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The yellow fever (YF) vaccine is usually produced with egg-based methods, which has limitations, including potential adverse effects and low production yields. Alternatively, producing the vaccine using Vero cells or HEK 293 cells can overcome some of these issues, but these methods are significantly more expensive. In the current study, the YF vaccine candidate 17DD virus was produced in primary chicken embryo fibroblast (CEF) cells. The primary CEF cells isolation from eggs was optimized through a two-step process. In the first step, the important parameters that contribute to the development of the egg embryo, such as egg position, relative humidity (RH), and incubation time are optimized. In second step, primary CEF release parameters namely; trypsin volume and incubation temperature are optimized. Both steps were optimized using statistical methods. Further, the seeding cell density of isolated CEF was also optimized. It was observed that 5 x 104 cells/cm2 gave the highest virus titer of 3.89 million PFU/ml. The 17DD yields achieved in primary CEFs are much higher than egg-based production and it is an economically viable method. [ABSTRACT FROM AUTHOR]

Mosquito-borne flaviviruses represent a public health challenge due to the high-rate endemic infections, severe clinical outcomes, and the potential risk of emerging global outbreaks. Flavivirus disease pathogenesis converges on cellular factors from vectors and hosts, and their interactions are still unclear. Exosomes and microparticles are extracellular vesicles released from cells that mediate the intercellular communication necessary for maintaining homeostasis; however, they have been shown to be involved in disease establishment and progression. This review focuses on the roles of extracellular vesicles in the pathogenesis of mosquito-borne flavivirus diseases: how they contribute to viral cycle completion, cell-to-cell transmission, and cellular responses such as inflammation, immune suppression, and evasion, as well as their potential use as biomarkers or therapeutics (antiviral or vaccines). We highlight the current findings concerning the functionality of extracellular vesicles in different models of dengue virus, Zika virus, yellow fever virus, Japanese encephalitis virus, and West Nile virus infections and diseases. The available evidence suggests that extracellular vesicles mediate diverse functions between hosts, constituting novel effectors for understanding the pathogenic mechanisms of flaviviral diseases. [ABSTRACT FROM AUTHOR]

Introduction: Japanese encephalitis virus (JEV) and Zika virus (ZIKV) are prevalent in over 80 countries or territories worldwide, causing hundreds of thousands of cases annually. But currently there is a lack of specific antiviral agents and effective vaccines. Methods: In the present study, to identify human neutralizing monoclonal antibody (mAb) against JEV or/and ZIKV, we isolated ZIKV-E protein-binding B cells from the peripheral venous blood of a healthy volunteer who had received the JEV live-attenuated vaccine and performed 10× Genomics transcriptome sequencing and BCR sequencing analysis, we then obtained the V region amino acid sequences of a novel mAb LZY3412. We expressed mAb LZY3412 and evaluated its ability to bind to E proteins of dengue virus, JEV and ZIKV, neutralize JEV and ZIKV infections in vitro , protect mice against lethal JEV or ZIKV attack. The epitopes on E proteins of JEV/ZIKV recognized by mAb LZY3412 were analyzed using molecular docking and constructing E protein mutants. Results: Our results show that recombinant mAb LZY3412 has high-affinity with the E proteins of three viruses, with the kinetically derived binding affinity (KD) values of 440 and 482.5 nM against JEV-E protein and ZIKV-E protein, respectively; recombinant mAb LZY3412 can efficiently neutralize JEV and ZIKV infections in vitro , with the NT50 values of 19.9 ng/mL and 631 ng/mL, respectively; application of recombinant mAb LZY3412 can significantly improve the percentage survival and reduce the serum viral loads of neonatal mice infected with JEV or ZIKV. Finally, two amino acid residues (Ala399 and Gly400) located in EDIII of JEV-E protein were potentially recognized by LZY3412 whereas two amino acid residues (Met15 and Thr406) out of EDIII of ZIKV-E proteins were recognized by LZY3412. Discussion: Considering mAb LZY3412 has neutralizing effects on various flaviviruses, this mAb is expected to be developed as therapeutic or preventive drug for these flaviviruses, and can also be used to guide the development of vaccines against these flaviviruses. [ABSTRACT FROM AUTHOR]

The European subtype of tick-borne encephalitis virus (TBEV-Eur; species Orthoflavivirus encephalitidis, family Flaviviridae) was the only tick-borne flavivirus present in central Europe known to cause neurologic disease in humans and several animal species. Here, we report a tick-borne flavivirus isolated from Alpine chamois (Rupicapra rupicapra rupicapra) with encephalitis and attached ticks, present over a wide area in the Alps. Cases were detected in 2017 in Salzburg, Austria, and 2023 in Lombardy and Piedmont, Italy. The virus strains exhibit 94.8–97.3% nucleotide identities to each other and are more closely related to Louping ill viruses (LIV; Orthoflavivirus loupingi; 90–92% identities) than to TBEV-Eur (less than 88%). The chamois-derived virus strains, tentatively termed "Alpine chamois encephalitis virus", form a well-supported independent genetic clade with Spanish goat encephalitis virus, clearly separated from other LIV. This supports its designation as a new virus subtype with the proposed shared taxonomic name "Spanish goat and Alpine chamois encephalitis virus subtype" within the species Orthoflavivirus loupingi. The zoonotic potential of this newly identified virus subtype as well as its host range in other animal species including farm animals needs to be further investigated. [ABSTRACT FROM AUTHOR]

Flaviviruses are a diverse group of viruses primarily transmitted through hematophagous insects like mosquitoes and ticks. Significant expansion in the geographic range, prevalence, and vectors of flavivirus over the last 50 years has led to a dramatic increase in infections that can manifest as hemorrhagic fever or encephalitis, leading to prolonged morbidity and mortality. Millions of infections every year pose a serious threat to worldwide public health, encouraging scientists to develop a better understanding of the pathophysiology and immune evasion mechanisms of these viruses for vaccine development and antiviral therapy. Extensive research has been conducted in developing effective antivirals for flavivirus. Various approaches have been extensively utilized in clinical trials for antiviral development, targeting virus entry, replication, polyprotein synthesis and processing, and egress pathways exploiting virus as well as host proteins. However, to date, no licensed antiviral drug exists to treat the diseases caused by these viruses. Understanding the mechanisms of host–pathogen interaction, host immunity, viral immune evasion, and disease pathogenesis is highly warranted to foster the development of antivirals. This review provides an extensively detailed summary of the most recent advances in the development of antiviral drugs to combat diseases. [ABSTRACT FROM AUTHOR]

Flaviviruses are arthropod-borne viruses primarily transmitted through the mosquito Aedes aegypti or Culex genus of mosquitos. These viruses are predominantly found in tropical and subtropical regions of the world with their geographical spread predicted to increase as global temperatures continue to rise. These viruses cause a variety of diseases in humans with the most prevalent being caused by dengue, resulting in hemorrhagic fever and associated sequala. Current approaches for therapeutic control of flavivirus infections are limited, and despite recent advances, there are no approved drugs. Vaccines, available for a few circulating flaviviruses, still have limited potential for controlling contemporary and future outbreaks. Mouse models provide us with a valuable tool to test the effectiveness of drugs and vaccines, yet for many flaviviruses, well-established mouse models are lacking. In this review, we highlight the current state of flavivirus vaccines and therapeutics, as well as our current understanding of mouse models for various flaviviruses. [ABSTRACT FROM AUTHOR]

Tick-borne encephalitis virus (TBEV) is flavivirus transmitted to the host via tick saliva which contains various molecules with biological impacts. One of such molecules is Iristatin, a cysteine protease inhibitor from Ixodes ricinus that has been shown to have immunomodulatory properties. To characterize Iristatin in the relation to TBEV, we investigate whether this tick inhibitor has any capacity to influence TBEV infection. Mice were intradermally infected by TBEV with or without Iristatin and the viral multiplication was determined in skin and brain tissues by RT-PCR two and 5 days after infection. The viral RNA was detected in both intervals in skin and increased by time. The application of Iristatin caused a reduction in viral RNA in skin but not in the brain of infected mice 5 days post-infection. Moreover, anti-viral effect of Iristatin on skin was accompanied by a significant decline of interferon-stimulated gene 15 gene expression. The effect of Iristatin on TBEV replication was tested also in vitro in primary macrophages and dendritic cells; however, no changes were observed suggesting no direct interference of Iristatin with virus replication. Still, the Iristatin caused a suppression of Erk1/2 phosphorylation in TBEV-infected dendritic cells and had the anti-apoptotic effect. This is the first report showing that a tick cystatin decreases the viral RNA in the host skin, likely indirectly through creating skin environment that is less supportive for TBEV replication. Assuming, that viral RNA reflects the amount of infectious virus, decline of TBEV in host skin could influence the tick biology or virus transmission during cofeeding. [ABSTRACT FROM AUTHOR]

Flaviviruses, a group of single-stranded RNA viruses spread by mosquitoes or ticks, include several significant neurotropic viruses, such as West Nile virus (WNV), Japanese encephalitis virus (JEV), tick-borne encephalitis virus (TBEV), and Zika virus (ZIKV). These viruses can cause a range of neurological diseases during acute infection, from mild, flu-like symptoms to severe and fatal encephalitis. A total of 20–50% of patients who recovered from acute flavivirus infections experienced long-term cognitive issues. Here, we discuss these major neurotropic flaviviruses-induced clinical diseases in humans and the recent findings in animal models and provide insights into the underlying disease mechanisms. [ABSTRACT FROM AUTHOR]

Background This study aimed to characterise the clinical and epidemiological profiles of dengue patients and their outcomes during an ongoing outbreak in a non-endemic region of Bangladesh. Methods This prospective observational study analysed 805 confirmed dengue cases during August–December 2023. Data on demographic, clinical and laboratory profiles, as well as outcomes, were gathered using a structured questionnaire. Statistical analysis was conducted using SPSS 25. Results The mean age of dengue patients was 31.5 (±12.2) y, with the majority being males (81.2%). All 805 patients experienced fever, 792 (98.4%) had headaches, 698 (86.7%) had myalgia, 601 (74.7%) had persistent vomiting and 598 (74.3%) had abdominal pain. Bleeding was observed in 191 (23.7%) patients and neurological symptoms were seen in 209 (25.9%) patients. Most patients (n=781, 97%) exhibited non-severe symptoms, while 3% (n=24) had severe symptoms. Among the 24 severe cases, four (16.7%) patients were reported to have encephalitis and one (4.2%) patient had meningoencephalitis. Moreover, 365 patients (45.3%) had travelled to an endemic region who were predominantly males (n=327, 89.6%). Most dengue patients recovered well with rapid fluid replacement therapy (n=754, 93.7%). Conclusions The 2023 dengue outbreak in a non-endemic area of Bangladesh primarily impacted males, young adults, with the majority presenting non-severe symptoms. Further studies are essential to validate and build upon these results. [ABSTRACT FROM AUTHOR]

AbstractThe Flaviviridae family includes many medically relevant members, such as dengue, Zika, West Nile, and hepatitis C viruses, that produce hundreds of millions of infections annually. There is a close relationship between these infections and inflammation triggering as an important part of the host’s immune response and of pathogenesis. These inflammatory processes are mediated by the activation of multiprotein complexes known as inflammasomes. Several inflammasomes have been described which differ in their composition and their activating stimuli. The NLRP3 inflammasome is the most studied. Its activation begins by the recognition of pathogen-associated molecular patterns such as viral RNA, potassium efflux, calcium flux, increased reactive oxygen species; and culminates in the maturation and secretion of pro-inflammatory cytokines such as IL-1β and IL-18, and cell death by pyroptosis. This review summarizes the most relevant aspects of NLRP3 inflammasome activation in relevant flavivirus infections from clinical and laboratory studies in biological models. Understanding the activation, mounting, and regulation of the inflammatory response during viral infections is a poorly exploited area of opportunity for the development of efficient and safe treatment strategies, which could include NLRP3 inflammasome inhibition. [ABSTRACT FROM AUTHOR]

Arthropod-borne viruses or arboviruses, including West Nile virus (WNV), dengue virus (DENV), and Zika virus (ZIKV) pose significant threats to public health. It is imperative to develop novel methods to control these mosquito-borne viral infections. We previously showed that insulin/insulin-like growth factor-1 signaling (IIS)-dependent activation of ERK and JAK-STAT signaling has significant antiviral activity in insects and human cells. Continuous immune pressure can lead to adaptive mutations of viruses during infection. We aim to elucidate how IIS-signaling in mosquitoes selects for West Nile virus escape variants, to help formulate future transmission blocking strategies. We hypothesize that passage of WNV under activation of IIS will induce adaptive mutations or escape variants in the infecting virus. To test our hypothesis, WNV was serially passaged through Culex quinquefasciatus Hsu cells in the presence or absence of bovine insulin to activate IIS antiviral pressure. We sequenced WNV genes encoding for E, NS2B, NS3, and NS5 and identified variants in E and NS5 arising from IIS antiviral pressure. In parallel to the genetic analyses, we also report differences in the levels of virus replication and Akt activation in human cells and mosquitoes using virus passaged in the presence or absence of insulin. Finally, using adult Culex quinquefasciatus , we demonstrated the enhancement of immune response gene expression in virus-infected mosquitoes fed on insulin, compared to control. Notably, virus collected from insulin-fed mosquitoes contained a non-synonymous mutation in NS3. These results contribute towards achieving our long-term goal of manipulating mosquito IIS-dependent antiviral immunity to reduce WNV or other flavivirus transmission to mammalian hosts. [ABSTRACT FROM AUTHOR]

The green peach aphid (Myzus persicae) is a generalist pest damaging crops and transmitting viral pathogens. Using Illumina sequencing of small (s)RNAs and poly(A)-enriched long RNAs, we analyzed aphid virome components, viral gene expression and antiviral RNA interference (RNAi) responses. Myzus persicae densovirus (family Parvoviridae), a single-stranded (ss)DNA virus persisting in the aphid population, produced 22 nucleotide sRNAs from both strands of the entire genome, including 5′- and 3′-inverted terminal repeats. These sRNAs likely represent Dicer-dependent small interfering (si)RNAs, whose double-stranded RNA precursors are produced by readthrough transcription beyond poly(A) signals of the converging leftward and rightward transcription units, mapped here with Illumina reads. Additionally, the densovirus produced 26–28 nucleotide sRNAs, comprising those enriched in 5′-terminal uridine and mostly derived from readthrough transcripts and those enriched in adenosine at position 10 from their 5′-end and mostly derived from viral mRNAs. These sRNAs likely represent PIWI-interacting RNAs generated by a ping-pong mechanism. A novel ssRNA virus, reconstructed from sRNAs and classified into the family Flaviviridae, co-persisted with the densovirus and produced 22 nucleotide siRNAs from the entire genome. Aphids fed on plants versus artificial diets exhibited distinct RNAi responses affecting densovirus transcription and flavivirus subgenomic RNA production. In aphids vectoring turnip yellows virus (family Solemoviridae), a complete virus genome was reconstituted from 21, 22 and 24 nucleotide viral siRNAs likely acquired with plant phloem sap. Collectively, deep-sequencing analysis allowed for the identification and de novo reconstruction of M. persicae virome components and uncovered RNAi mechanisms regulating viral gene expression and replication. [ABSTRACT FROM AUTHOR]

The Zika virus (ZIKV) epidemic elicited a rapid commitment to the development of animal models for ZIKV research. Non-human primates (NHPs) and mice have made significant contributions to this research, but NHPs are expensive, have a long gestation period, and are available only in small numbers; non-genetically modified mice are resistant to infection. To address these deficiencies, we have established the laboratory opossum, Monodelphis domestica, as a small animal model that complements the mouse and monkey models. We developed and validated an indirect ELISA for measuring antibodies to ZIKV in opossums, as well as an immunohistochemistry (IHC) method to detect ZIKV NS1 protein in tissue samples. Opossum pups inoculated intracerebrally as embryos, juveniles inoculated by several routes, and mothers that cannibalized inoculated pups became persistently infected with ZIKV. The virus spread to multiple organs and persisted for up to 38 weeks (the latest endpoint of the experiments). A robust humoral immune response was mounted, and high titers of antibodies also persisted for 38 weeks. The results establish M. domestica as a natural, non-genetically modified animal model in which ZIKV persists long-term after experimental exposure and as a unique animal model for research on the immune response to ZIKV. [ABSTRACT FROM AUTHOR]

Proteases are key enzymes in viral replication, and interfering with these targets is the basis for therapeutic interventions. We previously introduced a hypothesis about conformational selection in the protease of dengue virus and related flaviviruses, based on conformational plasticity noted in X-ray structures. The present work presents the first functional evidence for alternate recognition by the dengue protease, in a mechanism based primarily on conformational selection rather than induced-fit. Recognition of distinct substrates and inhibitors in proteolytic and binding assays varies to a different extent, depending on factors reported to influence the protease structure. The pH, salinity, buffer type, and temperature cause a change in binding, proteolysis, or inhibition behavior. Using representative inhibitors with distinct structural scaffolds, we identify two contrasting binding profiles to dengue protease. Noticeable effects are observed in the binding assay upon inclusion of a non-ionic detergent in comparison to the proteolytic assay. The findings highlight the impact of the selection of testing conditions on the observed ligand affinity or inhibitory potency. From a broader scope, the dengue protease presents an example, where the induced-fit paradigm appears insufficient to explain binding events with the biological target. Furthermore, this protein reveals the complexity of comparing or combining biochemical assay data obtained under different conditions. This can be particularly critical for artificial intelligence (AI) approaches in drug discovery that rely on large datasets of compounds activity, compiled from different sources using non-identical testing procedures. In such cases, mismatched results will compromise the model quality and its predictive power. [Display omitted] • Buffer type, pH, salt, and temperature influence ligand recognition. • Experimental conditions in binding and proteolytic assays affect the obtained data. • Ligand recognition in DENV protease involves mainly conformational selection. [ABSTRACT FROM AUTHOR]

A cellular protein, non-POU-domain-containing octamer binding protein (NONO), bound to the replication complex of Japanese encephalitis virus (JEV) by directly interacting with the viral 3' UTR RNA and NS3 protein. These interactions were also identified in West Nile virus (WNV) and Zika virus (ZIKV). The infection of JEV or the expression of JEV NS3 protein in cells could induce relocation of NONO protein from the nucleus to the cytoplasm. In JEV-infected cells, the NS3, NS5, and viral RNA could be concurrently detected in the immunoprecipitation by the NONO-specific antibody, suggesting that NONO could integrate into the replication complex of JEV. Further results of co-immunoprecipitation assays showed that NONO protein interacted with NS3 helicase domains 1 and 2 by its two RNA recognize motifs (RRMs). The knockdown and knockout of NONO in cells could significantly reduce the replication of JEV and ZIKV but had no effect on the replication of vesicular stomatitis virus (VSV). The effect of NONO protein on JEV proliferation occurred during the replication stage, rather than the attachment and entry stages. The level of viral positive-strand RNA in NONO knockout cells was significantly reduced than that in wild-type cells at 12-48 h post-JEV infection. However, the level of negative-strand virus RNA had no difference between NONO knockout and wild-type cells at 12-24 h post-infection. In summary, our study ident ified a cellular protein that bound to the replication complex of flavivirus and facilitated the synthesis of positive-strand RNA. [ABSTRACT FROM AUTHOR]

West Nile Virus (WNV) is a zoonotic, vector-borne pathogen affecting humans and animals, particularly in Europe. The virus is primarily transmitted through mosquitoes that infect birds, which serve as the main reservoirs. Humans and horses are incidental hosts. This review focuses on the epidemiology of WNV in southern Europe, particularly its increasing prevalence. Methods included an extensive literature review and analysis of recent outbreaks. WNV is largely asymptomatic in humans, but a small percentage can develop West Nile neuroinvasive disease (WNND), leading to severe neurological symptoms and fatalities. Horses can also suffer from neurological complications, with high mortality rates. Climate change, migratory birds, and mosquito population dynamics contribute to the virus spread across Europe. Control efforts focus on vector management, and while vaccines are available for horses, none has been approved for humans. Surveillance, particularly of bird and mosquito populations, and further research into the virus molecular structure are crucial for understanding and mitigating future outbreaks. [ABSTRACT FROM AUTHOR]

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As positive-sense RNA viruses, the genomes of flaviviruses serve as the template for all stages of the viral life cycle, including translation, replication, and infectious particle production. Yet, they encode just 10 proteins, suggesting that the structure and dynamics of the viral RNA itself helps shepherd the viral genome through these stages. Herein, we highlight advances in our understanding of flavivirus RNA structural elements through the lens of their impact on the viral life cycle. We highlight how RNA structures impact translation, the switch from translation to replication, negative- and positive-strand RNA synthesis, and virion assembly. Consequently, we describe three major themes regarding the roles of RNA structure in flavivirus infections: 1) providing a layer of specificity; 2) increasing the functional capacity; and 3) providing a mechanism to support genome compaction. While the interactions described herein are specific to flaviviruses, these themes appear to extend more broadly across RNA viruses.

One of the most recent advances in the analysis of viral RNA-cellular protein interactions is the Comprehensive Identification of RNA-binding Proteins by Mass Spectrometry (ChIRP-MS). Here, we used ChIRP-MS in mock-infected and Zika-infected wild-type cells and cells knockout for the zinc finger CCCH-type antiviral protein 1 (ZAP). We characterized ‘ZAP-independent’ and ‘ZAP-dependent’ cellular protein interactomes associated with flavivirus RNA and found that ZAP affects cellular proteins associated with Zika virus RNA. The ZAP-dependent interactome identified with ChIRP-MS provides potential ZAP co-factors for antiviral activity against Zika virus and possibly other viruses. Identifying the full spectrum of ZAP co-factors and mechanisms of how they act will be critical to understanding the ZAP antiviral system and may contribute to the development of antivirals.

Flaviviruses, including Zika virus (ZIKV) and Dengue virus (DENV), rely on their non-structural protein 5 (NS5) for both replication of viral genome and suppression of host IFN signaling. DENV and ZIKV NS5s were shown to facilitate proteosome-mediated protein degradation of human STAT2 (hSTAT2). However, how flavivirus NS5s have evolved for species-specific IFN-suppression remains unclear. Here we report structure-function characterization of the DENV serotype 2 (DENV2) NS5-hSTAT2 complex. The MTase and RdRP domains of DENV2 NS5 form an extended conformation to interact with the coiled-coil and N-terminal domains of hSTAT2, thereby promoting hSTAT2 degradation in cells. Disruption of the extended conformation of DENV2/ZIKV NS5, but not the alternative compact state, impaired their hSTAT2 binding. Our comparative structural analysis of flavivirus NS5s further reveals a conserved protein-interaction platform with subtle amino-acid variations likely underpinning diverse IFN-suppression mechanisms. Together, this study uncovers a conformational selection mechanism underlying species-specific hSTAT2 inhibition by flavivirus NS5.

Abstract Background Flavivirus is a highly prevalent and outbreak-prone disease, affecting billions of individuals annually and posing substantial public health challenges. Vaccination is critical to reducing the global impact of flavivirus infections, making the development of a safe and effective vaccine a top priority. The self-assembled pan-epitope vaccine presents key advantages for improving immunogenicity and safety without relying on external vectors or adding immunomodulatory elements, both of which are essential for successful vaccine development. Results In this study, the pan-epitope peptide TBT was combined with adjuvant CpG to form the TBT-CpG nanovaccine (TBT-CpG NaVs), which was found to be spherical, uniform in shape, and demonstrated strong serum stability. In vitro studies showed that the TBT-CpG NaVs were efficiently taken up and internalized by bone marrow-derived dendritic cells (BMDCs). Flow cytometry and transcriptomic analysis indicated that the antigens were effectively presented to antigen-presenting cells (APCs) via the MHC II pathway, which facilitated BMDCs maturation and promoted the release of pro-inflammatory cytokines IL-1β, TNF-α, and IL-6. In vivo studies confirmed that TBT-CpG NaVs enhanced antigen-specific IgG levels, significantly increased IFN-γ and IL-4 expression in spleen cells, and offered protective effects against Dengue virus (DENV) and Zika virus (ZIKV) infections. Safety evaluations revealed no hepatotoxicity and no significant organ damage in immunized mice. Conclusion The self-assembled candidate nanovaccine TBT-CpG NaVs effectively activates BMDCs and triggers a targeted immune response, providing antiviral effects against DENV and ZIKV. This vaccine demonstrates good immunogenicity and safety, establishing a promising foundation and a new strategy for the development of safe and effective vaccines. Graphical Abstract

Abstract Dengue virus (DENV) is a mosquito-borne virus that causes dengue fever, a significant public health concern in many tropical and subtropical regions. Dengue is endemic in more than 100 countries, primarily in tropical and subtropical regions of the world. Each year, up to 400 million people get infected with dengue. Approximately 100 million people get sick from infection, and 40,000 die from severe dengue. Unfortunately, dengue vaccine development is also marred with various complicating factors, as the forefront candidate vaccine performed unsatisfactorily. Moreover, the only licensed vaccine (Dengvaxia) for children 9 through 16 years of age is available in just a few countries. The treatment difficulties are compounded by the absence of an effective antiviral agent. Exploring plant-based therapeutics for dengue from the laboratory to clinical application involves a multi-stage process, encompassing various scientific disciplines. Individual investigators have screened a wide range of plant extracts or compounds for potential antiviral activity against DENV. In vitro studies help identify candidates that exhibit inhibitory effects on viral replication. Some of the most promising medicinal plants showing in vitro activity against DENV include Andrographis paniculate , Acorus calamus, and Cladogynos orientalis. Further laboratory studies, both in vitro and in animal models (in vivo), elucidate the mechanisms of action by which the identified compounds exert antiviral effects. Medicinal plants such as Carica papaya, Cissampelos pareira, and Ipomea batata exhibited potent platelet-enhancing activities while Azadirachta indica and Curcuma longa showed promising effects in both in vitro and in vivo studies. Based on positive preclinical results, researchers design clinical trials. This involves careful planning of trial phases, patient recruitment criteria, ethical considerations, and endpoints. The most important medicinal plants showing efficacy and safety in clinical trials include Carica papaya and Cissampelos pareira. This review suggests that several promising medicinal plants exist that have the potential to be turned into clinical drugs to treat dengue infection. However, in addition to developing synthetic and plant-based therapies against dengue infection, vector management strategies should be made robust, emphasizing the need to focus on reducing disease incidence.

Abstract The zinc finger antiviral protein 1 (ZAP) has broad antiviral activity. ZAP is an interferon (IFN)-stimulated gene, which itself may enhance type I IFN antiviral response. In a previous study, Zika virus was identified as ZAP-resistant and not sensitive to ZAP antiviral activity. Here, we found that ZAP was associated with the inhibition of Zika virus in Vero cells, in the absence of a robust type I IFN system because Vero cells are deficient for IFN-alpha and -beta. Also, quantitative RNA-seq data indicated that endogenous ZAP is associated with altered global gene expression both in the steady state and during Zika virus infection. Further studies are warranted to elucidate this IFN-alpha and -beta independent anti-Zika virus activity and involvement of ZAP.

Abstract Hepatitis C virus (HCV) infection poses a significant public health challenge and often leads to long-term health complications and even death. Parkinson’s disease (PD) is a progressive neurodegenerative disorder with a proposed viral etiology. HCV infection and PD have been previously suggested to be related. This work aimed to identify potential biomarkers and pathways that may play a role in the joint development of PD and HCV infection. Using BioOptimatics-bioinformatics driven by mathematical global optimization-, 22 publicly available microarray and RNAseq datasets for both diseases were analyzed, focusing on sex-specific differences. Our results revealed that 19 genes, including MT1H, MYOM2, and RPL18, exhibited significant changes in expression in both diseases. Pathway and network analyses stratified by sex indicated that these gene expression changes were enriched in processes related to immune response regulation in females and immune cell activation in males. These findings suggest a potential link between HCV infection and PD, highlighting the importance of further investigation into the underlying mechanisms and potential therapeutic targets involved.

Abstract Emerging infectious diseases are a major threat to biodiversity and an important public health issue. Flaviviruses are the cause of several emerging vector-borne zoonotic arboviruses whose distribution is currently increasing in Europe. The evidence that West Nile virus (WNV) circulates in resident and migratory species has implications for both animal and public health and should therefore be studied in depth. USUTU (USUV), Bagaza (BAGV) and tick-borne encephalitis virus (TBEV) are other viruses that are beginning to spread more widely. An integrated surveillance program, namely in birds, is essential for reducing the risk of infection in human populations within the One Health principles. In the present study, wild birds admitted to wildlife rehabilitation centers in Portugal were sampled. Two hundred eight blood samples were assayed serologically for antibodies to flaviviruses by using a commercial ELISA kit. An overall seroprevalence of 19.6% (95% confidence interval [CI]: 13.7–26.7%) was observed. Antibodies against flaviviruses were detected in 13 (35.1%) different species of wild birds. Accipitriformes (26.7%; 95% CI: 18.5–36.2%) and Strigiformes (26.7%; 95% CI: 14.6–42.0%) were the orders with the highest seroprevalence rates recorded. There were no statistically significant differences (p = 0.725) between the geographical regions (NUTS II) studied, but a statistically significant difference (p = 0.017) was found between sex (male: 34.4%; female: 4.8%). A higher seroprevalence was detected in adults (32.1%) than in juvenile birds (9.3%) (p = 0.014), and age was considered a risk factor for flavivirus infection in wild birds (odds ratio 1.4; 95% CI: 0.5–4.0). More epidemiological studies are needed in Portugal since the actual spread of the genus Flavivirus throughout the country is unknown.