Emanuela Roscetto

Biofilm-associated infections caused by Staphylococcus aureus are highly resistant to antimicrobial treatments. Low-intensity electric fields have shown promise as an antibiofilm strategy; however, the role of frequency remains poorly understood. In this study, mature S. aureus biofilms were exposed to low-intensity alternating electric fields (12.5 mV/cm) across broadband (10 Hz–10 MHz) frequency range and band-limited sub-ranges, as well as to selected single-tone frequencies within the interval 1–100 kHz. Antibiofilm activity was assessed in terms of cell culturability (CFU/mL) and biofilm biomass. A significant frequency-dependent effect was observed (p < 0.001), with maximal activity observed only at specific frequencies within the range 1–100 kHz, whereas other frequencies were ineffective. These findings demonstrate that the electrical antibiofilm effect is characterized by a frequency selective response, which is similar to a band-pass filter with a nearly flat shape inside the band-pass range and strongly attenuated effects outside such a band. This frequency-selective response supports the hypothesis of a biophysical mechanism involving membrane and matrix interactions and highlights the potential for developing targeted, frequency-optimized electroceutical strategies.

Enterobacter spp. and Giardia duodenalis are two resistant and globally distributed pathogens widely reported in humans and animals worldwide. To contrast the antimicrobial resistance phenomenon, this study aimed to evaluate the in vitro activity of antimicrobial peptides (AMPs), such as temporins, on the viability of Enterobacter spp. and G. duodenalis cysts isolated from naturally infected dogs. A total of 50 clinically healthy dogs were sampled, with 25 in each of the two sampling sessions. Rectal and oral swabs for Enterobacter spp. and fecal samples for G. duodenalis isolation were carried out. The AMPs were synthesized by a solid-phase peptide synthesis approach. Enterobacter ludwigii and E. hormaechei strains were isolated from two different dogs, while 16 dogs were positive for G. duodenalis. Subsequently, cysts and bacterial strains were treated with the following peptides: TL-34, TL-48, TL-42, TL-51, RB-71, and RB-58 at different concentrations (i.e., 250 mg/mL, 125 mg/mL, 62. mg/mL, and 31.25 for Enterobacter spp. strains and 0.15 mg/mL, 0.3 mg/mL, 0.6 mg/mL, and 1.2 mg/mL for G. duodenalis cysts). The results showed antimicrobial activity of TL-34, TL-48, and RB-71 against the two Enterobacter strains, while TL-51/TL-42 and RB-58 were not effective. In contrast, for G. duodenalis, TL-34, TL-48, TL-42, and RB-58 were effective at the two highest concentrations, inhibiting the viability of 100% of Giardia cysts, while the other two peptides (TL-51 and RB-71) showed less effectiveness. The AMPs used in this study have in vitro antimicrobial activity on cysts and bacterial strains and appear to have potential for the treatment of these infections in dogs.

Bacterial biofilms pose a significant challenge to antibiotic treatment. Conventional therapeutic interventions often fail to effectively eradicate mature biofilms due to the protective extracellular matrix and altered physiological states of the embedded bacteria. In this study, we show that low-intensity sinusoidal electric fields induce a time-limited window of vulnerability in mature Staphylococcus aureus biofilms, increasing their susceptibility to vancomycin. In particular, the simultaneous application of electric fields and vancomycin during a transient vulnerability window results in a potentiated effect, leading to an approximately 3-log ₁₀ reduction in culturable cell counts relative to untreated controls. Conversely, the effectiveness of the antibiotic is reduced when vancomycin is administered after the cessation of the time-limited vulnerability window associated to the electrical stimulation, indicating a time-dependent reversibility of the electrical effect on the biofilm. These results suggest that the electric fields can create a critical phase of increased susceptibility to antibiotics, establishing a bioelectric strategy for biofilm control and highlighting the importance of treatment timing.

Endophytic fungi (EF) are microorganisms that colonize the internal tissues of host plants, providing a range of benefits to them. In this symbiosis, they act as a reservoir of bioactive metabolites that are important for enhancing the host’s defense mechanisms as a resistance against pathogens. These molecules usually possess antimicrobial properties that can be exploited for application in agriculture and medicine. In this context, the current work was designed to evaluate the phytotoxic and antimicrobial properties of the endophytic fungus Curvularia inaequalis, isolated for the first time from the Iranian medicinal plant Echium khuzistanicum. Culture filtrates, their organic extracts, and isolated metabolites were tested against a series of plants to assess their phytotoxicity, as well as against a wide range of plant and human pathogens to evaluate their antimicrobial activity. The main compounds characterizing the organic extract of C. inaequalis have been identified as (R)-phomalactone, catenioblin A, and (-) asperpentyn (1–3) by using spectroscopic techniques, NMR mainly, and HR-ESI-MS. In the bioactivity evaluation carried out in this study, (R)-phomalactone (1) stood out as the most promising compound, exhibiting significant non-host phytotoxic activity on tomato leaves; potent antibacterial activity against a wide range of human pathogens, including methicillin-resistant Staphylococcus aureus (MRSA) strains; and marked antifungal activity against several economically important phytopathogens. (–)-Asperpentyn (3) also showed robust and selective antifungal activity against phytopathogens, while catenioblin A (2) exhibited only a slight phytotoxic effect and limited overall bioactivity in this study. These findings reveal that the isolated endophytic fungi hold considerable promise as an untapped source of bioactive metabolites with antibacterial, antifungal, and phytotoxic activities.

In this paper, the authors describe an experimental study carried out on biological samples consisting of a 96-h mature Methicillin-Resistant-Staphylococcus-Aureus biofilm. The initial objective was to electrically characterize the biofilm using impedance spectroscopy, by scanning a wide range of frequencies [1 Hz ÷ 10 MHz]. Concurrently, confocal microscopy observations, XTT assays, crystal violet staining method and colony-forming unit assay were performed to characterize the biological activity. The experimental investigation unexpectedly demonstrated that the reproducibility of measurement data was significantly affected by the destructive interaction between the electric field and the biofilm. This interaction was found to be strongly dependent on both the amplitude of the field and the exposure time. Moreover, a significative reduction of total biomass of the biofilm was found in a specific frequency range [10 kHz ÷ 100 kHz]. The results suggest several limitations of impedance spectroscopy as a tool for biofilm identification, since the “sample under test” must not be altered during the measurement process. Conversely, they demonstrate the deleterious impact of the electric field on the biofilm, thereby unveiling a potentially efficacious therapeutic paradigm for biofilm treatment. We expect that the open issues highlighted in this paper will be a source of inspiration for further understanding of the mechanism of interaction between the electric field and biofilm, both in terms of treatment and diagnosis.

Background: Carbapenem-resistant Acinetobacter baumannii (CRAB) poses significant challenges in healthcare due to its multidrug resistance and high mortality rates among critically ill patients. Results: We enrolled 45 patients. Cefiderocol was administered to 40% of patients, often (38.8%) in combination with other antibiotics. Colistin was administered to 60% of patients and always in combination, mostly with ampicillin–sulbactam. The overall ECS and OCS rates were 77.8% and 66.7%, respectively. Patients treated with an initial cefiderocol-based regimen showed a higher rate of ECS compared with patients initially treated with colistin-based regimens (100% vs. 63%, p < 0.05). Patients treated with cefiderocol alone showed a higher rate of ECS compared with patients treated with cefiderocol-based regimens (100% vs. 70.6%, p < 0.05). No differences in OCS rates were recorded depending on the treatment received. Additionally, cefiderocol regimens were associated with fewer ADRs compared to colistin-based treatment. Methods: This prospective observational study enrolled patients with CRAB infections from January 2022 to August 2023. Patients were treated with cefiderocol-based or colistin-based regimens and were monitored for 28 days to assess early clinical success (ECS), overall clinical success (OCS) and adverse drug reactions (ADRs). Conclusions: This study highlights the potential advantages of cefiderocol, even used as a monotherapy, in treating CRAB, especially when early clinical and laboratory response was assessed. This research contributes to the ongoing discussion on the most effective and safe treatments for combating CRAB infections, supporting the use of cefiderocol in clinical practice.

Lampedusa, the largest island of the Pelagie archipelago, Sicily, Italy, has proven to be a rich source of plants and shrubs used in folk medicine. These plants, often native to the island, have been very poorly investigated for their phytochemical composition and biological potential to be translated into pharmacological applications. To start achieving this purpose, a specimen of Limonium lopadusanum, a plant native to Lampedusa, was investigated for the first time. This manuscript reports the results of a preliminary biological assay, focused on antimicrobial activity, carried out using the plant organic extracts, and the isolation and chemical and biological characterization of the secondary metabolites obtained. Thus 3-hydroxy-4-methoxybenzoic acid methyl ester (syn: methyl isovanillate, (1), methyl syringate (2), pinoresinol (3), erythrinassinate C (4) and tyrosol palmitate (5) were isolated. Their antimicrobial activity was tested on several strains and compound 4 showed promising antibacterial activity against Enterococcus faecalis. Thus, this metabolite has antibiotic potential against the drug-resistant opportunistic pathogen E. faecalis.

The emergence of multidrug-resistant strains requires the urgent discovery of new antibacterial drugs. In this context, an antibacterial screening of a subset of anthelmintic avermectins against gram-positive and gram-negative strains was performed. Selamectin completely inhibited bacterial growth at 6.3 μg/mL concentrations against reference gram-positive strains, while no antibacterial activity was found against gram-negative strains up to the highest concentration tested of 50 μg/mL. Given its relevance as a community and hospital pathogen, further studies have been performed on selamectin activity against Staphylococcus aureus (S. aureus), using clinical isolates with different antibiotic resistance profiles and a reference biofilm-producing strain. Antibacterial studies have been extensive on clinical S. aureus isolates with different antibiotic resistance profiles. Mean MIC₉₀ values of 6.2 μg/mL were reported for all tested S. aureus strains, except for the macrolide-resistant isolate with constitutive macrolide-lincosamide-streptogramin B resistance phenotype (MIC₉₀ 9.9 μg/mL). Scanning Electron Microscopy (SEM) showed that selamectin exposure caused relevant cell surface alterations. A synergistic effect was observed between ampicillin and selamectin, dictated by an FIC value of 0.5 against methicillin-resistant strain. Drug administration at MIC concentration reduced the intracellular bacterial load by 81.3%. The effect on preformed biofilm was investigated via crystal violet and confocal laser scanning microscopy. Selamectin reduced the biofilm biomass in a dose-dependent manner with minimal biofilm eradication concentrations inducing a 50% eradication (MBEC₅₀) at 5.89 μg/mL. The cytotoxic tests indicated that selamectin exhibited no relevant hemolytic and cytotoxic activity at active concentrations. These data suggest that selamectin may represent a timely and promising macrocyclic lactone for the treatment of S. aureus infections.

Invasive fungal infections (IFIs) represent a severe complication of COVID-19, yet they are under-estimated. We conducted a retrospective analysis including all the COVID-19 patients admitted to the Infectious Diseases Unit of the Federico II University Hospital of Naples until the 1 July 2021. Among 409 patients, we reported seven cases of IFIs by Candida spp., seven of Pneumocystis jirovecii pneumonia, three of invasive pulmonary aspergillosis, and one of Trichosporon asahii. None of the cases presented underlying predisposing conditions, excluding one oncohematological patient treated with rituximab. Ten cases showed lymphopenia with high rates of CD4+ < 200/µL. All cases received high-dose steroid therapy (mean duration 33 days, mean cumulative dosage 1015 mg of prednisone equivalent), and seven cases had severe COVID-19 disease (OSCI ≥ 5) prior to IFI diagnosis. The cases showed a higher overall duration of hospitalization (63 vs 24 days) and higher mortality rate (23% vs. 7%) compared with the COVID-19 patients who did not developed IFIs. Cases showed a higher prevalence of high-dose steroid therapy and lymphopenia with CD4+ < 200/µL, primarily due to SARS-CoV-2 infection and not related to underlying comorbidities. IFIs strongly impact the overall length of hospitalization and mortality. Therefore, clinicians should maintain a high degree of suspicion of IFIs, especially in severe COVID-19 patients.