Multiplex detection of extensively drug resistant tuberculosis using binary deoxyribozyme sensors

Area of Science:

• Molecular Biology
• Diagnostic Technologies
• Antimicrobial Resistance

Background:

• Current Mycobacterium tuberculosis (Mtb) diagnostics suffer from low sensitivity, slow results, and high costs.
• The rise of multidrug-resistant (MDR) and extensively drug-resistant (XDR) Mtb strains poses a significant global health challenge.
• There is a critical need for accurate, affordable, and rapid point-of-care molecular diagnostic tests for Mtb.

Purpose of the Study:

• To establish proof-of-concept for a novel diagnostic platform, TB-DzT, for Mtb detection.
• To identify drug-resistant Mtb mutants using binary deoxyribozyme (BiDz) sensors.
• To develop a cost-effective and sensitive alternative for MDR- and XDR-TB molecular diagnostics.

Main Methods:

• Developed the TB-DzT platform combining multiplex PCR with single nucleotide polymorphism (SNP) detection.
• Utilized highly selective BiDz sensors targeting loci for species typing and resistance to rifampin, isoniazid, and fluoroquinolones.
• Validated the assay on clinical isolates for Mtb detection and identification of specific resistance mutations.

Main Results:

• Accurately detected Mtb and 5 key mutations associated with resistance to three major anti-TB drugs in clinical samples.
• Demonstrated the ability of TB-DzT to detect heteroresistance, a minority population of drug-resistant Mtb.
• Showcased the platform's capability to detect unknown mutations at target loci using combinatorial BiDz sensors.

Conclusions:

• The TB-DzT platform offers a foundation for developing accurate, sensitive, and cost-effective molecular diagnostics for Mtb.
• This novel approach provides a promising solution for the timely diagnosis of drug-resistant tuberculosis.
• The developed BiDz sensor technology enables precise detection of resistance mutations and heteroresistance patterns.