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Key Factors for Removing Bias in Viability PCR-Based Methods: A Review
F Codony1, M Dinh-Thanh2, G Agustí3
1Laboratori Municipal Aigües de Mataró, Carrer Pitàgores 1-7, 08304, Mataró, Spain. Francesc.codony@aiguesmataro.cat.
Abstract:
During the last decade, the viability PCR approach has been investigated intensively with the goal of detecting solely the DNA derived from intact cells by removing the bias resulting from dead or damaged cell artifacts. However, since the first published paper in 2003 and despite the various continuous improvements, the stubborn reality is that in most cases the goal has not been achieved. Complete neutralization of nucleic acids derived from dead microorganisms was not possible, at least without affecting live cells. However, recently published works in this field show how neutralization rates can be improved. In this communication, we review recent improvements that enable effective vPCR procedures.
Insights
Viability PCR (vPCR) aims to detect DNA from intact cells, but neutralizing dead cell DNA without harming live cells remains challenging. Recent advancements show improved neutralization rates for more effective vPCR procedures.
Area of Science:
- Microbiology
- Molecular Biology
- Environmental Science
Background:
- Viability PCR (vPCR) is designed to detect DNA from intact cells, excluding DNA from dead or damaged cells.
- Despite extensive research since 2003, effectively neutralizing DNA from non-viable cells without impacting viable cells has been a persistent challenge.
- Artifacts from dead cells can introduce bias in microbial community analysis.
Purpose of the Study:
- To review recent improvements in viability PCR (vPCR) techniques.
- To highlight advancements enabling more effective vPCR procedures.
- To address the challenge of complete neutralization of DNA from non-viable cells.
Main Methods:
- Review of recently published literature on viability PCR.
- Analysis of improvements in vPCR protocols and methodologies.
- Evaluation of methods for neutralizing nucleic acids from dead cells.
Main Results:
- While complete neutralization remains difficult, recent studies demonstrate improved neutralization rates in vPCR.
- New methods show promise in reducing bias from dead cell artifacts.
- Advancements allow for more effective discrimination between viable and non-viable cells.
Conclusions:
- Recent improvements have enhanced the effectiveness of viability PCR procedures.
- Further research continues to refine vPCR for accurate detection of DNA from intact cells.
- Effective vPCR is crucial for unbiased microbial analysis.
