Purpose: The aim of this study was to determine if prolonged cold stress exposure would promote the formation of Lm variants with enhanced cold tolerance and/or resistance to other food-related stresses.
Methods: BHI cultures of 11 Lm strains were grown to stationary phase at 4°C and then maintained at this temperature for one year. Survivors were enumerated monthly and screened for the presence of variants with enhanced cold tolerance. Additionally, the pH and membrane lipid compositions of the long-term cold storage strains were compared to those of the parent strains. Isolates demonstrating enhanced cold tolerance were also subsequently evaluated for their salt (BHI+6% NaCl, 25°C) and acid tolerance (BHI pH 5, 25°C).
Results: Ten Lm variants with enhanced cold tolerance were successfully isolated from one of the 11 strains after 84 days of storage. At 4°C, the parent strain had a maximum cell density of 7.21±0.10 log CFU/ml while the variants reached 8.96±0.02 CFU/ml. Similarly, the parent strain had a maximum growth rate (µmax) of 0.58±0.03 log CFU/ml/h while the variants had a µmax of 0.94±0.02. Two of these variants also exhibited significantly (P<0.05) increased salt and acid tolerance. After one year of storage the pH of the cultures decreased from 7.10±0.03 to 5.68±0.03 and the number of survivors ranged from 5.70 to 7.51 log CFU/ml.
Significance: Our results show that prolonged cold stress exposure either in an environmental or food processing setting can promote the development of Lm variants with enhanced tolerances to food-related stresses.