Amphiphilic peptide Mastoparan-B induces conformational changes within the AdeB efflux pump, down-regulates adeB gene expression, and restores antibiotic susceptibility in an MDR strain of Acinetobacter baumannii.

Mastoparan B (MP-B) is an amphiphilic peptide with a potent antimicrobial activity against most Gram-negative bacteria. However, there is little information available on the inhibition of the Acinetobacter baumannii resistance-nodulation-cell-division (RND) efflux pump using this antimicrobial peptide. Here, we carried out a series of in-silico experiments to find the mechanisms underlying the anti-efflux activity of MP-B using a multi-drug resistant (MDR) strain of A. baumannii (AB). According to our findings, MP-B demonstrated a potent antibacterial activity against an MDR-AB (minimum inhibitory concentration [MIC] = 1 µg/mL) followed by a 20-fold reduction in the adeB gene expression in the presence of sub-MIC of this peptide. Using Groningen Machine for Chemicals Simulation (GROMACS) via PyMOL Graphical User Interface (GUI), (we observed that, the AdeB transporter had conserved helix-turn-helix regions and a tight pore rich in Phe and Ala residues. To understand how inhibition of the AdeB is achieved, we generated 20 apo-MP-B poses using the InterPep and SiteMap tools. The high-quality model was created by homology modeling and used for docking via AutoDock/Vina to identify the MP-B binding sites. We established that the most apo-MP-B formed H-bonds to the backbone of five amino acids in the Helix-5. As a result, the dihedral angles of the involved amino acids shift by 9.0-9.6 Ǻ, causing a change in the conformation of the AdeB protein. This led to helix conformation stereoisomerization and block the AdeB activity. MP-B presumably has dual mechanisms. (1) It blocks the AdeB transporter by changing its conformation. (2) MP-B influences the adeB gene expression by binding to G-protein which laterally controls efflux regulators like MarA, RamA, SoxS, and Rob proteins.

Quercetin Effects on Cell Cycle Arrest and Apoptosis and Doxorubicin Activity in T47D Cancer Stem Cells.

BACKGROUNDS: Targeting breast cancer stem cells with the CD44+/CD24- phenotype is critical for complete eradication of cancer cells due to its Self-renewal, differentiation, and therapeutic resistance ability. Quercetin is a popular flavonoid with lower adverse effects and has anti-tumor properties. Therefore, we assessed the anticancer activity of Quercetin and Doxorubicin alone and in combination in the T47D cells of human breast cancer and their isolated Cancer stem cells (CSCs). MATERIALS AND METHODS: The human breast cancer cell line T47D was used for this experiment. T47D CSCs were isolated by magnetic bead sorting using the MACS system. The anticancer activity of Quercetin and Doxorubicin alone and in combination were evaluated using MTT cytotoxicity assay and cell cycle distribution and apoptosis induction by flow cytometry analysis. RESULTS: We have shown that almost 1% of T47D cell populations are made up of CD44+/CD24- cells, which considered as cancer stem cells. Quercetin and Doxorubicin alone or in combination inhibited cell proliferation and induced apoptosis in breast cancer T47D cells and in lower extent in CD44+/CD24- cells. Quercetin significantly strengthened Doxorubicin's cytotoxicity and apoptosis induction in both cell populations. Quercetin and Doxorubicin and their combination induced G2/M arrest in the T47D cells and to a lesser extent in isolated CSCs. A value of p < 0.05 was considered as indicating a statistically significant difference. CONCLUSION: These outcomes suggested that CSCs are a minor population of cancer cells, which play a significant role in drug resistance by being quiescent, slow cycling and resistance to apoptosis. Furthermore, our data showed that adding Quercetin to Doxorubicin is an effective approach for the treatment of both CSCs and bulk tumor cells.

The Antibacterial and Anti-biofilm Traits of the Novel BMAP-27-Melittin Conjugated Peptide Nanoparticle Against Streptococcus mutans: Clinical Isolates from Oral Cavity.

Background & Objective: The spread and development of drug-resistant bacterial strains has prompted the hunt for novel antibacterial polypeptides undergoing conformational changes to confer rapid bactericidal effects. The aim of this study was to evaluate the effect of novel BMAP27-Melittin conjugated peptide- nanoparticle (NP) against Streptococcus mutans as the primary pathogen from subgingival plaques. Methods: Sixty subgingival plaque samples were collected, and 39 S. mutans isolates were identified. The BMAP27-Melittin conjugated peptide was purchased from GenScript Company, USA. Minimum Inhibitory Concentration (MIC), Minimum Bactericidal Concentration (MBC), Biofilm Inhibitory Concentration (BIC), and Biofilm Eradication Concentration (BEC) of BMAP27-Melittin-NP were calculated using the microtiter method. Results: Thirty-nine infected subjects were reported, including 24 males and 15 females (P=0.299). MIC, MBC, BIC, and BEC of BMAP27-Melittin-NP against S. mutans were 1.8, 2.9, 2.1, and 3.8µg/mL, respectively. The mean MBC, BEC, and BIC values were significantly lower among clinical isolates than S. mutans ATCC 35688 standard strain (P=0.032, 0.001, and 0.001, respectively). Conclusion: BMAP27-Melittin-NP demonstrated significant antibacterial and anti-biofilm effects against clinical isolates of S. mutans which can be considered a promising compound to prevent or treat dental caries and eradicate the oral infections.

The Effect of Gold Nano Particles with Different Sizes on Streptococcus Species.

STATEMENT OF THE PROBLEM: Streptococcus mutans, Streptococcus sanguinis, and Streptococcus salivarius are most common etiologic bacteria for dental caries. Different sizes of gold nanoparticles may have different antibacterial effects on these species. PURPOSE: This study aimed to compare the antibacterial effect of chlorhexidine and three sizes of gold nano particles (25, 60, 90nm) against clinical and standard strains of Streptococcus mutans, Streptococcus sanguinis, and Streptococcus salivarius. MATERIALS AND METHOD: In this cross-sectional study, the specimens were collected from 75 children aged 3-5 years old. Antibacterial effect of chlorhexidine and three sizes of gold nano particles (25, 60, 90nm) were investigated by evaluating the minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) against three bacterial strains. RESULTS: The MIC and MBC of gold nanoparticles with different sizes against Streptococcus mutans, Streptococcus sanguinis, and Streptococcus salivarius were statistically different. The MIC and MBC of smaller gold nano particles (25nm) were significantly lower (p<0.001) than larger ones. Patient-derived bacteria had significantly higher values of MIC and MBC in comparison to standard species (p<0.001). CONCLUSION: The results of this study confirmed the significant size-dependency of gold nano particles for antibacterial activity. As the size of gold nano particles decrease, the antibacterial properties enhance.

New Strategy of Reducing Biofilm Forming Bacteria in Oral Cavity by Bismuth Nanoparticles.

OBJECTIVE: Enterococcus faecalis and Streptococcus salivarius are the most important species in dental decay and producing biofilm. Treatment with chlorhexidine 2% mouthwash for 7 days is the best way to eliminate these bacteria. However, due to the ability of these bacteria to survive in harsh environments, increasing emergence of bacterial resistance against available antibiotics, and favorable properties of nanoparticles including broad spectrum antimicrobial activity and lower toxicity, we decided to evaluate reducing biofilm forming bacteria in oral cavity by bismuth nanoparticles. MATERIALS AND METHODS: This was a cross-sectional study of 40 samples isolated from the patients visiting dental clinics in Shiraz in 2019. Samples, which showed growth, were cultured on blood agar plates and incubated for the PCR procedure. Nanoparticle powder was dissolved in high-purity water, and the final concentration of bismuth nanoparticles (BiNPs) was measured with a spectrophotometer. Minimum inhibitory concentration (MIC) of BiNPs against E. faecalis and S. salivarius was determined by the microbroth dilution method according to methods for antimicrobial susceptibility tests. Also, bactericidal assays were conducted in a Mueller-Hinton broth medium and reported as the concentration of BiNPs that reduced the viable bacterial count by 99.9%. Statistical analysis was carried out using SPSS 21 and one-way analysis of variance, and P values less than 0.05 were considered significant. RESULTS: MICs of BiNP suspension against Streptococcus salivarius and Enterococcus faecalis were 2.5 and 5 µg/ml, respectively. Minimum bactericidal concentrations (MBC) of BiNP suspension against Streptococcus salivarius and Enterococcus faecalis were 5 and 10 µg/ml, respectively. Antibacterial activity of BiNPs was compared with chlorhexidine 2%. MICs of BiNPs against Streptococcus salivarius and Enterococcus faecalis were one-twentieth less than those of chlorhexidine. MBC of BiNPs against both pathogens was one-tenth less than those of chlorhexidine. CONCLUSION: BiNPs were more effective than chlorhexidine, and MIC and MBC of bismuth nanoparticles are lower than those of chlorhexidine.

Assessment of the Antibacterial Effects of Bismuth Nanoparticles against Enterococcus faecalis.

INTRODUCTION: Enterococcus faecalis (E. faecalis) is the most important species in dentistry and plays a significant role in the etiology of persistent apical lesions after root canal treatment. Up to date, the intracanal application of 2% chlorhexidine for 7 days is the best way to eliminate E. faecalis. However, due to the ability of this bacterium to persist and survive in harsh environments, many studies have been directed towards finding an alternative strategy for prevention or eradication of it. This study was conducted to investigate the effect of bismuth nanoparticles on E. faecalis, as an etiologic factor in recurrent root canal infections. METHODS: Forty patients, referred to Endodontic Ward of Shiraz University of Medical Science for endodontic pretreatment, provided root canal samples. First, all samples were transferred in Enterococcosel broth and incubated. Then, samples which showed growth were plated on blood agar plates and incubated for further PCR procedure. Nanoparticle powder was dissolved in high-purity water, and the final concentration of bismuth nanoparticles (BiNPs) was measured by the spectrophotometer. Minimum inhibitory concentration (MIC) of BiNPs against E. faecalis was determined by microbroth dilution method according to methods for antimicrobial susceptibility tests. Also, bactericidal assays were conducted in Mueller-Hinton broth medium and reported as the concentration of BiNPs that reduced the viable bacterial count by 99.9%. RESULTS: Of all samples, 77.5% revealed the presence of E. faecalis by PCR. Also, E. faecalis growth inhibition was observed at concentrations ranging from 0.625 µg/ml to 20 µg/ml (geometric mean: 2.337 µg/ml), and the MBC values were between 1.25 µg/ml and 40 µg/ml (geometric mean: 4.781 µg/ml), which in comparison with chlorhexidine, these values were about one-eighth of chlorhexidine. CONCLUSION: The experimental data suggest that bismuth nanoparticles could be an interesting alternative to combat E. faecalis, which, in view of the advantages mentioned for bismuth nanoparticle like inhibiting Streptococcus mutans biofilm formation and higher antibacterial activity compared to chlorhexidine, can be suggested to be used in different fields of dentistry.

Evaluation of Antibiotic Resistance Pattern in Dental Bacteremia Detected by Multiplex PCR Technique.

The aim of this study was to detect oral bacteremia and offer the antibiotic resistance patterns. Bacterial resistance pattern was evaluated in 50 patients. A spectrophotometer device equipped with UV and electrophoresis of the extracted samples on agarose gel for antibiogram test were used. PCR test 15 minutes after tooth extraction showed that bacterial strains were extracted from 16 patients. Lactobacillus, Enterococcus faecalis (E. faecalis), Streptococcus sanguinis (S. sanguinis), Streptococcus salivarius (S. salivarius), and Streptococcus mutans (S. mutans) were extracted from 5, 4, 4, 4, and 6 patients. 100% of Lactobacillus, E. faecalis, S. sanguinis, S. salivarius, and S. mutans were sensitive to tigecycline. Most of the Lactobacillus antibiotic resistance was against tetracycline and ciprofloxacin. Antibiotic resistance in S. salivarius was observed in 75% of the cases against piperacillin-tazobactam, ciprofloxacin, and cefotaxime, while in S. mutans was 84% of the cases against ceftriaxone. The results of the current study showed that tooth extraction causes bacteremia before, during, and after tooth extraction. Generally, the highest antibiotic resistance occurred against tetracycline, ciprofloxacin, and ampicillin-sulbactam. In most cases, the bacteria showed partial resistance to these antibiotics; however, tigecycline showed 100% efficacy on all types of bacteria. Streptococcus strains (salivarius, mutans, and sanguinis) were sensitive to most of the antibiotics while antibiotic sensitivity was less evident in Lactobacillus and E. faecalis. Antibiotic resistance has become a critical issue, since it leads to treatment failure when there is a need for antibiotic therapy.

Multiplex polymerase chain reaction for detection of bacteremia during dental extraction.

AIM: Incidence of transient bacteremia following dental extractions ranges 30%-70% among adults and 58%-100% in children. This study aims to assess the multiplex polymerase chain reaction (PCR) technique in detection of transient bacteremia during dental extraction. METHODS: Twenty volunteers for dental extraction entered the study. Blood samples were taken at baseline and 30 seconds and 15 minutes after extraction. Five types of bacteria were selected, namely Streptococcus mutans, S. salivarius, S. sanguinis, Enterococcus faecalis and Lactobacillus. Blood samples were evaluated by microbial culture and multiplex PCR. RESULTS: Blood culture showed rates of 0%, 80% and 25% for bacteremia before, during and after dental extraction, respectively. A significant difference was observed between baseline and during extraction. Using PCR, bacteremia was deemed 20%, 100% and 30% before, during and after extraction, respectively, and a significant difference was witnessed between the baseline and during extraction. The highest incidence was at the 30-second mark after extraction, and the most prevalent type of bacteria was S. mutans. CONCLUSIONS: Multiplex PCR can be used to determine bacterial diversity with high accuracy during occurrence of bacteremia. Therefore, in cases where positive blood cultures only indicate the presence of one species, we recommend the use of this method to detect more diverse bacteria types.

Medical and dental applications of nanomedicines.

Nanoparticles are tiny materials with nanosized components less than 100 nm in at least one dimension with physicochemical properties, which make them very attractive for medical application. These compounds have been evaluated as potential medicines for several decades. Nanotechnology has provided advances in the various fields of health sciences such as diagnosis, prevention and treatment by application of the agents named nanomedicines, including proteins, polymers, micelles, dendrimers, liposomes, emulsions, nanocapsules and nanoparticles. These materials can act as a scaffold, gene/drug delivery, tumor suppressor, conjugated with surgical implant, etc. They can also use as a nanocomposite, artificial tooth and dental caries preventing agent in the dentistry science. This current review tries to summarize recent applications of nanomedicine in the medical and dental fields.

Antimicrobial and antibiofilm activity of silver, titanium dioxide and iron nano particles.

PURPOSE: To compare the antimicrobial effects of chlorhexidine, penicillin, erythromycin, clindamycin, tetracycline and vancomycin with silver, titanium dioxide and iron nanoparticles and also to consider the synergistic antibacterial and antibiofilm effects of nanoparticles in clinical and standard strains of Streptococcus mutans and Streptococcus sanguinis. METHODS: The specimens collected from 66 3-5 year-old children with detected S. mutans and S. sanguinis by PCR were then exposed to the antimicrobial activity of chlorhexidine, penicillin, erythromycin, clindamycin, tetracycline and vancomycin with silver, titanium dioxide and iron nanoparticles measured by microdilution and disc diffusion tests and the colony counted after 1 to 5 minutes. The antibiofilm activity was examined by microtiter test. RESULTS: Use of nanoparticles alone showed higher minimum inhibitory concentration (MIC) than using them synergistically. The most effective synergistic solution was the one containing TiO2, Ag and Fe3O4 showing 0.019 µg#47;ml in S. mutans and S. sanguinis. Furthermore, this solution had the lowest biofilm inhibitory concentration (BIC) and colony forming units than the other antibiotics and chlorhexidine. CLINICAL SIGNIFICANCE: The solution containing TiO2, Ag and Fe33O4 showed the lowest inhibitory and antibiofilm concentration against S. mutans and S. sanguinis compared to those of other nanoparticle containing solutions, antibiotics and chlorhexidine, thus it may be used for treating dental caries, dental plaque and oral infections.