277. Impact of Delays in Processing of Respiratory Specimens Prior to Real Time RT-PCR Testing on Presence and Amount of Amplifiable RNA
Session: Poster Abstract Session: Diagnostic Microbiology; Novel Molecular Methods
Thursday, October 3, 2013
Room: The Moscone Center: Poster Hall C
Background:   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 if sample integrity is compromised by delays in processing, as may occur frequently due to weekends and holidays.

Methods: Upper respiratory specimens were collected from patients at least 18 years of age hospitalized or seen in an acute care setting with acute respiratory illness (ARI) during 3 influenza seasons 2009-2012 and tested for internal control human RNase P (RNP), influenza A virus (FluA), and influenza B virus (FluB) RNA by rRT-PCR. Processing time was measured in hours from specimen collection to placement in lysis buffer.  Multivariable logistical and linear regression models were conducted to evaluate the association between outcome of test results or amount of amplifiable RNA respectively with processing time adjusting for age, and duration of illness. All analyses were performed using Stata 12.1.

Results: 5582 of 5633 (99%) samples had detectable RNP.   Of these, 635 (11.4%) were influenza positive; 469 (8.4%) FluA, 163 (2.9%) Flu B, and 3 (0.1%) both FluA and B.  Mean processing time was 11.5 hours (min 0.1h, max 105.2h).  There was no significant association between processing time and presence of RNP (OR=1.0, p=0.74), and no relation between processing time and detection of influenza (OR=1.0, p=0.06). Similarly, there was no association between processing time and mean cycle threshold (Ct) values of RNP (p= 0.80), FluA (p=0.65), and FluB (p=0.13) assays.  Older age (OR=0.99; p=<0.001) and longer duration of illness (OR=0.92, p=<0.001) were associated with a lower likelihood of influenza detection.  Older age was also associated with increased FluB Ct values (p=0.001), and longer duration of illness was associated with increased FluA Ct values (p=<0.001), both indicating the presence of less amplifiable RNA.  

Conclusion: Delays in processing time of upper respiratory specimens up to 105 hours was not associated with decreased detection of amplifiable RNA, suggesting specimen integrity is not compromised by such delays.

Ryan Dare, MD, MS1, Yuwei Zhu, MD, MS2, John Williams, MD3, Marie Griffin, MD, MPH2,4,5 and H. Keipp Talbot, MD, MPH2, (1)Internal Medicine, Vanderbilt University Medical Center, Nashville, TN, (2)Vanderbilt University School of Medicine, Nashville, TN, (3)Vanderbilt Univ. Med. Ctr., Nashville, TN, (4)Preventive Medicine, Vanderbilt University School of Medicine, Nashville, TN, (5)Veterans Affairs Tennessee Valley Health Care System, Nashville, TN


R. Dare, None

Y. Zhu, None

J. Williams, Quidel: Scientific Advisor, Consulting fee

M. Griffin, MedImmune: Grant Investigator, Research grant

H. K. Talbot, Sanofi Pasteur: Grant Investigator, Research grant
Wyeth (Pfizer): Grant Investigator, Research grant
MedImmune (AstraZeneca): Grant Investigator, Research grant
Protein Sciences: Grant Investigator, Research grant
Vaxxinate: Grant Investigator, Research grant

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