Improved recovery of SARS-CoV-2 from wastewater through application of RNA and DNA stabilising agents.
Stephen H BellDanielle M AllenMarina I ReyneJonathan F W LockArthur FitzgeraldAshley LevickasAndrew J LeeConnor G G BamfordDeirdre F GilpinJohn W McGrathPublished in: Letters in applied microbiology (2023)
Wastewater Based Epidemiology (WBE) has become an integral part of the public health effort to track the levels of SARS-CoV-2 within communities. Detection of SARS-CoV-2 in wastewater can be challenging due to relatively low levels of virus within the sample. The wastewater matrix is also comprised of commercial and domestically derived contaminants, as well as RNases, all of which can adversely affect RT-qPCR analysis. To improve SARS-CoV-2 detection within wastewater samples we investigated both the effect of template dilution (as a means to reduce RT-qPCR inhibition) and sample stabilisation via addition of DNA/RNA Shield™ and/or RNA Later™ (to prevent RNA degradation via RNases) as a means to improve viral fragment detection. Using both methodologies a significant improvement in SARS-CoV-2 detection from wastewater samples was observed. No adverse effects of stabilising agent addition on downstream Next Generation Sequencing workflows were detected.
Keyphrases
- sars cov
- wastewater treatment
- respiratory syndrome coronavirus
- public health
- anaerobic digestion
- loop mediated isothermal amplification
- circulating tumor
- label free
- nucleic acid
- real time pcr
- single molecule
- cell free
- gene expression
- ms ms
- liquid chromatography
- high resolution
- mass spectrometry
- simultaneous determination