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Minor groove binder modification of widely used TaqMan hydrolysis probe for detection of dengue virus reduces risk of false-negative real-time PCR results for serotype 4

Gray, ER; Heaney, J; Ferns, RB; Sequeira, PC; Nastouli, E; Garson, JA; (2019) Minor groove binder modification of widely used TaqMan hydrolysis probe for detection of dengue virus reduces risk of false-negative real-time PCR results for serotype 4. Journal of Virological Methods , 268 pp. 17-23. 10.1016/j.jviromet.2019.03.006.

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Abstract

Dengue is a vector-transmitted viral infection that is a significant cause of morbidity and mortality in humans worldwide, with over 50 million apparent cases and a fatality rate of 2.5 % of 0.5 million severe cases per annum in recent years. Four serotypes are currently co-circulating. Diagnosis of infection may be by polymerase chain reaction, serology or rapid antigen test for NS1. Both pan-serotype and serotype-specific genome detection assays have been described, however, achieving adequate sensitivity with pan-serotype assays has been challenging. Indeed, as we show here, inspection of components and cycling parameters of a pan-serotype RT-qPCR assay in use in laboratories worldwide revealed insufficient probe stability to accommodate potential nucleotide mismatches, resulting in false-negatives. A minor-groove binder (MGB)-modified version of the probe was designed and its performance compared with that of the original probe in 32 samples. Eight of the samples were undetected by the original probe but detected by the MGB modified probe and six out of seven of these that could be serotyped belonged to serotype 4. Sequencing of the region targeted by the probe in these samples revealed two mismatches which were also universally present in all other serotype 4 sequences in a public database. We therefore recommend adoption of this MGB modification in order to reduce the risk of false-negative results, especially with dengue serotype 4 infections.

Type: Article
Title: Minor groove binder modification of widely used TaqMan hydrolysis probe for detection of dengue virus reduces risk of false-negative real-time PCR results for serotype 4
Location: Netherlands
DOI: 10.1016/j.jviromet.2019.03.006
Publisher version: https://doi.org/10.1016/j.jviromet.2019.03.006
Language: English
Additional information: This version is the author accepted manuscript. For information on re-use, please refer to the publisher’s terms and conditions.
Keywords: Dengue, TaqMan, False-negative, Sensitivity, Serotype 4, Minor groove binder
UCL classification: UCL > Provost and Vice Provost Offices
UCL > Provost and Vice Provost Offices > School of Life and Medical Sciences
UCL > Provost and Vice Provost Offices > School of Life and Medical Sciences > Faculty of Medical Sciences
UCL > Provost and Vice Provost Offices > School of Life and Medical Sciences > Faculty of Medical Sciences > Div of Infection and Immunity
UCL > Provost and Vice Provost Offices > School of Life and Medical Sciences > Faculty of Pop Health Sciences > UCL GOS Institute of Child Health
UCL > Provost and Vice Provost Offices > School of Life and Medical Sciences > Faculty of Pop Health Sciences > UCL GOS Institute of Child Health > ICH Pop, Policy and Practice Prog
UCL > Provost and Vice Provost Offices > UCL BEAMS
UCL > Provost and Vice Provost Offices > UCL BEAMS > Faculty of Maths and Physical Sciences
UCL > Provost and Vice Provost Offices > UCL BEAMS > Faculty of Maths and Physical Sciences > London Centre for Nanotechnology
URI: http://discovery.ucl.ac.uk/id/eprint/10070968
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