Improvement of the Method for Inactivation and Extraction of SARS-CoV-2 Genetic Material
Name: JULIANA TEIXEIRA DUTRA FRAGA SPOSITO
Publication date: 25/02/2025
Examining board:
Name |
Role |
|---|---|
| CARLOS GRAEFF TEIXEIRA | Examinador Interno |
| MARIANE VEDOVATTI MONFARDINI SAGRILLO | Examinador Externo |
| SANDRA LUCIA VENTORIN VON ZEIDLER | Examinador Externo |
| VINÍCIUS SANTANA NUNES | Examinador Externo |
Summary: During the pandemic, the need for a rapid and accurate diagnosis became a major challenge due to the high demand for testing, which overwhelmed laboratories and hindered virus containment. SARS-CoV-2, identified in December 2019, quickly spread globally, requiring effective detection methods. Molecular diagnosis, predominantly performed using RT-PCR, is considered the gold standard but relies on the quality of viral RNA extraction, a critical process for test sensitivity. However, this technique faces significant challenges, such as the need for skilled professionals, high costs, and limitations associated with sample collection and storage.
This study aimed to improve the SARS-CoV-2 viral RNA extraction method through thermal inactivation and optimize molecular diagnosis. Nasopharyngeal secretion samples were analyzed under different storage and processing conditions. The results indicated that previously frozen samples required fewer amplification cycles in RT-PCR, suggesting better RNA preservation. Additionally, thermal inactivation at 95°C without commercial extraction kits proved effective, reducing the diagnostic process time. Sample agitation and the use of Proteinase K also influenced amplification efficiency, highlighting the importance of these factors in standardizing laboratory protocols.
The evaluation of storage time impact revealed that samples stored for prolonged periods required a higher number of amplification cycles, which could compromise virus detection in certain situations. The combination of thermal inactivation with Proteinase K proved to be a promising strategy for maintaining RNA integrity and ensuring reliable diagnosis. These findings reinforce the need to optimize laboratory protocols, considering both genetic material stability and operational feasibility in high-demand scenarios.
The implementation of these techniques could benefit laboratories in various settings, particularly those with limited infrastructure for conventional RNA extraction. Furthermore, optimizing sample processing may contribute to faster responses during outbreaks and pandemics, ensuring greater accessibility to molecular diagnosis. Thus, this study provides a valuable contribution to improving SARS-CoV-2 detection strategies, with potential impacts on epidemiological surveillance and infectious disease control
