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A Ratiometric pH Sensor for Gram-Positive and Gram-Negative Bacteria.

Dorothea Kossmann1, Aya Iizuka2,3, Nina Khanna2,4

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|January 20, 2026
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Summary
This summary is machine-generated.

Researchers developed a novel fluorescent probe to track bacterial cytoplasmic pH in real-time. This tool helps identify pH-sensitive phenotypes and understand bacterial adaptation in fluctuating environments.

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Area of Science:

  • Microbiology
  • Biochemistry
  • Cell Biology

Background:

  • Bacterial populations exhibit phenotypic heterogeneity, impacting their response to environmental stressors like antibiotics and immune system components.
  • Understanding single-cell physiology is crucial for analyzing bacterial adaptation to environmental changes, particularly pH stress.
  • Current methods for real-time cytoplasmic pH monitoring in bacteria are limited.

Purpose of the Study:

  • To develop and validate a ratiometric fluorescent probe for sensing bacterial cytoplasmic pH.
  • To assess the probe's utility in real-time monitoring of pH fluctuations and identifying pH-sensitive phenotypes.
  • To evaluate the probe's application in studying bacterial interactions with immune cells.

Main Methods:

  • Synthesis of hemicyanine dye-based ratiometric fluorescent probes.
  • Uptake and validation of probes in Gram-positive (e.g., Staphylococcus epidermidis, MRSA) and Gram-negative (e.g., Escherichia coli) bacteria.
  • Real-time monitoring of cytoplasmic pH changes in response to environmental conditions and during phagocytosis by immune cells.

Main Results:

  • The fluorescent probes demonstrated rapid and reversible responses to cytoplasmic pH changes across a biologically relevant range.
  • Probes were successfully validated for cytoplasmic pH sensing in multiple bacterial species, including clinically relevant strains like MRSA.
  • The probes facilitated the identification of pH-sensitive bacterial phenotypes and monitored phagocytosis of virulent strains by immune cells.

Conclusions:

  • The developed ratiometric fluorescent probes are a promising tool for analyzing bacterial cytoplasmic pH and phenotypic heterogeneity.
  • These probes can provide insights into the physiological states of bacterial populations, including antibiotic-resistant and persistent strains.
  • This technology offers a valuable approach for studying bacterial adaptation and host-pathogen interactions in real-time.