Designing Bacterial Chemotactic Receptors Guided by Photonic Femtoliter Well Arrays for Quantifiable, Label-Free Measurement of Bacterial Chemotaxis

Abstract

Whole cell bioreporters, such as bacterial cells, can be used for environmental and clinical sensing of specific analytes. However, the current methods implemented to observe such bioreporters in the form of chemotactic responses heavily rely on microscope analysis, fluorescent labels, and hard-to-scale microfluidic devices. Herein, we demonstrate that chemotaxis can be detected within minutes using intrinsic optical measurements of silicon femtoliter well arrays (FMAs). This is done via phase-shift reflectometric interference spectroscopic measurements (PRISM) of the wells, which act as silicon diffraction gratings, enabling label-free, real-time quantification of the number of trapped bacteria cells in the optical readout. By generating unsteady chemical gradients over the wells, we first demonstrate that chemotaxis toward attractants and away from repellents can be easily differentiated based on the signal response of PRISM. The lowest concentration of chemorepellent to elicit an observed bacterial response was 50 mM, whereas the lowest concentration of chemoattractant to elicit a response was 10 mM. Second, we employed PRISM, in combination with a computational approach, to rapidly scan for and identify a novel synthetic histamine chemoreceptor strain. Consequently, we show that by using a combined computational design approach, together with a quantitative, real-time, and label-free detection method, it is possible to manufacture and characterize novel synthetic chemoreceptors in Escherichia coli (E. coli).

Shining light in blind alleys: deciphering bacterial attachment in silicon microstructures

“…By utilizing MB, as opposed to a nutrient-rich growth medium, only events related to bacterial attachment are observed during the experiment, though cell motility is preserved. 51,65…”

“…45,46 In this method, bacterial cells colonize on diffractive, patterned Si microstructures, while zero-order reflectance spectra are continuously collected during illumination with a broadband white light source. 45–51 The resulting reflectance spectra exhibit optical interference fringes due to the reflection of the incident light at the two interfaces of the Si microstructures (top and bottom). As bacteria occupy the empty spaces within the microstructures, the refractive index of the filling medium increases, resulting in increased values of optical path difference.…”

Bacterial chemotaxis: a way forward to aromatic compounds biodegradation

“…However, these methods are laborious and time-consuming. In a recent study, Davidov et al [156] presented a novel method to quantify bacterial chemotaxis within minutes of using silicon femtoliter-well arrays by optical measurement. In this method, silicon well acts as a diffraction grating in PRISM (phase-shift reflectometric interference spectroscopic measurements) that enables label-free, real-time quantification of bacterial cells in an optical readout.…”

Antifungal susceptibility testing ofAspergillus nigeron silicon microwells by intensity-based reflectometric interference spectroscopy

“…1D-i and 1D-ii). Whereas in previous works [28][29][30][31][32][33] the values of 2nL were monitored over time, in this assay, the intensity of the reflected light is tracked, as A. niger tends to grow on top of the silicon microwells resulting in a decrease in peak intensity. The percent change in peak intensity of the fast Fourier spectrum, ΔI, of the reflected light over time is calculated as:…”

“…28 Previously, phase-shift reflectometric interference spectroscopic measurements (PRISM) was demonstrated to monitor antibiotic susceptibility and the behavior of bacteria within microstructured arrays. [29][30][31][32][33] Reflectance spectra of the arrays were collected over time in order to infer values of 2nL, in which n represents refractive index of the medium within the arrays and L represents the height of the microstructures. However, due to the different behavior and morphology of filamentous fungi compared to bacteria, herein we apply intensity-based PRISM, referred to as iPRISM, as a principle for the detection of microorganisms and as a tool for label-free, phenotypic antifungal susceptibility testing using fungal species A. niger as a model microorganism.…”