Development of a non-infectious control for viral hemorrhagic fever PCR assays.

Autor: Knox MA; Massey University, School of Veterinary Science, Palmerston North, Manawatu-Wanganui, New Zealand., Bromhead C; Massey University, School of Health Sciences, Wellington, New Zealand., Hayman DT; Massey University, School of Veterinary Science, Palmerston North, Manawatu-Wanganui, New Zealand.
Jazyk: angličtina
Zdroj: PLoS neglected tropical diseases [PLoS Negl Trop Dis] 2024 Apr 22; Vol. 18 (4), pp. e0011390. Date of Electronic Publication: 2024 Apr 22 (Print Publication: 2024).
DOI: 10.1371/journal.pntd.0011390
Abstrakt: Assay validation is an essential component of disease surveillance testing, but can be problematic in settings where access to positive control material is limited and a safety risk for handlers. Here we describe a single non-infectious synthetic control that can help develop and validate the PCR based detection of the viral causes of Crimean-Congo hemorrhagic fever, Ebola virus disease, Lassa fever, Marburg virus disease and Rift Valley fever. We designed non-infectious synthetic DNA oligonucleotide sequences incorporating primer binding sites suitable for five assays, and a T7 promotor site which was used to transcribe the sequence. Transcribed RNA was used as template in a dilution series, extracted and amplified with RT-PCR and RT-qPCR to demonstrate successful recovery and determine limits of detection in a range of laboratory settings. Our results show this approach is adaptable to any diagnostic assay requiring validation of nucleic acid extraction and/or amplification, particularly where sourcing reliable, safe material for positive controls is infeasible.
Competing Interests: The authors have declared that no competing interests exist.
(Copyright: © 2024 Knox et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.)
Databáze: MEDLINE
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