Mannitol Enhances Antibiotic Sensitivity of Persister Bacteria in Pseudomonas aeruginosa Biofilms

Abstract

The failure of antibiotic therapies to clear Pseudomonas aeruginosa lung infection, the key mortality factor for cystic fibrosis (CF) patients, is partly attributed to the high tolerance of P. aeruginosa biofilms. Mannitol has previously been found to restore aminoglycoside sensitivity in Escherichia coli by generating a proton-motive force (PMF), suggesting a potential new strategy to improve antibiotic therapy and reduce disease progression in CF. Here, we used the commonly prescribed aminoglycoside tobramycin to select for P. aeruginosa persister cells during biofilm growth. Incubation with mannitol (10–40 mM) increased tobramycin sensitivity of persister cells up to 1,000-fold. Addition of mannitol to pre-grown biofilms was able to revert the persister phenotype and improve the efficacy of tobramycin. This effect was blocked by the addition of a PMF inhibitor or in a P. aeruginosa mutant strain unable to metabolise mannitol. Addition of glucose and NaCl at high osmolarity also improved the efficacy of tobramycin although to a lesser extent compared to mannitol. Therefore, the primary effect of mannitol in reverting biofilm associated persister cells appears to be an active, physiological response, associated with a minor contribution of osmotic stress. Mannitol was tested against clinically relevant strains, showing that biofilms containing a subpopulation of persister cells are better killed in the presence of mannitol, but a clinical strain with a high resistance to tobramycin was not affected by mannitol. Overall, these results suggest that in addition to improvements in lung function by facilitating mucus clearance in CF, mannitol also affects antibiotic sensitivity in biofilms and does so through an active, physiological response.

The bactericidal activity of β-lactam antibiotics is increased by metabolizable sugar species

“…Our findings that survival during gentamicin exposure was improved by the presence of sugar metabolites is in sharp contrast to recent reports demonstrating the ability of b-Lactam potentiation by sugar metabolites specific carbohydrates to restore the aminoglycoside susceptibility of non-or slowly growing persister cells in planktonic cultures and in biofilms (Allison et al, 2011;Barraud et al, 2013). The authors attributed the effect to the generation of a proton motive force across the cytoplasmic membrane, which facilitates the uptake of aminoglycoside antibiotics in bacteria with otherwise low metabolic activity.…”

Lack of intracellular trehalose affects formation of Escherichia coli persister cells

“…Therefore, trehalose, which increases Amp-and Ofxtolerance, affects persisters most probably by a different mechanism. There is a growing interest in factors that sensitize persister bacteria to antibiotics and/or facilitate eradication of biofilms (Allison et al, 2011;Barraud et al, 2013;Conlon et al, 2013;Lebeaux et al, 2014;Kim et al, 2011;Leszczyń ska et al, 2013). We believe that the results presented here may help to find new strategies for more efficient antibacterial therapies.…”

Contribution of Stress Responses to Antibiotic Tolerance in Pseudomonas aeruginosa Biofilms

“…This hypothesis also predicts that genotype will be immaterial, as the pattern of gene expression in a cell is of no significance if the antibiotic never enters the cell in the first place. If reduced membrane potential is responsible for tobramycin tolerance, it could be possible to enhance antibiotic efficacy by adding a substrate that restores the proton motive force and membrane potential (79,80).…”