Staff Publications

Staff Publications

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    'Staff publications' is the digital repository of Wageningen University & Research

    'Staff publications' contains references to publications authored by Wageningen University staff from 1976 onward.

    Publications authored by the staff of the Research Institutes are available from 1995 onwards.

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Record number 318830
Title Development of a flow cytometric method to analyze subpopulations of bacteria in probiotic products and dairy starters
Author(s) Bunthof, C.J.; Abee, T.
Source Applied and Environmental Microbiology 68 (2002). - ISSN 0099-2240 - p. 2934 - 2942.
DOI http://dx.doi.org/10.1128/AEM.68.6.2934-2942.2002
Department(s) Food Microbiology Laboratory
VLAG
Publication type Refereed Article in a scientific journal
Publication year 2002
Abstract Flow cytometry (FCM) is a rapid and sensitive technique that can determine cell numbers and measure various physiological characteristics of individual cells by using appropriate fluorescent probes. Previously, we developed an FCM assay with the viability probes carboxyfluorescein diacetate (cFDA) and TOTO-1 {1'-(4,4,7,7-tetramethyl-4,7-diazaundecamethylene)-bis-4-[3-methyl-2,3dihydro(benzo-1,3-oxazole)-2-methylidene]-1-(3'-trimethylammoniumpropyl)-pyridinium tetraiodide} for (stressed) lactic acid bacteria (C. J. Bunthof, K. Bloemen, P. Breeuwer, F. M. Rombouts, and T. Abee, Appl. Environ. Microbiol. 67:2326-2335, 2001). cFDA stains intact cells with enzymatic activity, and TOTO-1 stains membrane-permeabilized cells. Here we used this assay to study the viability of bacterial suspensions in milk, dairy fermentation starters, and probiotic products. To facilitate FCM analysis of bacteria in milk, a commercially available milk-clearing solution was used. The procedure was optimized to increase the signal-to-noise ratio. FCM enumerations were accurate down to a concentration of 105 cells ml-1. The level of retrieval of Lactobacillus plantarum WCFS 1 suspended in milk was high, and viability was not affected by the procedure. The plate counts for cleared samples of untreated cell suspensions were nearly as high as the total FCM counts, and the correlation was strong (r > 0.99). In dairy fermentation starters and in probiotic products the FCM total cell counts were substantially higher than the numbers of CFU. Three functional populations could be distinguished: culturable cells, cells that are intact and metabolically active but not culturable, and permeabilized cells. The proportions of the populations differed in the products tested. This FCM method provides tools to assess the functionality of different populations in fermentation starters and probiotic products.
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