Organic Substances Interfere with Reverse Transcription-Quantitative PCR-Based Virus Detection in Water Samples

• DOI: 10.1128/aem.03082-14
• Publication Date: 2014-12-20T02:44:09Z
• Authors: Akihiko Hata; Hiroyuki Katayama; Hiroaki Furumai
• Keywords: 0303 health sciences; 03 medical and health sciences; Molecular Diagnostic Techniques; Reverse Transcriptase Polymerase Chain Reaction; Virology; Viruses; Enzyme Inhibitors; Organic Chemicals; Real-Time Polymerase Chain Reaction; Sensitivity and Specificity; 6. Clean water; 3. Good health

Reverse transcription (RT)-PCR-based virus detection from water samples is occasionally hampered by organic substances that are co-concentrated during concentration procedures. To characterize these substances, containing commercially available humic acid, which known to inhibit RT-PCR, and river were subjected adsorption-elution-based using an electronegative membrane. In this study, the before, during, after analyzed in terms of properties efficiencies. Two out three acid solutions resulted RT-quantitative PCR (qPCR) inhibition caused >3-log10-unit underestimation spiked poliovirus. Over 60% organics contained two recovered concentrate, while over uninhibited solution lost process. River concentrates also RT-qPCR. Organic concentrations increased 2.3 3.9 times procedure. The inhibitory fractions 10- 100-kDa size range, suspected be RT-PCR inhibitors. According excitation-emission matrices, acid-like protein-like concentrates, but did not seem affect detection. Our findings reveal detailed analyses effective characterizing substances.