Serum Levels of Arachidonic Acid, Interleukin-6, and C-Reactive Protein as Potential Indicators of Pulmonary Viral Infections: Comparative Analysis of Influenza A, Respiratory Syncytial Virus Infection, and COVID-19.

Acute respiratory tract infections, including influenza A (FluA), respiratory syncytial virus (RSV) infection, and COVID-19, can aggravate to levels requiring hospitalization, increasing morbidity and mortality. Identifying biomarkers for an accurate diagnosis and prognosis of these infections is a clinical need. We performed a cross-sectional study aimed to investigate the changes in circulating levels of arachidonic acid, interleukin 6 (IL-6), and C-reactive protein (CRP) in patients with FluA, RSV, or COVID-19, and to analyze the potential of these parameters as diagnosis or prognosis biomarkers. We analyzed serum samples from 172 FluA, 80 RSV, and 217 COVID-19 patients, and 104 healthy volunteers. Individuals with lung viral diseases showed reduced arachidonic acid concentrations compared to healthy people, with these differences being most pronounced in the order COVID-19 > RSV > FluA. Conversely, IL-6 and CRP levels were elevated across diseases, with IL-6 emerging as the most promising diagnostic biomarker, with areas under the curve (AUC) of the receiver operating characteristics plot higher than 0.85 and surpassing arachidonic acid and CRP. Moreover, IL-6 displayed notable efficacy in distinguishing between FluA patients who survived and those who did not (AUC = 0.80). These findings may provide useful tools for diagnosing and monitoring the severity of acute viral respiratory tract infections, ultimately improving patient outcomes.

Development and Validation of an Enzyme-Linked Immunosorbent Assay-Based Protocol for Evaluation of Respiratory Syncytial Virus Vaccines.

Recently, respiratory syncytial virus (RSV) vaccines based on the prefusion F (pre-F) antigen were approved in the United States. We aimed to develop an enzyme-linked immunosorbent assay (ELISA)-based protocol for the practical and large-scale evaluation of RSV vaccines. Two modified pre-F proteins (DS-Cav1 and SC-TM) were produced by genetic recombination and replication using an adenoviral vector. The protocol was established by optimizing the concentrations of the coating antigen (pre-F proteins), secondary antibodies, and blocking buffer. To validate the protocol, we examined its accuracy, precision, and specificity using serum samples from 150 participants across various age groups and the standard serum provided by the National Institute of Health. In the linear correlation analysis, coating concentrations of 5 and 2.5 µg/mL of DS-Cav1 and SC-TM showed high coefficients of determination (r > 0.90), respectively. Concentrations of secondary antibodies (alkaline phosphatase-conjugated anti-human immunoglobulin G, diluted 1:2000) and blocking reagents (5% skim milk/PBS-T) were optimized to minimize non-specific reactions. High accuracy was observed for DS-Cav1 (r = 0.90) and SC-TM (r = 0.86). Further, both antigens showed high precision (coefficient of variation < 15%). Inhibition ELISA revealed cross-reactivity of antibodies against DS-Cav1 and SC-TM, but not with the attachment (G) protein.

Peptide-Based electrochemical potential Scanning: A novel approach for disulfide manipulation in pediatric Respiratory syncytial virus detection.

Respiratory syncytial virus (RSV) poses a significant risk to children under two years old, necessitating rapid and accurate diagnostic methods. This study introduces an innovative approach using peptides and electrochemical potential scanning for RSV detection. By replacing enzymatic catalysis with electrochemical scanning, the method simplifies the process and reduces costs. Unbound peptides undergo potential-induced disulfide bridge opening, while target-bound peptides remain protected. After removing the target protein, copper ions and a reduced short peptide promote disulfide bridge formation, leading to crosslinking and passivation of the electrode surface. The degree of polymerization and passivation correlates with the target protein levels, generating a signal. This novel method offers enhanced sensitivity, specificity, and scalability, potentially revolutionizing RSV diagnostics in children under two years old. By addressing the limitations of traditional assays, it provides a cost-effective, rapid, and efficient approach for early RSV detection and improved clinical outcomes in this vulnerable population.

Severe Pneumonia Caused by Respiratory Syncytial Virus and Adenovirus in Children from 2 to 24 Months at Children's Hospital 1 in Ho Chi Minh City, Vietnam.

In Vietnam, due to the lack of facilities to detect respiratory viruses from patients' specimens, there are only a few studies on the detection of viral pathogens causing pneumonia in children, especially respiratory syncytial virus (RSV) and adenovirus (Adv). Here, we performed a cross-sectional descriptive prospective study on 138 children patients from 2 to 24 months old diagnosed with severe pneumonia hospitalized at the Respiratory Department of Children's Hospital 1 from November 2021 to August 2022. The number of patients selected in this study was based on the formula n = ([Z(1 - α/2)]2 × P [1 - P])/d2, with α = 0.05, p = 0.5, and d = 9%, and the sampling technique was convenient sampling until the sample size was met. A rapid test was used to detect RSV and Adv from the nasopharyngeal swabs and was conducted immediately after the patient's hospitalization. Laboratory tests were performed, medical history interviews were conducted, and nasotracheal aspirates were collected for multiplex real-time PCR (MPL-rPCR) to detect viral and bacterial pathogens. The results of the rapid test and the MPL-rPCR in the detection of both pathogens were the same at 31.9% (44/138) for RSV and 8.7% (7/138) for Adv, respectively. Using MPL-rPCR, the detection rate was 21% (29/138) for bacterial pathogens, 68.8% (95/138) for bacterial-viral co-infections, and 6.5% (9/138) for viral pathogens. The results showed few distinctive traits between RSV-associated and Adv-associated groups, and the Adv group children were more prone to bacterial infection than those in the RSV group. In addition, the Adv group experienced a longer duration of treatment and a higher frequency of re-hospitalizations compared to the RSV group. A total of 100% of Adv infections were co-infected with bacteria, while 81.82% of RSV co-infected with bacterial pathogens (p = 0.000009). This study might be one of the few conducted in Vietnam aimed at identifying viral pathogens causing severe pneumonia in children.

Development and validation of a respiratory syncytial virus multiplex immunoassay.

PURPOSE: Respiratory syncytial virus (RSV) is one of the leading causes of severe respiratory disease in infants and adults. While vaccines and monoclonal therapeutic antibodies either are or will shortly become available, correlates of protection remain unclear. For this purpose, we developed an RSV multiplex immunoassay that analyses antibody titers toward the post-F, Nucleoprotein, and a diverse mix of G proteins. METHODS: A bead-based multiplex RSV immunoassay was developed, technically validated to standard FDA bioanalytical guidelines, and clinically validated using samples from human challenge studies. RSV antibody titers were then investigated in children aged under 2 and a population-based cohort. RESULTS: Technical and clinical validation showed outstanding performance, while methodological developments enabled identification of the subtype of previous infections through use of the diverse G proteins for approximately 50% of samples. As a proof of concept to show the suitability of the assay in serosurveillance studies, we then evaluated titer decay and age-dependent antibody responses within population cohorts. CONCLUSION: Overall, the developed assay shows robust performance, is scalable, provides additional information on infection subtype, and is therefore ideally suited to be used in future population cohort studies.

Robust and sensitive amplicon-based whole-genome sequencing assay of respiratory syncytial virus subtype A and B.

Prevention of respiratory syncytial virus (RSV) infection is now a global health priority, with a long-acting monoclonal antibody and two RSV vaccines recently licenced for clinical use. Most licenced and candidate interventions target the RSV fusion (RSV-F) protein. New interventions may be associated with the spread of mutations, reducing susceptibility to antibody neutralization in RSV-F. There is a need for ongoing longitudinal global surveillance of circulating RSV strains. To achieve this large-scale genomic surveillance, a reliable, high-throughput RSV sequencing assay is required. Here we report an improved high-throughput RSV whole-genome sequencing (WGS) assay performed directly on clinical samples without additional enrichment, using a 4-primer-pool, short-amplicon PCR-tiling approach that is suitable for short-read sequencing platforms. Using upper respiratory tract (URT) RSV-positive clinical samples obtained from a sentinel network of primary care providers and from hospital patients (29.7% and 70.2%, respectively; n = 1,037), collected over the period 2019 to 2023, this assay had a threshold of approximately 4 × 103 to 8 × 103 copies/mL (RSV-B and RSV-A sub-types, respectively) as the lowest amount of virus needed in the sample to achieve >96% of whole-genome coverage at a high-quality level. Using a Ct value of 31 as an empirical cut-off, the overall assay success rate of obtaining >90% genome coverage at a read depth minimum of 20 was 96.83% for clinical specimens successfully sequenced from a total of 1,071. The RSV WGS approach described in this study has increased sensitivity compared to previous approaches and can be applied to clinical specimens without the requirement for enrichment. The updated approach produces sequences of high quality consistently and cost-effectively, suitable for implementation to underpin national programs for the surveillance of RSV genomic variation. IMPORTANCE: In this paper, we report an improved high-throughput respiratory syncytial virus (RSV) whole-genome sequencing (WGS) assay performed directly on clinical samples, using a 4-primer-pool, short-amplicon PCR-tiling approach that is suitable for short-read sequencing platforms. The RSV WGS approach described in this study has increased sensitivity compared to previous approaches and can be applied to clinical specimens without the requirement for enrichment. The updated approach produces sequences of high quality consistently and cost-effectively, suitable for implementation to underpin national and global programs for the surveillance of RSV genomic variation. The quality of sequence produced is essential for preparedness for new interventions in monitoring antigenic escape, where a single point mutation might lead to a reduction in antibody binding effectiveness and neutralizing activity, or indeed in the monitoring of retaining susceptibility to neutralization by existing and new interventions.

Vigilancia de virus sincicial respiratorio e influenza en niños escolarizados asistidos en un hospital pediátrico durante 2 meses del segundo semestre de 2021 / Respiratory syncytial virus and influenza surveillance in schoolchildren seen at a children's hospital over 2 months of the second semester of 2021

Introducción. Durante la pandemia de COVID-19, disminuyeron las notificaciones de infecciones respiratorias. El objetivo fue estimar la prevalencia de virus sincicial respiratorio (VSR) e influenza en niños escolarizados asistidos en un hospital pediátrico durante el retorno a la presencialidad. Métodos. Estudio transversal de casos sospechosos de COVID-19, de 3-18 años, con prueba negativa para SARSCoV-2, entre agosto y octubre de 2021. Se estratificó por nivel educativo. Se utilizó PCR para detectar VSR e influenza. Resultados. Se incluyeron 619 niños: 234 del nivel inicial, 224 del primario y 161 del secundario; 25,5 % (158) fueron positivos para VSR (36,3 % del nivel inicial versus 21 % del primario y 16 % del secundario); en adolescentes se asoció la infección al contacto escolar con caso sintomático (OR 2,5; IC95%: 1-6,80; p = 0,04). No se aisló virus influenza. Conclusión. VSR se aisló en un cuarto de la población estudiada, con mayor frecuencia en el nivel inicial; en adolescentes, se asoció con contacto escolar sintomático. No se detectaron casos de influenza

Introduction. Reporting of respiratory infections reduced during the COVID-19 pandemic. The objective was to estimate the prevalence of respiratory syncytial virus (RSV) and influenza in schoolchildren seen at a children's hospital during the return to school. Methods. Cross-sectional study of patients aged 3­18 years suspected of COVID-19 with a negative test for SARS-CoV-2 between August and October 2021. Participants were stratified by level of education. PCR was used to detect RSV and influenza. Results. A total of 619 children were included: 234 in preschool, 224 in primary and 161 in secondary school; 25.5% (158) tested positive for RS (36.3% in the pre-school level versus 21% in primary and 16% in secondary school). Infection among adolescents was associated with school contact with symptomatic cases (OR 2.5; 95%CI 1­6.80; p = 0.04). No case of influenza was detected. Conclusion. RSV was isolated in one fourth of the study population, with a higher frequency in pre-school; among adolescents, it was associated with school contact with symptomatic cases. No case of influenza was detected.

Evaluación de un método de amplificación isotérmica mediado por bucle para la detección rápida del virus sincicial respiratorio en niños con infección respiratoria aguda / Evaluation of a loop-mediated isothermal amplification method for the rapid detection of human respiratory syncytial virus in children with acute respiratory infection

Resumen Introducción. El virus sincicial respiratorio humano (hRSV) es la causa más frecuente de infección respiratoria aguda de las vías respiratorias inferiores en niños menores de cinco años. El desarrollo de técnicas moleculares para identificarlo es uno de los retos actuales en el campo de la investigación clínica. Objetivo. Evaluar un método de amplificación isotérmica para la detección rápida del hRSV en niños con infección respiratoria aguda. Materiales y métodos. Se extrajo el ARN viral de 304 muestras de hisopado nasal en niños con síntomas de infección respiratoria aguda atendidos en el servicio de urgencias del Hospital de la Universidad del Norte en Barranquilla entre abril del 2016 y julio del 2017. Se evaluó la prueba de amplificación isotérmica mediada por bucle mediante transcriptasa inversa de la proteína de la matriz (M) (Reverse Transcription Loop-Mediated Isothermal Amplification, RT-LAMP) comparada con técnicas moleculares como la reacción en cadena de la polimerasa mediante transcriptasa inversa múltiple anidada (Reverse Transcription- Polymerase Chain Reaction, RT-PCR), la cual se empleó como la prueba estándar, la PCR en tiempo real (quantitative PCR, qPCR) y la RT-LAMP de la proteína L (L) para la detección rápida del virus sincicial respiratorio (VSR), subtipo A y subtipo B. Resultados. La prueba de RT-LAMP (M) tuvo una sensibilidad de 93,59 %, una especificidad de 92,92 % y una concordancia de 0,83 ± 0,036 comparada con la prueba de RT-PCR anidada. El índice kappa del RT-LAMP (M) fue superior, y los valores del RT-LAMP (L) y la qPCR concordaron (0,75 ± 0,043 y 0,71 ± 0,045, respectivamente). Conclusiones. Estos resultados indican que la prueba RT-LAMP (M) puede considerarse como una herramienta de utilidad clínica para detectar el hSRVA, dado que el tiempo requerido para la obtención de resultados, así como los costos, es menor, y su desempeño es mejor que el de las otras pruebas moleculares evaluadas.

Abstract Introduction: Human respiratory syncytial virus (hRSV) is the most frequent cause of acute respiratory infection of the lower respiratory tract in children under the age of five. The development of molecular techniques able to identify hRSV is one of the current challenges in the field of clinical research. Objective: To evaluate the ability of an isothermal amplification method to rapidly detect hRSV in children with acute respiratory infection. Materials and methods: We collected 304 nasopharyngeal swab samples from children with symptoms of acute respiratory infection who attended the emergency unit at Hospital de la Universidad del Norte in Barranquilla from April, 2016, to July, 2017. After extracting viral RNA from the samples, we evaluated the ability of the reverse transcriptase-loop-mediated isothermal amplification (RT-LAMP) M assay to rapidly detect hRSVA and hRSVB compared to other molecular techniques: quantitative PCR (qPCR), reverse transcriptase-LAMP L assay, and as a standard, the multiplex nested reverse transcriptase polymerase chain reaction (nested RT-PCR). Results: The RT-LAMP M assay had a sensitivity of 93.59% and a specificity of 92.92%, and a concordance of 0.83 ± 0.036 as compared with the nested RT-PCR test. While the Kappa index of the RT-LAMP M assay was higher than the values for the RT-LAMP L assay and the qPCR, the values of the latter two methods were in agreement (0.75 ± 0.043 and 0.71 ± 0.045, respectively). Conclusion: Due to the shorter running times, lower costs and better performance of the RT-LAMP M assay, it can be considered as a useful clinical tool for the detection of RSVA.

Virus más frecuentemente aislados en la exacerbación de asma y su correlación con nivelesde eosinófilos e inmunoglobulina E sérica total / Most commonly isolated viruses in asthma exacerbation and their correlation with eosinophil and total serum immunoglobulin E levels

Antecedentes. Las exacerbaciones de asma continúan siendo una causa de hospitalización en el Servicio de Urgencias. Los desencadenantesson alérgenos e infecciones, principalmente, de tipo viral. El objetivo fue determinar la relación entre los virus detectados durante la exacerbación asmática y los niveles de eosinófilos e inmunoglobulina E (IgE) sérica en pacientes pediátricos. Población y métodos. Estudio transversal analítico. Se incluyeron niños de cinco a quince años atendidos en Urgencias de Pediatría con exacerbación de asma, en el período de marzo de 2013 a febrero de 2016. Se obtuvo ácido ribonucleico viral en el aspirado nasofaríngeo con el kit CLART PneumoVir. Se cuantificaron los eosinófilos en la sangre periférica y los niveles de IgE sérica total. Se consideró eosinofilia un conteo ≥ 0,4 x 103/mm3 e IgE elevada, ≥ 350 UI/L. Se realizó la correlación de Pearson. Se definió significancia con valor de p ≤ 0,05.Resultados. De 211 niños con exacerbación de asma, en el 20%, se aisló un virus. Los virus aislados más frecuentemente fueron el rinovirus, el enterovirus y el virus sincitial respiratorio. Se encontró una correlación entre los niveles de eosinófilos e IgE sérica total en los niños con exacerbación de asma y rinovirus de 0,89, con una p= 0,0001.Conclusiones. Las infecciones por rinovirus, enterovirus y virus sincitial respiratorio son más frecuentes en las exacerbaciones de asma en menores de 15 años. Se observó una correlación entre los niveles de eosinófilos e IgE en presencia de rinovirus.

Background. Asthma exacerbations are still a cause of hospitalization at the Emergency Department. The triggers of asthma exacerbations include allergens and infections ­mostly viral­. The objective of this study was to establish the relationship between viruses detected during an asthma exacerbation and eosinophil and serum immunoglobulin E (IgE) levels in pediatric patients. Population and methods. Cross-sectional. analytical study. Children aged 5-15 years seen at the Pediatric Emergency Department with an asthma exacerbation in the period between March 2013 and February 2016 were included. Viral ribonucleic acid was extracted from nasopharyngeal aspirates using the CLART Pneumo Vir kit. Eosinophil levels were measured in peripheral blood and total IgE levels, in serum. Eosinophilia was defined as a count ≥ 0.4 x 103/mm3 and high IgE. as a level ≥ 350 IU/L. The Pearson's correlation was carried out. A value of p ≤ 0.05 was considered significant.Results. Out of 211 children with asthma exacerbation, a virus was isolated in 20%. The most commonly isolated viruses were rhinovirus. enterovirus, and respiratory syncytial virus. A correlation of 0.89 was established between eosinophil and total serum IgE levels in children with asthma exacerbation and rhinovirus, with a p value of 0.0001. Conclusions. Rhinovirus, enterovirus, and respiratory syncytial virus were the most common viruses in asthma exacerbations in children younger than 15 years. A correlation was established between eosinophil and IgE levels in the presence of rhinovirus.

Infecciones respiratorias agudas bajas en niños menores de 2 años: hospitalizaciones durante el invierno del año 2014 / Low acute respiratory infections in children under 2 years: 2014 winter hospitalizations / Infecções agudas das vias respiratórias inferiores em crianças menores de 2 anos: hospitalizações em 2014

Durante el invierno las infecciones respiratorias agudas bajas (IRAB) determinan un incremento en la demanda asistencial, afectando sobre todo a los niños más pequeños. El objetivo de la investigación fue describir las características clínicas, modalidades de tratamiento y evolución de los menores de 2 años hospitalizados en el Hospital Pediátrico-Centro Hospitalario Pereira Rossell por IRAB de etiología viral durante el invierno de 2014. Se realizó un estudio descriptivo, retrospectivo, describiendo las características de los menores de 2 años hospitalizados del 9/6 al 21/9/2014 por IRAB de probable etiología viral. Se describieron las características epidemiológicas y clínicas, el tratamiento realizado y la evolución de los pacientes. En el período evaluado egresaron 742 niños (34% de los egresos de la institución). Tenían una mediana de edad de 4 meses; 18% presentaba al menos un factor de riesgo de IRAB grave. Se identificó al virus respiratorio sincicial en 59,6%. La estadía hospitalaria tuvo una mediana de 4 días. En las salas de cuidados moderados se aplicó ventilación no invasiva a 46 niños, y oxigenación de alto flujo a 129 niños, logrando una mejoría clínica en el 87,0% y el 87,6% respectivamente. Ingresaron a unidades de cuidados intensivos 217 niños, 54% requirió asistencia ventilatoria mecánica. Dos pacientes fallecieron. En el período evaluado los niños pequeños con IRAB representaron una importante proporción de los egresos, con importante carga asistencial. La mayoría eran niños sin factores de riesgo. La aplicación de las técnicas de tratamiento en cuidados moderados fue efectiva, permitiendo disminuir la demanda de camas de cuidados intensivos.

During the winter, low acute respiratory infections (LARI) determine an increase in care demand, especially affecting younger children. The objective of the research was to describe the clinical characteristics, treatment modalities and evolution of children under 2 years of age hospitalized at the Hospital Pediátrico-Centro Hospitalario Pereira Rossell for viral etiology LARI during the 2014 winter. A descriptive, retrospective study was conducted, describing the characteristics of children under 2 years hospitalized between 9/6 and 9/21/2014 for LARI of probable viral etiology. The following features were described: epidemiological and clinical characteristics, treatment performed and patient's evolution. During the period evaluated, 742 children were discharged (34% of the institution's admissions). They had a median age of 4 months; 18% had at least one risk factor for severe LARI. Respiratory syncytial virus was identified in 59.6%. The hospital stay had a median of 4 days. In moderate care rooms noninvasive ventilation was applied to 46 children, and high flow oxygenation to 129 children, achieving clinical improvement in 87.0% and 87.6%, respectively. Two hundred and seventeen children were admitted 54% required mechanical ventilation. Two patients died. In the evaluated period small children with LARI represented a significant proportion of the discharges, with an important burden of care. The majority were children without risk factors. The application of treatment techniques in moderate care was effective, allowed a decrease in the demand for intensive care beds.

Durante o inverno, as infecções respiratórias agudas baixas (IRAB) determinam o aumento da demanda de cuidados, afetando especialmente as crianças menores. O objetivo da pesquisa foi descrever as características clínicas, as modalidades de tratamento e a evolução de crianças menores de dois anos hospitalizados no Hospital Pediátrico-Centro Hospitalario Pereira Rossell devido a IRAB de etiologia viral, durante o inverno de 2014. Foi realizado um estudo descritivo e retrospectivo, descrevendo as características das crianças menores de 2 anos hospitalizadas entre 9/6 e 21/9/2014 devido a IRAB de provável etiologia viral. Descreveram-se: características epidemiológicas e clínicas, tratamento realizado e evolução dos pacientes. Durante o período avaliado, 742 crianças foram hospitalizadas (34% dos ingressos da instituição). Tinham uma idade média de 4 meses; 18% tinham pelo menos um fator de risco para IRAB severa. O vírus sincicial respiratório foi identificado em 59,6%. A estadia hospitalaria teve uma mediana de 4 dias. Em salas de cuidados moderados, a ventilação não invasiva foi aplicada a 46 crianças e a oxigenação de alto fluxo a 129 crianças, atingindo melhora clínica em 87,0% e 87,6%, respectivamente. Foram internadas em unidades de terapia intensiva 217 crianças, 54% necessitaram de ventilação mecânica. Dois pacientes faleceram. No período avaliado, as crianças pequenas com IRAB representaram uma proporção significativa das despesas, com um carga importante de atendimento. A maioria carecia de fatores de risco. A aplicação de técnicas de tratamento em cuidados moderados foi eficaz, permitindo uma diminuição da demanda por leitos de terapia intensiva.

El virus respiratorio sincicial: patógeno de niños... y de grandes / Respiratory syncytial virus: a pathogen for the small ones and the big ones

Desde el descubrimiento del virus respiratorio sincicial (VRS) en 1956, se ha demostrado en todo el mundo su impacto como el principal causante de infecciones respiratorias agudas bajas (IRAB) que requieren hospitalización en el lactante. Posteriormente se ha descrito que una inadecuada respuesta inmune favorece reinfecciones en la infancia. Más recientemente, numerosos trabajos epidemiológicos lo han asociado a IRAB en adultos, especialmente de tercera edad y en ciertos pacientes inmunocomprometidos. Se ha avanzado en el conocimiento de la estructura y función de los diferentes componentes del VRS, lo que ha permitido facilitar su diagnóstico y avanzar en estrategias de desarrollo de antivirales y vacunas. En efecto, el diagnóstico de laboratorio de VRS es muy simple en niños, por su alta excreción viral, pero para demostrar su participación en adultos se requieren técnicas de alta sensibilidad. La patogenia de la infección es muy compleja y muchos aspectos todavía no se han aclarado. Intervienen factores dependientes del virus -cepa, dosis infectiva, capacidad del virus de inhibir la respuesta inmune- y del hospedero humano, como edad, enfermedades concomitantes, integridad del aparato inmune y otros. Se menciona que otros factores como frío, humedad ambiental, contaminación aérea, hacinamiento, también actuarían en combinación con los inicialmente mencionados. Es necesario conocer los mecanismos responsables de la adquisición de inmunidad contra el VRS para entender las estrategias usadas en el intento de desarrollar vacunas, cuyos esfuerzos son todavía infructuosos. Actualmente se conoce bastante del VRS como patógeno de niños. Sin embargo, cada día se documenta más su participación en enfermedades de adultos, por lo que haremos un resumen para promover su consideración como posible patógeno respiratorio.

Since respiratory syncytial virus (RSV) was identified in 1956, its impact as the main cause of severe acute lower respiratory infections in infants has been shown. Studies about RSV immunopathogenesis have demonstrated that the host immune response is important in protecting from re-infections. The presence of RSV in exacerbation of chronic diseases as COPD and bronchial asthma in adults and its severity in cases with immunodeficiency has been also related to an inadequate response. The actual knowledge on the molecular structure and functions of the virus has allowed to improve diagnosis and to develop new strategies for vaccines and antiviral drugs. The etiologic diagnosis in children is easier than in adults due to the higher viral shedding; therefore techniques based on antibody reactions (immunofluorescence, immunocromatography, etc) are good enough in this group. By contrast, in adults, highly sensitive molecular techniques are needed. Although the advances in understanding the pathogenesis process in neonates and infants, many pathogenic factors still need to be elucidated. The virus strains, viral loads and immune response have been described as important players; however, the changes on the host immunity to RSV according to age and co-morbidities associated to severity of illness needs to be explored. RSV has been known as a children pathogen, nowadays this agent is being recognized as an important agent in adults, especially in those with chronic diseases, immunodeficiency and in immune-senescence.

Rapid antigen detection test for respiratory syncytial virus diagnosis as a diagnostic tool / Teste rápido de detecção de antígenos para o diagnóstico do Vírus Sincicial Respiratório como ferramenta de diagnóstico

Abstract Objective: The aim of this study was to evaluate the QuickVue® RSV Test Kit (QUIDEL Corp, CA, USA) as a screening tool for respiratory syncytial virus in children with acute respiratory disease in comparison with the indirect immunofluorescence assay as gold standard. In Brazil, rapid antigen detection tests for respiratory syncytial virus are not routinely utilized as a diagnostic tool, except for the diagnosis of dengue and influenza. Methods: The authors retrospectively analyzed 486 nasopharyngeal aspirate samples from children under age 5 with acute respiratory infection, between December 2013 and August 2014, the samples were analyzed by indirect immunofluorescence assay and QuickVue® RSV Test kit. Samples with discordant results were analyzed by real time PCR and nucleotide sequencing. Results: From 313 positive samples by immunofluorescence assays, 282 (90%) were also positive by the rapid antigen detection test, two were positive only by rapid antigen detection test, 33 were positive only by immunofluorescence assays, and 171 were positive by both methods. The 35 samples with discordant results were analyzed by real time PCR; the two samples positive only by rapid antigen detection test and the five positive only by immunofluorescence assays were also positive by real time PCR. There was no relation between the negativity by QuickVue® RSV Test and viral load or specific strain. The QuickVue® RSV Test showed sensitivity of 90%, specificity of 98.8%, predictive positive value of 99.3%, and negative predictive value of 94.6%, with accuracy of 93.2% and agreement κ index of 0.85 in comparison to immunofluorescence assay. Conclusions: This study demonstrated that the QuickVue® RSV Test Kit can be effective in early detection of Respiratory syncytial virus in nasopharyngeal aspirate and is reliable for use as a diagnostic tool in pediatrics.

Resumo Objetivo: Avaliar o teste QuickVue® RSV Test Kit (QUIDEL Corp, CA, EUA) para o diagnóstico rápido do vírus sincicial respiratório em crianças com doença respiratória aguda, comparandoo com a imunofluorescência indireta como padrão ouro. Visto que, no Brasil, testes rápidos para detecção de antígenos para vírus sincicial respiratório não são rotineiramente utilizados como ferramenta de diagnóstico, exceto para Dengue e Influenza. Métodos: Um total de 486 amostras de aspirado de nasofaringe de crianças menores de 5 anos com doença respiratória aguda, coletadas entre dezembro de 2013 e agosto de 2014, foram analisadas por imunofluorescência e pelo teste QuickVue®. Amostras com resultados discordantes entre os métodos foram submetidas a PCR em tempo real e sequenciamento. Resultados: Das 313 amostras positivas por IFI, 282 foram positivas no teste rápido (90%), 2 amostras foram positivas apenas no teste rápido (0.6%), 33 apenas na imunofluorescência (10.5%) e 171 foram negativas em ambos os métodos. As 35 amostras com resultados discordantes foram testadas por PCR em tempo real, sendo que duas que foram positivas apenas no teste rápido e 5 apenas na imunofluorescência confirmaram-se positivas. Não houve relação entre a ausência de positividade no teste QuickVue® com a carga ou com a cepa viral. O teste QuickVue® mostrou sensibilidade de 90.1%, especificidade 98.9%, valor preditivo positivo 99.3%, valor preditivo negativo de 94.6%, acurácia de 93.2% e índice de concordância de 0.85 em comparação à imunofluorescência. Conclusões: Nosso estudo demonstrou que o teste QuickVue® RSV pode ser efetivo na detecção precoce do vírus sincicial respiratório em amostras de aspirado de nasofaringe e é confiável como uma ferramenta de diagnósticos em pediatria.

Hospitalizaciones por IRAB de probable etiología viral en niños durante el año 2012: estrategia asistencial / Child hospitalizations for acute lower respiratory infections probably caused by virus in 2012: a healthcare strategy

Introducción: en el Hospital Pediátrico del Centro Hospitalario Pereira Rossell (HP-CHPR) durante los meses fríos, las infecciones respiratorias agudas bajas (IRAB) determinan gran demanda asistencial. Para afrontarlas se pone en práctica una estrategia de diagnóstico y tratamiento denominada Plan de Invierno (PI), que se planifica anualmente, en función de las características de los pacientes tratados en años anteriores. Objetivos: describir las características epidemiológicas, clínicas y evolutivas de los niños que requirieron admisión por IRAB en el HP-CHPR durante los meses fríos del 2012 y describir las principales acciones de la estrategia Plan Invierno del año 2012 (PI-2012). Metodología: se describieron las características epidemiológicas, clínicas y evolutivas de los menores de 24 meses que egresaron del HP-CHPR, en el marco de la estrategia PI-2012, por IRAB de probable etiología viral. Se describió las principales acciones de la estrategia de diagnóstico y tratamiento realizada. Resultados: durante el PI-2012 egresaron 887 niños (25,5% de los egresos hospitalarios). La mediana de edad fue 4 meses. Menos de 10% tenía factores de riesgo para enfermedad grave. El virus respiratorio sincitial (VRS) fue el más frecuentemente identificado. Se aplicó un protocolo de diagnóstico y tratamiento. Se pusieron en práctica técnicas especiales de tratamiento en salas de cuidados moderados. Se aplicó ventilación no invasiva (VNI) en 87 niños, con éxito en 80% de los casos. Se realizó oxigenación de alto flujo (OAF) a 22 niños, con éxito en 100%. Requirieron ingreso a terapia intensiva (CTI) 149 niños, 55,7% requirió asistencia ventilatoria mecánica (AVM). Dos pacientes fallecieron en salas de cuidados moderados, cuatro en CTI. Conclusiones: las IRAB representaron una causa muy importante de morbimortalidad durante el período descripto. Los niños que requirieron ingreso hospitalario eran en su mayoría pequeños. Un porcentaje importante cursó una enfermedad grave, requiriendo modalidades especiales de tratamiento. Es importante continuar en la búsqueda de la mejor estrategia terapéutica para estos niños.

Introduction: during the colder months, acute lower respiratory infections (ALRI) determine a considerable increase in the demand for care at the Pereira Rossell Pediatric Hospital (HP-CHPR). In order to respond to such demand the hospital implements a diagnosis and treatment strategy called Winter Plan (WP), which is planned annually based on the characteristics of patients treated in previous years. Objectives: to describe the epidemiological and clinical characteristics and evolution of children with ALRI who required hospitalization in the HP-CHPR during the cold months of 2012, and to describe the main actions of the 2012 Winter Plan strategy (2012 WP). Methodology: epidemiological, clinical and evolution characteristics of children under 24 months discharged from HP-CHPR within the 2012WP strategy were described. The strategy’s main action for diagnosis and treatment were described. Results: 887 children were discharged during the 2012 WP strategy, representing 25.5% of hospital discharges. Median age was 4 months. Less than 10% presented risk factors for severe disease. Respiratory syncytial virus (RSV) was the most frequently identified virus. A protocol for diagnosis and treatment was applied. Special techniques for treatment were implemented in transitional care rooms. 87 children were administered non-invasive ventilation, which was successful in 80% of cases. High-flow oxygen was used in 22 children, being this successful in all patients. 149 children required admission to intensive care unit (ICU) , 55.7% needed mechanical ventilation assistance. Two patients died in the medium unit and four in the ICU. Conclusions: ALRI represent a major cause of morbidity and mortality during the period described . Most children who required hospitalization were young. A significant percentage presented a severe disease, requiring special treatment modalities. It is important to continue searching for the best therapeutic strategy for these children.

Virus respiratorio sincicial: Patrón clínico epidemiológico en niños internados en un hospital pediátrico durante los años 2000-2013 / Respiratory syncytial virus: Clinical and epidemiological pattern in pediatric patients admitted to a children's hospital between 2000 and 2013

Introducción. El virus respiratorio sincicial (VRS) es el principal agente asociado a infección respiratoria aguda baja en niños. El objetivo de este estudio fue describir el patrón clínico-epidemiológico e identificar los factores de riesgo de infección por VRS. Población y métodos. Estudio prospectivo de cohorte de pacientes internados por infección respiratoria aguda baja en el Hospital de Niños Ricardo Gutiérrez, marzo-noviembre, 20002013. El diagnóstico viral para VRS, adenovirus, influenza y parainfluenza se realizó por inmunofluorescencia indirecta de aspirados nasofaríngeos. Resultados. Se incluyeron 12 555 niños; 38,2% (4798) presentaron rescate viral; el VRS representó el 81,8% (3924/4798) sin variaciones anuales significativas (71,2-88,1), con patrón epidémico estacional (mayo-julio); fue seguido por influenza (7,6%), parainfluenza (5,9%) y adenovirus (4,7%). Los casos con rescate de VRS (3924) tuvieron una mediana de edad de 7 meses (0-214 meses); 74,2% eran menores de 1 año; 43,1%, menores de 6 meses; 56,5%, varones; y la manifestación clínica más frecuente fue bronquiolitis (60,7%). El 41,6% tenía comorbilidades; las más frecuentes, enfermedad respiratoria crónica (74%), cardiopatías congénitas (14%) y enfermedad neurológica crónica (10,2%). El 25% presentó complicaciones. La letalidad fue 1,9% (74/3888). Los predictores independientes de infección por VRS fueron la edad < 3 meses OR 2,8 (2,14-3,67), p < 0,01 , la bronquiolitis como presentación clínica OR 1,54 (1,32-1,79), p < 0,01 y la presencia de hipoxemia al momento del ingreso OR 1,84 (1,42-2,37), p < 0,01 . Conclusiones. La infección por VRS presentó un patrón epidémico estacional y se asoció más a niños pequeños menores de tres meses con bronquiolitis e hipoxemia al momento del ingreso.

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Identificación de virus respiratorios en pacientes pediátricos / Identification of respiratory viruses in pediatric patients

Los virus son la principal causa de infecciones respiratorias agudas. Para la detección de los virus respiratorios se emplea la Inmunofluorescencia directa (IFD). Objetivos: Describir los resultados de las IFD positivas según edad de los pacientes y estacionalidad y determinar el porcentaje de positividad. Material y métodos: Estudio descriptivo retrospectivo. Se incluyeron todas las IFD realizadas en pacientes <15 años en el Hospital de Niños de Viña del Mar entre enero 2012 y noviembre 2013. Resultados: Se realizaron 32015 IFD en 9012 pacientes en ambos años. El virus más frecuente es el Virus Respiratorio Sincicial (VRS) (51 por ciento 2012, 63 porciento 2013), obteniendo un mayor número de casos en pacientes menores de 1año. El Adenovirus (ADV) ocupa el segundo lugar con una incidencia uniforme a lo largo del año. Se observó una disminución de los casos de Influenza A y B desde el año 2012 al 2013. El porcentaje de positividad global de los exámenes es muy bajo (4,25 por ciento 2012 y 3,19 por ciento 2013). El VRS alcanza la mayor positividad en invierno (31,8 por ciento 2012 y24,7 por ciento 2013) y llega al 0 por ciento durante el verano. Conclusiones: Nuestros resultados se asemejan a la epidemiología nacional principalmente en la distribución de los virus según edad y estacionalidad. El virus más frecuentemente encontrado es el VRS. Hubo menos casos de Influenza A durante el 2013 en comparación con los resultados en el país. Existe una solicitud aumentada de exámenes lo que lleva a un porcentaje de positividad muy bajo...

The main cause of acute respiratory infections is viral. The direct immunofluorescence (IF) is used in the detection of respiratory viruses. Objectives: To describe the results of positive IF in relationship to the patient´s age and seasonality and to determine the percentage of positive results. Material and Methods: A descriptive retrospective study that includes all IF performed in patients <15 years old in a private clinic in the V region from January 2012 to November 2013. Results: 32015 IF were conducted in 9012 patients during both years. The Respiratory Syncytial Virus (RSV) is the most common virus (51 percent 2012, 63 percent 2013), obtaining a larger number of cases in patients under the age of 1. The second most common virus is Adenovirus (ADV) with a uniform incidence during the year. A decrease in cases of Influenza A and B was observed from 2012 to 2013. The percent positivity of the tests is very low (4.25 percent and 3.19 percent 2012 2013). The RSV positive testing is higher during the winter (31.8 percent and 24.7 percent 2012 2013) and 0 percent during the summer. Conclusions: Our results are similar to the ones obtained by the national epidemiology in terms of age and season relationship. The RSV is the most common virus. There were fewer cases of influenza A in 2013 compared with results in the country. Due to high demand in IF testing, a very low percentage of positive results is observed...

A survey strategy for human respiratory syncytial virus detection among haematopoietic stem cell transplant patients: epidemiological and methodological analysis

Human respiratory syncytial virus (HRSV) causes severe infections among children and immunocompromised patients. We compared HRSV infections among Haematopoietic Stem Cell Transplant program (HSCT) patients and children using direct immunofluorescence (DFA), point-of-care RSV Bio Easy® and a polymerase chain reaction (PCR) assay. Overall, 102 samples from HSCT patients and 128 from children obtained positivity rate of 18.6% and 14.1% respectively. PCR sensitivity was highest mainly on samples collected after five days of symptoms onset. A combination of both DFA and reverse transcriptase-PCR methods for HSCT high-risk patients is the best diagnostic flow for HRSV diagnosis among these patients.

Predictores de coqueluche al ingreso en lactantes hospitalizados con infección respiratoria aguda baja / [Pertussis predictors in hospitalized infants with acute lower respiratory tract infection].

BACKGROUND: Pertussis, or whooping cough, caused by Bordetella pertussis (BP) is a re-emerging problem in our environment. Although generally considered that the disease is relatively easy to identify infections respiratory syncytial virus (RSV) can present with similar symptoms in infants remains difficult discrimination. OBJECTIVE: Compare clinical symptoms at admission and complementary studies in infants hospitalized with acute respiratory infection (ARI) and RSV by BP to establish markers that enable their early clinical prediction. MATERIALS AND METHODS: Observational, analytical, case-crossover cross comparing younger than 6 months hospitalized with suspected IRA and pertussis (2007-2012) in which BP identified (PCR and culture) and / or VRS (immunofluorescence in nasal secretions). Coinfections were excluded. Bivariate analysis was performed by calculating OR with 95

CI. Were considered significant at p <0.05. The variables studied were age, sex, hits cough, cyanosis, vomiting, apnea, wheezing and CBC with differential RESULTS: We included 174 infants, 72 (41

) BP and 102 (59

) VRS. Age 2 ± 1 months (range :1-6). In both groups was documented cough and wheeze (OR: 1.2 (0.9 to 1.5) p: 0.1 and OR = 0.9 (0.8 to 1.06) p: 0.2, respectively ). Cyanosis (87

, OR: 13.4 p <0.01) and vomiting (26

, OR: 3.4 p <0.01) were more frequent in infants with BP. The absolute lymphocyte count was significantly higher in children with BP (9387 ± 6317 vs. 5127 ± 2766, p <0.01). By ROC curve was identified at 9000 cells / ml as the best point to differentiate VSR BP (AUC = 0.73, 95

CI :0,64-0, 81). CONCLUSIONS: In infants under 6 months with IRA income presence of apnea, cyanosis and lymphocytosis allowing predict significantly differentiate between pertussis those with RSV infections.

Diagnóstico virológico de la infección por virus sincitial respiratorio / Virological diagnosis of respiratory syncytial virus

La enfermedad respiratoria aguda se define como un conjunto de infecciones del tracto respiratorio que pueden ser causadas por una gran variedad de microorganismos tanto virales como bacterianos, y que constituyen un importante problema de salud pública en el mundo. La infección por Virus Sincitial Respiratorio (VSR) está catalogada como una de las principales causas de enfermedad respiratoria aguda, presentándo se especialmente en niños menores de dos años. La falta de diagnóstico confiable de la etiología en las infecciones respiratorias, da como resultado un manejo inadecuado de los pacientes, lo cual puede originar varios tipos de complicaciones. Por tal razón, en esta revisión de la literatura nos enfocamos en presentar un panorama general de la situación de las infecciones respiratorias debidas a VSR en Latinoamérica y las principales dificultades que se presentan al realizar el diagnostico virológico. Para el caso puntual del VSR debido a que todos los agentes etiológicos producen signos y síntomas similares, estos no pueden ser tomados como referencia para distinguir el agente etiológico asociado, así que, en este trabajo se describen las estrategias para realizar el diagnostico de VSR como por ejemplo los que se encargan de detectar anticuerpos específicos en el suero y también los métodos de detección del virus directamente en la muestra de secreción respiratoria, es decir el aislamiento viral en cultivo celular, la detección de antígenos por fluorescencia y la detección de ácidos nucleicos.

Acute respiratory disease is defined as a set of respiratory tract infections that can be caused by a variety of both viral and bacterial microorganisms, which constitute a major public health problem in the world. Respiratory Syncytial Virus (RSV) infection is listed as a major cause of acute respiratory disease, occurring especially in children less than two years old. The lack of accurate diagnosis of respiratory infections etiology, results in an inadequate management of patients, which can cause lots of complications. Thus, this review is focused on presenting an overview of the status of respiratory infections due to RSV in Latin America and the main difficulties related with performing the virological diagnosis. For the specific case of RSV, since all etiologic agents produce similar symptoms, clinical signs cannot be taken as a reference to distinguish the etiological agent of the disease, so, in this paper we describe strategies for diagnosis of RSV such as those that detects specific antibodies in serum and also methods that directly detects the virus in the respiratory secretion sample, in other words, the viral isolation in cell culture, antigens and nucleic acid detection.

Respiratory syncytial virus, infants and intensive therapy

The aims of this study were to determine the presence of respiratory syncytial virus (RSV) and to assess the clinical features of the disease in infants with acute low respiratory tract infection hospitalized at pediatric intensive care units (PICU) of two university teaching hospitals in São Paulo State, Brazil. Nasopharyngeal secretions were tested for the RSV by the polymerase chain reaction. Positive and negative groups for the virus were compared in terms of evolution under intensive care (mechanical pulmonary ventilation, medications, invasive procedures, complications and case fatality). Statistical analysis was performed using the Mann Whitney and Fisher's exact tests. A total of 21 infants were assessed, 8 (38.1 percent) of whom were positive for RSV. The majority of patients were previously healthy while 85.7 percent required mechanical pulmonary ventilation, 20/21 patients presented with at least one complication, and the fatality rate was 14.3 percent. RSV positive and negative groups did not differ for the variables studied. Patients involved in this study were critically ill and needed multiple PICU resources, independently of the presence of RSV. Further studies involving larger cohorts are needed to assess the magnitude of the impact of RSV on the clinical evolution of infants admitted to the PICU in our settings.

Duplex-PCR assay for the detection of adenovirus and respiratory syncytial virus in nasopharyngeal samples

Human adenovirus (HAdV) and human respiratory syncytial virus (HRSV) are important etiologic agents of acute respiratory infections. In this study, a duplex polymerase chain reaction (PCR) assay was developed for the simultaneous detection of HAdV and HRSV in clinical samples. Sixty previously screened nasopharyngeal aspirates were used: 20 HAdV-positive, 20 HRSV-positive and 20 double-negative controls. Eight samples were positive for both viruses. The duplex PCR assay proved to be as sensitive and specific as single-target assays and also detected the mixed infections with certainty. The identification of both viruses in a single reaction offers a reduction in both cost and laboratory diagnostic time.