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Advanced target‐specific probe‐based real‐time loop‐mediated isothermal amplification assay for the rapid and specific detection of porcine circovirus 3
Author(s) -
Kim HyeRyung,
Lim DaRae,
Chae HaGyeong,
Park JiYoung,
Kim SeongHee,
Lee KyoungKi,
Lee Changhee,
Lyoo Young S.,
Park ChoiKyu
Publication year - 2020
Publication title -
transboundary and emerging diseases
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.392
H-Index - 63
eISSN - 1865-1682
pISSN - 1865-1674
DOI - 10.1111/tbed.13671
Subject(s) - porcine circovirus , detection limit , loop mediated isothermal amplification , microbiology and biotechnology , virology , real time polymerase chain reaction , polymerase chain reaction , biology , nucleic acid , recombinase polymerase amplification , dna , virus , chemistry , chromatography , genetics , gene
Porcine circovirus type 3 (PCV3) is an emerging viral pathogen that has been identified in pigs with various clinical signs. For rapid and specific detection of PCV3, an advanced real‐time loop‐mediated isothermal amplification (rLAMP) assay that uses both assimilating probes and swarm primers were developed and evaluated in this study. The assay specifically amplified PCV3 DNA, but it did not amplify other porcine viral nucleic acids. The limit of detection of rLAMP with swarm primers was 50 PCV3 DNA copies/reaction, which was comparable to that of the real‐time quantitative polymerase chain reaction (qPCR) and 10 times more sensitive than rLAMP without swarm primers. In an evaluation of clinical samples, the rLAMP assay was able to detect PCV3 DNA within 17.34 ± 4.45 min, which is more rapid than what has been previously reported for the standard qPCR assay (31.78 ± 4.60 min). Detection with rLAMP was largely in agreement with that of the qPCR with a kappa value (95% confidence interval) of 0.98 (0.95–1.00). Taken together, these results suggest that the rLAMP assay presented will be a valuable tool for rapid, specific and reliable diagnosis of PCV3 in clinical samples.