The method is based on that of Richards in which contaminants on membrane filters are incubated on nutrient medium and the microcolonies stained and observed. Beer samples (100 to 1000 ml) were filtered through 47 mm, 0.45 um, white grid membrane filters, which were then rinsed with an equal volume of sterile distilled water, transferred to Universal Beer Agar (UBA) and incubated at 25 to 30 degrees C for 16 to 48 h (aerobic) or 48 to 96 h (anaerobic). After incubation, the filters were placed on absorbent prefilters, heated for 5 min to 105 degrees C and stained by immersion in Loeffler's methylene blue, aqueous 0.5% safranin, Gram's safranin or 0.05% acridine orange in citrate buffer (pH 3.8). The stained filters were dried at 50 to 60 degrees C on absorbent paper and clarified on glass slides. Microcolonies were then counted directly using a stereo microscope (18 to 36 times magnification). As a standard comparison method, the microorganisms trapped on black grid membrane filters were incubated on UBA for 3 days (aerobic) or 7 days (anaerobic) and microcolonies were observed visually. The new method gave colony counts closely similar to those of the standard method, it was more convenient than other accelerated methods and gave results shown to be statistically as accurate as the membrane filtration method currently in use. Incubation periods could be reduced from 3 to 1 day (aerobic) and 7 to 4 days (anaerobic).
Keywords: aerobic anaerobic beer contamination detection filtration membrane microorganism stability