Development and evaluation of rapid detection of classical swine fever virus by reverse transcription loop-mediated isothermal amplification (RT-LAMP).
A one-step reverse transcription loop-mediated isothermal amplification (RT-LAMP) was developed for the detection of classical swine fever virus (CSFV) and a set of primers designed based on the E2 gene reference sequences of the CSFV. The assay was optimized to amplify CSFV RNA by incubation at 63 degrees C for 50 min. The RT-LAMP amplification products had a ladder-like appearance when electrophoresed on an agarose gel. The RT-LAMP assay showed higher sensitivity than the conventional RT-PCR, and no cross-reactivity appeared with other related porcine viruses including porcine circovirus type 2 (PCV2), porcine parvovirus (PPV), porcine pseudorabies virus (PRV), and porcine reproductive and respiratory syndrome virus (PRRSV). The positive predictive value (PPV) of CSFV RT-LAMP for 227 field tissue samples was 94.7%, the negative predictive value (NPV) was 30.8% and showed better for the conventional RT-PCR method. Thus, the RT-LAMP assay is extremely rapid, sensitive, and specific and has potential usefulness for rapid laboratory diagnosis and pen-side detection for CSFV detection in pigs.
Yin
Shuanghui
S
Lanzhou Veterinary Research Institute, Chinese Academy of Agricultural Sciences, State Key Laboratory of Veterinary Etiological Biology, Key Laboratory of Animal Virology of Ministry of Agriculture, Xujiaping No. 1, Lanzhou 730046, Gansu, China.
Shang
Youjun
Y
Zhou
Guangqing
G
Tian
Hong
H
Liu
Yanhong
Y
Cai
Xuepeng
X
Liu
Xiangtao
X
D000818:Animals; D006692:Classical Swine Fever Virus / Q000235:genetics / Q000302:isolation & purification*; D021141:Nucleic Acid Amplification Techniques / Q000379:methods*; D009928:Organ Specificity; D012367:RNA, Viral / Q000235:genetics / Q000302:isolation & purification; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000379:methods*; D012680:Sensitivity and Specificity; D013552:Swine; D014759:Viral Envelope Proteins / Q000235:genetics
D012367:RNA, Viral; D014759:Viral Envelope Proteins; C087086:glycoprotein E2, classical swine fever virus