Deciphering the core antiviral mechanism of Yinqiao powder: Inhibition of dengue virus adsorption mediated by wogonin via host receptor HSP90AA1 blockade.

Targeting dengue virus envelope protein: Insights from structural analysis and high-throughput screening.

A proximity induced strand displacement amplification and CRISPR/Cas12a-based SERS assay for ultrasensitive detection of dengue virus.

Mosquito-borne orthoflaviviruses, such as Japanese encephalitis virus (JEV), West Nile virus (WNV), dengue virus (DENV), and Zika virus (ZIKV), pose considerable public health challenges due to their zoonotic potential and the absence of specific therapeutics. Conventional detection methods exhibit critical limitations in multiplexing capability, prolonged turnaround time, and procedural complexity. These factors hinder rapid outbreak response and underscore the necessity for improved detection platforms. This study developed a quadruplex quantitative real-time PCR (qRT-PCR) assay utilizing Minor Groove Binder (MGB) probes for the simultaneous detection of JEV, WNV, DENV, and ZIKV. The MGB probe-based qPCR assay demonstrated high specificity in distinguishing these four orthoflaviviruses and exhibited significantly enhanced sensitivity compared to conventional TaqMan probes, achieving detection limit of 2 copies/reaction. This method was successfully applied to detect virus-positive samples in cell cultures, laboratory mosquitoes, and field-collected mosquitoes, thereby validating the practical utility of the MGB probe-based qPCR assay. Compared to conventional multiplex detection methods, our MGB-based approach provides a promising detection platform, thereby demonstrating its potential utility in the high-throughput surveillance of mosquito-borne orthoflaviviruses.

Dengue is caused by infection with 1 of 4 dengue virus (DENV) serotypes (DENV-1 to DENV-4). V181 and Butantan-Dengue Vaccine (Butantan-DV) are independently developed, single-dose, quadrivalent dengue vaccines based on the same attenuated parental strains. In this randomized, double-blind, active comparator-controlled, multisite, phase 2 trial, we compared the safety and immunogenicity of V181 and Butantan-DV in healthy adults in Brazil. The primary immunogenicity objectives were to compare vaccine-induced neutralizing antibody geometric mean titers (GMTs) and seroconversion rates for DENV-1 to DENV-4 at day 28 postvaccination. The primary and secondary safety objectives, respectively, were to assess vaccine-related serious adverse events and the proportion of participants experiencing solicited adverse events through day 28 postvaccination. Overall, 1364 participants were assigned (1:1) to receive V181 (n = 682) or Butantan-DV (n = 682). At day 28 postvaccination, V181 induced a noninferior immune response to Butantan-DV for DENV-1 and DENV-2 but not DENV-3 and DENV-4, as measured using GMT ratios. Noninferiority was established for all DENV serotypes based on seroconversion rates. Most participants developed a trivalent or tetravalent neutralizing antibody response. Vaccine viremia was detected for each DENV serotype. V181 and Butantan-DV were generally well tolerated. V181 was immunogenic in adults, and while immunogenicity was noninferior to Butantan-DV for DENV-1 and DENV-2, seroconversion rates were noninferior for DENV-1 to DENV-4. The safety profile of V181 is comparable to that of Butantan-DV and supports its continued evaluation as a single-dose quadrivalent vaccine for dengue prevention. NCT05710224.

Dengue fever is one of the most widely distributed vector-borne infectious diseases globally, prevalent in tropical and subtropical regions. An estimated 3.9 billion people in 128 countries are at risk of infection globally, and the 2024 outbreak was the most severe. As a tropical region, Hainan Province in China serves as a significant endemic area for dengue fever, where epidemic prevention and control remain critical. To analyze the transmission dynamics and pathogen variation characteristics of the 2024 dengue fever outbreak in Hainan Province. Acute-phase serum samples from 18 confirmed dengue cases during the 2024 outbreak in Hainan Province were collected and serotyped using TaqMan real-time PCR. The viral envelope (E) gene and representative whole genomes were sequenced and compared with domestic reference strains to assess epidemic trends. Phylogenetic and molecular clock analyses were performed based on E gene and full-genome sequences to infer spatiotemporal transmission dynamics. Sequence alignment and homology modeling were used to identify E gene mutations and evaluate their potential effects on E protein functional domains and viral pathogenicity. Among the 18 samples, 17 were dengue virus (DENV)-1, and 1 was DENV-3. Phylogenetic analysis revealed that the primary strains in this outbreak were highly homologous to those circulating in Guangdong Province; however, two strains exhibited significant genetic differences, suggesting possible independent introduction. Additionally, two unique DENV-1 E gene mutations (D147N and S338L) were identified, which we hypothesize may be associated with viral characteristics, subject to further functional verification. This study systematically reveals the epidemiological characteristics of the 2024 dengue fever outbreak and provides a scientific basis for formulating local control strategies and a research foundation for further exploring the molecular-level pathogenicity and transmissibility of DENV.

Seroprevalence of dengue virus infection in people residing in Vietnam: A systematic review and meta-analysis.

The burden of dengue disease in Germany is a growing public health concern. While travellers to dengue-endemic countries are at increased risk of contracting a dengue infection, data describing dengue-related healthcare utilization and outcomes in Germany are limited. This study investigated the healthcare burden of dengue disease in travellers returning to Germany. A retrospective analysis (InGef research database) was used to estimate dengue incidence using anonymized representative data from the German Statutory Health Insurance from approximately 7 million insured persons/year during 2015-23. Outpatient and inpatient healthcare use and dengue associated costs were investigated. From 2015 to 2023, among 10 151 240 insured persons, 887 dengue episodes were recorded. Patients had a median age of 33years (interquartile range 26-46) and 52% were male. Each year, 103-182 episodes were reported (COVID-19 pandemic period [2020-22]: 10-62). Excluding the pandemic years, incidence was 1.4-2.5/100 000 persons, with an estimated total of 1453-2639 cases/year in Germany. Of the 37% hospitalized persons, 4% required intensive care. Mean hospital stay was 3.3days; most patients were discharged after 1 overnight stay and no dengue related deaths were reported. Overall, 41% of patients took sick leave during their episode (mean sick leave for hospitalized patients: 8.8 working days); 38% of those not hospitalized took sick leave lasting 6.9 working days on average. Patients with dengue had 2-fold increased healthcare visits, resulting in a 162% increase in costs (mean costs/episode/quarter: 930€) compared with the pre-index period, with these costs mainly driven by outpatient care (23%) and hospitalization (64%). Hospitalized patients had a 4-fold rise in costs (quarterly costs/episode: 1700€; inpatient care accounting for 80%) compared with the pre-index period. Dengue virus infection in travellers returning to Germany impacted the healthcare system in Germany, leading to high costs, hospitalizations and physician visits and considerable sick leave.

Dengue virus (DENV) infection drives pathological inflammation through coordinated activation of pattern recognition receptors (PRRs) in myeloid cells, yet no approved immunomodulatory therapy exists to interrupt this process. CLEC5A and Toll-like receptor 2 (TLR2) have each been implicated in DENV-induced cytokine production, but whether their co-engagement represents a convergent and therapeutically targetable axis remains unclear. We define the cytokine landscape of human macrophages infected with all four DENV serotypes and reverse genetics DENV2 (rgDV2) strains carrying NS1 mutations derived from the severe 2015 Taiwan outbreak. We further demonstrate the potential therapeutic effect of bispecific antibodies simultaneously targeting CLEC5A and TLR2-engineered in both IgG1 and IgG4 formats. Human macrophages infected with all four DENV serotypes, as well as reverse genetics DENV2 (rgDV2), exhibit a consistently dominant pro-inflammatory program driven by CLEC5A-TLR2 co-signaling. Treatment with bispecific antibodies, in both IgG1 and IgG4 formats, potently suppress the production of TNF-α, IL-6, IL-8, and MCP-1 production across all viral strains tested. Notably, the IgG4 format preserves robust inhibitory efficacy while minimizing Fc receptor engagement, thereby offering a rational strategy to mitigate the risk of antibody-dependent enhancement (ADE). This dual-targeting approach also remains effective against NS1 mutant strains linked to enhanced virulence and cytokine storm. Together, these findings identify CLEC5A-TLR2 co-signaling as a convergent inflammatory axis in dengue pathogenesis and establish bispecific dual-receptor blockade as a mechanistically grounded, ADE-aware immunomodulatory strategy to mitigate cytokine-driven pathology.

Orthoflavivirus, such as West Nile Virus (WNV), dengue virus (DENV), and ZIKA virus (ZIKV), are globally distributed pathogens that pose substantial threats to human health. Currently, there are still no effective antiviral drugs for WNV or ZIKV. Despite the availability of two licensed DENV vaccines, their use remains limited due to potential risks, highlighting an urgent need for antiviral drug development. The highly conserved orthoflavivirus protease NS2B/NS3 is required for viral replication, making it a promising antiflavivirus target. A major challenge, however, is the highly charged active site of this enzyme, which requires charged chemical matters with low bioavailability. An alternative and more attractive strategy is to target potential allosteric sites or folding intermediate states of the protease. In this work, we employ the topology-based coarse-grained Go̅ modeling to explore the coupled binding and folding pathways of WNV NS2B/NS3 protease and study the effects of the widely used experimental construct with a G4SG4 linker between NS2B and NS3 on stability and folding. Our results provide a holistic conformational landscape of protease binding and folding, including several key intermediate states. We find that the presence of the G4SG4 linker alters the folding pathways and destabilizes the NS2B C-terminus. The latter is consistent with experimental observations that the G4SG4-linked protease has lower activity and adopts an open state without the substrate in crystal structures. Together, these findings provide, for the first time, a complete picture of the binding and folding of the NS2B/NS3 protease and identify important folding intermediate states that could be targeted for allosteric antiviral drug development.

Orthoflaviviruses are genetically related, yet cause distinct disease patterns ranging from hepatitis and vascular shock syndrome to encephalitis and congenital abnormalities. There is an incomplete understanding of the cellular pathways co-opted by orthoflaviviruses, and differences in host response to infection may underlie the diverse pathologies caused. We present a single-cell approach (Quantification of Infection and CRISPR guide sequencing; QIC-seq) that combines CRISPR/Cas9 knockout with virus-inclusive transcriptomics to systematically compare host factor requirements and host transcriptional response to orthoflaviviral challenge. Using a CRISPR library focused on select ER-proteostasis genes, we show that dengue and yellow fever viruses are strictly dependent on subunits of the oligosaccharyltransferase complex, while the more distantly related West Nile and Langat viruses are dependent on components of the ER-associated degradation machinery. Our data further shows virus-induced upregulation of interferon-stimulated genes, and activation of the unfolded protein response. QIC-seq enables quantitative comparisons of viral host factor utilization, which may inform development of host-directed antiviral therapies.

Airports are strategic targets for wastewater-based epidemiology because they concentrate highly mobile populations and may provide early signals of pathogen circulation. However, metagenomic investigations of airport wastewater remain limited, particularly in South America. Here, we present one of the first hybrid-capture target-enriched metagenomic investigations of airport wastewater in Brazil, integrating the detection of human-associated viruses and bacteriophage-derived host signatures to evaluate airports as sentinel surveillance sites. Seven untreated wastewater samples collected from a major Brazilian airport between December 2021 and March 2023 were concentrated, subjected to nucleic acid extraction, and analyzed using hybrid-capture target-enriched next-generation sequencing. Taxonomic analysis identified 615 viral and bacteriophage-associated taxa, including 440 viruses and 175 bacteriophages. Among the viral fraction, 21 human-associated viral taxa representing eight viral families were selected for detailed analysis. Norovirus GII was detected in all samples, while Mamastrovirus 1 and JC polyomavirus were detected in six of seven samples. SARS-CoV-2 and dengue virus type 1 were simultaneously detected in the March, 2023 sample. The bacteriophage fraction comprised 47 host-associated phage groups, with Streptococcus-associated phages predominating across samples. These findings demonstrate that airport wastewater can capture diverse human viral and bacteriophage-derived signatures associated with population mobility, supporting its application in environmental genomic surveillance and early-warning systems for emerging and circulating pathogens.

The bioactive constituents of ginger (Zingiber officinale Roscoe) display well-documented immunomodulatory and antimicrobial effects; however, the role of these components in combating dengue virus (DENV) and other flaviviruses infection remains unknown. Here, activity-guided molecular networking and antiviral screening identified 10-shogaol as the primary antiviral phytochemical in ginger, with a half-maximal inhibitory concentration (IC50) value of 2.502 μM. Mechanistically, 10-shogaol bound the n-octyl-β-d-glucopyranoside (β-OG) pocket of viral envelope protein (E), interfering with viral attachment and conferring broad antiviral properties against DENV and Zika virus (ZIKV). Additionally, 10-shogaol suppressed proinflammatory cytokines and mitigated tissue damage in mice by acting as a novel Nrf2 activator by inhibiting Keap1-mediated ubiquitination and proteasome degradation. Activation of Nrf2 by 10-shogaol induced antioxidant gene expression and inhibited inflammatory NF-κB signaling. Collectively, our study suggests that 10-shogaol acts as a broad-spectrum entry inhibitor against DENV and ZIKV as well as confers anti-inflammatory benefits for mitigating virus-induced inflammatory tissue damage.

Dengue virus (DENV) and Chikungunya virus (CHIKV) are significant arboviruses transmitted by Aedes mosquitoes. The concurrent circulation of these viruses complicates syndromic diagnosis and subsequently, the clinical management in endemic settings like India. The present study evaluated the prevalence and clinical features of dengue immunoglobulin M (IgM) and dual IgM seropositive patients (dengue and chikungunya) among a subset of patients presenting with clinical suspicion of dengue fever admitted in tertiary care settings in Belagavi, Northwestern Karnataka, India. A retrospective observational study was conducted using serum samples received from tertiary care hospitals in Belagavi, Karnataka between January 2024 to October 2025. Of 151 clinically suspected dengue cases, 94 patients with complete clinical records and available serum samples were included. Dengue virus (DENV) and chikungunya virus (CHIKV) IgM antibodies were detected using Indian Council of Medical Research - National Institute of Virology (ICMR-NIV) Immunoglobulin M Antibody Capture (MAC) Enzyme Linked Immunosorbent Assay (ELISA) kits. The reported sensitivity and specificity were 98.53% and 98.84% for DENV IgM ELISA and 95% and 98% for CHIKV IgM ELISA, respectively. Dual IgM seropositivity to dengue and chikungunya was observed in 19 patients (20.2%); whereas 73 (77.7%) were positive only for dengue IgM and 2 (2.1%) were negative for both assays. No isolated CHIKV IgM positivity was detected. Dual-positive patients exhibited predominantly dengue-like illness, characterized by myalgia (57.9%), thrombocytopenia (median platelet count, 70,000 cells/mm3) and elevated aspartate transaminase (AST) levels (median AST, 67 U/L). Dual IgM seropositivity for dengue and chikungunya was observed in 20.2% of suspected dengue cases. While IgM serology cannot confirm concurrent infection, these findings suggest co-circulation of both arboviruses and highlight the need for integrated arboviral surveillance in endemic regions.

Dengue remains a major public health challenge in tropical regions, where viral genetic diversity, vector ecology and human mobility contribute to recurrent outbreaks. In 2024, Brazil experienced its largest recorded dengue epidemic, with intense transmission across the Northeast region. This study characterizes the epidemiological, genomic and phylogeographic features of dengue virus (DENV) circulation during the 2024 outbreak in Alagoas, northeastern Brazil. Of 6102 suspected cases analysed by molecular diagnostics, 1607 (26.3%) were confirmed as DENV infections. Epidemiological analyses revealed widespread transmission across 79 municipalities, marked seasonality with peak incidence between March and August, and a disproportionate burden among adolescents and young adults. DENV-1 and DENV-2 were the predominant serotypes, accounting for 57.1% and 42.6% of confirmed cases, respectively; three DENV-3 infections and one DENV-1/DENV-2 co-infection were also identified. Whole-genome sequencing yielded high-quality genomic data supporting phylogenetic and phylogeographic analyses. DENV-1 sequences belonged to genotype V, classified into lineages E.1 and D.1.1, while DENV-2 sequences clustered within genotype II (lineage F.1.1.2) and genotype III (lineage C.1.1). Time-scaled phylogenetic analyses revealed multiple independent introductions of both serotypes from distinct Brazilian regions, followed by sustained local transmission and lineage expansion. Infections among non-residents, including an international case, further underscore the role of population mobility in viral dissemination. Collectively, these findings indicate that the 2024 outbreak was driven by recurrent viral introductions combined with the persistence of locally established transmission chains, highlighting the value of integrated genomic surveillance for outbreak preparedness and public health response.

Mathematical model of cellular and humoral immune responses of dengue virus infection with hematocrit dynamics.

Dengue fever, a mosquito-borne disease caused by dengue virus (DENV), has become a global health problem, and no FDA-approved drug is currently available. Qingwen Baidu Decoction (QBD) is used to treat the critical phase of dengue fever in China, but its mechanism of action remains unclear. In this work, we integrated bioinformatics analysis, machine learning, and network pharmacology to investigate the possible molecular targets and potential active chemical components of QBD. Common targets between differentially expressed genes from DENV infected samples and predicted targets of QBD were identified by bioinformatics analysis and refined by machine learning algorithms including LASSO, random forest and SVM-RFE. Three core genes, CXCL10, EZH2 and EPHB2 were significantly overexpressed in dengue fever patients, indicating their potential diagnostic and therapeutic value. Single cell transcriptome analysis further revealed that QBD primarily targets dendritic cells, monocytes and macrophages. Immune infiltration analysis using ssGSEA showed that these three core genes were significantly associated with CD4+ and CD8+ T cell subtypes, suggesting their involvement in host immune regulation. Molecular docking and molecular dynamics simulations identified eight chemical components of QBD as potential active ingredients. Based on these computational predictions, we hypothesize that QBD may exert its therapeutic effects through dual mechanisms, including directly binding to DENV proteins to inhibit viral replication, while also regulating the function of CXCL10 and EZH2 to alleviate DENV-induced inflammatory responses and modulate host immunity. These results provide a theoretical reference for future experimental validation and drug development.

Aedes aegypti is the principal vector of several medically important arboviruses, including dengue virus (DENV) and chikungunya virus (CHIKV), both of which remain endemic in Thailand. However, studies on arbovirus surveillance and mitochondrial genetic diversity of Ae. aegypti populations across broad geographic scales remain limited. This study aimed to assess the detection ofDENV and CHIKV andcharacterize the mitochondrial genetic diversity of Ae. aegypti across multiple regions of Thailand. A total of 303 adult Ae. aegypti mosquitoes collected from eight provinces across five geographic regions of Thailand were screened for DENV and CHIKV RNA. Mitochondrial cytochrome c oxidase subunit I (COI) genetic diversity was assessed using haplotype network analysis and analysis of molecular variance (AMOVA). CHIKV RNA was detected in 51 mosquitoes (16.8%) from four provinces, with the highest detection rates observed in Prachuap Khiri Khan and Bangkok. DENV RNA was not detected in any sample. COI analysis revealed moderate to high mitochondrial haplotype diversity and geographic variation in haplotype distribution among populations. Both widely shared and province-specific haplotypes were identified. AMOVA indicated that most genetic variation occurred within populations (93.19%), with significantgenetic differentiation among populations (ΦST = 0.068, P < 0.001). CHIKV RNA was detected in Ae. aegypti populations from multiple regions of Thailand, whereas DENV RNA was not detected. Mitochondrial COI analysis revealed substantial haplotype diversity and geographic variation among populations. These findings contribute to current knowledge of arbovirus occurrence and mosquito genetic diversity in Thailand and provide a foundation for future vector surveillance studies.

Dengue virus (DENV) continues to pose a significant global health threat, with increasing infection rates and limited treatment options. The viral NS2B-NS3 protease (NS2B-NS3pro), a highly conserved two-component enzyme essential for polyprotein processing and replication, is a key target for antiviral drug development. Here, we report the 2.25 Å-resolution crystal structure of DENV serotype 2 (DENV-2) NS2B-NS3pro, in which a PEG fragment is bound within a solvent-exposed pocket between β10, β11, β14 and β15 of NS3. This structure reveals a previously unrecognized solvent-exposed PEG-binding pocket, which may represent a putative ligand-binding surface for future validation and inhibitor-design studies. We also show that the glutathione-coated gold nanocluster (GSH-AuNC) directly inhibits DENV-2 NS2B-NS3pro. Bio-layer interferometry and fluorescence-based protease assays indicate that the nanocluster binds NS2B-NS3pro with a dissociation constant of 15.64 µM and inhibits its catalytic activity with an IC50 of 16.04 µM, consistent with a direct inhibitory effect at the in vitro enzymatic level. Molecular dynamics simulations further suggest that GSH-AuNC is predicted to interact with the catalytic triad of NS2B-NS3pro, forming stable electrostatic and van der Waals interactions that may interfere with substrate access. Collectively, these findings provide an in vitro structural and biochemical basis for targeting DENV-2 NS2B-NS3pro and may inform future development of small-molecule or nanomedicine-based inhibitors, although cell-based antiviral efficacy, cytotoxicity, and cellular uptake remain to be evaluated.

Ambient PM2.5 exposure alters mosquito feeding and reproductive traits during biomass-burning haze.