Vivian Tullio

Solid lipid nanoparticles as promising tool for intraocular tobramycin delivery: Pharmacokinetic studies on rabbits

Eye drops are widely accepted as formulations for targeting the anterior segment notwithstanding their limitations in terms of bioavailability. The unique structure of the eye requires specially-designed formulations able to favor the pharmacokinetic profile of administered drugs, mainly minimizing the influence of ocular barriers. Nanotechnology-based delivery systems lead to significant technological and therapeutical advantages in ophthalmic therapy. The aim of the present study was to determine whether tobramycin as ion-pair incorporated in mucoadhesive Solid Lipid Nanoparticles (SLN) reaches the inner parts of the eye favoring drug activity. After technological characterization of the tobramycin entrapped SLN formulation (Tobra-SLN), a pharmacokinetic study in rabbits after topical instillation and intravenous administration of the formulation has been carried out. In addition, the intracellular activity of Tobra-SLN formulation against phagocytosed Pseudomonas aeruginosa was investigated. The SLN were spherical in shape, and showed a hydrodynamic diameter of about 80nm, a negative zeta potential (-25.7mV) with a polydispersity index of 0.15, representative of a colloidal dispersion with high quality, characterized by an unimodal relatively narrow size distribution. As demonstrated by FTIR and DSC, tobramycin ion-pair could be concentrated into lipid inner core of SLN, without interaction with the stearic acid, thus promoting a slow and constant drug release profile in the dissolution medium. Surprisingly, the drug concentration was significantly higher in all ocular tissues after ocular and intravenous administration of Tobra-SLN formulation with respect to reference formulations and only Tobra-SLN allowed the penetration of drug into retina. Furthermore, the use of Tobra-SLN resulted in both higher intraphagocytic antibiotic concentrations in polymorphonuclear granulocytes and greater bactericidal activity against intracellular Pseudomonas aeruginosa, probably due to the ability of Tobra-SLN to penetrate either into phagocytic cells, or alternatively to cross bacterial barrier. The present study broadens the knowledge on the use of SLN as carriers for ocular drug delivery to the posterior chamber and might open new avenues for treatment of ocular infections, representing a strategy to overcome the microbial resistance.

Non-dermatophyte moulds as skin and nail foot mycosis agents: Phoma herbarum, Chaetomium globosum and Microascus cinereus

The increased prevalence of dermatomycoses along with the wide range of organisms now recognized as potential pathogens needs accurate laboratory isolation and identification of the aetiological agents. In this report three cases of foot dermatomycoses due to filamentous fungi commonly present in the environment with ubiquitous distribution are described in immunocompetent subjects. Skin and nail samples were collected, suspended in 20% KOH solution, examined under a light microscope and cultured in Mycobiotic agar and Sabouraud dextrose agar containing chloramphenicol to detect fungal growth. Phoma herbarum, Chaetomium globosum, and Microascus cinereus were isolated and identified.

In Vitro Activities of Fluconazole and Voriconazole against Clinical Isolates of Candida spp. Determined by Disk Diffusion Testing in Turin, Italy

ABSTRACT The in vitro activities of fluconazole and voriconazole against 1,024 clinical isolates of Candida spp. were determined by the agar disk diffusion test using the Clinical and Laboratory Standards Institute (CLSI) M44-A guidelines. The results of this investigation demonstrated the broad-spectrum in vitro activity of voriconazole, relative to that of fluconazole, against yeasts tested, in particular fluconazole-resistant isolates, such as Candida krusei that showed high susceptibility to voriconazole. The situation in Turin, Italy, is quite similar to that of the rest of Italy, reflecting the worldwide trend.

Schizophyllum commune: an unusual of agent bronchopneumonia in an immunocompromised patient

We report a case of bronchopneumonia due to Schizophyllum commune in an immunocompromised patient. While this fungus rarely causes disease in humans, it has been reported in association with several clinical entities and lung disorders. A 59-year-old white man with a gastric carcinoma was admitted to S. Giovanni Battista Hospital (Turin, Italy). Three days after the admission, he developed a bronchopneumonia, which was diagnosed through the use of X-ray and showed an abnormal infiltrative shadow. Samples of bronchial aspirate were collected for laboratory microbiological investigation. Direct microscopic examination of these specimens revealed the presence of numerous septate, hyaline hyphae and rare clamp connections. Sabouraud Dextrose Agar and Columbia agar plus 5% blood media inoculated with portions of the same specimens yielded, after 4-5 days of incubation at 25 degrees C and 37 degrees C, a cottony white mould. The fungus was identified on the basis of its macroscopic and microscopic morphology. The macroscopic examination of the colony showed raised, curved, fan-shaped and shell-like basidiocarps. The microscope examination revealed the presence of hyaline, septate hyphae with clamp connections and short, thin spicules. The fungal isolate was identified as S. commune. The patient was cured after therapy with intravenous fluconazole (600 mg twice daily for over six weeks).

Antimicrobial chitosan nanodroplets: new insights for ultrasound-mediated adjuvant treatment of skin infection

BACKGROUND Chronic wounds, characterized by hypoxia, inflammation and impaired tissue remodeling, are often worsened by bacterial/fungal infections. Intriguingly, chitosan-shelled/decafluoropentane-cored oxygen-loaded nanodroplets (OLNs) have proven effective in delivering oxygen to hypoxic tissues. AIM The present work aimed at investigating nanodroplet antimicrobial properties against methicillin-resistant Staphylococcus aureus (MRSA) or Candida albicans, toxicity on human keratinocytes (HaCaT) and ultrasound (US)-triggered transdermal delivery. MATERIALS & METHODS Nanodroplet antibacterial/antifungal properties, human cytotoxicity, and US-triggered transdermal delivery were measured through microbiological, biochemical, and sonophoresis assays, respectively. RESULTS OLNs and oxygen-free nanodroplets (OFNs) displayed short- or long-term cytostatic activity against MRSA or Candida albicans, respectively. OLNs were not toxic to keratinocytes, whereas OFNs slightly affected cell viability. Complementary US treatment promoted OLN transdermal delivery. CONCLUSION As such, US-activated chitosan-shelled OLNs appear as promising, nonconventional and innovative tools for adjuvant treatment of infected chronic wounds.

Synergistic effect of erythromycin on polymorphonuclear cell antibacterial activity against erythromycin-resistant phenotypes of Streptococcus pyogenes

To evaluate the synergistic activity of erythromycin and human polymorphonuclear cells (PMNs) on the binomial erythromycin-resistant (ERY(R)) Streptococcus pyogenes/host, the phagocytic and bactericidal activities of PMNs against ERY(R) streptococcal strains (cMLS(B), M, and iMLS(B) A, B and C phenotypes) were assessed in the presence of the macrolide. The results showed that when erythromycin, PMNs and streptococci [both erythromycin-sensitive (ERY(S)) and ERY(R)] were simultaneously present in the culture medium, PMN phagocytic activity was similar to that of drug-free controls. In contrast, the results emphasised a significant high increase in intracellular killing by PMNs in the presence of erythromycin not only for ERY(S) streptococci but also for ERY(R)S. pyogenes with high (cMLS(B), iMLS(B) A and iMLS(B) B phenotypes) and moderate (M and iMLS(B) C phenotypes) erythromycin resistance compared with controls without drug. From literature data it emerged that, even if intracellularly concentrated, erythromycin is relatively inactive because of its instability. The results indicate that the enhanced intra-PMN streptococcal killing detected is mainly attributable to PMN bactericidal systems that synergise with intracellular erythromycin in eradicating ERY(R)S. pyogenes strains (both with high and moderate resistance). These data confirm that the antibiotic resistance detected in vitro does not always imply a failure of antimicrobial treatment.

Synergy of Caspofungin with Human Polymorphonuclear Granulocytes for Killing Candida albicans

ABSTRACT The influence of caspofungin on polymorphonuclear leukocyte (PMN) phagocytosis and intracellular killing of Candida albicans was investigated. Caspofungin, at all of the concentrations tested (2, 3.2, and 8 μg/ml), significantly increased intracellular killing by PMNs through its direct action on both yeast cells and PMNs, indicating the potential ability of caspofungin to synergize with phagocytes for candidal killing. Caspofungin may therefore constitute an effective therapeutic option for the treatment of invasive fungal infections, including those refractory to conventional treatment with azole agents.

Role of caspofungin in restoring the impaired phagocyte-dependent innate immunity towards Candida albicans in chronic haemodialysis patients

Phagocyte-dependent cellular immunity in chronic kidney disease patients undergoing haemodialysis treatment is frequently impaired owing to the uraemic state, resulting in an intrinsic susceptibility to developing invasive fungal infections with high mortality rates. Since synergism between phagocytic cells and antifungal drugs may be crucial for successful therapy, the aim of this study was to evaluate the effects exerted by caspofungin (CAS) on the functional activities of polymorphonuclear cells (PMNs) in haemodialysed patients (HDs) towards Candida albicans compared with those of PMNs from healthy subjects (HSs). PMNs were separated from venous blood samples of 66 HDs and 30 HSs (as controls), and measurement of phagocytic and intracellular fungicidal activities of HD-PMNs and HS-PMNs was performed in the presence of CAS at the minimum inhibitory concentration (MIC) and at sub-MICs. CAS-free controls were also included. In the drug-free test condition, no significant difference between the phagocytic activity of HD-PMNs and HS-PMNs was detected. In contrast, a progressive decline in the intracellular killing activity of HD-PMNs against proliferating yeasts was observed. CAS at MIC and sub-MIC levels was able to improve significantly the intracellular fungicidal activity of HD-PMNs against C. albicans, restoring their functionality. These findings provide evidence that CAS exerts a synergistic effect on HD-PMNs against C. albicans, being able to strength the depressed intracellular killing activity. These results corroborate the use of CAS as an effective therapeutic option for the treatment of invasive fungal infections in HDs, in whom even a marginal influence of antifungal drugs on host response may have a relevant effect.

In vitro compared activity of telithromycin and azithromycin against northwest Italian isolates of Streptococcus pyogenes and Streptococcus pneumoniae with different erythromycin susceptibility.

Aims:  This study compared the in vitro activity of telithromycin with that of azithromycin against 438 Streptococcus pyogenes and 198 Streptococcus pneumoniae, isolated over the period 2005–2007 from specimens of different human origin obtained in three Piemonte Region’s hospitals.

Positive Interaction of Thyme (Red) Essential Oil with Human Polymorphonuclear Granulocytes in Eradicating Intracellular Candida albicans

The essential oils have started to be recognized for their potential antimicrobial role only in recent years. Clinical experience showed that the efficacy of antimicrobial agents depends not only on their direct effect on a given microorganism but also on the functional activity of the host immune system. Since data on the effects of essential oils on the innate immune system are scanty and fragmentary, the aim of this study was to evaluate the influence of thyme (red) essential oil (EO), at subinhibitory/inhibitory concentrations, on intracellular killing activity by human polymorphonuclear granulocytes (PMNs) against Candida albicans. In order to provide a frame of reference for the activity of this EO, its in vitro killing activity in the absence of PMNs was also evaluated.Results showed that EO at subminimal inhibitory (subMIC)/minimal inhibitory (MIC) concentrations significantly enhanced intracellular killing of C. albicans in comparison with EO-free controls and was comparable to the positive control (fluconazole). In in vitro killing assays without PMNs, we observed progressive growth of the yeast cells in the presence of EO subMIC/MIC concentrations. A positive antifungal interaction with phagocytes could explain why this EO, which appeared to be only fungistatic in time-kill assays, had efficacy in killing yeast cells once incubated with PMNs.