Differentiation of Pseudomonads by Amplification of Metabolic Profiles

Abstract

Cultures of four species of Pseudomonas grown in a complex medium were shifted down to a minimal medium for assaying aminopeptidases and metabolic end products. Cultural conditions were further modified to include both agitation and stationary conditions. Growth rates were determined for each condition to obtain maximal cell yield. Cells were then harvested for intracellular aminopeptidase assay by fluorometric analysis of enzyme activity, and a gas-liquid chromatography analysis was made for extracellular metabolic end products. A high reproducibility was obtained from the aminopeptidase profiles of strains of Pseudomonas fluorescens, P. putida, P. stutzeri, and P. pickettii, including a control consisting of a blind test of previous isolates. The shift down to the minimal medium increased enzyme activity in each case and induced characteristic new peaks in most instances. Gas-liquid chromatography analysis was concentrated on short-chain volatile fatty acids no longer than seven carbons. Butyl ester derivatives were prepared to enhance separation and increase percent recovery of these volatile components. The resulting change in profiles under different physiological conditions indicates a new method for the possible identification of Pseudomonas species.