Research articles
ScienceAsia 50 (2024):ID 2024105 1-7 |doi:
10.2306/scienceasia1513-1874.2024.105
Establishment and application of a multiplex PCR method for
the simultaneous detection of PRV, PCV2, PRRSV, CSFV, and
SIV associated with porcine respiratory disease
Yinming Mao, Deyuan Tang*, Song He, Piao Zhou, Zhiyong Zeng, Bin Wang, Tao Huang, Zhengbo Liao, Shenlin Yuan, Xu Chen
ABSTRACT: Swine respiratory tract disease is a significant issue in pig farming, causing substantial harm and loss. The
main pathogens involved in porcine viral respiratory diseases are porcine pseudorabies virus (PRV), porcine circovirus
type 2 (PCV2), porcine respiratory and reproductive syndrome virus (PRRSV), swine fever virus (CSFV), and swine
influenza virus (SIV). These pathogens often co-infect pigs, leading to increased morbidity and mortality, complicating
disease diagnosis and control. This study focused on designing specific primers for the gene sequences of PRV gB,
PCV2 ORF2, PRRSV ORF6, CSFV C, and SIV M genes to enable rapid diagnosis of multi-pathogen infections. The
optimized single and multiplex polymerase chain reaction (PCR) systems demonstrated high specificity, sensitivity, and
repeatability. A single PCR/RT-PCR method was successfully established for the rapid diagnosis of all tested pathogens,
laying the groundwork for a five-plex PCR method. The established five-plex PCR showed a minimum detection limit
at the picogram level with optimal annealing temperature at 56.5 ?C. Specificity tests confirmed detection of only the
target bands with no amplification of other viruses. Clinical sample analysis demonstrated quick and accurate detection
using this method.
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College of Animal Science, Gui Zhou University, Guiyang 550025 China |
* Corresponding author, E-mail: tdyuan@163.com
Received 3 Dec 2023, Accepted 29 Jul 2024
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