Detection of influenza by real time RT-PCR is not affected by delays in respiratory specimen processing.

Abstract

Sample processing for real-time reverse transcriptase polymerase chain reaction (rRT-PCR) based diagnostic assays requires stabilizing sample ribonucleic acid (RNA) in lysis buffer prior to testing. The stability of viral RNA prior to processing is difficult to assure. It is unknown whether clinical sample integrity is compromised by delays in processing, as may occur due to weekends and holidays. We sought to examine the integrity of respiratory specimens with variable processing times. Upper respiratory specimens were collected during three influenza seasons 2009-2012 and tested for influenza virus and internal control human RNase P (RNP) RNA by rRT-PCR. Time to processing was measured in hours from specimen collection to placement in lysis buffer. Six hundred thirty-five (11.4%) of 5,583 samples were influenza positive. Mean and median times to processing were 11.5 hr and 6.0 hr, respectively (min 0.1 hr, max 105.2 hr). There were no significant associations between time to processing and presence of RNP (OR = 1.0, P = 0.740), or detection of influenza (OR = 1.0, P = 0.060). Longer duration of illness was associated with a lower likelihood of influenza detection (OR = 0.92, P < 0.001) and with increased influenza A cycle threshold (Ct) values (P < 0.001), while older age was associated with increased influenza B Ct values (P = 0.001), indicating the presence of less amplifiable RNA. Delays in time to processing of upper respiratory specimens up to 105 hr were not associated with decreased detection of amplifiable RNA, suggesting specimen integrity is not compromised by such delays. J. Med. Virol. 88:1891-1895, 2016. © 2016 Wiley Periodicals, Inc.