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.
© 2011 The Authors. Journal of Applied Microbiology © 2011 The Society for Applied Microbiology.