Bacterial strains with pectinase activity were isolated from Zimbabwe tobacco strips using pectin as carbon source. ARDRA patterns of 16S rDNA combined with 16S rDNA sequence analysis, physiological and biochemical experiments were used to identify the isolated strains. Different conditions including incubating time, growth temperature, initial pH and inoculation quantity for the enzyme production were also studied. The results indicated that bacteria isolated from Zimbabwe tobacco strips with high pectin-degrading ability mainly belong to Bacillus subtilis (genus Bacillus) and Alcaligenes faecalis (genus Alcaligenes). Among these strains, Bacillus subtilis strain T10 possessed the highest pectinase activity (571 U/mg) and polygalacturonate lyase activity (297 U/mg) under its optimum enzyme fermantation conditions, which use 6% (V/V) culture fluid as the inoculum for enzyme production in the initial pH 7.5 fermentation medium at 35 °C for 48?56 h.