Evaluation of non-extracted genital swabs for real-time HSV PCR

Nucleic acid extraction of clinical samples is accepted as a key requirement in molecular diagnostics. At Barts Health NHS Trust, swabs taken from patients with clinical suspicion of HSV infection were routinely extracted on the Qiagen MDx BioRobot prior to testing with a real-time triplex PCR for HSV1, HSV2, and VZV. The aim of this study was to adapt an existing HSV1/HSV2/VZV real-time PCR by replacing VZV with phocine herpesvirus 1 (PhHV) as an internal control (IC) and evaluate whether this adapted assay required the nucleic acid extraction step for predominantly genital swabs. First 313 non-extracted and extracted swabs were tested in parallel with the existing triplex HSV1/HSV2/VZV real-time PCR. The second stage involved testing 176 non-extracted swabs using a triplex real-time PCR for HSV1, HSV2, and PhHV and comparing the results with the samples extracted and tested by the original triplex assay. The results correlated well when the existing assay was used, with only three non-extracted samples that would have been reported as negative compared to the extracted sample result (Cq s 33, 39, 35-two samples HSV1, one sample HSV2). In the evaluation using the adapted assay containing the IC, two of 176 samples were discordant, where a HSV negative non-extracted sample result would have been reported differently to the extracted sample result (Cq s 32, 33-both HSV1). This study demonstrated that it is feasible to test non-extracted swabs for HSV in a real-time PCR that includes an IC. J. Med. Virol. 87: 125-129, 2015. © 2014 Wiley Periodicals, Inc.