Cathelicidin-BF-15

Cathelicidin-BF, derived from the banded krait (Bungarus fasciatus), is a typically cationic, amphiphilic and α-helical antimicrobial peptide (AMP) with 30 amino acids that exerts powerful effects on multidrug-resistant (MDR) clinical isolates, including Pseudomonas aeruginosa, Acinetobacter baumannii, and Klebsiella pneumoniae, but whether it targets plasma membranes or intracellular targets to kill bacteria is still controversial. In the present study, we demonstrated that the disruption of bacterial membranes with high concentrations of cathelicidin-BF was the cause of bacterial death, as with conventional antibiotics at high concentrations. At lower concentrations, cathelicidin-BF did not cause bacterial plasma membrane disruption, but it was able to cross the membrane and aggregate at the nucleoid regions. Functional proteins of the transcription processes of P. aeruginosa and A. baumannii were affected by sublethal doses of cathelicidin-BF, as demonstrated by comparative proteomics using isobaric tags for relative and absolute quantification and subsequent gene ontology (GO) analysis. Analysis using the Kyoto Encyclopedia of Genes and Genomes showed that cathelicidin-BF mainly interferes with metabolic pathways related to amino acid synthesis, metabolism of cofactors and vitamins, metabolism of purine and energy supply, and other processes. Although specific targets of cathelicidin-BF must still be validated, our study offers strong evidence that cathelicidin-BF may act upon intracellular targets to kill superbugs, which may be helpful for further efforts to discover novel antibiotics to fight against them.

Antimicrobial peptides (AMPs) represent a promising therapeutic alternative for the treatment of antibiotic-resistant bacterial infections. The present study investigates the antimicrobial activity of new, rationally-designed derivatives of a short α-helical peptide, RR. From the peptides designed, RR4 and its D-enantiomer, D-RR4, emerged as the most potent analogues with a more than 32-fold improvement in antimicrobial activity observed against multidrug-resistant strains of Pseudomonas aeruginosa and Acinetobacter baumannii. Remarkably, D-RR4 demonstrated potent activity against colistin-resistant strains of P. aeruginosa (isolated from cystic fibrosis patients) indicating a potential therapeutic advantage of this peptide over several AMPs. In contrast to many natural AMPs, D-RR4 retained its activity under challenging physiological conditions (high salts, serum, and acidic pH). Furthermore, D-RR4 was more capable of disrupting P. aeruginosa and A. baumannii biofilms when compared to conventional antibiotics. Of note, D-RR4 was able to bind to lipopolysaccharide to reduce the endotoxin-induced proinflammatory cytokine response in macrophages. Finally, D-RR4 protected Caenorhabditis elegans from lethal infections of P. aeruginosa and A. baumannii and enhanced the activity of colistin in vivo against colistin-resistant P. aeruginosa.

Nonfermenting Gram-negative bacteria such as Pseudomonas aeruginosa and Acinetobacter baumannii are widespread in the environment and are increasingly associated with nosocomial infections, often associated with multidrug-resistance phenotypes. This study aimed to evaluate epidemiological, physiological, and molecular characteristics of carbapenem resistance in P. aeruginosa and A. baumannii. In total, 63 nonreplicated strains (44 A. baumannii and 19 P. aeruginosa) were isolated from hospitalized patients. Antimicrobial resistance patterns, biocide tolerance, oxidative stress, hemolytic activity, and biofilm formation were assessed. Genetic markers related to β-lactamase synthesis, efflux systems, and porin loss were screened by PCR. Epidemiological data of patients were analyzed. Advanced age, intensive care unit admission, invasive medical devices, treatment with fluoroquinolones or β-lactams/β-lactamase inhibitor combinations, and prolonged hospital stay were predisposing factors for infection. Colistin showed to be active in vitro against these bacteria. Carbapenem-resistant P. aeruginosa strains did not show hemolytic activity and were less tolerant to oxidative stress and biocides. However, increased ability of biofilm formation was observed, comparing to the carbapenem-susceptible isolates. Genetic markers related to oxacillinases synthesis (OXA-23 and OXA-143), oprD absence, and efflux pump (adeB) were detected in carbapenem-resistant A. baumannii. Screening for OXA-51-like gene was performed as confirmatory test for A. baumannii identification. In P. aeruginosa genes encoding efflux pumps (MexAB-OprM, MexCD-OprJ, MexEF-OprN, and MexXY-OprM) and SPM-1 were found; besides, oprD absence was also observed. Our results suggest that these organisms are well adapted to different environments and confirm the difficulty of therapeutic management of patients with infections associated with multidrug-resistant microorganisms, with direct impact on mortality and epidemiological control of these strains in health centers.