Detección y cuantificación de virus dengue 2 en lisado celular y plasma de niños por qPCR en tiempo real usando un estuche comercial y el equipo EcoTM System-Illumina

Detección y cuantificación de virus dengue 2 en lisado celular y plasma de niños por qPCR en tiempo real usando un estuche comercial y el equipo EcoTM System-Illumina

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Federico Perdomo Celis Universidad Surcolombiana, Neiva, Colombia
Piedad Perilla Universidad Surcolombiana, Neiva, Colombia
Doris M. Salgado Universidad Surcolombiana
Carlos F. Narváez Hospital Universitario de Neiva, Colombia
Abstract

Methods  for  Dengue  virus  (DENV)  diagnosis  in  endemic areas  are  greatly  needed.  One of  them  is  the  real-time polymerase  chain  reaction  (qPCR)  that  also  enables  to quantitate  the  viral  genome.  Kits of  qPCR  for  DENV  are expensive and restrict their use to a small number of qPCR devices, which limits the application of the technique. Here, we evaluated the performance of a commercial kit of qPCR to DENV-2 detection  on  a  locally  available  qPCR  device (EcoTM System, Illumina), not cited by the kit manufacturer.VERO-76 cells lysate and plasma from children, both with confirmed ongoing DENV-2 infection, were evaluated. As specificity control, cell lysates and plasma  from  children infected  with  DENV-1,  and  uninfected  lysate,  were  also included. The reactions were simultaneously evaluated in an Applied Bio systems  7300  device.  The standard  curve generated  by  EcoTM  was  robust  (R2=  0.99)  with  low variability in the replicates (<10%). The reaction efficiency was high (88.8%) and signal was only obtained in lysates and plasma infected  with  DENV-2.  There was  a  strong positive  correlation  (R2=  1.0,  P=  0.0028)  between  the number of copies of viral RNA in the samples detected by both  qPCR  devices.  Thus, the use  of  the  evaluate  kit  for detection of DENV-2 here tested can be extended to EcoTM. With  this  work,  technological  capacity  for  DENV  study  in an endemic zone is greatly strengthened.

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Author Biographies / See

Federico Perdomo Celis, Universidad Surcolombiana, Neiva, Colombia

Semillero de Formación SINEDIR, Grupo de Parasitología y Medicina Tropical, Programa de Medicina,

Piedad Perilla, Universidad Surcolombiana, Neiva, Colombia

Semillero de Formación SINEDIR, Grupo de Parasitología y Medicina Tropical, Programa de Medicina,

Doris M. Salgado, Universidad Surcolombiana

Semillero de Formación SINEDIR, Grupo de Parasitología y Medicina Tropical, Programa de Medicina,

Carlos F. Narváez, Hospital Universitario de Neiva, Colombia

Servicio de Pediatría,
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