Differentiation of PCV1 and PCV2 by a multiplex real-time PCR assay

Porcine circovirus (PCV), belonging to Circoviridae family, is a small, non-enveloped, icosahedral virus containing a circular single-stranded DNA genome (Tischer and others 1982). PCV is divided into two genotypes, PCV type 1 (PCV1) and PCV type 2 (PCV2). PCV1 is a contaminant of PK-15 cells that is non-pathogenic and exists in pigs (Tischer and others 1986, Puvanendiran and others 2011). PCV2 is associated with postweaning multisystemic wasting syndrome (PMWS), which has caused severe losses in the global swine industry in recent decades. Many PK-15 cell lines are contaminated with PCV1, which confers difficulty in using PK-15 cell line to isolate PCV2 for further research. Accordingly, this study aimed to establish a multiplex real-time PCR assay to differentiate and rapidly detect PCV1 and PCV2 in laboratory and clinical samples. PCV2 strain SD2 (GenBank Accession Number DQ478947) was originally isolated from a pig that clinically manifested PMWS (Zhou and others 2009). In the present study, PCV1 was cultured in our laboratory using PK-15 cells. PCV1 and PCV2 viral DNA were extracted by the proteinase K method according to the instructions(Sambrook and others 2001). Primer P15ʹ-GAACCGCGGGCTGGCTGA ACTTTTGAAAGT-3ʹ and …