pHEAA/pMA-DNA Hydrogel Sensing Platform for Detecting Drug-Resistant Gene Variants in Mycobacterium tuberculosis.

Mutations in the drug-resistant gene ofMycobacterium tuberculosiscan make it challenging to use drugs in clinical practice. Traditional genetic testing for resistance requires cell culture and susceptibility testing, which take 1-2 weeks. In this study, a DNA-sensitive hydrogel (pHEAA/pMA-DNA) has been developed with nucleic acid binding ability, water retention capacity, and high-temperature resistance, allowing it to work normally at 105 °C. Molecular dynamics simulations have been used to obtain the necessary physicochemical parameters. The DNA-sensitive hydrogel acts as a novel biosensor for detecting rifampicin- and isoniazid-resistant gene mutations in Mycobacterium tuberculosis. The microarray sensor's detection range is between 109 copies/ml and 101 copies/mL, and its stability coefficient of variation (CV) is 2.424%. The study demonstrates that there is no mutual interference in the gel lattice. In addition, experiments on actual nucleic acid samples reveal accurate detection of bacterial strains and drug-resistant gene mutations. The regression curve conforms to the kinetic characteristics of nucleic acid amplification, exhibiting a sigmoidal shape. The Four-Parameter Logistic Regression (4PL) equation was employed for fitting, achieving an excellent coefficient of determination (R2 = 0.99791 > 0.99). The method enables parallel detection of microarray biosensors in multidrug-resistant Mycobacterium tuberculosis. The sensors show high efficiency in detecting resistance mutation sites of Mycobacterium tuberculosis to rifampicin and isoniazid, paving the way for researchers to design different probes in in vitro detection fields.