Rapid and sensitive detection of Laem-Singh virus by reverse transcription loop-mediated isothermal amplification combined with a lateral flow dipstick

Abstract

Laem-Singh virus (LSNV) was discovered recently in Thailand in farmed Giant Tiger shrimp (Penaeus monodon) displaying signs of slow growth syndrome. Loop-mediated isothermal amplification (LAMP) allows DNA to be amplified rapidly at a constant temperature. Here a reverse transcription (RT)-LAMP method was combined with a chromatographic lateral-flow dipstick (LFD) to detect LSNV RNA rapidly and specifically. The reaction was optimized at 65°C for 30min and amplified DNA hybridized to an FITC-labeled oligonucleotide probe for 5min was detected at LFD test line 5min after application. Including 10min for rapid RNA extraction, test results could be generated within 1h and did not require electrophoresis. Compared to an existing RT-PCR method, the RT-LAMP-LFD was also ∼1000-fold more sensitive in detecting LSNV RNA.