RESUMO
Unbiased high-throughput sequencing (HTS) has enabled new insights into the diversity of agents implicated in central nervous system (CNS) infections. The addition of positive selection capture methods to HTS has enhanced the sensitivity while reducing sequencing costs and the complexity of bioinformatic analysis. Here we report the use of virus capture-based sequencing for vertebrate viruses (VirCapSeq-VERT) and bacterial capture sequencing (BacCapSeq) in investigating CNS infections. Thirty-four samples were categorized: (1) patients with definitive CNS infection by routine testing; (2) patients meeting clinically the Brighton criteria (BC) for meningoencephalitis; (3) patients with presumptive infectious etiology highest on the differential. RNA extracts from cerebrospinal fluid (CSF) were used for VirCapSeq-VERT, and DNA extracts were used for BacCapSeq analysis. Among 8 samples from known CNS infections in group 1, VirCapSeq and BacCapSeq confirmed 3 expected diagnoses (42.8%), were negative in 2 (25%), yielded an alternative result in 1 (11.1%), and did not detect 2 expected negative pathogens. The confirmed cases identified HHV-6, HSV-2, and VZV while the negative samples included JCV and HSV-2. In groups 2 and 3, 11/26 samples (42%) were positive for at least one pathogen; however, 27% of the total samples (7/26) were positive for commensal organisms. No microbial nucleic acids were detected in negative control samples. HTS showed limited promise for pathogen identification in presumed CNS infectious diseases in our small sample. Before conducting larger-scale prospective studies to assess the clinical value of this novel technique, clinicians should understand the benefits and limitations of using this modality.
Assuntos
Meningoencefalite , Vírus , Humanos , Estudos Prospectivos , Sequenciamento de Nucleotídeos em Larga Escala/métodos , Herpesvirus Humano 2/genéticaRESUMO
Background: Unbiased high-throughput sequencing (HTS) has enabled new insights into the diversity of agents implicated in central nervous system (CNS) infections. The addition of positive selection capture methods to HTS has enhanced the sensitivity while reducing sequencing costs and complexity of bioinformatic analysis. Here we report the use of virus capture based sequencing for vertebrate viruses (VirCapSeq-VERT) and bacterial capture sequencing (BacCapSeq) in investigating CNS infections. Design/Methods: Thirty-four samples were categorized: (1) Patients with definitive CNS infection by routine testing; (2) Patients meeting clinically Brighton Criteria (BC) for meningoencephalitis (3) Patients with presumptive infectious etiology highest on the differential. RNA extracts from cerebrospinal fluid (CSF) were used for VirCapSeq-VERT and DNA extracts were used for BacCapSeq analysis. Results: Among 8 samples from known CNS infections in group 1, VirCapSeq and BacCapSeq confirmed 3 expected diagnoses (42.8%), were negative in 2 (25%), yielded an alternative result in 1 (11.1%), and did not detect 2 expected negative pathogens. The confirmed cases identified HHV-6, HSV-2, and VZV while the negative samples included JCV and HSV-2. In groups 2 and 3,11/26 samples (42%) were positive for at least one pathogen, however 27% of the total samples (7/26) were positive for commensal organisms. No microbial nucleic acids were detected in negative control samples. Conclusions: HTS showed limited promise for pathogen identification in presumed CNS infectious diseases in our small sample. Before conducting larger-scale prospective studies to assess clinical value of this novel technique, clinicians should understand benefits and limitations of using this modality.
RESUMO
In this study, the syntheses of a BaCuO3 perovskite-structured oxide and rGO were conducted using a sol-gel method and ultrasonication process, respectively. Their physico-chemical characteristics were studied by powder X-ray diffraction (PXRD), Raman spectroscopy, and ultraviolet-visible (UV-vis) spectroscopy analyses. The perovskite type particles were found to present as a cubic crystal system with a crystal size in the range of 10-60 nm. Their catalytic performance was investigated using differential thermal analysis (DTA) measurements at varying heating rates via the thermal decomposition of ammonium perchlorate (AP) and 3-nitro-1,2,4-triazol-5-one (NTO). The kinetics and thermodynamics parameters, such as the average activation energy, pre-exponential factor, entropy, and Gibbs free energy, were also investigated, which showed a decrease in the decomposition peak temperature of AP and NTO in the presence of the catalysts rGO, BaCuO3, and rGO + BaCuO3 compared to pure AP and NTO.
RESUMO
Immunocompromised individuals are at risk of prolonged SARS-CoV-2 infection due to weaker immunity, co-morbidities, and lowered vaccine effectiveness, which may evolve highly mutated variants of SARS-CoV-2. Nonetheless, limited data are available on the immune responses elicited by SARS-CoV-2 infection, reinfections, and vaccinations with emerging variants in immunocompromised patients. We analyzed clinical samples that were opportunistically collected from eight immunocompromised individuals for mutations in SARS-CoV-2 genomes, neutralizing antibody (NAb) titers against different SARS-CoV-2 variants, and the identification of immunoreactive epitopes using a high-throughput coronavirus peptide array. The viral genome analysis revealed two SARS-CoV-2 variants (20A from a deceased patient and an Alpha variant from a recovered patient) with an eight amino-acid (aa) deletion within the N-terminal domain (NTD) of the surface glycoprotein. A higher NAb titer was present against the prototypic USA/WA1/2020 strain in vaccinated immunocompromised patients. NAb titer was absent against the Omicron variant and the cultured virus of the 20A variant with eight aa deletions in non-vaccinated patients. Our data suggest that fatal SARS-CoV-2 infections may occur in immunocompromised individuals even with high titers of NAb post-vaccination. Moreover, persistent SARS-CoV-2 infection may lead to the emergence of newer variants with additional mutations favoring the survival and fitness of the pathogen that include deletions in NAb binding sites in the SARS-CoV-2 surface glycoprotein.
Assuntos
COVID-19 , Vacinas Virais , Humanos , Hospedeiro Imunocomprometido , Glicoproteínas de Membrana , SARS-CoV-2/genéticaRESUMO
IMPORTANCE: We describe the first report to our knowledge of cutaneous and systemic pathogenicity of human polyomavirus 9 in solid organ transplant recipients. OBJECTIVE: Three solid organ transplant recipients developed a widespread, progressive, violaceous, and hyperkeratotic skin eruption. All died from pulmonary and multiorgan failure around 1 year from onset of the rash. Routine clinical diagnostic testing could not identify any causative agent; therefore, samples and autopsies were investigated for novel pathogens using high-throughput sequencing. DESIGN, SETTING, AND PARTICIPANTS: This case series, including 3 solid organ transplant recipients who developed characteristic pink, violaceous, or brown hyperkeratotic papules and plaques throughout the body, was conducted at the Columbia University Medical Center. Lesional skin biopsies were collected from all 3 patients and subjected to high-throughput illumina sequencing for identification of microbial pathogens. Human polyomavirus 9 was identified in lesional skin biopsies. We subsequently collected ocular swabs, oral swabs, urine samples, and blood samples from patients, and organ tissues at autopsy in 1 patient. We investigated these samples for the presence of human polyomavirus 9 using in situ hybridization and quantitative polymerase chain reaction (PCR) assays. MAIN OUTCOMES AND MEASURES: A description of the clinical and pathologic findings of 3 patients. RESULTS: This case series study found that human polyomavirus 9 was detected in the skin biopsies of all 3 patients by a capture-based high-throughput sequencing method platform (VirCapSeq-VERT). Human polyomavirus 9 was also detected in blood, oral, ocular swabs, and urine by real-time polymerase chain reaction (PCR) assay. In situ hybridization and quantitative PCR assays were performed on the skin biopsies from 3 patients and lung autopsy of 1 patient, which showed the presence of human polyomavirus 9 messenger RNA transcripts, indicating active viral replication and pathogenesis in the skin and lungs. CONCLUSIONS AND RELEVANCE: Human polyomavirus 9 was associated with the widespread cutaneous eruption. All 3 patients had progression of cutaneous disease, accompanied by clinical deterioration, pulmonary failure, and death. One patient underwent autopsy and human polyomavirus 9 was identified in the lungs and paratracheal soft tissue. These findings suggest that human polyomavirus 9 may be associated with cutaneous and possibly pulmonary infection and death in solid organ transplant recipients.
Assuntos
Exantema , Transplante de Órgãos , Infecções por Polyomavirus , Polyomavirus , Dermatopatias , DNA Viral/análise , Humanos , Pulmão , Transplante de Órgãos/efeitos adversos , Polyomaviridae , Polyomavirus/genética , Reação em Cadeia da Polimerase em Tempo Real , TransplantadosRESUMO
A 71-year-old woman previously on rituximab treatment for rheumatoid arthritis presented with 2 years of progressive neurologic symptoms. She was found to have persistent hypogammaglobulinemia and B cell depletion despite rituximab discontinuation a year prior. MRI revealed diffuse meningeal enhancement along the entire neuroaxis. LP showed a CSF lymphocytic pleocytosis, elevated protein, and presence of enterovirus by PCR. The patient was hospitalized several times for progressive clinical and radiologic decline, though she had transient improvements following treatment with immunoglobulin therapy. Her CSF remained positive for enterovirus PCR for at least 12 months. Though two brain biopsies were non-diagnostic, pan-Enterovirus was ultimately identified using a high-throughput next-generation sequencing technique. She was treated with compassionate-use pocapavir with clinical stabilization at 4-month follow-up; however, she expired 8 months later from a bacterial pneumonia.
Assuntos
Infecções por Enterovirus , Enterovirus , Meningoencefalite , Idoso , Enterovirus/genética , Infecções por Enterovirus/tratamento farmacológico , Feminino , Humanos , Imunização Passiva , Éteres Fenílicos , Rituximab/uso terapêuticoRESUMO
Serodiagnosis of SARS-CoV-2 infection is impeded by immunological cross-reactivity among the human coronaviruses (HCoVs): SARS-CoV-2, SARS-CoV-1, MERS-CoV, OC43, 229E, HKU1, and NL63. Here we report the identification of humoral immune responses to SARS-CoV-2 peptides that may enable discrimination between exposure to SARS-CoV-2 and other HCoVs. We used a high-density peptide microarray and plasma samples collected at two time points from 50 subjects with SARS-CoV-2 infection confirmed by qPCR, samples collected in 2004-2005 from 11 subjects with IgG antibodies to SARS-CoV-1, 11 subjects with IgG antibodies to other seasonal human coronaviruses (HCoV), and 10 healthy human subjects. Through statistical modeling with linear regression and multidimensional scaling we identified specific peptides that were reassembled to identify 29 linear SARS-CoV-2 epitopes that were immunoreactive with plasma from individuals who had asymptomatic, mild or severe SARS-CoV-2 infections. Larger studies will be required to determine whether these peptides may be useful in serodiagnostics.
Assuntos
COVID-19/imunologia , COVID-19/virologia , Mapeamento de Peptídeos , Peptídeos/imunologia , SARS-CoV-2/fisiologia , Sequência de Aminoácidos , Animais , COVID-19/sangue , Quirópteros , Epitopos/imunologia , Humanos , Imunoglobulina G/metabolismo , Peptídeos/química , Proteoma/metabolismoRESUMO
ABSTRACT Objective: To compare the effect of Maleic acid and Irritrol (combination of Chlorhexidine & Ethylenediaminetetraacetic acid) irrigation on the sealing ability of Biodentine when used as root-end filling material. Material and Methods: Thirty single-rooted human premolars were subjected to standardized root canal instrumentation and were irrigated with 2.5% Sodium Hypochlorite (NaOCl) after each instrument change. Apical root resections were done by removing 3 mm of the apex at a 90° angle to the long axis of the root with a diamond bur. The root end cavity preparation was done using ultrasonic tips at a low power setting under water coolant. They were randomly divided into 3 groups (n=10). Group 1 was irrigated with 7% Maleic acid, Group 2 was irrigated with Irritrol, and Group 3 was irrigated with 0.9% Saline (Control). Then, the root end cavities of all the samples were restored with Biodentine and were subjected to leakage analysis using a glucose filtration test. The mean values for each group were calculated and the obtained data was statistically analysed using ANOVA with post-hoc Tukey's test, ANOVA with post-hoc Games-Howell test and repeated measures ANOVA with post-hoc Bonferroni test. Results: The specimens irrigated with Irritrol showed least microleakage as compared to 7% Maleic acid and 0.9% Saline. Conclusion: A regimen involving irrigation of the root-end cavity with Irritrol followed by root-end filling with Biodentine can improve the apical seal.
Assuntos
Humanos , Ácidos , Preparo de Canal Radicular/instrumentação , Materiais Dentários , Cavidade Pulpar , Dente Pré-Molar , Radiografia Dentária/instrumentação , Análise de Variância , ÍndiaRESUMO
BACKGROUND: The second largest Ebola virus disease (EVD) outbreak began in the Democratic Republic of Congo in July 2018 in North Kivu Province. Data suggest the outbreak is not epidemiologically linked to the 2018 outbreak in Equateur Province, and that independent introduction of Ebola virus (EBOV) into humans occurred. We tested for antibodies to ebolaviruses in febrile patients seeking care in North Kivu Province prior to the EVD outbreak. METHODS: Patients were enrolled between May 2017 and April 2018, before the declared start of the outbreak in eastern DRC. Questionnaires were administered to collect demographic and behavioural information to identify risk factors for exposure. Biological samples were evaluated for ebolavirus nucleic acid, and for antibodies to ebolaviruses. Prevalence of exposure was calculated, and demographic factors evaluated for associations with ebolavirus serostatus. RESULTS: Samples were collected and tested from 272 people seeking care in the Rutshuru Health Zone in North Kivu Province. All patients were negative for filoviruses by PCR. Intial screening by indirect ELISA found that 30 people were reactive to EBOV-rGP. Results were supported by detection of ebolavirus reactive linear peptides using the Serochip platform. Differential screening of all reactive serum samples against the rGP of all six ebolaviruses and Marburg virus (MARV) showed that 29 people exhibited the strongest reactivity to EBOV and one to Bombali virus (BOMV), and western blotting confirmed results. Titers ranged from 1:100 to 1:12,800. Although both sexes and all ages tested positive for antibodies, women were significantly more likely to be positive and the majority of positives were in February 2018. CONCLUSIONS: We provide the first documented evidence of exposure to Ebola virus in people in eastern DRC. We detected antibodies to EBOV in 10% of febrile patients seeking healthcare prior to the declaration of the 2018-2020 outbreak, suggesting early cases may have been missed or exposure ocurred without associated illness. We also report the first known detection of antibodies to BOMV, previously detected in bats in West and East Africa, and show that human exposure to BOMV has occurred. Our data suggest human exposure to ebolaviruses may be more frequent and geographically widespread.
RESUMO
Maternal Zika virus (ZIKV) infection during pregnancy can have profound teratogenic effects including microcephaly and intracranial calcification. In adults, infection may cause acute inflammatory polyneuropathy. These complications appear after resolution of viremia. Thus, molecular assays of blood are typically insufficient to make the link between ZIKV infection and disease. An alternative approach to testing for ZIKV exposure is serology. However, specific serological implication of ZIKV can be confounded by cross-reactivity with closely related flaviviruses. We used high-density peptide arrays that tile the proteomes of a selection of arboviruses to search for ZIKV-specific linear epitopes that would enable the development of accurate serological tests. We identified a 20-amino-acid-long diagnostic peptide sequence in the NS2B protein of ZIKV that was immunoreactive with sera from patients with a confirmed history of infection with ZIKV but not other flaviviruses. We then established a ZIKV peptide ELISA (ZIKV-NS2B-concat ELISA) that enables sensitive and specific diagnosis of exposure to ZIKV.
Assuntos
Imunoglobulina G/isolamento & purificação , Testes Sorológicos/métodos , Proteínas não Estruturais Virais/imunologia , Infecção por Zika virus/diagnóstico , Zika virus/imunologia , Adulto , Anticorpos Antivirais/análise , Anticorpos Antivirais/sangue , Anticorpos Antivirais/isolamento & purificação , Especificidade de Anticorpos , Biotinilação , Ensaio de Imunoadsorção Enzimática/métodos , Epitopos/química , Epitopos/imunologia , Feminino , Humanos , Imunoglobulina G/sangue , Fragmentos de Peptídeos/química , Fragmentos de Peptídeos/imunologia , Gravidez , Complicações Infecciosas na Gravidez/sangue , Complicações Infecciosas na Gravidez/diagnóstico , Proteínas não Estruturais Virais/química , Infecção por Zika virus/sangue , Infecção por Zika virus/virologiaRESUMO
Rotaviruses are the most common infectious agents causing severe diarrheal diseases in young children globally. Three rare human rotavirus strains, two G3P[9] and one G3P[6], were detected in stool samples of children under 5 years of age hospitalized for gastroenteritis in Lebanon during the course of a surveillance study. Complete genomes of these strains were sequenced using VirCapSeq-VERT, a capture based high-throughput sequencing method. Genomic sequences were further characterized by using phylogenetic analyses with global RVA G3P[6]/P[9] strains, other vaccine and reference strains. Genetic analysis revealed that the G3P[6] strain emerged as a DS-1/Wa-like mono-reassortant strain with a potential Ethiopian origin. The two G3P[9] strains possessed a mixed DS-1/Wa/AU-1-like origin indicating that these may have evolved via multiple reassortment events involving feline, human and bovine rotaviruses. Furthermore, analysis of these strains revealed high antigenic variability compared to the vaccine strains. Additional studies are essential to fully understand the evolutionary dynamics of G3P[6]/P[9] strains spreading worldwide and their implications on vaccine effectiveness.
Assuntos
Infecções por Rotavirus/virologia , Rotavirus/genética , Pré-Escolar , Epitopos/imunologia , Gastroenterite/epidemiologia , Gastroenterite/virologia , Genoma Viral , Genótipo , Humanos , Lactente , Líbano/epidemiologia , Filogenia , Vírus Reordenados/genética , Rotavirus/imunologia , Rotavirus/isolamento & purificação , Infecções por Rotavirus/epidemiologia , Vacinas contra Rotavirus/imunologiaRESUMO
Borrelia miyamotoi is an emerging tick-borne spirochete transmitted by ixodid ticks. Current serologic assays for B. miyamotoi are impacted by genetic similarities to other Borrelia and limited understanding of optimal antigenic targets. In this study, we employed the TBD-Serochip, a peptide array platform, to identify new linear targets for serologic detection of B. miyamotoi. We examined a wide range of suspected B. miyamotoi antigens and identified 352 IgM and 91 IgG reactive peptides, with the majority mapping to variable membrane proteins. These included peptides within conserved fragments of variable membrane proteins that may have greater potential for differential diagnosis. We also identified reactive regions on FlaB, and demonstrate crossreactivity of B. burgdorferi s.l. C6 with a B. miyamotoi C6-like peptide. The panel of linear peptides identified in this study can be used to enhance serodiagnosis of B. miyamotoi.
Assuntos
Borrelia/fisiologia , Epitopos/isolamento & purificação , Testes Sorológicos/instrumentaçãoRESUMO
BACKGROUND: Recent emergence of Zika virus (ZIKV), and the global spread of dengue (DENV), chikungunya (CHIKV) and West Nile viruses (WNV) raised urgent need of accurate and affordable molecular diagnosis of these clinically indistinguishable arboviral infections. OBJECTIVES: We established a pentaplex real-time reverse transcription PCR (rRT-PCR) assay (CII-ArboViroPlex rRT-PCR) for specific and sensitive detection of the African and American genotypes of ZIKV, all four serotypes of DENV, CHIKV, WNV and a housekeeping gene as internal control in single reaction. STUDY DESIGN: Specific primers and probe sets were designed for ZIKV, DENV, CHIKV, WNV and RNase P (housekeeping gene) and tested for in-vitro transcribed RNA standards, virus cultures, clinical samples positive for ZIKV, DENV, CHIKV and WNV and limit of detection (LOD) were determined for each. Results Using ten-fold serially diluted in-vitro transcribed RNA, CII- ArboViroPlex rRT-PCR assay has LOD of 100 RNA copies/reaction (Rn) for ZIKV in serum or urine, 100 RNA copies/Rn for DENV in serum, and 10 RNA copies/Rn for CHIKV and WNV in serum. LODs from sera spiked with quantitated viral stocks were 2.6â¯×â¯102 GEQ/Rn for ZIKV, 2.2â¯×â¯101 GEQ/Rn for DENV-1, 9.4â¯×â¯100 GEQ/Rn for DENV-2, 2.3â¯×â¯102 GEQ/Rn for DENV-3, 1.4â¯×â¯103 GEQ/Rn for DENV-4, 2.7â¯×â¯102 GEQ/Rn for CHIKV, and 1.05â¯×â¯101 GEQ/Rn for WNV. CONCLUSIONS: The CII-ArboViroPlex rRT-PCR assay is a quantitative one-step pentaplex rRT-PCR assay for the molecular detection and differential diagnosis of ZIKV, DENV, CHIKV, WNV and a human housekeeping gene control in a single- PCR reaction.
Assuntos
Febre de Chikungunya/diagnóstico , Dengue/diagnóstico , Reação em Cadeia da Polimerase Multiplex/métodos , Febre do Nilo Ocidental/diagnóstico , Linhagem Celular , Vírus Chikungunya/genética , Vírus da Dengue/genética , Genes Essenciais , Humanos , Limite de Detecção , Técnicas de Diagnóstico Molecular/métodos , RNA Viral/sangue , RNA Viral/urina , Reação em Cadeia da Polimerase em Tempo Real , Vírus do Nilo Ocidental , Zika virus/genética , Infecção por Zika virus/diagnósticoRESUMO
Acute flaccid myelitis (AFM) has caused motor paralysis in >560 children in the United States since 2014. The temporal association of enterovirus (EV) outbreaks with increases in AFM cases and reports of fever, respiratory, or gastrointestinal illness prior to AFM in >90% of cases suggest a role for infectious agents. Cerebrospinal fluid (CSF) from 14 AFM and 5 non-AFM patients with central nervous system (CNS) diseases in 2018 were investigated by viral-capture high-throughput sequencing (VirCapSeq-VERT system). These CSF and serum samples, as well as multiple controls, were tested for antibodies to human EVs using peptide microarrays. EV RNA was confirmed in CSF from only 1 adult AFM case and 1 non-AFM case. In contrast, antibodies to EV peptides were present in CSF of 11 of 14 AFM patients (79%), significantly higher than controls, including non-AFM patients (1/5 [20%]), children with Kawasaki disease (0/10), and adults with non-AFM CNS diseases (2/11 [18%]) (P = 0.023, 0.0001, and 0.0028, respectively). Six of 14 CSF samples (43%) and 8 of 11 sera (73%) from AFM patients were immunoreactive to an EV-D68-specific peptide, whereas the three control groups were not immunoreactive in either CSF (0/5, 0/10, and 0/11; P = 0.008, 0.0003, and 0.035, respectively) or sera (0/2, 0/8, and 0/5; P = 0.139, 0.002, and 0.009, respectively).IMPORTANCE The presence in cerebrospinal fluid of antibodies to EV peptides at higher levels than non-AFM controls supports the plausibility of a link between EV infection and AFM that warrants further investigation and has the potential to lead to strategies for diagnosis and prevention of disease.
Assuntos
Anticorpos Antivirais/líquido cefalorraquidiano , Viroses do Sistema Nervoso Central/líquido cefalorraquidiano , Enterovirus Humano D/imunologia , Enterovirus Humano D/isolamento & purificação , Infecções por Enterovirus/líquido cefalorraquidiano , Mielite/líquido cefalorraquidiano , Doenças Neuromusculares/líquido cefalorraquidiano , Adolescente , Adulto , Anticorpos Antivirais/sangue , Anticorpos Antivirais/imunologia , Antígenos Virais/imunologia , Viroses do Sistema Nervoso Central/sangue , Criança , Enterovirus Humano D/genética , Infecções por Enterovirus/sangue , Feminino , Sequenciamento de Nucleotídeos em Larga Escala , Humanos , Masculino , Pessoa de Meia-Idade , Mielite/sangue , Doenças Neuromusculares/sangue , Análise Serial de Proteínas , RNA Viral/sangue , RNA Viral/líquido cefalorraquidiano , RNA Viral/genética , Adulto JovemRESUMO
Brazil, which is hyperendemic for dengue virus (DENV), has had recent Zika (ZIKV) and (CHIKV) Chikungunya virus outbreaks. Since March 2016, CHIKV is the arbovirus infection most frequently diagnosed in Rio de Janeiro. In the analysis of 1835 syndromic patients, screened by real time RT-PCR, 56.4% of the cases were attributed to CHIKV, 29.6% to ZIKV, and 14.1% to DENV-4. Sequence analyses of CHIKV from sixteen samples revealed that the East-Central-South-African (ECSA) genotype of CHIKV has been circulating in Brazil since 2013 [95% bayesian credible interval (BCI): 03/2012-10/2013], almost a year before it was detected by arbovirus surveillance program. Brazilian cases are related to Central African Republic sequences from 1980's. To the best of our knowledge, given the available sequence published here and elsewhere, the ECSA genotype was likely introduced to Rio de Janeiro early on 2014 (02/2014; BCI: 07/2013-08/2014) through a single event, after primary circulation in the Bahia state at the Northestern Brazil in the previous year. The observation that the ECSA genotype of CHIKV was circulating undetected underscores the need for improvements in molecular methods for viral surveillance.
Assuntos
Febre de Chikungunya/diagnóstico , Vírus Chikungunya/genética , Teorema de Bayes , Brasil/epidemiologia , Febre de Chikungunya/epidemiologia , Febre de Chikungunya/virologia , Vírus Chikungunya/classificação , Vírus Chikungunya/isolamento & purificação , Genótipo , Sequenciamento de Nucleotídeos em Larga Escala , Humanos , Filogenia , RNA Viral/química , RNA Viral/metabolismo , Análise de Sequência de RNARESUMO
Zika virus (ZIKV) is implicated in fetal stillbirth, microcephaly, intracranial calcifications, and ocular anomalies following vertical transmission from infected mothers. In adults, infection may trigger autoimmune inflammatory polyneuropathy. Transmission most commonly follows the bite of infected Aedes mosquitoes but may also occur through sexual intercourse or receipt of blood products. Definitive diagnosis through detection of viral RNA is possible in serum or plasma within 10 days of disease onset, in whole blood within 3 weeks of onset, and in semen for up to 3 months. Serological diagnosis is nonetheless critical because few patients have access to molecular diagnostics during the acute phase of infection and infection may be associated with only mild or inapparent disease that does not prompt molecular testing. Serological diagnosis is confounded by cross-reactivity of immune sera with other flaviviruses endemic in the areas where ZIKV has recently emerged. Accordingly, we built a high-density microarray comprising nonredundant 12-mer peptides that tile, with one-residue overlap, the proteomes of Zika, dengue, yellow fever, West Nile, Ilheus, Oropouche, and chikungunya viruses. Serological analysis enabled discovery of a ZIKV NS2B 20-residue peptide that had high sensitivity (96.0%) and specificity (95.9%) versus natural infection with or vaccination against dengue, chikungunya, yellow fever, West Nile, tick-borne encephalitis, or Japanese encephalitis virus in a microarray assay and an enzyme-linked immunosorbent assay (ELISA) of early-convalescent-phase sera (2 to 3 weeks after onset of symptomatic infection).IMPORTANCE The emergence of Zika virus (ZIKV) as a teratogen is a profound challenge to global public health. Molecular diagnosis of infection is straightforward during the 3-week period when patients are viremic. However, serological diagnosis thereafter of historical exposure has been confounded by cross-reactivity. Using high-density peptide arrays that tile the proteomes of a selection of flaviviruses to identify a ZIKV-specific peptide, we established two assays that enable sensitive and specific diagnosis of exposure to ZIKV. These assays may be useful in guiding clinical management of mothers at risk for potential exposure to ZIKV and enable insights into the epidemiology of ZIKV infections.