Browsing by Author "Gomez, Grace F."
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Item Effect of a Tea Polyphenol on Different Levels of Exposure of Nicotine and Tobacco Extract on Streptococcus mutans Biofilm Formation(Frontiers Media, 2021-12-01) Taylor, Emily S.; Gomez, Grace F.; Moser, Elizabeth A. S.; Sanders, Brian J.; Gregory, Richard L.; Biomedical and Applied Sciences, School of DentistryObjective: The purpose of this study was to compare the effects of different levels of nicotine and tobacco extract exposure on Streptococcus mutans biofilm formation and the inhibitory effect of the polyphenol epigallocatechin-3 gallate (EGCG) found in green tea. This study addressed the results of biofilm assays with EGCG and varying relative concentrations of nicotine and tobacco extract consistent with primary, secondary and tertiary levels of smoking exposure. Primary smoking exposure to nicotine has been demonstrated to significantly increase biofilm formation, while EGCG has been demonstrated to reduce S. mutans biofilm formation. Methods: S. mutans was treated with varying levels of nicotine or cigarette smoke condensate (CSC) concentrations (0–32 mg/ml and 0–2 mg/ml, respectively) in Tryptic Soy broth supplemented with 1% sucrose for different lengths of time simulating primary, secondary and tertiary smoking exposure with and without 0.25 mg/ml EGCG. The amount of total growth and biofilm formed was determined using a spectrophotometric crystal violet dye staining assay. Results: For both nicotine and CSC, primary exposure displayed overall significantly less growth compared to secondary exposure. For nicotine, secondary exposure demonstrated significantly greater growth than tertiary exposure levels. Overall, significantly greater total bacterial growth and biofilm formation in the presence of nicotine and CSC was observed in the absence of EGCG than in the presence of EGCG. However, biofilm growth was not significantly different among different concentrations of CSC. Conclusion: The results of this study help illustrate that nicotine-induced S. mutans biofilm formation is reduced by the presence of EGCG. This provides further evidence of the potential beneficial properties of polyphenols.Item Effect of phototherapy on the metabolism of Streptococcus mutans biofilm based on a colorimetric tetrazolium assay(JST, 2018-06) Gomez, Grace F.; Huang, Ruijie; Eckert, George; Gregory, Richard L.; Biomedical Sciences and Comprehensive Care, School of DentistryThe aim of this in vitro study was to determine the effect of violet-blue light on the metabolic activity of early Streptococcus mutans biofilm, reincubated at 0, 2, and 6 h after 5 min of violet-blue light treatment. S. mutans UA159 biofilm cells were cultured for 12 to 16 h in microtiter plates with Tryptic Soy broth (TSB) or TSB with 1% sucrose (TSBS) and irradiated with violet-blue light for 5 min. After irradiation, the plates were reincubated at 37°C for 0, 2, or 6 h in 5% CO2. Colorimetric tetrazolium salt reduction assay was used to investigate bacterial metabolic activity. Mixed model ANOVA was used to find the difference between the violet-blue light treated and nontreated groups. Bacterial metabolic activity was significantly lower in the violet-blue light group for TSB than in the nontreated group (P < 0.0001) regardless of recovery time. However, the differences between metabolic activity in the treated groups without sucrose decreased over time. For TSBS, metabolic activity was significantly lower with violet-blue light at 0 and 2 h. Violet-blue light inhibited the metabolic activity of S. mutans biofilm cells in the light-treated group. This finding may present a unique treatment method for patients with active caries.Item Evaluating hop extract concentrations found in commercial beer to inhibit Streptococcus mutans biofilm formation(Wiley, 2022) Gregory, Eric R.; Bakhaider, Renad F.; Gomez, Grace F.; Huang, Ruijie; Moser, Elizabeth A.S.; Gregory, Richard L.; Biomedical Sciences and Comprehensive Care, School of DentistryAims: The purpose of this study was to compare the effect of hop extracts with diverse β-acid concentrations on Streptococcus mutans biofilm formation. Methods and results: Ten different hop extracts, with α-acid concentrations similar to those found in commercial beer products and β-acid concentrations ranging from 2.6 to 8.1%, were added to distilled water to make standardized concentrations. S. mutans isolates were treated with hop extract dilutions varying from 1:2 to 1:256. The minimum inhibitory, minimum bactericidal and minimum biofilm inhibitory concentrations were determined and the optical density was evaluated. Live/dead staining confirmed the bactericidal effects. Biofilm formation of several strains of S. mutans was significantly inhibited by hop extract dilutions of 1:2, 1:4, 1:8, 1:16 and 1:32. Strong negative correlations were observed between α- and β-acid concentrations of the hop extracts and S. mutans total growth and biofilm formation. Conclusions: The use of hop extracts prepared similarly to commercial beer decreased S. mutans biofilm formation. Significance and impact of the study: The inclusion of hops in the commercial beer products may provide beneficial health effects. Further studies are warranted to determine an effect in vivo on the development of dental caries.Item The Impact of Nicotine and Cigarette Smoke Condensate on Metabolic Activity and Biofilm Formation of Candida albicans on Acrylic Denture Material(Wiley, 2018) Alzayer, Yasmin Mohammed; Gomez, Grace F.; Eckert, George J.; Levon, John A.; Gregory, Richard L.; Biomedical and Applied Sciences, School of DentistryPurpose Smokers have increased denture stomatitis caused primarily by Candida albicans. The primary aim of this study was to demonstrate the impact of a wide range of nicotine and cigarette smoke condensate (CSC) concentrations on biofilm formation and metabolic activity of C. albicans on acrylic denture material. Materials and Methods C. albicans (ATCC strain 10231) was used. Standardized denture acrylic (PMMA) specimens (total of 135 specimens) were incubated with C. albicans and exposed to nicotine and CSC at different concentrations (0, 0.25, 0.5, 1, 2, 4, 8, 16, and 32 mg/ml) and (0, 0.25, 0.5, 1, 2, and 4 mg/ml), respectively. For each experiment, 3 samples per nicotine and CSC concentration and a total of 45 specimens (27 specimens for the nicotine and 18 specimens for the CSC‐treated samples) were used and were selected randomly for each group. The control group consisted of 0 mg/ml of nicotine or CSC. The viability of C. albicans was measured using spiral plating on blood agar plates. The effect of nicotine and CSC concentrations on planktonic cells was were measured using a microplate reader. Metabolic activity of 24‐hour‐old established C. albicans biofilm exposed to nicotine and CSC for 24 hours in microtiter plates was determined using a 2,3‐bis (2‐methoxy‐4‐nitro‐5‐sulfophenyl)‐2H‐tetrazolium‐carboxanilide (XTT) reduction assay. Results The viability of C. albicans increased concomitant with increasing concentrations of CSC and nicotine, particularly at 0.5 and 2 mg/ml, respectively. Concentrations of CSC and nicotine above this resulted in an inhibitory effect on C. albicans viability. CSC and nicotine at 4 and 16 mg/ml, respectively, increased C. albicans biofilm metabolic activity. Conclusion Nicotine and CSC up to certain concentrations caused increases in biofilm formation, metabolic activity, viability, and planktonic cell absorbance of C. albicans. This in vitro study demonstrates the effectiveness of tobacco on promoting the growth of C. albicans and suggests their potential contributing factor in C. albicans biofilm related infections in smokers.Item Nicotine Upregulates Coaggregation of Candida albicans and Streptococcus mutans(Wiley, 2017) Ashkanane, Ali; Gomez, Grace F.; Levon, John; Windsor, Jack; Eckert, George J.; Gregory, Richard L.; Biomedical and Applied Sciences, School of DentistryPurpose Denture stomatitis is a condition of painless inflammation of denture-bearing mucosa. Reports indicate that nicotine, the major psychoactive ingredient in tobacco, increases growth of Streptococcus mutans and Candida albicans in denture biofilm. The purpose of this study was to determine the in vitro effects of nicotine on coaggregation of C. albicans with S. mutans. Material and Methods C. albicans strain ATCC 10231, S. mutans strain UA159 (ATTC 700610), and nicotine dilutions (ranging from 0 to 32 mg/ml) were used for this study. Both microorganisms were grown for 24 hours in dilutions of nicotine (0 to 32 mg/ml) made in tryptic soy broth (TSB) or TSB supplemented with 1% sucrose (TSBS; S. mutans) or yeast peptone dextrose broth (YPD; C. albicans). Suspensions of the nicotine-treated cells were prepared, mixed together and incubated for up to 24 hours to determine if there was an increase in coaggregation of nicotine-treated cells compared to the no nicotine control cells. Qualitative analysis of coaggregation was performed using a visual aggregation assay and light microscopic observation. A spectrophotometric assay was used to provide a quantitative analysis of the coaggregation. Results The visual aggregation assay indicated a significant increase in coaggregation between C. albicans and S. mutans with increasing incubation time (0 to 24 hours) and nicotine concentrations (0 to 4 mg/ml). Microbial growth in nicotine at 4 mg/ml demonstrated a significant increase in coaggregation after 24 hours of incubation. The numbers of coaggregated S. mutans/C. albicans cells exhibited a significant increase with incubation time and nicotine concentrations when the samples were examined microscopically. More coaggregation of S. mutans and C. albicans was observed with incubation time and increased nicotine compared to the 0 mg/ml nicotine group. There was a noticeable increase of coaggregation when cells were grown in TSBS compared to TSB. Absorbance of nicotine-treated cells (0.25 to 4 mg/ml) exhibited a decrease in values compared to 0 mg/ml at 0 hours of incubation, confirming increased coaggregation. Conclusion These results demonstrated the effect of nicotine in increasing the coaggregation of S. mutans with C. albicans. Coaggregation increased with incubation time and nicotine concentration. Coaggregation was increased with S. mutans grown in TSBS compared to TSB, suggesting that growth in sucrose media leads to an increase in receptors responsible for coaggregation.Item Photo Inactivation of Streptococcus mutans Biofilm by Violet-Blue light(Springer, 2016-09) Gomez, Grace F.; Huang, Ruijie; MacPherson, Meoghan; Zandona, Andrea G. Ferreira; Gregory, Richard L.; Department of Biomedical and Applied Sciences, IU School of DentistryAmong various preventive approaches, non-invasive phototherapy/photodynamic therapy is one of the methods used to control oral biofilm. Studies indicate that light at specific wavelengths has a potent antibacterial effect. The objective of this study was to determine the effectiveness of violet-blue light at 380–440 nm to inhibit biofilm formation of Streptococcusmutans or kill S. mutans. S. mutans UA159 biofilm cells were grown for 12–16 h in 96-well flat-bottom microtiter plates using tryptic soy broth (TSB) or TSB with 1 % sucrose (TSBS). Biofilm was irradiated with violet-blue light for 5 min. After exposure, plates were re-incubated at 37 °C for either 2 or 6 h to allow the bacteria to recover. A crystal violet biofilm assay was used to determine relative densities of the biofilm cells grown in TSB, but not in TSBS, exposed to violet-blue light. The results indicated a statistically significant (P < 0.05) decrease compared to the non-treated groups after the 2 or 6 h recovery period. Growth rates of planktonic and biofilm cells indicated a significant reduction in the growth rate of the violet-blue light-treated groups grown in TSB and TSBS. Biofilm viability assays confirmed a statistically significant difference between violet-blue light-treated and non-treated groups in TSB and TSBS. Visible violet-blue light of the electromagnetic spectrum has the ability to inhibit S.mutans growth and reduce the formation of S.mutans biofilm. This in vitro study demonstrated that violet-blue light has the capacity to inhibit S. mutans biofilm formation. Potential clinical applications of light therapy in the future remain bright in preventing the development and progression of dental caries.Item Role of Hemin and Growth Media on the Autofluorescence of Streptococcus mutans(Office of the Vice Chancellor for Research, 2015-04-17) Gomez, Grace F.; Eckert, George J.; Gregory, Richard L.Abstract Carious lesions fluoresce under blue light. The primary cariogenic bacterium Streptococcus mutans has been shown previously to fluoresce within blue light wavelengths. In this study we wanted to determine the role of hemin and various growth media on the fluorescing properties of S. mutans under planktonic (total biomass) and biofilm (biofilm mass) growth conditions. UA159 was grown for 24 h at 37ºC in Tryptic Soy Broth (TSB), Brain Heart Infusion (BHI) and Todd Hewitt broth (THB) with and without hemin in 5% CO2. Biofilm was grown for 24 h in a 96 well sterile microplate in the above described media with and without hemin. A stock solution of Protoporphyrin –IX was prepared and diluted to concentrations ranging from 1.6-3.1×10-4 g/ml. A SpectraMax (M3) was used to determine the fluorescence from UA159. RFU of total biomass and biofilm mass was assessed by exciting at fixed wavelengths of 385 and 405 nm at a spectral band width of 10 nm. Emission spectra at 770 nm were observed with 385 nm and an emission of 800 and 810 nm with 405 nm. ANOVA on the ranks of the measurements was used, with four different factors including wavelength (770, 800 and 810 nm); total biomass or biofilm mass; various growth media (TSB, BHI, THB) and the presence/absence of hemin and interactions among the factors. The analysis allowed each media-hemin combination to have different variances. Hemin decreased the amount of fluorescence; regardless of the levels of the other factors (p≤0.0001). Without hemin, BHI had more fluorescence than THB (p≤0.0003) and TSB (p≤0.0001). However with hemin, THB had more fluorescence than BHI (p≤0.0001) and TSB (p≤0.0001). The role of hemin and porphyrin-related compounds in the metabolism of S. mutans should be elucidated.