Polyamines Are Essential for the Formation of Plague Biofilm

Abstract

We provide the first evidence for a link between polyamines and biofilm levels in Yersinia pestis , the causative agent of plague. Polyamine-deficient mutants of Y. pestis were generated with a single deletion in speA or speC and a double deletion mutant. The genes speA and speC code for the biosynthetic enzymes arginine decarboxylase and ornithine decarboxylase, respectively. The level of the polyamine putrescine compared to the parental speA ⁺ speC ⁺ strain (KIM6+) was depleted progressively, with the highest levels found in the Y. pestis Δ speC mutant (55% reduction), followed by the Δ speA mutant (95% reduction) and the Δ speA Δ speC mutant (>99% reduction). Spermidine, on the other hand, remained constant in the single mutants but was undetected in the double mutant. The growth rates of mutants with single deletions were not altered, while the Δ speA Δ speC mutant grew at 65% of the exponential growth rate of the speA ⁺ speC ⁺ strain. Biofilm levels were assayed by three independent measures: Congo red binding, crystal violet staining, and confocal laser scanning microscopy. The level of biofilm correlated to the level of putrescine as measured by high-performance liquid chromatography-mass spectrometry and as observed in a chemical complementation curve. Complementation of the Δ speA Δ speC mutant with speA showed nearly full recovery of biofilm to levels observed in the speA ⁺ speC ⁺ strain. Chemical complementation of the double mutant and recovery of the biofilm defect were only observed with the polyamine putrescine.