Effects of Environmental and Nutritional Factors on Production of the Polyketide Phytotoxin Coronatine by Pseudomonas syringae pv. Glycinea

Abstract

Pseudomonas syringae pv. glycinea PG4180 produces the polyketide phytotoxin coronatine. The effects of environmental, nutritional, and host factors on growth and coronatine production by PG4180 were examined by varying the components of a defined basal medium which contained the following nutrients per liter: glucose (10 g), NH 4 Cl (1 g), MgSO 4 · 7H 2 O (0.2 g), KH 2 PO 4 (4.1 g), K 2 HPO 4 · 3H 2 O (3.6 g), and FeCl 3 (2 μM). Bacterial growth was recorded as dry weight, and coronatine production was measured by high-performance liquid chromatography. Both growth and the quantity of coronatine synthesized were significantly affected by carbon source, nutrient levels (glucose, NH 4 Cl, phosphate, Mg, and SO 4 ), amino acid supplements, and the presence of complex carbon and nitrogen sources. The yield of coronatine generally declined when conditions were varied from those in the basal medium. Coronatine production and growth were not affected when the pH was adjusted from 6.5 to 7.8. Increases in the osmolarity of the basal medium significantly decreased coronatine production without affecting growth. The addition of plant extracts, plant-derived secondary metabolites, or zinc did not affect growth or coronatine production, while the addition of millimolar levels of KNO 3 or micromolar levels of FeCl 3 significantly enhanced coronatine production. The yield of coronatine was maximized after a 7-day incubation at 18°C and 280 rpm. The results of the present study were used to formulate a medium which allowed for enhanced coronatine production in nearly all strains of P. syringae tested. A rapid method for extracting coronatine from small volumes of culture supernatant was also developed.