Propidium monoazide combined with real-time quantitative PCR underestimates heat-killed Listeria innocua

Area of Science:

• Microbiology
• Molecular Biology

Background:

• Accurate differentiation between viable and dead bacteria is crucial in food safety and clinical diagnostics.
• Quantitative PCR (qPCR) combined with propidium monoazide (PMA) is a common method for this differentiation.
• However, the reliability of PMA-qPCR needs validation against established methods.

Purpose of the Study:

• To evaluate the accuracy of propidium monoazide quantitative PCR (PMA-qPCR) for determining viable Listeria innocua.
• To compare PMA-qPCR results with traditional plate counts and confocal fluorescence microscopy.

Main Methods:

• Bacterial samples of Listeria innocua were treated with PMA.
• DNA was extracted and amplified using qPCR.
• Results were compared with those obtained from standard plate counts and confocal fluorescence microscopy.

Main Results:

• PMA-qPCR significantly overestimated the proportion of viable Listeria innocua cells.
• Discrepancies were noted when compared to both plate counts and confocal fluorescence microscopy.
• This suggests potential inaccuracies in PMA's ability to differentiate live and dead cells in this context.

Conclusions:

• The combination of PMA and qPCR may not reliably distinguish between viable and dead Listeria innocua.
• Caution is advised when employing PMA-qPCR as a sole analytical tool for viability assessment.
• Further research is needed to optimize or validate PMA-qPCR for bacterial viability studies.