In vitro activity of N-acetylcysteine against Stenotrophomonas maltophilia and Burkholderia cepacia complex grown in planktonic phase and biofilm.

Stenotrophomonas maltophilia and Burkholderia cepacia complex (Bcc) have been increasingly recognized as relevant pathogens in hospitalized, immunocompromised and cystic fibrosis (CF) patients. As a result of complex mechanisms, including biofilm formation and multidrug resistance phenotype, S. maltophilia and Bcc respiratory infections are often refractory to therapy, and have been associated with a worse outcome in CF patients. Here we demonstrate for the first time that N-acetylcysteine (NAC), a mucolytic agent with antioxidant and anti-inflammatory properties, may exhibit antimicrobial and antibiofilm activity against these pathogens. The antimicrobial and antibiofilm activity of high NAC concentrations, potentially achievable by topical administration, was tested against a collection of S. maltophilia (n = 19) and Bcc (n = 19) strains, including strains from CF patients with acquired resistance traits. Minimum Inhibitory Concentrations (MICs) and Minimum Bactericidal Concentrations (MBCs) ranged from 16 to 32 mg/ml and from 32 to >32 mg/ml, respectively. Sub-MIC concentrations (i.e., 0.25 × MIC) slowed down the growth kinetics of most strains. In time-kill assays, 2-day-old biofilms were more affected than planktonic cultures, suggesting a specific antibiofilm activity of NAC against these pathogens. Indeed, a dose- and time-dependent antibiofilm activity of NAC against most of the S. maltophilia and Bcc strains tested was observed, with a sizable antibiofilm activity observed also at 0.5 and 1 × MIC NAC concentrations. Furthermore, at those concentrations, NAC was also shown to significantly inhibit biofilm formation with the great majority of tested strains.

Sealing effectiveness against Staphylococcus aureus of five different implant-abutment connections.

PURPOSE: To compare the sealing effectiveness of four different implant-abutment connections against Staphylococcus aureus (S. aureus). The null hypotheses stated that there was no difference on sealing ability among the implant-abutment connections tested. METHODS: Five diverse commercially available dental implants were used to investigate the degree of microleakage at the implant-abutment junction (IAJ): Group 1: Torque Type conical implant with double conic connection - TTc (Winsix); Group 2: Torque Type conical implant with Cone Morse connection - TTcm (Winsix); Group 3: Free Lock connection - K type implant (Winsix); Group 4: Internal double hexagon - OsseoSpeed; Group 5: Internal hexagon - Aadva Implant. Nine implants were tested in each group and one group was used as the negative control (Group 4). The abutments were connected to implants according to manufacturers' recommendations. All procedures involving connection and disconnection of implants were performed in sterile conditions in a laminar flow biological safety cabinet. S. aureus ATCC 6538, a methicillin susceptible reference strain, was chosen for the experiments to test the degree of microleakage. Statistical analysis was performed in order to find significant differences among the five groups regarding sealing capability of the implant-abutment connections tested. The recorded data were statistically analyzed. RESULTS: One implant from Group 4 was excluded from the study because of the growth of a contaminant after 48 hours of incubation in all three wells (i.e. Paenibacillus pabuli, environmental Gram-positive bacteria). Wells A and B (i.e. wells where the samples were passed before being located in the final well C) of all other samples (n = 46) remained sterile over the 72 hours of incubation, indicating the lack of external contamination during implant-abutment connection. Similarly, no bacterial growth was observed in the five negative controls (i.e. one implant for each type), which had been inoculated with sterile saline and processed as the others. Bacterial microleakage was demonstrated with three samples, including one sample of Group 1, one of Group 3 and one of Group 5, in which growth of S. aureus in wells C after 48 hours of incubation was demonstrated (Table 1). No statistically significant difference between groups was noticed (P> 0.05). CLINICAL SIGNIFICANCE: Within the limitations of the present in vitro model, the results obtained suggest a tendency toward a better sealing capability for conical connections and internal hexagon.

Effects of photodynamic laser and violet-blue led irradiation on Staphylococcus aureus biofilm and Escherichia coli lipopolysaccharide attached to moderately rough titanium surface: in vitro study.

Effective decontamination of biofilm and bacterial toxins from the surface of dental implants is a yet unresolved issue. This study investigates the in vitro efficacy of photodynamic treatment (PDT) with methylene blue (MB) photoactivated with λ 635 nm diode laser and of λ 405 nm violet-blue LED phototreatment for the reduction of bacterial biofilm and lipopolysaccharide (LPS) adherent to titanium surface mimicking the bone-implant interface. Staphylococcus aureus biofilm grown on titanium discs with a moderately rough surface was subjected to either PDT (0.1% MB and λ 635 nm diode laser) or λ 405 nm LED phototreatment for 1 and 5 min. Bactericidal effect was evaluated by vital staining and residual colony-forming unit count. Biofilm and titanium surface morphology were analyzed by scanning electron microscopy (SEM). In parallel experiments, discs coated with Escherichia coli LPS were treated as above before seeding with RAW 264.7 macrophages to quantify LPS-driven inflammatory cell activation by measuring the enhanced generation of nitric oxide (NO). Both PDT and LED phototreatment induced a statistically significant (p < 0.05 or higher) reduction of viable bacteria, up to -99 and -98% (5 min), respectively. Moreover, besides bactericidal effect, PDT and LED phototreatment also inhibited LPS bioactivity, assayed as nitrite formation, up to -42%, thereby blunting host inflammatory response. Non-invasive phototherapy emerges as an attractive alternative in the treatment of peri-implantitis to reduce bacteria and LPS adherent to titanium implant surface without causing damage of surface microstructure. Its efficacy in the clinical setting remains to be investigated.

Effect of High N-Acetylcysteine Concentrations on Antibiotic Activity against a Large Collection of Respiratory Pathogens.

The effect of high N-acetylcysteine (NAC) concentrations (10 and 50 mM) on antibiotic activity against 40 strains of respiratory pathogens was investigated. NAC compromised the activity of carbapenems (of mostly imipenem and, to lesser extents, meropenem and ertapenem) in a dose-dependent fashion. We demonstrated chemical instability of carbapenems in the presence of NAC. With other antibiotics, 10 mM NAC had no major effects, while 50 mM NAC sporadically decreased (ceftriaxone and aminoglycosides) or increased (penicillins) antibiotic activity.

The effects of diode laser on Staphylococcus aureus biofilm and Escherichia coli lipopolysaccharide adherent to titanium oxide surface of dental implants. An in vitro study.

Effective decontamination of biofilm and bacterial toxins from the surface of dental implants is a yet unresolved issue. This in vitro study aims at providing the experimental basis for possible use of diode laser (λ 808 nm) in the treatment of peri-implantitis. Staphylococcus aureus biofilm was grown for 48 h on titanium discs with porous surface corresponding to the bone-implant interface and then irradiated with a diode laser (λ 808 nm) in noncontact mode with airflow cooling for 1 min using a Ø 600-µm fiber. Setting parameters were 2 W (400 J/cm2) for continuous wave mode; 22 µJ, 20 kHz, 7 µs (88 J/cm2) for pulsed wave mode. Bactericidal effect was evaluated using fluorescence microscopy and counting the residual colony-forming units. Biofilm and titanium surface morphology were analyzed by scanning electron microscopy (SEM). In parallel experiments, the titanium discs were coated with Escherichia coli lipopolysaccharide (LPS), laser-irradiated and seeded with RAW 264.7 macrophages to quantify LPS-driven inflammatory cell activation by measuring the enhanced generation of nitric oxide (NO). Diode laser irradiation in both continuous and pulsed modes induced a statistically significant reduction of viable bacteria and nitrite levels. These results indicate that in addition to its bactericidal effect laser irradiation can also inhibit LPS-induced macrophage activation and thus blunt the inflammatory response. The λ 808-nm diode laser emerges as a valuable tool for decontamination/detoxification of the titanium implant surface and may be used in the treatment of peri-implantitis.

Characterization of KPC-encoding plasmids from two endemic settings, Greece and Italy.

OBJECTIVES: Global dissemination of KPC-type carbapenemases is mainly associated with the spread of high-risk clones of Klebsiella pneumoniae and of KPC-encoding plasmids. In this study, we explored the population structure of KPC-encoding plasmids from the recent epidemics of KPC-producing K. pneumoniae (KPC-Kp) in Greece and Italy, the two major European endemic settings. METHODS: Thirty-four non-replicate clinical strains of KPC-Kp representative of the early phases (2008-11) of the Greek (n = 22) and Italian (n = 12) epidemics were studied. Isolates were typed by MLST, and blaKPC-carrying plasmids were characterized by S1 profiling, PCR-based replicon typing and RFLP. Transfer experiments by conjugation or transformation were carried out with Escherichia coli recipients. Eleven plasmids, representative of all different restriction profiles, were completely sequenced. RESULTS: The representative Greek strains belonged to 14 sequence types (STs), with a predominance of ST258. The representative Italian strains belonged to three STs, with a predominance of clonal complex 258 (ST258, ST512). The 34 strains carried plasmids of variable size (78-166 kb), either with blaKPC-2 or blaKPC-3 gene embedded in a Tn4401a transposon. Plasmids from Greek strains were mostly of a single RFLP type (A) and resembled the archetypal pKpQIL KPC-encoding plasmid, while plasmids from Italian strains belonged to a more heterogeneous population, showing five RFLP profiles (A, C-F). Types A and C resembled pKpQIL or deletion derivatives thereof, while types D-F included plasmids with hybrid structures between pKpQIL, pKPN3 and pKPN101-IT. CONCLUSIONS: pKpQIL-like plasmids played a major role in the dissemination of blaKPC in Greece and Italy, but evolved with different dynamics in these endemic settings.

Highly bactericidal Ag nanoparticle films obtained by cluster beam deposition.

The recent emergence of bacterial pathogens resistant to most or all available antibiotics is among the major global public health problems. As indirect transmission through contaminated surfaces is a main route of dissemination for most of such pathogens, the implementation of effective antimicrobial surfaces has been advocated as a promising approach for their containment, especially in the hospital settings. However, traditional wet synthesis methods of nanoparticle-based antimicrobial materials leave a number of key points open for metal surfaces: such as adhesion to the surface and nanoparticle coalescence. Here we demonstrate an alternative route, i.e. supersonic cluster beam deposition, to obtain antimicrobial Ag nanoparticle films deposited directly on surfaces. The synthesized films are simple to produce with controlled density and thickness, are stable over time, and are shown to be highly bactericidal against major Gram positive and Gram negative bacterial pathogens, including extensively drug-resistant strains. FROM THE CLINICAL EDITOR: The use of silver nanoparticle in health care is getting more widespread. The authors here describe the technique of cluster beam deposition for spraying silver on surfaces used in health care sectors. This may open a new avenue for future anti-bacterial coatings.