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Related Experiment Video

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Phage Phenomics: Physiological Approaches to Characterize Novel Viral Proteins
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Dissecting Bacterial Growth Variants by Cellular Phenotypic Profiling and Backtracing.

Jonathan Hira1, Christian S Lentz2

  • 1Department of Medical Biology and Centre for New Antibacterial Strategies (CANS), UiT-The Arctic University of Norway, Tromsø, Norway.

Methods in Molecular Biology (Clifton, N.J.)
|January 1, 2026
PubMed
Summary

This study introduces a new method, Cellular Phenotypic Profiling and backTracing (CPPB), to distinguish bacterial cell states. CPPB links single-cell growth fate with phenotypic traits, aiding the study of bacterial heterogeneity.

Keywords:
Fluorescence-activated cell sortingFluorescent probePersisterPhenotypic heterogeneityStaphylococcus aureusViable-but non-culturable (VBNC)

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Area of Science:

  • Microbiology
  • Cell Biology
  • Molecular Biology

Background:

  • Bacterial populations exhibit diverse phenotypes, including growing, persistent, and viable-but-non-culturable cells.
  • Differentiating these states is difficult due to a lack of biomarkers and the dynamic nature of cell phenotypes.
  • Understanding these states is crucial for controlling bacterial infections and developing new treatments.

Purpose of the Study:

  • To present a novel technique, Cellular Phenotypic Profiling and backTracing (CPPB), for differentiating bacterial cell phenotypes.
  • To provide step-by-step protocols for implementing the CPPB method.
  • To demonstrate the utility of CPPB in studying bacterial dormancy and antibiotic tolerance.

Main Methods:

  • CPPB utilizes index sorting to link single-cell growth fate with phenotypic characterization.
  • Fluorescent probes are employed for detailed cellular phenotypic profiling.
  • The method was validated using Staphylococcus aureus under low pH dormancy-inducing conditions with live-dead staining.

Main Results:

  • The study successfully demonstrated the application of CPPB in differentiating bacterial cell states.
  • CPPB effectively connected single-cell growth trajectories with their specific phenotypic profiles.
  • The technique proved adaptable to different bacterial species and various fluorescent probes.

Conclusions:

  • CPPB offers a powerful tool for dissecting bacterial phenotypic heterogeneity.
  • This method facilitates the study of bacterial dormancy, persistence, and antibiotic tolerance.
  • CPPB is a versatile technique with broad applicability in microbiology and infectious disease research.