Fluorescent nanosensors reveal dynamic pH gradients during biofilm formation

Abstract

Understanding the dynamic environmental microniches of biofilms will permit us to detect, manage and exploit these communities. The components and architecture of biofilms have been interrogated in depth; however, little is known about the environmental microniches present. This is primarily because of the absence of tools with the required measurement sensitivity and resolution to detect these changes. We describe the application of ratiometric fluorescent pH-sensitive nanosensors, as a tool, to observe physiological pH changes in biofilms in real time. Nanosensors comprised two pH-sensitive fluorophores covalently encapsulated with a reference pH-insensitive fluorophore in an inert polyacrylamide nanoparticle matrix. The nanosensors were used to analyse the real-time three-dimensional pH variation for two model biofilm formers: (i) opportunistic pathogen Pseudomonas aeruginosa and (ii) oral pathogen Streptococcus mutans. The detection of sugar metabolism in real time by nanosensors provides a potential application to identify therapeutic solutions to improve oral health.

Monitoring and imaging pH in biofilms utilizing a fluorescent polymeric nanosensor

“…However, many nanosensors reported so far can only map very narrow pH ranges or have a relatively high aggregation tendency in biological systems, which makes them unsuitable for this task 20 , 31 . Moreover, the broad application of such nanosensors for pH measurements in biofilms requires either commercial systems, which are not yet available, or at least sensor particles that can be easily prepared from commercial components without the need for an elaborate synthesis 26 , 32 .…”

Plasmonic Au@Ag@mSiO2 Nanorattles for In Situ Imaging of Bacterial Metabolism by Surface-Enhanced Raman Scattering Spectroscopy

“…In this context, various fluorescence-based approaches have been developed to noninvasively monitor pH changes in bacterial populations. 62 The aforementioned analytical approaches have limited multiplex capabilities and generally only allow monitoring a single parameter (e.g., pH). As shown herein, the plasmonic Au@Ag@mSiO 2 nanorattles enabled not only the sensing of pH in bacterial colonies but also the detection of secreted metabolites (e.g., pyocyanin) in bacterial cultures, highlighting the promising potential of the nanosensor for multiplex sensing of bacterial metabolism by SERS.…”

“…However, these methods are generally destructive as they involve the preparation of cellular extracts for biochemical analysis. In this context, various fluorescence-based approaches have been developed to noninvasively monitor pH changes in bacterial populations . The aforementioned analytical approaches have limited multiplex capabilities and generally only allow monitoring a single parameter (e.g., pH).…”

Microbial Biofilms: Structural Plasticity and Emerging Properties

“…As an example, pH variation in environmental microniches in Pseudomonas aeruginosa and Streptococcus mutans biofilms was monitored in real time using pH-sensitive nanosensors. These fluorescent nanosensors revealed pH gradients during the building of 3D structures that helped to better understand the dynamic modulations of extracellular matrices during the developmental process and to potentially identify targets for biofilm control [31].…”