Nanoporous layer fiber biosensing platform for real time culture- and separation-free detecting bacterial pathogens and measuring their susceptibility to antibiotics

•Nanoporous layer fiber biosensing platform has been developed for pathogens detection.•NFB integrates evanescent wave fluorescence technology and size effect of nanoporous layerl In situ etched nano-layer of fiber probe can effectively distinguish free aptasensor and aptasensor bound with bacteria.•NFB can rapidly quantify E.coli O157:H7 and S. typhimurium in real samples with high selectivity and sensitivity l Portable NFB can serve as a powerful tool for real time measurement of the response of bacteria to antibiotics. Simple, rapid, and sensitive detection of bacterial pathogens in environmental and clinical samples is a key requirement because of the ensuing high morbidity and mortality. In this study, a nanoporous layer fiber biosensing platform (called NFB) was developed for the rapid culture- and separation-free detection of bacterial pathogens through integrating evanescent wave fluorescence technology, the size effect of nanoporous layer, and real-time aptasensors. The in-situ etched nanoporous layer of the fiber probe can not only effectively distinguish free Cy5.5-aptasensors and Cy5.5-aptasensors bound to pathogens but also enhance evanescent wave fluorescence. NFB can selectively and sensitively quantify Escherichia coli O157:H7 and Salmonella typhimurium species in environmental and blood samples within 18 min and 32 min, respectively. NFB detected a significant increase in fluorescence when the mixture of pathogens and Cy5.5-aptasensors was exposed to antibiotics. Our results clearly demonstrated that the portable NFB can serve as a powerful tool for real time detection of pathogens and measurement of their response to antibiotics.