Amplification of viral RNA from drinking water using TransPlex™ whole‐transcriptome amplification

Parker, J.K.; Chang, T.‐Y.; Meschke, J.S.

Aims: Viral pathogens in environmental media are generally highly diffuse, yet small quantities of pathogens may pose a health risk. This study evaluates the ability of TransPlex™ whole transcriptome amplification (WTA) to amplify small quantities of RNA viruses from complex environmental matrices containing background nucleic acids. Methods and Results: DNA extracts from mock drinking water samples containing mixed microbial populations were spiked with small quantities of echovirus type 13 (EV) RNA. Samples were amplified using a Transplex™ WTA kit, and EV‐specific quantitative reverse transcription polymerase chain reaction (qRT‐PCR) was used to quantify target pathogens before and after application of WTA. Samples amplified by WTA demonstrated a decreased limit of detection. The log‐linear relationship between serial dilutions was maintained following amplification by WTA. Conclusions: WTA is able to increase the quantity of target organism RNA in mixed populations, while maintaining log linearity of amplification across different target concentrations. Significance and Impact of the Study: WTA may serve as an effective preamplification step to increase the levels of RNA prior to detection by other molecular methods such as PCR, microarrays and sequencing.