Investigations of psychrotrophic microorganisms have been limited even though the dominant environment of the Earth is cold and enzymes with high activities at low temperatures could have commercial uses. We have isolated and characterized three psychrotrophic strains with beta-galactosidase activities. The isolates, B7, D2, and D5, were gram-positive, catalase-positive, obligate aerobes. Cells observed with a scanning electron microscope appeared as rods during the early stages of growth but became coccoid during the stationary phase. An analysis of the amino acid composition of the cell walls demonstrated the presence of lysine as the predominant diamino acid in all three isolates. The cell cycle morphology and cell wall composition suggest that the three isolates are members of the genus Arthrobacter. The beta-galactosidase activities in whole cells were labile when incubated at 40 degrees C and had temperature optima about 20 degrees C below that of the enzyme encoded by the lacZ gene of Escherichia coli. Electrophoresis of extracts from the isolates in nondenaturing polyacrylamide gels detected at least two protein bands that hydrolyzed 5-bromo-4-chloro-3-indolyl-beta-D-galactopyranoside (X-Gal), suggesting the presence of beta-galactosidase isozymes.