Analysis of real time PCR amplification efficiencies from three genomic region of dengue virus.
Invest Clin
; 54(1): 5-19, 2013 Mar.
Article
em En
| MEDLINE
| ID: mdl-23781709
Early diagnosis of dengue virus (DENV) infection represents a key factor in preventing clinical complications attributed to the disease. The aim of this study was to evaluate the amplification efficiencies of an in-house quantitative real time-PCR (qPCR) assay of DENV, using the non-structural conserved genomic region protein-5 (NS5) versus two genomic regions usually employed for virus detection, the capsid/pre-membrane region (C-prM) and the 3'-noncoding region (3'NC). One-hundred sixty seven acute phase serum samples from febrile patients were used for validation purposes. Results showed that the three genomic regions had similar amplification profiles and correlation coefficients (0.987-0.999). When isolated viruses were used, the NS5 region had the highest qPCR efficiencies for the four serotypes (98-100%). Amplification from acute serum samples showed that 41.1% (67/167) were positive for the universal assay by at least two of the selected genomic regions. The agreement rates between NS5/C-prM and NS5/3'NC regions were 56.7% and 97%, respectively. Amplification concordance values between C-prM/NS5 and NS5/3'NC regions showed a weak (kappa = 0.109; CI 95%) and a moderate (kappa = 0.489; CI 95%) efficiencies in amplification, respectively. Serotyping assay using a singleplex NS5-TaqMan format was much more sensitive than the C-prM/SYBR Green I protocol (76%). External evaluation showed a high sensitivity (100%), specificity (78%) and high agreement between the assays. According to the results, the NS5 genomic region provides the best genomic region for optimal detection and typification of DENV in clinical samples.
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Coleções:
01-internacional
Base de dados:
MEDLINE
Assunto principal:
RNA Viral
/
Genoma Viral
/
Proteínas não Estruturais Virais
/
Regiões 3' não Traduzidas
/
Dengue
/
Vírus da Dengue
/
Proteínas do Capsídeo
/
Reação em Cadeia da Polimerase em Tempo Real
Tipo de estudo:
Diagnostic_studies
/
Evaluation_studies
/
Guideline
/
Screening_studies
Limite:
Humans
Idioma:
En
Revista:
Invest Clin
Ano de publicação:
2013
Tipo de documento:
Article
País de afiliação:
Venezuela
País de publicação:
Venezuela