Clinical evaluation of multiplex RT-PCR assays for the detection of influenza A/B and respiratory syncytial virus using a high throughput system

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J Virol Methods. 2019 Apr 1. pii: S0166-0934(18)30631-1. doi: 10.1016/j.jviromet.2019.03.015. [Epub ahead of print]
Clinical evaluation of multiplex RT-PCR assays for the detection of influenza A/B and respiratory syncytial virus using a high throughput system.

Eigner U¹, Reucher S², Hefner N³, Staffa-Peichl S³, Kolb M³, Betz U³, Holfelder M³, Spier G⁴, Pfefferle S², L?tgehetmann M².
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Abstract

BACKGROUND:

Lower respiratory tract infections are a major threat to public health systems worldwide, with RSV and influenza being the main agents causing hospitalization. In outbreak situations, high-volume respiratory testing is needed. In this study, we evaluated the analytical and clinical performance of a pre-designed primer/probe set for the simultaneous multiplex detection of both viruses on a high-throughput platform, the cobas 6800, using the "open channel" of the system for integration of lab-developed assays for the detection of influenza and RSV.
RESULTS:

Using the influenza/RSV qPCR Assay with swabs, LoD (95%) in TCID50/mL for influenza-A was 0.009, influenza-B 0.003, RSV-A 0.202, and RSV-B 0.009. Inter-run variability (3xLoD) was low (<1 Ct for all targets). Of 371 clinical respiratory specimens analyzed, results were concordant for 358 samples. The calculated sensitivity and specificity of the assay were 98.3% and 98.4% for Flu-A, 100% and 98.5% for Flu-B, and 98.6% and 99.7% for RSV. All quality assessment panel specimens (N = 63, including avian influenza strains) were correctly identified. None of the tested microorganisms showed cross-reactivity.
CONCLUSION:

Compared with CE-IVD assays, the assay evaluated here showed good analytical and clinical sensitivity and specificity with broad coverage of different virus strains. It offers high-throughput capacity with low hands-on time, facilitating the laboratory management of large respiratory outbreaks.
Copyright ? 2019. Published by Elsevier B.V.


KEYWORDS:

Influenza, Human; Multiplex Real-time PCR; Respiratory Syncytial Viruses

PMID: 30946852 DOI: 10.1016/j.jviromet.2019.03.015