SINGLE CELL VARIABILITY AND POPULATION DYNAMICS OF LISTERIA MONOCYTOGENES AND SALMONELLA TYPHIMURIUM IN FRESH-CUT SALADS AND THEIR STERILE LIQUID OR SOLIDIFIED EXTRACTS

摘要

Within a homogeneous microbial population, great variability in lag time and probability of growth among individual cells has been observed. In a complex food environment, the variety of stress factors may increase this variability and, subsequently, affect the safety and the quality of the products. We aimed (a) to evaluate the growth variability of Listeria monocytogenes and Salmonella Typhimurium single cells in leafy-vegetable salads, identifying uncertainty sources in risk assessment, and (b) evaluate whether broth-based growth simulations may approximate the average outgrowth of a population from single cells in foods Samples (10 g) of freshly cut lettuce and cabbage were inoculated with 1-4 or 1000 cells/sample and stored at 8°C. Their liquid or solidified sterile extracts were also inoculated with the above cell numbers, in order to evaluate the behavior of pathogens in the presence or absence of indigenous flora. Storage at 8 and 10°C was tested to simulate marginal temperature fluctuations. At least ten independent samples of each case were analyzed per sampling day. Population of 1000 cells increased up to 3 logs with limited variation (SD <1 log CFU/g). Conversely, great variability (0.5-3 log CFU/g increase) in growth of single cells was observed on lettuce. The percentage of lettuce samples exceeding the microbiological criterion of 100 CFU/g for I. monocytogenes varied from 5 to 70%, depending on storage time; however, such variability was not observed in sterile extracts, suggesting that the natural flora of salads is a stress factor. Cabbage did not support growth of Salmonella. Conversely, L. monocytogenes single cells did not increase in sterile cabbage extracts, whereas they increased from 1 to 3.5 logs in cabbage salad, probably due to the commensal behavior of indigenous flora. Salmonella showed no growth at 8°C but increased 4 logs at 10°C, illustrating the impact of boundary conditions on food safety. Results of Monte Carlo simulation of bacterial growth based on broth data markedly deviated from the observed responses in salads, but agreed well with those in sterile extracts of lettuce. Good agreement was also obtained with growth of 103 cells/sample, assuming that the behavior of the population is approximated by the cumulative behavior of individual cells. The probability of growth of individual pathogenic cells on leafy greens is highly affected by the indigenous microflora, the storage temperature and the available nutrients. These parameters should be considered during growth simulations in risk assessment.