Development and validation of a multiplex conventional PCR assay for simultaneous detection and grouping of porcine bocaviruses

Xiaowen Zheng, Gaopeng Liu, Tanja Opriessnig, Zining Wang, Zongqi Yang, Yonghou Jiang

Research output: Contribution to journalArticlepeer-review

Abstract

Porcine bocavirus (PBoV), a newly described porcine parvovirus, has received attention because it can be commonly identified in clinically affected pigs including pigs with post-weaning multisystemic wasting syndrome (PWMS) and pigs with diarrhea. In recent years, novel PBoVs have been identified and were classified into three genogroups, but the ability to detect and classify these novel PBoVs is not comprehensive to date. In this study, a multiplex conventional PCR assay for simultaneous detection and grouping of PBoVs was developed by screening combinations of mixed primer pairs followed by optimization of the PCR conditions. This method exclusively amplifies targeted fragments of 531 bp from the VP1 gene of PBoV G1, 291 bp from the NP1 gene of PBoV G2, and 384 bp from the NP1/VP1 gene of PBoV G3. The assay has a detection limit of 1.0 × 103 copies/μL for PBoV G1 4.5 × 103 for PBoV G2 and 3.8 × 103 for PBoV G3 based on testing mixed purified plasmid constructs containing the specific viral target fragments. The performance of the multiplex PCR assay was comparable to that of the single PCRs which used the same primer pairs. Using the newly established multiplex PCR assay, 227 field samples including faeces, serum and tissue samples from pigs were investigated. All three PBoV genogroups were detected in the clinical samples with a detection rate of 1.3%, 2.6% and 12.3%, respectively for PBoV G1, G2 and G3. Additionally, coinfections with two or more PBoV were detected in 1.7% of the samples investigated. These results indicate the multiplex PCR assay is specific, sensitive and rapid, and can be used for the detection and differentiation of single and multiple infections of the three PBoV genogroups in pigs.
Original languageEnglish
Pages (from-to)164-169
JournalJournal of Virological Methods
Volume236
Early online date19 Jul 2016
DOIs
Publication statusPublished - Oct 2016

Keywords

  • multiplex PCR
  • detection
  • grouping
  • porcine bocaviruses
  • prevalence rate

Fingerprint Dive into the research topics of 'Development and validation of a multiplex conventional PCR assay for simultaneous detection and grouping of porcine bocaviruses'. Together they form a unique fingerprint.

Cite this