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Flow cytometric assessment of the protectants for enhanced in vitro survival of probiotic lactic acid bacteria through simulated human gastro-intestinal stresses

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Abstract

The aim of this study was to apply flow cytometric (FCM) analysis to assess the use of sucrose and lecithin vesicles for the protection of probiotic lactic acid bacteria in response to the challenge of gastric acidity and bile salts. FCM analysis in combination with fluorescent probes carboxyfluorescein (cF) and propidium iodide was used to reveal the physiological heterogeneity in the stressed bacteria population. Three subpopulations (intact, stressed, and damaged) were differentiated by FCM in all six examined strains. Significant changes were observed in the presence of the selected protectants. The addition of 20 mM sucrose in the simulated gastric fluid substantially increased the number of intact cells over 20 folds and reduced the damaged subpopulation by half. The presence of 2 % (w/v) lecithin vesicles was shown to protect 50 % more intact cells from the challenge of bile salts. The improved survival as evaluated by FCM analysis was further assessed for the proliferation capacity by sorting a number of cells from each subpopulation on nutrient agar plate. The result confirmed conformity between the proliferation-based cultivability and the probe-indicated viability in the samples of the intact and the damaged subpopulations. However, it also revealed the complexities of the stressed (injured) subpopulation. In conclusion, FCM analysis confirmed that the selected protectants could improve the survival of the probiotic strains in the simulated GI environments. The FCM analysis also proved to be a useful analytical tool for the probiotics research.

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Acknowledgments

This work was supported by Technology for Industry Fellowship (TIF) New Zealand grant DPAC0702 and by the Drapac Trust, New Zealand. We thank the team of Food Safety & Preservation—Plant and Food Research, the team of AnQual Lab—the University of Auckland, and Ms. Vivien Zhang for their kind help and emotional support throughout the project. We are also grateful for Mrs. Amanda Collocott’s kind help on critical reading of the manuscript.

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Authors and Affiliations

  1. School of Pharmacy, University of Auckland, Auckland, New Zealand

    Song Chen & Sanjay Garg

  2. AsureQuality, Auckland, New Zealand

    Yu Cao

  3. Department of Nutrition, School of Medical Sciences, University of Auckland, Auckland, New Zealand

    Lynnette R. Ferguson

  4. Bioactives Research New Zealand, Auckland, New Zealand

    Quan Shu

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  1. Song Chen
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  2. Yu Cao
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  4. Quan Shu
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  5. Sanjay Garg
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Correspondence to Song Chen.

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Chen, S., Cao, Y., Ferguson, L.R. et al. Flow cytometric assessment of the protectants for enhanced in vitro survival of probiotic lactic acid bacteria through simulated human gastro-intestinal stresses. Appl Microbiol Biotechnol 95, 345–356 (2012). https://doi.org/10.1007/s00253-012-4030-3

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