The number of antibiotic-resistant microbial infections is dramatically increasing, while the discovery of new antibiotics is significantly declining. Furthermore, the activity of antibiotics is negatively influenced by the ability of bacteria to form sessile communities, called biofilms, and by the microenvironment of the infection, characterized by an acidic pH, especially in the lungs of patients suffering from cystic fibrosis (CF). Antimicrobial peptides represent interesting alternatives to conventional antibiotics, and with expanding properties. Here, we explored the effects of an acidic pH on the antimicrobial and antibiofilm activities of the AMP Esc(1-21) and we found that it slightly lost activity (from 2- to 4-fold) against the planktonic form of a panel of Gram-negative bacteria, with respect to a ≥ 32-fold of traditional antibiotics. Furthermore, it retained its activity against the sessile form of these bacteria grown in media with a neutral pH, and showed similar or higher effectiveness against the biofilm form of bacteria grown in acidic media, simulating a CF-like acidic microenvironment, compared to physiological conditions.

Clostridioides difficile infection (CDI) is one of the most common healthcare-associated infections, contributing to significant morbidity and mortality, particularly in hospitalized patients. Its incidence is influenced by infection control measures, antimicrobial stewardship, and hospital hygiene protocols. The Coronavirus disease 2019 (COVID-19) pandemic led to the widespread adoption of stringent infection prevention strategies. These measures may have influenced the incidence of CDI. At the university hospital Policlinico Umberto I in Rome, we conducted a retrospective analysis of CDI cases across three periods: preCOVID-19, COVID-19 and postCOVID-19. Our findings revealed a statistically significant decrease in CDI cases during the COVID-19 period compared to the preCOVID-19 period, as well as in the postCOVID-19 period compared to the COVID-19 period. Additionally, we explored the potential association between CDI and multidrug-resistant organisms (MDRO). Our findings indicate a reduction in MDRO infections among patients with CDI in the COVID-19 and postCOVID-19 periods compared to the preCOVID-19 period. These results highlight the potential impact of infection control strategies on CDI incidence and suggest a possible relationship between CDI and MDRO infection. Further research is needed to assess the long-term effects of these measures and their implications for future infection prevention strategies.

Background/Objectives: Developing novel strategies to combat respiratory infections caused by multidrug-resistant “priority pathogens” like the ESKAPEE Pseudomonas aeruginosa and Staphylococcus aureus is an urgent priority. Methods: We investigated two shortened variants of the proline-rich antimicrobial peptide (PrAMP) B7-005, B7-006 (15-mer) and B7-007 (13-mer). Evaluation included MIC assays against laboratory and clinical multidrug-resistant isolates, mechanistic studies of membrane permeabilization, cytotoxicity testing on BEAS-2B bronchial epithelial cells, and proteolytic stability assays in human elastase and sputum. Results: Despite their reduced size, lower positive charge, and decreased proline content, both variants retained full antimicrobial activity against clinical pathogens with consistent MIC values ≤ 25 µM. These variants exhibit membrane permeabilization in P. aeruginosa but may also relay on a hybrid mode of action involving also intracellular targets. Notably, B7-006 and B7-007 displayed low cytotoxicity compared to the lytic peptide BMAP-18. While B7-007 showed greater susceptibility to proteolytic degradation than its parent B7-005, it preserved partial integrity during the initial hours of exposure. Conclusions: Overall, these findings demonstrate that the B7 scaffold tolerates substantial truncation while preserving potency and selectivity, identifying a minimal 13-amino-acid active core. This work provides critical insights into structure–activity relationships and supports the development of compact, mechanistically versatile antimicrobial peptides to address the growing threat of multidrug-resistant respiratory pathogens.

To evaluate the in vitro activity of zosurabalpin (ZAB), a novel tethered macrocyclic peptide antibiotic, developed by Roche, against carbapenem-resistant Acinetobacter baumannii (CRAB) isolates. A total of 100 clinical CRAB isolates were obtained from respiratory samples and blood cultures of patients who were hospitalized at the University Hospital Policlinico Umberto I in Rome. Carbapenem resistance was confirmed by meropenem susceptibility testing with the MicroScan WalkAway system (Beckman Coulter). These isolates were tested for their susceptibility to ZAB and cefiderocol (FDC). ZAB MICs were determined by broth microdilution according to Clinical and Laboratory Standards Institute guidelines, using homemade cation-adjusted Mueller-Hinton broth (CAMHB) supplemented with 20% heat-inactivated horse serum. Instead, antimicrobial susceptibility to FDC was tested using the microdilution method with homemade iron-depleted CAMHB. The concentrations of both antibiotics ranged from 32 mg/L to ≤0.015 mg/L. Clinical break points for FDC and meropenem were defined according to the European Committee on Antimicrobial Susceptibility Testing 2025 guidelines. ZAB demonstrated potent in vitro activity against all clinical CRAB isolates, with an MIC₉₀ value of 0.25 mg/L and MIC values ranging from ≤0.015 to 0.5 mg/L. Eleven of the 100 clinical isolates showed resistance to FDC, with MIC values between 4 and >32 mg/L. Interestingly, these strains were susceptible to ZAB, with MIC values between 0.06 and 0.5 mg/L. These findings indicate that ZAB exhibits strong in vitro activity against CRAB, including strains resistant to last-line commercially available antibiotics such as FDC.

Carbapenemase-producing Klebsiella pneumoniae represents a major clinical threat due to the multidrug-resistant phenotype of this Gram-negative bacterium and to the limited available therapeutic options. Currently, antimicrobial peptides are regarded as promising candidates for the development of novel anti-infective strategies. In this study, the antimicrobial activity of the frog skin-derived antimicrobial peptide Esc(1-21) and its three analogs, an Aib-containing peptide and two variants bearing d-amino acids (i.e., the diastereomer Esc(1-21)-1c and [Aib8]-Esc(1-21)-1c isoform), was evaluated against a panel of clinical carbapenemase-producing Klebsiella pneumoniae isolates. In addition, the ability of these d-amino acid-containing peptides to potentiate the effectiveness of ceftazidime, a widely used third-generation cephalosporin for treatment of severe Gram-negative bacterial infections, was investigated. Mechanistic analyses revealed that Esc(1-21)-1c promotes outer membrane permeabilization, thereby facilitating ceftazidime entry into the periplasmic space. Notably, the peptide/ceftazidime combination treatment resulted in enhanced outer membrane destabilization and increased cytoplasmic membrane perturbation compared to the peptide alone. These findings support a self-reinforcing mechanism in which ceftazidime-mediated cell wall damage further destabilizes the bacterial envelope, promoting additional peptide uptake and amplifying cytoplasmic membrane disruption. Time–kill assays confirmed the superior bactericidal activity of the peptide/ceftazidime combination compared to either agent used alone. Importantly, no outer membrane perturbation was observed in a strain lacking a synergistic response, suggesting that membrane susceptibility is a key determinant of this cooperative interaction. Overall, these results highlight the potential of Esc-derived peptides as adjuvants capable of restoring β-lactam efficacy against multidrug-resistant carbapenemase-producing Klebsiella pneumoniae, offering a promising strategy to combat antimicrobial resistance.