The stainat a final concentration of 10 lgml)1was added to 1 mlof second-diluted natural whey starter, previously washedtwice with sterilized distilled water and then incubatedfor 30 min at room temperature in the dark. After incu-bation, the samples were filtered on a black polycarbonatemembrane (Millipore Corp., Billerica, MA, USA); themembrane was air-dried and mounted on a glass slide inCitifluor solution (Citifluor Ltd, London, UK). The bacte-ria on DAPI-stained membranes were enumerated bycounting the total number of blue-fluorescing bacteria.The number of bacteria was estimated from counts of 20microscopic fields (at ·1000) and calculated as follows:N ¼C Aa V Dwhere N is the number of cells per ml; C is the number ofcells per observation field; A is the filtration area (mm2);a is the observation field area (mm2); V is the volume offiltered sample (ml) and D is the dilution factor.