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Reviews of "Laboratory Validation of a Simplified DNA Extraction Protocol Followed by a Portable qPCR Detection of M. Tuberculosis DNA Suitable for Point of Care Settings"

Reviewers: W Sabiiti (University of St Andrews) | 📒📒📒 ◻️◻️ • H C Schindler (Instituto Aggeu Magalhães) | 📘📘📘📘📘

Published onMay 08, 2024
Reviews of "Laboratory Validation of a Simplified DNA Extraction Protocol Followed by a Portable qPCR Detection of M. Tuberculosis DNA Suitable for Point of Care Settings"
key-enterThis Pub is a Review of
Laboratory validation of a simplified DNA extraction protocol followed by a portable qPCR detection of M. tuberculosis DNA suitable for point of care settings
Laboratory validation of a simplified DNA extraction protocol followed by a portable qPCR detection of M. tuberculosis DNA suitable for point of care settings

Tuberculosis, caused by Mycobacterium tuberculosis, is a treatable and curable disease, and yet remains one of the leading causes of death worldwide. Diagnosis is essential to reducing the number of cases and starting treatment, but costly tests and equipments that require complex infrastructure hamper their widespread use as a tool to contain the disease in vulnerable populations as well countries lacking resources. Therefore, it becomes necessary to develop new technological approaches to molecular methods as well as screening tests that can be rapidly conducted among people presenting to a health facility to differentiate those who should have further diagnostic evaluation for TB from those who should undergo further investigation for non-TB diagnoses. The present study aimed to evaluate two experimental DNA extraction methods from clinical samples (FTA card versus sonication) followed by analysis in a portable qPCR instrument (the Q3-plus). The FTA card-based protocol showed 100% sensitivity and specificity, while the sonication protocol showed 80% sensitivity and 89% specificity when compared to the traditional gold standard culture. The portable protocol, comprised by the FTA card method and the portable instrument Q3-Plus, showed sensitivity and specificity of 92% and 61%, respectively, when compared to culture, and 75% and 81%, respectively, when compared to the standard TB case classification. The ROC curve showed an AUC of 0.78 (p<0.001) for the portable protocol and 0.93 (p<0.001) for the GeneXpert MTB/RIF. The limit of detection (LOD) for Mycobacterium tuberculosis (H37Rv strain) detection in spiked samples obtained using the portable protocol (FTA card and Q3-Plus) was 19.3 CFU/mL. As an added benefit, using the FTA card facilitates sample handling, transport, and storage. It is concluded that the use of the FTA card protocol and the Q3-Plus yields similar sensitivity and specificity as the gold standard diagnostic tests and case classification. We suggest that the platform is suitable to use as a point of care tool, assisting in the screening of tuberculosis in hard-to-reach or resource-limited areas.

To read the original manuscript, click the link above.

Summary of Reviews: Soares et al. present a simplified DNA extraction protocol (FTA) followed by portable qPCR detection (Q3-Plus) of M. tuberculosis DNA, tailored for point-of-care settings. Reviewers note that rigorous validation of the sensitivity/specificity and further field studies evaluating effectiveness/practicality are required before widespread implementation. Regardless, reviewers highlight that this innovative approach not only shortens turnaround times but also underscores the importance of investing in practical and cost-effective diagnostic technologies to improve healthcare access and outcomes in resource-limited areas.

Reviewer 1 (Wilber S…) | 📒📒📒 ◻️◻️

Reviewer 2 (Haiana Charifker S…) | 📘📘📘📘📘

RR:C19 Strength of Evidence Scale Key

📕 ◻️◻️◻️◻️ = Misleading

📙📙 ◻️◻️◻️ = Not Informative

📒📒📒 ◻️◻️ = Potentially Informative

📗📗📗📗◻️ = Reliable

📘📘📘📘📘 = Strong

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