(J-393) The pili system controls Acinetobacter baumannii wettability, electrophoretic mobilities, adhesion, biofilm formation and drug resistance to Tazobactam

Introduction:: Acinetobacter baumannii is a Gram-negative nosocomial bacterial pathogen that has multidrug resistance (MDR) to antibiotics. A. baumannii claims 34% of all Gram-negative bacilli infections and has a high mortality rate for elderly patients in hospital settings. The bacterial’s ability to cause infections and resist antibiotics is linked to their abilities to form biofilms. A. baumannii’s biofilm formation is facilitated by molecular structures that are present in pili such as the Type IV Pili (T4P) system represented by the major pilin subunits, PilA, PilB and PilD. While their role in biofilm formation and cell adhesion has been acknowledged, the mechanistic means through which they do that are largely missing. As such, the goal of this study was to explore the roles of the T4P system subunits in controlling bacterial wettability, electrophoretic mobilities, biofilm formation, and MDR.

Materials and Methods:: Four A. baumannii 5075 strains including a wildtype and three strains deficient in pilus-forming genes (ΔPilA, ΔPilB, and ΔPilD) were investigated. The strains were cultured at 37 ^oC and 150 rpm in Luria Bertani (LB) broth until late exponential phase of growth. The cells were treated with Tazobactam (TAZ), b-lactamase inhibitor, at a range of concentrations. For all strains, the minimum inhibitory concentration (MIC) was determined. Contact angle measurements of wettability and electrophoretic mobilities measurements as a function of ionic strength and antibiotic exposure were carried out in a goniometer and zeta sizer according to our published protocols¹. Soft-particle analysis of the Derjaguin-Landau-Verwey-Overbeek (DLVO) theory was used to predict the likelihood of A. baumannii to form biofilms on a variety of hospital materials. All measurements were done in triplicates. One-Way ANOVA was used to compare results.


Results, Conclusions, and Discussions:: Results and Discussion: The doubling time for the DPilA mutant was 2/3 smaller than that of all other strains (Table 1). Prior to any exposure to antibiotics, all strains were hydrophilic as evident from contact angles in water that are below 90^o with DPilA mutant being the most hydrophobic. All untreated strains were also negatively charged (Table 1). When compared, the DPilA mutant formed most biofilms (Fig. 1A). After treatment with 32 mg/mL TAZ (MIC) for 10 hours, strains became more negatively charged except for DPilD and more hydrophobic (Table 1). Treatment with TAZ at MIC for 36 hours almost completely inhibited biofilm formation for all strains (Fig. 1B). When results were modeled using SOFT-DLVO, our results indicated that the untreated A. baumannii strains have attractive energies to Teflon, Polyethylene, and Polyvinylchloride; all common hospital materials.

Conclusions: The strains investigated varied in their properties with DPilA being the most hydrophobic and the highest biofilm former; confirming the importance of pilus-forming genes in controlling cellular properties and biofilm formation. Further, van der Waals energies dominated the energy profiles of A. baumannii to common hospital materials; suggesting that design of effective antibiotics should target hydrophobic cell domains. The use of TAZ at MIC was able to suppress biofilms for all strains. Currently, TAZ is not clinically used on its own to treat A. baumannii infections. Our results suggest that TAZ’s use as an antibiotic should be explored.




Acknowledgements (Optional): : The AB strains were supplied through collaboration with Dr. Jieh-Juen Yu at The University of Texas at San Antonio.


References (Optional): : References: 1) A.O. Eskhan and N.I. Abu-Lail, Force-averaging DLVO model predictions of the adhesion strengths quantified for pathogenic Listeria monocytogenes EGDe grown under variable pH stresses, Langmuir, 2020, 36, 30, 8947–8964.