Chitinolytic activity of Pseudomonas fluorescens isolates from barley and sugar beet rhizosphere

Twelve isolates of Pseudomonas fluorescens, isolated from barley and sugar beet rhizosphere as antagonists towards the plant pathogenic microfungi Rhizoctonia solani and Pythium ultimum, showed chitinolytic activity in batch cultures when grown in media without exogenous chitin. Enzyme tests in cultures demonstrated a complete array of chitinolytic enzymes. Endochitinase and chitobiosidase activities seemed to be tightly coupled in the isolates, while there was no relationship to N-acetyl-glucosaminidase activity. Endochitinase activity, showing hydrolysis of chitin polymers, was found to be extracellular in all isolates, although the most active isolates also retained a cell-bound fraction. Isoelectric focusing gel electrophoresis of supernatants containing extracellular enzyme activity showed that all isolates produced one native endochitinase in logarithmic phase. This enzyme was subsequently modified into several isozymes by extracellular processing as the cultures aged in stationary phase. The 12 isolates could be grouped into seven isozymic patterns. Detailed studies of three selected isolates showed that extracellular release of endochitinase activity also took place in stationary phase. The results, however, indicated that in stationary phase regulation of both the overall production of native enzyme and the subsequent formation of isozymes were different among the P. fluorescens isolates.