Stress- and growth rate-related differences between plate count and real-time PCR data during growth of Listeria monocytogenes.
To assess the overestimation of bacterial cell counts in real-time PCR in relation to stress and growth phase, four different strains of L. monocytogenes were exposed to combinations of osmotic stress (0.5 to 8% [vol/vol] NaCl) and acid stress (pH 5 to 7) in a culture model at a growth temperature of 10 degrees C or were grown under optimal conditions. Growth curves obtained from real-time PCR, optical density, and viable count data were compared. As expected, optical density data revealed entirely different growth curves. Good to moderate growth conditions yielded good correlation of real-time PCR data and plate count data (r(2) = 0.96 and 0.99) with similar cell counts. When growth conditions became worse, the numbers of CFU decreased during the stationary phase, whereas real-time-PCR-derived bacterial cell equivalents differed in this regard; the correlation worsened (r(2) = 0.84). However, fitted growth curves revealed that maximum growth rates calculated from real-time PCR data were not significantly different from those derived from plate count data. The overestimation of bacterial cell counts by real-time PCR observed in the stationary phase under higher-stress conditions might be explained by the accumulation of viable but nonculturable bacteria or dead bacteria and extracellular DNA. Considering these results, real-time PCR data collected from naturally contaminated samples should be viewed with caution.
Reichert-Schwillinsky
Franziska
F
Institute for Milk Hygiene, Milk Technology, and Food Safety, Department for Farm Animals and Veterinary Public Health, University of Veterinary Medicine, Veterinaerplatz 1, 1210 Vienna, Austria.
Pin
Carmen
C
Dzieciol
Monika
M
Wagner
Martin
M
Hein
Ingeborg
I
D000143:Acids / Q000494:pharmacology; D000900:Anti-Bacterial Agents / Q000494:pharmacology; D015169:Colony Count, Microbial / Q000379:methods*; D008089:Listeria monocytogenes / Q000187:drug effects / Q000254:growth & development*; D050296:Microbial Viability; D009997:Osmotic Pressure; D016133:Polymerase Chain Reaction / Q000379:methods*; D013223:Statistics as Topic; D013312:Stress, Physiological
D000143:Acids; D000900:Anti-Bacterial Agents