Development and comparison of a rapid RT-qPCR assay with qiastat-Dx respiratory panel for the co-detection of SARS-CoV-2, influenza A virus and respiratory syncytial virus

Buket Baddal; Gokce Akan; Gülten Tuncel; Melis Kalaycı; Aysegul Bostancı; Tamer Sanlidag

Development and comparison of a rapid RT-qPCR assay with qiastat-Dx respiratory panel for the co-detection of SARS-CoV-2, influenza A virus and respiratory syncytial virus

Abstract

Molecular multiplex assays for simultaneous detection of respiratory viral pathogens are central for the efficient identification of multiple pathogens with overlapping clinical presentation. Following the COVID-19 pandemic, influenza viruses, respiratory syncytial virus (RSV) and SARS-CoV-2 began to co-circulate and health authorities issued a warning against a triple epidemic. In this study, a rapid and low-cost multiplex RT-qPCR assay was developed for the detection of influenza A virus, SARS-CoV-2 and RSV A/B. The assay was designed to include two mastermixes (MMs), MM-1 containing primer-probe mix to target SARS-CoV-2 and influenza A virus, and MM-2 containing primer-probe mix for RSV A and B. The assay was validated using 80 clinical oro-nasopharyngeal specimens and the results were compared to Qiastat-Dx respiratory panel. The four-target RT-qPCR assay was able to detect all of the SARS-CoV-2 positive (n=33) specimens and RSV A/B positive specimens (n=22), while the assay was unable to detect 2 of the influenza A positive samples (n=25) in comparison to Qiastat-Dx Analyzer, which had Ct > 35. All of the negative samples were detected correctly (n=60). Overall, PPA (95% CI) in SARS-CoV-2 samples was 100 % (87-100 %), 92 % (72.5-98.6 %) in influenza A samples and 100 % (81.5-100 %) for RSV A/B samples. In conclusion, a rapid, reliable and cost-effective multiplex RT-qPCR assay was developed for the simultaneous detection of four pathogens from nasopharyngeal swabs. This assay has the potential to enhance laboratory diagnostic capabilities and improve public health responses during future epidemics, enabling timely interventions.

Keywords

Ethical Statement

The study protocol was approved by the Institutional Review Board at Near East University (YDU/2023/116-1762). Informed consent has been taken from the individuals who voluntarily participated for the optimization of the in-house assay.

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Details

Primary Language

English

Subjects

Infectious Diseases

Journal Section

Research Article

Authors

Buket Baddal ^*
0000-0003-3319-2179
Cyprus

Gokce Akan
0000-0002-3878-1135
Cyprus

Gülten Tuncel
0000-0002-3332-471X
Cyprus

Melis Kalaycı
0000-0002-5779-5532
Cyprus

Aysegul Bostancı
0000-0003-4319-9032
Cyprus

Tamer Sanlidag
0000-0002-6015-8032
Cyprus

Publication Date

December 31, 2025

Submission Date

January 20, 2025

Acceptance Date

October 21, 2025

Published in Issue

Year 2025 Volume: 42 Number: 4

IZ

https://izlik.org/JA55EH39UD

Cite

RIS / Bibtex

APA

Baddal, B., Akan, G., Tuncel, G., Kalaycı, M., Bostancı, A., & Sanlidag, T. (2025). Development and comparison of a rapid RT-qPCR assay with qiastat-Dx respiratory panel for the co-detection of SARS-CoV-2, influenza A virus and respiratory syncytial virus. Deneysel Ve Klinik Tıp Dergisi, 42(4), 404-410. https://izlik.org/JA55EH39UD

AMA

1.Baddal B, Akan G, Tuncel G, Kalaycı M, Bostancı A, Sanlidag T. Development and comparison of a rapid RT-qPCR assay with qiastat-Dx respiratory panel for the co-detection of SARS-CoV-2, influenza A virus and respiratory syncytial virus. J. Exp. Clin. Med. 2025;42(4):404-410. https://izlik.org/JA55EH39UD

Chicago

Baddal, Buket, Gokce Akan, Gülten Tuncel, Melis Kalaycı, Aysegul Bostancı, and Tamer Sanlidag. 2025. “Development and Comparison of a Rapid RT-QPCR Assay With Qiastat-Dx Respiratory Panel for the Co-Detection of SARS-CoV-2, Influenza A Virus and Respiratory Syncytial Virus”. Deneysel Ve Klinik Tıp Dergisi 42 (4): 404-10. https://izlik.org/JA55EH39UD.

EndNote

Baddal B, Akan G, Tuncel G, Kalaycı M, Bostancı A, Sanlidag T (December 1, 2025) Development and comparison of a rapid RT-qPCR assay with qiastat-Dx respiratory panel for the co-detection of SARS-CoV-2, influenza A virus and respiratory syncytial virus. Deneysel ve Klinik Tıp Dergisi 42 4 404–410.

IEEE

[1]B. Baddal, G. Akan, G. Tuncel, M. Kalaycı, A. Bostancı, and T. Sanlidag, “Development and comparison of a rapid RT-qPCR assay with qiastat-Dx respiratory panel for the co-detection of SARS-CoV-2, influenza A virus and respiratory syncytial virus”, J. Exp. Clin. Med., vol. 42, no. 4, pp. 404–410, Dec. 2025, [Online]. Available: https://izlik.org/JA55EH39UD

ISNAD

Baddal, Buket - Akan, Gokce - Tuncel, Gülten - Kalaycı, Melis - Bostancı, Aysegul - Sanlidag, Tamer. “Development and Comparison of a Rapid RT-QPCR Assay With Qiastat-Dx Respiratory Panel for the Co-Detection of SARS-CoV-2, Influenza A Virus and Respiratory Syncytial Virus”. Deneysel ve Klinik Tıp Dergisi 42/4 (December 1, 2025): 404-410. https://izlik.org/JA55EH39UD.

JAMA

1.Baddal B, Akan G, Tuncel G, Kalaycı M, Bostancı A, Sanlidag T. Development and comparison of a rapid RT-qPCR assay with qiastat-Dx respiratory panel for the co-detection of SARS-CoV-2, influenza A virus and respiratory syncytial virus. J. Exp. Clin. Med. 2025;42:404–410.

MLA

Baddal, Buket, et al. “Development and Comparison of a Rapid RT-QPCR Assay With Qiastat-Dx Respiratory Panel for the Co-Detection of SARS-CoV-2, Influenza A Virus and Respiratory Syncytial Virus”. Deneysel Ve Klinik Tıp Dergisi, vol. 42, no. 4, Dec. 2025, pp. 404-10, https://izlik.org/JA55EH39UD.

Vancouver

1.Buket Baddal, Gokce Akan, Gülten Tuncel, Melis Kalaycı, Aysegul Bostancı, Tamer Sanlidag. Development and comparison of a rapid RT-qPCR assay with qiastat-Dx respiratory panel for the co-detection of SARS-CoV-2, influenza A virus and respiratory syncytial virus. J. Exp. Clin. Med. [Internet]. 2025 Dec. 1;42(4):404-10. Available from: https://izlik.org/JA55EH39UD