Accessible, robust ultrahigh-throughput microbiome analysis via integration of uniform droplet-templated emulsification and FACS
- 1Department of Biotechnology, Center for Microbial Ecology and Technology (CMET), Ghent University, Ghent, Belgium
- 2Department of Microbiology, Immunology and Transplantation, Laboratory of Molecular Bacteriology (Rega Institute), KU Leuven, Leuven, Belgium
- 3Centre for Advanced Process Technology for Urban Resource recovery (CAPTURE), Ghent, Belgium
Abstract
Microbial phenotypes vary at the single-cell level, shaping key community traits like resilience and adaptability. Yet, current methods either lack resolution (e.g., culturing, sequencing), are too costly, or technically complex, limiting widespread use. To address this gap, we introduce and validate a workflow called DE-SWIRL (Double Emulsion–Sorting Workflow with sImple, Rapid emuLsification), an accessible, low-cost workflow enabling ultrahigh-throughput (~107 microcultures/experiment) screening of individual microbial cells. DE-SWIRL integrates a published droplet-templated emulsification protocol producing uniform double emulsions from monodisperse single emulsions with Fluorescence-Activated Cell Sorting (FACS). We validated that double emulsions of 6 and 24 pL can be reliably formed, with ~45% droplet survival. To address persistent large-particle contaminants, we validated a gating strategy and show it enables accurate screening and sorting. When starting from a monodisperse single emulsion population, oil layer variability is higher for droplet-templated emulsification than for on-chip microfluidics, but maintains a comparably uniform core emulsion while offering substantial time savings. We demonstrate DE-SWIRL's utility by isolating viable strains from a synthetic community with up to 99% sorting purity and isolating droplet cocultures from a mixed community. This workflow provides a fast, accessible, and affordable workflow for screening entire microbiomes at a single-cell level using a fluorescent assay of interest.
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BibTeX
@article{nauwynck2026accessible,
title = {{Accessible, robust ultrahigh-throughput microbiome analysis via integration of uniform droplet-templated emulsification and FACS}},
author = {Wannes Nauwynck and Karoline Faust and Nico Boon},
journal = {PLOS One},
year = {2026},
month = sep,
doi = {10.1371/journal.pone.0356887},
url = {https://doi.org/10.1371/journal.pone.0356887},
note = {CC BY 4.0},
}
RIS
TY - JOUR
TI - Accessible, robust ultrahigh-throughput microbiome analysis via integration of uniform droplet-templated emulsification and FACS
AU - Wannes Nauwynck
AU - Karoline Faust
AU - Nico Boon
PY - 2026
DA - 2026/09/09
JO - PLOS One
DO - 10.1371/journal.pone.0356887
UR - https://doi.org/10.1371/journal.pone.0356887
AB - Microbial phenotypes vary at the single-cell level, shaping key community traits like resilience and adaptability. Yet, current methods either lack resolution (e.g., culturing, sequencing), are too costly, or technically complex, limiting widespread use. To address this gap, we introduce and validate a workflow called DE-SWIRL (Double Emulsion–Sorting Workflow with sImple, Rapid emuLsification), an accessible, low-cost workflow enabling ultrahigh-throughput (~107 microcultures/experiment) screening of individual microbial cells. DE-SWIRL integrates a published droplet-templated emulsification protocol producing uniform double emulsions from monodisperse single emulsions with Fluorescence-Activated Cell Sorting (FACS). We validated that double emulsions of 6 and 24 pL can be reliably formed, with ~45% droplet survival. To address persistent large-particle contaminants, we validated a gating strategy and show it enables accurate screening and sorting. When starting from a monodisperse single emulsion population, oil layer variability is higher for droplet-templated emulsification than for on-chip microfluidics, but maintains a comparably uniform core emulsion while offering substantial time savings. We demonstrate DE-SWIRL's utility by isolating viable strains from a synthetic community with up to 99% sorting purity and isolating droplet cocultures from a mixed community. This workflow provides a fast, accessible, and affordable workflow for screening entire microbiomes at a single-cell level using a fluorescent assay of interest.
SN - 1932-6203
ER -