Abstract
BackgroundPorcine Deltacoronavirus (PDCoV) is a newly emerged enteropathogenic coronavirus that causes diarrhea and mortality in neonatal piglets. PDCoV has spread to many countries around the world, leading to significant economic losses in the pork industry. Therefore, a rapid and sensitive method for detection of PDCoV in clinical samples is urgently needed.ResultsIn this study, we developed a single-tube one-step reverse transcription loop-mediated isothermal amplification (RT-LAMP) assay specific for nucleocapsid gene to diagnose and monitor PDCoV infections. The detection limit of RT-LAMP assay was 1 × 101 copies of PDCoV, which was approximately 100-fold more sensitive than gel-based one-step reverse transcription polymerase chain reaction (RT-PCR). This assay could specifically amplify PDCoV and had no cross amplification with porcine epidemic diarrhea virus (PEDV), transmissible gastroenteritis virus (TGEV), porcine kobuvirus (PKoV), porcine astrovirus (PAstV), porcine reproductive and respiratory syndrome virus (PRRSV), classic swine fever virus (CSFV), and porcine circovirus type 2 (PCV2). By screening a panel of clinical specimens (N = 192), this method presented a similar sensitivity with nested RT-PCR and was 1–2 log more sensitive than conventional RT-PCR in detection of PDCoV.ConclusionsThe RT-LAMP assay established in this study is a potentially valuable tool, especially in low-resource laboratories and filed settings, for a rapid diagnosis, surveillance, and molecular epidemiology investigation of PDCoV infections. To the best of our knowledge, this is the first work for detection of newly emerged PDCoV with LAMP technology.
Highlights
Porcine Deltacoronavirus (PDCoV) is a newly emerged enteropathogenic coronavirus that causes diarrhea and mortality in neonatal piglets
We developed and evaluated a specific and sensitive Reverse transcription loopmediated isothermal amplification (RT-LAMP) assay for visual detection of PDCoV, which might be a good tool for the diagnosis of PDCoV in field samples
Optimization of RT‐LAMP assay for detection of PDCoV The RT-LAMP assay for detection of PDCoV was successfully developed
Summary
Porcine Deltacoronavirus (PDCoV) is a newly emerged enteropathogenic coronavirus that causes diarrhea and mortality in neonatal piglets. Available methods for detection of PDCoVs include conventional reverse transcription—polymerase chain reaction (RT-PCR), nested RT-PCR, real-time RTPCR, and ELISA [6, 10,11,12,13] These techniques have some shortcomings, such as higher requirements for equipment, high cost, extended detection period, and/or low sensitivity [14]. Reverse transcription loopmediated isothermal amplification (RT-LAMP) provides a potential effective tool for rapid and accurate identification of viral pathogens, which amplifies nucleic acids under isothermal conditions with high sensitivity and specificity. This novel gene detection technique is costeffective and time-saving, and only requires a constant temperature water bath. We developed and evaluated a specific and sensitive RT-LAMP assay for visual detection of PDCoV, which might be a good tool for the diagnosis of PDCoV in field samples
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