Development and evaluation of single-tube nested PCR (STNPCR) for the detection of porcine circovirus type 2 (PCV2).
Porcine circovirus 2 (PCV2) is a common virus in pig population and is associated with the postweaning multisystemic wasting disease (PMWS). In this study, it was developed and evaluated the single-tube nested PCR (STNPCR) method for the detection of PCV2 DNA. PCV2 reference controls and swine tissue samples were used, and primers were selected for targeting specific regions of the viral genome. In comparison of the methods, STNPCR was 10 times more sensitive than conventional PCR and showed the same sensitivity to nested PCR (NPCR), but with reduction in the risk of cross-contamination. In clinical application, 55 tissue samples were analysed by conventional PCR and resulted in 67% (37/55) of positive reactions, while the NPCR and STNPCR were able to identify the presence of viral DNA in 100% (55/55) of the samples. The high sensitivity combined with the elimination of cross-contamination makes the STNPCR method suitable for the epidemiological studies of PCV2 and can aid in the diagnosis of PMWS.
Laboratory of Molecular Studies and Experimental Therapy, Department of Genetics, Center for Biological Sciences, Federal University of Pernambuco, Pernambuco, Brazil.
N E
NE
Pontes
C N
CN
Barbosa
A L S
AL
Jesus
J G
JG
Silva
A C
AC
Freitas
D000818:Animals; D001483:Base Sequence; D017925:Circovirus / Q000235:genetics / Q000302:isolation & purification*; D019295:Computational Biology; D017931:DNA Primers / Q000235:genetics; D004279:DNA, Viral / Q000235:genetics; D005260:Female; D016679:Genome, Viral / Q000235:genetics; D008969:Molecular Sequence Data; D016133:Polymerase Chain Reaction / Q000379:methods / Q000662:veterinary*; D053570:Porcine Postweaning Multisystemic Wasting Syndrome / Q000175:diagnosis* / Q000821:virology*; D012680:Sensitivity and Specificity; D017422:Sequence Analysis, DNA / Q000662:veterinary; D013552:Swine
D017931:DNA Primers; D004279:DNA, Viral