A COMPARISON OF THREE STAINING METHODS FOR DIRECT MICROSCOPIC COUNTING OF BACTERIA IN MILK

Abstract

The Levowitz-Weber (LW) stain was compared collaboratively with two new stains, the pH 4 toluidine blue (pH 4 TBO), and the periodic acid-bisulfite-toluidine blue (PST), for direct microscopic counting of bacteria in nonfat dry milk (NDM). Mean counts for 10 samples (18 determinations each) were for the PST, pH 4 TBO, and LW stains, respectively, 89.1, 65.1, and 64.8 million/g. The mean count for the PST stain was significantly higher than the other two at the 5% level. The higher counts with the PST stain were as expected since the PST stain is designed to improve staining of heat plasmolyzed cells such as are found in NDM. Interlaboratory variation in counts was high. This variation was independent of the stain used or the laboratory preparing the stained smear. More work is needed on techniques for direct microscopic counting of bacteria to reduce interlaboratory variability.