Biological properties and spectrum of activity of tolaasin, a lipodepsipeptide toxin produced by the mushroom pathogen Pseudomonas tolaasii

Abstract The biological properties and spectrum of activity of tolaasin, a lipodepsipeptide toxin produced by the mushroom pathogen Pseudomonas tolaasii , were investigated. An erythrocyte lysis bioassay was developed and used to assay tolaasin activity. Tolaasin-induced haemolysis was dose dependent, maximal between pH 6·0 and 7·0, increased with temperature and was inhibited by the addition of divalent metal ions. Tolaasin production commenced during exponential growth of P. tolaasii and continued into stationary phase. Culture filtrate from wild type P. tolaasii was able to pit mushroom tissue and lyse erythrocytes, whereas culture filtrate from a tolaasin defective mutant had no lytic activity. Ultrastructural studies showed disruption of the Agaricus bisporus plasma membrane and vacuole membranes by both P. tolaasii and tolaasin, but not by a tolaasin defective mutant. Pre-incubation of tolaasin with multilamellar liposomes resulted in loss of lytic activity and osmotic protectants prevented tolaasin-induced haemolysis, demonstrating that tolaasin partitions into membranes, forms pores in erythrocyte membranes and causes lysis by a colloid osmotic mechanism. Tolaasin was phytotoxic when infiltrated into leaves of Nicotiana tabacum and was shown to be active against a range of basidiomycetes and Gram-positive bacteria. Gram-negative bacteria were resistant to tolaasin, but became susceptible when treated concomitantly with sub-minimal inhibitory concentrations of polymyxin B. P. tolaasii was resistant to tolaasin even in the presence of polymyxin B.