OBJECTIVE: To investigate the inhibitory effects of levofloxacin (LEV) combined with cellulase against bacille CalmetteGuerin (BCG) biofilms in vitro. METHODS: The mature growth cycle of BCG biofilms was determined using the XTT method and crystal violet staining. BCG planktonic bacteria and BCG biofilms were treated with different concentrations of LEV and cellulose alone or jointly, and the changes in biofilm biomass were quantified with crystal violet staining. The mature BCG biofilm was then treated with cellulase alone for 24 h, and after staining with SYTO 9 and Calcofluor White Stain, the number of viable bacteria and the change in cellulose content in the biofilm were observed with confocal laser scanning microscopy. The structural changes of the treated biofilm were observed under scanning electron microscopy. RESULTS: The MIC, MBC and MBEC values of LEV determined by broth microdilution method were 4 μg/mL, 8 μg/mL and 1024 μg/mL, respectively. The combined treatment with 1/4×MIC LEV and 2.56, 5.12 or 10.24 U/mL cellulase resulted in a significant reduction in biofilm biomass (P < 0.001). Cellulase treatments at the concentrations of 10.24, 5.12 and 2.56 U/mL all produced significant dispersion effects on mature BCG biofilms (P < 0.001). CONCLUSION LEV combined with cellulose can effectively eradicate BCG biofilm infections, suggesting the potential of glycoside hydrolase therapy for improving the efficacy of antibiotics against biofilmassociated infections caused by Mycobacterium tuberculosis.
Levofloxacin/pharmacology* ; Gentian Violet/pharmacology* ; BCG Vaccine/pharmacology* ; Anti-Bacterial Agents/pharmacology* ; Biofilms ; Cellulases/pharmacology* ; Microbial Sensitivity Tests

OBJECTIVE: This study aimed to investigate the regulation of histone-like nucleoid structuring protein (H-NS) on biofilm formation and cyclic diguanylate (c-di-GMP) synthesis in Vibrio parahaemolyticus RIMD2210633. METHODS: Regulatory mechanisms were analyzed by the combined utilization of crystal violet staining, quantification of c-di-GMP, quantitative real-time polymerase chain reaction, LacZ fusion, and electrophoretic-mobility shift assay. RESULTS: The deletion of hns enhanced the biofilm formation and intracellular c-di-GMP levels in V. parahaemolyticus RIMD2210633. H-NS can bind the upstream promoter-proximal DNA regions of scrA, scrG, VP0117, VPA0198, VPA1176, VP0699, and VP2979 to repress their transcription. These genes encode a group of proteins with GGDEF and/or EAL domains associated with c-di-GMP metabolism. CONCLUSION One of the mechanisms by which H-NS represses the biofilm formation by V. parahaemolyticus RIMD2210633 may be via repression of the production of intracellular c-di-GMP.
Bacterial Proteins/metabolism* ; Biofilms ; Cyclic GMP/analogs & derivatives* ; Gene Expression Regulation, Bacterial ; Gentian Violet ; Histones/metabolism* ; Vibrio parahaemolyticus/genetics*

PURPOSE: The aim of this study is to evaluate the effectiveness of MnO₂-diatom microbubbler (DM) on the surface of prosthetic materials as a mouthwash by comparing the biofilm removal effect with those previously used as a mouthwash in dental clinic.MATERIALS AND METHODS: DM was fabricated by doping manganese dioxide nanosheets to the diatom cylinder surface. Scanning electron microscopy (SEM) was used to observe the morphology of DM and to analyze the composition of doped MnO₂. Stereomicroscope was used to observe the reaction of DM in 3% hydrogen peroxide. Non-precious metal alloys, zirconia and resin specimens were prepared to evaluate the effect of biofilm removal on the surface of prosthetic materials. And then Streptococcus mutans and Porphyromonas gingivalis biofilms were formed on the specimens. When 3% hydrogen peroxide solution and DM were treated on the biofilms, the decontamination effect was compared with chlorhexidine gluconate and 3% hydrogen peroxide solution by crystal violet staining.RESULTS: Manganese dioxide was found on the surface of the diatom cylinder, and it was found to produce bubble of oxygen gas when added to 3% hydrogen peroxide. For all materials used in the experiments, biofilms of the DM-treated groups got effectively removed compared to the groups used with chlorhexidine gluconate or 3% hydrogen peroxide alone.CONCLUSION: MnO₂-diatom microbubbler can remove bacterial membranes on the surface of prosthetic materials more effectively than conventional mouthwashes.
Alloys ; Biofilms ; Chlorhexidine ; Decontamination ; Dental Clinics ; Dental Plaque ; Diatoms ; Gentian Violet ; Hydrogen Peroxide ; In Vitro Techniques ; Manganese ; Membranes ; Microscopy, Electron, Scanning ; Mouthwashes ; Oral Hygiene ; Oxygen ; Porphyromonas gingivalis ; Streptococcus mutans

OBJECTIVES: Biofilm formation is critical to dental caries initiation and development. The aim of this study was to investigate the effects of nicotine exposure on Streptococcus mutans (S. mutans) biofilm formation concomitantly with the inhibitory effects of sodium chloride (NaCl), potassium chloride (KCl) and potassium iodide (KI) salts. This study examined bacterial growth with varying concentrations of NaCl, KCl, and KI salts and nicotine levels consistent with primary levels of nicotine exposure. MATERIALS AND METHODS: A preliminary screening experiment was performed to investigate the appropriate concentrations of NaCl, KCl, and KI to use with nicotine. With the data, a S. mutans biofilm growth assay was conducted using nicotine (0–32 mg/mL) in Tryptic Soy broth supplemented with 1% sucrose with and without 0.45 M of NaCl, 0.23 M of KCl, and 0.113 M of KI. The biofilm was stained with crystal violet dye and the absorbance measured to determine biofilm formation. RESULTS: The presence of 0.45 M of NaCl, 0.23 M of KCl, and 0.113 M of KI significantly inhibited (p < 0.05) nicotine-induced S. mutans biofilm formation by 52%, 79.7%, and 64.1%, respectively. CONCLUSIONS: The results provide additional evidence regarding the biofilm-enhancing effects of nicotine and demonstrate the inhibitory influence of these salts in reducing the nicotine-induced biofilm formation. A short-term exposure to these salts may inhibit S. mutans biofilm formation.
Biofilms ; Dental Caries ; Gentian Violet ; Mass Screening ; Nicotine ; Potassium Chloride ; Potassium Iodide ; Salts ; Sodium Chloride ; Streptococcus mutans ; Streptococcus ; Sucrose

BACKGROUND/AIMS: Cronobacter sakazakii, an emergent pathogen is considered as a major concern to infants and neonates fed on reconstituted powdered infant milk formula. In conjunction with many other factors, biofilm forming capacity adds to its pathogenic potential. In view of the facts that infants are at highest risk to C. sakazakii infections, and emerging antibiotic resistance among pathogens, it is imperative to evaluate probiotic cultures for their efficacy against C. sakazakii. Therefore, pure probiotic strains were isolated from commercial probiotic products and tested for their antimicrobial and anti-biofilm activities against C. sakazakii. METHODS: A total of 6 probiotic strains were tested for their antibiotic susceptibility followed by antimicrobial activity using cell-free supernatant (CFS) against C. sakazakii. The inhibitory activity of CFS against biofilm formation by C. sakazakii was determined using standard crystal violet assay and microscopic observations. RESULTS: All the probiotic strains were sensitive to ampicillin, tetracycline, vancomycin and carbenicillin whereas most of the strains were resistant to erythromycin and novobiocin. Four of the 6 probiotic derived CFS possessed antimicrobial activity against C. sakazakii at a level of 40 μL. A higher biofilm inhibitory activity (>80%) was observed at initial stages of biofilm formation with weaker activity during longer incubation upto 48 hours (50%–60%). CONCLUSIONS: The study indicated the efficacy of isolated commercial probiotics strains as potential inhibitor of biofilm formation by C. sakazakii and could be further explored for novel bioactive molecules to limit the emerging infections of C. sakazakii.
Ampicillin ; Biofilms ; Carbenicillin ; Cronobacter sakazakii ; Cronobacter ; Drug Resistance, Microbial ; Erythromycin ; Gentian Violet ; Humans ; Infant ; Infant, Newborn ; Milk ; Novobiocin ; Probiotics ; Tetracycline ; Vancomycin

OBJECTIVES: The purpose of this study is to determine methods of dental caries prevention by investigating the use of compounds of Diospyros kaki (D. kaki) peel, Momordica charantia (M. charantia), and Canavalia gladiata (C. gladiata) extracts to limit the cariogenic traits of Streptococcus mutans (S. mutans), such as their ability to proliferate and adhere to the tooth surface. METHODS: Broth microdilution and the agar spreading assay were used to determine the antimicrobial effect and minimum inhibitory concentration (MIC) of S. mutans extracts. In order to identify the adhesive ability of S. mutans at varying concentrations, culture plates were first stained with 1 ml of 0.01% crystal violet for 15 minutes at room temperature, and then eluted with 1 ml of EtOH:Acetone (8:2) solution for 15 minutes in a 37℃ incubator. Eluted solutions were then evaluated by use of a spectrophotometer at 575 nm. RESULTS: Experiments were conducted in order to investigate the effectiveness of D. kaki peel, M. charantia, and C. gladiata extracts on limiting the proliferation of S. mutans. The MIC was measured as an indication of whether the antibacterial activity of D. kaki peel, M. charantia, and C. gladiata extracts had a significant bacteriostatic effect on S. mutans. M. charantia extract was effective for growth inhibition on S. mutans at a minimum concentration of 0.25%. From the adhesion ability assay, M. charantia extract had an anti-adhesive effect. CONCLUSIONS: These results indicate that M. charantia extract demonstrates antibacterial activity and has an anti-adhesive effect on S. mutans. Due to these properties, M. charantia extract may be used to prevent dental caries.
Adhesives ; Agar ; Canavalia ; Dental Caries ; Diospyros ; Gentian Violet ; Incubators ; Microbial Sensitivity Tests ; Momordica charantia ; Momordica ; Streptococcus mutans ; Streptococcus ; Thiram ; Tooth

OBJECTIVE: Nerve growth factor (NGF) is a member of the neurotrophic factor family and plays a vital role in the physiological processes of organisms, especially in the nervous system. Many recent studies have reported that NGF is also involved in the regulation of tumourigenesis by either promoting or suppressing tumor growth, which depends on the location and type of tumor. However, little is known regarding the effect of NGF on interspinal schwannoma (IS). In the present study, we aimed to explored whether mouse nerve growth factor (mNGF), which is widely used in the clinic, can influence the growth of interspinal schwannoma cells (ISCs) isolated from IS in vitro.METHODS: ISCs were isolated, cultured and identified by S-100 with immunofluorescence analysis. S-100-positive cells were divided into five groups, and separately cultured with various concentrations of mNGF (0 [phosphate buffered saline, PBS], 40, 80, 160, and 320 ng/mL) for 24 hours. Western blot and quantantive real time polymerase chain reaction (PCR) were applied to detect tyrosine kinase A (TrkA) receptor and p75 neurotrophin receptor (p75(NTR)) in each group. Crystal violet staining was selected to assess the effect of mNGF (160 ng/mL) on ISCs growth.RESULTS: ISCs growth was enhanced by mNGF in a dose-dependent manner. The result of crystal violet staining revealed that it was significantly strengthened the cells growth kinetics when cultured with 160 ng/mL mNGF compared to PBS group. Western blot and quantantive real time PCR discovered that TrkA receptor and mRNA expression were both up-regualated under the condition of mNGF, expecially in 160 ng/mL, while the exoression of p75(NTR) demonstrated no difference among groups.CONCLUSION: From these data, we conclude that exogenous mNGF can facilitate ISC growth by activating both TrkA receptor and p75(NTR). In addition, patients who are suffering from IS should not be administered mNGF in the clinic.
Animals ; Blotting, Western ; Fluorescent Antibody Technique ; Gentian Violet ; Humans ; In Vitro Techniques ; Kinetics ; Mice ; Nerve Growth Factor ; Nervous System ; Neurilemmoma ; Physiological Processes ; Protein-Tyrosine Kinases ; Real-Time Polymerase Chain Reaction ; Receptor, Nerve Growth Factor ; Receptor, trkA ; Receptors, Nerve Growth Factor ; RNA, Messenger

PURPOSE: The goal of this study was to develop and validate a standardized in vitro pathogenic biofilm attached onto saliva-coated surfaces. METHODS: Fusobacterium nucleatum (F. nucleatum) and Porphyromonas gingivalis (P. gingivalis) strains were grown under anaerobic conditions as single species and in dual-species cultures. Initially, the bacterial biomass was evaluated at 24 and 48 hours to determine the optimal timing for the adhesion phase onto saliva-coated polystyrene surfaces. Thereafter, biofilm development was assessed over time by crystal violet staining and scanning electron microscopy. RESULTS: The data showed no significant difference in the overall biomass after 48 hours for P. gingivalis in single- and dual-species conditions. After adhesion, P. gingivalis in single- and dual-species biofilms accumulated a substantially higher biomass after 7 days of incubation than after 3 days, but no significant difference was found between 5 and 7 days. Although the biomass of the F. nucleatum biofilm was higher at 3 days, no difference was found at 3, 5, or 7 days of incubation. CONCLUSIONS: Polystyrene substrates from well plates work as a standard surface and provide reproducible results for in vitro biofilm models. Our biofilm model could serve as a reference point for studies investigating biofilms on different surfaces.
Bacterial Adhesion ; Biofilms ; Biomass ; Fusobacterium nucleatum ; Fusobacterium ; Gentian Violet ; In Vitro Techniques ; Microscopy, Electron, Scanning ; Polystyrenes ; Porphyromonas gingivalis ; Porphyromonas

From dye-assisted conventional chromoendoscopy to novel virtual chromoendoscopy, image-enhanced endoscopy (IEE) is continuously evolving to meet clinical needs and improve the quality of colonoscopy. Dye-assisted chromoendoscopy using indigo carmine or crystal violet, although slightly old-fashioned, is still useful to emphasize the pit patterns of the colonic mucosa and predict the histological structures of relevant lesions. Equipment-based virtual chromoendoscopy has the advantage of being relatively easy to use. There are several types of virtual chromoendoscopy that vary depending on the manufacturer and operating principle. IEE plays distinctive roles with respect to histologic characterization of colorectal polyps and prediction of the invasion depth of colorectal cancers. In addition, the newest models of IEE have the potential to increase adenoma and polyp detection rates in screening colonoscopy.
Adenoma ; Colon ; Colonoscopy ; Colorectal Neoplasms ; Endoscopy* ; Gastrointestinal Diseases* ; Gentian Violet ; Image Enhancement ; Indigo Carmine ; Mass Screening ; Mucous Membrane ; Polyps

PURPOSE: To report a case of irregular astigmatism caused by a free flap during laser-assisted in situ keratomileusis (LASIK) surgery that was treated with a flap rotation based on postoperative topography. CASE SUMMARY: A 21-year-old female underwent LASIK, which was complicated by a free cap on her right eye. Because the gentian violet markings were no longer present, the exact orientation of the cap was unknown. At 3 months after surgery, the astigmatism of the right eye was −3.00 diopters (D) with an uncorrected visual acuity (UCVA) of 0.4, and the astigmatism of the left eye was −0.75 D with an UCVA of 1.0. The corneal topography was analyzed in order to return to the existing position. Free cap repositioning was performed and irregular astigmatism was corrected to improve the UCVA to 1.0. CONCLUSIONS: If the preoperative markings cannot be identified on a free flap during LASIK, secondary postoperative corneal topographic analysis can be performed to restore the corneal free flap to its original position to minimize astigmatism with good visual outcomes.
Astigmatism ; Corneal Topography ; Female ; Free Tissue Flaps ; Gentian Violet ; Humans ; Keratomileusis, Laser In Situ ; Visual Acuity ; Young Adult