Comparative analysis of different TaqMan real-time RT-PCR assays for the detection of swine Hepatitis E virus and integration of Feline calicivirus as internal control.
J Appl Microbiol
; 106(4): 1360-9, 2009 Apr.
Article
en En
| MEDLINE
| ID: mdl-19187137
AIMS: The aim of this study was to compare the performance of four TaqMan RT-PCR assays with a commonly used nested RT-PCR and to include the Feline calicivirus (FCV) as an internal control. METHODS AND RESULTS: RNA extracted from 87 swine faecal samples and 103 swine blood samples was subjected to different detection systems. Faecal samples naturally contaminated with Hepatitis E virus (HEV) and negative samples were artificially inoculated with 3.2 x 10(3) PFU of FCV. Detection results obtained on faecal and plasma samples were 35.6% and 4.9% with the nested RT-PCR assay, 8.0% and 0%, 0% and 0%, 13.8% and 0% and 36.8% and 3.9% with TaqMan systems A, B, C and D respectively. The Ct means obtained with the multiplex TaqMan assay were 30.11 and 30.43 for the detection of FCV with HEV contaminated samples and negative samples. CONCLUSIONS: The TaqMan system D was more suitable for the detection of swine HEV strains than the three others and FCV was integrated successfully as an internal control. SIGNIFICANCE AND IMPACT OF THE STUDY: FCV was demonstrated as an efficient control to monitor the RNA extraction process and HEV amplification procedure in a multiplex HEV/FCV TaqMan assay. This control would be helpful in limiting false negative results.
Texto completo:
1
Colección:
01-internacional
Base de datos:
MEDLINE
Asunto principal:
Enfermedades de los Porcinos
/
Virus de la Hepatitis E
/
Reacción en Cadena de la Polimerasa de Transcriptasa Inversa
Tipo de estudio:
Diagnostic_studies
Límite:
Animals
Idioma:
En
Revista:
J Appl Microbiol
Asunto de la revista:
MICROBIOLOGIA
Año:
2009
Tipo del documento:
Article
País de afiliación:
Canadá
Pais de publicación:
Reino Unido