IMPATTO DI RINOVIRUS ED ENTEROVIRUS NELLA POLMONITE DI COMUNITÀ IN ETÀ PEDIATRICA: STUDIO DEI CEPPI VIRALI CIRCOLANTI E CORRELAZIONE CON IL QUADRO CLINICO.

Background: Community-acquired pneumonia (CAP) is one of the most important causes of childhood mortality in developing countries and, despite the availability of effective, preventive and therapeutic measures, remains a significant cause of morbidity and hospitalization in the industrialized world. Little in known about the prevalence of viral infections in children with CAP. Detailed information concerning the etiology of CAP in required in order to formulate treatment reccomendations and implement preventive measures. Evaluating the role played by each potential pathogen may also contribute to improving our understanding of the pathogenesis of the disease. Objectives: To describe the clinical and virological data collected from children with radiographically confirmed CAP in whom samples from respiratory secretions were analyzed for 17 respiratory viruses. Patients and method: a nasopharyngeal swab was taken from children admitted to hospital because of CAP during five consecutive winter and early spring seasons (2007-2012). Samples were tested using the Luminex xTAG Respiratory Virus Panel Fast assay, which detects influenza A virus, influenza B virus, respiratory syncytial virus (RSV) -A and -B, parainfluenzavirus-1, -2, -3, and -4, adenovirus, human metapneumovirus, coronaviruses 229E, NL63, OC43, and HKU1, enterovirus/rhinovirus (RV), and bocavirus. A real-time reverse transcriptase polymerase chain reaction (RT-PCR) was used to identify the RV in the enterovirus/RV positive swabs. Samples which were positive for RV were used for sequencing analysis and to reconstruct the phylogenetic tree. In addition, a total of twelve enterovirus C 104 (EV-C104) strains were analyzed for a complete genome sequence. Nasopharyngeal swabs positive for RSV and RV were analyzed by means of RT-PCR for autolysin-A and wzg genes of Streptooccus pneumoniae. Results: A total of 435 children (73,5%) were positive for at least one virus: the most frequently detected was RSV, which was found in 188 (31,7%), followed by RV (n = 144, 24,3%), bocavirus (n = 60, 10,1%), influenza viruses (n = 57, 9,6%), and human metapneumovirus (n = 49, 8,2%). Viral co-infections were found in 117 children (19,7% of the enrolled children; 26,9% of those with viral infections). Marginal differences were found between the infections owing to a single virus. Co-infections showed radiographic evidence of alveolar pneumonia significantly more frequently than single infections. RV was identified in 198 samples (42.2%), and the VP4/VP2 region was successfully amplified in 151 (76.3%). RV-A was identified in 78 samples (51.6%), RV-B in 14 (9.3%) and HRV-C in 59 (39.1%). Forty-seven (31.1%) of the children with RV infection were aged <1 year, 71 (47.0%) were aged 1-3 years, and 33 (21.9%) were aged ≥4 years. Blast and phylogenetic analyses showed that the RV strains were closely related to a total of 66 reference genotypes, corresponding to 29 RV-A, 9 RV-B and 28 RV-C strains. The genome length of the seven strains was 7406 nucleotides (nt). The seven genomes showed 91.0–96.9% nucleotide identity with respect to other available EV-C104 complete genomes. The P1 and P2 regions of the Italian strains were closely related to EV-C104 identified in Switzerland, while the P3 region was closely related to the EV-C117 strain. Sixty-five ofthe 126 RSV-positive children (51,6%) were colonized with S. pneumoniae. Mean bacterial load was significantly higher in patients with alveolar involvement . Serotypes 5 and 19A were almost exclusively indentified in the children with RSV and alveolar CAP, although the difference was significant only for serotype 19A. Serotypes1, 5 and 19A were indentified in children with RV and alveolar CAP, although the difference was significant only for serotype 5. Conclusions: The findings of the study highlights the importance of respiratory viruses in children with CAP and show the characteristics of both single infections and co-infections associated with the disease. Moreover, this study shows that, RV-A, but also RV-B, and, more rarely, RV-C, can be detected in children with CAP. A large number of genotypes may be involved in causing pediatric CAP and can vary year to year. The same genotype can circulate for a prolonged period of time, being associated with a number of CAP episodes in different years. Regarding the genome characterization of EV-C104 strains, it was possible to establish the phylogenetic tree and to understand the relationship between the isolated strains and other known enterovirus C strains. The determination of nasopharyngeal pneumococcal bacterial load and identification of serotypes in children positive for RSV or RV infection, contribute to the diagnosis of mixed lung infection.