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Publication Details
AFRICAN RESEARCH NEXUS
SHINING A SPOTLIGHT ON AFRICAN RESEARCH
immunology and microbiology
A novel multiplex real-time RT-PCR assay with FRET hybridization probes for the detection and quantitation of 13 respiratory viruses
Journal of Virological Methods, Volume 165, No. 2, Year 2010
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Description
Quantitative multiplex real-time RT-PCR assays utilizing fluorescence resonance energy transfer (FRET) hybridization probes were developed for the detection of 13 respiratory viruses, including well recognized viral causes (respiratory syncytial virus, influenza viruses A and B, parainfluenza viruses types 1, 2, and 3, adenovirus) as well as viruses described recently as causes of acute respiratory tract infections (human coronaviruses NL63, HKU1, 229E, and OC43, human bocavirus, and human metapneumovirus). FRET probes have an improved toleration for single base mismatches than other probe chemistries, reducing the chances of missing highly variable RNA viruses. The assay could detect 2.5-25 DNA/RNA copies/μl (2.5×103-2.5×104 copies/ml). Validation on 91 known positive respiratory specimens indicated similar specificity as commercial direct immunofluorescence assays (IFA) or single-round PCRs used in initial identification. Screening of 270 IFA negative respiratory specimens identified new viruses in 40/270 (14.8%) cases and additional 79/270 (29.3%) well recognized viruses missed by routine diagnostic assays including 6.7% co-infections. All viruses could be detected in the clinical screening panel. The assays demonstrates an improved sensitivity and scope of detecting respiratory viruses relative to routine antigen detection assays while the quantitative utility may facilitate investigation of the role of co-infections and viral load in respiratory virus pathogenesis. © 2010 Elsevier B.V.
Authors & Co-Authors
Lassaunière, Ria
South Africa, Pretoria
Faculty of Health Sciences
Kresfelder, Tina Louise
South Africa, Pretoria
Faculty of Health Sciences
Venter, Marietjie
South Africa, Pretoria
Faculty of Health Sciences
South Africa, Johannesburg
National Health Laboratory Service
Statistics
Citations: 74
Authors: 3
Affiliations: 2
Identifiers
Doi:
10.1016/j.jviromet.2010.02.005
ISSN:
01660934
Research Areas
Genetics And Genomics
Study Approach
Quantitative