In silico design of peptide inhibitors for Dengue virus to treat Dengue virus-associated infections.

Dengue virus is a single positive-strand RNA virus that is composed of three structural proteins including capsid, envelope, and precursor membrane while seven non-structural proteins (NS1, NS2A, NS2B, NS3A, NS3B, NS4, and NS5). Dengue is a viral infection caused by the dengue virus (DENV). DENV infections are asymptomatic or produce only mild illness. However, DENV can occasionally cause more severe cases and even death. There is no specific treatment for dengue virus infections. Therapeutic peptides have several important advantages over proteins or antibodies: they are small in size, easy to synthesize, and have the ability to penetrate the cell membranes. They also have high activity, specificity, affinity, and less toxicity. Based on the known peptide inhibitor, the current study designs peptide inhibitors for dengue virus envelope protein using an alanine and residue scanning technique. By replacing I21 with Q21, L14 with H14, and V28 with K28, the binding affinity of the peptide inhibitors was increased. The newly designed peptide inhibitors with single residue mutation improved the binding affinity of the peptide inhibitors. The inhibitory capability of the new promising peptide inhibitors was further confirmed by the utilization of MD simulation and free binding energy calculations. The molecular dynamics simulation demonstrated that the newly engineered peptide inhibitors exhibited greater stability compared to the wild-type peptide inhibitors. According to the binding free energies MM(GB)SA of these developed peptides, the first peptide inhibitor was the most effective against the dengue virus envelope protein. All peptide derivatives had higher binding affinities for the envelope protein and have the potential to treat dengue virus-associated infections. In this study, new peptide inhibitors were developed for the dengue virus envelope protein based on the already reported peptide inhibitor.

Antiviral activity of pimecrolimus against dengue virus type 2 infection in vitro and in vivo.

Dengue virus (DENV) infection is a public health concern in several countries and is associated with severe diseases, such as dengue hemorrhagic fever and dengue shock syndrome. DENVs are transmitted to humans via the bites of infected Aedes mosquitoes, and no antiviral therapeutics are currently available. In this work, we aimed to identify antiviral drugs against DENV type 2 (DENV2) infections and selected pimecrolimus as a potential antiviral drug candidate. Pimecrolimus significantly inhibited DENV2-mediated cell death and replication in vitro. We also confirmed a decrease in the number of plaques formed as well as in the envelope protein levels of DENV2. The time-of-addition and course experiments revealed that pimecrolimus inhibited DENV2 infection during the early stages of the virus replication cycle. In an experimental mouse model, orally administered pimecrolimus alleviated body weight loss and lethality caused by DENV2 infection, which we used as readouts of the drug's antiviral potency. Furthermore, pimecrolimus significantly inhibited the DENV2 load and ameliorated focal necrosis in the liver and spleen. Taken together, our in vitro and in vivo findings suggest that pimecrolimus is a promising antiviral drug candidate for the treatment of DENV2 infection.

Reduced protein-coding transcript diversity in severe dengue emphasises the role of alternative splicing.

Dengue fever, a neglected tropical arboviral disease, has emerged as a global health concern in the past decade. Necessitating a nuanced comprehension of the intricate dynamics of host-virus interactions influencing disease severity, we analysed transcriptomic patterns using bulk RNA-seq from 112 age- and gender-matched NS1 antigen-confirmed hospital-admitted dengue patients with varying severity. Severe cases exhibited reduced platelet count, increased lymphocytosis, and neutropenia, indicating a dysregulated immune response. Using bulk RNA-seq, our analysis revealed a minimal overlap between the differentially expressed gene and transcript isoform, with a distinct expression pattern across the disease severity. Severe patients showed enrichment in retained intron and nonsense-mediated decay transcript biotypes, suggesting altered splicing efficiency. Furthermore, an up-regulated programmed cell death, a haemolytic response, and an impaired interferon and antiviral response at the transcript level were observed. We also identified the potential involvement of the RBM39 gene among others in the innate immune response during dengue viral pathogenesis, warranting further investigation. These findings provide valuable insights into potential therapeutic targets, underscoring the importance of exploring transcriptomic landscapes between different disease sub-phenotypes in infectious diseases.

Phylodynamics unveils invading and diffusing patterns of dengue virus serotype-1 in Guangdong, China from 1990 to 2019 under a global genotyping framework.

BACKGROUND: The strong invasiveness and rapid expansion of dengue virus (DENV) pose a great challenge to global public health. However, dengue epidemic patterns and mechanisms at a genetic scale, particularly in term of cross-border transmissions, remain poorly understood. Importation is considered as the primary driver of dengue outbreaks in China, and since 1990 a frequent occurrence of large outbreaks has been triggered by the imported cases and subsequently spread to the western and northern parts of China. Therefore, this study aims to systematically reveal the invasion and diffusion patterns of DENV-1 in Guangdong, China from 1990 to 2019. METHODS: These analyses were performed on 179 newly assembled genomes from indigenous dengue cases in Guangdong, China and 5152 E gene complete sequences recorded in Chinese mainland. The genetic population structure and epidemic patterns of DENV-1 circulating in Chinese mainland were characterized by phylogenetics, phylogeography, phylodynamics based on DENV-1 E-gene-based globally unified genotyping framework. RESULTS: Multiple serotypes of DENV were co-circulating in Chinese mainland, particularly in Guangdong and Yunnan provinces. A total of 189 transmission clusters in 38 clades belonging to 22 subgenotypes of genotype I, IV and V of DENV-1 were identified, with 7 Clades of Concern (COCs) responsible for the large outbreaks since 1990. The epidemic periodicity was inferred from the data to be approximately 3 years. Dengue transmission events mainly occurred from Great Mekong Subregion-China (GMS-China), Southeast Asia (SEA), South Asia Subcontinent (SASC), and Oceania (OCE) to coastal and land border cities respectively in southeastern and southwestern China. Specially, Guangzhou was found to be the most dominant receipting hub, where DENV-1 diffused to other cities within the province and even other parts of the country. Genome phylogeny combined with epidemiological investigation demonstrated a clear local consecutive transmission process of a 5C1 transmission cluster (5C1-CN4) of DENV-1 in Guangzhou from 2013 to 2015, while the two provinces of Guangdong and Yunnan played key roles in ongoing transition of dengue epidemic patterns. In contextualizing within Invasion Biology theories, we have proposed a derived three-stage model encompassing the stages of invasion, colonization, and dissemination, which is supposed to enhance our understanding of dengue spreading patterns. CONCLUSIONS: This study demonstrates the invasion and diffusion process of DENV-1 in Chinese mainland within a global genotyping framework, characterizing the genetic diversities of viral populations, multiple sources of importation, and periodic dynamics of the epidemic. These findings highlight the potential ongoing transition trends from epidemic to endemic status offering a valuable insight into early warning, prevention and control of rapid spreading of dengue both in China and worldwide.

Dengue virus exploits autophagy vesicles and secretory pathways to promote transmission by human dendritic cells.

Dengue virus (DENV), transmitted by infected mosquitoes, is a major public health concern, with approximately half the world's population at risk for infection. Recent decades have increasing incidence of dengue-associated disease alongside growing frequency of outbreaks. Although promising progress has been made in anti-DENV immunizations, post-infection treatment remains limited to non-specific supportive treatments. Development of antiviral therapeutics is thus required to limit DENV dissemination in humans and to help control the severity of outbreaks. Dendritic cells (DCs) are amongst the first cells to encounter DENV upon injection into the human skin mucosa, and thereafter promote systemic viral dissemination to additional human target cells. Autophagy is a vesicle trafficking pathway involving the formation of cytosolic autophagosomes, and recent reports have highlighted the extensive manipulation of autophagy by flaviviruses, including DENV, for viral replication. However, the temporal profiling and function of autophagy activity in DENV infection and transmission by human primary DCs remains poorly understood. Herein, we demonstrate that mechanisms of autophagosome formation and extracellular vesicle (EV) release have a pro-viral role in DC-mediated DENV transmission. We show that DENV exploits early-stage canonical autophagy to establish infection in primary human DCs. DENV replication enhanced autophagosome formation in primary human DCs, and intrinsically-heightened autophagosome biogenesis correlated with relatively higher rates of DC susceptibility to DENV. Furthermore, our data suggest that viral replication intermediates co-localize with autophagosomes, while productive DENV infection introduces a block at the late degradative stages of autophagy in infected DCs but not in uninfected bystander cells. Notably, we identify for the first time that approximately one-fourth of DC-derived CD9/CD81/CD63+ EVs co-express canonical autophagy marker LC3, and demonstrate that DC-derived EV populations are an alternative, cell-free mechanism by which DCs promote DENV transmission to additional target sites. Taken together, our study highlights intersections between autophagy and secretory pathways during viral infection, and puts forward autophagosome accumulation and viral RNA-laden EVs as host determinants of DC-mediated DENV infection in humans. Host-directed therapeutics targeting autophagy and exocytosis pathways thus have potential to enhance DC-driven resistance to DENV acquisition and thereby limit viral dissemination by initial human target cells following mosquito-to-human transmission of DENV.

Serotype and genomic diversity of dengue virus during the 2023 outbreak in Pakistan reveals the circulation of genotype III of DENV-1 and cosmopolitan genotype of DENV-2.

Dengue, a mosquito-borne viral disease, poses a significant public health challenge in Pakistan, with a significant outbreak in 2023, prompting our investigation into the serotype and genomic diversity of the dengue virus (DENV). NS-1 positive blood samples from 153 patients were referred to the National Institute of Health, Pakistan, between July and October 2023. Among these, 98 (64.1%) tested positive using multiplex real-time PCR, with higher prevalence among males (65.8%) and individuals aged 31-40. Serotyping revealed DENV-1 as the predominant serotype (84.7%), followed by DENV-2 (15.3%). Whole-genome sequencing of 18 samples (DENV-1 = 17, DENV-2 = 01) showed that DENV-1 (genotype III) samples were closely related (>99%) to Pakistan outbreak samples (2022), and approx. > 98% with USA (2022), Singapore and China (2016), Bangladesh (2017), and Pakistan (2019). The DENV-2 sequence (cosmopolitan genotype; clade IVA) shared genetic similarity with Pakistan outbreak sequences (2022), approx. > 99% with China and Singapore (2018-2019) and showed divergence from Pakistan sequences (2008-2013). No coinfection with dengue serotypes or other viruses were observed. Comparisons with previous DENV-1 sequences highlighted genetic variations affecting viral replication efficiency (NS2B:K55R) and infectivity (E:M272T). These findings contribute to dengue epidemiology understanding and underscore the importance of ongoing genomic surveillance for future outbreak responses in Pakistan.

Potential of MALDI-TOF MS biotyping to detect deltamethrin resistance in the dengue vector Aedes aegypti.

Insecticide resistance in mosquitoes is spreading worldwide and represents a growing threat to vector control. Insecticide resistance is caused by different mechanisms including higher metabolic detoxication, target-site modification, reduced penetration and behavioral changes that are not easily detectable with simple diagnostic methods. Indeed, most molecular resistance diagnostic tools are costly and labor intensive and then difficult to use for routine monitoring of insecticide resistance. The present study aims to determine whether mosquito susceptibility status against the pyrethroid insecticides (mostly used for mosquito control) could be established by the protein signatures of legs and/or thoraxes submitted to MALDI-TOF Mass Spectrometry (MS). The quality of MS spectra for both body parts was controlled to avoid any bias due to unconformity protein profiling. The comparison of MS profiles from three inbreeds Ae. aegypti lines from French Guiana (IRF, IR03, IR13), with distinct deltamethrin resistance genotype / phenotype and the susceptible reference laboratory line BORA (French Polynesia), showed different protein signatures. On both body parts, the analysis of whole protein profiles revealed a singularity of BORA line compared to the three inbreeding lines from French Guiana origin, suggesting that the first criteria of differentiation is the geographical origin and/or the breeding history rather than the insecticide susceptibility profile. However, a deeper analysis of the protein profiles allowed to identify 10 and 11 discriminating peaks from leg and thorax spectra, respectively. Among them, a specific peak around 4870 Da was detected in legs and thoraxes of pyrethroid resistant lines compared to the susceptible counterparts hence suggesting that MS profiling may be promising to rapidly distinguish resistant and susceptible phenotypes. Further work is needed to confirm the nature of this peak as a deltamethrin resistant marker and to validate the routine use of MS profiling to track insecticide resistance in Ae. aegypti field populations.

DengueSeq: a pan-serotype whole genome amplicon sequencing protocol for dengue virus.

BACKGROUND: The increasing burden of dengue virus on public health due to more explosive and frequent outbreaks highlights the need for improved surveillance and control. Genomic surveillance of dengue virus not only provides important insights into the emergence and spread of genetically diverse serotypes and genotypes, but it is also critical to monitor the effectiveness of newly implemented control strategies. Here, we present DengueSeq, an amplicon sequencing protocol, which enables whole-genome sequencing of all four dengue virus serotypes. RESULTS: We developed primer schemes for the four dengue virus serotypes, which can be combined into a pan-serotype approach. We validated both approaches using genetically diverse virus stocks and clinical specimens that contained a range of virus copies. High genome coverage (>95%) was achieved for all genotypes, except DENV2 (genotype VI) and DENV 4 (genotype IV) sylvatics, with similar performance of the serotype-specific and pan-serotype approaches. The limit of detection to reach 70% coverage was 10-100 RNA copies/µL for all four serotypes, which is similar to other commonly used primer schemes. DengueSeq facilitates the sequencing of samples without known serotypes, allows the detection of multiple serotypes in the same sample, and can be used with a variety of library prep kits and sequencing instruments. CONCLUSIONS: DengueSeq was systematically evaluated with virus stocks and clinical specimens spanning the genetic diversity within each of the four dengue virus serotypes. The primer schemes can be plugged into existing amplicon sequencing workflows to facilitate the global need for expanded dengue virus genomic surveillance.

The alteration of NK cells phenotypes related to the functions and dengue disease outcomes.

Natural killer cells (NK cells) are the front line of immune cells to combat pathogens and able to influence the subsequent adaptive immune responses. One of the factors contributing to pathogenesis in dengue hemorrhagic fever (DHF) disease is aberrant immune activation during early phase of infection. This study explored the profile of NK cells in dengue infected pediatric patients with different degrees of disease severity. DHF patients contained higher frequency of activated NK cells but lower ratio of CD56dim:CD56bright NK subsets. Activated NK cells exhibited alterations in several NK receptors. Interestingly, the frequencies of NKp30 expressing activated NK cells were more pronounced in dengue fever (DF) than in DHF pediatric patients. In vitro functional analysis indicated that degranulation of NK cells in responding to dengue infected dendritic cells (DCs) required cell-cell contact and type I IFNs. Meanwhile, Interferon gamma (IFN-γ) production initially required cell-cell contact and type I IFNs followed by Interleukin-12 (IL-12), Interleukin-15 (IL-15) and Interleukin-18 (IL-18) resulting in the amplification of IFN-γ producing NK cells over time. This study highlighted the complexity and the factors influencing NK cells responses to dengue virus. Degree of activation, phenotypes of activated cells and the crosstalk between NK cells and other immune cells, could modulate the outcome of NK cells function in the dengue disease.

Dynamics of emergence and genetic diversity of dengue virus in Reunion Island from 2012 to 2022.

BACKGROUND: Dengue is a major public health concern in Reunion Island, marked by recurrent epidemics, including successive outbreaks of dengue virus serotypes 1 and 2 (DENV1 and DENV2) with over 70,000 cases confirmed since 2017. METHODOLOGY/PRINCIPAL FINDINGS: In this study, we used Oxford Nanopore NGS technology for sequencing virologically-confirmed samples and clinical isolates collected between 2012 and 2022 to investigate the molecular epidemiology and evolution of DENV in Reunion Island. Here, we generated and analyzed a total of 499 DENV1, 360 DENV2, and 18 DENV3 sequences. By phylogenetic analysis, we show that different genotypes and variants of DENV have circulated in the past decade that likely originated from Seychelles, Mayotte and Southeast Asia and highly affected areas in Asia and Africa. CONCLUSIONS/SIGNIFICANCE: DENV sequences from Reunion Island exhibit a high genetic diversity which suggests regular introductions of new viral lineages from various Indian Ocean islands. The insights from our phylogenetic analysis may inform local health authorities about the endemicity of DENV variants circulating in Reunion Island and may improve dengue management and surveillance. This work emphasizes the importance of strong local coordination and collaboration to inform public health stakeholders in Reunion Island, neighboring areas, and mainland France.

Introduction of New Dengue Virus Lineages of Multiple Serotypes after COVID-19 Pandemic, Nicaragua, 2022.

Major dengue epidemics throughout Nicaragua's history have been dominated by 1 of 4 dengue virus serotypes (DENV-1-4). To examine serotypes during the dengue epidemic in Nicaragua in 2022, we performed real-time genomic surveillance in-country and documented cocirculation of all 4 serotypes. We observed a shift toward co-dominance of DENV-1 and DENV-4 over previously dominant DENV-2. By analyzing 135 new full-length DENV sequences, we found that introductions underlay the resurgence: DENV-1 clustered with viruses from Ecuador in 2014 rather than those previously seen in Nicaragua; DENV-3, which last circulated locally in 2014, grouped instead with Southeast Asia strains expanding into Florida and Cuba in 2022; and new DENV-4 strains clustered within a South America lineage spreading to Florida in 2022. In contrast, DENV-2 persisted from the formerly dominant Nicaragua clade. We posit that the resurgence emerged from travel after the COVID-19 pandemic and that the resultant intensifying hyperendemicity could affect future dengue immunity and severity.

CRISPR Screen Reveals PACT as a Pro-Viral Factor for Dengue Viral Replication.

The dengue virus is a single-stranded, positive-sense RNA virus that infects ~400 million people worldwide. Currently, there are no approved antivirals available. CRISPR-based screening methods have greatly accelerated the discovery of host factors that are essential for DENV infection and that can be targeted in host-directed antiviral interventions. In the present study, we performed a focused CRISPR (Clustered Regularly Interspaced Palindromic Repeats) library screen to discover the key host factors that are essential for DENV infection in human Huh7 cells and identified the Protein Activator of Interferon-Induced Protein Kinase (PACT) as a novel pro-viral factor for DENV. PACT is a double-stranded RNA-binding protein generally known to activate antiviral responses in virus-infected cells and block viral replication. However, in our studies, we observed that PACT plays a pro-viral role in DENV infection and specifically promotes viral RNA replication. Knockout of PACT resulted in a significant decrease in DENV RNA and protein abundances in infected cells, which was rescued upon ectopic expression of full-length PACT. An analysis of global gene expression changes indicated that several ER-associated pro-viral genes such as ERN1, DDIT3, HERPUD1, and EIF2AK3 are not upregulated in DENV-infected PACT knockout cells as compared to infected wildtype cells. Thus, our study demonstrates a novel role for PACT in promoting DENV replication, possibly through modulating the expression of ER-associated pro-viral genes.

The Dynamic Relationship between Dengue Virus and the Human Cutaneous Innate Immune Response.

Dengue virus (DENV) is a continuing global threat that puts half of the world's population at risk for infection. This mosquito-transmitted virus is endemic in over 100 countries. When a mosquito takes a bloodmeal, virus is deposited into the epidermal and dermal layers of human skin, infecting a variety of permissive cells, including keratinocytes, Langerhans cells, macrophages, dermal dendritic cells, fibroblasts, and mast cells. In response to infection, the skin deploys an array of defense mechanisms to inhibit viral replication and prevent dissemination. Antimicrobial peptides, pattern recognition receptors, and cytokines induce a signaling cascade to increase transcription and translation of pro-inflammatory and antiviral genes. Paradoxically, this inflammatory environment recruits skin-resident mononuclear cells that become infected and migrate out of the skin, spreading virus throughout the host. The details of the viral-host interactions in the cutaneous microenvironment remain unclear, partly due to the limited body of research focusing on DENV in human skin. This review will summarize the functional role of human skin, the cutaneous innate immune response to DENV, the contribution of the arthropod vector, and the models used to study DENV interactions in the cutaneous environment.

Regional Variation of the CD4 and CD8 T Cell Epitopes Conserved in Circulating Dengue Viruses and Shared with Potential Vaccine Candidates.

As dengue expands globally and many vaccines are under trials, there is a growing recognition of the need for assessing T cell immunity in addition to assessing the functions of neutralizing antibodies during these endeavors. While several dengue-specific experimentally validated T cell epitopes are known, less is understood about which of these epitopes are conserved among circulating dengue viruses and also shared by potential vaccine candidates. As India emerges as the epicenter of the dengue disease burden and vaccine trials commence in this region, we have here aligned known dengue specific T cell epitopes, reported from other parts of the world with published polyprotein sequences of 107 dengue virus isolates available from India. Of the 1305 CD4 and 584 CD8 epitopes, we found that 24% and 41%, respectively, were conserved universally, whereas 27% and 13% were absent in any viral isolates. With these data, we catalogued epitopes conserved in circulating dengue viruses from India and matched them with each of the six vaccine candidates under consideration (TV003, TDEN, DPIV, CYD-TDV, DENVax and TVDV). Similar analyses with viruses from Thailand, Brazil and Mexico revealed regional overlaps and variations in these patterns. Thus, our study provides detailed and nuanced insights into regional variation that should be considered for itemization of T cell responses during dengue natural infection and vaccine design, testing and evaluation.

Toward a Categorization of Virus-ncRNA Interactions in the World of RNA to Disentangle the Tiny Secrets of Dengue Virus.

In recent years, the function of noncoding RNAs (ncRNAs) as regulatory molecules of cell physiology has begun to be better understood. Advances in viral molecular biology have shown that host ncRNAs, cellular factors, and virus-derived ncRNAs and their interplay are strongly disturbed during viral infections. Nevertheless, the folding of RNA virus genomes has also been identified as a critical factor in regulating canonical and non-canonical functions. Due to the influence of host ncRNAs and the structure of RNA viral genomes, complex molecular and cellular processes in infections are modulated. We propose three main categories to organize the current information about RNA-RNA interactions in some well-known human viruses. The first category shows examples of host ncRNAs associated with the immune response triggered in viral infections. Even though miRNAs introduce a standpoint, they are briefly presented to keep researchers moving forward in uncovering other RNAs. The second category outlines interactions between virus-host ncRNAs, while the third describes how the structure of the RNA viral genome serves as a scaffold for processing virus-derived RNAs. Our grouping may provide a comprehensive framework to classify ncRNA-host-cell interactions for emerging viruses and diseases. In this sense, we introduced them to organize DENV-host-cell interactions.

Dengue Virus and Zika Virus Seroprevalence in the South Pacific Populations of the Cook Islands and Vanuatu.

Arboviral diseases are serious threats to global health with increasing prevalence and potentially severe complications. Significant arthropod-borne viruses are the dengue viruses (DENV 1-4), the Zika virus (ZIKV), and the chikungunya virus (CHIKV). Among the areas most affected is the South Pacific Region (SPR). Here, arboviruses not only cause a high local burden of disease, but the region has also proven to contribute to their global spread. Outpatient serum samples collected between 08/2016 and 04/2017 on three islands of the island states of Vanuatu and the Cook Islands were tested for anti-DENV- and anti-ZIKV-specific antibodies (IgG) using enzyme-linked immunosorbent assays (ELISA). ELISA test results showed 89% of all test sera from the Cook Islands and 85% of the Vanuatu samples to be positive for anti-DENV-specific antibodies. Anti-ZIKV antibodies were identified in 66% and 52%, respectively, of the test populations. Statistically significant differences in standardized immunity levels were found only at the intranational level. Our results show that in both the Cook Islands and Vanuatu, residents were exposed to significant Flavivirus transmission. Compared to other seroprevalence studies, the marked difference between ZIKV immunity levels and previously published CHIKV seroprevalence rates in our study populations is surprising. We propose the timing of ZIKV and CHIKV emergence in relation to recurrent DENV outbreaks and the impact of seasonality as explanatory external factors for this observation. Our data add to the knowledge of arboviral epidemics in the SPR and contribute to a better understanding of virus spread, including external conditions with potential influence on outbreak dynamics. These data may support preventive and rapid response measures in the affected areas, travel-related risk assessment, and infection identification in locals and returning travelers.

Secreted dengue virus NS1 from infection is predominantly dimeric and in complex with high-density lipoprotein.

Severe dengue infections are characterized by endothelial dysfunction shown to be associated with the secreted nonstructural protein 1 (sNS1), making it an attractive vaccine antigen and biotherapeutic target. To uncover the biologically relevant structure of sNS1, we obtained infection-derived sNS1 (isNS1) from dengue virus (DENV)-infected Vero cells through immunoaffinity purification instead of recombinant sNS1 (rsNS1) overexpressed in insect or mammalian cell lines. We found that isNS1 appeared as an approximately 250 kDa complex of NS1 and ApoA1 and further determined the cryoEM structures of isNS1 and its complex with a monoclonal antibody/Fab. Indeed, we found that the major species of isNS1 is a complex of the NS1 dimer partially embedded in a high-density lipoprotein (HDL) particle. Crosslinking mass spectrometry studies confirmed that the isNS1 interacts with the major HDL component ApoA1 through interactions that map to the NS1 wing and hydrophobic domains. Furthermore, our studies demonstrated that the sNS1 in sera from DENV-infected mice and a human patient form a similar complex as isNS1. Our results report the molecular architecture of a biological form of sNS1, which may have implications for the molecular pathogenesis of dengue.

Human movement and environmental barriers shape the emergence of dengue.

Understanding how emerging infectious diseases spread within and between countries is essential to contain future pandemics. Spread to new areas requires connectivity between one or more sources and a suitable local environment, but how these two factors interact at different stages of disease emergence remains largely unknown. Further, no analytical framework exists to examine their roles. Here we develop a dynamic modelling approach for infectious diseases that explicitly models both connectivity via human movement and environmental suitability interactions. We apply it to better understand recently observed (1995-2019) patterns as well as predict past unobserved (1983-2000) and future (2020-2039) spread of dengue in Mexico and Brazil. We find that these models can accurately reconstruct long-term spread pathways, determine historical origins, and identify specific routes of invasion. We find early dengue invasion is more heavily influenced by environmental factors, resulting in patchy non-contiguous spread, while short and long-distance connectivity becomes more important in later stages. Our results have immediate practical applications for forecasting and containing the spread of dengue and emergence of new serotypes. Given current and future trends in human mobility, climate, and zoonotic spillover, understanding the interplay between connectivity and environmental suitability will be increasingly necessary to contain emerging and re-emerging pathogens.

Diagnostic performance of the bioline dengue duo rapid test on symptomatic patients assisted at Armed Forces Hospital (Hfa) in Brasília, Brazil.

Dengue necessitates accurate diagnosis. Rapid tests such as Bioline™ DENGUE DUO have gained traction, but validation in specific populations is essential. This study aimed to evaluate the performance of the Bioline™ test, alongside assessing the socio-epidemiological profile of symptomatic patients in a Brasília Military Hospital. The serum of 404 symptomatic patients was analyzed by the Bioline™ DENGUE DUO test, followed by Dengue virus detection and discrimination of the four serotypes by RT-qPCR. Accuracy was assessed using parameters including sensitivity (S), specificity (E), positive and negative predictive values (PPV and NPV), and positive (RV +) and negative (RV-) likelihood ratios. The NS1 component exhibited a sensitivity of 70.37%, a specificity of 97.30%, and an overall efficiency of 90.10% when compared to RT-qPCR as the gold standard. The IgM component demonstrated a sensitivity of 26.85%, a specificity of 89.53%, and an overall efficiency of 72.77% when compared to RT-qPCR as the gold standard. The IgG component demonstrated a sensitivity of 23.15%, a specificity of 68.92%, and an overall efficiency of 56.68% when compared to RT-qPCR as the gold standard. Several rapid tests are commercially available. However, considering variations across regions and demographic groups, it is important to question their accuracy in specific populations. Rapid tests are important screening tools, but they can have limitations for the certainty of diagnosis. Bioline™ DENGUE DUO displayed good specificity, but sensitivity was slightly below optimal levels. While helpful for confirming dengue, improvements are needed to effectively rule out the disease.

Role of angiotensin II in cellular entry and replication of dengue virus.

Previous studies have demonstrated the relevance of several soluble molecules in the pathogenesis of dengue. In this regard, a possible role for angiotensin II (Ang II) in the pathophysiology of dengue has been suggested by the observation of a blockade of Ang II in patients with dengue, increased expression of molecules related to Ang II production in the plasma of dengue patients, increased expression of circulating cytokines and soluble molecules related to the action of Ang II, and an apparent relationship between DENV, Ang II effects, and miRNAs. In addition, in ex vivo experiments, the blockade of Ang II AT1 receptor and ACE-1 (angiotensin converting enzyme 1), both of which are involved in Ang II production and its function, inhibits infection of macrophages by DENV, suggesting a role of Ang II in viral entry or in intracellular viral replication of the virus. Here, we discuss the possible mechanisms of Ang II in the entry and replication of DENV. Ang II has the functions of increasing the expression of DENV entry receptors, creation of clathrin-coated vesicles, and increasing phagocytosis, all of which are involved in DENV entry. This hormone also modulates the expression of the Rab5 and Rab7 proteins, which are important in the endosomal processing of DENV during viral replication. This review summarizes the data related to the possible involvement of Ang II in the entry of DENV into cells and its replication.