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Asthmatics are highly susceptible to respiratory viral infections, possibly due to impaired innate immunity. However, the exact mechanisms of susceptibility are likely to differ amongst viruses. Therefore, we infected primary nasal epithelial cells (NECs) from adults with mild-to-moderate asthma, with respiratory syncytial virus (RSV) or human metapneumovirus (hMPV) in vitro and investigated the antiviral response. NECs from these asthmatics supported elevated hMPV but not RSV infection, compared to non-asthmatic controls. This correlated with reduced apoptosis and reduced activation of caspase-9 and caspase-3/7 in response to hMPV, but not RSV. The expression of heat shock protein 70 (HSP70), a known inhibitor of caspase activation and subsequent apoptosis, was amplified in response to hMPV infection. Chemical inhibition of HSP70 function restored caspase activation and reduced hMPV infection in NECs from asthmatic subjects. There was no impairment in the production of IFN by NECs from asthmatics in response to either hMPV or RSV, demonstrating that increased infection of asthmatic airway cells by hMPV is IFN-independent. This study demonstrates, for the first time, a mechanism for elevated hMPV infection in airway epithelial cells from adult asthmatics and identifies HSP70 as a potential target for antiviral and asthma therapies.

It is well established that glycosaminoglycans (GAGs) function as attachment factors for human metapneumovirus (HMPV), concentrating virions at the cell surface to promote interaction with other receptors for virus entry and infection. There is increasing evidence to suggest that multiple receptors may exhibit the capacity to promote infectious entry of HMPV into host cells; however, definitive identification of specific transmembrane receptors for HMPV attachment and entry is complicated by the widespread expression of cell surface GAGs. pgsA745 Chinese hamster ovary (CHO) cells are deficient in the expression of cell surface GAGs and resistant to HMPV infection. Here, we demonstrate that the expression of the Ca2+-dependent C-type lectin receptor (CLR) DC-SIGN (CD209L) or L-SIGN (CD209L) rendered pgsA745 cells permissive to HMPV infection. Unlike infection of parental CHO cells, HMPV infection of pgsA745 cells expressing DC-SIGN or L-SIGN was dynamin dependent and inhibited by mannan but not by pretreatment with bacterial heparinase. Parental CHO cells expressing DC-SIGN/L-SIGN also showed enhanced susceptibility to dynamin-dependent HMPV infection, confirming that CLRs can promote HMPV infection in the presence or absence of GAGs. Comparison of pgsA745 cells expressing wild-type and endocytosis-defective mutants of DC-SIGN/L-SIGN indicated that the endocytic function of CLRs was not essential but could contribute to HMPV infection of GAG-deficient cells. Together, these studies confirm a role for CLRs as attachment factors and entry receptors for HMPV infection. Moreover, they define an experimental system that can be exploited to identify transmembrane receptors and entry pathways where permissivity to HMPV infection can be rescued following the expression of a single cell surface receptor.

Human metapneumovirus (hMPV) is a significant cause of acute respiratory infection (ARI) in hospitalized children. In Paraguay, ARI is one of the leading causes of childhood hospitalization, with viruses being the primary causal agents. However, a large number of cases of unknown etiology remain. The influenza pandemic of 2009 led to intensified vigilance concerning respiratory infections and more thorough efforts to confirm the presence of viruses such as hMPV. Our objective was to detect hMPV in children hospitalized for ARI in Paraguay during 2009. We studied respiratory samples from 240 children age <5 years hospitalized for IRAs in Paraguay during 2009 who had tested negative for other respiratory viruses. We used the hMPV-reactive LightMix® kit from TIB MOLBIOL for the detection of the nucleoprotein (N) gene by real-time PCR according to manufacturer-specified procedures. Of the 240 samples studied, 29 (12%) were positive for hMPV, with the highest rates detected in July and August (winter) predominating in children over 1 year of age. The most common signs and symptoms were cough, fever, and respiratory distress; while the most common complications were pneumonia and bronchiolitis. These results provide the first evidence concerning the prevalence of hMPV in children hospitalized for ARI in Paraguay, El Metapneumovirus humano (hMPV) constituye una importante causa de Infecciones Respiratorias Agudas (IRAs) en niños hospitalizados. En Paraguay, las IRAs están entre las primeras causas de hospitalización durante la infancia, siendo los virus los principales agentes causales. Sin embargo, aún persiste una alta proporción de casos sin etiología identificada. La pandemia de Influenza en el año 2009, condujo a una intensificación de la vigilancia de las infecciones respiratorias, lo cual permitió al mismo tiempo la búsqueda de otros virus como el hMPV. Nuestro objetivo fue detectar hMPV en niños hospitalizados por IRAs en Paraguay durante el año 2009. Fueron estudiadas 240 muestras respiratorias de niños <5 años internados por IRAs en Paraguay durante el año 2009, que habían resultado negativas para otros virus respiratorios. Fue utilizado el reactivo LightMix Kit human MPV de TIBMOLBIOL, para la detección del gen N de hMPV por PCR en Tiempo Real, siguiendo el procedimiento indicado por el fabricante. De las 240 muestras estudiadas, 29 (12%) resultaron positivas para hMPV, con la mayor detección en julio y agosto; predominando en mayores de 1 año. Los principales signos y síntomas fueron tos, fiebre y dificultad respiratoria; y las complicaciones más frecuentes neumonía y bronquiolitis. Estos resultados proveen las primeras evidencias en Salud Pública, de la importancia del hMPV asociado a niños hospitalizados por IRAs en Paraguay.

Human metapneumovirus (hMPV) is a significant cause of acute respiratory infection (ARI) in hospitalized children. In Paraguay, ARI is one of the leading causes of childhood hospitalization, with viruses being the primary causal agents. However, a large number of cases of unknown etiology remain. The influenza pandemic of 2009 led to intensified vigilance concerning respiratory infections and more thorough efforts to confirm the presence of viruses such as hMPV. Our objective was to detect hMPV in children hospitalized for ARI in Paraguay during 2009. We studied respiratory samples from 240 children age <5 years hospitalized for IRAs in Paraguay during 2009 who had tested negative for other respiratory viruses. We used the hMPV-reactive LightMix® kit from TIB MOLBIOL for the detection of the nucleoprotein (N) gene by real-time PCR according to manufacturer-specified procedures. Of the 240 samples studied, 29 (12%) were positive for hMPV, with the highest rates detected in July and August (winter) predominating in children over 1 year of age. The most common signs and symptoms were cough, fever, and respiratory distress; while the most common complications were pneumonia and bronchiolitis. These results provide the first evidence concerning the prevalence of hMPV in children hospitalized for ARI in Paraguay, El Metapneumovirus humano (hMPV) constituye una importante causa de Infecciones Respiratorias Agudas (IRAs) en niños hospitalizados. En Paraguay, las IRAs están entre las primeras causas de hospitalización durante la infancia, siendo los virus los principales agentes causales. Sin embargo, aún persiste una alta proporción de casos sin etiología identificada. La pandemia de Influenza en el año 2009, condujo a una intensificación de la vigilancia de las infecciones respiratorias, lo cual permitió al mismo tiempo la búsqueda de otros virus como el hMPV. Nuestro objetivo fue detectar hMPV en niños hospitalizados por IRAs en Paraguay durante el año 2009. Fueron estudiadas 240 muestras respiratorias de niños <5 años internados por IRAs en Paraguay durante el año 2009, que habían resultado negativas para otros virus respiratorios. Fue utilizado el reactivo LightMix Kit human MPV de TIBMOLBIOL, para la detección del gen N de hMPV por PCR en Tiempo Real, siguiendo el procedimiento indicado por el fabricante. De las 240 muestras estudiadas, 29 (12%) resultaron positivas para hMPV, con la mayor detección en julio y agosto; predominando en mayores de 1 año. Los principales signos y síntomas fueron tos, fiebre y dificultad respiratoria; y las complicaciones más frecuentes neumonía y bronquiolitis. Estos resultados proveen las primeras evidencias en Salud Pública, de la importancia del hMPV asociado a niños hospitalizados por IRAs en Paraguay.