Quantifying the Impact of Biological and Experimental Variability Near the Growth Boundaries on the Stochastic Responses of Growth, Gene Transcription and Acid Resistance of Listeria monocytogenes

AbstractHereby, we showed that the experimental and biological variability in culture preparation had little effect on the stochastic outcome of: (i) growth, (ii) relative transcription of stress- (gad2, sigB) and virulence- (prfA) associated genes and (iii) subsequent acid resistance of Listeria monocytogenes, across growth/ no growth boundaries regarding combinations of pH (4.8-7.2) (HCl) and NaCl (0-8% w/v) at 7oC. Variability of bacterial response, as described by the coefficient of variation (CV) and root mean square error (RMSE) was affected mainly by the previously pH and NaCl conditions that the pathogen had experienced. High biological variability regarding growth potential of the pathogen, was observed in conditions across growth boundaries (at 7oC), such as pH 5.5-6.4 and NaCl 2-8% w/v, as manifested by the highly ranged growth parameters (CVareas 18.3% - 49%, RMSEareas 8.7-13.9, RMSElag time 9.3-31.2), while acid resistance (pH 2.0, HCl, 37 °C) was highly variable when pathogen habituated pH (5.0-5.2) and NaCl (2% w/v) resulting in average D1 (3-6min) and D2 (14-16min) of high variability (CVD1 28-35%; CVD2 35-60%). Moreover at the same conditions the highest upregulation of gad2 was observed with high biological variability of RMSEgad2 2.8, while relative transcription levels of pfrA ranged from 0.60 to 4.22, indicating the potential risk derived from the stochastic bacterial response (up- or down- regulation) regarding induction of virulence mechanisms in growth boundary conditions