Prevención de Infecciones Respiratorias por VSR: Consenso Interinstitucional

Las infecciones por el VSR en constituyen una causa importante de morbilidad, hospitalización, ausentismo escolar y laboral, así como de mortalidad, en el mundo, así como en el Paraguay.En la actualidad, existen herramientas para la prevención del VSR. En el año 2012, el Paraguay, ha incorporado el palivizumab (MedImmune, EE. UU.), anticuerpo monoclonal, producido por tecnología de DNA recombinante. Este anticuerpo se administra en 5 dosis, cada 30 días y está indicado en lactantes nacidos prematuros y aquellos con trastornos cardiopulmonares. Por otro lado, actualmente se cuenta con una nueva herramienta para la prevención del VSR. El anticuerpo monoclonal de vida media prolongada Nirsevimab, específico para la conformación de prefusión de la proteína F, aprobado por EMA y FDA. Este monoclonal, está indicado para la prevención de las Infecciones respiratorias agudas bajas, causada por VSR en recién nacidos y lactantes nacidos durante o al ingresar a su primera temporada de VSR, en prematuros y lactantes hasta los 24 meses de edad que siguen siendo vulnerables a la enfermedad grave por VSR durante su segunda temporada de VSR. Luego de analizar la situación epidemiológica del VSR en el país así como la evidencia de eficacia, eficiencia y efectividad de este monoclonal; instituciones académicas, sociedades científicas, organizaciones no gubernamentales y gubernamentales se reunieron y elaboraron un consenso interinstitucional para la prevención de las infecciones por VSR, sugiriendo la incorporación del Nirsevimab, en menores de 12 meses de edad antes de su primera temporada de VSR y en reemplazo del Palivizumab, debido a que el nuevo monoclonal tiene el potencial de cambiar el panorama de las infecciones por VSR en el lactante y producir un impacto en la reducción la mortalidad y morbilidad infantil; reduciendo la carga al sistema de salud, en lo que se refiere a la disminución de la ocupación de camas hospitalarias tanto en sala como en unidades de cuidados intensivos, así como la disminución de la carga para los cuidadores, médicos y proveedores de atención médica y la mortalidad infantil.

Respiratory syncytial virus (RSV) infections constitute a significant cause of morbidity, hospitalization, school and work absenteeism, as well as mortality worldwide, including in Paraguay. Currently, tools for RSV prevention are available. In 2012, Paraguay approved the use of palivizumab (MedImmune, USA), a monoclonal antibody produced through recombinant DNA technology. This antibody is administered in 5 doses, every 30 days and is indicated in infants born prematurely and those with cardiopulmonary disorders. Furthermore, a novel tool for RSV prevention has recently become available. Nirsevimab, a long-acting monoclonal antibody specific to the prefusion conformation of the F protein, has been approved by both the European Medicines Agency (EMA) and the Food and Drug Administration (FDA). This monoclonal antibody is indicated for the prevention of acute lower respiratory tract infections caused by RSV in newborns and infants born during or entering their first RSV season, as well as in premature infants and infants up to 24 months of age who remain vulnerable to severe RSV disease during their second RSV season. After analyzing the epidemiological situation of RSV in our country and evaluating the evidence of efficacy, efficiency, and effectiveness of this monoclonal antibody, academic institutions, scientific societies, and non-governmental and governmental organizations developed consensus guidelines on a new prevention alternative for the prevention of RSV infections, suggesting the incorporation of Nirsevimab in children under 12 months of age before their first RSV season and replacing Palivizumab. The new monoclonal has the potential to change the panorama of RSV infections in infants and produce an impact on the reduction of infant mortality and morbidity reducing the burden on the health system by decreasing hospital bed occupancy both in wards and in intensive care units, as well as the decrease in the burden on caregivers, physicians and health care providers.

Gravedad y complicaciones en lactantes con infección por el virus respiratorio sincitial posterior a la pandemia por SARS-CoV-2 / Severity and complications in infants with respiratory syncytial virus infection after the SARS-CoV-2 pandemic

Resumen Introducción: Posterior a la pandemia por SARS-CoV-2 se ha observado un incremento en la hospitalización por virus respiratorio sincitial (VRS), con mayores complicaciones. Se han encontrado alteraciones extrapulmonares asociadas, disfunción biventricular y lesión renal aguda, entre otras. El objetivo de este estudio fue analizar la evolución y las complicaciones en niños hospitalizados con enfermedad respiratoria de vías bajas secundaria a infección por VRS tras la pandemia de COVID-19. Métodos: Se incluyeron todos los menores de 2 años que ingresaron al servicio de urgencias con infección por VRS. Se analizaron las características clínicas, la necesidad de oxígeno suplementario, el uso de aminas, el índice de angina renal y el requerimiento de terapia de sustitución renal. Se realizó ecografía pulmonar al ingreso. En el análisis estadístico, para las variables cuantitativas se determinaron la media y la desviación estándar, y para las variables cualitativas la frecuencia y el porcentaje. Se evaluaron las diferencias de la distribución con la prueba exacta de Fisher. Resultados: Hubo 45 pacientes con infección por VRS. El 26.7% requirieron ventilación mecánica invasiva y el 11.1% diálisis peritoneal. La letalidad fue de cuatro casos, tres de ellos menores de 12 meses con puntuación de LUS > 7; esto contrasta con el 90.2% de los sobrevivientes con puntaje < 7 (p = 0.0004). Conclusiones: Se observó un aumento en la incidencia de bronquiolitis tras la pandemia, en más de la mitad de los casos con cuadros de moderados a graves, y todos requirieron oxígeno suplementario al ingreso. La lesión renal aguda fue la manifestación extrapulmonar más frecuente.

Abstract Background: After the SARS-CoV-2 pandemic, there has been an increase in hospitalization for lower respiratory infection secondary to respiratory syncytial virus (RSV), with greater complications. Associated extrapulmonary alterations, biventricular dysfunction, acute kidney injury, among others, have been found. The objective of this study was to analize the evolution and complications in hospitalized children with lower respiratory infection secondary to RSV after COVID-19 pandemic. Methods: All pediatric patients under 2 years of age admitted to the emergency department with RSV infection were included. Clinical characteristics, need for supplemental oxygen, use of amines, renal angina index, and requirement for renal replacement therapy were analyzed. Lung ultrasound was performed upon admission. Statistical analysis was carried out for the quantitative variables by means of mean and standard deviation, and qualitative variables by frequency and percentage. Differences in the distribution were evaluated with Fisher’s exact distribution. Results: 45 patients with RSV infection were identified, 26.7% required invasive mechanical ventilation and 11.1% requiered peritoneal dialysis. Fatality was observed in four cases, three of these younger than 12 months with a LUS score > 7; contrasts with 90.2% of survivors with a score < 7 (p = 0.0004). Conclusions: An increase in the incidence of bronchiolitis after pandemic was observed, with more than half having moderate to severe symptoms and requiring supplemental oxygen support in all patients upon admission. Acute kidney injury is the most common extrapulmonary manifestation.

Actualización de las recomendaciones de vacunación en adultos con enfermedades respiratorias. Documento de la Asociación Argentina de Medicina Respiratoria para los neumonólogos, 2023 / Recommendations in Adults with Respiratory Diseases. Document from the Argentinian Association of Respiratory Medicine for Pulmonologists, 2023

RESUMEN Los adultos fumadores, con comorbilidades, y los ancianos tienen mayor riesgo de contraer infecciones pulmonares y de tener peor evolución. La neumonía adquirida en la comunidad debida a virus, neumococo, además de otras bacterias y microorga nismos "atípicos" afecta tanto a adultos sanos como enfermos. La vacuna antigripal se diseña el verano anterior orientada a las cepas esperadas para la temporada siguiente. Su eficacia depende fundamentalmente de la variante viral que finalmente sea la responsable del brote. La vacuna anti-neumocócica polisacárida existe desde 1983 y será inexorablemente reemplazada por vacunas conjugadas de mayor eficacia, que previenen la infección por los serotipos presentes en la vacuna. La inmunización contra SARS-CoV-2 aceleró la reducción del contagio y la gravedad de COVID-19 notablemente. La vacuna acelular para Bordetella pertussis no está en el calendario de adultos, aun cuando vacunarlos fortalece el control del contagio infantil. La vacunas doble bacteriana (difteria y tétanos), y triple (doble + pertusis), y contra sarampión, varicela, rubeola, HPV, Haemophylus influenzae, meningococo, herpes zóster, fiebre hemorrágica argentina y fiebre amarilla están disponibles, pero son de uso limitado. Nuevas vacunas, como la recientemente aprobada por los CDC contra el virus sincicial respiratorio, pronto estarán disponibles.

ABSTRACT Adult smokers, subjects with comorbidities, and the elderly are at higher risk of pulmo nary infections and worse outcomes. Community-acquired pneumonia due to viruses, pneumococcus, other bacteria, and "atypical" microorganisms affects healthy and sick adults. The flu vaccine is designed the previous summer for the strains expected for the following season. Its effectiveness depends fundamentally on the viral variant ultimately responsible for the outbreak. The anti-pneumococcal polysaccharide vaccine has been available since 1983 and it is expected to be replaced by conjugate vaccines which are more effective in preventing infection due to serotypes present in the vaccine. Immuniza tion against SARS-CoV-2 diminished contagion and severity of COVID-19 remarkably. The acellular vaccine for Bordetella pertussis is not on the schedule for all adults, even when vaccinating them strengthens the control of contagion in children. Double bacterial (diphtheria and tetanus), triple (double + pertussis), measles, varicella, rubella, HPV, Haemophilus influenzae, meningococcal, herpes zoster, Argentine hemorrhagic fever, and yellow fever vaccines are available, but their use is limited. New vaccines such as the one recently approved by the CDC against respiratory syncytial virus will soon be available.

Preparation and immunization effect evaluation of recombinant F protein vaccine of respiratory syncytial virus / 中国生物制品学杂志

@#Objective To prepare recombinant F protein vaccine of respiratory syncytial virus(RSV) and evaluate its immunization effect.Methods Two RSV vaccines based on RSV F protein were prepared:one was a mucosal vaccine with bacterial like particle(BLP)as adjuvant and the other was an injectable vaccine with aluminium hydroxide as adjuvant.Forty female BALB/c mice were randomly divided into four groups:BLP-F,BLP control,AL-F and AL control group,with 10mice in each group.BLP-F and BLP control group were inoculated intranasally,and AL-F and AL control group were inoculated subcutaneously.The mice were immunized once each at day 0,14 and 28,respectively.Two weeks after the last immunization,the titers of serum IgG antibody and IgA antibody in nasal lotion were detected by ELISA,and the titers of neutralizing antibody were detected by plaque test.Results Both vaccines induced high levels of serum binding antibodies and neutralizing antibodies,and the induction capacity of injected vaccine was stronger than that of mucosal vaccine.The injected vaccine induced the increase of IgG in serum,which was about 10 times higher than the mucosal immune response,but could not induce the increase of IgA.However,the mucosal vaccine induced the high level of mucosal IgA,but the serum IgG antibody was relatively low.Conclusion Both vaccines based on RSV F protein are promising candidates,and each vaccine has its own advantages.Follow-up studies will evaluate the feasibility of these two vaccines as immunogens using a combination immunization approach to simultaneously enhance systemic and mucosal immune responses against RSV.

Tribute to early pioneering research: A review of original studies on respiratory syncytial virus infection in pediatric patients in China / 中华微生物学和免疫学杂志

Respiratory syncytial virus (RSV) is the leading cause of lower respiratory tract infections in infants and young children worldwide. The study on RSV infection in children dates back to the early 1970s. Looking back at the history, we can see a close association between scientific research and pediatric clinical needs, and how solid data are obtained by strictly following the classic methodology for viral etiological study. The original studies on respiratory syncytial virus infection in children lay the foundation for RSV research in China.

Intranasal immunization with single-dose vaccine based on recombinant influenza virus H1N1 expressing the extracellular domain of respiratory syncytial virus G protein induces robust immunity and protection in mice / 中华微生物学和免疫学杂志

Objective:To construct a novel respiratory syncytial virus (RSV) vaccine based on a recombinant influenza virus vector and evaluate its immune protective effects in mice.Methods:A recombinant H1N1 influenza A virus (IAV) expressing the extracellular domain (Gecto) of RSV A2 G protein was constructed and rescued, named as PR8NAGecto/WSN. After in vitro verification of the Gecto expression and PR8NAGecto/WSN growth kinetics, a single dose of PR8NAGecto/WSN was used to immunize BALB/c mice through intranasal administration to evaluate the efficacy of PR8NAGecto/WSN by assessing humoral (IgG, neutralizing antibody), mucosal (IgA) and cellular immunity (IFN-γ ELISPOT). Four weeks after immunization, the mice were challenged with RSV A2 or RSV B9320 to evaluate the protective effects of PR8NAGecto/WSN by analyzing mouse body weight changes, lung tissue virus titers and pathological changes. Results:A single-dose intranasal immunization with PR8NAGecto/WSN induced robust humoral, mucosal and cellular immunity in mice. Moreover, the mice in the immunized group had lower lung virus loads and mild lung pathological damages following the challenge with RSV A or RSV B subtype as compared with the control group.Conclusions:A single-dose intranasal immunization with PR8NAGecto/WSN induces robust immunity and provide protection against RSV A and B challenges in mice. This study provides new ideas and reference for the development of novel mucosal vaccines against RSV.

Preparation and immunogenicity evaluation of two PreF trimer recombinant protein vaccines against respiratory syncytial virus / 中华微生物学和免疫学杂志

Objective:To construct and purify four respiratory syncytial virus (RSV) PreF proteins through gene sequence design and optimization and evaluate their immunogenicity.Methods:Coronin-1A and T4 trimer protein gene sequences were optimized with Human and CHO codons, and then added to RSV F protein sequence. The above plasmids were transfected into Expi293F cells for protein expression. After purification by nickel column, four trimer proteins were prepared. SDS-PAGE and Western blot were performed for protein identification. BALB/c mice were immunized at week 0 and week 3, and blood samples were collected to measure the activities of binding and neutralizing antibodies in serum.Results:SDS-PAGE and Western blot showed that the four proteins had stable trimer structure. Antigen-antibody affinity test showed that the four trimer proteins had strong affinity with RSV-specific monoclonal antibodies 8897, D25, Motavizumab, AM14 and Palivizumab. The titers of antibodies induced by the two T4 trimers were higher after the initial immunization, while there was a substantial increase in the titers of antibodies induced by Human codon-optimized trimer protein after the second immunization.Conclusions:PreF trimer protein can be prepared by adding any of the two different heterotrimer motifs, and induce effective binding and neutralizing antibodies in mice.

Immunogenicity of PreF and PostF recombinant protein vaccines against respiratory syncytial virus / 中华微生物学和免疫学杂志

Objective:To compare the immunogenicity of the prefusion (PreF) and postfusion (PostF) conformations of the respiratory syncytial virus (RSV) F protein.Methods:The expression of PreF and PostF recombinant proteins was analyzed by SDS-PAGE and Western blot. The binding affinity between F protein and its specific antibodies was detected by Octet. The binding antibodies and neutralizing antibodies in immune serum were detected after immunizing mice with PreF or PostF recombinant protein.Results:PreF protein was stable in the form of a trimer after modification with higher binding affinity with monoclonal antibodies such as D25, 8897, AM14, Palivizumab and Motavizumab. PostF protein lacked the antigenic site ? and showed a monomer conformation. Besides, it was unable to bind to D25, 8897 and AM14 antibodies. Animal experiments showed that AS01 adjuvant was better than aluminum adjuvant in inducing binding antibodies and neutralizing antibodies against RSV Long strains. The binding antibodies induced by PreF and PostF recombinant proteins had similar binding ability to PreF protein, while the binding antibodies induced by PostF recombinant protein showed stronger binding ability to PostF than to PreF.Conclusions:PreF has more epitopes and the trimer form of PreF recombinant protein after modification is more stable and can induce stronger neutralizing antibodies. Moreover, the immunopotentiating effect of AS01 adjuvant is better than that of aluminum adjuvant. Therefore, stabilization-based trimer structure modification of PreF and the development of adjuvants are crucial for the development of RSV vaccines.

A new variant of respiratory syncytial virus glycoprotein gene and clinical features of infected children / 中华微生物学和免疫学杂志

Objective:To analyze the epidemiological features of respiratory syncytial virus (RSV) infection in Beijing, and monitor the sequence variations in RSV glycoprotein (G) gene and clinical features of infected children.Methods:Respiratory tract specimens were collected from children with acute respiratory infection in the Children′s Hospital Affiliated to Capital Institute of Pediatrics from January 1, 2023 to December 31, 2023. RSV-positive specimens screened by multiple nucleic acid testing were subjected to PCR to amplify the full-length RSV G gene. A phylogenetic tree was constructed after gene sequencing to analyze RSV subtypes and trace G gene variants. Clinical data were retrieved from the medical record system to analyze the clinical features of children with RSV infection in Beijing.Results:A total of 5 489 respiratory specimens were collected from 3 046 male patients and 2 443 female patients. The average age of the patients was 4.36 years. A total of 589 RSV-positive specimens (10.7%, 589/5 489) were detected with 349 from male patients and 240 from female patients. The average age of children with RSV infection was (2.51±2.78) years and the median age was 0.48 years. RSV had been circulating among children in Beijing since March 2023 with two epidemic peaks in May (24.6%, 122/496) and December (18.2%, 126/693). The predominant subtype of RSV in the first half of 2023 was subtype A, but it was replaced by subtype B from November 2023. Phylogenetic analysis revealed a novel G gene of RSV subtype B (RSV-B-BA9-954bp) with a length of 954 bp, which belonged to a new cluster in the phylogenetic tree. The percentage of patients admitted to the Intensive Care Unit (ICU) was higher in children with new variant of RSV subtype B infection than in those with common RSV subtype B infection [44.1% (15/34) vs 25.2% (31/123), χ 2=4.600, P=0.032], while the counts of white blood cells and the levels of C-reactive protein were lower in the children with new variant infection ( P<0.05). Conclusions:RSV has been prevalent among children in Beijing since March 2023 with two epidemic peaks. The predominant A subtype is gradually replaced by to B subtype. A new variant of RSV B G gene (RSV-B-BA9-954bp) is detected among the children.

Transcriptomic analysis of A549 cells infected with ON1 genotype of human respiratory syncytial virus subtype A isolated in Beijing / 中华微生物学和免疫学杂志

Objective:To analyze the differentially expressed genes of human respiratory syncytial virus (RSV) subtype A genotype ON1, a predominant genotype in Beijing, after infecting A549 cells using transcriptomic sequencing, and provide potential targets for RSV prevention and treatment.Methods:A local strain (61397-ON1) identified by whole-genome sequencing as ON1 genotype of RSV subtype A was selected to infect A549 cells. Total mRNA was extracted, and the differentially expressed genes in infected and uninfected A549 cells were screened by transcriptomic sequencing. GO analysis and KEGG pathway analysis were performed. Besides, six genes with differential expression ratio greater than two times were randomly selected for qRT-PCR verification.Results:There were 1 632 differentially expressed genes between infected and uninfected A549 cells, of which 807 genes were up-regulated and 825 genes were down-regulated. The differentially expressed genes were mainly involved in immune response-related biological processes such as cytokine response and positive regulation of MAPK cascades, and were enriched in MAPK signaling pathway, NOD-like receptor signaling pathway, p53 signaling pathway, TNF signaling pathway, IL-17 signaling pathway and NF-κB signaling pathway. The results of qRT-PCR for six differentially expressed genes were consistent with the trend of transcriptome data.Conclusions:The differentially expressed genes of RSV A subtype ON1 genotype after infecting A549 cells are mainly involved in cytokine response and immune-related signaling pathways. This study provides basic data for further study of the molecular mechanism of RSV infection and the development of prevention and treatment strategies.

Molecular epidemiological characteristics of respiratory syncytial virus in hospitalized children with acute respiratory tract infection in Shanghai / 中华微生物学和免疫学杂志

Objective:To investigate the epidemiological characteristics, genotypes and genetic evolution of respiratory syncytial virus (RSV) isolated in Shanghai from April 2020 to December 2021, which was a period from the COVID-19 outbreak to the phase of regular epidemic prevention and control.Methods:This retrospective study collected the nasopharyngeal secretions or nasopharyngeal aspirates of children with acute respiratory tract infection (ARTI) admitted to the Shanghai Children′s Hospital from April 2020 to December 2021. PCR-capillary electrophoresis and RT-PCR were used for virus identification and the amplification of the gene fragment of the second hypervariable region of RSV G protein. Homology analysis and phylogenetic analysis were conducted using bioinformatics software. Chi-square test was used to compare the detection rates of RSV. Results:A total of 6 211 samples were collected and 13.62% (846/6 211) of them were positive for RSV. The positive rates of RSV in male and female patients were 14.07% (503/3 574) and 13.01% (343/2 637), respectively, with no significant gender difference (χ 2=1.467, P=0.226). The highest detection rate of RSV was found in children ≤6 months of age, and the rate of RSV infection decreased gradually with age (χ 2=352.942, P<0.001). No RSV-positive specimens were detected from April 2020 to August 2020, after which the detection rate of RSV gradually increased with two epidemic peaks occurring from December 2020 to February 2021 and from August to October 2021. The predominant epidemic subtype was RSV subtype B in 2020 and the first 9 months of 2021, which was gradually replaced by RSV subtype A in the last 3 months of 2021. The 176 strains of RSV subtype A obtained in this study were all ON1 genotype, and the nucleotide homology of the Shanghai epidemic strains was 90.20%-99.50%. All of the 250 strains of RSV subtype B were BA9 genotype, and the nucleotide homology of the Shanghai epidemic strains was 90.10%-100.00%. Conclusions:From April 2020 to December 2021, with the regular prevention and control of COVID-19, there is a change in the epidemic season of RSV. The prevalent genotypes of RSV subtypes A and B are ON1 and BA9, respectively, and the subtype A gradually replaces subtype B as the most prevalent subtype.

Epidemiological characteristics of respiratory syncytial virus in children in Hebei from 2019 to 2023 / 中华微生物学和免疫学杂志

Objective:To analyze the epidemiological characteristics of respiratory syncytial virus (RSV) among children in Hebei from 2019 to 2023.Methods:A total of 46 576 lower respiratory tract specimens were collected from hospitalized children in the Children′s Hospital of Hebei Province from 2019 to 2023. Multiple RT-PCR and capillary electrophoresis were used to detect 13 common respiratory pathogens in the specimens, and the results were statistically analyzed.Results:The overall positive rate of RSV was 18.76%(8 739/46 576). The overall positive rates of RSV in male and female children were 18.84%(5 174/27 462) and 18.65%(3 565/19 114), respectively, showing no statistically significant difference between genders (χ 2=0.916, P=0.339). A linear relationship was found between the positive rate of RSV and age ( P<0.01). There was a significant difference in the positive rates of RSV in different years (χ 2=723.71, P<0.01). The positive rate of RSV peaked in the period from December to February from 2019 to 2021. In 2019 and 2020, the positive rates of RSV were very low from May to October, while the positive rate of RSV was above 10% throughout the whole year of 2021 and small off-season epidemics occurred in May and August. The positive rate of RSV was low in 2022, and no significant seasonal change was observed. The rate of RSV infections peaked from April to June in 2023. There were significant differences in the rates of RSV infections before, during and after the COVID-19 epidemic in each age group ( P<0.01). The rate of mixed infections was 29.20%(2 522/8 739), and the most common other respiratory pathogen was human rhinovirus (52.29%, 1 342/2 552 ). Conclusions:RSV is a common pathogen causing respiratory tract infections in children in Hebei, especially in children under 3 years old. After the COVID-19 epidemic, there are off-season RSV epidemics. Given the variations in the epidemiological features of RSV, it is necessary to carry out continuous monitoring of RSV to provide scientific data for the prevention and control of related diseases.

Clinical epidemiological characteristics of respiratory syncytial virus infection in children in single center in Tianjin / 中华微生物学和免疫学杂志

Objective:To analyze the epidemiological characteristics of respiratory syncytial virus (RSV) infection in children in Tianjin, and provide reference for the prevention and treatment of RSV infection in children.Methods:Clinical data of 2 743 children with acute respiratory infections treated at the Tianjin Fifth Central Hospital from January 2022 to January 2024 were collected. Multiplex fluorescent PCR was used to detect the nucleic acid fragments of six respiratory pathogens including Mycoplasma pneumoniae, human rhinovirus, adenovirus, influenza A virus, influenza B virus and RSV in the throat swabs of the patients. A retrospective analysis was performed on the epidemiological and clinical data of RSV-RNA positive cases. Results:The positive rate of RSV-RNA in the 2 743 children was 15.09% (414/2 743). The positive rate of RSV-RNA was 9.29% (73/786) in 2022 and 16.53% (302/1 827) in 2023, with a statistically significant difference between the two years (χ 2=23.45, P<0.05). The incidence of RSV infection in winter and spring was significantly different from that in summer and autumn (χ 2=19.46, P<0.05). The highest and the lowest infection rates of RSV were found in winter (19.32%, 193/999) and autumn (9.43%, 45/477), respectively. There was a significant difference in RSV infection rate among different age groups (χ 2=71.38, P<0.05), with the highest infection rate in the age group of 0-2 years (21.18%, 230/1 086), and the lowest infection rate in the age group of 6-8 years (6.29%, 27/429). Among the 414 children with RSV infection, 359 cases (84.97%) were infected with RSV alone, while the other 55 cases (13.29%) were infected with mixed pathogens. Fifty-two cases had co-infection of RSV and one other pathogen. The most common pathogens in co-infection cases were human rhinovirus (4.83%, 20/414) and Mycoplasma pneumoniae (6.04%, 25/414). Conclusions:The RSV infection rate among children with acute respiratory tract infection in Tianjin from 2022 to 2024 was 15.09%, with the highest infection rate in spring and the lowest infection rate in autumn. RSV infection can occur in children of all ages, with the highest infection rate in children aged 0-2 years and the lowest infection rate in children aged 6-8 years. RSV infection is often complicated by other respiratory pathogens, and the most common pathogens are human rhinovirus and Mycoplasma pneumoniae.

Outbreak and clinical features of respiratory syncytial virus in Chengde from 2022 to 2023 / 中华微生物学和免疫学杂志

Objective:To analyze the prevalence and clinical features of respiratory syncytial virus (RSV) in Chengde city.Methods:From August 2022 to June 2023, throat swabs and clinical data of 478 hospitalized children with respiratory tract infection in the Chengde Central Hospital were collected. Real-time quantitative PCR was used to detect the molecular epidemiology of RSV-A and RSV-B subtypes and analyze the clinical features of patients with RSV infection.Results:Among the hospitalized children, 67.57% (323/478) tested positive for RSV. The outbreak of RSV infection was caused by RSV-A subtype. The peaks of RSV-A infection occurred from November to December, 2022 and May to June, 2023. There were 86.07% (278/323) of the RSV-A-positive cases had mixed infection with other pathogens, primarily bacterial pathogens with Streptococcus pneumoniae being the most common, followed by Klebsiella pneumoniae. Influenza virus A was the most common viral pathogens causing mixed infection. The level of lactate dehydrogenase was higher in the patients with single RSV-A infection than in those with mixed infection ( Z=2.396, P=0.017), and higher than the normal upper limit. Compared with the single infection group, the mixed infection group had higher white blood cell count ( Z=2.417, P=0.016), neutrophil ratio ( Z=3.218, P=0.001), C-reactive protein level ( Z=1.998, P=0.046) and creatinine level ( Z=2.107, P=0.035), and lower lymphocyte ratio ( Z=3.205, P=0.001), but they were all within the normal range. There were no significant differences in the clinical features between RSV-A-positive patients co-infected with bacteria or other viruses (all P>0.05). Conclusions:RSV-A is the leading cause of respiratory tract infection in children in Chengde from 2022 to 2023, and often co-detected with bacteria. The mixed infection with other respiratory pathogens is related to the clinical features of patients with RSV-A infection.

Establishing N-ELISA-based method for rapid detection of neutralizing antibodies against respiratory syncytial virus / 中华微生物学和免疫学杂志

Objective:To prepare rabbit polyclonal antibodies against respiratory syncytial virus (RSV) N protein and use them as the detection antibodies to establish a N-ELISA-based method for rapid detection of neutralizing antibodies.Methods:A plasmid of pET30a-N for the expression of RSV N protein was constructed. After purification, the protein was immunized into New Zealand rabbits to prepare polyclonal antibodies, which were used as the detection antibodies. Positive serum samples were diluted and used to neutralize RSV (100 TCID 50/well). Hep-2 cells were inoculated and cultured, and then the cells were fixed with 80% acetone. ELISA was performed to detect RSV N protein in infected cells. When the absorbance value of a well was below the cut-off value, it was regarded as the positive well in the neutralization test. The highest dilution of a positive well serum was the neutralizing antibody titer. After optimizting the antibody dilution, detection time, cell density and the duration of neutralization, the method for neutralizing antibody detection was established based on N-ELISA. The established method was verified by analyzing the influences of different cell generations and edge effects, and calculating the accuracy, repeatability and precision. The correlation between the established method and microneutralization method was analyzed by detecting human RSV IgG-positive serum. Results:The plasmid pET30a-N was successfully constructed, and the expressed N protein showed high purity and good specificity. After the third immunization, the antibody titer in rabbit serum was 1∶51 200, and the antibodies could specifically bind to RSV. The prepared rabbit anti-RSV N polyclonal antibodies had a titer of 1∶51 200, and showed good specificity. The neutralizing antibodies could be detected on day 4 with the established method, and the duration of neutralization was shortened to 30 min. Cell generations and the position of wells in the 96-well plate (edge well and non-edge well) had no significant effect on the method, and the repeatability, precision and accuracy of the method were good. In the detection of 64 RSV IgG-positive human serum samples by the established method and microneutralization method, the correlation coefficient was 0.929 6, indicating a good positive correlation between the two methods.Conclusions:A N-ELISA-based method for rapid neutralizing antibody detection is successfully established, which can be used to evaluate the serum antibody level after RSV vaccination.

Verification of Yunshi Ganmao Heji efficacy against respiratory syncytial virus based on network pharmacology and animal experiments / 中华微生物学和免疫学杂志

Objective:To investigate the mechanism of Yunshi Ganmao Heji against respiratory syncytial virus (RSV) infection based on network pharmacology and in vivo experiments. Methods:Network pharmacological prediction: Several databases including TCMSP and GeneCards were used to predict the active ingredients and targets of Yunshi Ganmao Heji in the intervention of RSV infection. Cytoscape 3.2.1 software was used to construct the traditional Chinese medicine component-disease target network diagram. The interactions between proteins were analyzed by STRING database. GO functional enrichment analysis and KEGG pathway enrichment analysis were performed using Metascape database. Molecular docking technology was used to verify the results of network pharmacology. Experimental verification of Yunshi Ganmao Heji for the intervention of RSV infection: A mouse model of RSV infection was established through intranasal infection. After the administration of Yunshi Ganmao Heji, blood routine test results, lung indexes and pathological changes in lung tissue were analyzed. Peripheral blood T cell subsets were detected by flow cytometry. The levels of TNF-α, IL-6 and IL-1β in serum were detected by ELISA. RT-PCR was used to detect the relative expression of TLR4, NF-κB and RSV-N gene at mRNA level in lung tissues.Results:A total of 41 active ingredients of Yunshi Ganmao Heji and 111 drug targets for RSV infection were obtained. Besides, 167 signaling pathways mainly including PI3K/AKT, MAPK and Toll-like receptor signaling pathways were obtained. Molecular docking results showed that the binding energies of luteotin, kaempferol and quercetin, three active ingredients of Yunshi Ganmao Heji, with RSV-G, RSV-F, PI3K, AKT1 and Bcl-2 were less than 0 kcal/mol. In vivo experiment results showed that compared with RSV group, the counts of white blood cells and lymphocytes increased and the lung index decreased in high-dose Yunshi Ganmao Heji group, with statistically significant difference ( P<0.05). HE staining showed pulmonary hyperplasia, thickened alveolar wall and inflammatory cell infiltration in interstitium in RSV group. Alveoli in ribavirin group as well as low-dose, medium-dose and high-dose Yunshi Ganmao Heji groups tended to be of uniform size, and the alveolar walls was roughly uniform in thickness. Compared with RSV group, the low-dose, medium-dose and high-dose Yunshi Ganmao Heji groups showed significantly increased numbers of CD3 +, CD4 + and CD8 + T lymphocytes, decreased CD4 + /CD8 + T cell ratio, lower levels of TNF-α, IL-6, IL-1β in serum, and reduced viral load and inhibited expression of TLR4 and NF-κB at mRNA level in lung tissues ( P<0.05). Conclusions:Yunshi Ganmao Heji can regulate RSV infection by targeting multiple targets and pathways with several active ingredients. Its main functions are to alleviate pathological injury in lung tissues and reduce inflammatory response, and the possible mechanism underlying the antiviral functions may be related to its inhibitory effect on the activation of TLR4/NF-κB pathway.

Research progess in respiratory syncytial virus receptors / 中华微生物学和免疫学杂志

Respiratory syncytial virus (RSV) is one of the major pathogens of acute respiratory infections, becoming a huge global burden. Virus-receptor interactions play a key role in the pathogenesis of RSV infection. The distribution of receptors influences the cellular and the tissue tropism of RSV as well as the viral replication and proliferation in the host. However, the RSV receptors are currently unknown, which is one of the reasons that hinders the development of RSV vaccines and therapeutic drugs. In this study, the existing RSV receptors are reviewed with the hope to provide ideas for the research of RSV vaccines and therapeutic drugs.

Preparation and immunization effect evaluation of recombinant F protein vaccine of respiratory syncytial virus / 中国生物制品学杂志

@#Objective To prepare recombinant F protein vaccine of respiratory syncytial virus(RSV) and evaluate its immunization effect.Methods Two RSV vaccines based on RSV F protein were prepared:one was a mucosal vaccine with bacterial like particle(BLP)as adjuvant and the other was an injectable vaccine with aluminium hydroxide as adjuvant.Forty female BALB/c mice were randomly divided into four groups:BLP-F,BLP control,AL-F and AL control group,with 10mice in each group.BLP-F and BLP control group were inoculated intranasally,and AL-F and AL control group were inoculated subcutaneously.The mice were immunized once each at day 0,14 and 28,respectively.Two weeks after the last immunization,the titers of serum IgG antibody and IgA antibody in nasal lotion were detected by ELISA,and the titers of neutralizing antibody were detected by plaque test.Results Both vaccines induced high levels of serum binding antibodies and neutralizing antibodies,and the induction capacity of injected vaccine was stronger than that of mucosal vaccine.The injected vaccine induced the increase of IgG in serum,which was about 10 times higher than the mucosal immune response,but could not induce the increase of IgA.However,the mucosal vaccine induced the high level of mucosal IgA,but the serum IgG antibody was relatively low.Conclusion Both vaccines based on RSV F protein are promising candidates,and each vaccine has its own advantages.Follow-up studies will evaluate the feasibility of these two vaccines as immunogens using a combination immunization approach to simultaneously enhance systemic and mucosal immune responses against RSV.

IL2rg-/- rats support prolonged infection of human RSV / 中国实验动物学报

Objective To overcome the limitations of existing human respiratory syncytial virus(hRSV)animal models,such as semi-permissiveness and short duration of infection,this study established an IL2rg gene knockout(IL2rg-/-)rat model using TALEN gene editing technology.Methods The animal model was infected with hRSV intranasally.Clinical characteristics,body weight,and temperature changes were observed over the infection period(0～35 days).The total viral loads in respiratory organs,such as the nasal tissue,trachea,and lungs,were measured at various time points(4,11,20,and 35 days post-infection).Pathological analysis was conducted on target organs at the endpoint of observation(35 days post-infection).Changes in peripheral blood T,B,NK,and NKT cells and various cytokines were assessed at various time points(4,20,and 35 days post-infection).Results(1)IL2rg/-knockout rats sustained high viral loads in the nasal cavity upon intranasal inoculation with hRSV.The average peak titer rapidly reached 1 × 1010 copies/g in nasal tissue and 1 × 107 copies/g up to 5 weeks post-infection.(2)However,no significant pathological changes were noted in nasal,tracheal,or lung tissues.(3)An increase was observed in the content of peripheral blood B cells in hRSV-infected IL2rg--rats.(4)IL-6 and MCP-1 were increased in the early stage of infection and then decreased at the end of the observation period.Conclusions This study established a new IL2rg-/-rat model using TALEN technology and found that this model effectively supported high-level replication and long-term infection of hRSV,providing a good basis for antiviral drug screening and in vivo efficacy evaluation of anti-hRSV antibodies.

Characteristics of respiratory syncytial virus infections in children with severe acute respiratory tract infections in Huzhou City， Zhejiang Province from 2018 to 2022 / 上海预防医学

ObjectiveTo understand the prevalence of respiratory syncytial virus （RSV） infections among children hospitalized with severe acute respiratory infections （SARI） in Huzhou， Zhejiang Province， and to explore the epidemiological characteristics and influencing factors of RSV. MethodsNasopharyngeal swab samples from children under 15 years of age， newly admitted to the Huzhou First People's Hospital from 2018 to 2022 and who met the SARI surveillance definition， were collected for respiratory virus nucleic acid testing. SPSS 25.0 and Excel 2016 software were used for statistical analysis of the data. ResultsA total of 1 076 samples were tested for nucleic acids， with an overall positivity rate of 11.2%. The positivity rate of infants under 5 years old was relatively high （16.7%）. RSV infection showed a seasonal distribution， mainly concentrated in autumn and winter， with the highest positivity rate in December （22.3%）. A total of 23 cases of respiratory mixed infection were detected， with a mixed infection rate of 2.13%， among which 16 were RSV-related mixed infections. Logistic regression analysis indicated that season and age were risk factors for RSV infection. ConclusionFrom 2018 to 2022， RSV showed different epidemiological characteristics across different ages and seasons in SARI cases in Huzhou， with significant differences， providing a scientific basis for future prevention of RSV infections.