Evaluation of droplet digital qRT-PCR (dd qRT-PCR) for quantification of SARS CoV-2 RNA in stool and urine specimens of COVID-19 patients

Abstract

IntroductionThere have been a few reports of viral load detection in stool and urine samples of patients with coronavirus disease 2019 (COVID-19), and the transmission of the virus through faecal oral route. For clinical diagnosis and treatment, the widely used reverse transcription-polymerase chain reaction (qRT-PCR) method has some limitations.MethodsThe aim of our study to assess the presence and concentration of SARS CoV-2 RNA in stool and urine samples from COVID-19 patients with mild, moderate, and severe disease, we compared a traditional qRT-PCR approach with a ddPCR. ddPCR and qRT-PCR-based target gene analysis were performed on 107 COVID-19-confirmed patients paired samples (N1 and N2). The MagMax magnetic beads base method was used to isolate RNA. Real-time qRT-PCR and dd PCR were performed on all patients.Results and DiscussionThe average cycle threshold (Ct) of qRT-PCR was highly correlated with the average copy number of 327.10 copies/l analyzed in ddPCR. In ddPCR, urine samples showed 27.1% positivity while for stool it was 100%.ConclusionThis study’s findings not only show that SARS CoV-2 is present in urine and faeces, but also suggest that low concentrations of the viral target ddPCR make it easier to identify positive samples and help resolve for cases of inconclusive diagnosis.