Prevalence and genotypes of the adenovirus infection as well detection of co-infection with bocavirus in Mexican immunosuppressed and non-immunosuppressed children with pneumonia.

BACKGROUND: Adenovirus (AdV) causes respiratory infection; recent observations suggest that some subtypes have more ability to develop fatal disease. AdV infection has been associated with co-infection with human bocavirus (HBoV). We analysed the frequency of AdV infection, its subtypes and the presence of co-infection with HBoV, as well the clinical characteristics of such co-infection in Mexican paediatric immunosuppressed (IP) and non-immunosuppressed patients (non-IP) diagnosed with pneumonia. METHODS: A total of 5185 nasopharyngeal swabs from two groups of children with pneumonia, one IP and the other non-IP, were analysed for the detection of AdV by immunofluorescence and confirmed by PCR and culture. HBoV was identified by PCR. Positive samples for AdV and AdV/HBoV were typed using PCR sequencing, the clinical characteristics of the AdV/HBoV co-infection were analysed. RESULTS: Thirty-seven of the 5185 (0.71%) samples were positive for AdV, of those 27/37 (73%) were detected in non-IP and 10/37 (27%) in the IP group. Twelve were typed as follows: 9/12 (75%) as Species B1 subtype 3, of those 8/9 (88.9%) in non-IP and 1/9 in the IP group. One of twelve AdV2 subtype B11a was identified in one non-IP and the remaining two out of 12 successfully typed, were identified as Species C subtypes 2 and 6 in the group of non-IP. The presence of both AdV and HBoV1 in co-infection was observed in 2/37 (5.4%) non-IP with a syndrome like influenza. CONCLUSIONS: In this 5 year analysis of samples from non-IP and IP hospitalized paediatric patients with a diagnosis of pneumonia, a low incidence of AdV was found. B1 was the most frequent subtype and frequently found in non-IP, and two cases of co-infection AdV/HBoV1 were detected in two non-IP with a influenza-like syndromes. This is the first report of HBoV and AdV co-infection in Mexico. The frequency of AdV and HBoV co-infection was lower than that reported in other populations.

incorporación de la técnica de RT-PCR para la detección del virus sincitial respiratorio / Incorporation of the RT-PCR technique to the detection of respiratory syncitial virus

Objetivo: Estandarizar e incorporar técnicas de biología molecular, RT-PCR para la detección del virus sincitial respiratorio que complementen y enriquezcan el diagnóstico.Material y métodos: se utilizaron cepas de referencia del virus sincitial respiratorio de los grupos A y B (virus stock ATCC), las cuales se propagaron y titularon en células HEp-2. Se realizaron pruebas de inmunofluorescencia indirecta y de la prueba RT-PCR. Para ello se extrajo el ARN del virus con trizol, se realizó una transcripción reversa para obtener ADNc y, para la PCR se utilizaron oligonucleótidos que amplifican un fragmento del gen de la proteína G del virus sincitial respiratorio. Para comprobar la especificidad de la prueba se utilizaron virus de la misma familia (sarampión y parainfluenza) y virus de diferentes familias (influenza y adenovirus). Al evaluar la sensibilidad se utilizaron diferentes diluciones del ADNc viral.Resultados: en el cultivo, el efecto citopático se puede observar claramente del cuarto al octavo día. La prueba de inmunofluorescencia como se sabe es una técnica sensible y específica para el virus. La RT-PCR que se desarrollo, fue efectiva para la amplificación del ADNc; además, mostró ser específica para el virus sincitial respiratorio, ya que no amplifica material genético de otros virus incluyendo a aquellos que pertenecen a la misma familia. La sensibilidad de la prueba es alta, alcanzando a amplificar hasta picogramos de ADNc.Conclusiones: la rapidez de la inmunofluorescencia, permite dar un diagnóstico presuntivo que después puede ser comprobado por el aislamiento y propagación del virus en cultivo celular. La técnica de RT-PCR con los oligonucleótidos que utilizamos fue sensible, específica y rápida, por tanto debe ser tomada en cuenta para el apoyo al diagnóstico clínico. Con lo anterior, no se trata de sustituir las técnicas tradicionales, sino contar con mayores opciones y además reforzarlas; de esta manera, se podrá fundamentar mejor el diagnóstico de las infecciones virales.