Development of a non-infectious control for viral hemorrhagic fever PCR assays.
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Date
2024-04-22
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PLOS
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(c) 2024 The Author/s
CC BY 4.0
CC BY 4.0
Abstract
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.
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Knox MA, Bromhead C, Hayman DT. (2024). Development of a non-infectious control for viral hemorrhagic fever PCR assays.. PLoS Negl Trop Dis. 18. 4. (pp. e0011390-).