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1.
Rev Bras Epidemiol ; 24: e210020, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-33825776

RESUMO

OBJECTIVE: To describe the entry of Dengue virus (DENV) serotypes in Brazil and its federative units. METHODS: A systematic review of studies published between 1980 and 2018 in databases and in the gray literature was performed using descriptors related to the years of entry of the DENV serotypes. Additionally, experts and official sources of information (Brazilian Ministry of Health) were consulted. RESULTS: From 100 publications selected for the systematic review, 26 addressed the entry of DENV serotypes in the North region of the country, 33 in the Northeast, 24 in the Southeast, 14 in the Central-West, and five in the South. DENV-1 and DENV-4 were introduced in the North region in 1981. DENV-2 was introduced in the Southeast in 1990. DENV-3 was introduced in the North in 1999. CONCLUSION: The rapid expansion of dengue throughout the Brazilian territory was verified from the second half of the 1980s, with the gradual entry of the four serotypes, which resulted in the emergence of epidemics of arbovirus, which are currently verified in the country. Considering the epidemiology of the disease, more information should be disseminated and published in the wide-ranging scientific literature for a better understanding of the spread and circulation of DENV serotypes.


Assuntos
Vírus da Dengue , Dengue , Brasil/epidemiologia , Dengue/epidemiologia , Vírus da Dengue/genética , Geografia , Humanos , Sorogrupo
2.
Nat Commun ; 12(1): 1810, 2021 03 22.
Artigo em Inglês | MEDLINE | ID: mdl-33753725

RESUMO

For most pathogens, transmission is driven by interactions between the behaviours of infectious individuals, the behaviours of the wider population, the local environment, and immunity. Phylogeographic approaches are currently unable to disentangle the relative effects of these competing factors. We develop a spatiotemporally structured phylogenetic framework that addresses these limitations by considering individual transmission events, reconstructed across spatial scales. We apply it to geocoded dengue virus sequences from Thailand (N = 726 over 18 years). We find infected individuals spend 96% of their time in their home community compared to 76% for the susceptible population (mainly children) and 42% for adults. Dynamic pockets of local immunity make transmission more likely in places with high heterotypic immunity and less likely where high homotypic immunity exists. Age-dependent mixing of individuals and vector distributions are not important in determining spread. This approach provides previously unknown insights into one of the most complex disease systems known and will be applicable to other pathogens.


Assuntos
Algoritmos , Vírus da Dengue/genética , Dengue/transmissão , Modelos Teóricos , Adulto , Aedes/virologia , Animais , Criança , Dengue/epidemiologia , Dengue/virologia , Vírus da Dengue/classificação , Vírus da Dengue/fisiologia , Genoma Viral/genética , Interações Hospedeiro-Patógeno , Humanos , Mosquitos Vetores/virologia , Filogenia , Filogeografia/métodos , Filogeografia/estatística & dados numéricos , Dinâmica Populacional , Tailândia/epidemiologia
3.
Arch Virol ; 166(4): 1103-1112, 2021 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-33575893

RESUMO

Dengue virus (DV) is a mosquito-borne virus that is endemic to many tropical and subtropical areas. Recently, the annual incidence of DV infection has increased worldwide, including in Korea, due to global warming and increased global travel. We therefore sought to characterize the molecular and evolutionary features of DV-1 and DV-4 isolated from Korean overseas travelers. We used phylogenetic analysis based on the full coding region to classify isolates of DV-1 in Korea into genotype I (43251, KP406802), genotype IV (KP406803), and genotype V (KP406801). In addition, we found that strains of DV-4 belonged to genotype I (KP406806) and genotype II (43257). Evidence of positive selection in DV-1 strains was identified in the C, prM, NS2A, and NS5 proteins, whereas DV-4 showed positive selection only in the non-structural proteins NS2A, NS3, and NS5. The substitution rates per site per year were 5.58 × 10-4 and 6.72 × 10-4 for DV-1 and DV-4, respectively, and the time of the most recent common ancestor was determined using the Bayesian skyline coalescent method. In this study, the molecular, phylogenetic, and evolutionary characteristics of Korean DV-1 and DV-4 isolates were evaluated for the first time.


Assuntos
Vírus da Dengue/genética , Dengue/virologia , Evolução Molecular , Viagem , Sequência de Aminoácidos , Substituição de Aminoácidos , Sequência de Bases , Dengue/epidemiologia , Vírus da Dengue/classificação , Vírus da Dengue/isolamento & purificação , Genótipo , Humanos , Filogenia , RNA Viral/genética , República da Coreia/epidemiologia , Seleção Genética , Sorogrupo , Proteínas Virais/genética
4.
Mem Inst Oswaldo Cruz ; 115: e200287, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-33533869

RESUMO

BACKGROUND: The heat-labile nature of Dengue virus (DENV) in serum samples must be considered when applying routine diagnostic tests to avoid issues that could impact the accuracy of test results with direct implications for case management and disease reporting. OBJECTIVES: To check if pre-analytical variables, such as storage time and temperature, have an impact on the accuracy of the main routine diagnostic tests for dengue. METHODS: Virus isolation, reverse transcription real-time polymerase chain reaction (RT-PCR) and NS1 enzyme-linked immunosorbent assay (ELISA) were evaluated using 84 samples submitted to different pre-analytical conditions. FINDINGS: Sensitivity and negative predictive value were directly affected by sample storage conditions. RT-PCR and virus isolation showed greater dependence on well-conserved samples for an accurate diagnosis. Interestingly, even storage at -30ºC for a relatively short time (15 days) was not adequate for accurate results using virus isolation and RT-PCR tests. On the other hand, NS1 ELISA showed no significant reduction in positivity for aliquots tested under the same conditions as in the previous tests. MAIN CONCLUSIONS: Our results support the stability of the NS1 marker in ELISA diagnosis and indicate that the accuracy of routine tests such as virus isolation and RT-PCR is significantly affected by inadequate transport and storage conditions of serum samples.


Assuntos
Antígenos Virais/sangue , Vírus da Dengue/isolamento & purificação , Dengue/diagnóstico , Ensaio de Imunoadsorção Enzimática/métodos , Testes Imunológicos/métodos , Reação em Cadeia da Polimerase Via Transcriptase Reversa/métodos , Proteínas não Estruturais Virais/imunologia , Anticorpos Antivirais/sangue , Antígenos Virais/imunologia , Dengue/sangue , Dengue/virologia , Vírus da Dengue/genética , Vírus da Dengue/imunologia , Humanos , Valor Preditivo dos Testes , Sensibilidade e Especificidade , Proteínas não Estruturais Virais/genética
5.
BMC Infect Dis ; 21(1): 166, 2021 Feb 10.
Artigo em Inglês | MEDLINE | ID: mdl-33568111

RESUMO

BACKGROUND: An unexpected dengue outbreak occurred in Hunan Province in 2018. This was the first dengue outbreak in this area of inland China, and 172 cases were reported. METHODS: To verify the causative agent of this outbreak and characterise the viral genes, the genes encoding the structural proteins C/prM/E of viruses isolated from local residents were sequenced followed by mutation and phylogenetic analysis. Recombination, selection pressure, potential secondary structure and three-dimensional structure analyses were also performed. RESULTS: Phylogenetic analysis revealed that all epidemic strains were of the cosmopolitan DENV-2 genotype and were most closely related to the Zhejiang strain (MH010629, 2017) and then the Malaysia strain (KJ806803, 2013). Compared with the sequence of DENV-2SS, 151 base substitutions were found in the sequences of 89 isolates; these substitutions resulted in 20 non-synonymous mutations, of which 17 mutations existed in all samples (two in the capsid protein, six in the prM/M proteins, and nine in the envelope proteins). Moreover, amino acid substitutions at the 602nd (E322:Q → H) and 670th (E390: N → S) amino acids may have enhanced the virulence of the epidemic strains. One new DNA binding site and five new protein binding sites were observed. Two polynucleotide binding sites and seven protein binding sites were lost in the epidemic strains compared with DENV-2SS. Meanwhile, five changes were found in helical regions. Minor changes were observed in helical transmembrane and disordered regions. The 429th amino acid of the E protein switched from a histamine (positively charged) to an asparagine (neutral) in all 89 isolated strains. No recombination events or positive selection pressure sites were observed. To our knowledge, this study is the first to analyse the genetic characteristics of epidemic strains in the first dengue outbreak in Hunan Province in inland China. CONCLUSIONS: The causative agent is likely to come from Zhejiang Province, a neighbouring province where dengue fever broke out in 2017. This study may help clarify the intrinsic geographical relatedness of DENV-2 and contribute to further research on pathogenicity and vaccine development.


Assuntos
Vírus da Dengue/genética , Dengue/diagnóstico , Proteínas do Envelope Viral/genética , Sítios de Ligação , Proteínas do Capsídeo/química , Proteínas do Capsídeo/genética , Proteínas do Capsídeo/metabolismo , China/epidemiologia , Dengue/epidemiologia , Dengue/virologia , Vírus da Dengue/classificação , Vírus da Dengue/isolamento & purificação , Surtos de Doenças , Genótipo , Humanos , Mutação , Filogenia , Estrutura Terciária de Proteína , RNA Viral/química , RNA Viral/metabolismo , Análise de Sequência de RNA , Proteínas do Envelope Viral/química , Proteínas não Estruturais Virais/genética , Proteínas não Estruturais Virais/metabolismo
6.
Arch Virol ; 166(3): 863-870, 2021 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-33495898

RESUMO

A dengue virus serotype 1 (DENV-1) epidemic occurred from October to December 2018 in Xishuangbanna, Yunnan, Southwest China, neighboring Myanmar, Laos, and Vietnam. In this study, we investigated the molecular characteristics, evolution, and potential source of DENV from Xishuangbanna. The C (capsid), prM (premembrane), and E (envelope) genes of DENV isolated from 87 serum samples obtained from local patients were amplified and sequenced, and the sequences were evaluated by identification of mutations, phylogenetic and homologous recombination analysis, and secondary structure prediction. Phylogenetic analysis showed that all of the epidemic DENV strains from Xishuangbanna could be grouped in a branch with DENV-1 isolates, and were most similar to the Fujian 2005 (China, DQ193572) and Singapore 2016 (MF314188) strains. When compared with DENV-1SS (the standard strain), there were 31 non-synonymous mutations, but no obvious homologous recombination signal was found. Secondary structure prediction showed that some changes had occurred in a helical region in proteins of the MN123849 and MN123854 strains, but there were few changes in the disordered region. This study reveals the molecular characteristics of the structural genes of the Xishuangbanna epidemic strains in 2018 and provides a reference for molecular epidemiology, infection, and pathogenicity research and vaccine development.


Assuntos
Proteínas do Capsídeo/genética , Vírus da Dengue/genética , Dengue/epidemiologia , Proteínas do Envelope Viral/genética , Sequência de Aminoácidos , China/epidemiologia , Vírus da Dengue/classificação , Vírus da Dengue/isolamento & purificação , Surtos de Doenças , Genótipo , Humanos , Epidemiologia Molecular , Filogenia , RNA Viral/genética , Alinhamento de Sequência , Análise de Sequência de RNA , Sorogrupo
7.
BMC Infect Dis ; 21(1): 104, 2021 Jan 22.
Artigo em Inglês | MEDLINE | ID: mdl-33482756

RESUMO

BACKGROUND: Dengue virus (DENV) infection is increasingly common in southern China and can be transmitted through blood transfusion but is not currently part of donor screening throughout the region. We assessed DENV prevalence among donors at the Xishuangbanna Blood Center, Yunnan, to support development of DENV screening strategies. METHODS: Blood samples were collected randomly between June 2019 and August 2019. These were screened for anti-DENV IgG and IgM using enzyme-linked immunosorbent assay (ELISA). Then, all reactive samples and some randomly-chosen non-reactive samples were used to detect DENV RNAs using real-time polymerase-chain-reaction (RT-PCR) assays. After RT-PCR, samples were further tested for soluble nonstructural protein 1 (NS1) using the colloidal gold method. Donors demographics were also collected and assessed. RESULTS: Over the study period, 2254 donor samples were collected and tested for anti-DENV IgG and IgM by ELISA. This revealed 598 anti-DENV IgG and/or IgM reactive samples, a serological prevalence of 26.53%. Of these, 26 were RT-PCR positive and/or NS1 positive. Significant differences in DENV prevalence were noted by occupation (P = 0.001), education (P < 0.001), and ethnicity (P = 0.026). CONCLUSION: The prevalence of DENV in Xishuangbanna Blood Center was higher than most other blood centers that have implemented DENV donor screening. Our study provides first-hand data about the prevalence of DENV and allows the development of a screening strategy for clinical use.


Assuntos
Doadores de Sangue , Vírus da Dengue/isolamento & purificação , Dengue/diagnóstico , Dengue/epidemiologia , Programas de Rastreamento/métodos , Adulto , Anticorpos Antivirais/sangue , China/epidemiologia , Dengue/sangue , Vírus da Dengue/genética , Vírus da Dengue/imunologia , Feminino , Humanos , Masculino , Programas de Rastreamento/estatística & dados numéricos , Pessoa de Meia-Idade , Prevalência , RNA Viral/genética , Proteínas não Estruturais Virais/genética , Adulto Jovem
8.
BMC Infect Dis ; 20(1): 948, 2020 Dec 11.
Artigo em Inglês | MEDLINE | ID: mdl-33308178

RESUMO

BACKGROUND: Dengue patients develop different disease severity ranging from mild (dengue fever [DF]) to severe forms (dengue hemorrhagic fever [DHF] and the fatal dengue shock syndrome [DSS]). Host genetics are considered to be one factor responsible for the severity of dengue outcomes. To identify genes associated with dengue severity that have not been studied yet, we performed genetic association analyses of interferon lambda 3 (IFNL3), CD27, and human leukocyte antigen-DPB1 (HLA-DPB1) genes in Thai dengue patients. METHODS: A case-control association study was performed in 877 children (age ≤ 15 years) with dengue infection (DF, n = 386; DHF, n = 416; DSS, n = 75). A candidate single nucleotide polymorphism of each of IFNL3, CD27, and HLA-DPB1 was selected to be analyzed. Genotyping was performed by TaqMan real-time PCR assay, and the association with dengue severity was examined. RESULTS: The rs9277534 variant of HLA-DPB1 was weakly associated with DHF. The genotype GG and G allele conferred protection against DHF (p = 0.04, odds ratio 0.74 for GG genotype, p = 0.03, odds ratio 0.79 for G allele). The association became borderline significant after adjusting for confounders (p = 0.05, odds ratio 0.82). No association was detected for IFNL3 or CD27. CONCLUSIONS: The present study demonstrated the weak association of the rs9277534 variant of HLA-DPB1 with protection against DHF. This variant is in the 3' untranslated region and affects HLA-DPB1 surface protein expression. Our finding suggests that HLA-DPB1 may be involved in DHF pathogenesis.


Assuntos
Vírus da Dengue/genética , Vírus da Dengue/imunologia , Cadeias beta de HLA-DP/genética , Interferons/genética , Dengue Grave/epidemiologia , Dengue Grave/genética , Índice de Gravidade de Doença , Membro 7 da Superfamília de Receptores de Fatores de Necrose Tumoral/genética , Regiões 3' não Traduzidas/genética , Adolescente , Alelos , Estudos de Casos e Controles , Criança , Vírus da Dengue/isolamento & purificação , Feminino , Estudos de Associação Genética , Genótipo , Humanos , Masculino , Razão de Chances , Polimorfismo de Nucleotídeo Único , Dengue Grave/virologia , Tailândia/epidemiologia
9.
Zhonghua Liu Xing Bing Xue Za Zhi ; 41(12): 2119-2124, 2020 Dec 10.
Artigo em Chinês | MEDLINE | ID: mdl-33378826

RESUMO

Objective: To analyze the epidemiological and etiological characteristics of a dengue fever outbreak in Hunan province in 2018. Methods: Real-time PCR assay was performed for the laboratory diagnosis of 8 suspected dengue fever cases. Etiological surveillance was performed in 186 suspected dengue fever cases and fever cases who had close contacts with dengue fever patients. C6/36 cells was used for the virus isolation from acute phase serum. By sequencing the full length of E genes of 15 dengue virus strains, phylogenetic analysis was performed based on the sequences obtained, including reference sequences from the NCBI GenBank database, the serotypes and gene subtypes of the virus were analyzed to trace the possible source of transmission. An emergency monitoring of vector density and a retrospective survey of sero-epidemiology in healthy population were conducted in the epidemic area. Results: In the serum samples of 8 suspected patients, 6 were dengue virus RNA positive, and 4 were NS1 antigen positive. In 186 suspected patients, 96 were dengue virus nucleic acid, NS1 antigen or antibody positive in etiological test. A total of 64 dengue virus strains were isolated. The phylogenetic analysis showed that all the dengue virus strains belonged to type 2, which might be from Guangdong or Zhejiang provinces. The Bretub index was up to 65, indicating an extremely high risk of transmission. The positive rate of the dengue virus IgG antibody was 0.53%(2/377) in retrospective survey of 377 healthy people. Conclusion: The field epidemiologic and the molecular genetics analyses showed the outbreak of dengue fever in Hunan in 2018 was caused by imported cases and dengue virus 2.


Assuntos
Vírus da Dengue , Dengue , Surtos de Doenças , China/epidemiologia , Dengue/epidemiologia , Dengue/virologia , Vírus da Dengue/genética , Humanos , Filogenia , Estudos Retrospectivos
10.
BMC Infect Dis ; 20(1): 835, 2020 Nov 11.
Artigo em Inglês | MEDLINE | ID: mdl-33176708

RESUMO

BACKGROUND: The spatial distribution and burden of dengue in sub-Saharan Africa remains highly uncertain, despite high levels of ecological suitability. The goal of this study was to describe the epidemiology of dengue among a cohort of febrile children presenting to outpatient facilities located in areas of western Uganda with differing levels of urbanicity and malaria transmission intensity. METHODS: Eligible children were first screened for malaria using rapid diagnostic tests. Children with a negative malaria result were tested for dengue using a combination NS1/IgM/IgG rapid test (SD Bioline Dengue Duo). Confirmatory testing by RT-PCR was performed in a subset of participants. Antigen-capture ELISA was performed to estimate seroprevalence. RESULTS: Only 6 of 1416 (0.42%) children had a positive dengue rapid test, while none of the RT-PCR results were positive. ELISA testing demonstrated reactive IgG antibodies in 28 (2.2%) participants with the highest prevalence seen at the urban site in Mbarara (19 of 392, 4.9%, p < 0.001). CONCLUSIONS: Overall, these findings suggest that dengue, while present, is an uncommon cause of non-malarial, pediatric febrile illness in western Uganda. Further investigation into the eocological factors that sustain low-level transmission in urban settings are urgently needed to reduce the risk of epidemics.


Assuntos
Vírus da Dengue/genética , Vírus da Dengue/imunologia , Dengue/diagnóstico , Dengue/epidemiologia , Febre/diagnóstico , Adolescente , Criança , Pré-Escolar , Dengue/virologia , Testes Diagnósticos de Rotina/efeitos adversos , Ensaio de Imunoadsorção Enzimática , Feminino , Humanos , Imunoglobulina G/imunologia , Imunoglobulina M/imunologia , Malária/diagnóstico , Malária/epidemiologia , Malária/parasitologia , Masculino , Plasmodium/imunologia , Plasmodium/isolamento & purificação , Prevalência , Estudos Prospectivos , Reação em Cadeia da Polimerase em Tempo Real , Estudos Soroepidemiológicos , Uganda/epidemiologia
11.
PLoS One ; 15(10): e0227239, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-33064724

RESUMO

Species of the genus Flavivirus are widespread in Brazil and are a major public health concern. The country's largest city, São Paulo, is in a highly urbanized area with a few forest fragments which are commonly used for recreation. These can be considered to present a potential risk of flavivirus transmission to humans as they are home simultaneously to vertebrate hosts and mosquitoes that are potential flavivirus vectors. The aim of this study was to conduct flavivirus surveillance in field-collected mosquitoes in the Capivari-Monos Environmental Protection Area (EPA) and identify the flavivirus species by sequence analysis in flavivirus IFA-positive pools. Monthly mosquito collections were carried out from March 2016 to April 2017 with CO2-baited CDC light traps. Specimens were identified morphologically and grouped in pools of up to 10 individuals according to their taxonomic category. A total of 260 pools of non-engorged females were inoculated into C6/36 cell culture, and the cell suspensions were analyzed by indirect immunofluorescence assay (IFA) after the incubation period. IFA-positive pools were tested by qRT-PCR with genus-specific primers targeting the flavivirus NS5 gene to confirm IFA-positive results and sequenced to identify the species. Anopheles cruzii (19.5%) and Wyeomyia confusa (15.3%) were the most frequent vector species collected. IFA was positive for flaviviruses in 2.3% (6/260) of the sample pools. This was confirmed by qRT-PCR in five pools (83.3%). All five flavivirus-positive pools were successfully sequenced and the species identified. DENV serotype 2 (DENV-2) was detected in Culex spp. and Culex vaxus pools, while ZIKV was identified in An. cruzii, Limatus durhamii and Wy. confusa pools. To the best of our knowledge, detection of flavivirus species of medical importance has never previously been reported in these species of wild-caught mosquitoes. The finding of DENV-2 and ZIKV circulating in wild mosquitoes suggests the existence of an enzootic cycle in the area. In-depth studies of DENV-2 and ZIKV, including investigation of mosquito infection, vector competence and infection in sylvatic hosts, are needed to shed light on the transmission dynamics of these important viruses and the potential risk of future outbreaks of DENV-2 and ZIKV infections in the region.


Assuntos
Vírus da Dengue/isolamento & purificação , Mosquitos Vetores/virologia , Proteínas não Estruturais Virais/genética , Zika virus/isolamento & purificação , Animais , Anopheles/virologia , Brasil/epidemiologia , Culex/virologia , Vírus da Dengue/genética , Feminino , Vigilância da População , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Meio Selvagem , Zika virus/genética
12.
PLoS One ; 15(10): e0238609, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-33112881

RESUMO

INTRODUCTION: Although immune responses to the Japanese Encephalitis virus (JEV), and the dengue viruses (DENV) have a potential to modulate the immune responses to each other, this has been poorly investigated. Therefore, we developed an ELISA to identify JEV specific, DENV non cross-reactive antibody responses by identifying JEV specific, highly conserved regions of the virus and proceeded to investigate if the presence of JEV specific antibodies associate with dengue disease severity. METHODOLOGY AND RESULTS: 22 JEV specific peptides were identified from highly conserved regions of the virus and the immunogenicity and specificity of these peptides were assessed in individuals who were non-immune to JEV and DENV (JEV-DENV-, N = 30), those who were only immune to the JEV and not DENV (JEV+DENV-, N = 30), those who were only immune to DENV(JEV-DENV+, N = 30) and in those who were immune to both viruses (JEV+DENV+, N = 30). 7/22 peptides were found to be highly immunogenic and specific and these 7 peptides were used as a pool to further evaluate JEV-specific responses. All 30/30 JEV+DENV- and 30/30 JEV+DENV+ individuals, and only 3/30 (10%) JEV-DENV+ individuals responded to this pool. We further evaluated this pool of 7 peptides in patients following primary and secondary dengue infection during the convalescent period and found that the JEV-specific peptides, were unlikely to cross react with DENV IgG antibodies. We further compared this in-house ELISA developed with the peptide pool with an existing commercial JEV IgG assay to identify JEV-specific IgG following vaccination, and our in-house ELISA was found to be more sensitive. We then proceeded to investigate if the presence of JEV-specific antibodies were associated with dengue disease severity, and we found that those who had past severe dengue (n = 175) were significantly more likely (p<0.0001) to have JEV-specific antibodies than those with past non-severe dengue (n = 175) (OR 5.3, 95% CI 3.3 to 8.3). CONCLUSIONS: As our data show that this assay is highly sensitive and specific for detection of JEV-specific antibody responses, it would be an important tool to determine how JEV seropositivity modulate dengue immunity and disease severity when undertaking dengue vaccine trials.


Assuntos
Anticorpos Antivirais/sangue , Vírus da Encefalite Japonesa (Espécie)/imunologia , Ensaio de Imunoadsorção Enzimática/métodos , Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , Sequência de Aminoácidos , Especificidade de Anticorpos , Criança , Sequência Conservada , Reações Cruzadas , Dengue/imunologia , Dengue/virologia , Vírus da Dengue/classificação , Vírus da Dengue/genética , Vírus da Dengue/imunologia , Vírus da Encefalite Japonesa (Espécie)/genética , Encefalite Japonesa/epidemiologia , Encefalite Japonesa/imunologia , Encefalite Japonesa/virologia , Ensaio de Imunoadsorção Enzimática/estatística & dados numéricos , Feminino , Voluntários Saudáveis , Humanos , Imunoglobulina G/sangue , Masculino , Pessoa de Meia-Idade , Fragmentos de Peptídeos/genética , Fragmentos de Peptídeos/imunologia , Sensibilidade e Especificidade , Estudos Soroepidemiológicos , Sorogrupo , Sri Lanka/epidemiologia , Vacinação , Proteínas Virais/genética , Proteínas Virais/imunologia , Adulto Jovem
13.
Sci Adv ; 6(39)2020 09.
Artigo em Inglês | MEDLINE | ID: mdl-32978154

RESUMO

Detection of viruses is critical for controlling disease spread. Recent emerging viral threats, including Zika virus, Ebola virus, and SARS-CoV-2 responsible for coronavirus disease 2019 (COVID-19) highlight the cost and difficulty in responding rapidly. To address these challenges, we develop a platform for low-cost and rapid detection of viral RNA with DNA nanoswitches that mechanically reconfigure in response to specific viruses. Using Zika virus as a model system, we show nonenzymatic detection of viral RNA with selective and multiplexed detection between related viruses and viral strains. For clinical-level sensitivity in biological fluids, we paired the assay with sample preparation using either RNA extraction or isothermal preamplification. Our assay requires minimal laboratory infrastructure and is adaptable to other viruses, as demonstrated by quickly developing DNA nanoswitches to detect SARS-CoV-2 RNA in saliva. Further development and field implementation will improve our ability to detect emergent viral threats and ultimately limit their impact.


Assuntos
Betacoronavirus/genética , Infecções por Coronavirus/diagnóstico , DNA de Cadeia Simples/genética , Eletroforese em Gel de Ágar/métodos , Pneumonia Viral/diagnóstico , RNA Viral/genética , Análise de Sequência de RNA/métodos , Sequência de Bases , Linhagem Celular Tumoral , Infecções por Coronavirus/virologia , Dengue/diagnóstico , Dengue/virologia , Vírus da Dengue/genética , Eletroforese em Gel de Ágar/economia , Humanos , Limite de Detecção , Pandemias , Pneumonia Viral/virologia , Saliva/virologia , Análise de Sequência de RNA/economia , Zika virus/genética , Infecção por Zika virus/diagnóstico , Infecção por Zika virus/virologia
14.
Nucleic Acids Res ; 48(17): 9872-9885, 2020 09 25.
Artigo em Inglês | MEDLINE | ID: mdl-32890404

RESUMO

The ribosomal stalk proteins, RPLP1 and RPLP2 (RPLP1/2), which form the ancient ribosomal stalk, were discovered decades ago but their functions remain mysterious. We had previously shown that RPLP1/2 are exquisitely required for replication of dengue virus (DENV) and other mosquito-borne flaviviruses. Here, we show that RPLP1/2 function to relieve ribosome pausing within the DENV envelope coding sequence, leading to enhanced protein stability. We evaluated viral and cellular translation in RPLP1/2-depleted cells using ribosome profiling and found that ribosomes pause in the sequence coding for the N-terminus of the envelope protein, immediately downstream of sequences encoding two adjacent transmembrane domains (TMDs). We also find that RPLP1/2 depletion impacts a ribosome density for a small subset of cellular mRNAs. Importantly, the polarity of ribosomes on mRNAs encoding multiple TMDs was disproportionately affected by RPLP1/2 knockdown, implying a role for RPLP1/2 in multi-pass transmembrane protein biogenesis. These analyses of viral and host RNAs converge to implicate RPLP1/2 as functionally important for ribosomes to elongate through ORFs encoding multiple TMDs. We suggest that the effect of RPLP1/2 at TMD associated pauses is mediated by improving the efficiency of co-translational folding and subsequent protein stability.


Assuntos
Fosfoproteínas/metabolismo , Proteínas Ribossômicas/metabolismo , Proteínas do Envelope Viral/genética , Células A549 , Animais , Chlorocebus aethiops , Vírus da Dengue/genética , Vírus da Dengue/metabolismo , Humanos , Proteínas de Membrana/química , Proteínas de Membrana/genética , Proteínas de Membrana/metabolismo , Fosfoproteínas/genética , Domínios Proteicos , Proteínas Ribossômicas/genética , Ribossomos/metabolismo , Células Vero , Proteínas do Envelope Viral/química , Proteínas do Envelope Viral/metabolismo
15.
Euro Surveill ; 25(36)2020 09.
Artigo em Inglês | MEDLINE | ID: mdl-32914745

RESUMO

In August 2020, during the coronavirus disease (COVID-19) pandemic, five locally acquired cases of dengue virus type 1 were detected in a family cluster in Vicenza Province, North-East Italy where Aedes albopictus mosquitoes are endemic. The primary case was an importation from West Sumatra, Indonesia. This is the first outbreak of autochthonous dengue reported in Italy. During the COVID-19 pandemic, screening of febrile travelers from endemic countries is crucial in areas where competent vectors are present.


Assuntos
Vírus da Dengue/isolamento & purificação , Dengue/diagnóstico , Viagem , Adulto , Pré-Escolar , Dengue/epidemiologia , Dengue/imunologia , Dengue/virologia , Vírus da Dengue/genética , Surtos de Doenças , Transmissão de Doença Infecciosa , Feminino , Febre/etiologia , Humanos , Imunoglobulina G/sangue , Imunoglobulina M/sangue , Indonésia , Itália/epidemiologia , Masculino , Pessoa de Meia-Idade , RNA Viral/sangue , Reação em Cadeia da Polimerase Via Transcriptase Reversa
16.
Am J Trop Med Hyg ; 103(5): 2054-2058, 2020 11.
Artigo em Inglês | MEDLINE | ID: mdl-32876014

RESUMO

The incidence and spread of dengue virus (DENV) have increased rapidly in recent decades. Dengue is underreported in Africa, but recent outbreaks and seroprevalence data suggest that DENV is widespread there. A lack of ongoing surveillance limits knowledge about its spatial reach and hinders disease control planning. We sought to add data on dengue distribution in Kenya through diagnostic testing of serum specimens from persons with an acute febrile illness (AFI) attending an outpatient clinic in rural western Kenya (Asembo) during rainy seasons. Patients with symptoms not likely to be misclassified as dengue (e.g., diarrhea and anemia), those with a positive diagnostic laboratory results which explained their febrile illness, or those with serum collected more than 5 days after fever onset were excluded. However, febrile patients with a positive malaria smear were included in the study. We used reverse transcription polymerase chain reaction (RT-PCR) to test for DENV and IgM anti-DENV to test for recent infection. Of the 615 serum specimens available for testing, none were dengue positive by either RT-PCR or IgM anti-DENV testing. Dengue did not appear to be a cause of febrile illness in this area of western Kenya, although our relatively small sample size may not have identified DENV infections occurring at low incidence. A more widespread AFI surveillance system that includes dengue diagnostic testing by RT-PCR and antibody-based methods is required to more definitively gauge the size and geographic distribution of DENV infection in western Kenya.


Assuntos
Vírus da Dengue/imunologia , Dengue/epidemiologia , Doença Aguda/epidemiologia , Adolescente , Adulto , Criança , Pré-Escolar , Dengue/virologia , Vírus da Dengue/genética , Monitoramento Epidemiológico , Feminino , Febre , Humanos , Lactente , Quênia/epidemiologia , Masculino , Adulto Jovem
17.
Proc Natl Acad Sci U S A ; 117(37): 22823-22832, 2020 09 15.
Artigo em Inglês | MEDLINE | ID: mdl-32868439

RESUMO

Conjugation of RNAs with nanoparticles (NPs) is of significant importance because of numerous applications in biology and medicine, which, however, remains challenging especially for large ones. So far, the majority of RNA labeling relies on solid-phase chemical synthesis, which is generally limited to RNAs smaller than 100 nucleotides (nts). We, here, present an efficient and generally applicable labeling strategy for site-specific covalent conjugation of large RNAs with a gold nanoparticle (Nanogold) empowered by transcription of an expanded genetic alphabet containing the A-T/U and G-C natural base pairs (bps) and the TPT3-NaM unnatural base pair (UBP). We synthesize an amine-derivatized TPT3 (TPT3A), which is site specifically incorporated into a 97-nt 3'SL RNA and a 719-nt minigenomic RNA (DENV-mini) from Dengue virus serotype 2 (DENV2) by in vitro T7 transcription. The TPT3A-modified RNAs are covalently conjugated with mono-Sulfo-N-hydroxysuccinimidyl (NHS)-Nanogold NPs via an amine and NHS ester reaction and further purified under nondenaturing conditions. TPT3 modification and Nanogold labeling cause minimal structural perturbations to the RNAs by circular dichroism, small angle X-ray scattering (SAXS), and binding activity assay. We demonstrate the application of the Nanogold-RNA conjugates in large RNA structural biology by an emerging molecular ruler, X-ray scattering interferometry (XSI). The internanoparticle distance distributions in the 3'SL and DENV-mini RNAs derived from XSI measurements support the hypothetical model of flavivirus genome circularization, thus, validate the applicability of this labeling strategy. The presented strategy overcomes the size constraints in conventional RNA labeling strategies and is expected to have wide applications in large RNA structural biology and RNA nanotechnology.


Assuntos
Vírus da Dengue/genética , Ouro/química , Nanopartículas Metálicas/química , RNA Viral/química , RNA Viral/genética , Vírus da Dengue/química , Espalhamento a Baixo Ângulo , Transcrição Genética
18.
Braz J Infect Dis ; 24(5): 452-454, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32866435

RESUMO

Many regions of the world where dengue epidemics are seasonal are also facing the COVID-19 pandemic. This is a medical concern because both diseases are difficult to distinguish since they have similar clinical symptoms and laboratory findings, and because they have different clinical management. So far, co-infection of SARS-CoV-2 and dengue virus (DENV) has not been studied. Herein we report the first case of a patient with co-infection of COVID-19 and dengue. Both infections were simultaneously laboratory confirmed by positive RT-qPCR for SARS-CoV-2 and RT-qPCR for DENV, NS1, IgM and IgG antibody tests for dengue. The patient had a favorable clinical improvement, without severe symptoms. This case emphasize that, in pandemic era, having a diagnostic of one infection does not rule out the possibility of having another infection concomitantly. In addition, underscores the importance of an accurate and timely diagnosis to prevent the spread of COVID-19.


Assuntos
Coinfecção , Infecções por Coronavirus , Vírus da Dengue , Dengue , Pandemias , Pneumonia Viral , Betacoronavirus , Técnicas de Laboratório Clínico , Dengue/complicações , Dengue/diagnóstico , Vírus da Dengue/genética , Humanos
19.
PLoS One ; 15(8): e0237384, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32764809

RESUMO

Dengue fever is one of the major public health problems in Lao PDR. Over the last decade, dengue virus (DENV) epidemics were characterized by a novel predominant serotype accompanied by at least two other serotypes. Since 2008, DENV-2 circulated at a low level in Lao PDR but its epidemiologic profile changed at the end of 2018. Indeed, the number of confirmed DENV-2 cases suddenly increased in October 2018 and DENV-2 became predominant at the country level in early 2019. We developed a Genotype Screening Protocol (GSP) to determine the origin(s) of the Lao DENV-2 and study their genetic polymorphism. With a good correlation with full envelope gene sequencing data, this molecular epidemiology tool evidence the co-circulation of two highly polymorphic DENV-2 genotypes, i.e. Asian I and Cosmopolitan genotypes, over the last five years, suggesting multiple introductions of DENV-2 in the country. GSP approach provides relevant first line information that may help countries with limited laboratory resources to reinforce their capabilities to DENV-2 and to follow the epidemics progresses and assess situations at the regional level.


Assuntos
Vírus da Dengue/genética , Técnicas de Genotipagem/métodos , Inquéritos e Questionários , Vírus da Dengue/isolamento & purificação , Vírus da Dengue/fisiologia , Humanos , Laos , Sorotipagem , Fatores de Tempo , População Urbana/estatística & dados numéricos
20.
PLoS One ; 15(7): e0234959, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32663230

RESUMO

The economic and social impacts due to diseases transmitted by mosquitoes in the latest years have been significant. Currently, no specific treatment or commercial vaccine exists for the control and prevention of arboviruses, thereby making entomological characterization fundamental in combating diseases such as dengue, chikungunya, and Zika. The morphological identification of mosquitos includes a visual exam of the samples. It is time consuming and requires adequately trained professionals. Accordingly, the development of a new automated method for realizing mosquito-perception and -classification is becoming increasingly essential. Therefore, in this study, a computational model based on a convolutional neural network (CNN) was developed to extract features from the images of mosquitoes and then classify the species Aedes aegypti, Aedes albopictus, and Culex quinquefasciatus. In addition, the model was trained to detect the mosquitoes of the genus Aedes. To train CNNs to perform the automatic morphological classification of mosquitoes, a dataset, which included 7,561 images of the target mosquitoes and 1,187 images of other insects, was acquired. Various neural networks, such as Xception and DenseNet, were used for developing the automatic-classification model based on images. A structured optimization process of random search and grid search was developed to select the hyperparameters set and increase the accuracy of the model. In addition, strategies to eliminate overfitting were implemented to increase the generalization of the model. The optimized model, during the test phase, obtained the balanced accuracy (BA) of 93.5% in classifying the target mosquitoes and other insects and the BA of 97.3% in detecting the mosquitoes of the genus Aedes in comparison to Culex. The results provide fundamental information for performing the automatic morphological classification of mosquito species. Using a CNN-embedded entomological tool is a valuable and accessible resource for health workers and non-taxonomists for identifying insects that can transmit infectious diseases.


Assuntos
Arbovirus/classificação , Culicidae/classificação , Processamento de Imagem Assistida por Computador/métodos , Aedes/virologia , Animais , Automação Laboratorial/métodos , Febre de Chikungunya/transmissão , Vírus Chikungunya/genética , Culex/virologia , Culicidae/genética , Dengue/transmissão , Vírus da Dengue/genética , Feminino , Masculino , Mosquitos Vetores/virologia , Zika virus/genética , Infecção por Zika virus/transmissão
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