The development of biosensors for the rapid identification of pathogenic bacteria with high sensitivity and specificity is highly desired for environmental monitoring, biomedical diagnostics, and homeland security. Current detection methods, such as bacteriologic culture, serological, and PCR tests, are either time consuming or require the use of high-end instruments and species-specific antibodies, greatly complicating their engineering transition to real world applications. Biosensors based on surface-enhanced Raman spectroscopy (SERS) for direct detection and discrimination of bacteria offers many advantages over the current detection methods such as being reagent-less, multiplexing and the potential for on-line and infield applications. However, conventional SERS is a near-field effect which severely limits the detection of bacteria. Intellectual Merit: In this work, the PI challenge the current pathogenic bacteria detection methods by proposing to develop a new biosensor platform based on long-rang SERS (LRSERS) for the sensitive detection and rapid identification of pathogenic bacteria. The LR-SERS concept is supported by the fundamental physics in that the surface plasmon resonance (SPR) can be extended to a long distance from the metal surface by the coupling of SPRs at both metal/dielectric interfaces of a metal thin film. This work is aimed to rationally design fundamentally new LR-SERS substrates using theoretical electromagnetic calculations and to develop a microarray biosensor based on the LR-SERS platform. Electromagnetic finite-difference time-domain (FDTD) calculations will be performed first in order to rationally design the plasmonic nanostructures with the strongest local electric field away from the metal surface to ~ 50 nm, the length covering bacterial cell walls. Subsequently, the LR-SERS substrates will be fabricated via electron beam lithography (EBL) for the detection of bacteria. While the LRSERS microarray biosensor developed in this project is targeted for rapid identification of any pathogenic bacteria, we will use the isolates of vibrio species for testing because the pathogenic marine bacterium V. parahaemolyticus is the leading cause of seafood-borne bacterial illness in the word. The LR-SERS spectra of isolates of V. parahaemolyticus from different phylogenetic groups and sources (environment and clinical) will be collected and analyzed to determine the correlation of LR-SERS spectral characteristic peaks to the gene encoding outer membrane and cell wall proteins of pathogenic strains. A method for the quick identification of pathogenic V.parahaemolyticus from a mixture of bacteria strains will be established using LR-SERS barcode and principle component analysis (PCA). And finally, LR-SERS microarray biosensors will be integrated with a microfluidic system for the in-situ, sensitive, and high-throughput detection and identification of pathogenic bacteria. Broader Impact: The LR-SERS microarray biosensors developed in this project provide a new approach strategy that can lead to, for example, the development of biosensor networks deployed off-coast for health early warning systems. Results from this project will make significant impact on the related fields such as analytical chemistry, biomedicine, pharmacology, forensics, food safety, agriculture, bio-fuel research, environmental monitoring, and bio-defense. Graduate, undergraduate, and community college students will receive training and participate in this highly interdisciplinary research project, especially students from underrepresented groups such as female. The participation of international exchange students will promote the international collaborations. The knowledge gained from this work will be disseminated through the lectures the PI provides for the courses every year and via outreach programs established by the UW Center for Nanotechnology and the Seattle Pacific Science Center.
