RESUMO
The prompt and accurate identification of the etiological agents of viral respiratory infections is a critical measure in mitigating outbreaks. In this study, we developed and clinically evaluated a novel melting-curve-based multiplex real-time PCR (M-m-qPCR) assay targeting the RNA-dependent RNA polymerase (RdRp) and nucleocapsid phosphoprotein N of SARS-CoV-2, the Matrix protein 2 of the Influenza A virus, the RdRp domain of the L protein from the Human Respiratory Syncytial Virus, and the polyprotein from Rhinovirus B genes. The analytical performance of the M-m-qPCR underwent assessment using in silico analysis and a panel of reference and clinical strains, encompassing viral, bacterial, and fungal pathogens, exhibiting 100% specificity. Moreover, the assay showed a detection limit of 10 copies per reaction for all targeted pathogens using the positive controls. To validate its applicability, the assay was further tested in simulated nasal fluid spiked with the viruses mentioned above, followed by validation on nasopharyngeal swabs collected from 811 individuals. Among them, 13.4% (109/811) tested positive for SARS-CoV-2, and 1.1% (9/811) tested positive for Influenza A. Notably, these results showed 100% concordance with those obtained using a commercial kit. Therefore, the M-m-qPCR exhibits great potential for the routine screening of these respiratory viral pathogens.
RESUMO
OBJECTIVES: We aimed to evaluate the distribution, clinical presentations and severity of common acute respiratory infections (ARI) viruses in infants across 3 clinical settings. STUDY DESIGN: In a prospective virus surveillance study, infants under 1 year with fever and/or respiratory symptoms were enrolled from outpatient, emergency department, and inpatient settings from December 16, 2019 through April 30, 2020. Demographic and clinical characteristics were collected through parent/guardian interviews, medical chart abstractions, and follow-up surveys. Nasal swabs were collected and tested for viruses using quantitative reverse-transcription polymerase chain reaction. RESULTS: We enrolled 366 infants and tested nasal swabs on 360 (98%); median age was 6.3 months, 50% male. In total, 295 (82%) had at least 1 virus detected; rhinovirus/enterovirus (RV/EV; 42%), respiratory syncytial virus (RSV; 34%), and influenza (15%) were the most common. RSV was the most frequently detected virus in the inpatient (63%) and emergency department (37%) settings, and RV/EV was most frequently detected virus in the outpatient setting (54%). RSV-positive infants had a lower median age (4.9 months) and were more likely to have respiratory distress, and RV/EV-positive infants were less likely to have respiratory distress. Influenza-positive infants had a higher median age (8 months) and were more likely to have systemic symptoms. RSV infection and younger age were associated with higher odds of hospitalization in multivariable logistic regression. CONCLUSIONS: Across 3 clinical settings, and combining virologic, patient, and health-system information, our results highlight the burden of viral ARI among infants. Overall, RSV, RV/EV, and influenza were most commonly detected, with RSV having the highest disease severity.
Assuntos
Influenza Humana/epidemiologia , Infecções por Vírus Respiratório Sincicial/epidemiologia , Estudos de Coortes , Feminino , Humanos , Lactente , Masculino , Orthomyxoviridae/isolamento & purificação , Estudos Prospectivos , Vírus Sinciciais Respiratórios/isolamento & purificaçãoRESUMO
Abstract Objectives: To report epidemiological features, clinical characteristics, and outcomes of human rhinovirus (HRV) infections in comparison with other community acquired respiratory virus (CRV) infections in patients hospitalized for two consecutive years. Methods: This was a cross-sectional study. Clinical, epidemiological, and laboratory data of patients hospitalized with acute respiratory syndrome in a tertiary care hospital from 2012 to 2013 were reviewed. Results: HRV was the most common CRV observed (36%, 162/444) and was present in the majority of viral co-detections (69%, 88/128), mainly in association with human enterovirus (45%). Most HRV-infected patients were younger than 2 years (57%). Overall, patients infected with HRV had a lower frequency of severe acute respiratory infection than those infected with other CRVs (60% and 84%, respectively, p = 0.006), but had more comorbidities (40% and 27%, respectively; p = 0.043). However, in the adjusted analysis this association was not significant. The mortality rate within the HRV group was 3%. Detection of HRV was more prevalent during autumn and winter, with a moderately negative correlation between viral infection frequency and temperature (r = −0.636, p < 0.001) but no correlation with rainfall (r = −0.036, p = 0.866). Conclusion: HRV is usually detected in hospitalized children with respiratory infections and is often present in viral co-detections. Comorbidities are closely associated with HRV infections. These infections show seasonal variation, with predominance during colder seasons.
Resumo Objetivos: Relatar as características epidemiológicas, as características clínicas e os resultados das infecções por rinovírus humano (RVH) em comparação a outras infecções por vírus respiratórios adquiridos na comunidade (VRCs) em pacientes internados por dois anos consecutivos. Métodos: Este foi um estudo transversal. Foram revisados os dados clínicos, epidemiológicos e laboratoriais de pacientes internados com síndrome respiratória aguda em um hospital terciário de 2012 a 2013. Resultados: O RVH foi o VRC mais comum observado (36%, 162/444) e esteve presente na maior parte das codetecções virais (69%, 88/128), principalmente em associação ao enterovírus humano (45%). A maioria dos pacientes infectados por RVH possuía menos de 2 anos (57%). De modo geral, os pacientes com RVH apresentaram uma menor frequência de infecção respiratória aguda grave que os pacientes infectados por outros VRCs (60% e 84%, respectivamente, p = 0,006), porém mais comorbidades (40% e 27%, respectivamente; p = 0,043). Contudo, em uma análise ajustada, essa associação não foi significativa. A taxa de mortalidade no grupo RVH foi 3%. A detecção de RVH foi mais prevalente durante o outono e inverno, com uma correlação negativa moderada entre a frequência de infecção viral e a temperatura (r = -0,636, p < 0,001), porém nenhuma correlação com a precipitação (r = −0,036, p = 0,866). Conclusão: O RVH é normalmente detectado em crianças internadas com infecções respiratórias e normalmente está presente em codetecções virais. As comorbidades estão estreitamente associadas a infecções por RVH. Essas infecçõesmostram variação sazonal, com predominância durante as estações mais frias.
Assuntos
Humanos , Masculino , Feminino , Pré-Escolar , Criança , Adolescente , Adulto , Pessoa de Meia-Idade , Adulto Jovem , Infecções Respiratórias/epidemiologia , Rhinovirus/isolamento & purificação , Infecções por Picornaviridae/epidemiologia , Infecções Respiratórias/diagnóstico , Infecções Respiratórias/virologia , Rhinovirus/classificação , Estações do Ano , Brasil/epidemiologia , Prevalência , Estudos Transversais , Infecções por Picornaviridae/diagnóstico , Infecções por Picornaviridae/virologia , HospitalizaçãoRESUMO
OBJECTIVES: To report epidemiological features, clinical characteristics, and outcomes of human rhinovirus (HRV) infections in comparison with other community acquired respiratory virus (CRV) infections in patients hospitalized for two consecutive years. METHODS: This was a cross-sectional study. Clinical, epidemiological, and laboratory data of patients hospitalized with acute respiratory syndrome in a tertiary care hospital from 2012 to 2013 were reviewed. RESULTS: HRV was the most common CRV observed (36%, 162/444) and was present in the majority of viral co-detections (69%, 88/128), mainly in association with human enterovirus (45%). Most HRV-infected patients were younger than 2 years (57%). Overall, patients infected with HRV had a lower frequency of severe acute respiratory infection than those infected with other CRVs (60% and 84%, respectively, p=0.006), but had more comorbidities (40% and 27%, respectively; p=0.043). However, in the adjusted analysis this association was not significant. The mortality rate within the HRV group was 3%. Detection of HRV was more prevalent during autumn and winter, with a moderately negative correlation between viral infection frequency and temperature (r=-0.636, p<0.001) but no correlation with rainfall (r=-0.036, p=0.866). CONCLUSION: HRV is usually detected in hospitalized children with respiratory infections and is often present in viral co-detections. Comorbidities are closely associated with HRV infections. These infections show seasonal variation, with predominance during colder seasons.
Assuntos
Infecções por Picornaviridae/epidemiologia , Infecções Respiratórias/epidemiologia , Rhinovirus/isolamento & purificação , Adolescente , Adulto , Brasil/epidemiologia , Criança , Pré-Escolar , Estudos Transversais , Feminino , Hospitalização , Humanos , Masculino , Pessoa de Meia-Idade , Infecções por Picornaviridae/diagnóstico , Infecções por Picornaviridae/virologia , Prevalência , Infecções Respiratórias/diagnóstico , Infecções Respiratórias/virologia , Rhinovirus/classificação , Estações do Ano , Adulto JovemRESUMO
BACKGROUND: Viral infections play a significant role in causing acute respiratory infections (ARIs) and exacerbations of chronic diseases. Acute respiratory infections are now the leading cause of mortality in children worldwide, especially in developing countries. Recently, human rhinovirus (HRV) infection has been emerged as an important cause of pneumonia and asthma exacerbation. OBJECTIVES: To determine the role of several viral agents principally, respiratory syncytial virus, and HRV in children with ARIs and their relationship with asthma exacerbation and pneumonia. METHODS: Between October 2011 and March 2014, 432 nasopharyngeal samples of children <15 years of age with ARI hospitalized at a referral hospital for respiratory diseases were tested for the presence of respiratory viruses using a multiplex RT-qPCR. Clinical, epidemiological, and demographic data were collected and associated with symptomatology and viral infections. RESULTS: Viral infections were detected in at least 59·7% of the enrolled patients, with HRV (26·6%) being the most frequently detected. HRV infections were associated with clinical features of asthma and difficulty in breathing such as wheezing (P = 0·0003), supraesternal (P = 0·046), and xiphoid retraction (P = 0·030). HRV subtype C (HRV-C) infections were associated with asthma (P = 0·02). CONCLUSIONS: Human rhinovirus was the virus most commonly detected in pediatric patients with ARI. There is also an association of HRV-C infection with asthma exacerbation, emphasizing the relevance of this virus in severe pediatric respiratory disease.
RESUMO
Los métodos moleculares para diagnosticar rinovirus humanos (RVH) han aumentado la sensibilidad de detección. Esto ha permitido documentar la asociación entre los RVH y las infecciones respiratorias agudas (IRA) altas y bajas. La infección por RVH durante la infancia se asoció con posterior desarrollo de asma. Se estudió la frecuencia de RVH en 186 niños menores de 6 años ambulatorios con IRA (alta o baja), durante 2 años consecutivos (1/6/2008 - 31/5/2010). Se correlacionó la presencia de RVH con los antecedentes y características clínico-epidemiológicas. La detección de RVH se realizó con una RT-PCR en tiempo real que amplifica parte de la región 5' no codificante del genoma. Los virus respiratorios clásicos se estudiaron por inmunofluorescencia. En el 61% de los niños se detectó etiología viral. Las frecuencias fueron: RVH 27%, virus sincicial respiratorio (VSR) 16%, influenza A y B 9%, parainfluenza 8%, metapneumovirus 7% y adenovirus 0.5%. Se observaron coinfecciones duales en 8 casos, siendo RVH el más frecuente (en 4 de ellos). Los RVH circularon durante todo el período estudiado, con picos en invierno y primavera. No se observaron diferencias clínico-epidemiológicas significativas entre pacientes con o sin RVH, excepto un mayor porcentaje de niños afebriles con RVH. Los RVH fueron los virus más detectados en niños ambulatorios, principalmente en menores de 2 años, los segundos virus asociados a bronquiolitis, luego del VSR, y detectados tres veces más en los niños expuestos a tabaquismo pasivo (OR: 2,91; p = 0.012) que en el resto. Fueron identificados como único agente en el 28% de las bronquiolitis.
Molecular methods for human rhinoviruses (HRV) have increased the sensitivity in their diagnosis. HRV may cause acute respiratory infections (ARI) of the upper and lower respiratory tract. HRV infection during childhood is a predictor of asthma development. In this study, the HRV frequency in outpatient children with ARI was determined, and their clinical features and previous conditions were evaluated. A total of 186 respiratory samples of children under 6 year old attending the CEMIC pediatric emergency room from June 1, 2008 to May 31, 2010, were studied. Classical respiratory viruses were detected by immunofluorescence. A real time RT-PCR that amplifies part of the 5' non coding genomic region was used for HRV detection. Viral detection was obtained in 61% of children. The frequency was: 27% for HRV, 16% for respiratory syncytial virus (RSV), 9% for influenza, 8% for parainfluenza, 7% for metapneumovirus and 0.5% for adenovirus. Dual coinfection was detected in 8 children and HRV were the most frequent, detected in 4 of them. HRV circulated during the two year period of the study, with peaks during winter and spring. No clinical difference was observed between patients with or without HRV, except an increase percent of children with HRV without fever. HRV were the most frequent viruses detected in this population, mainly in children under 2 year old, the second cause of bronchiolitis after RSV and more frequently detected in children exposed to passive smoking (OR = 2.91; p = 0.012), and were detected as the sole etiologic agent in 28% of bronchiolitis.
Assuntos
Criança , Feminino , Humanos , Lactente , Recém-Nascido , Masculino , Infecções por Picornaviridae/epidemiologia , Infecções Respiratórias/virologia , Rhinovirus/isolamento & purificação , Doença Aguda , Distribuição por Idade , Argentina/epidemiologia , Bronquiolite/diagnóstico , Bronquiolite/virologia , Estudos Transversais , Faringite/diagnóstico , Faringite/virologia , Infecções Respiratórias/epidemiologia , Rinite/diagnóstico , Rinite/virologia , Estações do Ano , Distribuição por SexoRESUMO
Los métodos moleculares para diagnosticar rinovirus humanos (RVH) han aumentado la sensibilidad de detección. Esto ha permitido documentar la asociación entre los RVH y las infecciones respiratorias agudas (IRA) altas y bajas. La infección por RVH durante la infancia se asoció con posterior desarrollo de asma. Se estudió la frecuencia de RVH en 186 niños menores de 6 años ambulatorios con IRA (alta o baja), durante 2 años consecutivos (1/6/2008 - 31/5/2010). Se correlacionó la presencia de RVH con los antecedentes y características clínico-epidemiológicas. La detección de RVH se realizó con una RT-PCR en tiempo real que amplifica parte de la región 5 no codificante del genoma. Los virus respiratorios clásicos se estudiaron por inmunofluorescencia. En el 61% de los niños se detectó etiología viral. Las frecuencias fueron: RVH 27%, virus sincicial respiratorio (VSR) 16%, influenza A y B 9%, parainfluenza 8%, metapneumovirus 7% y adenovirus 0.5%. Se observaron coinfecciones duales en 8 casos, siendo RVH el más frecuente (en 4 de ellos). Los RVH circularon durante todo el período estudiado, con picos en invierno y primavera. No se observaron diferencias clínico-epidemiológicas significativas entre pacientes con o sin RVH, excepto un mayor porcentaje de niños afebriles con RVH. Los RVH fueron los virus más detectados en niños ambulatorios, principalmente en menores de 2 años, los segundos virus asociados a bronquiolitis, luego del VSR, y detectados tres veces más en los niños expuestos a tabaquismo pasivo (OR: 2,91; p = 0.012) que en el resto. Fueron identificados como único agente en el 28% de las bronquiolitis.(AU)
Molecular methods for human rhinoviruses (HRV) have increased the sensitivity in their diagnosis. HRV may cause acute respiratory infections (ARI) of the upper and lower respiratory tract. HRV infection during childhood is a predictor of asthma development. In this study, the HRV frequency in outpatient children with ARI was determined, and their clinical features and previous conditions were evaluated. A total of 186 respiratory samples of children under 6 year old attending the CEMIC pediatric emergency room from June 1, 2008 to May 31, 2010, were studied. Classical respiratory viruses were detected by immunofluorescence. A real time RT-PCR that amplifies part of the 5 non coding genomic region was used for HRV detection. Viral detection was obtained in 61% of children. The frequency was: 27% for HRV, 16% for respiratory syncytial virus (RSV), 9% for influenza, 8% for parainfluenza, 7% for metapneumovirus and 0.5% for adenovirus. Dual coinfection was detected in 8 children and HRV were the most frequent, detected in 4 of them. HRV circulated during the two year period of the study, with peaks during winter and spring. No clinical difference was observed between patients with or without HRV, except an increase percent of children with HRV without fever. HRV were the most frequent viruses detected in this population, mainly in children under 2 year old, the second cause of bronchiolitis after RSV and more frequently detected in children exposed to passive smoking (OR = 2.91; p = 0.012), and were detected as the sole etiologic agent in 28% of bronchiolitis.(AU)
Assuntos
Criança , Feminino , Humanos , Lactente , Recém-Nascido , Masculino , Infecções por Picornaviridae/epidemiologia , Infecções Respiratórias/virologia , Rhinovirus/isolamento & purificação , Doença Aguda , Distribuição por Idade , Argentina/epidemiologia , Bronquiolite/diagnóstico , Bronquiolite/virologia , Estudos Transversais , Faringite/diagnóstico , Faringite/virologia , Infecções Respiratórias/epidemiologia , Rinite/diagnóstico , Rinite/virologia , Estações do Ano , Distribuição por SexoRESUMO
Los métodos moleculares para diagnosticar rinovirus humanos (RVH) han aumentado la sensibilidad de detección. Esto ha permitido documentar la asociación entre los RVH y las infecciones respiratorias agudas (IRA) altas y bajas. La infección por RVH durante la infancia se asoció con posterior desarrollo de asma. Se estudió la frecuencia de RVH en 186 niños menores de 6 años ambulatorios con IRA (alta o baja), durante 2 años consecutivos (1/6/2008 - 31/5/2010). Se correlacionó la presencia de RVH con los antecedentes y características clínico-epidemiológicas. La detección de RVH se realizó con una RT-PCR en tiempo real que amplifica parte de la región 5 no codificante del genoma. Los virus respiratorios clásicos se estudiaron por inmunofluorescencia. En el 61% de los niños se detectó etiología viral. Las frecuencias fueron: RVH 27%, virus sincicial respiratorio (VSR) 16%, influenza A y B 9%, parainfluenza 8%, metapneumovirus 7% y adenovirus 0.5%. Se observaron coinfecciones duales en 8 casos, siendo RVH el más frecuente (en 4 de ellos). Los RVH circularon durante todo el período estudiado, con picos en invierno y primavera. No se observaron diferencias clínico-epidemiológicas significativas entre pacientes con o sin RVH, excepto un mayor porcentaje de niños afebriles con RVH. Los RVH fueron los virus más detectados en niños ambulatorios, principalmente en menores de 2 años, los segundos virus asociados a bronquiolitis, luego del VSR, y detectados tres veces más en los niños expuestos a tabaquismo pasivo (OR: 2,91; p = 0.012) que en el resto. Fueron identificados como único agente en el 28% de las bronquiolitis.(AU)
Molecular methods for human rhinoviruses (HRV) have increased the sensitivity in their diagnosis. HRV may cause acute respiratory infections (ARI) of the upper and lower respiratory tract. HRV infection during childhood is a predictor of asthma development. In this study, the HRV frequency in outpatient children with ARI was determined, and their clinical features and previous conditions were evaluated. A total of 186 respiratory samples of children under 6 year old attending the CEMIC pediatric emergency room from June 1, 2008 to May 31, 2010, were studied. Classical respiratory viruses were detected by immunofluorescence. A real time RT-PCR that amplifies part of the 5 non coding genomic region was used for HRV detection. Viral detection was obtained in 61% of children. The frequency was: 27% for HRV, 16% for respiratory syncytial virus (RSV), 9% for influenza, 8% for parainfluenza, 7% for metapneumovirus and 0.5% for adenovirus. Dual coinfection was detected in 8 children and HRV were the most frequent, detected in 4 of them. HRV circulated during the two year period of the study, with peaks during winter and spring. No clinical difference was observed between patients with or without HRV, except an increase percent of children with HRV without fever. HRV were the most frequent viruses detected in this population, mainly in children under 2 year old, the second cause of bronchiolitis after RSV and more frequently detected in children exposed to passive smoking (OR = 2.91; p = 0.012), and were detected as the sole etiologic agent in 28% of bronchiolitis.(AU)
Assuntos
Criança , Feminino , Humanos , Lactente , Recém-Nascido , Masculino , Infecções por Picornaviridae/epidemiologia , Infecções Respiratórias/virologia , Rhinovirus/isolamento & purificação , Doença Aguda , Distribuição por Idade , Argentina/epidemiologia , Bronquiolite/diagnóstico , Bronquiolite/virologia , Estudos Transversais , Faringite/diagnóstico , Faringite/virologia , Infecções Respiratórias/epidemiologia , Rinite/diagnóstico , Rinite/virologia , Estações do Ano , Distribuição por SexoRESUMO
INTRODUCTION: Acute respiratory tract infections are the most common illness in all individuals. Rhinoviruses have been reported as the etiology of more than 50 percent of respiratory tract infections worldwide. The study prospectively evaluated 47 elderly individuals from a group of 384 randomly assigned for acute respiratory viral infections (cold or flu) and assessed the occurrence of human rhinovirus (HRV), influenza A and B, respiratory syncytial virus and metapneumovirus (hMPV) in Botucatu, State of São Paulo, Brazil. METHODS: Forty-nine nasal swabs collected from 47 elderly individuals following inclusion visits from 2002 to 2003 were tested by GenScan RT-PCR. HRV-positive samples were sequenced for phylogenetic analysis. RESULTS: No sample was positive for influenza A/B or RSV. HRV was detected in 28.6 percent (14/47) and hMPV in 2 percent (1/47). Of 14 positive samples, 9 isolates were successfully sequenced, showing the follow group distribution: 6 group A, 1 group B and 2 group C HRVs. CONCLUSIONS: The high incidence of HRV during the months of the influenza season requires further study regarding HRV infection impact on respiratory complications among this population. Infection caused by HRV is very frequent and may contribute to increasing the already high demand for healthcare during the influenza season.
INTRODUÇÃO: Infecções agudas do trato respiratório estão entre as doenças mais comuns em todas as pessoas. Os rinovírus têm sido descritos como agente etiológico de mais de 50 por cento das infecções do trato respiratório ao redor do mundo. O objetivo deste trabalho foi avaliar a ocorrência de rinovírus humano (HRV), influenza vírus A e B, vírus respiratório sincicial humano e metapneumovírus (hMPV) em uma população de idosos que apresentava sintomas de gripe ou resfriado, e que residiam na Cidade de Botucatu, Estado de São Paulo, Brasil. MÉTODOS: Foram coletados swabs nasais de 47 idosos após visitas de inclusão, entre os anos de 2002 e 2003 e que foram testadas através de GeneScan RT-PCR. RESULTADOS: HRV foi detectado em 28.6 por cento (14/47) e hMPV em 2 por cento (1/47). De 14 amostras positivas para HRV, 9 foram sequenciadas, mostrando a seguinte distribuição de grupos: grupo A: 6 amostras, grupo B: 1 amostra e grupo C: 2 amostras. CONCLUSÕES: A alta incidência de HRV durante os meses de ocorrência de gripe necessita de estudos posteriores para avaliar o impacto desse vírus entre os idosos. A alta frequência de HRV pode contribuir para o aumento da demanda por serviços de saúde durante a estação de influenza.