CONICET | Buscador de Institutos y Recursos Humanos

Fusaric acid (FA) is a fungal metabolite produced by severalspecies of Fusarium that contributes to wilt and rot diseases of plants. Manyfluorescent pseudomonads can prevent wilt diseases caused by these fungi. This study was undertaken to evaluate the effect of FA on P.protegens Pf-‐5 and to elucidate the mechanisms that enable the bacterium tosurvive in the presence of the mycotoxin. P. protegens strains were cultured at 30oC in minimalE2 medium supplemented with 0.6 % (wt/vol) glucose. When required cultureswere amended with different concentrations of FA. Swimming and swarmingmotility were evaluated in media solidified with 0.3% or 0.6% (wt/vol) Bacto-‐Agar, respectively. Pyoverdine in the supernatants of cultures grown for 16 h at1000 rpm in 96-‐well microtiter plates was quantified by measuring fluorescenceemission at 485 nm after excitation at 420 nm. Proteins of the cytosolic andmembrane fractions of cultures grown with or without 2 mM FA were analyzedby two or one dimensional gel electrophoresis, respectively. Proteins expressedat visibly different levels in cells exposed or not to FA were identified by peptidefingerprint analysis using a MALDI-‐TOF mass spectrometry system. Deletionmutants of P. protegens were constructed by gene replacement using thesuicide plasmid pNPTS138-‐R6KT. UV absorption spectra of FA alone and FAincubated with increasing amounts of Fe2+, Fe3+, Cu2+, Mn2+ and Zn2+ wereanalyzed over the wavelength interval between 200 and 300 nm.The results confirm that FA negatively affects growth and motility of P.protegens. Moreover, a notable increase in secretion of the siderophorepyoverdine was observed when P. protegens was grown in the presence of FA. Concomitantly, levels of enzymes involved in the biosynthesis of pyoverdineand enantio-‐pyochelin, the second siderophore encoded by P. protegens, increased markedly. Moreover, whereas P. protegens mutant strains defective in either pyoverdine or enantio-‐pyochelin synthesis still displayed resistance to FA,double mutants lacking siderophores were highly sensitive to the mycotoxin.This effect was not observed when the double mutant was grown underconditions of iron excess. Spectrophotometric titrations revealed that FA bindsnot only Fe2+ and Fe3+, but also Zn2+, Mn2+ and Cu2+, with high affinity.The mycotoxin FA exerts a toxic effect on P. protegens Pf‐5 mainly by sequestering iron. As a result of iron chelation by FA, siderophoresynthesis is induced. At all events, the ability of P. protegens Pf‐5 to producesiderophores markedly increases its resistance to FA, and thus confers a clearadaptive advantage in soils in which FA producers are present.