Rapid, Culture-Free Platform for Detecting Bacteria Using Surface-Enhanced Raman Spectroscopy and its Application Toward Detection of Vaginal Microbes

Abstract

Bacteria play a significant role in our health through their contributions to various microbiomes, yet certain bacteria are pathogenic, causing infectious diseases in humans. Despite these roles, clinical methods for detection and identification of bacteria at the point of care (POC) are limited. Currently, lengthy and complex procedures inhibit routine microbiome testing and pathogen identification. The goal of this dissertation is to develop a rapid, culture-free platform for bacterial detection using surface-enhanced Raman spectroscopy (SERS). Our goal is also to demonstrate the utility of the platform in a clinical application of bacterial detection in the vaginal microbiome, a vital microbial community that protects the female reproductive tract. To accomplish these goals, limitations of current SERS substrates such as cost, difficulty in handling, and accessibility, are addressed via the investigation of two low-cost substrates. First, we developed a paper-based membrane via a simple fabrication process to enable bacterial detection at physiologically relevant concentrations, using the coffee ring effect. Next, we investigated the use of two types of commercially available aluminum foils, with varying surface properties, to passively concentrate bacteria at concentrations present in many microbiomes. Toward clinical feasibility, the aluminum foil substrate was then used in conjunction with SERS for biochemical characterization and discrimination between healthy vaginal Lactobacillus in pure and complex solutions. Multivariate statistics were employed to determine the dominant Lactobacillus species in solution with low error and strong goodness of prediction. This project culminated in a proof-of-concept clinical study aimed at detecting bacteria in human vaginal fluid. The SERS spectra of vaginal fluid collected from patients receiving routine care were grouped by bacterial presence, and significant differences were identified in spectral features relating to proteins, lipids, and organic acids. Finally, toward POC use, we compared the performance of a benchtop Raman microscope and portable spectrometer and found that the portable system provided comparable results, highlighting the potential for clinical translation of the developed method. Together, these studies progressed the use of SERS for culture-free detection of bacteria in clinically relevant samples.