Open AccessStress- and Growth Rate-Related Differences between Plate Count and Real-Time PCR Data during Growth of Listeria monocytogenes
Author(s) -
Franziska Reichert-Schwillinsky,
Carmen Pin,
Monika Dzieciol,
Martin Wagner,
Ingeborg Hein
Publication year - 2009
Publication title -
applied and environmental microbiology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.552
H-Index - 324
eISSN - 1070-6291
pISSN - 0099-2240
DOI - 10.1128/aem.01796-08
Subject(s) - bacterial growth , listeria monocytogenes , real time polymerase chain reaction , bacteria , biology , viable count , generation time , growth curve (statistics) , plate count , growth rate , optical density , mathematics , biochemistry , statistics , population , genetics , gene , medicine , demography , geometry , sociology , ophthalmology
To assess the overestimation of bacterial cell counts in real-time PCR in relation to stress and growth phase, four different strains ofL. 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°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.