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Assessment of Gram- and Viability-Staining Methods for Quantifying Bacterial Community Dynamics Using Flow Cytometry
Aurore Duquenoy1, Samuel Bellais2, Cyrielle Gasc1
1MaaT Pharma, Lyon, France.
Frontiers in Microbiology
|July 18, 2020
Summary
Flow cytometry offers a rapid, cost-effective method for analyzing gut microbiota dynamics. This technique successfully identified culturable anaerobic bacteria and differentiated Gram-positive from Gram-negative species in complex ecosystems.
Area of Science:
- Microbiology
- Biotechnology
- Analytical Chemistry
Background:
- Gut microbiota research is crucial for understanding human diseases.
- Current metagenomics profiling is time-consuming and requires expertise.
- There's a need for faster, simpler tools for microbial ecosystem analysis.
Purpose of the Study:
- To evaluate flow cytometry for analyzing microbial communities.
- To assess the culturability of anaerobic bacteria sorted by flow cytometry.
- To develop and validate staining methods for discriminating bacterial types.
Main Methods:
- Live/Dead staining and flow cytometry sorting of strictly anaerobic bacteria.
- Use of labeled vancomycin and Wheat Germ Agglutinin (WGA) for Gram staining.
- Application of staining methods on complex ecosystems and validation with 16S rRNA sequencing.
Main Results:
- 10-80% of "viable" anaerobic bacteria sorted by flow cytometry were culturable.
- Staining methods successfully differentiated Gram-positive and Gram-negative bacteria.
- Minor pH variations significantly impacted microbial community structure.
Conclusions:
- Flow cytometry is a rapid and efficient tool for analyzing complex microbial ecosystems.
- The developed staining method facilitates the study of bacterial community dynamics.
- This approach supports applications in research and industrial process control.
