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flavivirus - Top 30 Publications

Prolonged Detection of Japanese Encephalitis Virus in Urine and Whole Blood in a Returned Short-term Traveler.

We describe a fatal case of Japanese encephalitis virus infection following short-term travel to Thailand. Viral RNA was detected in urine and whole blood out to 26 and 28 days, respectively, after the onset of symptoms. Live virus was isolated from a urine specimen from day 14.

Dengue fever in a south Asian metropolis: a report on 219 cases.

Yearly epidemics of Dengue fever occur post-monsoon in India's capital, Delhi. A prospective observational study was conducted during the outbreak months to understand the epidemiology and outcome of this infection and its economic impact.

Differences in prevalence of symptomatic Zika virus infection by age and sex-Puerto Rico, 2016.

During the Zika virus (ZIKV) outbreak in Puerto Rico in 2016, non-pregnant women aged 20-39 years were disproportionately identified with ZIKV disease. We used household-based cluster investigations to determine if this disparity was associated with age- or sex-dependent differences in the rate of ZIKV infection or reporting symptoms.

Suppression of Zika virus infection and replication in endothelial cells and astrocytes by PKA inhibitor PKI 14-22.

The recent outbreak of Zika virus (ZIKV), a re-emerging flavivirus, and its associated neurological disorders, such as Guillain-Barré (GB) syndrome and microcephaly, have generated an urgent need for developing effective ZIKV vaccines and therapeutic agents. Here, we used human endothelial cells and astrocytes, both of which represent key cell types for ZIKV infection, to identify potential inhibitors for ZIKV replication. Because several pathways, including AMP-activated protein kinase (AMPK), protein kinase A (PKA), and mitogen-activated protein kinase (MAPK) signaling pathways, have been reported to play important roles in flavivirus replication, we tested inhibitors or agonists of these pathways for their effects on ZIKV replication. We identified PKA inhibitor, PKI 14-22 (PKI), as a potent inhibitor of ZIKV replication. PKI effectively suppressed the replication of ZIKV from both African and Asian/American lineages with high efficiency and minimal cytotoxicity. While ZIKV infection did not induce PKA activation, endogenous PKA activity was essential for supporting ZIKV replication. Interestingly, in addition to PKA, PKI also inhibited other unknown target(s) to block ZIKV replication. PKI inhibited ZIKV replication at the post-entry stage by preferentially affecting negative-sense RNA synthesis as well as viral protein translation. Together, these results have identified a potential inhibitor of ZIKV replication, which could be further explored for future therapeutic application.ImportanceThere is an urgent need to develop effective vaccines and therapeutic agents against Zika virus (ZIKV) infection, a re-emerging flavivirus associated with neurological disorders including Guillain-Barré (GB) syndrome and microcephaly. By screening for inhibitors of several cellular pathways, we have identified PKA inhibitor PKI 14-22 (PKI) as a potent inhibitor of ZIKV replication. We have shown that PKI effectively suppresses the replication of ZIKV of all the strains tested with minimal cytotoxicity in human endothelial cells and astrocytes, two key cell types of ZIKV infection. Furthermore, we have shown that PKI inhibits ZIKV negative-sense RNA synthesis and viral protein translation. This study has identified a potent inhibitor of ZIKV infection, which could be further explored for future therapeutic application.

Production of Monoclonal Antibody That Recognizes Zika Virus and Other Flaviviruses in Serum-Free Conditions.

With the recent outbreaks of Zika and Dengue virus infections in various countries worldwide, production of vaccines or diagnostic kits is an urgent public health demand. Production of a monoclonal antibody (mAb) that specifically binds to a common antigen shared by the Flavivirus genus will be necessary for new diagnostic kits or characterization and viral identity tests during vaccine development. This study aimed to cultivate, in serum-free conditions, the 4G2 hybridoma that produces an mAb, which recognizes a shared epitope from the Flavivirus genus. We compared 4G2 hybridoma growth and biochemical profiles between cells cultivated in batch mode over 10 days in roller bottles containing Dulbecco's modified Eagle's medium high glucose containing 10% fetal bovine serum medium or hybridomas directly adapted to Ex-Cell serum-free medium. Cellular parameters such as specific growth rate (μ), maximum cell concentration, specific l-lactate, and glucose and IgG rates were evaluated. Thereafter, we also compared total mAb volumetric productivity, purification yield, and mAb staining of Vero cells infected with Zika and Dengue-2 virus. Direct adaptation to serum-free conditions did not change hybridoma growth rate and mAb production under the conditions tested. Instead, serum-free mAb purification showed a higher yield with no alterations on mAb structure or mAb staining of Zika and Dengue Vero-infected cells.

Metagenomic analysis reveals Hepatitis A virus in suspected yellow fever cases in Brazil.

Using a metagenomic approach, we identified hepatitis A virus among cases of acute febrile illnesses that occurred in 2008-2012 in Brazil suspected as yellow fever. These findings reinforce the challenge facing routine clinical diagnosis in complex epidemiological scenarios.

Ribosomal stress and Tp53-mediated neuronal apoptosis in response to capsid protein of the Zika virus.

We report here that in rat and human neuroprogenitor cells as well as rat embryonic cortical neurons Zika virus (ZIKV) infection leads to ribosomal stress that is characterized by structural disruption of the nucleolus. The anti-nucleolar effects were most pronounced in postmitotic neurons. Moreover, in the latter system, nucleolar presence of ZIKV capsid protein (ZIKV-C) was associated with ribosomal stress and apoptosis. Deletion of 22 C-terminal residues of ZIKV-C prevented nucleolar localization, ribosomal stress and apoptosis. Consistent with a casual relationship between ZIKV-C-induced ribosomal stress and apoptosis, ZIKV-C-overexpressing neurons were protected by loss-of-function manipulations targeting the ribosomal stress effector Tp53 or knockdown of the ribosomal stress mediator RPL11. Finally, capsid protein of Dengue virus, but not West Nile virus, induced ribosomal stress and apoptosis. Thus, anti-nucleolar and pro-apoptotic effects of protein C are flavivirus-species specific. In the case of ZIKV, capsid protein-mediated ribosomal stress may contribute to neuronal death, neurodevelopmental disruption and microcephaly.

Imaging findings in congenital Zika virus infection syndrome: an update.

Zika virus (ZIKV) is a neurotropic and neurotoxic RNA Flavivirus prompt to cause severe fetal brain dysmorphisms during pregnancy, a period of rapid and critical central nervous system development. A wide range of clinico-radiological findings of congenital ZIKV infections were reported in the literature, such as microcephaly, overlapping sutures, cortical migrational and corpus callosum abnormalities, intracranial calcifications, ventriculomegaly, brain stem and cerebellar malformations, spinal cord involvement, and joint contractures. ZIKV is also related to other severe neurological manifestations in grown-up individuals such as Guillain-Barré syndrome and encephalomyelitis.

Economics of One Health: Costs and benefits of integrated West Nile virus surveillance in Emilia-Romagna.

Since 2013 in Emilia-Romagna, Italy, surveillance information generated in the public health and in the animal health sectors has been shared and used to guide public health interventions to mitigate the risk of West Nile virus (WNV) transmission via blood transfusion. The objective of the current study was to identify and estimate the costs and benefits associated with this One Health surveillance approach, and to compare it to an approach that does not integrate animal health information in blood donations safety policy (uni-sectoral scenario). Costs of human, animal, and entomological surveillance, sharing of information, and triggered interventions were estimated. Benefits were quantified as the averted costs of potential human cases of WNV neuroinvasive disease associated to infected blood transfusion. In the 2009-2015 period, the One Health approach was estimated to represent a cost saving of €160,921 compared to the uni-sectoral scenario. Blood donation screening was the main cost for both scenarios. The One Health approach further allowed savings of €1.21 million in terms of avoided tests on blood units. Benefits of the One Health approach due to short-term costs of hospitalization and compensation for transfusion-associated disease potentially avoided, were estimated to range from €0 to €2.98 million according to the probability of developing WNV neuroinvasive disease after receiving an infected blood transfusion.

Development of a quantitative NS1-capture enzyme-linked immunosorbent assay for early detection of yellow fever virus infection.

Yellow fever is an arboviral disease that causes thousands of deaths every year in Africa and the Americas. However, few commercial diagnostic kits are available. Non-structural protein 1 (NS1) is an early marker of several flavivirus infections and is widely used to diagnose dengue virus (DENV) infection. Nonetheless, little is known about the dynamics of Yellow fever virus (YFV) NS1 expression and secretion, to encourage its use in diagnosis. To tackle this issue, we developed a quantitative NS1-capture ELISA specific for YFV using a monoclonal antibody and recombinant NS1 protein. This test was used to quantify NS1 in mosquito and human cell line cultures infected with vaccine and wild YFV strains. Our results showed that NS1 was detectable in the culture supernatants of both cell lines; however, a higher concentration was maintained as cell-associated rather than secreted into the extracellular milieu. A panel of 73 human samples was used to demonstrate the suitability of YFV NS1 as a diagnostic tool, resulting in 80% sensitivity, 100% specificity, a 100% positive predictive value and a 95.5% negative predictive value compared with RT-PCR. Overall, the developed NS1-capture ELISA showed potential as a promising assay for the detection of early YF infection.

Zika clinical updates: implications for pediatrics.

Zika virus (ZIKV), a mosquito-borne flavivirus, has gained recognition over the past few years as an important new cause of congenital infection. As a result, it is critical that pediatricians understand its epidemiology, clinical presentation, clinical sequelae, and management.

Differential immune-related gene expression in the spleens of duck Tembusu virus-infected goslings.

Flaviviruses pose a significant threat to public health worldwide. Recently, a novel flavivirus, duck Tembusu virus (TMUV), was identified as the causative agent of a serious duck viral disease in Asia. Its rapid spread and expanded host range have raised substantial concerns regarding its potential threat to non-avian hosts, including humans. However, the specific molecular host responses to this virus are poorly understood. In this study, we used the RNA-sequencing technique to analyse the differential gene expression in the spleens of infected goslings 5days post-infection. In total, 2878 upregulated unigenes and 2943 downregulated unigenes were identified. The Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analyses revealed that different pattern recognition receptor (PRR) signalling pathways simultaneously participated in the sensing of the pathogen-associated molecular patterns (PAMPs) of TMUV, and the antigen presentation pathway and acquired immunity were activated. Then, the signals were transduced by the NF-kappa B (NF-κB) or the Janus kinase/signal transducers and activators of transcription (JAK/STAT) pathways, resulting in the enormous production of various cytokines and interferon-stimulated genes (ISGs). We further investigated the immune response patterns in the liver and brain tissue using RT-qPCR. The bacterial peptidoglycan sensor nucleotide-binding oligomerization domain-containing protein 1 (NOD1) receptor was significantly upregulated, especially in the brain tissue, suggesting that NOD1 likely induces an inflammatory response by interacting with dsRNA, which is similar to its actions during hepatitis C viral (HCV) infection. However, major histocompatibility complex II (MHCII) was downregulated only in the spleen, indicating that the downregulation of MHCII in the spleen may be an immune evasion strategy of TMUV to facilitate pathogenesis during infection. Here, we are the first to report a transcriptome analysis of the host immune response to TMUV infection, and the data reported herein may help elucidate the molecular mechanisms of the gosling-TMUV interaction.

Acute acalculous cholecystitis during Zika virus infection in an immunocompromised patient.

In silico design of a Zika virus non-structural protein 5 aiming vaccine protection against zika and dengue in different human populations.

The arboviruses Zika virus (ZIKV) and Dengue virus (DENV) have important epidemiological impact in Brazil and other tropical regions of the world. Recently, it was shown that previous humoral immunity to DENV enhances ZIKV replication in vitro, which may lead to more severe forms of the disease. Thus, traditional approaches of vaccine development aiming to control viral infection through neutralizing antibodies may induce cross-reactive enhancing antibodies. In contrast, cellular immune response was shown to be capable of controlling DENV infection independently of antibodies. The aim of the present study was to design a flavivirus NS5 protein capable of inducing a cellular immune response against DENV and ZIKV.

NS3 helicase from dengue virus specifically recognizes viral RNA sequence to ensure optimal replication.

The protein-RNA interactions within the flavivirus replication complex (RC) are not fully understood. Our structure of dengue virus NS3 adenosine triphosphatase (ATPase)/helicase bound to the conserved 5' genomic RNA 5'-AGUUGUUAGUCU-3' reveals that D290 and R538 make specific interactions with G2 and G5 bases respectively. We show that single-stranded 12-mer RNA stimulates ATPase activity of NS3, however the presence of G2 and G5 leads to significantly higher activation. D290 is adjacent to the DEXH motif found in SF2 helicases like NS3 and interacts with R387, forming a molecular switch that activates the ATPase site upon RNA binding. Our structure guided mutagenesis revealed that disruption of D290-R387 interaction increases basal ATPase activity presumably as a result of higher conformational flexibility of the ATPase active site. Mutational studies also showed R538 plays a critical role in RNA interactions affecting translocation of viral RNA through dynamic interactions with bases at positions 4 and 5 of the ssRNA. Restriction of backbone flexibility around R538 through mutation of G540 to proline abolishes virus replication, indicating conformational flexibility around residue R538 is necessary for RNA translocation. The functionally critical sequence-specific contacts in NS3 RNA binding groove in subdomain III reveals potentially novel allosteric anti-viral drug targets.

Detection and quantification of dengue virus using a novel biosensor system based on dengue NS3 protease activity.

The traditional methods, plaque assays and immuno-focus assays, used to titrate infectious dengue virus (DENV) particles are time consuming and labor intensive. Here, we developed a DENV protease activity detection system (DENPADS) to visualize DENV infection in cells based on dengue protease activity.

Zika virus infection followed by a first episode of psychosis: another flavivirus leading to pure psychiatric symptomatology.

A Call to Introduce Structured Zika Surveillance in India.

India has the climatic conditions conducive to year-round transmission of Zika virus, and a structured disease surveillance program should be implemented to prevent an outbreak. Such a program should (i) start screening before an outbreak arises; (ii) collect baseline data to assess future disease risk and monitor potential birth defects; and (iii) provide new insights into the ecology of the disease and inform public health policy following the one health concept.

West Nile disease in Tunisia: an overview of 60 years.

West Nile virus (WNV) is an arthropod borne virus of public health importance. The virus is a member of the genus Flavivirus and belongs to the Japanese encephalitis virus (JEV) antigenic complex under the Flaviviridae family. The WNV is continuously spreading across Tunisia especially in the coastal and Southern area of the country. The first human West Nile disease (WND) epidemic in Tunisia occurred in 1997, since then, the quantity and the extension of spillover effects increased constantly. However, the existing knowledge of factors triggering such events continues to be rather poor. The last epidemic WNV human meningitis and meningoencephalitis recorded in 2012, with 86 cases and 6 deaths, confirmed the failure of the current system in predicting new cases. This review, based on analysis of scientific papers published between 1970 and 2015, summarises the state of knowledge on WNV in Tunisia and highlights the existing knowledge and research gaps that need to be addressed.

Distribution of Usutu Virus in Germany and Its Effect on Breeding Bird Populations.

Usutu virus (USUV) is an emerging mosquitoborne flavivirus with an increasing number of reports from several countries in Europe, where USUV infection has caused high avian mortality rates. However, 20 years after the first observed outbreak of USUV in Europe, there is still no reliable assessment of the large-scale impact of USUV outbreaks on bird populations. In this study, we identified the areas suitable for USUV circulation in Germany and analyzed the effects of USUV on breeding bird populations. We calculated the USUV-associated additional decline of common blackbird (Turdus merula) populations as 15.7% inside USUV-suitable areas but found no significant effect for the other 14 common bird species investigated. Our results show that the emergence of USUV is a further threat for birds in Europe and that the large-scale impact on population levels, at least for common blackbirds, must be considered.

Peromyscus leucopus mouse brain transcriptome response to Powassan virus infection.

Powassan virus (POWV) is a tick-borne Flavivirus responsible for life-threatening encephalitis in North America and some regions of Russia. The ticks that have been reported to transmit the virus belong to the Ixodes species, and they feed on small-to-medium-sized mammals, such as Peromyscus leucopus mice, skunks, and woodchucks. We previously developed a P. leucopus mouse model of POWV infection, and the model is characterized by a lack of clinical signs of disease following intraperitoneal or intracranial inoculation. However, intracranial inoculation results in mild subclinical encephalitis from 5 days post infection (dpi), but the encephalitis resolves by 28 dpi. We used RNA sequencing to profile the P. leucopus mouse brain transcriptome at different time points after intracranial challenge with POWV. At 24 h post infection, 42 genes were significantly differentially expressed and the number peaked to 232 at 7 dpi before declining to 31 at 28 dpi. Using Ingenuity Pathway Analysis, we determined that the genes that were significantly expressed from 1 to 15 dpi were mainly associated with interferon signaling. As a result, many interferon-stimulated genes (ISGs) were upregulated. Some of the ISGs include an array of TRIMs (genes encoding tripartite motif proteins). These results will be useful for the identification of POWV restriction factors.

Ocular flutter following Zika virus infection.

Zika virus (ZIKV) is an emerging flavivirus which has been linked to a number of neurologic manifestations such as Guillain-Barré syndrome (GBS), transverse myelitis, and meningo-encephalitis. Ophthalmologic manifestations are increasingly being reported; however, ocular dyskinesias have not been described in this context to date. Herein, we report a case of a 22-year-old female who presented with ocular flutter and associated Guillain-Barré syndrome following acute ZIKV infection. We speculate that although such symptoms may have originated from a direct viral insult, a post-infectious autoimmune mechanism may not be excluded. Physicians should include ZIKV as well as other flaviviruses in their diagnostic workup for all patients with ocular flutter/opsoclonus, after excluding other non-infectious causes of central nervous system pathology. To the best of our knowledge, this is the first report on the association of ocular flutter, GBS, and ZIKV infection.

Dengue fever : Symptoms, epidemiology, entomology, pathogen diagnosis and prevention.

Dengue infections are among the most frequent causes of febrile disease in tropical climates. Infections are caused by a flavivirus transmitted by the Aedes mosquito. Aedes aegypti mosquitos are the main transmitters of Dengue viruses. Since these insects are both diurnal and container breeders, particular prevention and control measures are required. Symptom severity varies and can range from a mild, flu-like clinical picture to severe hemorrhage and shock. The most common symptoms experienced by travelers include fever, muscular pain, headaches and skin rash. Depending on the stage of infection, either rapid point-of-care tests or-during the viraemic phase-direct molecular detection of pathogens can be used as diagnostic methods. Serological diagnosis is challenging in terms of interpretation due to serological cross reactions with other flaviviruses.

Behavioral, climatic, and environmental risk factors for Zika and Chikungunya virus infections in Rio de Janeiro, Brazil, 2015-16.

The burden of arboviruses in the Americas is high and may result in long-term sequelae with infants disabled by Zika virus infection (ZIKV) and arthritis caused by infection with Chikungunya virus (CHIKV). We aimed to identify environmental drivers of arbovirus epidemics to predict where the next epidemics will occur and prioritize municipalities for vector control and eventual vaccination. We screened sera and urine samples (n = 10,459) from residents of 48 municipalities in the state of Rio de Janeiro for CHIKV, dengue virus (DENV), and ZIKV by molecular PCR diagnostics. Further, we assessed the spatial pattern of arbovirus incidence at the municipal and neighborhood scales and the timing of epidemics and major rainfall events. Lab-confirmed cases included 1,717 infections with ZIKV (43.8%) and 2,170 with CHIKV (55.4%) and only 29 (<1%) with DENV. ZIKV incidence was greater in neighborhoods with little access to municipal water infrastructure (r = -0.47, p = 1.2x10-8). CHIKV incidence was weakly correlated with urbanization (r = 0.2, p = 0.02). Rains began in October 2015 and were followed one month later by the largest wave of ZIKV epidemic. ZIKV cases markedly declined in February 2016, which coincided with the start of a CHIKV outbreak. Rainfall predicted ZIKV and CHIKV with a lead time of 3 weeks each time. The association between rainfall and epidemics reflects vector ecology as the larval stages of Aedes aegypti require pools of water to develop. The temporal dynamics of ZIKV and CHIKV may be explained by the shorter incubation period of the viruses in the mosquito vector; 2 days for CHIKV versus 10 days for ZIKV.

Zika virus: An updated review of competent or naturally infected mosquitoes.

Zika virus (ZIKV) is an arthropod-borne virus (arbovirus) that recently caused outbreaks in the Americas. Over the past 60 years, this virus has been observed circulating among African, Asian, and Pacific Island populations, but little attention has been paid by the scientific community until the discovery that large-scale urban ZIKV outbreaks were associated with neurological complications such as microcephaly and several other neurological malformations in fetuses and newborns. This paper is a systematic review intended to list all mosquito species studied for ZIKV infection or for their vector competence. We discuss whether studies on ZIKV vectors have brought enough evidence to formally exclude other mosquitoes than Aedes species (and particularly Aedes aegypti) to be ZIKV vectors. From 1952 to August 15, 2017, ZIKV has been studied in 53 mosquito species, including 6 Anopheles, 26 Aedes, 11 Culex, 2 Lutzia, 3 Coquillettidia, 2 Mansonia, 2 Eretmapodites, and 1 Uranotaenia. Among those, ZIKV was isolated from 16 different Aedes species. The only species other than Aedes genus for which ZIKV was isolated were Anopheles coustani, Anopheles gambiae, Culex perfuscus, and Mansonia uniformis. Vector competence assays were performed on 22 different mosquito species, including 13 Aedes, 7 Culex, and 2 Anopheles species with, as a result, the discovery that A. aegypti and Aedes albopictus were competent for ZIKV, as well as some other Aedes species, and that there was a controversy surrounding Culex quinquefasciatus competence. Although Culex, Anopheles, and most of Aedes species were generally observed to be refractory to ZIKV infection, other potential vectors transmitting ZIKV should be explored.

Modelling the effects of phylogeny and body size on within-host pathogen replication and immune response.

Understanding how quickly pathogens replicate and how quickly the immune system responds is important for predicting the epidemic spread of emerging pathogens. Host body size, through its correlation with metabolic rates, is theoretically predicted to impact pathogen replication rates and immune system response rates. Here, we use mathematical models of viral time courses from multiple species of birds infected by a generalist pathogen (West Nile Virus; WNV) to test more thoroughly how disease progression and immune response depend on mass and host phylogeny. We use hierarchical Bayesian models coupled with nonlinear dynamical models of disease dynamics to incorporate the hierarchical nature of host phylogeny. Our analysis suggests an important role for both host phylogeny and species mass in determining factors important for viral spread such as the basic reproductive number, WNV production rate, peak viraemia in blood and competency of a host to infect mosquitoes. Our model is based on a principled analysis and gives a quantitative prediction for key epidemiological determinants and how they vary with species mass and phylogeny. This leads to new hypotheses about the mechanisms that cause certain taxonomic groups to have higher viraemia. For example, our models suggest that higher viral burst sizes cause corvids to have higher levels of viraemia and that the cellular rate of virus production is lower in larger species. We derive a metric of competency of a host to infect disease vectors and thereby sustain the disease between hosts. This suggests that smaller passerine species are highly competent at spreading the disease compared with larger non-passerine species. Our models lend mechanistic insight into why some species (smaller passerine species) are pathogen reservoirs and some (larger non-passerine species) are potentially dead-end hosts for WNV. Our techniques give insights into the role of body mass and host phylogeny in the spread of WNV and potentially other zoonotic diseases. The major contribution of this work is a computational framework for infectious disease modelling at the within-host level that leverages data from multiple species. This is likely to be of interest to modellers of infectious diseases that jump species barriers and infect multiple species. Our method can be used to computationally determine the competency of a host to infect mosquitoes that will sustain WNV and other zoonotic diseases. We find that smaller passerine species are more competent in spreading the disease than larger non-passerine species. This suggests the role of host phylogeny as an important determinant of within-host pathogen replication. Ultimately, we view our work as an important step in linking within-host viral dynamics models to between-host models that determine spread of infectious disease between different hosts.

Visualization of West Nile Virus in Urine Sediment using Electron Microscopy and Immunogold up to Nine Years Postinfection.

West Nile virus (WNV) is an important emerging flavivirus in North America. Experimental studies in animals infer the development of persistent infection in the kidneys. In humans, recent studies suggest the possibility of persistent renal infection and chronic kidney disease. Considering the discrepancies between published studies on viral RNA detection in urine of convalescing WNV-positive patients, we explored the use of electron microscopy (EM) with anti-WNV E protein antibody immunogold labeling to detect virus in the urine sediment from a subset of study participants in the Houston WNV cohort. In 42% of evaluated study participants had visible sediment present in urine after centrifugation; viral particles consistent with the size and morphology of WNV were successfully detected using EM in the urine sediment up to 9 years postinfection. The anti-WNV immunogold labeling bound to virus envelope in the sediment allowed for enhanced detection when compared with PCR and provide a new technique for understanding kidney disease in WNV patients. These results provide further evidence of persistent infection in at least a subset of individuals infected with WNV. These findings present a novel tool to diagnose persistent WNV infection and its possible link with progressive renal pathology.

Resource Use and Costs of Dengue: Analysis of Data from Phase III Efficacy Studies of a Tetravalent Dengue Vaccine.

A tetravalent dengue vaccine (CYD-TDV) has recently been approved in 12 countries in southeast Asia and Latin America for individuals aged 9-45 years or 9-60 years (age indication approvals vary by country) living in endemic areas. Data on utilization of medical and nonmedical resources as well as time lost from school and work were collected during the active phase of two phase III efficacy studies performed in 10 countries in the Asia-Pacific region and Latin America (NCT01373281; NCT01374516). We compared dengue-related resource utilization and costs among vaccinated and nonvaccinated participants. Country-specific unit costs were derived from available literature. There were 901 virologically confirmed dengue episodes among participants aged ≥ 9 years (N = 25,826): corresponding to 373 episodes in the CYD-TDV group (N = 17,230) and 528 episodes in the control group (N = 8,596). Fewer episodes in the CYD-TDV group resulted in hospitalization than in the control group (7.0% versus 13.3%; P = 0.002), but both had a similar average length of stay of 4 days. Overall, a two-thirds reduction in resource consumption and missed school/work days was observed in the CYD-TDV group relative to the control group. The estimated direct and indirect cost (2014 I$) associated with dengue episodes per participant in the CYD-TDV group was 73% lower than in the control group (I$6.72 versus I$25.08); representing a saving of I$I8.36 (95% confidence interval [CI]:17.05-19.78) per participant with vaccination. This is the first study providing information on dengue costs among vaccinated individuals and direct confirmation that vaccination has the potential to reduce dengue illness costs.

Guillain-Barré Syndrome, Acute Disseminated Encephalomyelitis and Encephalitis Associated with Zika Virus Infection in Brazil: Detection of Viral RNA and Isolation of Virus during Late Infection.

Zika virus (ZIKV) emerged in Brazil in 2015, which was followed by an increase of Guillain-Barre Syndrome (GBS) cases. We report the epidemiological, clinical, and laboratory findings of the first six neurological cases associated with ZIKV in Brazil seen in a reference neurology hospital in Pernambuco, Brazil. In all cases, ZIKV was detected in serum and/or cerebrospinal fluid (CSF) samples. In this case series, four cases were defined as GBS, one as acute disseminated encephalomyelitis (ADEM) and the other as encephalitis. ZIKV was detected in all cases by RT-PCR and virus isolation was successful in two patients. The time between ZIKV acute symptoms and the development of neurological manifestations varied from 3 to 13 days and ZIKV was detected between 15 and 34 days after the initial symptoms. Our results highlight the need to include ZIKV as a differential diagnosis for neurological syndromes in countries with circulation of this arbovirus. Because the viremia in these patients appears to persist longer, direct diagnostic techniques such as RT-PCR and viral isolation should be considered even if it is after the acute phase of viral infection.

Functionalized 2,1-benzothiazine 2,2-dioxides as new inhibitors of Dengue NS5 RNA-dependent RNA polymerase.

Over recent years, many RNA viruses have been "re-discovered", including life-threatening flaviviruses, such as Dengue, Zika, and several encephalitis viruses. Since no specific inhibitors are currently available to treat these infections, there is a pressing need for new therapeutics. Among the flaviviral proteins, NS5 RNA-dependent RNA polymerase (RdRp) represents a validated target being essential for viral replication and it has no human analog. To date, few NS5 RdRp inhibitor chemotypes have been reported and no inhibitors are currently in clinical development. In this context, after an in vitro screening against Dengue 3 NS5 RdRp of our in-house HCV NS5B inhibitors focused library, we found that 2,1-benzothiazine 2,2-dioxides are promising non-nucleoside inhibitors of flaviviral RdRp with compounds 8 and 10 showing IC50 of 0.6 and 0.9 μM, respectively. Preliminary structure-activity relationships indicated a key role for the C-4 benzoyl group and the importance of a properly functionalized C-6 phenoxy moiety to modulate potency. Compound 8 acts as non-competitive inhibitor and its proposed pose in the so-called N pocket of the RdRp thumb domain allowed to explain the key contribution of the benzoyl and the phenoxy moieties for the ligand binding.