Assessment of Antibiofilm Potencies of Nervonic and Oleic Acid against Acinetobacter baumannii Using In Vitro and Computational Approaches

Abstract

Acinetobacter baumannii is a nosocomial pathogen, and its biofilms are tolerant to desiccation, nutrient starvation, and antimicrobial treatment on biotic and abiotic surfaces, tissues, and medical devices. Biofilm formation by A. baumannii is triggered by a quorum sensing cascade, and we hypothesized that fatty acids might inhibit its biofilm formation by interfering with quorum sensing. Initially, we investigated the antibiofilm activities of 24 fatty acids against A. baumannii ATCC 17978 and two clinical isolates. Among these fatty acids, two unsaturated fatty acids, nervonic and oleic acid, at 20 μg/mL significantly inhibited A. baumannii biofilm formation without affecting its planktonic cell growth (MICs were >500 μg/mL) and markedly decreased the motility of A. baumannii but had no toxic effect on the nematode Caenorhabditis elegans. Interestingly, molecular dynamic simulations showed that both fatty acids bind to the quorum sensing acyl homoserine lactone synthase (AbaI), and decent conformational stabilities of interactions between the fatty acids and AbaI were exhibited. Our results demonstrate that nervonic and oleic acid inhibit biofilm formation by A. baumannii strains and may be used as lead molecules for the control of persistent A. baumannii infections.

Keywords: AbaI; Acinetobacter baumannii; antibiofilm agents; biofilm formation; computational studies; fatty acid; nervonic acid; quorum sensing; virulence.

Figure 1

Impacts of various fatty acids on biofilm formation by the three A. baumannii strains in the presence of fatty acids at 100 μg/mL (A), The structures and antibiofilm activities of the selected fatty acids against the three strains of A. baumannii were quantified against A. baumannii ATCC 17978 (B), A. baumannii BAA 1709 (C), and A. baumannii A 550 (D). Error bars indicate standard deviations. *, p < 0.05 versus non-treated controls.

Figure 2

A. baumannii ATCC 17978 cell growth was investigated in the presence of nervonic, oleic, or myristoleic acid at 20 μg/mL (A), A. baumannii ATCC 17978 motility was assessed in the presence of fatty acid at 20 μg/mL after incubation for 24 h (B), the cytotoxicity of fatty acid was evaluated against C. elegans (C), A. baumannii ATCC 17978 biofilm formations on nylon membrane were observed by SEM in the presence of fatty acid at 20 μg/mL. Red arrows indicate dents in cells and scale bars represent 1 µm (D). Error bars indicate standard deviations. *, p < 0.05 versus non-treated controls.

Figure 3

The putative predicted active pocket of ligand binding on the rigid surface of AbaI is shown in red (A), stable conformation of the ligand–receptor complex (B), and 25 molecular docking runs of nervonic acid with AbaI receptor (C).

Figure 4

Interactions of ligands with AbaI receptor protein with nervonic (A), oleic (B), and myristoleic acid (C).

Figure 5

Ligand–AbaI complex stability studies to reveal the interactions. Binding energies (A–C) and RMSD of nervonic, oleic, and myristoleic acid, respectively, as determined by MD simulation of YASARA (D–F). Ligand–AbaI complex comparative binding positions of nervonic (G), oleic (H), and myristoleic acid (I) with AbaI as displayed before and after 100.3 ns MD simulations.

Figure 6

Plausible mechanism for the inhibition of A. baumannii quorum sensing by nervonic acid.