The olive knot disease is a bacterial disease that primarily attacs the olive. It is caused by Gram negative flourescent bacterium called Pseudomonas svastanoi. The disease is constantly present in the olive groves in the Istrian region which disturbs their production to a great extent. In the infected tissue, the bacterium causes abnormal multiplication of the meristemic cells, and the appearance of visible tumorous knots on different olive organs (leaves, shoots, branches, petioles). Apart from the fact that the disease causes drying of the branches and olive shoots, it also has an impact on the crop and the quality of the fruit. Since the olive knot disease can cause significant damage in the olive production, the aim of the study was to isolate bacterial colonies from tumorous knots on shoots and olive branches, and to determine their pathogenicity in order to try to prevent further spread of the pathogen. The isolation of bacterial colonies was done through perial decimal dilution and by putting chopped plant tissue pieces on KB substrates. Bacterial isolates were obtained through a serie of subcultivations on nutrient KB substrates, and their basic bacteriological features were tested (Gram reaction and production of fluorescent pigment on KB medium). Pathogenicity of the bacterial isolate that passed earlier tests was verified through the biotest on tobacco (Nicotiana tabacum var. Samsun). The study confirmed the pathogenicity of one bacterial isolate from olive knots and its background in Pseudomonas genus.