Rapid detection of human coronavirus NL63 by isothermal reverse transcription recombinase polymerase amplification
Background: Human coronaviruses are one of the leading causes for respiratory tract infections and for frequent primary care consultation. The human coronavirus NL63 (HCoV..µNL63) is one representative of the seasonal coronaviruses and capable of infecting the upper and lower respiratory tract and c...
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| Format: | Article |
| Language: | English |
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Elsevier
2022-11-01
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| Series: | Journal of Clinical Virology Plus |
| Subjects: | |
| Online Access: | http://www.sciencedirect.com/science/article/pii/S2667038022000540 |
| _version_ | 1811187624151351296 |
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| author | Aline Dorendorf Iris Bachmann Martin Spiegel Ahmed Abd El Wahed Gregory Dame Frank Hufert |
| author_facet | Aline Dorendorf Iris Bachmann Martin Spiegel Ahmed Abd El Wahed Gregory Dame Frank Hufert |
| author_sort | Aline Dorendorf |
| collection | DOAJ |
| description | Background: Human coronaviruses are one of the leading causes for respiratory tract infections and for frequent primary care consultation. The human coronavirus NL63 (HCoV..µNL63) is one representative of the seasonal coronaviruses and capable of infecting the upper and lower respiratory tract and causative agent for croup in children. Objectives: For fast detection of HCoV-NL63, we developed an isothermal reverse transcription recombinase polymerase amplification (RT-RPA) assay. Study design: The analytical sensitivities of the RT-RPA assay were identified for in vitro transcribed ribonucleic acid (RNA) and for genomic viral RNA from cell culture supernatant. Moreover, specificity was tested with nucleic acids from other human coronaviruses and a variety of clinically relevant respiratory viruses. Finally, a clinical nasopharyngeal swab sample with spiked genomic viral HCoV-NL63 RNA was analyzed. Results: Our HCoV-NL63 RT-RPA assay is highly specific and has an analytical sensitivity of 13 RNA molecules/reaction for in vitro transcribed RNA. For genomic viral RNA from cell culture supernatant spiked into a clinical nasopharyngeal swab sample the assay...s analytical sensitivity is 170 RNA molecules/reaction. The assay shows amplification of the lowest detectable target copy number after 8 minutes and 7 minutes, respectively. Conclusions: We were able to design a sensitive and specific RT-RPA assay for the detection of HCoV-NL63. Additionally, the assay is characterized by short duration, isothermal amplification, and simple instrumentation. |
| first_indexed | 2024-04-11T14:05:28Z |
| format | Article |
| id | doaj.art-07d1f3d9cc674894a255bb6bd2e6aa81 |
| institution | Directory Open Access Journal |
| issn | 2667-0380 |
| language | English |
| last_indexed | 2024-04-11T14:05:28Z |
| publishDate | 2022-11-01 |
| publisher | Elsevier |
| record_format | Article |
| series | Journal of Clinical Virology Plus |
| spelling | doaj.art-07d1f3d9cc674894a255bb6bd2e6aa812022-12-22T04:19:53ZengElsevierJournal of Clinical Virology Plus2667-03802022-11-0124100115Rapid detection of human coronavirus NL63 by isothermal reverse transcription recombinase polymerase amplificationAline Dorendorf0Iris Bachmann1Martin Spiegel2Ahmed Abd El Wahed3Gregory Dame4Frank Hufert5Institute of Microbiology and Virology, Brandenburg Medical School Theodor Fontane, Neuruppin, GermanyInstitute of Microbiology and Virology, Brandenburg Medical School Theodor Fontane, Neuruppin, GermanyInstitute of Microbiology and Virology, Brandenburg Medical School Theodor Fontane, Neuruppin, Germany; Infection Biology Unit, German Primate Center–Leibniz Institute for Primate Research, Göttingen, GermanyInstitute of Animal Hygiene and Veterinary Public Health, University of Leipzig, GermanyInstitute of Microbiology and Virology, Brandenburg Medical School Theodor Fontane, Neuruppin, Germany; Corresponding author.Institute of Microbiology and Virology, Brandenburg Medical School Theodor Fontane, Neuruppin, Germany; Brandenburg University of Technology Cottbus, Senftenberg, Germany; Faculty of Health Sciences, Joint Faculty of Brandenburg University of Technology Cottbus – Senftenberg, Brandenburg Medical School Theodor Fontane, Neuruppin and University of Potsdam, Potsdam, GermanyBackground: Human coronaviruses are one of the leading causes for respiratory tract infections and for frequent primary care consultation. The human coronavirus NL63 (HCoV..µNL63) is one representative of the seasonal coronaviruses and capable of infecting the upper and lower respiratory tract and causative agent for croup in children. Objectives: For fast detection of HCoV-NL63, we developed an isothermal reverse transcription recombinase polymerase amplification (RT-RPA) assay. Study design: The analytical sensitivities of the RT-RPA assay were identified for in vitro transcribed ribonucleic acid (RNA) and for genomic viral RNA from cell culture supernatant. Moreover, specificity was tested with nucleic acids from other human coronaviruses and a variety of clinically relevant respiratory viruses. Finally, a clinical nasopharyngeal swab sample with spiked genomic viral HCoV-NL63 RNA was analyzed. Results: Our HCoV-NL63 RT-RPA assay is highly specific and has an analytical sensitivity of 13 RNA molecules/reaction for in vitro transcribed RNA. For genomic viral RNA from cell culture supernatant spiked into a clinical nasopharyngeal swab sample the assay...s analytical sensitivity is 170 RNA molecules/reaction. The assay shows amplification of the lowest detectable target copy number after 8 minutes and 7 minutes, respectively. Conclusions: We were able to design a sensitive and specific RT-RPA assay for the detection of HCoV-NL63. Additionally, the assay is characterized by short duration, isothermal amplification, and simple instrumentation.http://www.sciencedirect.com/science/article/pii/S2667038022000540human coronavirus NL63recombinase polymerase amplification |
| spellingShingle | Aline Dorendorf Iris Bachmann Martin Spiegel Ahmed Abd El Wahed Gregory Dame Frank Hufert Rapid detection of human coronavirus NL63 by isothermal reverse transcription recombinase polymerase amplification Journal of Clinical Virology Plus human coronavirus NL63 recombinase polymerase amplification |
| title | Rapid detection of human coronavirus NL63 by isothermal reverse transcription recombinase polymerase amplification |
| title_full | Rapid detection of human coronavirus NL63 by isothermal reverse transcription recombinase polymerase amplification |
| title_fullStr | Rapid detection of human coronavirus NL63 by isothermal reverse transcription recombinase polymerase amplification |
| title_full_unstemmed | Rapid detection of human coronavirus NL63 by isothermal reverse transcription recombinase polymerase amplification |
| title_short | Rapid detection of human coronavirus NL63 by isothermal reverse transcription recombinase polymerase amplification |
| title_sort | rapid detection of human coronavirus nl63 by isothermal reverse transcription recombinase polymerase amplification |
| topic | human coronavirus NL63 recombinase polymerase amplification |
| url | http://www.sciencedirect.com/science/article/pii/S2667038022000540 |
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