Biophysical Fluorescence Laboratory
Center For Bioimaging Sciences
Departments of Biological Sciences & Chemistry

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Bacteria and biofilms

Bacteria shield themselves with their membranes and, in communities, with biofilms. We use imaging FCS to measure how molecules move through these barriers, and how antimicrobial compounds change them.

3D rendering of a bacterial biofilm imaged by light-sheet microscopy.

Biofilms

Biofilms are dense communities of bacteria held together by a self-produced matrix, and that matrix makes them much harder to treat than free-swimming bacteria. Z-scanning light-sheet imaging FCS lets us measure how fluorescent molecules such as dextrans, antibiotics and DNA nanoparticles diffuse in and around live biofilms.

The resulting diffusion maps show molecules slowing sharply as they enter the biofilm, revealing where and how strongly its structure limits penetration.

People

In collaboration with Mark Schembri, The University of Queensland, Australia.

Z-scanning light-sheet FCS measuring diffusion inside and outside a bacterial biofilm
Z-scanning light-sheet FCS maps the diffusion coefficient of fluorescent molecules outside and inside a biofilm.
ITIR-FCS screening assay: bacteria image, segmentation mask, diffusion coefficient distributions for control and EHG-treated bacteria, and diffusion coefficient over time
(A) Bacteria imaged by ITIR-FCS and (B) automatically segmented. (C) Membrane diffusion coefficients in untreated (Ctrl) and EHG-treated bacteria. (D) Change in diffusion coefficient over incubation time.

Bacterial screening assay

There is a growing need for new antimicrobials in personal hygiene products. Compounds that attack bacterial membranes are attractive because bacteria cannot easily change their membrane composition, but existing screening methods are slow.

We combined imaging total internal reflection FCS (ITIR-FCS) with a segmentation algorithm that automatically finds bacterial cells, and measure how compounds change membrane dynamics in live bacteria across concentrations and incubation times. The assay reports membrane activity in under 30 minutes and produces dose–response curves within 2 hours, detecting activity at lower concentrations and an order of magnitude faster than commonly used susceptibility tests.

People

In collaboration with Procter & Gamble, Singapore.

Recent publications

  1. The biofilm matrix of uropathogenic Escherichia coli contributes to antimicrobial resistance by reducing molecular transportKulkarni R, Lian ZJ, Nhu NTK, Blaskovich MAT, Wohland T, Schembri MAnpj Biofilms and Microbiomes (2026). doi.org/10.1038/s41522-026-01174-6Biofilms
  2. Decoding interactions between biofilms and DNA nanoparticlesSousa A, Kulkarni R, Johannessen M, Wohland T, Skalko-Basnet N, Obuobi SBiofilm (2025). doi.org/10.1016/j.bioflm.2025.100260Biofilms
  3. A live bacterial screening assay for membrane-active antimicrobial compounds using imaging fluorescence correlation spectroscopyZhu S, Wang Y, Chong SMS, Wohland TAnalytical Chemistry (2025). doi.org/10.1021/acs.analchem.4c05698Screening assay
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