Abstract
Concerns about acquisition of antibiotic resistance have led to increasing demand for new antimicrobial therapies. OligoG CF-5/20 is an alginate oligosaccharide previously shown to have antimicrobial and antibiotic potentiating activity. We investigated the structural modification of the bacterial cell wall by OligoG CF-5/20 and its effect on membrane permeability. Binding of OligoG CF-5/20 to the bacterial cell surface was demonstrated in Gram-negative bacteria. Permeability assays revealed that OligoG CF-5/20 had virtually no membrane-perturbing effects. Lipopolysaccharide (LPS) surface charge and aggregation were unaltered in the presence of OligoG CF-5/20. Small angle neutron scattering and circular dichroism spectroscopy showed no substantial change to the structure of LPS in the presence of OligoG CF-5/20, however, isothermal titration calorimetry demonstrated a weak calcium-mediated interaction. Metabolomic analysis confirmed no change in cellular metabolic response to a range of osmolytes when treated with OligoG CF-5/20. This data shows that, although weak interactions occur between LPS and OligoG CF-5/20 in the presence of calcium, the antimicrobial effects of OligoG CF-5/20 are not related to the induction of structural alterations in the LPS or cell permeability. These results suggest a novel mechanism of action that may avoid the common route in acquisition of resistance via LPS structural modification.
Highlights
Biofilm growth of P. aeruginosa[3,8,9], and that this effect was independent of an interaction with the P. aeruginosa mexAB-oprM efflux pump system[3], this study sought to investigate whether OligoG cystic fibrosis (CF)-5/20 exerts its antibiotic potentiation effects via direct interaction with the bacterial cell
This study focused on the nanoscale interaction of OligoG CF-5/20 with the Gram-negative cell surface, following strong, irreversible binding to the cell wall after centrifugation
OligoG CF-5/20 has previously been shown to remain bound to the pseudomonal cell surface, leading to cellular aggregation, even following exposure to hydrodynamic shear[8]
Summary
Biofilm growth of P. aeruginosa[3,8,9], and that this effect was independent of an interaction with the P. aeruginosa mexAB-oprM efflux pump system[3], this study sought to investigate whether OligoG CF-5/20 exerts its antibiotic potentiation effects (up to 512-fold) via direct interaction with the bacterial cell. Cell-surface oligosaccharides such as the hydrophilic O-antigen component of LPS in Gram-negative bacteria[16] play a role in facilitating biofilm attachment. The outer membrane of Gram-negative bacteria is selectively resistant to noxious agents due to its effective permeability barrier function (enabling hydrophobic drugs to diffuse across the lipid bilayer, whilst small hydrophilic drugs use the porins to gain access to the cell). Both Gram-positive and Gram-negative bacteria have an overall negative electrostatic surface charge. Isothermal titration calorimetry has previously been employed to elucidate the mechanisms by which novel antimicrobials interact with the cell surface target, LPS35,36
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