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Rapid susceptibility testing for nontuberculosis mycobacteria using flow cytometry

S E Bownds¹, T A Kurzynski, M A Norden

¹Wisconsin State Laboratory of Hygiene, University of Wisconsin, Madison, 53706, USA.

Journal of Clinical Microbiology

|June 1, 1996

Summary

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Rapid susceptibility testing for mycobacteria is now possible using fluorescein diacetate (FDA) staining and flow cytometry. This method quickly differentiates drug-treated from untreated mycobacteria without requiring organism multiplication.

Area of Science:

Microbiology
Clinical Diagnostics
Flow Cytometry

Background:

Susceptibility testing for mycobacteria is crucial for effective treatment.
Traditional methods can be time-consuming, delaying appropriate therapy.
Rapid diagnostic tools are needed to combat mycobacterial infections.

Purpose of the Study:

To evaluate the efficacy of fluorescein diacetate (FDA) staining and flow cytometry for rapid mycobacterial susceptibility testing.
To determine if this method can differentiate drug-treated from untreated mycobacteria across various species.
To assess the assay's performance within 24 hours.

Main Methods:

Mycobacterial species (M. tuberculosis, M. avium, M. fortuitum, M. gordonae, M. marinum) were incubated with various antituberculosis agents.

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Fluorescein diacetate (FDA) staining was applied to assess cell viability.

Flow cytometry was used to quantify FDA hydrolysis and differentiate treated from untreated mycobacteria.

Main Results:

Assay suspensions treated with antimycobacterial agents showed reduced FDA hydrolysis compared to drug-free controls.
Clear differentiation between treated and untreated mycobacteria was achieved at 6 and 24 hours.
Mycobacterial multiplication was not necessary for accurate differentiation.
The assay demonstrated simplicity, reproducibility, and speed.

Conclusions:

FDA staining combined with flow cytometry provides a rapid and reliable method for mycobacterial susceptibility testing.
This technique allows for timely differentiation of drug effects on mycobacteria.
The assay is applicable to a range of mycobacterial species and various antimicrobial agents.