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Item Analyses of the Effects of Arginine, Nicotine, Serotype and Collagen-Binding Proteins on Biofilm Development by 33 Strains of Streptococcus mutans(Frontiers Media, 2021-11-25) Wagenknecht, Dawn R.; Gregory, Richard L.; Biomedical Sciences and Comprehensive Care, School of DentistryStreptococcus mutans serotype k strains comprise <3% of oral isolates of S. mutans but are prominent in diseased cardiovascular (CV) tissue. Collagen binding protein (CBP) genes, cbm and cnm, are prevalent in serotype k strains and are associated with endothelial cell invasion. Nicotine increases biofilm formation by serotype c strains of S. mutans, but its effects on serotype k strains and strains with CBP are unknown. Saliva contains arginine which alters certain properties of the extracellular polysaccharides (EPS) in S. mutans biofilm. We examined whether nicotine and arginine affect sucrose-induced biofilm of S. mutans serotypes k (n = 23) and c (n = 10) strains with and without CBP genes. Biofilm mass, metabolism, bacterial proliferation, and EPS production were assessed. Nicotine increased biomass and metabolic activity (p < 0.0001); arginine alone had no effect. The presence of a CBP gene (either cbm or cnm) had a significant effect on biofilm production, but serotype did not. Nicotine increased bacterial proliferation and the effect was greater in CBP + strains compared to strains lacking CBP genes. Addition of arginine with nicotine decreased both bacterial mass and EPS compared to biofilm grown in nicotine alone. EPS production was greater in cnm + than cbm + strains (p < 0.0001). Given the findings of S. mutans in diseased CV tissue, a nicotine induced increase in biofilm production by CBP + strains may be a key link between tobacco use and CV diseases.Item Antibacterial Activities of Methanol and Aqueous Extracts of Salvadora persica against Streptococcus mutans Biofilms: An In Vitro Study(MDPI, 2021-12-01) Balhaddad, Abdulrahman A.; Mokeem, Lamia; Melo, Mary Anne S.; Gregory, Richard L.; Biomedical Sciences and Comprehensive Care, School of DentistryThe use of herbal products in oral hygiene care has a long history, and their use is popular today. A tree stick, named Salvadora persica (S. persica), is commonly used to remove dental plaque and clean teeth in many countries. In addition, extracts of S. persica can be used as a mouthwash, as they demonstrate antimicrobial properties. This study aimed to investigate the antibacterial effect of S. persica methanol and aqueous extracts against Streptococcus mutans (S. mutans) biofilm. A S. mutans biofilm formation assay was conducted using different concentrations of S. persica methanol or water extracts in tryptic soy broth (TSB) supplemented with 1% sucrose. The biofilm was stained with crystal violet dye, and the absorbance was assessed to examine biofilm formation. One-way analysis of variance (ANOVA) and Tukey tests were used to analyze the results. The S. persica methanol extract displayed a significant inhibition (p ≤ 0.001) against the S. mutans biofilm. The 10 mg/mL concentration of the S. persica methanol extract was determined as the minimum biofilm inhibitory concentration (MBIC). The used methanol concentration, mixed with TSB supplemented with 1% sucrose and without the S. persica extract, did not inhibit the S. mutans biofilm. The S. persica aqueous extract did not demonstrate any biofilm inhibition at any concentration (p ≥ 0.05). The findings of this study suggest the potential of using S. persica methanol extract as a mouthwash or adjunctive to oral hygiene tools.Item The anticariogenic effect of xylitol on seven Streptococcus mutans strains(Discovery Scientific Society, 2021-07) Nassar, Hani M.; Alhazzazi, Turki Y.; Hazzazi, Loai W.; Gregory, Richard L.; Biomedical Sciences and Comprehensive Care, School of DentistryIntroduction: Xylitol can affect caries-inducing bacteria; however, different Streptococcus mutans strains might respond differently. Aim: To investigate the effect of xylitol on biofilm formation and metabolic activity of seven S. mutans strains. Methods: Seven S. mutans strains (UA159, A32-2, NG8, 10449, UA130, LM7, and OMZ175) were inoculated into 96-well microtiter plates and were tested with various xylitol concentrations (0.0, 0.0016, 0.0031, 0.0063, 0.0125, 0.025, 0.05, 0.1, 0.2, 0.4 and 0.8 g/mL) for inhibition of biofilm formation and bacterial metabolic activity by recording absorbance values. Lactate dehydrogenase and extracellular polysaccharide assays were conducted at 0.0, 0.1, 0.2, 0.4, and 0.8 g xylitol/mL. Data were analyzed by one-way analysis of variance, Tukey’s, paired t, and LSD tests at 0.05 significance level. Results: Xylitol produced a significant decrease in bacterial biofilm formation compared to controls at 0.4 g/mL, with almost complete lack of biofilm formation at 0.8 g/mL. This was consistent with metabolic activity which demonstrated a significant activity reduction occurring for all strains at 0.4 g/mL, and a complete lack of activity at 0.8 g/mL for all seven strains. There was a trend for lower LDH and EPS production with the increase in xylitol concentration especially with UA159, UA130, and NG8. Conclusion: Xylitol has a clear anticariogenic effect on S. mutans which was slightly different depending on the tested strain confirming that the benefit of xylitol might vary from one patient to another. The effect is more apparent at concentrations of 0.4 g/mL and higher.Item Combined Effects of Soda Drinks and Nicotine on Streptococcus Mutans Metabolic Activity and Biofilm Activity(2019) Mokeem, Lamia Sami; Gregory, Richard; Cook, Norman Blaine; Windsor, Jack; Eckert, GeorgeItem Combined effects of soft drinks and nicotine on Streptococcus mutans metabolic activity and biofilm formation(J-STAGE, 2021-01) Mokeem, Lamia S.; Willis, Lisa H.; Windsor, L. Jack; Cook, N. Blaine; Eckert, George; Gregory, Richard L.; Cariology, Operative Dentistry and Dental Public Health, School of DentistryThe purpose of this study was to explore the effects of nicotine on the activity of Streptococcus mutans (S. mutans) in soft drinks. Regular soft drinks contain large proportions of high-fructose corn syrup (HFCS), which increases the activity of S. mutans resulting in high-caries risk compared with sugar-free soft drinks. Nicotine use exhibits a strong correlation with increased S. mutans biofilm formation. The soft drinks chosen were (Coca-Cola Classic, Diet Coke, Coca-Cola Zero Sugar, Caffeine-Free Coca-Cola, Caffeine-Free Diet Coke, Caffeine-Free Coca-Cola Zero Sugar). S. mutans was grown overnight in tryptic soy broth; nicotine was diluted in tryptic soy broth supplemented with 1.0% sucrose followed by soft drinks in dilution of 1:3. Total growth absorbance and biofilm growth were determined by spectrophotometry, absorbance measured to determine biofilm formation, and metabolic activity quantified. One-way ANOVA showed a considerable effect for HFCS and caffeine in the presence of nicotine and their interaction in all measures. Results showed sugar-free caffeinated colas demonstrated significant effect in inhibiting S. mutans biofilm formation and metabolic activity with nicotine. Nicotine-induced S. mutans increased biofilm formation and metabolic activity in the presence of HFCS and caffeine in soft drinks. In conclusion, smokers should consider sugar-free caffeinated versions to minimize the chance of developing dental caries dut to the reduction of biofilm formation.Item A comparison of IgA antibody levels in caries-resistant and caries-susceptible children(1993) Rose, Paul Todd; Hughes, Christopher V.; Avery, David R.; Sanders, Brian J.; Branca, Ronald A.; Gregory, Richard L.Secretory immunity is believed to play a role in natural resistance to dental caries. Although dental caries has dramatically decreased in children in the United States, there remains a population of caries-susceptible children even in fluoridated communities. Previous studies have shown a positive correlation between salivary immunoglobulin A (sIgA) antibody levels to Streptococcus mutans and caries resistance in adults. In the present study, an enzyme-linked immunosorbent assay (ELISA) was used to compare IgA antibody levels to S. mutans in saliva from 20 caries susceptible (DMFS greater than 5) and 20 caries-resistant (DMFS less than or equal to 1) children (ages 7-11). All subjects resided in fluoridated communities. Salivary S. mutans numbers were significantly higher (p ≤ 0.05) in the caries susceptible (31.2 percent of total streptococci) group than in the caries resistant (1.6 percent of total streptococci) group. Whole saliva from caries-resistant children had significantly higher (p = 0.05) levels of IgA antibodies to S. mutans than saliva from caries-susceptible children. However, whole saliva from caries-resistant children had similar levels of IgA1 or IgA2 antibodies against S. mutans to saliva from caries-susceptible children. These results suggest that IgA antibody to S. mutans may play a role in natural protection from dental caries in children and confirm previous reports indicating a role for salivary IgA antibodies to S. mutans in mediation of caries.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 Sciences and Comprehensive Care, 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 Caffeine on the Growth of Streptococcus mutans(Office of the Vice Chancellor for Research, 2015-04-17) DuBois, Aubrey E.; Gregory, Richard L.Caffeine consumption is a staple of the typical adult diet. Previous research has demonstrated many possible health benefits of regular consumption of caffeine-containing beverages such as coffee and tea. Coffee may contain up 200 mg caffeine/cup (84 μg/ml). This study investigated the correlation between oral health and caffeine consumption by observing the effects of the compound on the growth of a leading contributor to tooth decay, Streptococcus mutans. Assays were performed to examine the effect of different concentrations of caffeine on both the planktonic and biofilm growth of the bacteria. Caffeine concentrations of 200 and 400 μg/ml demonstrated significant biofilm formation enhancement (p<0.05). Contrastingly, concentrations from 31.25 through 100 μg/ml caused a slight, significant inhibition in biofilm formation. Planktonic growth of S. mutans was marginally inhibited in concentrations of 31.25 through 200 μg/ml. The results of this study indicate a potential for adverse side effects to oral health when caffeine is consumed in high concentrations. Lower concentrations such as those naturally found in coffee and tea may inhibit formation of biofilm and dental plaque, thereby promoting good oral health.Item Effect of Green Tea on Streptococcus mutans Metabolic Activity, Planktonic Growth, and Biofilm Activity in the Presence of Nicotine(Office of the Vice Chancellor for Research, 2013-04-05) Gardner, R.; Foltz, J.; Li, Mingyun; Huang, Ruijie; Gregory, Richard L.Streptococcus mutans is the main bacterial cause of dental caries, and it has been proven by previous research that its growth is affected by various concentrations of nicotine and other agents. The amount of S. mutans in the mouth is directly proportional to the number of dental cavities. Studies have shown that smokers have an increased amount of caries, much of which is due to the low concentrations of nicotine the mouth is exposed to. It is known that S. mutans thrives in low-moderate concentrations of nicotine, and that nicotine is a promoting agent for S. mutans. S. mutans has also been proven as a contributor to atherosclerosis, resulting from dental plaque entering the bloodstream. Green Tea is a commonly consumed beverage, which has been known to reduce the number of dental cavities. Previous research has concluded that green tea contains polyphenols, which have antimicrobial effects, including an inhibitory effect on S. mutans. The objective of this research is to observe how green tea affects S. mutans metabolic activity, as well as biofilm and planktonic growth, in the presence of nicotine. The experiments compared S. mutans treated with nicotine concentrations (0-8 mg/ml), and S. mutans treated with a 2.5 g/200 mL concentration of Sencha Jade Reserve Japanese green tea in conjunction with the various nicotine concentrations. The assays were performed in a microtiter plate; the XTT and biofilm assays measured absorbance, and the planktonic assay measured kinetic growth. The experiments conclude that green tea has an inhibitory effect on nicotine-treated S. mutans metabolic activity and planktonic growth, with higher concentrations of green tea inhibiting more effectively. It was also concluded that green tea increases biofilm formation. These conclusions provide evidence of the inhibitory effect green tea has on nicotine-treated S. mutans, and may indicate a way to reduce the incidence of caries and atherosclerosis.Item Effect of nicotine on biofilm formation of Streptococcus mutans isolates from smoking and non-smoking subjects(Taylor and Francis, 2019-01-01) El-Ezmerli, Nasreen F.; Gregory, Richard L.; Biomedical Sciences and Comprehensive Care, School of DentistryObjectives: To investigate effects of nicotine on biofilm formation of Streptococcus mutans isolates from oral washes of smoker and non-smoker human subjects.Materials and methods: This study was conducted using 60 S. mutans isolates with three S. mutans isolates collected from oral washes of ten smoking subjects and ten from non-smoking subjects. Biofilm was formed by culturing each S. mutans strain (10 μl) in 190 μl of TSB supplemented with 1% sucrose (TSBS) containing 0, 0.25, 0.5, 1.0, 2.0, 4.0, 8.0, 16.0, and 32.0 mg/ml of nicotine for 24 h in 5% CO2 at 37°C in 96 well microtiter plates. The absorbance values of biofilm were measured at 490 nm in a microplate spectrophotometer.Results: There was a significant effect (p-value < 0.05) of nicotine concentrations and smoking on the growth of biofilm, planktonic cells, and total absorbance, for all strains of S. mutans. Isolates from smokers had significantly more biofilm at 0–16 mg/ml of nicotine compared to those from non-smokers (p-value < 0.0001).Conclusion: S. mutans smoker isolates are more affected by high nicotine concentrations than non-smoker isolates.Clinical Relevance: The use of nicotine products increases the growth of S. mutans and may place tobacco users at risk for dental decay.