Proanthocyanidin Interferes with Intrinsic Antibiotic Resistance Mechanisms of Gram-Negative Bacteria

Antibiotic resistance is spreading at an alarming rate among pathogenic bacteria in both medicine and agriculture. Interfering with the intrinsic resistance mechanisms displayed by pathogenic bacteria has the potential to make antibiotics more effective and decrease the spread of acquired antibiotic resistance. Here, it is demonstrated that cranberry proanthocyanidin (cPAC) prevents the evolution of resistance to tetracycline in Escherichia coli and Pseudomonas aeruginosa, rescues antibiotic efficacy against antibiotic-exposed cells, and represses biofilm formation. It is shown that cPAC has a potentiating effect, both in vitro and in vivo, on a broad range of antibiotic classes against pathogenic E. coli, Proteus mirabilis, and P. aeruginosa. Evidence that cPAC acts by repressing two antibiotic resistance mechanisms, selective membrane permeability and multidrug efflux pumps, is presented. Failure of cPAC to potentiate antibiotics against efflux pump-defective mutants demonstrates that efflux interference is essential for potentiation. The use of cPAC to potentiate antibiotics and mitigate the development of resistance could improve treatment outcomes and help combat the growing threat of antibiotic resistance

Evaluation of anti-biofilm activity of acidic amino acids and synergy with ciprofloxacin on Staphylococcus aureus biofilms

Acidic amino acids, aspartic acid (Asp) and glutamic acid (Glu) can enhance the solubility of many poorly soluble drugs including ciprofloxacin (Cip). One of the mechanisms of resistance within a biofilm is retardation of drug diffusion due to poor penetration across the matrix. To overcome this challenge, this work set to investigate novel counter ion approach with acidic amino acids, which we hypothesised will disrupt the biofilm matrix as well as simultaneously improve drug effectiveness. The anti-biofilm activity of D-Asp and D-Glu was studied on Staphylococcus aureus biofilms. Synergistic effect of combining D-amino acids with Cip was also investigated as a strategy to overcome anti-microbial resistance in these biofilms. Interestingly at equimolar combinations, D-Asp and D-Glu were able to significantly disperse (at 20 mM and 40 mM) established biofilms and inhibit (at 10 mM, 20 mM and 40 mM) new biofilm formation in the absence of an antibiotic. Moreover, our study confirmed L-amino acids also exhibit anti-biofilm activity. The synergistic effect of acidic amino acids with Cip was observed at lower concentration ranges (<40 mM amino acids and <90.54 µM, respectively), which resulted in 96.89% (inhibition) and 97.60% (dispersal) reduction in CFU with exposure to 40 mM amino acids. Confocal imaging indicated that the amino acids disrupt the honeycomb-like extracellular DNA (eDNA) meshwork whilst also preventing its formation

Targeting biofilms using phages and their enzymes

"Available online 10 March 2021"The complex biofilm architecture composed of extracellular polymeric structures (EPS) provides a protective shield to physiologically diverse bacterial cells immersed in its structure. The evolutionary interplay between bacteria and their viruses (phages) forced the latter ones to develop specific strategies to overcome the biofilm defensive barriers and kill sessile cells. Phages are equipped with a wide panel of enzyme-degrading EPS macromolecules which together are powerful weapons to combat biofilms. Antibiofilm performance can be achieved by combining phages or phage-borne enzymes with other antimicrobials such as antibiotics. Nevertheless, a variety of enzymes encoded in phage genomes still need to be explored. To advance in biofilm control strategies we must deepen the understanding of the biofilm biology itself, as well as discover and better exploit the unlimited antibacterial potential of phages.JA was supported by the Portuguese Foundation for Science and Technology (FCT) under the scope of the strategic funding of UIDB/04469/2020 unit. PG was supported by grants from Innovative Training Networks (ITN) Marie Skłodowska-Curie Actions H2020-MSCA-ITN2018. Reference 813439, and PID2019-105311RB-I00 (MICIU/AEI/FEDER, UE, Spain). ZDK was supported by the National Science Centre of Poland (Narodowe Centrum Nauki), grant numbers 2016/21/B/NZ6/01157 and 2017/26/M/NZ1/00233.info:eu-repo/semantics/publishedVersio

Targeted disruption of the extracellular polymeric network of Pseudomonas aeruginosa biofilms by alginate oligosaccharides

Acquisition of a mucoid phenotype by Pseudomonas sp. in the lungs of cystic fibrosis (CF) patients, with subsequent over-production of extracellular polymeric substance (EPS), plays an important role in mediating the persistence of multi-drug resistant (MDR) infections. The ability of a low molecular weight (Mn=3200 g mol-1) alginate oligomer (OligoG CF-5/20) to modify biofilm structure of mucoid Pseudomonas aeruginosa (NH57388A) was studied in vitro using scanning electron microscopy (SEM), confocal laser scanning microscopy (CLSM) with Texas Red (TxRd®)-labelled OligoG and EPS histochemical staining. Structural changes in treated biofilms were quantified using COMSTAT image-analysis software of CLSM z-stack images, and nanoparticle diffusion. Interactions between the oligomers, Ca2+ and DNA were studied using molecular dynamics simulations (MDS), Fourier transform infrared spectroscopy (FTIR) and isothermal titration calorimetry (ITC). Imaging demonstrated that OligoG treatment (&#62;0.5%) inhibited biofilm formation, demonstrating a significant reduction in both biomass and biofilm height (17.8 vs. 5.5 µm; P &#60;0.05). TxRd®-labelled oligomers readily diffused into established (24 h) biofilms. OligoG treatment (≥2%) induced alterations in the EPS of established biofilms; significantly reducing the structural quantities of sugar residues, and extracellular (e)DNA (P &#60;0.05) with a corresponding increase in nanoparticle diffusion (P&#60;0.05) and antibiotic efficacy against established biofilms. ITC demonstrated an absence of rapid complex formation between DNA and OligoG and confirmed the interactions of OligoG with Ca2+ evident in FTIR and MDS. The ability of OligoG to diffuse into biofilms, potentiate antibiotic activity, disrupt DNA-Ca2+-DNA bridges and biofilm EPS matrix highlights its potential for the treatment of biofilm-related infections