Applied Research for Liquid-phase Chip Detection of Foodborne Salmonella Carrying Plasmid-mediated Quinolone Resistance Genes

Objective: To establish a method to detect four genes: qnrS, aac(6')-Ib-cr, oqxA, oqxB, among the plasmid-mediated quinolone resistance (PMQR) genes carried by foodborne Salmonella by applying a novel liquid-phase chip technique. Methods: For the four PMQR genes carried by foodborne Salmonella: qnrS, aac(6')-Ib-cr, oqxA, and oqxB, corresponding primers and microspheres were designed, and liquid-phase microarray technology was used to perform specificity experiments on seven standard strains. Reproducibility and sensitivity experiments were performed on four foodborne Salmonella strains containing one PMQR gene each. 71 quinolone resistant Salmonella strains from foodborne risk monitoring were tested, and a comparison experiment with ordinary PCR was carried out. Results: A method for the detection of four PMQR genes, qnrS, aac(6')-Ib-cr, oqxA and oqxB, in foodborne Salmonella was successfully established by liquid-phase microarray technology. The LOD (limit of detection) of qnrS, aac(6')-Ib-cr, oqxA and oqxB were 5, 25, 10 and 10 CFU/mL respectively. The median fluorescence values (MFI) of all positive determinations were ≥5 times those of the negative control group. The coefficients of variation (CV) of the repeatability experiments were less than 5%. In the specificity experiments, all quinolone resistant Salmonella strains were detected, and no cross-reactivity with other non-target bacteria was observed. The detection rate of qnrS, aac(6')-Ib-cr, oqxA and oqxB were 29.6% (21/71), 35.2% (25/71), 28.2% (20/71), 23.9% (17/71) respectively. The coincidence rate between the liquid chip technology and PCR was 100%. Conclusion: The experimentally established liquid-phase chip technique for the detection of foodborne Salmonella plasmid-mediated quinolone resistance genes qnrS, aac(6')-Ib-cr, oqxA and oqxB is characterized by high sensitivity, good specificity, stability, and accurate results, which can provide technical support for the detection of foodborne Salmonella PMQR genes and the monitoring of drug resistance.