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

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.