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Updated: Sep 14, 2025
Author Spotlight: Advancing VRL Diagnosis Using Cell-Free DNA Extraction from Vitreous Humor
Published on: January 12, 2024
Evaluation of Mycobacterium Tuberculosis Derived Cell-Free DNA-Based Multi-Targeted Real-Time PCR from Vitreous Fluid
Kusum Sharma1, Megha Sharma2, Himanshu Joshi1
1Department of Medical Microbiology and Internal Medicine, Postgraduate Institute of Medical Education and Research, Chandigarh, India.
Background:
Ocular tuberculosis (OTB) is an important cause of vision loss in India. Rapid and accurate diagnosis of OTB is paramount to decreasing morbidity and saving the eyes from blindness. DNA present in cell-free form is expected to give a higher detection yield. Therefore, the present study aimed to evaluate the utility of Cell-free DNA (CF-DNA)-based Multi-targeted real-time PCR (Mrt-PCR) for diagnosing OTB and compare it with Truenat MTB plus and GeneXpert MTB/RIF Ultra (Xpert Ultra).
Methods:
CF-DNA-based Mrt-PCR (using IS6110, IS1081, and nrdZ genes), Truenat MTB plus, and Xpert Ultra were carried out on 50 vitreous fluid (VF) samples [2 confirmed OTB, 22 clinically suspected of OTB, and 25 controls]. rpoB gene sequencing was carried out in all the positive cases to detect Rifampicin susceptibility.
Results:
CF-DNA-based Mrt-PCR, Truenat MTB plus, and Xpert Ultra were positive in 19/25(76%), 16/25(64%), and 13/25(56%) patients of OTB, respectively, and negative in all the 25 control group subjects. There were three cases detected only by IS1081, two only by IS6110 and one only by nrdZ gene. Truenat MTB RIF and Xpert Ultra detected RIF resistance in three cases, which were confirmed by rpoB gene sequencing.
Conclusion:
CF-DNA-based Mrt-PCR is a robust and promising technique for diagnosing OTB, yielding a higher detection rate than contemporary platforms -Truenat MTB plus and Xpert Ultra.
