Pseudomonas aeruginosa, a Gram-negative bacterium, is a ubiquitous opportunistic pathogen able to form biofilms that aid its survival in a variety of environments. In previous work, we demonstrated that the specific interaction between bacterioferritin (BfrB), the main iron storage protein in P. aeruginosa, and its associated ferredoxin (Bfd) is required to establish a dynamic equilibrium between Fe3+ compartmentalized in BfrB and cytosolic free iron (Fe2+). This equilibrium buffers the concentration of intracellular iron and enables regulation by the ferric uptake regulator (Fur). In the present study we investigated the implications of perturbing the BfrB:Bfd interaction on the formation and integrity of P. aeruginosa biofilms on abiotic surfaces and at the air-liquid interface (pellicles). We compared the establishment and stability of biofilms formed by the wild type strain with mutant strains where the BfrB:Bfd interaction was blocked, either by in frame deletion of the bfd gene or in a variant with the bfrB allele encoding BfrB (L68A/E81A). The effects of the mutations were evaluated by a time-dependent microscopic inspection of the pellicles and surface-attached biofilms, and by quantitative determination of (i) total biofilm mass, (ii) concentration of iron and pyoverdine in the spent culture media, (iii) intracellular iron levels, (iv) production of quorum sensing molecules and rhamnolipids, and (iv) swarming motility. Our results revealed that inhibiting the BfrB:Bfd interaction triggers an iron starvation response that weakens the biofilm structure, causing early cell detachment and concomitant dissolution. We also showed that blockade of the BfrB:Bfd complex causes a decrease in the levels of free cytosolic iron in planktonic and pellicle-embeded cells, which results in exacerbated production of pyoverdine, quorum sensing molecules, and rhamnolipids, and increased bacterial motility. These findings suggest that a rational approach to dysregulating bacterial iron homeostasis is a promising strategy to combat P. aeruginosa biofilms.

Disruption of iron homeostasis impairs Pseudomonas aeruginosa biofilms

2019 Fall National Meeting

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