Development and Validation of LAMP Assays for Distinguishing MPXV Clades with Fluorescent and Colorimetric Readouts
Human monkeypox (Mpox) is a zoonotic disease caused by the Monkeypox virus (MPXV). As of 14 August 2024, the World Health Organization (WHO) has declared it a global health emergency. For Mpox, this was the second public health emergency of global significance in the past two years. MPXV belongs to...
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2025-01-01
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| author | Nazente Atceken Sara Asghari Dilmani Ahmed Choukri Abdullah Mutlu Sarıkaya Defne Yigci Gozde Korkmaz Savas Tasoglu |
| author_facet | Nazente Atceken Sara Asghari Dilmani Ahmed Choukri Abdullah Mutlu Sarıkaya Defne Yigci Gozde Korkmaz Savas Tasoglu |
| author_sort | Nazente Atceken |
| collection | DOAJ |
| description | Human monkeypox (Mpox) is a zoonotic disease caused by the Monkeypox virus (MPXV). As of 14 August 2024, the World Health Organization (WHO) has declared it a global health emergency. For Mpox, this was the second public health emergency of global significance in the past two years. MPXV belongs to the <i>Poxviridae</i> family and is phylogenetically and epidemically divided into two clades: the Congo Basin (Clade-I) and the West African (Clade-II) clades. Clade-I has been associated with more severe disease progression and higher mortality compared to Clade-II, and thus the differentiation between clades can play an important role in predicting disease prognosis. The LAMP technique has the advantages of not requiring thermal cycling and achieving higher amplification in a shorter time compared to qPCR. Different types of LAMP assays were developed in this study to benefit from these advantages. We report the development of LAMP-1 and LAMP-2 assays using the LAMP method to detect MPXV Clade-I and Clade-II, respectively. The LAMP-1 assay includes both fluorescence and visible colorimetric readout tests developed with sensitivities of 10<sup>3</sup> and 10<sup>7</sup> copies, respectively. For the LAMP-2 assay, a probe-based test utilizing the Novel R-Duplex DARQ probe was developed, offering fluorescence detection at a sensitivity of 10<sup>3</sup> copies. As a result, we successfully developed three highly specific molecular diagnostic tests that distinctly differentiate between MPXV clades, delivering essential tools for the precise diagnosis and effective control of Mpox. |
| format | Article |
| id | doaj-art-9f646c44948c42349ce7dba62cd02d85 |
| institution | Kabale University |
| issn | 2079-6374 |
| language | English |
| publishDate | 2025-01-01 |
| publisher | MDPI AG |
| record_format | Article |
| series | Biosensors |
| spelling | doaj-art-9f646c44948c42349ce7dba62cd02d852025-01-24T13:25:28ZengMDPI AGBiosensors2079-63742025-01-011512310.3390/bios15010023Development and Validation of LAMP Assays for Distinguishing MPXV Clades with Fluorescent and Colorimetric ReadoutsNazente Atceken0Sara Asghari Dilmani1Ahmed Choukri Abdullah2Mutlu Sarıkaya3Defne Yigci4Gozde Korkmaz5Savas Tasoglu6School of Biomedical Sciences and Engineering, Koç University, 34450 Istanbul, TurkeySchool of Biomedical Sciences and Engineering, Koç University, 34450 Istanbul, TurkeyDepartment of Mechanical Engineering, Koç University, 34450 Istanbul, TurkeyDepartment of Biochemistry Faculty of Pharmacy, Ankara University, 06560 Ankara, TurkeySchool of Medicine, Koç University, 34450 Istanbul, TurkeyKoç University Translational Medicine Research Center (KUTTAM), Koç University, 34450 Istanbul, TurkeySchool of Biomedical Sciences and Engineering, Koç University, 34450 Istanbul, TurkeyHuman monkeypox (Mpox) is a zoonotic disease caused by the Monkeypox virus (MPXV). As of 14 August 2024, the World Health Organization (WHO) has declared it a global health emergency. For Mpox, this was the second public health emergency of global significance in the past two years. MPXV belongs to the <i>Poxviridae</i> family and is phylogenetically and epidemically divided into two clades: the Congo Basin (Clade-I) and the West African (Clade-II) clades. Clade-I has been associated with more severe disease progression and higher mortality compared to Clade-II, and thus the differentiation between clades can play an important role in predicting disease prognosis. The LAMP technique has the advantages of not requiring thermal cycling and achieving higher amplification in a shorter time compared to qPCR. Different types of LAMP assays were developed in this study to benefit from these advantages. We report the development of LAMP-1 and LAMP-2 assays using the LAMP method to detect MPXV Clade-I and Clade-II, respectively. The LAMP-1 assay includes both fluorescence and visible colorimetric readout tests developed with sensitivities of 10<sup>3</sup> and 10<sup>7</sup> copies, respectively. For the LAMP-2 assay, a probe-based test utilizing the Novel R-Duplex DARQ probe was developed, offering fluorescence detection at a sensitivity of 10<sup>3</sup> copies. As a result, we successfully developed three highly specific molecular diagnostic tests that distinctly differentiate between MPXV clades, delivering essential tools for the precise diagnosis and effective control of Mpox.https://www.mdpi.com/2079-6374/15/1/23loop-mediated isothermal amplification technologymonkeypox virus clade detectionnovel reverse-duplex detection of amplification by release of quenching probe |
| spellingShingle | Nazente Atceken Sara Asghari Dilmani Ahmed Choukri Abdullah Mutlu Sarıkaya Defne Yigci Gozde Korkmaz Savas Tasoglu Development and Validation of LAMP Assays for Distinguishing MPXV Clades with Fluorescent and Colorimetric Readouts Biosensors loop-mediated isothermal amplification technology monkeypox virus clade detection novel reverse-duplex detection of amplification by release of quenching probe |
| title | Development and Validation of LAMP Assays for Distinguishing MPXV Clades with Fluorescent and Colorimetric Readouts |
| title_full | Development and Validation of LAMP Assays for Distinguishing MPXV Clades with Fluorescent and Colorimetric Readouts |
| title_fullStr | Development and Validation of LAMP Assays for Distinguishing MPXV Clades with Fluorescent and Colorimetric Readouts |
| title_full_unstemmed | Development and Validation of LAMP Assays for Distinguishing MPXV Clades with Fluorescent and Colorimetric Readouts |
| title_short | Development and Validation of LAMP Assays for Distinguishing MPXV Clades with Fluorescent and Colorimetric Readouts |
| title_sort | development and validation of lamp assays for distinguishing mpxv clades with fluorescent and colorimetric readouts |
| topic | loop-mediated isothermal amplification technology monkeypox virus clade detection novel reverse-duplex detection of amplification by release of quenching probe |
| url | https://www.mdpi.com/2079-6374/15/1/23 |
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