Identification of mechanisms modulating chlorhexidine and octenidine susceptibility in Proteus mirabilis.

AIMS: We aimed to identify mechanisms underlying the tolerance of Proteus mirabilis-a common cause of catheter associated urinary tract infection-to the clinically used biocides chlorhexidine (CHD) and octenidine (OCT). METHODS AND RESULTS: We adapted three clinical isolates to grow at concentrations of 512 µg ml-1 CHD and 128 µg ml-1 OCT. Genetic characterization and complementation studies revealed mutations inactivating the smvR repressor and increasing smvA efflux expression were associated with adaptation to both biocides. Mutations in mipA (encoding the MltA interacting protein) were less prevalent than smvR mutations and only identified in CHD adapted populations. Mutations in the rppA response regulator were exclusive to one adapted isolate and were linked with reduced polymyxin B susceptibility and a predicted gain of function after biocide adaptation. Biocide adaptation had no impact on crystalline biofilm formation. CONCLUSIONS: SmvR inactivation is a key mechanism in both CHD and OCT tolerance. MipA inactivation alone confers moderate protection against CHD, and rppA showed no direct role in either CHD or OCT susceptibility.

An In Vitro Bladder Model for Studying Catheter-Associated Urinary Tract Infection and Associated Analysis of Biofilms.

Urethral catheters are among the most widely used medical devices, applied to manage a wide range of conditions in hospital, community, and care home settings. In long-term catheterized individuals, infection with Proteus mirabilis frequently complicates the care of patients owing to formation of extensive crystalline biofilms. Here we describe the use of an in vitro bladder model of the catheterized urinary tract and associated analyses to study P. mirabilis crystalline biofilm formation. The model originally described by Stickler et al. (1999, 310:494-501, Methods Enzymol) replicates a complete sterile closed drainage system as used in clinical practice, and permits formation of biofilms directly on catheters under conditions representative of those encountered in vivo. Models may be used to replicate either established infection or early stage colonization, and we describe a range of associated methods for quantification and visualization of biofilms formed on catheters. These methods are also easily adapted to study catheter-associated biofilm formation by other urinary tract pathogens.

Fluoxetine and thioridazine inhibit efflux and attenuate crystalline biofilm formation by Proteus mirabilis.

Proteus mirabilis forms extensive crystalline biofilms on indwelling urethral catheters that block urine flow and lead to serious clinical complications. The Bcr/CflA efflux system has previously been identified as important for development of P. mirabilis crystalline biofilms, highlighting the potential for efflux pump inhibitors (EPIs) to control catheter blockage. Here we evaluate the potential for drugs already used in human medicine (fluoxetine and thioridazine) to act as EPIs in P. mirabilis, and control crystalline biofilm formation. Both fluoxetine and thioridazine inhibited efflux in P. mirabilis, and molecular modelling predicted both drugs interact strongly with the biofilm-associated Bcr/CflA efflux system. Both EPIs were also found to significantly reduce the rate of P. mirabilis crystalline biofilm formation on catheters, and increase the time taken for catheters to block. Swimming and swarming motilies in P. mirabilis were also significantly reduced by both EPIs. The impact of these drugs on catheter biofilm formation by other uropathogens (Escherichia coli, Pseudomonas aeruginosa) was also explored, and thioridazine was shown to also inhibit biofilm formation in these species. Therefore, repurposing of existing drugs with EPI activity could be a promising approach to control catheter blockage, or biofilm formation on other medical devices.