Combining Phenotypic and Genotypic Methods for the Rapid Detection of Antibiotic-Resistant Bacteria in Wastewater...

Antimicrobial resistance (AMR) has emerged as a critical global health threat, leading to untreatable infectious diseases and significantly increasing human morbidity and mortality. Rapid, portable, and user-friendly detection of antibiotic-resistant bacteria (ARB) in the environment plays a key role in controlling the spread of diseases and combating the emergence of AMR. In this study, we report a portable microfluidic centrifugal disc (CD) assay incorporating both phenotypic and genotypic methods for ARB detection. This assay significantly reduces the time required for bacterial culturing from days to hours while maintaining the reliability of phenotypic methods. Moreover, the CD assay eliminates the need for a centralized laboratory and well-trained personnel. The on-CD ARB detection can be completed in less than 3 h after sample and reagent loading. The results demonstrated that the on-CD assay provided performance comparable to that of the benchtop ARB detection assay. Analysis of variance (ANOVA) comparing the benchtop and on-CD assays yielded a p-value of 0.08 for the bacterial growth rate and a p-value of 0.73 for the cell recovery rate after the sample purification, confirming no significant differences between the two approaches. The ARB detection assay was able to identify ampicillin-resistant spiked in wastewater at concentrations as low as 10 CFU/μL. Additionally, the CD assay detected indigenous ampicillin-resistant in wastewater, demonstrating the potential of this platform for environmental applications.