Microfluidic Chip-based Enrichment and Nucleic Acid Extraction for Quantitative Detection of Mycobacterium Smegmatis in Aerosols.

Abstract

Tuberculosis (TB) is ranked as the third most prevalent infectious disease globally. Early detection and treatment are crucial for effective management. Conventional diagnostic methods primarily rely on sputum samples, which present challenges in accessibility and have limited accuracy in certain populations such as children, individuals with HIV, and those with extrapulmonary TB. To address the need for point-of-care diagnostics, this study introduces a rapid diagnostic approach for TB using exhaled breath aerosol as a more easily obtainable specimen. Mycobacterium smegmatis, a non-pathogenic bacterium genetically similar to Mycobacterium tuberculosis, is used as a surrogate organism. The method involves the use of microfluidic chips for concentrating and electrolyzing mycobacteria in the aerosol, followed by extracting and quantifying nucleic acids using real-time fluorescence quantitative PCR. Notably, successful enrichment and quantification of bacterial content were achieved even at a minimal bacterial aerosol concentration of 10⁴ CFU/mL. The developed chips are characterized by their cost-effectiveness, ease of use, high bacterial enrichment, efficient nucleic acid extraction, and low detection threshold (4.4 × 10^-18 mol/L). This innovative approach offers a promising method for early TB screening and opens avenues for the rapid identification of other aerosol-transmitted diseases.