Dataset Information

Elucidating the mechanism by which synthetic helper peptides sensitize Pseudomonas aeruginosa to multiple antibiotics.

ABSTRACT: The emergence and rapid spread of multi-drug resistant (MDR) bacteria pose a serious threat to the global healthcare. There is an urgent need for new antibacterial substances or new treatment strategies to deal with the infections by MDR bacterial pathogens, especially the Gram-negative pathogens. In this study, we show that a number of synthetic cationic peptides display strong synergistic antimicrobial effects with multiple antibiotics against the Gram-negative pathogen Pseudomonas aeruginosa. We found that an all-D amino acid containing peptide called D-11 increases membrane permeability by attaching to LPS and membrane phospholipids, thereby facilitating the uptake of antibiotics. Subsequently, the peptide can dissipate the proton motive force (PMF) (reducing ATP production and inhibiting the activity of efflux pumps), impairs the respiration chain, promotes the production of reactive oxygen species (ROS) in bacterial cells and induces intracellular antibiotics accumulation, ultimately resulting in cell death. By using a P. aeruginosa abscess infection model, we demonstrate enhanced therapeutic efficacies of the combination of D-11 with various antibiotics. In addition, we found that the combination of D-11 and azithromycin enhanced the inhibition of biofilm formation and the elimination of established biofilms. Our study provides a realistic treatment option for combining close-to-nature synthetic peptide adjuvants with existing antibiotics to combat infections caused by P. aeruginosa.

SUBMITTER: Xia Y

PROVIDER: S-EPMC8445441 | biostudies-literature |

REPOSITORIES: biostudies-literature

ACCESS DATA

Similar Datasets

Project description:Bacterial colonization and biofilm development on medical devices can lead to infection. Antimicrobial peptide-coated surfaces may prevent such infections. Melimine and Mel4 are chimeric cationic peptides showing broad-spectrum antimicrobial activity once attached to biomaterials and are highly biocompatible in animal models and have been tested in Phase I and II/III human clinical trials. These peptides were covalently attached to glass using an azidobenzoic acid linker. Peptide attachment was confirmed using X-ray photoelectron spectroscopy and amino acid analysis. Mel4 when bound to glass was able to adopt a more ordered structure in the presence of bacterial membrane mimetic lipids. The ability of surface bound peptides to neutralize endotoxin was measured along with their interactions with the bacterial cytoplasmic membrane which were analyzed using DiSC(3)-5 and Sytox green, Syto-9, and PI dyes with fluorescence microscopy. Leakage of ATP and nucleic acids from cells were determined by analyzing the surrounding fluid. Attachment of the peptides resulted in increases in the percentage of nitrogen by 3.0% and 2.4%, and amino acid concentrations to 0.237 nmole and 0.298 nmole per coverslip on melimine and Mel4 coated surfaces, respectively. The immobilized peptides bound lipopolysaccharide and disrupted the cytoplasmic membrane potential of Pseudomonas aeruginosa within 15 min. Membrane depolarization was associated with a reduction in bacterial viability by 82% and 63% for coatings melimine and Mel4, respectively (p < 0.001). Disruption of membrane potential was followed by leakage of ATP from melimine (1.5 ± 0.4 nM) or Mel4 (1.3 ± 0.2 nM) coated surfaces compared to uncoated glass after 2 h (p < 0.001). Sytox green influx started after 3 h incubation with either peptide. Melimine coatings yielded 59% and Mel4 gave 36% PI stained cells after 4 h. Release of the larger molecules (DNA/RNA) commenced after 4 h for melimine (1.8 ± 0.9 times more than control; p = 0.008) and after 6 h with Mel4 (2.1 ± 0.2 times more than control; p < 0.001). The mechanism of action of surface bound melimine and Mel4 was similar to that of the peptides in solution, however, their immobilization resulted in much slower (approximately 30 times) kinetics.

Project description:Microorganisms continuously monitor their surroundings and adaptively respond to environmental cues. One way to cope with various stress-related situations is through the activation of the stringent stress response pathway. In Pseudomonas aeruginosa this pathway is controlled and coordinated by the activity of the RelA and SpoT enzymes that metabolize the small nucleotide secondary messenger molecule (p)ppGpp. Intracellular ppGpp concentrations are crucial in mediating adaptive responses and virulence. Targeting this cellular stress response has recently been the focus of an alternative approach to fight antibiotic resistant bacteria. Here, we examined the role of the stringent response in the virulence of P. aeruginosa PAO1 and the Liverpool epidemic strain LESB58. A ?relA/?spoT double mutant showed decreased cytotoxicity toward human epithelial cells, exhibited reduced hemolytic activity, and caused down-regulation of the expression of the alkaline protease aprA gene in stringent response mutants grown on blood agar plates. Promoter fusions of relA or spoT to a bioluminescence reporter gene revealed that both genes were expressed during the formation of cutaneous abscesses in mice. Intriguingly, virulence was attenuated in vivo by the ?relA/?spoT double mutant, but not the relA mutant nor the ?relA/?spoT complemented with either gene. Treatment of a cutaneous P. aeruginosa PAO1 infection with anti-biofilm peptides increased animal welfare, decreased dermonecrotic lesion sizes, and reduced bacterial numbers recovered from abscesses, resembling the phenotype of the ?relA/?spoT infection. It was previously demonstrated by our lab that ppGpp could be targeted by synthetic peptides; here we demonstrated that spoT promoter activity was suppressed during cutaneous abscess formation by treatment with peptides DJK-5 and 1018, and that a peptide-treated relA complemented stringent response double mutant strain exhibited reduced peptide susceptibility. Overall these data strongly indicated that synthetic peptides target the P. aeruginosa stringent response in vivo and thus offer a promising novel therapeutic approach.

Dataset Information

Elucidating the mechanism by which synthetic helper peptides sensitize Pseudomonas aeruginosa to multiple antibiotics.

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

Tweets