Vital fluorescent stains for detection of stress in Pythium ultimum and Rhizoctonia solani challenged with viscosinamide from Pseudomonas fluorescensDR54

Viscosinamide from Pseudomonas fluorescens DR54 exerts a biocontrol effect on the plant pathogenic microfungi Pythium ultimum and Rhizoctonia solani. Evidence for physiological changes in these and four other fungi was obtain ed by vital fluorescent staining with Calcofluor white, Nile red, Syto 13, 3,3'-diheptyloxacarbocyanine, 5-(and-6)-carboxy FDA and 5-(and-6)-carboxy S NARF-1, acetoxymethyl, acetate. Based on studies of the Fungi grown on soli d agar medium and Calcofluor white staining, all isolates responded to visc osinamide treatment by an increased branching, sometimes including hyphal s welling and an increased septation. Further, the staining pattern indicated changes in the esterase activity, intracellular pH and mitochondrial organ ization and activity. Several of the fungi showed a reduced Nile red staini ng indicating a general decrease in hydrophobicity of cellular membranes or cell walls. In contrast, only Pythium spp. were stainable with the nucleic acid stain Syto 13 and showed changes in the nuclei in response to viscosi namide. Finally, it was shown that Calcofluor while, Nile red and Syto 13 c ould also be used to study fungal stress in situ using soil microcosms. The study demonstrated that combinations of vital fluorescent stains are suita ble for detailed studies of fungal stress as induced by, e.g., bacterial an tibiotics. (C) 1999 Federation of European Microbiological Societies. Publi shed by Elsevier Science B.V. All rights reserved.