Delayed Sequelae of Neonatal Respiratory Syncytial Virus Infection Are Dependent on Cells of the Innate Immune System

Infection with respiratory syncytial virus (RSV) in neonatal mice leads to exacerbated disease if mice are reinfected with the same virus as adults. Both T cells and the host major histocompatibility complex genotype contribute to this phenomenon, but the part played by innate immunity has not been defined. Since macrophages and natural killer (NK) cells play key roles in regulating inflammation during RSV infection of adult mice, we studied the role of these cells in exacerbated inflammation following neonatal RSV sensitization/adult reinfection. Compared to mice undergoing primary infection as adults, neonatally sensitized mice showed enhanced airway fluid levels of interleukin-6 (IL-6), alpha interferon (IFN-α), CXCL1 (keratinocyte chemoattractant/KC), and tumor necrosis factor alpha (TNF-α) at 12 to 24 h after reinfection and IL-4, IL-5, IFN-γ, and CCL11 (eotaxin) at day 4 after reinfection. Weight loss during reinfection was accompanied by an initial influx of NK cells and granulocytes into the airways and lungs, followed by T cells. NK cell depletion during reinfection attenuated weight loss but did not alter T cell responses. Depletion of alveolar macrophages with inhaled clodronate liposomes reduced both NK and T cell numbers and attenuated weight loss. These findings indicate a hitherto unappreciated role for the innate immune response in governing the pathogenic recall responses to RSV infection.     

Respiratory Syncytial Virus Infection of the Newborn

In an outbreak of respiratory syncytial (R.S.) virus infection in a maternity hospital the respiratory illness was of a mild nature and the virus was not found in infants without respiratory symptoms. This confirms the suggestion that R.S. virus can infect infants at a very early age. Rapid diagnosis was achieved by applying the direct fluorescent antibody technique to cells in nasal secretions. This proved to be more sensitive than culture techniques where there was delay between the onset of respiratory symptoms and submission of specimens to the laboratory.     

抗呼吸道合胞病毒的免疫预防

呼吸道合胞病毒(RSV)感染是一个影响婴幼儿健康的全球性的问题,目前尚未有令人满意的治疗药物,免疫预防就显得尤为重要。近年研究表明,免疫预防在疫苗、单克隆抗体以及免疫球蛋白等领域均取得较大进展。     

Bacteremia in Children Hospitalized with Respiratory Syncytial Virus Infection

Background

The risk of bacteremia is considered low in children with acute bronchiolitis. However the rate of occult bacteremia in infants with RSV infection is not well established. The aim was to determine the actual rate and predictive factors of bacteremia in children admitted to hospital due to confirmed RSV acute respiratory illness (ARI), using both conventional culture and molecular techniques.

Methods

A prospective multicenter study (GENDRES-network) was conducted between 2011–2013 in children under the age of two admitted to hospital because of an ARI. Among those RSV-positive, bacterial presence in blood was assessed using PCR for Meningococcus, Streptococcus pneumoniae, Haemophilus influenzae, Streptococcus pyogenes, Klebsiella pneumoniae, Pseudomonas aeruginosa, Escherichia coli, and Staphylococcus aureus, in addition to conventional cultures.

Results

66 children with positive RSV respiratory illness were included. In 10.6% patients, bacterial presence was detected: H. influenzae (n = 4) and S. pneumoniae (n = 2). In those patients with bacteremia, there was a previous suspicion of bacterial superinfection and had received empirical antibiotic treatment 6 out of 7 (85.7%) patients. There were significant differences in terms of severity between children with positive bacterial PCR and those with negative results: PICU admission (100% vs. 50%, P-value = 0.015); respiratory support necessity (100% vs. 18.6%, P-value < 0.001); Wood-Downes score (mean = 8.7 vs. 4.8 points, P-value < 0.001); GENVIP scale (mean = 17 vs. 10.1, P-value < 0.001); and length of hospitalization (mean = 12.1 vs. 7.5 days, P-value = 0.007).

Conclusion

Bacteremia is not frequent in infants hospitalized with RSV respiratory infection, however, it should be considered in the most severe cases.     

Prospects For the Use of Peptides against Respiratory Syncytial Virus

Molecular Biology - The human respiratory syncytial virus (RSV) is one of the most common viral pathogens that affects the lower respiratory tract and could be a reason of bronchiolitis and/or...     

裸鼠呼吸道合胞病毒感染的动物模型

呼吸道合胞病毒(RSV)是全世界婴幼儿下呼吸道感染的首位病毒病原体,免疫缺陷个体容易发生严重感染,目前尚无理想RSV感染动物模型用于研究.我们用细胞免疫缺陷裸鼠感染RSV,旨在建立理想的动物模型,为RSV感染的防治研究奠定基础.裸鼠滴鼻感染RSV后肺组织分离到病毒,直接免疫荧光检测到支气管肺泡灌洗液RSV抗原阳性,空斑形成实验检测肺组织病毒滴度在感染后第3天达高峰,并持续到第9天仍能检测到病毒.免疫组化检测RSV抗原主要分布在细支气管、毛细支气管和肺泡上皮细胞胞浆内.肺组织病理学显示RSV感染导致裸鼠淋巴细胞浸润为主的肺间质性炎症,电镜分析超微结构可见到细胞内病毒颗粒和气血屏障的破坏.支气管肺泡灌洗液白细胞计数显示裸鼠RSV感染炎症高峰在感染后第9天.裸鼠RSV感染的病毒复制和病理改变特点与人相似,病毒持续高水平复制,是客观而实用的评价抗RSV制剂效果的小鼠模型.     

裸鼠呼吸道合胞病毒感染的动物模型

呼吸道合胞病毒(RSV)是全世界婴幼儿下呼吸道感染的首位病毒病原体,免疫缺陷个体容易发生严重感染,目前尚无理想RSV感染动物模型用于研究。我们用细胞免疫缺陷裸鼠感染RSV,旨在建立理想的动物模型,为RSV感染的防治研究奠定基础。裸鼠滴鼻感染RSV后肺组织分离到病毒,直接免疫荧光检测到支气管肺泡灌洗液RSV抗原阳性,空斑形成实验检测肺组织病毒滴度在感染后第3天达高峰,并持续到第9天仍能检测到病毒。免疫组化检测RSV抗原主要分布在细支气管、毛细支气管和肺泡上皮细胞胞浆内。肺组织病理学显示RSV感染导致裸鼠淋巴细胞浸润为主的肺间质性炎症,电镜分析超微结构可见到细胞内病毒颗粒和气血屏障的破坏。支气管肺泡灌洗液白细胞计数显示裸鼠RSV感染炎症高峰在感染后第9天。裸鼠RSV感染的病毒复制和病理改变特点与人相似,病毒持续高水平复制,是客观而实用的评价抗RSV制剂效果的小鼠模型。     

树鼩呼吸道合胞病毒感染动物模型的建立

探讨用灵长类动物树鼠句建立呼吸道合胞病毒(RSV)感染动物模型的可行性。28只树鼠句随机分为7组,每组4只,鼻内滴入106PFU RSV,感染后每24h检测1组。另取7只树鼠句鼻内滴入100μl Hep-2细胞培养液,滴鼻后每24h检测1只作对照。无菌取树鼠句肺组织进行病毒分离、空斑形成实验检测病毒滴度、病理检查和RT-PCR检测肺组织内RSV mRNA表达。结果显示:树鼠句感染RSV后,无明显呼吸道感染症状;树鼠句肺组织匀浆接种于Hep-2细胞上,第3、4、5实验组出现细胞病变效应(CPE);树鼠句感染RSV后肺部病理改变在3~7d较为明显,主要为间质改变;在106PFU感染浓度下树鼠句肺组织内RSV处于低水平复制,复制的高峰期在3~5d,峰值在第4d。提示:树鼠句可以用来建立RSV感染的动物模型。     

Speculation on Pathogenesis in Death from Respiratory Syncytial Virus Infection

The lungs of three infants, two with bronchiolitis and one with pneumonia, were examined by fluorescent antibody techniques for the distribution of respiratory syncytial (R.S.) virus, and also for the presence of human globulin. In bronchiolitis the lungs contained little virus, whereas in pneumonia virus was abundant and widespread; and, paradoxically, while in bronchiolitis human globulin had the same scanty distribution as virus it was absent in pneumonia. It is suggested that the essential process in bronchiolitis is a widespread type 1 allergic reaction dependent on a second encounter with R.S. virus antigen, whereas in R.S. virus pneumonia the mucosal necrosis and alveolar and interstitial inflammation are the result of direct virus damage to the lungs. The alternative explanation put forward is that the process may be a type 3 allergic reaction.     

Matrix Metalloproteinase 9 Exerts Antiviral Activity against Respiratory Syncytial Virus

Increased lung levels of matrix metalloproteinase 9 (MMP9) are frequently observed during respiratory syncytial virus (RSV) infection and elevated MMP9 concentrations are associated with severe disease. However little is known of the functional role of MMP9 during lung infection with RSV. To determine whether MMP9 exerted direct antiviral potential, active MMP9 was incubated with RSV, which showed that MMP9 directly prevented RSV infectivity to airway epithelial cells. Using knockout mice the effect of the loss of Mmp9 expression was examined during RSV infection to demonstrate MMP9’s role in viral clearance and disease progression. Seven days following RSV infection, Mmp9 ^-/- mice displayed substantial weight loss, increased RSV-induced airway hyperresponsiveness (AHR) and reduced clearance of RSV from the lungs compared to wild type mice. Although total bronchoalveolar lavage fluid (BALF) cell counts were similar in both groups, neutrophil recruitment to the lungs during RSV infection was significantly reduced in Mmp9 ^-/- mice. Reduced neutrophil recruitment coincided with diminished RANTES, IL-1β, SCF, G-CSF expression and p38 phosphorylation. Induction of p38 signaling was required for RANTES and G-CSF expression during RSV infection in airway epithelial cells. Therefore, MMP9 in RSV lung infection significantly enhances neutrophil recruitment, cytokine production and viral clearance while reducing AHR.     

Temperature-sensitive Mutants of Respiratory Syncytial Virus

Four conditional-lethal temperature-sensitive mutants of RS virus were detected among the progeny of 454 plaques derived from virus grown in the presence of 10(-4)m 5-fluorouridine. These mutants were stable (reversion frequency, 10(-5.0) or less and failed to produce plaques at 38 or 39 C. Plaquing efficiency was depressed 100-fold or more at 37 C. Variable suppression of growth at the restrictive temperature of 39 C was observed, ranging from 16-fold to complete suppression. The temperature-sensitive defect of three of the mutants appeared to affect functions which were expressed late in the replicative cycle. One of the mutants produced atypical nonsyncytial plaques.     

Roflumilast Inhibits Respiratory Syncytial Virus Infection in Human Differentiated Bronchial Epithelial Cells

Respiratory syncytial virus (RSV) causes acute exacerbations in COPD and asthma. RSV infects bronchial epithelial cells (HBE) that trigger RSV associated lung pathology. This study explores whether the phosphodiesterase 4 (PDE4) inhibitor Roflumilast N-oxide (RNO), alters RSV infection of well-differentiated HBE (WD-HBE) in vitro. WD-HBE were RSV infected in the presence or absence of RNO (0.1-100 nM). Viral infection (staining of F and G proteins, nucleoprotein RNA level), mRNA of ICAM-1, ciliated cell markers (digital high speed videomicroscopy, β-tubulin immunofluorescence, Foxj1 and Dnai2 mRNA), Goblet cells (PAS), mRNA of MUC5AC and CLCA1, mRNA and protein level of IL-13, IL-6, IL-8, TNFα, formation of H₂O₂ and the anti-oxidative armamentarium (mRNA of Nrf2, HO-1, GPx; total antioxidant capacity (TAC) were measured at day 10 or 15 post infection. RNO inhibited RSV infection of WD-HBE, prevented the loss of ciliated cells and markers, reduced the increase of MUC5AC and CLCA1 and inhibited the increase of IL-13, IL-6, IL-8, TNFα and ICAM-1. Additionally RNO reversed the reduction of Nrf2, HO-1 and GPx mRNA levels and consequently restored the TAC and reduced the H₂O₂ formation. RNO inhibits RSV infection of WD-HBE cultures and mitigates the cytopathological changes associated to this virus.