E-DNA detection of rpoB gene resistance in Mycobacterium tuberculosis in real samples using Fe3O4/polypyrrole nanocomposite

Abstract

In this work, we achieved the selective detection of wild and mutated rpoB gene in M. tuberculosis using an electrochemical DNA (E-DNA) sensor based on polypyrrole/Fe₃O₄ nanocomposite bearing redox naphthoquinone tag on PAMAM (spaNQ/PAMAM/PPy/Fe₃O₄). The hybridization between a given probe and the complementary DNA target induced a large decrease in the naphthoquinone redox signal as measured by SWV and no cross-hybridization with single nucleotide mismatch DNA target occurred. Thanks to the catalytic properties of iron oxide nanoparticles combined with conducting properties of polypyrrole platform, we demonstrated that the transducing system allowed the detection of 1 fM of DNA target in a 50-µL drop corresponding to 3 × 10⁴ copies of DNA. The sensor was able to detect the rpoB gene in PCR-amplified samples of genomic DNA and could also discriminate between the wild type rpoB gene and a single nucleotide mutated rpoB gene that provides resistance to rifampicin. Furthermore, the sensor could selectively detect the wild and mutant DNA in genomic samples without PCR amplification.

Keywords: E-DNA sensor; Fe(3)O(4); Mycobacterium tuberculosis; Naphthoquinone; PAMAM; Polypyrrole.