Selective inhibition of novel biofilm-associated proteases in Acinetobacter baumannii

: a potential antibiotic adjuvant approach

Abstract

Bacterial biofilms are complex microbial communities that are either attached to surfaces and embedded within a self-produced extracellular matrix. This mode of bacterial growth represents the predominant phenotype in nature, particularly, in chronic infections. Biofilms are very different from their planktonic correspondents and are distinguished by altered growth rates and elevated tolerance to antimicrobials and normal immune clearance, and often require minimum cidal concentrations of antibiotics up to 1000 times that of their equivalent planktonic or freely suspended bacteria. One of the most important biofilm-forming pathogens is Acinetobacter baumannii, an opportunistic, often hospital-acquired Gram-negative bacilli. A. baumannii biofilms exhibit an enhanced antibiotic tolerance profile and pose challenges to existing therapeutic options. Biofilm formation is an effective survival strategy in A. baumannii and results in biofilm-related contamination of medical devices, it is widely distributed in clinical settings especially Intensive Care Units (ICUs) and surgical wards. Recent research has shown that a number of proteases can be targeted as virulence factors in the process of bacterial biofilm formation and that selective inhibition may lead to biofilm reduction and increased sensitivity to conventional antimicrobials. The inhibition of bacterial proteases is emerging as a potential therapeutic target, which may facilitate the dissemination of bacteria from the biofilm. In this research, we studied the formation of A. baumannii biofilm under conventional environments and under the effect of different cysteine protease inhibitors. We utilized the Microtiter Dish Biofilm assay for the detection of biofilm biomass reduction using lead inhibitors in the presence or absence of conventional antibiotics, to examine the role of protease inhibitors as an antibiotic adjuvant and whether synergy or enhancement of activity exists by using protease inhibitor- antibiotic combinations, which may enhance the sensitivity of A. baumannii to antibiotics. This work developed a high throughput screening of 26 cysteine protease inhibitors by the Williams laboratory (QUB) and revealed that five compounds significantly reduced biofilm formation in A. baumannii. Then, we tested 5 commercially available antibiotics (Gentamicin, Imipenem, Meropenem Tobramycin, Tigecycline) on A. baumannii bacteria and determined the MIC and MBEC value for each. After obtaining the MBEC values, we started running synergy studies by treating the bacteria with combination of protease inhibitors at a fixed concentration (lOmM), and antibiotics with the lowermost MBEC values: Tigecycline and Tobramycin antibiotics, at a range of concentrations. The empirical findings showed that a significant reduction of A. baumannii biofilms reported only by the presence of 15.6 ;

Date of Award	Dec 2019
Original language	English
Awarding Institution	Queen's University Belfast
Supervisor	Brendan Gilmore (Supervisor)