from hamsters for the continued antigenic characterization of SARS-CoV-2.

HA antigenic variation and phylogenetic analysis of influenza B virus in Shiraz, Iran.

BACKGROUND: Influenza infection is highly contagious respiratory illness triggered by the influenza virus, bearing substantial implications for global health. Influenza B viruses, specifically the Victoria and Yamagata lineages, have contributed to the disease burden, and the mismatch between circulating strains and vaccine strains has led to increased mortality and economic costs. Understanding the global epidemiology, seasonal variations, and genetic characteristics of influenza B is crucial for effective prevention and control strategies. METHODS: The study investigated influenza B viruses in Shiraz, Iran during the Oct 2017 to Jan 2018. Throat swabs were collected from 235 individuals under 15 with influenza-like symptoms including fever and cough. Samples were stored at -80°C and transported to the lab for further analysis. Viral RNA was extracted and analyzed using Real-time PCR. The hemagglutinin (HA) gene of positive samples was sequenced, and phylogenetic trees were constructed. Amino acids indicative of adaptive mutations were identified using global sequence data. RESULTS: 23 of 235 samples (9.7 %) were positive for influenza B virus. The most common clinical manifestations were rhinorrhea and myalgia, with 20 individuals (87 % of the 23 infected people) each showing these symptoms. The phylogenetic analysis of the HA gene showed that the Victoria isolates were close to the B/Brisbane/60/2008 strain (12.5 % of the positive samples) and belonged to clade-1A, while the Yamagata isolates were close to the B/Phuket/3037/2013 strain (87.5 % of the positive samples) and belonged to clade-3. CONCLUSION: The study highlights the need for importance vaccine coverage in the Shiraz region to address limited genetic diversity and strain mismatch. Continuous surveillance of mutations in the HA gene resulting in amino acid substitutions and their impact on vaccine efficacy is crucial. This study showed that the circulation of influenza B in Shiraz matched with the recommended Yamagata vaccine strain. These findings contribute to the understanding of influenza B dynamics and emphasize the importance of region-specific prevention and control strategies.

N-glycosylation on hemagglutinin head reveals inter-branch antigenic variability of avian influenza virus H5-subtypes.

H5-subtype avian influenza virus (AIV) is globally prevalent and undergoes frequent antigenic drift, necessitating regular updates to vaccines. One of the many influencing elements that cause incompatibility between vaccinations and epidemic strains is the dynamic alteration of glycosylation sites. However, the biological significance of N-glycosylation in the viral evolution and antigenic changes is unclear. Here, we performed a systematic analysis of glycosylation sites on the HA1 subunit of H5N1, providing insights into the changes of primary glycosylation sites, including 140 N, 156 N, and 170 N within the antigenic epitopes of HA1 protein. Multiple recombinant viruses were then generated based on HA genes of historical vaccine strains and deactivated for immunizing SPF chickens. Inactivated recombinant strains showed relatively closer antigenicity compared to which has identical N-glycosylation patterns. The N-glycosylation modification discrepancy highlights the inter-branch antigenic diversity of H5-subtype viruses in avian influenza and serves as a vital foundation for improving vaccination tactics.

A speed limit on serial strain replacement from original antigenic sin.

Many pathogens evolve to escape immunity, yet it remains difficult to predict whether immune pressure will lead to diversification, serial replacement of one variant by another, or more complex patterns. Pathogen strain dynamics are mediated by cross-protective immunity, whereby exposure to one strain partially protects against infection by antigenically diverged strains. There is growing evidence that this protection is influenced by early exposures, a phenomenon referred to as original antigenic sin (OAS) or imprinting. In this paper, we derive constraints on the emergence of the pattern of successive strain replacements demonstrated by influenza, SARS-CoV-2, seasonal coronaviruses, and other pathogens. We find that OAS implies that the limited diversity found with successive strain replacement can only be maintained if [Formula: see text] is less than a threshold set by the characteristic antigenic distances for cross-protection and for the creation of new immune memory. This bound implies a "speed limit" on the evolution of new strains and a minimum variance of the distribution of infecting strains in antigenic space at any time. To carry out this analysis, we develop a theoretical model of pathogen evolution in antigenic space that implements OAS by decoupling the antigenic distances required for protection from infection and strain-specific memory creation. Our results demonstrate that OAS can play an integral role in the emergence of strain structure from host immune dynamics, preventing highly transmissible pathogens from maintaining serial strain replacement without diversification.

Circulating rotavirus strains in children with acute gastroenteritis in Iran, 1986 to 2023 and their genetic/antigenic divergence compared to approved vaccines strains (Rotarix, RotaTeq, ROTAVAC, ROTASIIL) before mass vaccination: Clues for vaccination policy makers.

In the present study, first, rotaviruses that caused acute gastroenteritis in children under five years of age during the time before the vaccine was introduced in Iran (1986 to 2023) are reviewed. Subsequently, the antigenic epitopes of the VP7 and VP4/VP8 proteins in circulating rotavirus strains in Iran and that of the vaccine strains were compared and their genetic differences in histo-blood group antigens (HBGAs) and the potential impact on rotavirus infection susceptibility and vaccine efficacy were discussed. Overall data indicate that rotavirus was estimated in about 38.1 % of samples tested. The most common genotypes or combinations were G1 and P[8], or G1P[8]. From 2015 to 2023, there was a decline in the prevalence of G1P[8], with intermittent peaks of genotypes G3P[8] and G9P[8]. The analyses suggested that the monovalent Rotarix vaccine or monovalent vaccines containing the G1P[8] component might be proper in areas with a similar rotavirus genotype pattern and genetic background as the Iranian population where the G1P[8] strain is the most predominant and has the ability to bind to HBGA secretors. While the same concept can be applied to RotaTeq and RotasIIL vaccines, their complex vaccine technology, which involves reassortment, makes them less of a priority. The ROTASIIL vaccine, despite not having the VP4 arm (P[5]) as a suitable protection option, has previously shown the ability to neutralize not only G9-lineage I strains but also other G9-lineages at high titers. Thus, vaccination with the ROTASIIL vaccine may be more effective in Iran compared to RotaTeq. However, considering the rotavirus genotypic pattern, ROTAVAC might not be a good choice for Iran. Overall, the findings of this study provide valuable insights into the prevalence of rotavirus strains and the potential effectiveness of different vaccines in the Iranian and similar populations.

The Pathogenesis of Foot-and-Mouth Disease Virus Infection: How the Virus Escapes from Immune Recognition and Elimination.

Foot-and-mouth disease virus (FMDV) is a highly contagious and economically devastating pathogen that affects cloven-hoofed animals worldwide. FMDV infection causes vesicular lesions in the mouth, feet, and mammary glands, as well as severe systemic symptoms such as fever, salivation, and lameness. The pathogenesis of FMDV infection involves complex interactions between the virus and the host immune system, which determine the outcome of the disease. FMDV has evolved several strategies to evade immune recognition and elimination, such as antigenic variation, receptor switching, immune suppression, and subversion of innate and adaptive responses. This review paper summarizes the current knowledge on the pathogenesis of FMDV infection and the mechanisms of immune evasion employed by the virus. It also discusses the challenges and opportunities for developing effective vaccines and therapeutics against this important animal disease.

Seasonal antigenic prediction of influenza A H3N2 using machine learning.

Antigenic characterization of circulating influenza A virus (IAV) isolates is routinely assessed by using the hemagglutination inhibition (HI) assays for surveillance purposes. It is also used to determine the need for annual influenza vaccine updates as well as for pandemic preparedness. Performing antigenic characterization of IAV on a global scale is confronted with high costs, animal availability, and other practical challenges. Here we present a machine learning model that accurately predicts (normalized) outputs of HI assays involving circulating human IAV H3N2 viruses, using their hemagglutinin subunit 1 (HA1) sequences and associated metadata. Each season, the model learns an updated nonlinear mapping of genetic to antigenic changes using data from past seasons only. The model accurately distinguishes antigenic variants from non-variants and adaptively characterizes seasonal dynamics of HA1 sites having the strongest influence on antigenic change. Antigenic predictions produced by the model can aid influenza surveillance, public health management, and vaccine strain selection activities.

The antigenic landscape of human influenza N2 neuraminidases from 2009 until 2017.

Human H3N2 influenza viruses are subject to rapid antigenic evolution which translates into frequent updates of the composition of seasonal influenza vaccines. Despite these updates, the effectiveness of influenza vaccines against H3N2-associated disease is suboptimal. Seasonal influenza vaccines primarily induce hemagglutinin-specific antibody responses. However, antibodies directed against influenza neuraminidase (NA) also contribute to protection. Here, we analysed the antigenic diversity of a panel of N2 NAs derived from human H3N2 viruses that circulated between 2009 and 2017. The antigenic breadth of these NAs was determined based on the NA inhibition (NAI) of a broad panel of ferret and mouse immune sera that were raised by infection and recombinant N2 NA immunisation. This assessment allowed us to distinguish at least four antigenic groups in the N2 NAs derived from human H3N2 viruses that circulated between 2009 and 2017. Computational analysis further revealed that the amino acid residues in N2 NA that have a major impact on susceptibility to NAI by immune sera are in proximity of the catalytic site. Finally, a machine learning method was developed that allowed to accurately predict the impact of mutations that are present in our N2 NA panel on NAI. These findings have important implications for the renewed interest to develop improved influenza vaccines based on the inclusion of a protective NA antigen formulation.

Two proteins, the hemagglutinin and the neuraminidase, protrude from the surface of the influenza virus. Their detection by the immune system allows the host organism to mount defences against the viral threat. The virus evolves in response to this pressure, which manifests as changes in the appearance of its hemagglutinin and neuraminidase. This process, known as antigenic drift, leads to the proteins evading detection. It is also why flu vaccines require frequent updates, as they rely on 'training' the immune system to recognise the most important strains in circulation ­ primarily by exposing it to appropriate versions of hemagglutinin. While the antigenic drift of hemagglutinin has been extensively studied, much less is known about how the neuraminidase accumulates mutations, and how these affect the immune response. To investigate this question, Catani et al. selected 43 genetically distant neuraminidases from human viral samples isolated between 2009 and 2017. Statistical analyses were applied to define their relatedness, revealing that a group of closely related neuraminidases predominated from 2009 to 2015, before they were being taken over by a second group. A third group, which was identified in viruses isolated in 2013, was remarkably close to the neuraminidase of strains that circulated in the late 1990s. The fourth and final group of neuraminidases was derived from influenza viruses that normally circulate in pigs but can also occasionally infect humans. Next, Catani et al. examined the immune response that these 43 neuraminidases could elicit in mice, as well as in ferrets ­ the animal most traditionally used in influenza research. This allowed them to pinpoint which changes in the neuraminidase sequences were important to escape recognition by the host. Data obtained from the two model species were comparable, suggesting that these experiments could be conducted on mice going forward, which are easier to work with than ferrets. Finally, Catani et al. used machine learning to build a computational model that could predict how strongly the immune system would respond to a specific neuraminidase variant. These findings could help guide the development of new vaccines that include neuraminidases tailored to best prime and train the immune system against a larger variety of strains. This may aid the development of 'supra-seasonal' vaccines that protect against a broad range of influenza viruses, reducing the need for yearly updates.

Plasmodium falciparum AMA1 and CSP antigen diversity in parasite isolates from southern Ghana.

Introduction: Diversity in malarial antigens is an immune evasion mechanism that gives malaria parasites an edge over the host. Immune responses against one variant of a polymorphic antigen are usually not fully effective against other variants due to altered epitopes. This study aimed to evaluate diversity in the Plasmodium falciparum antigens apical membrane antigen 1 (PfAMA1) and circumsporozoite protein (PfCSP) from circulating parasites in a malaria-endemic community in southern Ghana and to determine the effects of polymorphisms on antibody response specificity. Methods: The study involved 300 subjects, whose P. falciparum infection status was determined by microscopy and PCR. Diversity within the two antigens was evaluated by msp2 gene typing and molecular gene sequencing, while the host plasma levels of antibodies against PfAMA1, PfCSP, and two synthetic 24mer peptides from the conserved central repeat region of PfCSP, were measured by ELISA. Results: Of the 300 subjects, 171 (57%) had P. falciparum infection, with 165 of the 171 (96.5%) being positive for either or both of the msp2 allelic families. Gene sequencing of DNA from 55 clonally infected samples identified a total of 56 non-synonymous single nucleotide polymorphisms (SNPs) for the Pfama1 gene and these resulted in 44 polymorphic positions, including two novel positions (363 and 365). Sequencing of the Pfcsp gene from 69 clonal DNA samples identified 50 non-synonymous SNPs that resulted in 42 polymorphic positions, with half (21) of these polymorphic positions being novel. Of the measured antibodies, only anti-PfCSP antibodies varied considerably between PCR parasite-positive and parasite-negative persons. Discussion: These data confirm the presence of a considerable amount of unique, previously unreported amino acid changes, especially within PfCSP. Drivers for this diversity in the Pfcsp gene do not immediately seem apparent, as immune pressure will be expected to drive a similar level of diversity in the Pfama1 gene.

Evolution and Antigenic Differentiation of Avian Influenza A(H7N9) Virus, China.

We characterized the evolution and molecular characteristics of avian influenza A(H7N9) viruses isolated in China during 2021-2023. We systematically analyzed the 10-year evolution of the hemagglutinin gene to determine the evolutionary branch. Our results showed recent antigenic drift, providing crucial clues for updating the H7N9 vaccine and disease prevention and control.

Mapping Genetic Markers Associated with Antigenicity and Host Range in H9N2 Influenza A Viruses Infecting Poultry in Pakistan.

The aim of the current study was to map the genetic diversity in the haemagglutinin (HA) glycoprotein of influenza A viruses (IAVs) of the H9N2 subtype. Twenty-five H9N2 IAVs were isolated from broiler chickens from March to July 2019. The HA gene was amplified, and phylogenetic analysis was performed to determine the evolutionary relationship. Important antigenic amino acid residues of HA attributed to immune escape and zoonotic potential were compared among H9N2 IAVs. Phylogenetic analysis revealed that sublineage B2 under the G1 lineage in Pakistan was found to be diversified, and newly sequenced H9N2 isolates were nested into two clades (A and B). Mutations linked to the antigenic variation and potential immune escape were observed as G72E (1/25, 4%), A180T (3/25, 12%), and A180V (1/25, 4%). A twofold significant reduction (P < 0.01) in log2 hemagglutination inhibition titers was observed with H9N2 IAV naturally harboring amino acid V180 instead of A180 in HA protein. Moreover, in the last 20 years, complete substitution at residues (T127D, D135N, and L150N) and partial substitution at residues (72, 74, 131, 148, 180, 183, 188, 216, 217, and 249, mature H9 HA numbering) associated with changes in antigenicity were observed. The presence of L216 in all H9N2 IAV isolates and T/V180 in four isolates in the receptor-binding site reveals the potential of these viruses to cross the species barrier to infect human or mammals. The current study observed the circulation of antigenically diverse H9N2 IAV variants that possess potential mutations that can escape the host immune system.

Nota de investigación- Mapeo de marcadores genéticos asociados con la antigenicidad y el rango de huéspedes en los virus de la influenza tipo A subtipo H9N2 que infectan a la avicultura en Pakistán. El objetivo del presente estudio fue mapear la diversidad genética en la glicoproteína hemaglutinina (HA) de los virus de la influenza A (IAV) del subtipo H9N2. Se aislaron veinticinco virus de influenza H9N2 de pollos de engorde de marzo a julio del 2019. Se amplificó el gene HA y se realizó un análisis filogenético para determinar la relación evolutiva. Se compararon importantes residuos de aminoácidos antigénicos de la hemaglutinina atribuidos al escape inmunológico y al potencial zoonótico entre los virus de la influenza aviar H9N2. El análisis filogenético reveló que el sublinaje B2 bajo el linaje G1 en Pakistán estaba diversificado, y los aislados de H9N2 recién secuenciados se agruparon en dos clados (A y B). Se observaron mutaciones relacionadas con la variación antigénica y el posible escape inmunológico como los residuos de aminoácidos G72E (1/25, 4%), A180T (3/25, 12%) y A180V (1/25, 4%). Se observó una reducción significativa al doble (P < 0.01) en los títulos de inhibición de la hemaglutinación log2 cuando el virus de la influenza aviar H9N2 albergaba naturalmente el aminoácido V180 en lugar del A180 en la proteína HA. Además, en los últimos 20 años, sustitución completa en los residuos (T127D, D135N y L150N) y sustitución parcial en los residuos (72, 74, 131, 148, 180, 183, 188, 216, 217 y 249, de acuerdo con la numeración de la HA subtipo madura) asociados con cambios en la antigenicidad. La presencia del residuo L216 en todos los aislados de influenza aviar H9N2 y T/V180 en cuatro aislados en el sitio de unión al receptor revela el potencial de estos virus para cruzar la barrera de las especies para infectar a humanos o mamíferos. El estudio actual observó la circulación de variantes antigénicamente diversas del virus de influenza aviar H9N2 que poseen mutaciones potenciales que pueden escapar del sistema inmunológico del huésped.

Prediction of the antigenic regions in eight RhD variants identified by computational biology.

BACKGROUND AND OBJECTIVES: Changes in RHD generate variations in protein structure that lead to antigenic variants. The classical model divides them into quantitative (weak and Del) and qualitative (partial D). There are two types of protein antigens: linear and conformational. Computational biology analyses the theoretical assembly of tertiary protein structures and allows us to identify the 'topological' differences between isoforms. Our aim was to determine the theoretical antigenic differences between weak RhD variants compared with normal RhD based on structural analysis using bioinformatic techniques. MATERIALS AND METHODS: We analysed the variations in secondary structures and hydrophobicity of RHD*01, RHD*01W.1, W2, W3, RHD*09.03.01, RHD*09.04, RHD*11, RHD*15 and RHD*21. We then modelled the tertiary structure and calculated their probable antigenic regions, intra-protein interactions, displacement and membrane width and compared them with Rhce. RESULTS: The 10 proteins are similar in their secondary structure and hydrophobicity, with the main differences observed in the exofacial coils. We identified six potential antigenic regions: one that is unique to RhD (R3), one that is common to all D (R6), three that are highly variable among RhD isoforms (R1, R2 and R4), one that they share with Rhce (R5) and two that are unique to Rhce (Ra and Rbc). CONCLUSION: The alloimmunization capacity of these subjects could be explained by the variability of the antigen pattern, which is not necessarily recognized or recognized with lower intensity by the commercially available antibodies, and not because they have a lower protein concentration in the membrane.

Comprehensive analysis of antigenic variations and genomic properties of hepatitis B virus in clinical samples in the mid-north east region of Bangladesh.

This investigation delineates an exhaustive analysis of the clinical, immunological, and genomic landscapes of hepatitis B virus (HBV) infection across a cohort of 22 verified patients. The demographic analysis unveiled a pronounced male bias (77.27%), with patient ages spanning 20 to 85 years and durations of illness ranging from 10 days to 4 years. Predominant clinical manifestations included fever, fatigue, anorexia, abdominal discomfort, and arthralgia, alongside observed co-morbidities such as chronic renal disorders and hepatocellular carcinoma. Antigenic profiling of the HBV envelope proteins elucidated significant heterogeneity among the infected subjects, particularly highlighted by discordances in the detection capabilities of small and large HBsAg assays, suggesting antigenic diversity. Quantitative assessment of viral loads unveiled a broad spectrum, accompanied by atypical HBeAg reactivity patterns, challenging the reliability of existing serological markers. Correlative studies between viral burden and antigenicity of the envelope proteins unearthed phenomena indicative of diagnostic evasion. Notably, samples demonstrating robust viral replication were paradoxically undetectable by the large HBsAg ELISA kit, advocating for more sophisticated diagnostic methodologies. Genotypic examination of three HBV isolates classified them as genotype D (D2), with phylogenetic alignment to strains from various global origins. Mutational profiling identified pivotal mutations within the basic core promoter and preS2/S1 regions, associated with an augmented risk of hepatocellular carcinoma. Further, mutations discerned in the small HBsAg and RT/overlap regions were recognized as contributors to vaccine and/or diagnostic escape mechanisms. In summation, this scholarly discourse elucidates the intricate interplay of clinical presentations, antigenic diversity, and genomic attributes in HBV infection, accentuating the imperative for ongoing investigative endeavors to refine diagnostic and therapeutic modalities.

Intra-serotypic antigenic diversity of dengue virus serotype 3 in Thailand during 2004-2015.

In addition to the well-known differences among the four dengue serotypes, intra-serotypic antigenic diversity has been proposed to play a role in viral evolution and epidemic fluctuation. A replacement of genotype II by genotype III of dengue virus serotype 3 (DENV3) occurred in Thailand during 2007-2014, raising questions about the role of intra-serotypic antigenic differences in this genotype shift. We characterized the antigenic difference of DENV3 of genotypes II and III in Thailand, utilizing a neutralizing antibody assay with DENV3 vaccine sera and monotypic DENV3 sera. Although there was significant antigenic diversity among the DENV3, it did not clearly associate with the genotype. Our data therefore do not support the role of intra-serotypic antigenic difference in the genotype replacement. Amino acid alignment showed that eight positions are potentially associated with diversity between distinct antigenic subgroups. Most of these amino acids were found in envelope domain II. Some positions (aa81, aa124, and aa172) were located on the surface of virus particles, probably involving the neutralization sensitivity. Notably, the strains of both genotypes II and III showed clear antigenic differences from the vaccine genotype I strain. Whether this differencewill affect vaccine efficacy requires further studies.

Genomic Epidemiology of Treponema pallidum and Circulation of Strains With Diminished tprK Antigen Variation Capability in Seattle, 2021-2022.

BACKGROUND: The incidence of syphilis continues to increase in the United States, yet little is known about Treponema pallidum genomic epidemiology within American metropolitan areas. METHODS: We performed whole-genome sequencing and tprK deep sequencing of 28 T. pallidum-containing specimens, collected mostly from remnant Aptima swab specimens from 24 individuals from Seattle Sexual Health Clinic during 2021-2022. RESULTS: All 12 individuals infected with Nichols-lineage strains were men who have sex with men, while a specific SS14 cluster (mean, 0.33 single-nucleotide variant) included 1 man who has sex with women and 5 women. All T. pallidum strains sequenced were azithromycin resistant via 23S ribosomal RNA A2058G mutation. Identical T. pallidum genomic sequences were found in pharyngeal and rectal swab specimens taken concurrently from the same individuals. The tprK sequences were less variable between patient-matched specimens and between epidemiologically linked clusters. We detected a 528-base pair deletion in the tprK donor site locus, eliminating 9 donor sites, in T. pallidum genomes of 3 individuals with secondary syphilis, associated with diminution of TprK diversity. CONCLUSIONS: We developed an end-to-end workflow for public health genomic surveillance of T. pallidum from remnant Aptima swab specimens. tprK sequencing may assist in linking cases beyond routine T. pallidum genome sequencing. T. pallidum strains with deletions in tprK donor sites currently circulate and are associated with diminished TprK antigenic diversity.

Development of an accurate and rapid method for whole genome characterization of canine parvovirus.

Canine parvovirus is a highly contagious pathogen affecting domestic dogs and other carnivores globally. Monitoring CPV through continuous genomic surveillance is crucial for mapping variability and developing effective control measures. Here, we developed a method using multiplex-PCR-next-generation sequencing to obtain full-length CPV genomes directly from clinical samples. This approach utilizes tiling and tailed amplicons to amplify overlapping fragments of roughly 250 base pairs. This enables the creation of Illumina libraries by conducting two PCR reaction runs. We tested the assay in 10 fecal samples from dogs diagnosed with CPV and one CPV-2 vaccine strain. Furthermore, we applied it to a feline sample previously diagnosed with the feline panleukopenia virus. The assay provided 100 % genome coverage and high sequencing depth across all 12 samples. It successfully provided the sequence of the coding regions and the left and right non-translated regions, including tandem and terminal repeats. The assay effectively amplified viral variants from divergent evolutionary groups, including the antigenic variants (2a, 2b, and 2c) and the ancestral CPV-2 strain included in vaccine formulations. Moreover, it successfully amplified the entire genome of the feline panleukopenia virus found in cat feces. This method is cost-effective, time-efficient, and does not require lab expertise in Illumina library preparation. The multiplex-PCR-next-generation methodology facilitates large-scale genomic sequencing, expanding the limited number of complete genomes currently available in databases and enabling real-time genomic surveillance. Furthermore, the method helps identify and track emerging CPV viral variants, facilitating molecular epidemiology and control. Adopting this approach can enhance our understanding of the evolution and genetic diversity of Protoparvovirus carnivoran1.