Rapid visual and direct detection of Acinetobacter baumannii and carbapenem resistance by oligoDNA-gold nanoparticles biosensor in clinical samples

Acinetobacter baumannii is a major nosocomial pathogen responsible for extensively drug-resistant (XDR) infections in healthcare settings and is ranked as a critical priority one pathogen on World Health Organisation list. However, clinical diagnosis generally is given as (Acinetobacter calcoaceticus-baumannii) ACB complex, including A. baumannii, A. calcoaceticus, A. nosocomialis, and A. pittii, and among these, A. baumannii is the most clinically significant species. Further the conventional diagnostic methods take 4-48 h and many of them require sophisticated infrastructure, highlighting the urgent need for rapid and economical detection methods both for A. baumannii and its carbapenem resistance determinants. Among the carbapenem-resistant A. baumannii, bla_OXA-23 and bla_NDM are the predominant β-lactamase genes, with New Delhi metallo-β-lactamase (NDM) conferring broad-spectrum carbapenem resistance and severely limiting the therapeutic options. In the present study, oligoDNA-gold nanoparticle (AuNP)-based biosensors were developed for the direct, rapid and visual detection of A. baumannii, bla_OXA-23 and bla_NDM in clinical respiratory samples without the need for DNA purification or amplification. The biosensor for the detection of A. baumannii targeted a conserved region of the Outer Membrane Nuclease encoding nucAb gene, enabling its accurate identification. bla_OXA-23 biosensor detected bla_OXA-23 gene, which is the highly prevalent carbapenem resistance gene in A. baumannii while the other biosensor, based on the conserved region of bla_NDM gene, detected bla_NDM-1 not only in A. baumannii but also its variants in Klebsiella pneumoniae, Escherichia coli and Pseudomonas aeruginosa. These biosensors produced visual results within 30 min, with a detection limit of 0.5 ng/µL DNA or 10³ CFU/mL to support early clinical decision-making for optimised therapy and antimicrobial resistance surveillance.