A comparison of nucleic acid amplification techniques for the assessment of bacterial viability

Aims: The ability to determine the presence and viability status of bacteria by molecular methods could offer significant advantages to the food, environmental and health sectors, in terms of improved speed and sensitivity of detection. Methods and Results: In this study, we have assessed three amplification techniques, PCR, RT-PCR and NASBA, for their ability to detect nucleic acid persistence in an E. coli strain following heat-killing. NASBA offered the greatest sensitivity of the three methods tested. The presence of residual DNA and mRNA could be detected by PCR and NASBA, respectively, for up to 30 h postdeath, by which time cell death had been confirmed by culture methods. Thus a single quantitative measurement based on nucleic acid amplification did not permit unequivocal determination of cell viability. Conclusions, Significance and Impact of the Study: The correlation between cell viability and persistence of nucleic acids must be well characterized for a particular analytical situation before molecular techniques can be substituted for traditional culture methods.