J Appl Biomed 22:115-122, 2024 | DOI: 10.32725/jab.2024.013

RT-qPCR investigation of post-mortem tissues during COVID-19

Zhanna Berdygulova^{1, 2, *}, Elina Maltseva^{1, 2, 3, *}, Yuliya Perfilyeva^{1, 2}, Anna Nizkorodova^{1, 2}, Andrey Zhigailov^{1, 2}, Dinara Naizabayeva^{1, 2, 3}, Yekaterina O. Ostapchuk^{1, 2}, Saltanat Kuatbekova¹, Zhaniya Dosmagambet^{1, 4}, Moldir Kuatbek^{1, 4}, Akerke Bissenbay^{1, 2}, Alena Cherusheva¹, Akzhigit Mashzhan¹, Nurshat Abdolla^{1, 2}, Sanzhar Ashimbekov⁴, Gulnara Ismagulova^{1, 2}, Andrey Dmitrovskiy¹, Seidigapbar Mamadaliyev¹, Yuriy Skiba^{1, 2, 3}

¹ Almaty Branch of the National Center for Biotechnology, Central Reference Laboratory, Almaty, Kazakhstan

² M. A. Aitkhozhin Institute of Molecular Biology and Biochemistry, Almaty, Kazakhstan

³ Tethys Scientific Society, Almaty, Kazakhstan

⁴ Asfendiyarov Kazakh National Medical University, Almaty, Kazakhstan

In 2020, there were numerous cases in Kazakhstan with clinical symptoms of COVID-19 but negative PCR results in nasopharyngeal and oropharyngeal swabs. The diagnosis was confirmed clinically and by CT scans (computed tomography). The problem with such negative PCR results for SARS-CoV-2 infection confirmation still exists and indicates the need to confirm the diagnosis in the bronchoalveolar lavage in such cases. There is also a lack of information about confirmation of SARS-CoV-2 infection in deceased patients. In this study, various tissue materials, including lungs, bronchi, and trachea, were examined from eight patients who died, presumably from SARS-CoV-2 infection, between 2020 and 2022. Naso/oropharyngeal swabs taken from these patients in hospitals tested PCR negative for SARS-CoV-2. This study presents a modified RNA isolation method based on a comparison of the most used methods for RNA isolation in laboratories: QIAamp Viral RNA Mini Kit and TRIzol-based method. This modified nucleic acid extraction protocol can be used to confirm SARS-CoV-2 infection by RT-qPCR in the tissues of deceased patients in disputed cases. RT-qPCR with RNA of SARS-CoV-2 re-extracted with such method from post-mortem tissues that were stored at -80 °C for more than 32 months still demonstrated high-yielding positive results.

Keywords: COVID-19; Post-mortem tissues; RNA extraction; RT-qPCR; SARS-CoV-2

Grants and funding:

This work was supported by the Committee of Science of the Ministry of Science and Higher Education of the Republic of Kazakhstan [state grant #AP09259103 “Genomic and subgenomic characterization of SARS-CoV-2 strains, circulating in Kazakhstan, for the development of scientific approaches to COVID-19 diagnostics”].

Conflicts of interest:

The authors have no conflict of interest to declare.

Received: September 7, 2023; Revised: June 6, 2024; Accepted: June 20, 2024; Prepublished online: June 21, 2024; Published: June 24, 2024  Show citation