Development of a loop-mediated isothermal amplification (LAMP) assay for rapid visual detection of porcine circovirus type 2 (PCV2) and its application.

ABSTRACT

A cost effective, simple and rapid method is critical for detection of porcine circovirus type 2 (PCV2) infection in pigs. The present study reports the development and evaluation a loop-mediated isothermal amplification (LAMP) assay for rapid visual detection of PCV2 in pigs. The time and temperature conditions for amplification of PCV2 genes were optimized to be 30 min at 67 °C. The developed assay was 10 fold more sensitive than conventional PCR with analytical sensitivity of 5 pg and 50 pg, respectively. The developed LAMP assay had a sensitivity of 100%, specificity of 85.45% and overall accuracy of 89.70%. This is perhaps the most rapid of all LAMP reports for PCV2 detection available globally. The assay did not cross-react with porcine parvovirus or classical swine fever virus. DNA sequencing was done to ensure accuracy of LAMP assay results. The assay was assembled into a kit of 20 reactions and validated in different laboratories in India. The developed LAMP assay was proved to be a specific, sensitive and rapid method for visual detection of PCV2 which does not require costly equipments.