Natural photosensitizers potentiate the targeted antimicrobial photodynamic therapy as the Monkeypox virus entry inhibitors: An in silico approach.

BACKGROUND: Monkeypox is a viral zoonotic disease that has emerged as a threat to public health. Currently, there is no treatment approved specifically targeting Monkeypox disease. Hence, it is essential to identify and develop therapeutic approaches to the Monkeypox virus. In the current in silico paper, we comprehensively involve using computer simulations and modeling to insights and predict hypotheses on the potential of natural photosensitizers-mediated targeted antimicrobial photodynamic therapy (aPDT) against D8L as a Monkeypox virus protein involved in viral cell entry. MATERIALS AND METHODS: In the current study, computational techniques such as molecular docking were combined with in silico ADMET predictions to examine how Curcumin (Cur), Quercetin (Qct), and Riboflavin (Rib) as the natural photosensitizers bind to the D8L protein in Monkeypox virus, as well as to determine pharmacokinetic properties of these photosensitizers. RESULTS: The three-dimensional structure of the D8L protein in the Monkeypox virus was constructed using homology modeling (PDB ID: 4E9O). According to the physicochemical properties and functional characterization, 4E9O was a stable protein with the nature of a hydrophilic structure. The docking studies employing a three-dimensional model of 4E9O with natural photosensitizers exhibited good binding affinity. D8L protein illustrated the best docking score (-7.6 kcal/mol) in relation to the Rib and displayed good docking scores in relation to the Cur (-7.0 kcal/mol) and Qct (-7.5 kcal/mol). CONCLUSIONS: The findings revealed that all three photosensitizers were found to obey the criteria of Lipinski's rule of five and displayed drug-likeness. Moreover, all the tested photosensitizers were found to be non-hepatotoxic and non-cytotoxic. In summary, our investigation identified Cur, Qct, and Rib could efficiently interact with D8L protein with a strong binding affinity. It can be concluded that aPDT using these natural photosensitizers may be considered an adjuvant treatment against Monkeypox disease.

Virtual screening and computational simulation analysis of antimicrobial photodynamic therapy using propolis-benzofuran A to control of Monkeypox.

BACKGROUND: Monkeypox is a viral zoonotic disease and there are no available treatments that specifically target the monkeypox virus. Antimicrobial photodynamic therapy (aPDT) is a non-invasive approach that has been introduced as a targeted adjuvant treatment against various microbial infections. In this study, we used a computational strategy to investigate the potential of aPDT using propolis-benzofuran A against the Monkeypox virus. METHODS: In this in silico study, the evaluation of drug-likeness, molecular properties, and bioactivity of propolis-benzofuran A was carried out using SwissADME. Pro-Tox II and OSIRIS servers were used to identify the organ toxicities and toxicological endpoints of propolis-benzofuran A. Molecular docking approach was employed to screen the potential binding modes of propolis-benzofuran A ligand with the Monkeypox virus A48R protein (PDB ID: 2V54). RESULTS: The results of the computational investigation revealed that propolis-benzofuran A obeyed all the criteria of Lipinski's rule of five and exhibited drug-likeness. The photosensitizing agent tested was categorized as toxicity class-5 and was found to be non-hepatotoxic, non-carcinogenic, non-mutagenic, and non-cytotoxic. The docking studies employing a predicted three-dimensional model of Monkeypox virus A48R protein with propolis-benzofuran A ligand exhibited good binding affinity (-7.84 kcal/mol). DISCUSSION: The computational simulation revealed that propolis-benzofuran A had a strong binding affinity with the Monkeypox virus A48R protein. Hence, aPDT based on this natural photosensitizer can be proposed as an adjuvant treatment against the Monkeypox virus.

Molecular Modeling and Simulation Analysis of Antimicrobial Photodynamic Therapy Potential for Control of COVID-19.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) can enter the host cells by binding the viral surface spike glycoprotein (SG) to angiotensin-converting enzyme 2. Since antiviral photodynamic therapy (aPDT) has been described as a new method for inhibiting viral infections, it is important to evaluate whether it can be used as a photoactivated disinfectant to control COVID-19. In this in silico study, SARS-CoV-2-SG was selected as a novel target for curcumin as a photosensitizer during aPDT to exploit its physicochemical properties, molecular modeling, hierarchical nature of protein structure, and functional analysis using several bioinformatics tools and biological databases. The results of a detailed computational investigation revealed that SARS-CoV-2-SG is most similar to 6VXX_A, with 100% query cover and identity. The predicted structure of SARS-CoV-2-SG displayed that it is a protein with a positive charge and random coil dominates other secondary structures located outside the viral cell. The protein-protein interaction network showed that SARS-CoV-2-SG interacted with ten potential interacting partners. In addition, primary screening of binding modes through molecular docking showed that curcumin desires to bind and interact with residues of SARS-CoV-2-SG as the main site to enhance the yield of aPDT. Overall, the computer simulation reveals that SARS-CoV-2-SG can be a suitable target site for interaction with curcumin during aPDT.

The synergistic effect of Nano-propolis and curcumin-based photodynamic therapy on remineralization of white spot lesions: An ex vivo study.

OBJECTIVE: One of the major problems in fixed orthodontic treatments is the control of enamel demineralization, white spots lesions (WSL), and dental caries around the brackets, which can be stopped by strengthening the remineralization process. The aim of this study was to investigate the synergistic effect of propolis nanoparticles (NPro) and nano-curcumin-based photodynamic therapy (NCur-PDT) in the remineralization of WSL ex vivo. MATERIALS AND METHODS: Experimental period was 5.5 months. After synthesis and characterization of NPro, the extracted bovine teeth were demineralized using a demineralization solution. They were divided into 7 groups (n=10) and treated in the following groups: 1) Negative control (artificial saliva), 2) Positive control or control of treatment (2% neutral sodium fluoride gel; NSF), 3) Nano-curcumin (NCur), 4) NPro, 5) Diode laser irradiation (light), 6) NCur with irradiation (NCur-PDT) and 7) NPro plus NCur-PDT (NPro+NCur-PDT). The treatment duration was 3 months and each treatment was conducted on T1 (the end of the third week), T2 (the end of the sixth week), T3 (the end of the ninth week), and T4 (the end of the twelfth week). Then, the microhardness, surface changes, and surface topography of the enamel were examined using digital hardness tester, DIAGNOdent Pen Reading, and scanning electron microscope (SEM), respectively. RESULTS: In all groups, after demineralization, the microhardness of tooth enamel decreased and after each of the treatments, the amount of microhardness significantly increased over time except light and artificial saliva groups. The earliest significant change in microhardness improvement was observed in the NPro+NCur-PDT group at T2 (P<0.05). Also, at T2, DIAGNODent Pen revealed the earliest significant improved change in the level of mineralization degree in the NPro+NCur-PDT group. NCur and light irradiation groups alone, like the control group (artificial saliva), did not lead to remineralization (P>0.05). Also, the NPro+NCur-PDT treatment results obtained from SEM after the last treatment in the twelfth week showed that remineralization in that group has significantly improved compared to other groups. CONCLUSION: The results of this study showed that combined use of NPro and NCur-PDT had more enamel remineralization efficacy in a shorter period. Simultaneous application of NPro and NCur-PDT had also a stronger therapeutic effect after 3 months.

Contribution of antimicrobial photo-sonodynamic therapy in wound healing: an in vivo effect of curcumin-nisin-based poly (L-lactic acid) nanoparticle on Acinetobacter baumannii biofilms.

BACKGROUND: The biofilm-forming ability of Acinetobacter baumannii in the burn wound is clinically problematic due to the development of antibiotic-resistant characteristics, leading to new approaches for treatment being needed. In this study, antimicrobial photo-sonodynamic therapy (aPSDT) was used to assess the anti-biofilm efficacy and wound healing activity in mice with established A. baumannii infections. METHODS: Following synthesis and confirmation of Curcumin-Nisin-based poly (L-lactic acid) nanoparticle (CurNisNp), its cytotoxic and release times were evaluated. After determination of the sub-significant reduction (SSR) doses of CurNisNp, irradiation time of light, and ultrasound intensity against A. baumannii, anti-biofilm activity and the intracellular reactive oxygen species (ROS) generation were evaluated. The antibacterial and anti-virulence effects, as well as, histopathological examination of the burn wound sites of treated mice by CurNisNp-mediated aPSDTSSR were assessed and compared with silver sulfadiazine (SSD) as the standard treatment group. RESULTS: The results showed that non-cytotoxic CurNisNp has a homogeneous surface and a sphere-shaped vesicle with continuous release until the 14th day. The dose-dependent reduction in cell viability of A. baumannii was achieved by increasing the concentrations of CurNisNp, irradiation time of light, and ultrasound intensity. There was a time-dependent reduction in biofilm growth, changes in gene expression, and promotion in wound healing by the acceleration of skin re-epithelialization in mice. Not only there was no significant difference between aPSDTSSR and SSD groups in antibacterial and anti-virulence activities, but also wound healing and re-epithelialization occurred more efficiently in aPSDTSSR than in the SSD group. CONCLUSIONS: In conclusion, CurNisNp-mediated aPSDT might be a promising complementary approach to treat burn wound infections.

Antimicrobial action of photodynamic therapy on Enterococcus faecalis biofilm using curing light, curcumin and riboflavin.

The aim of this study was to assess the effect of antimicrobial photodynamic therapy (aPDT) with curcumin and riboflavin on three-week Enterococcus faecalis biofilm. At first the 15-mm root canals of 65 single rooted extracted human teeth (including maxillary incisors, mandibular and maxillary canines and mandibular premolars) were separated from the crown and were prepared with ProTaper instruments. After autoclave sterilisation, samples were inoculated with E. faecalis suspension, and incubated for three weeks. After ensuring biofilm formation by scanning electron microscopy (SEM) in two teeth, the remaining 63 teeth were randomly divided into seven groups (n = 9): aPDT + curcumin, aPDT + riboflavin, LED, curcumin, riboflavin, 5.25% NaOCl (positive control) and no intervention (negative control). For light source a LED unit with 390-480 nm wavelength (peak of 460 nm), power density of 1000 ± 100 mW cm-2 and mean energy density of 60 J cm-2 was used. The roots were horizontally sectioned into coronal, middle and apical thirds each with 5 mm thicknesses. Dentin chips with equal weight (1 ± 0.005 g) were collected from the root canal walls with Gates Glidden drills and were transferred into microtubes containing 1 mL of sterile saline and vortexed for 30 s. Next, 10 µL of the contents of each tube was serially diluted and eventually, 10 µL of each solution was cultured on BHI agar. The number of colony-forming units was determined. Data were analysed using the Kruskal-Wallis and Friedman tests. The colony reduction was not significantly different between NaOCl and either riboflavin + LED or Curcumin + LED. The 5.25% NaOCl group showed maximum reduction in colony count, compared with the negative control (P = 0.00). Groups with aPDT with Curcumin + LED (P = 0.005), and with riboflavin + LED (P = 0.011) showed significant reduction in colony count in all three canal thirds (P < 0.05) without any difference with one another. With significant reduction of E. faecalis colony count, aPDT with Curcumin and riboflavin can serve as an adjunct to routine root canal disinfection method.

Robust antimicrobial photodynamic therapy with curcumin-poly (lactic-co-glycolic acid) nanoparticles against COVID-19: A preliminary in vitro study in Vero cell line as a model.

BACKGROUND: In this study, the ability of antimicrobial photodynamic therapy (aPDT) as a treatment approach and adjuvant therapy using curcumin-poly (lactic-co-glycolic acid) nanoparticles (Cur@PLGA-NPs) to inactivate Coronavirus disease 2019 (COVID-19) in plasma was investigated. Furthermore, to verify whether the quality requirement of aPDT-treated plasma is acceptable, the differences of the levels of clotting factors, total plasma proteins, and anti-A and/or anti-B antibodies titrations in plasma of patient before and after aPDT treatment were investigated. MATERIALS AND METHODS: Cur@PLGA-NPs was synthesized using Electrospinning process and characterized by different analysis including Scanning Electron Microscope (SEM), Transmission Electron Microscope (TEM), and Fourier Transform Infrared (FTIR) spectroscopy assays. The presence of the SARS-CoV-2 in the plasma samples of patients suspected of having COVID-19 was confirmed by real-time reverse transcription-polymerase chain reaction (RT-PCR) assay. Then, the treated plasma samples with Cur@PLGA-NPs plus blue laser were exposed to Vero cells. Eventually, cell cytotoxicity and apoptotic effects of treated Vero cells were evaluated. Levels of clotting factors including prothrombin time (PT) and activated partial thromboplastin time (APTT), total plasma proteins, and anti-A and/or anti-B antibodies measurements were performed using the coagulometer, method of Bradford, and titration procedure, respectively. RESULTS: The presence of SARS-CoV-2 was positive in 84.3 % of samples. Different concentrations of Cur@PLGA-NPs (3, 5, 7, and 10 % wt.), the irradiation times of blue laser (1, 3, and 5 min), and aPDT with the maximum dosed of blue laser light (522.8 J/cm2) plus 10 % wt. Cur@PLGA-NPs had no cytotoxicity. Although there were significant cell degradation and apoptotic effects in treated Vero cells with treated plasma using 10 % wt. Cur@PLGA-NPs, and a blue laser at an energy density of 522.8 J/cm2, no visible changes in cells and apoptosis were observed following aPDT. Total plasma protein content, PT, APTT, and anti-A and/or anti-B antibodies titers showed no significant changes (P > 0.05 for all comparisons) in treated plasma as compared to untreated plasma. CONCLUSION: aPDT exhibited in vitro anti-COVID-19 activities in the treated plasma containing SARS-COV-2 without Vero cell apoptosis and any adverse effects on plasma quality in aPDT-exposed plasma.

In Vitro Antimicrobial and Cytotoxicity Activities of Some Medicinal Plant Extracts against Oral Microbial Pathogens.

INTRODUCTION: Medicinal plants have long been of great interest to scientists in the search for the best treatment of diseases, especially the infectious diseases. In recent years, the use of herbal medicines has become more well-known because of their antimicrobial, antifungal, anti-cancer and less side effects. AIM: The aim of this study was to investigate the antimicrobial and antifungal effects of Urtica dioica, Equisetum arvense, and Punica Granatum peel extracts on two common oral microorganisms, Streptococcus mutans and Candida albicans. MATERIALS AND METHODS: The study investigated the hydro-alcoholic extract of the plants. The antimicrobial activity of the extracts was evaluated using the method of measuring the inhibition of microorganisms, and the minimum inhibitory concentration (MIC) and the minimum bactericidal concentration (MBC) were determined using different concentrations of the extracts and also biofilm assay and SEM were determined. Also cell viability was assessed by MTT assay on human gingival fibroblast cells. RESULTS: The lowest MIC against S. mutants and C. albicans was related to the hydro-alcoholic extract of U. dioica. There was a significant reduction in the microbial biofilms by all three extracts. Among them, U. dioica could decrease the biofilms of S. mutans and C. albicans more than other extracts. In addition, the best results for growth inhibition zone were the hydro-alcoholic extracts of E. arvense and U. dioica with 35 and 30 mm growth zone, respectively. The results of SEM showed that P. granatum peel, U. dioica and E. arvense could destroy microbial biofilms without exerting any cytotoxic effects on HGF cell. CONCLUSIONS: The results of the study suggest that U. dioica, E. arvense, and P. Granatum peel extracts can be used as mouthwash with the least significant difference with routine mouthwashes. Also, the plant-based mouthwashes may be more suitable substitutes for chemical types in the future.

In vitro antibacterial activity and durability of a nano-curcumin-containing pulp capping agent combined with antimicrobial photodynamic therapy.

BACKGROUND: Considering the antibacterial properties of nano-curcumin (nCur) reinforced with antimicrobial photodynamic therapy (aPDT), this study aimed to assess the antibacterial activity and durability of Activa BioActive Base/Liner (ABBL) containing nCur (nCur-ABBL) as a pulp capping agent against Streptococcus mutans, the most common cause of secondary caries. MATERIALS AND METHODS: In this in vitro experimental study, ABBL discs containing 0.5 %, 1%, 2%, and 5% (w/w) concentrations of nCur were fabricated. After aPDT using light emitting diode (LED) at 435 ± 20 nm wavelength for 5 min, the discs were undergone aging in artificial saliva for 90 days. The antibacterial activity of the discs against S. mutans was evaluated by the disc agar diffusion test, and the number of bacterial colonies present in the biofilm formed on the disc surfaces was counted after 0, 15, 30, and 60 days of aging. RESULTS: The maximum growth inhibition zone was noted around the 5% nCur-ABBL discs. Increasing the concentration of nCur from 0.5 % to 5% combined with aPDT significantly decreased the number of S. mutans colonies in the biofilm over time (P < 0.05). nCur-ABBL discs containing 2% and 5% nCur had no difference in antibacterial activity at any time point up to 60 days (P > 0.05). CONCLUSION: According to our data, 5% nCur-ABBL revealed the largest growth inhibition zone in S. mutans culture. Moreover, 5% nCur can serve as an excellent ABBL additive in aPDT producer against S. mutans biofilms up to 60 days of aging period.

Propolis nanoparticle enhances the potency of antimicrobial photodynamic therapy against Streptococcus mutans in a synergistic manner.

Less invasive removal approaches have been recommended for deep caries lesions. Antimicrobial photodynamic therapy (aPDT) and propolis nanoparticle (PNP) are highlighted for the caries management plan. Evidence is lacking for an additive effect of combination PNP with photosensitizer (PS) in aPDT. This study aimed to investigate the individual and synergistic effects of chlorophyllin-phycocyanin mixture (PhotoActive+) and toluidine blue O (TBO) as PSs in combination with PNP in the aPDT process (aPDTplus) against major important virulence factors of Streptococcus mutans. Following characterization, biocompatibility of the PSs alone, or in combination with PNP were investigated on human gingival fibroblast cell. The in vitro synergy of PhotoActive+ or TBO and PNP was evaluated by the checkerboard method. The bacteria's virulence properties were surveyed in the presence of the PSs, individually as well as in combination. When the PSs were examined in combination (synergistic effect, FIC Index < 0.5), a stronger growth inhibitory activity was exhibited than the individual PSs. The biofilm formation, as well as genes involved in biofilm formation, showed greater suppression when the PSs were employed in combination. Overall, the results of this study suggest that the combination of PhotoActive+ or TBO with PNP with the least cytotoxicity effects and the highest antimicrobial activites would improve aPDT outcomes, leading to synergistic effects and impairing the virulence of S. mutans.

Anti-biofilm and anti-metabolic effects of antimicrobial photodynamic therapy using chlorophyllin-phycocyanin mixture against Streptococcus mutans in experimental biofilm caries model on enamel slabs.

BACKGROUND: Partial (selective) removal of dental caries is a suitable manner to treat deep carious lesions in vital teeth with asymptomatic pulps. Antimicrobial photodynamic therapy (aPDT) was proposed as a promising ancillary approach for reduction of the residual bacteria from the cavity. Therefore, the focus of this study was to investigate the influence of aPDT using diode laser (DL) plus PhotoActive+ (chlorophyllin-phycocyanin mixture [CHL-PC]) as photosensitizer (PS) on metabolic activity and the reduction in the number of living bacteria within the preformed biofilm caries model on enamel slabs of Streptococcus mutans. MATERIALS AND METHODS: The lethal and sub-significant inhibitory (SSI) potential of aPDT using CHL-PC and 635 nm DL against experimental biofilm caries model on enamel slabs and metabolic activity of S. mutans was analyzed using crystal violet and XTT reduction assays, respectively. Intracellular ROS formation by DCFH-DA assay was measured in CHL-PC mediated aPDT treated bacterial samples. Tooth discoloration and cell cytotoxicity of CHL-PC were assessed in the CIEL*a*b* color space and neutral red assay, respectively. RESULTS: In this study aPDT at a maximum concentration level of CHL-PC (5000 µg/mL) with 3 min DL irradiation time (103.12 J/cm2) reduced the ex-vivo cariogenic biofilm of S. mutans by 36.93 % (P <  0.05). Although chlorhexidine (CHX) had an anti-biofilm effect about 1.7 fold compared to CHL-PC mediated aPDT, this difference was not significant (36.93 in comparison to 63.05 %; P >  0.05). CHL-PC mediated aPDT demonstrated a significant reduction in bacterial metabolic activity, with rates of 77 % at a SSI dose (using 156 µg/mL of CHL-PC and 3 min DL irradiation time with the energy density of 103.12 J/cm2). The treated bacterial cells exhibited significant (P < 0.05) increment in the ROS generation. The least color change (ΔE) was found using CHL-PC at a concentration of 156 µg/mL (ΔE = 2.74). CHL-PC in different concentrations showed no significant reduction in human gingival fibroblasts (HGFs) cell survival (P >  0.05). CONCLUSION: CHL-PC mediated aPDT not only reduces the number of living bacteria within the biofilms of S. mutans in an experimental biofilm caries model on enamel slabs but also its influences microbial virulence by reducing the metabolic activity of the S. mutants.

Dual wavelength irradiation antimicrobial photodynamic therapy using indocyanine green and metformin doped with nano-curcumin as an efficient adjunctive endodontic treatment modality.

BACKGROUND: Indocyanine green (ICG) doped with nano-curcumin (N-CUR) can increase photosensitivity in antimicrobial photodynamic therapy (aPDT). Since metformin (Met) provides a synergistic advantage with photosensitivity, it was conjugated with N-CUR@ICG. Aim of the study was to evaluate the photosensitizing effect of N-CUR@ICG-Met used as a new photosensitizer in dual wavelengths irradiation (diode laser and light-emitting diode, LED) aPDT in root canals infected with Enterococcus faecalis biofilm. MATERIALS AND METHODS: Following synthesis and confirmation of N-CUR@ICG-Met by Scanning electrone microscope (SEM), Fourier transform infrared (FT-IR) spectroscopy, X-ray diffraction (XRD) and Zetasizer analysis, the mature microbial biofilm was formed. The quantitative and qualitative evaluations of E. faecalis biofilm were made using microbial viability and SEM analysis of the following groups of treatment modalities (n = 5): 1- N-CUR, 2- ICG, 3- Met, 4- N-CUR@ICG, 5- N-CUR@ICG-Met, 6- Diode laser, 7- LED, 8- aPDT/diode, 9- aPDT/LED, 10- aPDT/diode laser + LED, 11- aPDT/LED + diode laser, 12- 2.5 % sodium hypochlorite (NaOCl). In addition, E. faecalis not treated served as negative control. Data were statistically analyzed using One-way Analysis of Variance (ANOVA) and the post-Hoc Bonferroni tests with a level of significance set at P < 0.05. RESULTS: SEM, FT-IR, XRD, Zetasizer analysis confirmed successful doping of ICG-Met onto/into N-CUR. The treatment modality of N-CUR@ICG-Met mediated aPDT/diode laser, aPDT/LED, aPDT/diode laser + LED, and aPDT/LED + diode laser statistically decreased the cell viability of E. faecalis for 69.40 %, 75.52 %, 82.74 %, and 83.84 %, respectively compared with the negative control group (P < 0.05). The double wavelengths irradiation technique, which exposed the N-CUR@ICG-Met with diode laser irradiation and immediately after with LED (i.e. group No. 10), as well as, N-CUR@ICG-Met with LED irradiation and immediately after with diode laser (i.e. group No. 11) reduced the viable E. faecalis count in biofilm structures statistically more than the other treatment modalities (P < 0.05). CONCLUSIONS: N-CUR@ICG-Met as a new photosensitizer in dual wavelengths irradiation method may improve anti-biofilm activity of aPDT against E. faecalis.

Antimicrobial properties of acrylic resins doped with Undaria pinnatifida exposed to light-emitting diode: In silico and in vitro assessments on multispecies biofilm-producing microbiota.

BACKGROUND: This study sought to evaluates the efficiency of anti-microbial activity of acrylic resins doped with different concentrations of Undaria pinnatifida after activation with light-emitting diode (LED) at producing photodynamic damage to multispecies biofilm-producing microbiome. MATERIAL AND METHODS: In this study, bioinformatics tools and computer simulation molecular modeling were used to evaluate the capacity of ferredoxin (FDX), an electron acceptor in metabolic pathways of U. pinnatifida, which can discharge electrons produced from photo-excited chlorophyll-a (Chl-a) by LED irradiation. Acrylic resin discs containing different concentration of U. pinnatifida (0, 0.5, 1, and 2%) were fabricated and were subjected to LED irradiation immediately before each experiment. After continuously rinsed (up to 30 days), the antimicrobial activity of acrylic resins doped with U. pinnatifida following photo-activation was determined by disc agar diffusion, biofilm formation inhibition, and eluted component assays versus bacterial species linked to caries that constitute a mixed biofilm including Streptococcus mutans, S. sanguinis, and Lactobacillus acidophilus, as well as Candida albicans as main etiology of candidal stomatitis. RESULTS: Modeling and a virtual screening analysis of FDX indicated that it is a stable protein with an iron-sulfur center that can discharge electrons produced from photo-excited Chl-a and transfers them to FDX-NADP+ reductase for NADP+ reduction in photosystem I, which is essential in the Calvin cycle for carbon assimilation. FDX acts as an electron transfer agent in the redox reactions. The results showed that growth inhibition zones were not seen around acrylic resin discs in any group. In biofilm test, the colony counts of all test microorganisms significantly decreased (36%-87%) by an increase in the percentage of U. pinnatifida in acrylic resins after photo-activation (P < 0.05). Acrylic resins doped with 2% wt. U. pinnatifida following photo-activation using LED was inhibited biofilm formation by the test microorganisms, up to 30 days of rinsing. CONCLUSION: Based on the results presented here, an acrylic resin containing U. pinnatifida, even at the lowest concentration, following photo-activation using LED have antimicrobial properties against planktonic and biofilm forms of the cariogenic microorganisms as well as C. albicans.

Changes of microbial cell survival, metabolic activity, efflux capacity, and quorum sensing ability of Aggregatibacter actinomycetemcomitans due to antimicrobial photodynamic therapy-induced bystander effects.

BACKGROUND: The bystander effects, whereby naive (bystander) microbial cells near microbial cells directly exposed to certain treatment show responses that would not have happened in the absence of the directly targeted microbial cells, is recently documented in the field of microbiology. In this article, we discuss that substantial bystander responses are also observed after antimicrobial photodynamic therapy (aPDT) using curcumin (Cur). MATERIALS AND METHODS: Bystander effects induced by whole bacterial cell suspension (WBCST), cell-free supernatants fluid (CFSFT), and bacterial cell pellet (BCPT) obtained from A. actinomycetemcomitans culture treated with Cur-aPDT on cell survival, quorum sensing (QS) ability, metabolic activity and efflux capacity of A. actinomycetemcomitans were determined using microbial viability assay, Escherichia coli-based bioassay, XTT reduction method, and ethidium bromide (EtBr) accumulation assay, respectively. RESULTS: A. actinomycetemcomitans cell survival reduced by 82.7% (P = 0.001) and 76.2% (P = 0.01) after exposure to WBCST and CFSFT, respectively. The A. actinomycetemcomitans population increased by 5.5% (P = 0.7) after exposure to BCPT. Bacterial metabolic activity decreased by 42.6% (P = 0.02), 35.3% (P = 0.03), and 9.4% (P = 0.5) after exposure to WBCST, CFSFT, and BCPT, respectively. A. actinomycetemcomitans exposed to WBCST, CFSFT, and BCPT showed a reduction of 83.2% (P = 0.001), 77.2% (P = 0.01) and 21.9% (P = 0.09) in the QS mediator compared to the WBCSU, CFSFU, and BCPU of untreated A. actinomycetemcomitans, respectively. No significant change of the EtBr accumulation was observed in the three preparations of the Cur-aPDT-treated culture (i.e. WBCST, CFSFT, and BCPT) compared to their respective controls. CONCLUSIONS: The results of the current study revealed that Cur-aPDT could significantly reduce microbial cell survival, cell metabolic activity, efflux capacity, and QS ability through the bystander effects. As a result, the bystander effects of Cur-aPDT along with the direct effect of Cur-aPDT can enhance the efficiency of aPDT as an adjunct therapeutic strategy for treatment of local infections.

Physico-mechanical and antimicrobial properties of an orthodontic adhesive containing cationic curcumin doped zinc oxide nanoparticles subjected to photodynamic therapy.

BACKGROUND: Potential complications on the crown level during fixed orthodontic procedures are white spot, enamel demineralization and tooth decay. This study evaluated the antimicrobial properties of an orthodontic adhesive incorporating cationic curcumin doped zinc oxide nanoparticles (cCur/ZnONPs), which can have the highest concentration of cCur/ZnONPs and shear bond strength (SBS) value simultaneously, against cariogenic bacteria including Streptococcus mutans, Streptococcus sobrinus, and Lactobacillus acidophilus. MATERIALS AND METHODS: Following synthesis and confirmation of cCur/ZnONPs, SBS and adhesive remnant index (ARI) of the test adhesives containing cCur/ZnONPs (1.2, 2.5, 5, 7.5, and 10% wt.) were measured using universal testing machine and stereomicroscope, respectively. After continuously rinsed (up to 180 day), the residual antimicrobial ability of modified adhesives which can have the highest concentration of cCur/ZnONPs and SBS value simultaneously were determined by disc agar diffusion (DAD), biofilm formation inhibition, and metabolic activity assays following photo-activation using light-emitting diode (LED) for 5 min against multispecies cariogenic biofilm-producing bacteria. RESULTS: Adhesive with 7.5% wt. cCur/ZnONPs showed the highest concentration of cCur/ZnONPs and SBS value (14.89 ± 3.26 MPa, P < 0.05) simultaneously. No significant differences in ARI scores were found between the modified adhesive and control (Transbond XT without the cCur/ZnONPs). 7.5% wt. cCur/ZnONPs following photo-activation was not colonized by the test microorganisms and suppressed 100% metabolic activity of the test microorganisms up to 90 day compared to the control group (cCur/ZnONPs free LED irradiation; P < 0.05). In DAD assay, the reduction of photodynamic disinfection of the 7.5% wt. cCur/ZnONPs against test bacteria was positively associated to the time, in such a way that it was decreased significantly after 60 day. From days 120 onwards, microbial biofilm formation and metabolic activity was progressively increased on 7.5% wt. cCur/ZnONPs adhesive discs compared to the control group (cCur/ZnONPs free LED irradiation). CONCLUSIONS: Our findings highlight the photo-activated 7.5% wt. cCur/ZnONPs can serve as an orthodontic adhesive additive to control the cariogenic multispecies biofilm, and also to reduce their metabolic activity.

The effect of antimicrobial photodynamic therapy on the expression of biofilm associated genes in Staphylococcus aureus strains isolated from wound infections in burn patients.

PURPOSE: Burn patients are particularly susceptible to microbial infection. Staphylococcus aureus causes burn wound, impetigo and cellulitis. Although sub-lethal antimicrobial photodynamic therapy (aPDT) would not result in microorganism killing, it can considerably influence microbial virulence factor. METHODS: Twelve methicillin-resistant S. aureus (MRSA) and methicillin-sensitive S. aureus (MSSA) isolated from burns patients. To determine the sub-lethal dose of aPDT, 12 clinical isolates of S. aureus photosensitized with 100 µg ml -1 toluidine blue O (TBO) and irradiated by light emitting diode (LED) with a wavelength of 630 ± 10 nm and energy densities of 52.0, 104.1, and 156.2 J/cm2, then bacterial viability was measured. The effects of sub-lethal aPDT on the expression levels of ica ABCD and ica R genes were assessed by quantitative Real-time PCR (qRT-PCR) method. RESULT: Fifty and 100 µg ml-1 of TBO significantly reduced the mean cell survival in the MRSA (2.5 - 3 log10) and MSSA (2.75-3.1 log10) isolates. The average expression levels of icaA, ica B, ica C, and ica D in the MRSA and MSSA isolates were decreased by (12, 14, 11, and 9) and (13, 14.5, 12, and 9.5) fold change, respectively (P < 0.05). However, the expression of ica R gene was decreased by 6 and 8 folds change in MRSA and MSSA, respectively. CONCLUSION: The potential of TBO-mediated aPDT could reduce the expression of ica ABCD as important genes involved in biofilm formation and ica R gene as a repressor of the ica operon. Therefore, the use of aPDT agents as a complementary therapy in wound infections of burn patients is recommended.

Antibacterial and Antibiofilm Efficacy of Antimicrobial Photodynamic Therapy Against Intracanal Enterococcus faecalis: An In Vitro Comparative Study with Traditional Endodontic Irrigation Solutions.

OBJECTIVES: Enterococcus faecalis (E. faecalis), an infecting microorganism of the root canals, is difficult to eliminate during endodontic therapy. In this study, the effect of root canal disinfection with sodium hypochlorite (NaOCl) and chlorhexidine (CHX) was evaluated on planktonic and biofilm forms of E. faecalis in comparison with antimicrobial photodynamic therapy (aPDT) as an alternative strategy for root canal disinfection. MATERIALS AND METHODS: In this study, E. faecalis (ATCC 29212) was used. The experimental procedures included aPDT with curcumin (CUR) and indocyanine green (ICG) as photosensitizers, irrigation with 5.25% NaOCl, 0.2% and 2.0% CHX solutions as traditional endodontic irrigating solutions, and the control group. The antibacterial and anti-biofilm potentials were assessed by counting the colony forming units and also using the crystal violet assay, respectively. RESULTS: According to the results, E. faecalis biofilm was disrupted by 65.3%, 81.0% and 92.6% using 0.2% CHX, 2.0% CHX, and 5.25% NaOCl, respectively (P<0.05). In addition, CUR- and ICG-mediated aPDT displayed a significant reduction in E. faecalis count (90.2% and 82.5%, respectively) and its biofilm (83.6% and 75.2%, respectively) in comparison to the control group (P<0.05). CONCLUSIONS: APDT has a high potential for elimination of E. faecalis and is almost equivalent to NaOCl and CHX. It can be used as an adjucnt to conventional endodontic irrigating solutions.

Exploring different photosensitizers to optimize elimination of planktonic and biofilm forms of Enterococcus faecalis from infected root canal during antimicrobial photodynamic therapy.

BACKGROUND: Despite the high success rate of endodontic treatment, failure may occur in some cases. In this case, Enterococcus faecalis is the most common species in endodontic treatment failure and post-treatment apical periodontitis. Therefore, a new adjunctive strategy is needed for the prevention of endodontic infections due to E. faecalis. The aim of the present study was to compare the antimicrobial and anti-biofilm activities of different common photosensitizers (PSs) for use in antimicrobial photodynamic therapy (aPDT) against E. faecalis. MATERIALS AND METHODS: E. faecalis strain ATCC 29212 was used as the tested strain and methylene blue (MB), toluidine blue O (TBO), indocyanine green (ICG), and curcumin (CUR) were used as PSs. Irradiation was carried out using diode laser and light emitting diode (LED) at wavelengths related to the above PSs. Then, antimicrobial and anti-biofilm activities were measured using the microbial viability assay and crystal violet test, respectively. RESULTS: aPDT with using the above PSs significantly decreased the CFU/mL count of E. faecalis compared to the control group (P < 0.05). The killing percentage of E. faecalis via PS mediated aPDT was 99.6%, 98.2%, 85.1%, and 65.0% for CUR, ICG, TBO, and MB, respectively. aPDT using the above PSs significantly decreased the biofilm formation ability of E. faecalis compared to the control group (P < 0.05). The biofilm reduction percentage of the PSs was 68.4%, 62.9%, 59.0%, and 47.6% for CUR, ICG, TBO, and MB, respectively. CONCLUSION: CUR and ICG mediated aPDT exhibited considerably more antimicrobial activity than other PSs, while TBO and MB demonstrated weaker anti-biofilm effects against E. faecalis compared to other PSs.

Inhibitory Effects of Antimicrobial Photodynamic Therapy with Curcumin on Biofilm-Associated Gene Expression Profile of Aggregatibacter actinomycetemcomitans.

OBJECTIVES: Periodontitis is an inflammation of periodontal tissues that is caused by the biofilm of periodontal pathogens. Aggregatibacter actinomycetemcomitans (A. actinomycetemcomitans) is an opportunistic periodontopathogen that can be the cause of periodontal diseases via fimbriae as a virulence factor. In this study, we aimed to determine the expression level of A. actinomycetemcomitans rcpA gene as a virulence factor associated with biofilm formation after antimicrobial photodynamic therapy (aPDT) as a relatively new therapeutic modality. MATERIALS AND METHODS: To determine sub-lethal doses of aPDT against A. actinomycetemcomitans ATCC 33384 strain, we used curcumin (CUR) as a photosensitizer at a final concentration of 40 µmol/ml, which was excited with a light-emitting diode (LED) at the wavelength of 450 nm. Quantitative real-time polymerase chain reaction (qRT-PCR) was then applied to monitor rcpA gene expression in A. actinomycetemcomitans. RESULTS: 10-40 µmol/ml of CUR caused a significant reduction in the growth of A. actinomycetemcomitans compared to control group (P<0.05). Also, the cell viability of A. actinomycetemcomitans was significantly decreased after more than four minutes of LED irradiation. Therefore, the sub-lethal dose of aPDT against A. actinomycetemcomitans was 5 µmol/ml of CUR with three minutes of LED irradiation at a fluency of 180-240 J/cm2, which reduced the expression of the rcpA gene by approximately 8.5-fold. CONCLUSIONS: aPDT with CUR leads to decreased cell survival and virulence of A. actinomycetemcomitans. Thus, CUR-aPDT can be used as an alternative approach for the successful treatment of periodontitis in vivo.

Association of virulence gene expression with colistin-resistance in Acinetobacter baumannii: analysis of genotype, antimicrobial susceptibility, and biofilm formation.

BACKGROUND: Acinetobacter baumannii causes difficult-to-treat nosocomial infections, which often lead to morbidity due to the development of antimicrobial drug resistance and expression of virulence genes. Data regarding the association of resistance to colistin, a last treatment option, and the virulence gene expression of A. baumannii is scarce. METHODS: We evaluated the MLVA genotype, antimicrobial resistance, and biofilm formation of 100 A. baumannii isolates from burn patients, and further compared the in vitro and in vivo expression of four virulence genes among five colistin-resistant A. baumannii (Cst-R-AB) isolates. Five Cst-R-AB isolates were tested; one from the present study, and four isolated previously. RESULTS: Our results showed that reduced expression of recA, along with increased in vivo expression of lpsB, dnaK, and blsA; are associated with colistin resistance among Cst-R-AB isolates. Differences in virulence gene expressions among Cst-R-AB isolates, may in part explain common discrepant in vitro vs. in vivo susceptibility data during treatment of infections caused by Cst-R-AB. CONCLUSIONS: Our findings highlight the intricate relationship between colistin-resistance and virulence among A. baumannii isolates, and underscore the importance of examining the interactions between virulence and antimicrobial resistance toward efforts to control the spread of multidrug-resistant A. baumannii (MDR-AB) isolates, and also to reduce disease severity in burn patients with MDR-AB infection.