Chromosomal DNA of nine strains of the Lactobacillus acidophilus (Hansen and Mocquot) group in which heterogeneity had previously been revealed by biochemical characteristics and DNA-DNA hybridization studies were further investigated by restriction analysis, Southern hybridization, conventional and pulsed-field gel electrophoresis (PFGE) analyses. Industrial strains were characterized and identified as Lact. acidophilus. For this species, the number of rRNA gene clusters was estimated to be at least four from analyses of rRNA gene restriction patterns. The chromosome size of type-strains of Lact. acidophilus and Lact. gasseri was estimated from PFGE analysis to be 1.85 and 2.02 Mb respectively and Lact. gasseri strains were found to contain large-sized linear plasmids.