Optimization of the elution buffer and concentration method for detecting hepatitis E virus in swine liver using a nested reverse transcription-polymerase chain reaction and real-time reverse transcription-polymerase chain reaction.
J Virol Methods
; 206: 99-104, 2014 Sep.
Article
in En
| MEDLINE
| ID: mdl-24907649
ABSTRACT
The aim of this study was to develop an optimal technique for detecting hepatitis E virus (HEV) in swine livers. Here, three elution buffers and two concentration methods were compared with respect to enhancing recovery of HEV from swine liver samples. Real-time reverse transcription-polymerase chain reaction (RT-PCR) and nested RT-PCR were performed to detect HEV RNA. When phosphate-buffered saline (PBS, pH 7.4) was used to concentrate HEV in swine liver samples using ultrafiltration, real-time RT-PCR detected HEV in 6 of the 26 samples. When threonine buffer was used to concentrate HEV using polyethylene glycol (PEG) precipitation and ultrafiltration, real-time RT-PCR detected HEV in 1 and 3 of the 26 samples, respectively. When glycine buffer was used to concentrate HEV using ultrafiltration and PEG precipitation, real-time RT-PCR detected HEV in 1 and 3 samples of the 26 samples, respectively. When nested RT-PCR was used to detect HEV, all samples tested negative regardless of the type of elution buffer or concentration method used. Therefore, the combination of real-time RT-PCR and ultrafiltration with PBS buffer was the most sensitive and reliable method for detecting HEV in swine livers.
Key words
Full text:
1
Collection:
01-internacional
Database:
MEDLINE
Main subject:
Swine Diseases
/
Polymerase Chain Reaction
/
Hepatitis E virus
/
Hepatitis E
/
Reverse Transcriptase Polymerase Chain Reaction
/
Real-Time Polymerase Chain Reaction
/
Liver
Type of study:
Diagnostic_studies
/
Evaluation_studies
Limits:
Animals
Language:
En
Journal:
J Virol Methods
Year:
2014
Document type:
Article