Design of Cinnamaldehyde- and Gentamicin-Loaded Double-Layer Corneal Nanofiber Patches with Antibiofilm and Antimicrobial Effects.

In this study, two-layer poly(vinyl alcohol)/gelatin (PVA/GEL) nanofiber patches containing cinnamaldehyde (CA) in the first layer and gentamicin (GEN) in the second layer were produced by the electrospinning method. The morphology, chemical structures, and thermal temperatures of the produced pure (PVA/GEL), CA-loaded (PVA/GEL/CA), GEN-loaded (PVA/GEL/GEN), and combined drug-loaded (PVA/GEL/CA/GEN) nanofiber patches were determined by scanning electron microscopy (SEM), Fourier transform infrared spectroscopy, and differential scanning calorimetry, respectively. Their mechanical properties, swelling and degradation behavior, and drug release kinetics were investigated. SEM images showed that both drug-free and drug-loaded nanofiber patches possess smooth and monodisperse structures, and nanofiber size increase occurred as the amount of drug increased. The tensile test results showed that the mechanical strength decreased as the drug was loaded. According to the drug release results, CA release ended at the 96th hour, while GEN release continued until the 264th hour. The antibacterial and antibiofilm activities of PVA/GEL, PVA/GEL/CA, PVA/GEL/GEN, and PVA/GEL/CA/GEN nanofiber patches against Pseudomonas aeruginosa and Staphylococcus aureus were evaluated. Results showed that PVA/GEL/GEN and PVA/GEL/CA/GEN nanofiber patches have excellent antibacterial and antibiofilm activities. Moreover, all materials were biocompatible, with no cytotoxic effects in the mammalian cell model for 8 days. PVA/GEL/GEN nanofiber patches were the most promising material for a high cell survival ratio, which was confirmed by SEM images. This research aims to develop an alternative method to stop and treat the rapid progression of bacterial keratitis.

Zeolite 4A as a jammer of bacterial communication in Chromobacterium violaceum and Pseudomonas aeruginosa.

Aim: To investigate the hypothesis that zeolites interfere with quorum-sensing (QS) systems of Chromobacterium violaceum and Pseudomonas aeruginosa by adsorbing N-acyl homoserine lactone (AHL) signal molecules. Methods: QS inhibition by zeolite 4A was investigated using an AHL-based bioreporter assay. The adsorption of the AHLs was evaluated by performing inductively coupled plasma-optical emission spectroscopy and confirmed by Monte Carlo and molecular dynamic simulations. Results: Zeolite 4A reduced the violacein production in C. violaceum by over 90% and the biofilm formation, elastase and pyocyanin production in P. aeruginosa by 87, 68 and 98%, respectively. Conclusion: Zeolite 4A disrupts the QS systems of C. violaceum and P. aeruginosa by means of adsorbing 3-oxo-C6-AHL and 3-oxo-C12-AHL signaling molecules and can be developed as a novel QS jammer to combat P. aeruginosa-related infections.

Quorum sensing and virulence of Pseudomonas aeruginosa during urinary tract infections.

INTRODUCTION: In the opportunistic pathogen Pseudomonas aeruginosa, the production of several virulence factors depends on quorum sensing (QS) involving N-acylhomoserine lactone signal molecules. In vitro studies have suggested that the QS system is crucial in the pathogenesis of P. aeruginosa. However, it is unclear whether QS systems of P. aeruginosa play the same role during infections. METHODOLOGY: In this study, to explore the contribution of QS systems to the pathogenesis of P. aeruginosa during urinary tract infections, we collected 82 clinical isolates. Detection of N-acyl-homoserine lactones (C12-HSL and C4-HSL) was performed on agar plates employing biosensor strains C. violaceum. Elastase and biofilm production were determined spectrophotometrically. QS genes were detected by PCR and subsequently underwent sequencing. RESULTS AND CONCLUSION: Six isolates were found to be negative in the production of both C12-HSL and C4-HSL and all virulence factors tested.  PCR analysis of these isolates revealed that four isolates contained all four QS genes while one isolate was negative for lasR gene, and one isolate negative for lasI, lasR and rhlR genes. Sequence analyses of these isolates showed that the lasR, lasI, rhlR and rhlI genes had point mutations. The combination of these mutations probably explains their C12-HSL, C4-HSL and virulence factor deficiencies. Results of this study suggest that QS deficient clinical isolates occur and are still capable of causing clinical infections in humans. 

Tocopherol, carotene, phenolic contents and antibacterial properties of rose essential oil, hydrosol and absolute.

The antioxidant and antibacterial activities, and total phenolic contents of Rosa damascena Mill. flower extracts (absolute, essential oil and hydrosol) were investigated. The chemical compositions of these extracts were analysed by GC-MS. Phenylethyl alcohol (78.38%) was found to be the main constituent of rose absolute, while citrenellol and geraniol were the major compounds (>55%) of rose essential oil and hydrosol. Tocopherol and carotene levels were determined by high performance liquid chromatography (HPLC) analysis. The levels of beta carotene (422.3+/-35.6 ppm), alpha tocopherol (2397.1+/-72.5 ppm) and gamma tocopherol (343.1+/-28.4 ppm) of rose absolute were found to be higher than that of essential oil and hydrosol. Their total phenolic contents were also evaluated. The total phenolic content of the tested extracts varied from 5.2 to 2134.3 GAE/mg L(-1). Rose absolute and essential oil contained high levels of phenolics and demonstrated strong antibacterial activity against Escherichia coli (ATCC 25922), Pseudomonas aeruginosa (ATCC 27853), Bacillus subtilis (ATCC 6633), Staphylococcus aureus (ATCC 6538), Chromobacterium violaceum (ATCC 12472) and Erwinia carotovora (ATCC 39048) strains.

Characterization of N-butanoyl-L-homoserine lactone (C4-HSL) deficient clinical isolates of Pseudomonas aeruginosa.

In the opportunistic pathogen Pseudomonas aeruginosa, the production of several virulence factors such as elastase, rhamnolipids and pyocyanin depends on cell-to-cell signaling or quorum sensing (QS) involving N-acylhomoserine lactone (AHL) signal molecules. In vitro studies with laboratory strains and virulence studies in animals with these same strains have demonstrated the contribution of QS to the pathogenesis of P. aeruginosa. However, the importance of P. aeruginosa QS systems in the development of human infections is not clearly known. In order to determine if deficiency within the QS system compromises the ability of P. aeruginosa to cause infections in humans, we collected 50 P. aeruginosa clinical isolates. Phenotypic characterization showed that isolates I-457, I-458, I-459 and I-461 were defective in the production of N-butanoyl-l-homoserine lactone (C4-HSL) signaling molecule and virulence factors elastase, protease, pyocyanin and rhamnolipids. Analysis of the sequences of the lasR, lasI, rhlR and rhlI genes of these four isolates showed that two of the four isolates had mutational defects in both rhlR and rhlI genes while other two isolates were only mutated in the rhlI gene. The combination of rhlR and rhlI mutations or only rhlI mutation probably explains their C4-HSL and virulence factors deficiencies. These observations suggest that QS deficient P. aeruginosa clinical isolates are able to cause infections and that in addition to known virulence factors, factors yet unidentified may contribute to the pathogenesis of P. aeruginosa.

Inhibition of quorum sensing-regulated behaviors by Scorzonera sandrasica.

Many Gram-negative bacteria use N-acyl homoserine lactone signal molecules to monitor their own population density and coordinate gene regulation in a process called quorum sensing (QS). Increasing evidence implies that certain eukaryotes produce QS-inhibitory compounds. In this work, we tested 46 terrestrial plants materials for their ability to inhibit QS-regulated behaviors in different bacterial species. Plant materials were dried and extracted using different solvents. The chloroform-soluble compounds extracted from Scorzonera sandrasica were found to inhibit violacein production, a QS-regulated behavior in Chromobacterium violaceum. In addition, the chloroform extract was also able to inhibit QS-regulated carbapenem antibiotic production in Erwinia carotovora. Because the regulation of many bacterial processes is controlled by QS systems, the finding of natural compounds acting as QS inhibitors suggests an attractive tool to control and handle detrimental infections caused by human, animal, and plant pathogens.

N-butanoyl-L-homoserine lactone (BHL) deficient Pseudomonas aeruginosa isolates from an intensive care unit.

Acylated homoserine lactones (AHLs) are self-generated diffusible signal molecules that mediate population density dependent gene expression (quorum sensing) in a variety of Gram-negative bacteria, and several virulence genes of human pathogens are known to be controlled by AHLs. In this study, strains of Pseudomonas aeruginosa, Acinetobacter baumannii, Escherichia coli and Klebsiella pneumoniae, isolated from intensive care patients, were screened for AHL production by using AHL responsive indicator strains of Chromobacterium violaceum CV026 and Agrobacterium tumefaciens NT1. Positive reactions were recorded for all 50 isolates of P. aeruginosa and 10 isolates of Acinetobacter baumannii with Agrobacterium tumefaciens NT1. Surprisingly, most P. aeruginosa isolates gave negative results with C. violaceum CV026 in contrast to previous reports. This suggests that the new isolates of P. aeruginosa either failed to make short chain AHLs or the level of the signal molecule is very low.