Role of the biosurfactant viscosin in broccoli head rot caused by a pectolytic strain of Pseudomonas fluorescens

Abstract

The lipopeptidic biosurfactant viscosin was examined as a pathogenicity factor of a pectolytic strain of Pseudomonas fluorescens that causes broccoli head rot. The critical micellar concentration (CMC) of viscosin was 4 𝛍g/mL in a 2 mM phosphate buffer (pH 7.0), and the surface tension was reduced from 71 mN/m to 25 mN/m. When broccoli florets were immersed in viscosin solutions of increasing concentration, the tissues became wetted at 10 𝛍g/mL and electrolytes were induced to leak at a concentration between 10 and 25 𝛍g/mL. Erythrocytes were lysed at concentrations of 10 𝛍g/mL and above. Since membrane effects occurred above the CMC, it appears that viscosin does not act as a membrane toxin, but rather as a nonspecific detergent. A viscosin deficient mutant, induced by Tn5 mutagenesis, caused decay of wounded florets only, but the decay failed to spread to adjacent nonwounded florets as had occurred with a wild strain. When the mutant strain (1 x 10⁷ cfu/mL) was incubated with viscosin (25 𝛍g/mL), it was able to induce electrolyte leakage and decay of broccoli florets. Leakage and decay failed to occur when florets were immersed in an extract of pectolytic enzymes alone, but rapid leakage and decay occurred when viscosin was added (25 𝛍g/mL). The mutant strain was unable to colonize the surface of intact broccoli florets, as shown by scanning electron microscope studies, but in the presence of viscosin (20 𝛍g/mL), it was able to colonize surfaces and penetrate stomata, Epicuticular wax was substantially altered on florets immersed in a viscosin solution of 20 𝛍g/mL and appeared to dissolve with 200 𝛍g/mL viscosin. Taken together, these results show that viscosin, as a powerful surfactant, is an important pathogenicity factor of this pectolytic strain of P. fluorescens that causes decay of the difficult-to-wet waxy surfaces of broccoli heads.