Original scientific paper

https://doi.org/10.2478/aiht-2025-76-3973

The impact of heat inactivation on RT-qPCR detection of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2): an experience from the University Clinical Centre of Vojvodina, Serbia

Jelena Stojčević Maletić orcid.org/0000-0002-9962-0810 ; University of Novi Sad Faculty of Medicine, Novi Sad, Serbia 2 University Clinical Centre of Vojvodina, Novi Sad, Serbia
Iva Barjaktarović orcid.org/0000-0003-2460-3458 ; University of Novi Sad Faculty of Medicine, Novi Sad, Serbia 2 University Clinical Centre of Vojvodina, Novi Sad, Serbia
Ljiljana Andrijević ; University of Novi Sad Faculty of Medicine, Novi Sad, Serbia
Katarina Bačulov orcid.org/0000-0003-1827-2824 ; University of Novi Sad Faculty of Medicine, Novi Sad, Serbia 2 University Clinical Centre of Vojvodina, Novi Sad, Serbia
Slobodanka Bogdanović Vasić orcid.org/0009-0000-6555-7924 ; Šabac Academy of Vocational Studies, Šabac, Serbia
Diandra Pintać Šarac ; University of Novi Sad Faculty of Medicine, Novi Sad, Serbia

Abstract

Handling clinical samples from patients suspected of SARS-CoV-2 infection puts healthcare workers at risk of exposure to infectious particles. To reduce this risk, samples are often heat-inactivated before nucleic acid isolation, but this procedure may affect the analytical sensitivity of the test. The aim of this study was therefore to evaluate the effects of heat inactivation (56 °C for 30 min) on RT-qPCR results of samples taken from nasopharyngeal and oropharyngeal (NP/OP) swabs collected from 200 symptomatic patients. Each sample was split into two aliquots – one subjected to heat inactivation and the other stored at 4 °C – followed by nucleic acid isolation and RTqPCR analysis using the GeneFinder COVID-19 nucleic acid test. Heat inactivation did not significantly affect the overall SARS-CoV-2 detection rate (55.5 % vs. 55.0 % in untreated and heat-treated groups; χ²=0.01; p=0.91). However, discrepancies occurred in 15.3 % of samples, all with quantification cycle (Cq) values >31, including target loss, gain, or complete signal disappearance after heat treatment. Heat inactivation also slightly decreased Cq values for the RNA-dependent RNA polymerase (RdRp) and envelope (E) genes and increased those for the nucleocapsid (N) gene, with significant changes in strongly positive samples (Cq≤33). In positive samples (Cq≤40), the human ribonuclease (RNase) P gene also exhibited significantly higher Cq values after heat treatment. In the strongly positive subgroup, correlation analysis showed moderate correlation for RdRp and very strong correlation for the N and E genes, and a weaker correlation for weakly positive samples. In conclusion, heat inactivation at 56 °C for 30 min does not significantly affect viral gene detection but may diminish it in samples with low viral load.

Keywords

COVID-19; GeneFinder nucleic acid test; human RNase P gene; nasopharyngeal swabs; oropharyngeal swabs; quantification cycle; real-time quantitative polymerase chain reaction; RNA isolation

Hrčak ID:

342187

URI

https://hrcak.srce.hr/342187

Publication date:

23.12.2025.

Article data in other languages: serbian

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