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Measuring pH of the Coxiella burnetii Parasitophorous Vacuole
Dhritiman Samanta1, Stacey D Gilk1
1Department of Microbiology and Immunology, Indiana University School of Medicine, Indianapolis, Indiana.
Abstract:
Coxiella burnetii is the causative agent of human Q fever, a zoonotic disease that can cause a debilitating, flu-like illness in acute cases, or a life-threatening endocarditis in chronic patients. An obligate intracellular bacterial pathogen, Coxiella survives and multiplies in a large lysosome-like vacuole known as the Coxiella parasitophorous vacuole (CPV). A unique characteristic of the CPV is the acidic environment (pH ∼5.0), which is required to activate Coxiella metabolism and the Coxiella type 4 secretion system (T4SS), a major virulence factor required for intracellular survival. Further, inhibiting or depleting vacuolar ATPase, a host cell protein that regulates lysosomal pH, inhibits intracellular Coxiella growth. Together, these data suggest that CPV pH is an important limiting factor for Coxiella growth and virulence. This unit describes a method to determine CPV pH using live cell microscopy of a pH-sensitive fluorophore conjugated to dextran. This technique is useful to measure changes in CPV pH during infection or in response to drug treatment. © 2017 by John Wiley & Sons, Inc.
Insights
Measuring the acidic environment of Coxiella parasitophorous vacuole (CPV) is key to understanding Coxiella burnetii virulence. This study details a microscopy method to determine CPV pH, crucial for Q fever research.
Area of Science:
- Microbiology
- Cell Biology
- Infectious Diseases
Background:
- Coxiella burnetii causes Q fever, a zoonotic disease with acute and chronic manifestations.
- Coxiella burnetii resides within a unique lysosome-like vacuole, the Coxiella parasitophorous vacuole (CPV).
- The acidic pH (∼5.0) of the CPV is essential for Coxiella metabolism and virulence, particularly for activating the type 4 secretion system (T4SS).
Purpose of the Study:
- To describe a method for determining the pH of the Coxiella parasitophorous vacuole (CPV).
- To provide a tool for measuring changes in CPV pH during infection or in response to therapeutic interventions.
Main Methods:
- Utilizes live cell microscopy.
- Employs a pH-sensitive fluorophore conjugated to dextran to probe vacuolar acidity.
- Focuses on quantifying the acidic environment within the CPV.
Main Results:
- The described method allows for the measurement of CPV pH.
- This technique can assess how CPV pH is altered during infection.
- The method is applicable to evaluating the impact of drug treatments on CPV pH.
Conclusions:
- CPV pH is a critical factor influencing Coxiella burnetii growth and virulence.
- The developed microscopy technique offers a valuable approach to study CPV pH dynamics.
- Understanding and manipulating CPV pH may present therapeutic strategies against Q fever.
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