Photodynamic Inactivation of Legionella pneumophila Biofilm Formation by Cationic Tetra- and Tripyridylporphyrins in Waters of Different Hardness

The bacterium Legionella pneumophila is still one of the probable causes of waterborne diseases, causing serious respiratory illnesses. In the aquatic systems, L. pneumophila exists inside free-living amoebae or can form biofilms. Currently developed disinfection methods are not sufficient for complete eradication of L. pneumophila biofilms in water systems of interest. Photodynamic inactivation (PDI) is a method that results in an antimicrobial effect by using a combination of light and a photosensitizer (PS). In this work, the effect of PDI in waters of natural origin and of different hardness, as a treatment against L. pneumophila biofilm, was investigated. Three cationic tripyridylporphyrins, which were previously described as efficient agents against L. pneumophila alone, were used as PSs. We studied how differences in water hardness affect the PSs’ stability, the production of singlet oxygen, and the PDI activity on L. pneumophila adhesion and biofilm formation and in biofilm destruction. Amphiphilic porphyrin showed a stronger tendency for aggregation in hard and soft water, but its production of singlet oxygen was higher in comparison to tri- and tetracationic hydrophilic porphyrins that were stable in all water samples. All three studied porphyrins were shown to be effective as PDI agents against the adhesion of the L. pneumophila to polystyrene, against biofilm formation, and in the destruction of the formed biofilm, in their micromolar concentrations. However, a higher number of dissolved ions, i.e., water hardness, generally reduced somewhat the PDI activity of all the porphyrins at all tested biofilm growth stages.     

In vitro Streptococcus mutans biofilm formation on surfaces of chlorhexidine-containing dentin bonding systems

This in vitro study evaluated the influence of chlorhexidine diacetate (CDA) when blended within dentin bonding systems (DBSs) on Streptococcus mutans (S. mutans) biofilm formation.One commercially available 0.2% wt CDA-containing DBS (Peak Universal Bond) and five experimental 0.2% wt CDA-containing DBS formulations (experimental Adper Scotchbond 1XT plus experimental resins, R₂, R₃, R₄, R₅) were assessed vs their no-CDA containing counterparts. Twenty-eight DBSs disks were prepared for each group (6.4 mm×1.0 mm) and cured for 80 s at 800 mW/cm² in a nitrogen atmosphere. A modified Drip-Flow Reactor was used to grow S. mutans biofilms on specimen surfaces for 24 h and adherent, viable biomass was evaluated using a tetrazolium salt assay (MTT). Two specimens from each of the tested materials were processed with LIVE/DEAD stain and observed using laser confocal microscopy (CLSM) while two disks from each group were examined by using scanning electron microscopy (SEM).MTT assay, CLSM and SEM observations showed that CDA addition decreased, increased or did not change S. mutans biofilm formation. The lowest biofilm formation was obtained with Peak Universal Bond and R₅ (with and without CDA).It may be concluded that the chemical composition of DBSs determines their ability to promote or hamper biofilm formation. Therefore, CDA addition may be helpful in modulating biofilm formation provided that DBS formulation is tuned and optimized.     

Antimicrobial activity,effects on Streptococcus mutans biofilm and interfacial bonding of adhesive systems with and without antibacterial agent

ObjectivesThis study evaluated the antimicrobial activity (by agar disk diffusion test, AD), viability of S. mutans biofilm (VB), and effect on resin-dentin interface (RDI) of six adhesive systems.MethodsThree adhesives containing antibacterial components (Gluma 2Bond (G2B)/glutaraldehyde, Clearfil SE Protect (CSP)/MDPB and Peak Universal Bond (PUB)/chlorhexidine) and the corresponding adhesives with similar composition, but without antibacterial agents (Gluma Comfort Bond, GCB; Clearfil SE Bond, CSB and Peak LC Bond, PLB) were evaluated. AD was determined measuring the extent of halo formation following application of adhesives and control groups (light cured or not) to cultures of four strict anaerobic and four facultative bacteria. For VB, a UA159 biofilm was grown on adhesive-coated hydroxyapatite discs for five days, collected and processed to count the number of viable cells. For RDI analysis, adhesives were applied according to manufacturers' recommendations and teeth were restored with resin composite, sectioned to obtain bonded slices and visualized by SEM.ResultsAn inhibition halo was observed for G2B (strict anaerobic/light cured and not light cured), CSP (strict anaerobic and facultative/light cured and not light cured) and PUB (strict anaerobic and facultative/not light cured). PUB when light cured produced an inhibition halo on L. casei and S. mutans only. G2B and CSP significantly reduced the viability of S. mutans. Adhesives containing antimicrobial compounds had no detectable effect on RDI.ConclusionThe MDPB-containing bonding agent showed better results of inhibition for all oral pathogens tested and a decrease of viability of Streptococcus mutans biofilm, among the adhesives tested.     

Optimization and Evaluation of a Chitosan/Hydroxypropyl Methylcellulose Hydrogel Containing Toluidine Blue O for Antimicrobial Photodynamic Inactivation

Photodynamic inactivation (PDI) combined with chitosan has been shown as a promising antimicrobial approach. The purpose of this study was to develop a chitosan hydrogel containing hydroxypropyl methylcellulose (HPMC), chitosan and toluidine blue O (TBO) to improve the bactericidal efficacy for topical application in clinics. The PDI efficacy of hydrogel was examined in vitro against the biofilms of Staphylococcus aureus (S. aureus) and Pseudomonas aeruginosa (P. aeruginosa). Confocal scanning laser microscopy (CSLM) was performed to investigate the penetration level of TBO into viable S. aureus biofilms. Incorporation of HMPC could increase the physicochemical properties of chitosan hydrogel including the hardness, viscosity as well as bioadhesion; however, higher HMPC concentration also resulted in reduced antimicrobial effect. CSLM analysis further demonstrated that higher HPMC concentration constrained TBO diffusion into the biofilm. The incubation of biofilm and hydrogel was further performed at an angle of 90 degrees. After light irradiation, compared to the mixture of TBO and chitosan, the hydrogel treated sample showed increased PDI efficacy indicated that incorporation of HPMC did improve antimicrobial effect. Finally, the bactericidal efficacy could be significantly augmented by prolonged retention of hydrogel in the biofilm as well as in the animal model of rat skin burn wounds after light irradiation.     

Cytotoxicity,genotoxicity and antibiofilm activity on Streptococcus mutans of an experimental self-etching adhesive system containing natural Butia capitata oil

This study evaluated the biocompatibility and antibiofilm effects on Streptococcus mutans of an experimental self-etching adhesive system formulated with a natural vegetable product. The cytotoxicity of Clearfil Protect Bond (CPB), Clearfil SE Bond (CSEB), Adper SE Plus (AP), an experimental adhesive containing Butia capitata oil (EA) and an oil-free experimental adhesive as control (CA) was tested on 3T3/NIH mouse fibroblasts. Genotoxicity was indicated by micronuclei formation, and cell alterations were analyzed using light microscopy. To evaluate the effect of the adhesives on S. mutans biofilm, biomass samples were collected for dry weight and bacterial viability analyses, and the pH of the culture media was determined daily as an indicator of biofilm acidogenicity. The CA primer and EA uncured bond adhesive were the least toxic. No statistical difference was observed between EA and the untreated control, and EA showed similar effect to CPB. These findings suggest EA is biocompatible and presents activity against S. mutans biofilm.     

Antibacterial Effect of Dental Adhesive Containing Dimethylaminododecyl Methacrylate on the Development of Streptococcus mutans Biofilm

Antibacterial bonding agents and composites containing dimethylaminododecyl methacrylate (DMADDM) have been recently developed. The objectives of this study were to investigate the antibacterial effect of novel adhesives containing different mass fractions of DMADDM on Streptococcus mutans (S. mutans) biofilm at different developmental stages. Different mass fractions of DMADDM were incorporated into adhesives and S. mutans biofilm at different developmetal stages were analyzed by MTT assays, lactic acid measurement, confocal laser scanning microscopy and scanning electron microscopy observations. Exopolysaccharides (EPS) staining was used to analyze the inhibitory effect of DMADDM on the biofilm extracellular matrix. Dentin microtensile strengths were also measured. Cured adhesives containing DMADDM could greatly reduce metabolic activity and lactic acid production during the development of S. mutans biofilms (p < 0.05). In earlier stages of biofilm development, there were no significant differences of inhibitory effects between the 2.5% DMADDM and 5% DMADDM group. However, after 72 h, the anti-biofilm effects of adhesives containing 5% DMADDM were significantly stronger than any other group. Incorporation of DMADDM into adhesive did not adversely affect dentin bond strength. In conclusion, adhesives containing DMADDM inhibited the growth, lactic acid production and EPS metabolism of S. mutans biofilm at different stages, with no adverse effect on its dentin adhesive bond strength. The bonding agents have the potential to control dental biofilms and combat tooth decay, and DMADDM is promising for use in a wide range of dental adhesive systems and restoratives.     

Flavonoid Baicalein Suppresses Oral Biofilms and Protects Enamel Hardness to Combat Dental Caries

The objectives of this study were to investigate the effects of a novel method using flavonoids to inhibit Streptococcus mutans (S. mutans), Candida albicans (C. albicans) and dual-species biofilms and to protect enamel hardness in a biofilm-based caries model for the first time. Several flavonoids, including baicalein, naringenin and catechin, were tested. Gold-standard chlorhexidine (CHX) and untreated (UC) groups served as controls. Optimal concentrations were determined by cytotoxicity assay. Biofilm MTT, colony-forming-units (CFUs), biofilm biomass, lactic acid and polysaccharide production were evaluated. Real-time-polymerase-chain reaction (qRT-PCR) was used to determine gene expressions in biofilms. Demineralization of human enamel was induced via S. mutans-C. albicans biofilms, and enamel hardness was measured. Compared to CHX and UC groups, the baicalein group achieved the greatest reduction in S. mutans, C. albicans and S. mutans-C. albicans biofilms, yielding the least metabolic activity, polysaccharide synthesis and lactic acid production (p < 0.05). The biofilm CFU was decreased in baicalein group by 5 logs, 4 logs, 5 logs, for S. mutans, C. albicans and S. mutans-C. albicans biofilms, respectively, compared to UC group. When tested in a S. mutans-C. albicans in vitro caries model, the baicalein group substantially reduced enamel demineralization under biofilms, yielding an enamel hardness that was 2.75 times greater than that of UC group. Hence, the novel baicalein method is promising to inhibit dental caries by reducing biofilm formation and protecting enamel hardness.     

Anti-Bacterial Effect of Cannabidiol against the Cariogenic Streptococcus mutans Bacterium: An In Vitro Study

Dental caries is caused by biofilm-forming acidogenic bacteria, especially Streptococcus mutans, and is still one of the most prevalent human bacterial diseases. The potential use of cannabidiol (CBD) in anti-bacterial therapies has recently emerged. Here we have studied the anti-bacterial and anti-biofilm activity of CBD against S. mutans. We measured minimum inhibitory concentration (MIC) and minimum biofilm inhibitory concentration (MBIC). The bacterial growth and changes in pH values were measured in a kinetic study. The biofilm biomass was assessed by Crystal Violet staining and 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2H-tetrazolium bromide (MTT) metabolic assay. Spinning Disk Confocal Microscopy (SDCM) was used to assess biofilm structure, bacterial viability and extracellular polysaccharide (EPS) production. CBD inhibited S. mutans planktonic growth and biofilm formation in a dose-dependent manner, with similar MIC and MBIC values (5 µg/mL). CBD prevented the bacteria-mediated reduction in pH values that correlated with bacterial growth inhibition. SDCM showed a decrease of 50-fold in live bacteria and EPS production. CBD significantly reduced the viability of preformed biofilms at 7.5 µg/mL with an 80 ± 3.1% reduction of metabolic activity. At concentrations above 20 µg/mL, there was almost no bacterial recovery in the CBD-treated preformed biofilms even 48 h after drug withdrawal. Notably, precoating of the culture plate surfaces with CBD prior to incubation with bacteria inhibited biofilm development. Additionally, CBD was found to induce membrane hyperpolarization in S. mutans. Thus, CBD affects multiple processes in S. mutans including its cariogenic properties. In conclusion, we show that CBD has a strong inhibitory effect against cariogenic bacteria, suggesting that it is a potential drug adjuvant for reducing oral pathogenic bacterial load as well as protecting against dental caries.     

Molasses-Silver Nanoparticles: Synthesis,Optimization, Characterization,and Antibiofilm Activity

Biofilms are matrix-enclosed communities of bacteria that are highly resistant to antibiotics. Adding nanomaterials with antibacterial activity to the implant surfaces may be a great solution against biofilm formation. Due to its potent and widespread antibacterial effect, silver nanoparticles were considered the most potent agent with different biological activities. In the present investigation, silver nanoparticles (AgNPs) were newly synthesized as antibiofilm agents using sugarcane process byproduct (molasses) and named Mo-capped AgNPs. The synthesized nanoparticles showed promising antimicrobial activity against S. aureus ATCC 6538 and C. albicans DAY185. Statistically designed optimization through response surface methodology was evaluated for maximum activity and better physical characteristics, namely the nanoparticles’ size and polydispersity index (PDI), and it was revealed that molasses concentration was the main effective factor. Minimal biofilm eradication concentration (MBEC) of Mo-capped AgNPs against S. aureus ATCC 6538 and C. albicans DAY185 was 16 and 32 µg/mL, respectively. Scanning electron microscope study of Mo-capped AgNP-treated biofilm revealed that AgNPs penetrated the preformed biofilm and eradicated the microbial cells. The optimally synthesized Mo-capped AgNPs were spherically shaped, and the average size diameter ranged between 29 and 88 nm with high proportions of Ag⁺ element (78.0%) recorded. Fourier-transform infrared spectroscopy (FTIR) analysis indicated the importance of molasses ingredients in capping and stabilizing the produced silver nanoparticles.     

Evaluation of the effects of different remineralizing agents on Streptococcus mutans biofilm adhesion

The aim of the study was to compare the effects of different remineralization methods that are well established in clinical and daily use on S. mutans biofilm adhesion. In this study 72 human third molars were used. From each tooth two pieces of 4?mm x 7?mm enamel blocks were acquired. The samples were divided into 6 groups in which include 10 samples per time period (24h and 48?h) and for each remineralization method; control, flouride, ozone, CPP-ACP, arginine, novamin. After remineralization procedures, enamel surfaces were covered with saliva. 10⁵ CFU/mL of active S. mutans culture were inoculated onto the samples. S. mutans colonies were counted with Plate Count Agar (PCA) decimal dilution method. Micromorphologic effects of different remineralization methods were observed by Scanning Electron Microscopy (SEM). The most S. mutans biofilm formation for both time periods was observed in the control group whereas the less biofilm adhesion was showed in the arginine group. There were no statistically significant differences among remineralization agents (p?>?0.05). In the control group there was statistical difference between 24?h and 48?h (p?<?0.005) but in the other study groups there were no significant difference between the time periods (p?>?0.05). Remineralization agents did not significant differ on S. mutans biofilm adhesion.     

In Silico Selection and In Vitro Evaluation of New Molecules That Inhibit the Adhesion of Streptococcus mutans through Antigen I/II

Streptococcus mutans is the main early colonizing cariogenic bacteria because it recognizes salivary pellicle receptors. The Antigen I/II (Ag I/II) of S. mutans is among the most important adhesins in this process, and is involved in the adhesion to the tooth surface and the bacterial co-aggregation in the early stage of biofilm formation. However, this protein has not been used as a target in a virtual strategy search for inhibitors. Based on the predicted binding affinities, drug-like properties and toxicity, molecules were selected and evaluated for their ability to reduce S. mutans adhesion. A virtual screening of 883,551 molecules was conducted; cytotoxicity analysis on fibroblast cells, S. mutans adhesion studies, scanning electron microscopy analysis for bacterial integrity and molecular dynamics simulation were also performed. We found three molecules ZINC19835187 (ZI-187), ZINC19924939 (ZI-939) and ZINC19924906 (ZI-906) without cytotoxic activity, which inhibited about 90% the adhesion of S. mutans to polystyrene microplates. Molecular dynamic simulation by 300 nanoseconds showed stability of the interaction between ZI-187 and Ag I/II (PDB: 3IPK). This work provides new molecules that targets Ag I/II and have the capacity to inhibit in vitro the S. mutans adhesion on polystyrene microplates.     

Superhydrophobic PDMS/SiNPs/T-ZnOw coating with reduced adhesion of Streptococcus mutans for dental caries prevention

The prevention of dental caries is based mainly on killing the cariogenic bacteria Streptococcus mutans. Prevention of S. mutans adhesion through the development of physical structures is rarely utilized. In this study, a superhydrophobic PDMS/SiNPs/T-ZnOw (PST) coating was prepared for use on a bovine tooth by mixing polydimethylsiloxane (PDMS), silicon dioxide nanoparticles (SiNPs), and tetrapod-like zinc oxide whiskers (T-ZnOw) using one-pot solution and spray methods. The results showed that the superhydrophobicity and roughness of the coating, affected by the PDMS content, were positively correlated with the anti-adhesive effect on S. mutans. The PST coating with PDMS, SiNPs, and T-ZnOw at a ratio of 2.5:1:1 exhibited the highest water contact angle (161°) and the best anti-adhesion effect (97.2% at 4 h and 98.1% at 12 h). The anti-adhesion property towards S. mutans was attributed to its needle-like structure, and the biofilm live-dead staining test showed that the coating had no bactericidal effect. In addition, the coating exhibited favorable durability and biocompatibility, providing a solid foundation for application in the human oral cavity. Thus, this study provides an effective method for caries prevention.     

Combined application of 13C NMR spectroscopy and confocal laser scanning microscopy—Investigation on biofilm structure and physico-chemical properties

Long-term biofilm processes are influenced by the interplay of biofilm accumulation and detachment, which in turn depend partially on the biofilm structure and composition. In this study a combination of confocal laser scanning microscopy (CLSM) and nuclear magnetic resonance (NMR) spectroscopy was applied to analyze biofilm structure, composition and molecular mobility. Whereas CLSM delivers information about the structure of biofilms the NMR measurement provides detailed but not locally resolved information about the chemical composition of biofilm constituents. Heterotrophic mixed-species biofilms were cultivated in rotating annular reactors exposed to different flow conditions and glucose concentrations in order to obtain biofilms with diverse architectural structures. The growth state of the biofilms appeared to influence the composition of biofilm and detached biomass. The difference in the ¹³C NMR spectra between the differently structured biofilms or between biofilm and detached biomass was small, except for the still exponential growing biofilm supplied with the highest glucose concentration. More information was gained from the mobility of specific molecular groups within the biofilm biomass. Molecules within the biofilm biomass of the non-filamentous biofilms were more strongly bound than the molecules within the respective detached biomass. Glucose starvation resulted in a reduction in the biofilm molecular mobility. The opposite was observed in the filamentous biofilm. In this case, the molecular mobility in the biofilm increased after starvation and the molecules in the detached biomass were bound more strongly than in the respective biofilm biomass. It could be shown that the combination of CLSM and ¹³C NMR spectroscopy is a promising approach to analyze the interactions between biofilm architecture, composition or growth state and biofilm detachment.     

Biodegradation of caries-affected dentin bonding interface of fluoride and MDPB-containing adhesive system

The aim of this study was to evaluate the effect of a fluoride/MDPB (12-methacryloyloxydodecylpyridinium bromide)-containing adhesive system on the durability of a bond to permanent artificially induced caries-affected dentin (CAD) exposed to Streptococcus mutans culture and water storage. Twelve third molars were selected. Flat dentin surfaces were submitted to artificial caries development in S. mutans and Broth heart infusion (BHI). Caries-infected dentin was removed with burs according to clinical criteria and CAD cavities were restored with Adper Scotchbond Multi-Purpose (SBM) and Clearfil Protect Bond (CPB). Nontrimmed resin–dentin bonded interfaces (1 mm²) were stored in S. mutans+BHI for 3 days, in deionized water for 3 months, and afterwards subjected to microtensile bond strength test (μTBS). The control group was not submitted to storage and immediate μTBS testing was performed. Fractographic analysis was performed after μTBS testing. Four molars were restored as described, and morphological evaluation hybrid layer (HL) was performed by scanning electron microscopy (SEM). Two-way ANOVA with split-plot design and Tukey′s tests were performed. No difference was found between μTBS values of SBM and CPB irrespective of groups. Significant decrease was observed in μTBS values after S. mutans culture and water storage, but without difference between them. CPB had more homogenous hybrid layer than SBM. Fluoride/MDPB-containing adhesive system did not prevent degradation of CAD bond strength in both degradation methods.     

The Interaction of CuS and Halothiobacillus HT1 Biofilm in Microscale Using Synchrotron Radiation-Based Techniques

In order to investigate the microbe-mineral interaction in the micro scale, spatial distribution and speciation of Cu and S in Halothiobacillus HT1 biofilm formed on a CuS surface was examined using synchrotron-based X-ray techniques. Confocal laser scanning microscope (CLSM) results indicated that Halothiobacillus HT1 biofilm formation gave rise to distinct chemical and redox gradients, leading to diverse niches in the biofilm. Live cells were distributed at the air-biofilm and membrane-biofilm interface. CuS was oxidized by Halothiobacillus HT1 biofilm, and copper penetrated into the biofilm. Sulfide was oxidized to cysteine (77.3%), sulfite (3.8%) and sulfonate (18.9%). Cu-cysteine-like species were involved in the copper homeostasis. These results significantly improve our understanding of the interfacial properties of the biofilm-mineral interface.     

Cationic Phenosafranin Photosensitizers Based on Polyhedral Oligomeric Silsesquioxanes for Inactivation of Gram-Positive and Gram-Negative Bacteria

The high photodynamic effect of the Newman strain of the S. aureus and of clinical strains of S. aureus MRSA 12673 and E. coli 12519 are observed for new cationic light-activated phenosafranin polyhedral oligomeric silsesquioxane (POSS) conjugates in vitro. Killing of bacteria was achieved at low concentrations of silsesquioxanes (0.38 µM) after light irradiation (λ_{em. max} = 522 nm, 10.6 mW/cm²) for 5 min. Water-soluble POSS-photosensitizers are synthesized by chemically coupling a phenosafranin dye (PSF) (3,7-diamino-5-phenylphenazine chloride) to an inorganic silsesquioxane cage activated by attachment of succinic anhydride rings. The chemical structure of conjugates is confirmed by ¹H, ¹³C NMR, HRMS, IR, fluorescence spectroscopy and UV-VIS analyzes. The APDI and daunorubicin (DAU) synergy is investigated for POSSPSFDAU conjugates. Confocal microscopy experiments indicate a site of intracellular accumulation of the POSSPSF, whereas iBuPOSSPSF and POSSPSFDAU accumulate in the cell wall or cell membrane. Results from the TEM study show ruptured S. aureus cells with leaking cytosolic mass and distorted cells of E. coli. Bacterial cells are eradicated by ROS produced upon irradiation of the covalent conjugates that can kill the bacteria by destruction of cellular membranes, intracellular proteins and DNA through the oxidative damage of bacteria.     

The Effect of a Silver Nanoparticle Polysaccharide System on Streptococcal and Saliva-Derived Biofilms

In this work, we studied the antimicrobial properties of a nanocomposite system based on a lactose-substituted chitosan and silver nanoparticles: Chitlac-nAg. Twofold serial dilutions of the colloidal Chitlac-nAg solution were both tested on Streptococcus mitis, Streptococcus mutans, and Streptococcus oralis planktonic phase and biofilm growth mode as well as on saliva samples. The minimum inhibitory and bactericidal concentrations of Chitlac-nAg were evaluated together with its effect on sessile cell viability, as well as both on biofilm formation and on preformed biofilm. In respect to the planktonic bacteria, Chitlac-nAg showed an inhibitory/bactericidal effect against all streptococcal strains at 0.1% (v/v), except for S. mitis ATCC 6249 that was inhibited at one step less. On preformed biofilm, Chitlac-nAg at a value of 0.2%, was able to inhibit the bacterial growth on the supernatant phase as well as on the mature biofilm. For S. mitis ATCC 6249, the biofilm inhibitory concentration of Chitlac-nAg was 0.1%. At sub-inhibitory concentrations, the Streptococcal strains adhesion capability on a polystyrene surface showed a general reduction following a concentration-dependent-way; a similar effect was obtained for the metabolic biofilm activity. From these results, Chitlac-nAg seems to be a promising antibacterial and antibiofilm agent able to hinder plaque formation.     

Rapid Hormetic Responses of Photosystem II Photochemistry of Clary Sage to Cadmium Exposure

Five-day exposure of clary sage (Salvia sclarea L.) to 100 μM cadmium (Cd) in hydroponics was sufficient to increase Cd concentrations significantly in roots and aboveground parts and affect negatively whole plant levels of calcium (Ca) and magnesium (Mg), since Cd competes for Ca channels, while reduced Mg concentrations are associated with increased Cd tolerance. Total zinc (Zn), copper (Cu), and iron (Fe) uptake increased but their translocation to the aboveground parts decreased. Despite the substantial levels of Cd in leaves, without any observed defects on chloroplast ultrastructure, an enhanced photosystem II (PSII) efficiency was observed, with a higher fraction of absorbed light energy to be directed to photochemistry (Φ_PSΙΙ). The concomitant increase in the photoprotective mechanism of non-photochemical quenching of photosynthesis (NPQ) resulted in an important decrease in the dissipated non-regulated energy (Φ_NO), modifying the homeostasis of reactive oxygen species (ROS), through a decreased singlet oxygen (¹O₂) formation. A basal ROS level was detected in control plant leaves for optimal growth, while a low increased level of ROS under 5 days Cd exposure seemed to be beneficial for triggering defense responses, and a high level of ROS out of the boundaries (8 days Cd exposure), was harmful to plants. Thus, when clary sage was exposed to Cd for a short period, tolerance mechanisms were triggered. However, exposure to a combination of Cd and high light or to Cd alone (8 days) resulted in an inhibition of PSII functionality, indicating Cd toxicity. Thus, the rapid activation of PSII functionality at short time exposure and the inhibition at longer duration suggests a hormetic response and describes these effects in terms of “adaptive response” and “toxicity”, respectively.     

Efficacy of a Topical Wound Agent Methanesulfonic Acid and Dimethylsulfoxide on In Vitro Biofilms

A topical desiccating wound agent containing methanesulfonic acid, dimethylsulfoxide and amorphous silica was evaluated in three in vitro models for its efficacy against biofilms produced by Pseudomonas aeruginosa (ATCC-15442) and Staphylococcus aureus (ATCC-6538). The in vitro biofilm models used were; the MBEC Assay^®, Centre for Disease Control (CDC) Biofilm Reactor^® and a Semi-solid biofilm model. A 30-s exposure of a topical wound desiccating agent was used in each model. A complete eradication of viable cells was demonstrated in all models for both strains (p < 0.0001). Imaging with scanning electron microscopy (SEM) was performed where possible. All three models demonstrated complete eradication of viable cells with a 30 s application of a topical wound desiccating agent.