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J Clin Microbiol. 2019 Aug 21. pii: JCM.00735-19. doi: 10.1128/JCM.00735-19. [Epub ahead of print]

Comparison of Three Adenovirus Quantitative PCR Assays with ATCC Reference Strains and Clinical Samples.

Author information

1
Department of Laboratory Medicine, University of Washington Medical Center, Seattle, Washington.
2
Vaccine and Infectious Diseases Division, Fred Hutchinson Cancer Research Center, Seattle, WA.
3
Department of Laboratory Medicine, University of Washington Medical Center, Seattle, Washington Lincook@uw.edu.

Abstract

Adenoviruses (AdV) have been associated with a variety of human diseases and are recognized as causing significant morbidity and mortality in immunocompromised or transplant patients. Quantification of AdV DNA in plasma is notoriously difficult due to the genetic diversity of the 71 different serotypes identified to date. There is no WHO standard available to harmonize quantitative data so results between labs vary widely. In this study we compared an laboratory-developed multiplex PCR assay with primers and probes specific for each group (A-G) and subgroup E4 (Octaplex) to one with a single primer and probe set (Jothikumar, 2005) and one utilizing bisulfite pre-treatment of DNA to reduce variation prior to amplification (Genetic Signatures). Our Octaplex assay detected all low copy-number clinical samples while the other two assays had subsets of samples that did not amplify. The Jothikumar assay failed to efficiently amplify 3 of the high copy number cultured strains, while the Genetic Signatures (GS) 3base™ assay had a positive bias, resulting in higher copies/mL (> 0.5 log10) for all culture fluids tested. All three assays resulted in end-point detection of the available 51 AdV types. Using two different materials to generate a standard curve revealed that the OctaplexTaqMan assay and the Jothikumar assay both consistently gave adenovirus levels lower than the commercial platform for AdV culture fluids, but not patient samples. This study highlights the differences in detection of AdV between laboratories that can be attributed to both the PCR method as well as the reference material used for quantitation.

PMID:
31434723
DOI:
10.1128/JCM.00735-19

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