Mutans Isolates Research Articles

Evaluation of Anti-biofilm Activity of Silica nanoparticles against dental Streptococcus mutans: In-Vitro Study

Background: Dental caries is a prevalent chronic infection caused by cariogenic bacteria that cling to teeth, mainly Streptococcus mutans. These bacteria break down sugars into acid over time, demineralizing the tooth structure. Biofilm as an early colonizer and is the most important bacterium in the formation of dental caries . Materials and Methods: The present study concentrates on isolation and identification of the S.mutans pathogen from patients suffering from dental caries. On specific media, the isolates were cultivated. From human teeth, one hundred dental caries samples are gathered. Using MS-agar (Mitis Salivarius agar), only fifty samples are regarded as positive bacterial isolates to identify the S.mutans isolates, their morphological, cultural, biochemical, and VITEK features were examined. . The microtiter plate method was used to detect the development of biofilms by comparing the characteristics of the isolates. In this work, the median values of optical density (OD) at 630 nm were utilised to employ ELISA to distinguish between S. mutans isolates that developed biofilms and those that did not. To confirm the type of bacteria using molecular identification. Objectives: The purpose of this work is to determine the subminimum and minimum inhibitory concentrations of silica nanoparticles that function as anti-biofilm agents, as they have an impact on bacterial growth and details of silica nanoparticle characterization. Results: In this study considered to be S.mutans are twenty isolates the morphological, cultural, biochemical, and VITEK properties of the bacterial isolates were used to identify them . in this study molecular identification to more confirm of S.mutans the result of PCR showed that 20/20(100%) isolates were PCR positive for 16r RNA gene . in this investigation different isolates were evaluated for their capacity to create biofilm; nine (45%) were judged to be strong producers, seven (35%), to be moderate producers, and four (20%) to be weak producers. the sol-gel method was used in this study to synthesis silica nanoparticles, which were then characterised by XRD, FE-SEM, EDX, and TEM. In this study, Si-NPs were tested against isolates of S.mutans by using a resazurin mediated microtiter plate the results have shown that silica nanoparticles has inhibitory action (MIC) against S. mutans with concentration 0,78 mg/ml and sub-MIC with concentration 0.39 mg/ml. .The anti-biofilm effect of silica nanoparticles on isolates of S. mutans in the current investigation revealed that the biofilm mean of control (biofilm development without silica nanoparticles) was 0.583, whereas the findings for anti-biofilm utilizing sub-MIC of NPs, was 0.185 in microtiter plate method when the (p value) less than 0.05 this proves the NPs effected on the biofilm formation by S.murans and inhibition it . Conclusion: Silica nanoparticles' anti-biofilm action on S.mutans isolates demonstrated the importance of biofilm inhibition.

Biofilm Production Activity and Antibiotics Susceptibility of Streptococcus mutans Isolates from Patients with Dental Caries

Dental caries, a prevalent chronic infectious ailment, commences with the adherence of cariogenic bacteria to the teeth, particularly Streptococcus mutans. These microorganisms utilize sugars to produce acid, resulting in the gradual demineralization of the dental structure. Streptococcus mutans a gram positive bacterium is found to play the most significant role in the dental caries in terms of being the earliest colonizer in the formation of caries. The current research focuses on the isolation and identification of the Streptococcus mutans pathogen in individuals affected by dental caries. The isolated strains were cultured on specific media, with one hundred samples of dental caries obtained from human teeth. Among these samples, fifty were identified as positive bacterial isolates utilizing MS-agar (Mitist Salivares agar), with ten isolates categorized under the group Streptococci and twenty isolates identified as S. mutans. Identification of bacterial isolates was carried out based on morphological and cultural characteristics, biochemical tests, and VITEK-II. Additionally, an antibiogram assay was conducted to assess the susceptibility and resistance of the pathogen to various drugs. The findings revealed that all locally isolated S. mutans strains displayed 100% resistance to Cefepime, while showing different levels of resistance and susceptibility to other antibiotics such as Doxycycline, Amoxicillin-clavulanic acid, Trimethoprim-sulfamethoxazole, Vancomycin, Tetracycline, Ampicillin, and Meropenem. In this study, ELISA was employed to differentiate between Streptococcus. mutans isolates that produced biofilms and those that did not by utilizing the optical density at 630 nm. Bacterial biofilms contribute to chronic diseases that pose challenges in treatment. Twenty distinct isolates were evaluated for their biofilm-forming capabilities, with 9 (45%) identified as strong biofilm producers, 7 (35%) classified as moderate biofilm producers, and 4 (20%) categorized as weak biofilm producers

Antibacterial and Antibiofilm Potential of Chlorophyllin Against Streptococcus mutans In Vitro and In Silico.

Streptococcus mutans is a leading causative agent of dental caries and exerts pathogenicity by forming biofilms. Dental caries continues to be a significant public health issue worldwide, affecting an estimated 2.5 billion people, showing a 14.6% increase over the past decade. Herein, the antibacterial potential of Chlorophyllin extracted from Spinacia oleracea was evaluated against biofilm-forming S. mutans via in vitro and in silico studies. The antimicrobial activity of chlorophyllin extract against S. mutans isolates was tested using the agar well diffusion method. Chlorophyllin extract was also tested against biofilm-forming isolates of S. mutans. Chlorophyllin was docked with the antigen I/II (AgI/II) protein of S. mutans to evaluate its antimicrobial mechanism. The chemical structure and canonical SMILES format of Chlorophyllin were obtained from PubChem. Additionally, adsorption, distribution, metabolism, excretion, and toxicity (ADMET) analyses of Chlorophyllin were performed using ADMETlab 2.0 to assess its pharmacokinetic properties. An agar well diffusion assay revealed that all S. mutans isolates were susceptible to Chlorophyllin extract and showed a variety of inhibition zones ranging from 32 to 41 mm. Chlorophyllin reduces the biofilm strength of four isolates from strong to moderate and six from strong to weak. The antibiofilm potential of Chlorophyllin was measured by a reduction in the number of functional groups observed in the Fourier Transform Infrared Spectrometer (FTIR) spectra of the extracellular polymeric substance (EPS) samples. Chlorophyllin showed binding with AgI/II proteins of S. mutans, which are involved in adherence to the tooth surface and initiating biofilm formation. The ADMET analysis revealed that the safety of Chlorophyllin exhibited favorable pharmacokinetic properties. Chlorophyllin stands out as a promising antibacterial and antibiofilm agent against biofilm-forming S. mutans, and its safety profile highlights its potential suitability for further investigation as a therapeutic agent.

Intraspecies interactions of Streptococcus mutans impact biofilm architecture and virulence determinants in childhood dental caries.

Early childhood dental caries (ECC) is the most common chronic disease among children, especially among low socioeconomic populations. Streptococcus mutans is most frequently associated with initiation of ECC. Although many studies report children with multiple S. mutans strains (i.e., genotypes) have greater odds of developing ECC, studies investigating intraspecies interactions in dental caries are lacking. This study investigates the impact of intraspecies interactions on cariogenic and fitness traits of clinical S. mutans isolates using in vitro and in vivo approaches. Association analysis evaluated if presence of multiple S. mutans genotypes within the first year of colonization was associated with caries. Initially, clinical S. mutans isolates from 10 children were evaluated. S. mutans strains (G09 and G18, most prevalent) isolated from one child were used for subsequent analysis. Biofilm analysis was performed for single and mixed cultures to assess cariogenic traits, including biofilm biomass, intra-polysaccharide, pH, and glucan. Confocal laser scanning microscopy (CLSM) and time-lapse imaging were used to evaluate spatial and temporal biofilm dynamics, respectively. A Drosophila model was used to assess colonization in vivo. Results showed the mean biofilm pH was significantly lower in co-cultured biofilms versus monoculture. Doubling of S. mutans biofilms was observed by CLSM and in vivo colonization in Drosophila for co-cultured S. mutans. Individual strains occupied specific domains in co-culture and G09 contributed most to increased co-culture biofilm thickness and colonization in Drosophila. Biofilm formation and acid production displayed distinct signatures in time-lapsed experiments. This study illuminates that intraspecies interactions of S. mutans significantly impacts biofilm acidity, architecture, and colonization.IMPORTANCEThis study sheds light on the complex dynamics of a key contributor to early childhood dental caries (ECC) by exploring intraspecies interactions of different S. mutans strains and their impact on cariogenic traits. Utilizing clinical isolates from children with ECC, the research highlights significant differences in biofilm architecture and acid production in mixed versus single genotype cultures. The findings reveal that co-cultured S. mutans strains exhibit increased cell density and acidity, with individual strains occupying distinct domains. These insights, enhanced by use of time-lapsed confocal laser scanning microscopy and a Drosophila model, offer a deeper understanding of ECC pathogenesis and potential avenues for targeted interventions.

Association between the biofilm formation of streptococcus mutans, dental caries experience, and resistance to antibiotics in adult patients

Objectives: The aim of this study was to consider the potential association between the formation of bucall mucusa streptococcus biofilms and a high DMFT index, as well as the occurrence of antibiotic resistance among adult patients in Sana'a, Yemen. Study design: A total of 100; 34-85 year old. Clinical examinations of patients were performed to estimate dental caries experience with the Silness-Loe index, as well as bucall mucusa swabs were collected to assess biofilm production by the phenotypic method, i.e., tissue culture palate methods (TCPM). Finally, the antibiogram susceptibility pattern of isolated S. mutans was done by the Kirby-Bauer disc diffusion method for L-Lactam antibiotics (as ampicillin and penicillin) and non-L-Lactam antibiotics (clindamycin, erythromycin, lincomycin, and vancomycin). Results: When isolated S. mutans were exposed to biofilm detection by the TCP method, 1 (1.2%) showed strong biofilm formation capacity, 71 (86.6%) showed moderate biofilm formation capacity, and 10 (12.2%) showed non/weak formation capacity of biofilm. There was an escalation in the rate of formation of S. mutans biofilms with an increased degree of caries index. The S. mutans biofilms positively showed a higher rate of resistance than non/weak biofilm formation, e.g., ampicillin (91.1% versus 8.9%, p <0x7E> 0.0001), tetracycline (87.8% versus 12.2%, p <0x7E> 0.0001), and co-trimoxazole (90%% versus 10%, p <0x7E> 0.0001), etc. Conclusion: The present study proved that S. mutans is still the major bacteria isolated from the oral cavity, but few persons might not have a significant number of S. mutans in the oral cavity. The S. mutans biofilm producers were more able to cause dental caries compared to the S. mutans biofilm non-producers. Drug-resistant factor in the S. mutans isolates was found to be associated with S. mutans biofilm formation.

Detection of SpaP and vicR Genes in Non-biofilm Producing Streptococcus mutans Isolated from Dental Caries and Plaque

Evidence accumulated over many decades has clearly shown that Streptococcus mutans is a major agent in dental caries. S. mutans SpaP gene (Cell Surface Antigen) mediates the binding of these cariogenic bacteria to tooth surfaces. The aim of this study was to detect the presence of SpaP and vicR genes in non-biofilm forming Streptococcus mutant isolated from dental plaques. In this study, 120 clinical samples were obtained from different dental clinics and hospitals in Baghdad. 26 Bacterial isolates identified as Streptococcus mutans by using selective media (Mitis-salivarius agar) and microscopic examination after gram staining then confirmed S. mutans by molecular detection using SpaP genes. Interestingly all 26 isolates were non-biofilm forming after microtiter plate assay based on the crystal violet staining method, and negatively resulted for the detection of vicR gene which responsible for biofilm formation. 10 antibiotics (Doxycycline, Ofloxacin, Tetracycline, Erythromycin, Vancomycin, Clindamycin, Rifampicin, Imipenem, Amikacin and Cefepime) were used to examine sensitivity of S. mutans isolates, all isolates were multi-drugs resist (MDR) for all 10 antibiotics used. We can conclude that absence of biofilm forming ability does not affecting antibiotic resistance and virulence of Streptococcus mutance.

Comparative Assessment of Antimicrobial Activity of Propolis and Chlorhexidine on Salivary Isolates of Candida albicans and Streptococcus mutans in Children with Severe Early Childhood Caries: An In Vitro Study.

Streptococcus mutans and Candida albicans are the chief microbes associated with severe early childhood caries (S-ECC). Diverse antimicrobial agents are widely used to prevent ECC, and a quest for newer natural products has been on the rise in the recent past. To estimate the antimicrobial activity of propolis with chlorhexidine on salivary specimens from children with S-ECC in vitro. A total of 60 children with S-ECC were designated. Salivary samples of 30 children (group I) were inoculated onto mitis salivarius agar (MSA) to isolate S. mutans. Another 30 samples (group II) were inoculated on sabouraud's dextrose agar and subcultured on HiCrome Candida differential agar to isolate C. albicans. Sensitivity testing for 0.2% chlorhexidine and 10% propolis extract was done using the agar well diffusion technique using Mueller-Hinton agar medium. The antimicrobial effect was evaluated by calculating the diameter of the zone of inhibition surrounding the well. All saliva samples collected from groups I and II showed growth of S. mutans and C. albicans, respectively. All cultured microbes were sensitive to 0.2% chlorhexidine and 10% propolis extract. The mean inhibition zone for S. mutans with chlorhexidine was 14.57 ± 0.63 mm, and with propolis, 11.93 ± 0.52 mm. The mean zone of inhibition for C. albicans with chlorhexidine was 12.83 ± 0.59 mm, and with propolis, 9.50 ± 0.73 mm. Chlorhexidine consistently showed statistically significantly larger zones of inhibition and hence appeared to be a more potent antimicrobial agent than propolis extract for both S. mutans and C. albicans. However, propolis has irrefutable action against both S. mutans and C. albicans. Propolis may be an acceptable substitute for chlorhexidine for long-term use as it has demonstrated antimicrobial activity and fewer side effects. Hence, this Association of Physicians of India herbal drug can be incorporated into mouthwashes and toothpaste to reduce microbial counts. Kodgi V, Shetty P, Thimmaiah C, et al. Comparative Assessment of Antimicrobial Activity of Propolis and Chlorhexidine on Salivary Isolates of Candida albicans and Streptococcus mutans in Children with Severe Early Childhood Caries: An In Vitro Study. Int J Clin Pediatr Dent 2024;17(5):591-595.

Bacteriophage Endolysins Antibacterial Effect against S. mutans Isolated from Dental Caries: An In-Vitro Study

Background: Many bacterial infections including dental caries that caused by Streptococcus mutans (S. mutans) were difficult to be treated due to the unavailability of effective antibiotics. Therefore, it requires to focus the attention on endolysin as an antibacterial agent. This study was designed to evaluate the In-vitro effectiveness of specific bacteriophage endolysins against S. mutans isolated from dental caries.Methods: Twenty-five isolates of S. mutans were collected from patients with dental caries attended to Al-Imamain Al-Kadhimain, Medical City Hospital, and AL-Ameen Health Center, during the period from December 2021 to May 2022. The S. mutans isolates were identified by culturing them on their specific mitis salivarius sucrose bacitracin tellurite media (MS-SBT agar), morphological characteristics, gram staining, biochemical tests and VITEK 2 compact system. Seven of these S. mutans isolates were used for the in-vitro study.Results: The bacteriophage endolysin enzyme extracted from the corresponding S. mutans isolate was effective as In-vitro antibacterial agent with minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) ranging between (13.63 – 17.83) µg/ml and (17.04-22.28) µg/ml respectively.Conclusion: The endolysin was effective as antibacterial agent against multi-drug resistant (MDR) S. mutans upon growing and testing on culture media.

Probiotics, A New Frontier in Oral Health: Unraveling the Efficacy of Probiotic Metabolites vs. Chlorhexidine on Streptococcus mutans in Dental Plaque

The battle against dental caries has long been fought with conventional antimicrobial agents, with Chlorhexidine leading the charge. However, recent research has delved into the potential of probiotic metabolites as an alternative or adjunctive therapy in the quest for better oral health. In this editorial, we explore the emerging evidence surrounding the efficacy of probiotic metabolites compared to Chlorhexidine, specifically targetting local clinical isolates of Streptococcus mutans within dental plaque. Streptococcus mutans, a key player in the formation of dental plaque, has been a prime target for antimicrobial interventions due to its significant role in tooth decay. Chlorhexidine, a widely used antiseptic, has been a stalwart in dental care. However, its efficacy and potential side effects have prompted researchers to seek alternative approaches. Probiotic metabolites, byproducts of beneficial bacteria, have emerged as a promising avenue, with studies suggesting their ability to promote oral health by modulating the microbial balance.1 Recent studies have begun to unravel the intricate interactions between probiotic metabolites and Streptococcus mutans. These investigations have not only explored the direct antimicrobial effects but also the potential for probiotics to influence the overall microbial environment in the oral cavity. The comparison with Chlorhexidine, a gold standard in dental care, adds depth to our understanding of these alternative therapies.2 The evidence supporting the efficacy of probiotic metabolites in controlling Streptococcus mutans is not only encouraging but also challenges the status quo. A study by Gong SG et al. (2023) demonstrated that probiotic metabolites significantly reduced the growth of Streptococcus mutans in vitro, highlighting their potential as a potent antimicrobial agent.3 This finding is particularly promising given the increasing concerns about antibiotic resistance and the need for sustainable alternatives.
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Antibiofilm and Antibacterial Properties of Herbal Extracts as Alternatives to Current Treatment Approaches: A Narrative Review

Bacterial biofilm formation poses significant challenges in the healthcare sector due to increased antibiotic resistance and persistent infections. This literature review explores the potential of some herbs and their extracts as alternative approaches to combat biofilm formation and multidrug-resistant bacteria. A detailed literature search was conducted across databases for published studies till 2023, to identify studies on medicinal plants' anti-biofilm and antibacterial properties. Key compounds within plant extracts showing anti-biofilm activity and their mechanisms of action were highlighted. A combination of keywords, MeSH terms, and Boolean operators were used to formulate the search strategy. Numerous studies demonstrated the efficacy of medicinal plants in inhibiting biofilm formation and combating multidrug-resistant bacteria. Active compounds such as benzyl (6Z,9Z,12Z)-6,9,12-octadecatrienoate, 3-benzyloxy-1-nitro-butan-2-ol, Pyridine, 3-(1-methyl-2-pyrrolidinyl)-(S), and others exhibited anti-biofilm and antibacterial potential. Extracts from Berginia ciliata, Clematis grata, and Clematis viticella showed over 80% inhibition of biofilm formation, while mango leaf extracts interfered with quorum sensing mechanisms in Pseudomonas aeruginosa PAO1. Salvadora persica extracts displayed significant biofilm inhibition against cariogenic Streptococcus mutans isolates. Medicinal plants and their extracts hold promise as alternative strategies to combat bacterial biofilms and multidrug-resistant bacteria. The identification of active compounds provides opportunities for further research and drug development. Molecular docking studies are crucial for understanding the molecular interactions between these compounds and bacterial targets, guiding the design of effective antibacterial agents based on natural compounds. Further research, including preclinical and clinical trials, is essential to validate the safety and efficacy of these extracts and their compounds for practical application in healthcare.

Comparison of the Antibacterial Efficacy of Bamboo Shoot Ethanol Extract With Chlorhexidine Mouth Rinse Against Salivary Streptococcus mutans and Lactobacillus acidophilus: An Ex Vivo Study.

Background Dental caries is the most prevalent polymicrobial oral infectious disease tormenting individuals' healthy lifestyles and presents a significant public health problem. The objective of this study was to evaluate and compare the antibacterial properties of different concentrations of bamboo shoot ethanol extract with chlorhexidine mouth rinse on isolates of salivary Streptococcus mutans (S. mutans) and Lactobacillus acidophilus (L. acidophilus). Materials and methods Non-stimulated salivary samples from 30 young adults were treated ex vivo with bamboo shoot ethanolic extract at concentrations of 30 µg/ml, 40 µg/ml, 50 µg/ml, and 60 µg/ml. The colony-forming units were quantified by measuring the number of viable bacterial cells. Inhibition zones were evaluated using the agar diffusion method. One-way ANOVA and post-hoc test were used to analyze the significant difference between variables using SPSS version 22.0 (IBM Corp., Armonk, NY). Results The mean zone of inhibition with bamboo shoot ethanolic extract against salivary S. mutans (23.00 ± 0.816) and L. acidophilus (22.00 ± 0.816) total counts was closest to the control chlorhexidine (S. mutans =22.00 ± 0.876 and L. acidophilus =21.10 ± 0.876). A greater activity against S. mutans and L. acidophilus is seen in the zone of inhibition of the 60 µg/ml experimental concentration of bamboo shoot ethanolic extract, with a significant difference in the disc diffusion assay. Conclusion The treatment with bamboo shoot extract was equivalent effective in the mentioned bacterial species. Clinical relevance It can be assured that preventive measures like mouth rinse and dentifrices compromising bamboo shoots, a potential dental biomaterial, would be optimistic agents for caries control, including the cariostatic effect.

Inhibition of Quorum Sensing Controlled Virulence Factors and Biofilm Formation of Streptococcus mutans Isolated From Orthodontic Subjects by 4-Hydroxycinnamic Acid.

Introduction Dental plaque biofilms are a collection of microorganisms that are adhered to the tooth enamel surface. Inhibition of plaque biofilms is required to prevent dental caries and periodontitis and currently, there are many chemical and herbal products in use for inhibition of biofilms but with limited success. Materials and methods Dental plaque collection was done from subjects undergoing orthodontic therapy followed by isolation of Streptococcus mutans. Isolated S. mutans were subjected to disk diffusion assay with 4-HCA (baseline 10mg/mL) for the zone of inhibition and broth micro-dilution to evaluate the minimum inhibitory concentration (MIC) and sub-MIC. Crystal violet staining was done for biofilm inhibition assay. Results Growth of S. mutans was inhibited by 4-HCA at concentrations as low as 0.31 mg/mL. 4-HCA (40μL) inhibited the bacterial growth and a clear zone (15 mm) was observed. 4-Hydroxycinnamic acids treated culture showed progressive reduction in the biofilm production at the concentration of 0.01 mg/mL. The 4-HCA concentration as low as 4 mg and 2 mg has remarkably inhibited biofilm formation of 49.3% and 34.3%, respectively. Conclusion The anti-quorum sensing and anti-biofilm activity of 4-Hydroxycinnamic acid against S. mutans isolated from subjects undergoing orthodontic treatment showed a remarkable result.

THE EFFECT OF DENTAL IMPLANTS ON AEROBIC BACTERIA COLONIZATION IN THE ORAL CAVITY AND THE ANTIBIOTIC PROFILE OF COMMON ISOLATED AEROBIC BACTERIA

Background and aims: The mouth's microflora may alter as a result of dental implants. The purpose of this study was to examine the composition of aerobic bacteria in patients with dental implants and those who had natural teeth (without implants) as well as the response of those bacteria to antibiotic treatment. Methods: Bacteriological tests were performed on 72 patients (36 dental implants and 36 natural teeth) who visited dental clinics run by Sana'a University's Faculty of Dentistry and private dental clinics. Antibiotic susceptibility tests and culture trials were carried out at the National Center for Public Health Laboratories (NCPHL) in Sana'a, Yemen. Swabs were taken from the mucous membrane of the palate and the dorsum of the tongue from both groups, and cultured on selective and non-selective solid medium, in addition to culturing the swabs on medium enriched with 5% blood, then all cultured plates were incubated to microaerophilic conditions (5% CO2) and oxygenated in 37oC for 48 hours. Then bacterial growth was identified by standard methods. Results: In implant patients, the rate of bacterial isolates from the palate and tongue was slightly higher for potentially harmful bacteria such as E. coli (8.3% in tongue implant patients vs. 2.8% in non-implant patients) and Pseudomonas aeruginosa (5.6% versus 0%). While in viridians Streptococcus including S. mutans, there was a higher colonization rate in implants patients (83.3% in the palate verses, 75% in the palate of individuals without implants). A low level of oxacillin resistance (5.1%) in S. mutans isolates but S. mutans had a substantial level of tetracycline resistance (55.93%), 11.9% for co-trimoxazole, 10.2% for erythromycin, and just 1.7% for clindamycin. Conclusion: The study found that pathogenic bacteria like E. coli and Pseudomonas aeruginosa were isolated from the palate and the back of the tongue swabs at a slightly elevated rate in implant patients; also colonization rates of Streptococcus viridians, including S. mutans, were higher in implant patients compared to those without implants. There was a significant levels of antibiotics resistance in S. aureus, CoNs and S. viridians oral isolates in both groups of tested individuals. Peer Review History: Received: 14 June 2023; Revised: 13 July; Accepted: 17 August, Available online: 15 September 2023 Academic Editor: Dr. A.A. Mgbahurike, University of Port Harcourt, Nigeria, amaka_mgbahurike@yahoo.com Received file: Reviewer's Comments: Average Peer review marks at initial stage: 6.0/10 Average Peer review marks at publication stage: 8.0/10 Reviewers: Dr. Rola Jadallah, Arab American University, Palestine, rola@aauj.edu Dr. Bilge Ahsen KARA, Ankara Gazi Mustafa Kemal Hospital, Turkey, ahsndkyc@gmail.com

Antibacterial and Anti-biofilm Effects of Chitosan Nanoparticles on Streptococcus Mutans Isolates

Background: Dental caries is an infectious disease caused by bacterial colonization and biofilm formation. Streptococcus mutans (S. mutans) is mainly responsible for dental caries development. Considering the side effects of synthetic antibacterial agents, attempts are ongoing to find antimicrobial agents with minimal or no side effects for preventing dental caries. Based on the reported antibacterial activity of chitosan, this in vitro study aimed to assess the antibacterial and anti-biofilm effects of chitosan nanoparticles on S. mutans clinical isolates.
 Methods: S. mutans isolates were isolated from supragingival plaque and carious lesions of patients by standard biochemical tests and Polymerase Chain Reaction (PCR) of the gtfB gene. The antibacterial activity and Minimum Inhibitory Concentration (MIC), Minimum Bactericidal Concentration (MBC) of chitosan nanoparticles against S. mutans was evaluated by the agar well-plate and broth micro-dilution test, respectively. Also, the effect of chitosan nanoparticles on biofilm formation was evaluated using micro-titer plate method. Data were analyzed using ANOVA.Methods: S. mutans isolates were isolated from supragingival plaque and carious lesions of patients by standard biochemical tests and Polymerase Chain Reaction (PCR) of the gtfB gene. The antibacterial activity and Minimum Inhibitory Concentration (MIC), Minimum Bactericidal Concentration (MBC) of chitosan nanoparticles against S. mutans was evaluated by the agar well-plate and broth micro-dilution test, respectively. Also, the effect of chitosan nanoparticles on biofilm formation was evaluated using micro-titer plate method. Data were analyzed using ANOVA.
 Results: Fifteen S. mutans isolates were collected from patients. The chitosan nanoparticles synthesized had a diameter of 20–30 nm. The chitosan nanoparticles showed antibacterial activity against S. mutans isolates. MICs and MBCs ranged from 0.625-2.5 µg/ml and 1.25-5 µg/ml, respectively. All isolates evaluated in this study were biofilm-forming and 5 of these produced a strong biofilm. The chitosan nanoparticles inhibited biofilm formation at 0.75 µg/ml concentration.
 Conclusion: Chitosan nanoparticles had antibacterial and antibiofilm activity on S. mutans clinical isolates. This study suggests the potential of chitosan nanoparticles as antimicrobial agents against cariogenic Streptococci.

The Effect of Diode Laser on Viability and Antibiotic Sensitivity of Streptococcus mutans Isolated From Dental Caries

Streptococcus mutans is one of the major cariogenic microbial flora. In an attempt to determine the mutagenic effect of diode laser on the viability and antibiotic sensitivity of this bacteria; A total of 30 samples were collected from dental caries. The isolates were identified using by conventional identification methods and confirmed using VITEK2 system. Twenty-one isolates were recorded as Streptococcus spp and ten of them were identified as Streptococcus mutans. Antibiotic susceptibility profile for Streptococcus mutans isolates against ten antibiotics was tested. The results revealed that all the isolates were resistant to cefixime and cephalothin, nine of them were resistant to erythromycin, ampicillin, and nalidixic acid, seven isolates were resistant to tetracycline and bacitracin, whereas all isolates were sensitive to chloramphenicol, ciprofloxacin and gentamycin. The effect of the diode laser on the isolates (S5 and S10) was tested at different time intervals (1, 2, 5, and 10) min. The viability of Streptococcus mutans isolates was affected by the obvious change in the behaviour of the bacteria from resistance to sensitivity after Laser exposure

Effect of Non-Nutritive Sweeteners on Antibacterial Activity of Black and Green Tea Aqueous Extracts against Salivary Mutans Streptococci (in-vitro Study).

Herbal medicines, such as plants and their constituents, have been used globally to treat and cure disorders since antiquity, long before the discovery of modern drugs. Some of these items require an addition to make them more appealing to consumers. This study is an in vitro evaluation of the antibacterial activity of tea (black and green tea aqueous extracts) against salivary Mutans streptococci, followed by an analysis of the effect of non-nutritive sweeteners on the antibacterial activity of these extracts against salivary Mutans streptococci. The examined bacteria were sensitive to various doses of black and green tea aqueous extract, with the inhibition zone expanding as the concentration of the extracts rose. At a dosage of 225mg/ml for black tea extracts and 200mg/ml for green tea extracts, all Mutans isolates were destroyed. In this trial, 1% stevia or sucralose did not inhibit the antibacterial activity of any tea extract, nor did 5% stevia inhibit the antimicrobial activity of black tea extract. In addition, this concentration inhibits the antimicrobial properties of green tea extracts. In this investigation, it found that increasing the content of nonnutritive sweeteners interfered with the antibacterial activity of black and green tea aqueous extract against salivary Mutans streptococci.

Prevention of Bacterial Infiltration in Class 1 Temporary Restorations Using Zinc Oxide/Calcium Sulphate Filling Materials: An In Vitro Study

This study evaluated sealing properties of zinc oxide/calcium sulphate filling material (Plastor©) to prevent bacterial infiltration in simulated Class 1 cavities with and without the presence of a spacer apically the restoration. Twenty-eight experimental Eppendorf tubes containing Tryptic Soy Agar were prepared and divided into seven groups: group A (Plastor©), group B (Plastor© + cotton pellet), group C (Plastor© + cotton pellet soaked of m-cresyl acetate), group D (Plastor© + cotton pellet soaked of eugenol), group E (Plastor© + PTFE pellet), group F (positive control), and group G (negative control). All prepared vials were individually immersed in tubes containing 5 mL of a suspension of Streptococcus mutans (105 CFU/mL) previously prepared and then incubated at 37 °C for 5 and 10 days. Subsequently, agar was collected and tested for S.mutans using real-time PCR. Aliquots of each agar samples were separately seeded on TSA for S. mutans isolation. The real-time PCR tests were negative for S. mutans on all the vials at both T5 and T10, except for positive control resulted positive at both T5 and T10. The isolation tests showed bacterial growth of S. mutans only with the agar samples collected from the vials of positive control, confirming real-time PCR tests. This in vitro study evidenced complete sealing ability of zinc oxide/calcium sulphate in temporary restorations of Class 1 cavities without mechanical loading at 10 days and complete immersion in a bacterial suspension. Moreover, sealing ability is not influenced by presence of spacers.

The Effect of Nonnutritive Sweeteners on the Antimicrobial Activity of Eucalyptus Extracts against Salivary Mutans Streptococci (in-vitro Study).

The eucalyptus tree is an excellent source of antimicrobial agents; it is used in many oral cure products. The bitter taste of these agents could compromise their usage. Therefore, fortifying the extracts with non-nutritive sweeteners could be a promising procedure for masking their unpleasant taste. This study was an in vitro evaluation of the antimicrobial activity of eucalyptus (alcoholic and aqueous) extracts against salivary Streptococci mutans. It aimed to investigate the effect of non-nutritive sweeteners on the antimicrobial activity of these extracts against salivary S. mutans. The test microbes were sensitive to different concentrations of eucalyptus alcoholic and aqueous extract, and the inhibition zone increased as the concentration of the extracts increased. All the Mutans isolates were killed at a concentration of 75 mg/ml for the alcoholic extract and 175 mg/ml for the aqueous extracts. In this experiment, the concentration of up to 15% stevia and up to 5% sucralose did not affect the antimicrobial activity of eucalyptus alcoholic extract. While the concentration of up to 1% of stevia and sucralose did not interfere with the antimicrobial activity of aqueous eucalyptus extract against salivary S. mutans.An increase in the concentration of non-nutritive sweeteners in this experiment appeared to interfere with the antimicrobial activity of eucalyptus extract against salivary S. mutans.

The potency of herbal extracts and its green synthesized nanoparticle formulation as antibacterial agents against Streptococcus mutans associated biofilms

This study aims to determine the effects of the extracts of Streblus asper, Cymbopogon citratus, Syzygium aromaticum and its formulation of green synthesized silver nanoparticle (AgNPs) on Streptococcus mutans growth and biofilm formation. The ethanolic extracts of S. asper, C. citratus, S. aromaticum, and a mix of the three herbs demonstrated antibacterial activity against S. mutans isolates by reducing bacterial biofilm formation and decreasing bacterial cell surface hydrophobicity. The formulated AgNPs from the ethanolic extracts could enhance the antibacterial activities of the plant extracts. Molecular docking found the best interaction between luteolin isolated from C. citratus and glucosyltransferase protein (GtfB), assuming the promising anti-biofilm activity. The scanning electron microscopy revealed morphological changes in the biofilm structure and a significant decrease in the biofilm area of the AgNPs treated. The study suggested that the extracts and its application could be used as natural alternative agents with multi-action against S. mutans infections.

Periapical lesion following Cnm-positive Streptococcus mutans pulp infection worsens cerebral hemorrhage onset in an SHRSP rat model.

Cerebral hemorrhage severely affects the daily life of affected individuals. Streptococcus mutans and its adhesion factor Cnm increase the adverse effects of cerebral hemorrhages. However, the mechanism by which Cnm-positive bacteria migrate from apical lesions to cerebral hemorrhage sites is unclear. Therefore, we established an S. mutans-infected apical lesion in a rat model of hypertension and investigated the neurological symptoms associated with cerebral hemorrhage. Eighteen 12-week-old stroke-prone spontaneously hypertensive rats were randomly divided into three groups, i.e. the no infection (control), dental infection with S. mutans KSM153 wild type (Cnm positive), and KSM153 Δcnm groups. Immunofluorescent staining was performed to visualize S. mutans protein. Serum interleukin-1β levels were measured. The adhesion of S. mutans to the extracellular matrix and human fibroblast cells was also analyzed. Serum antibody titers against S. mutans were comparable between Cnm positive and knockout mutants. However, 3-10 days post-infection, neurological symptom scores and cerebral hemorrhage scores were higher in Cnm-positive rats than in knockout mutants. The localization of S. mutans-derived protein was observed in the vicinity of disrupted blood vessels. Serum interleukin-1β levels significantly increased post-KSM153 WT infection. Cnm-positive S. mutans clinical isolates showed increased adhesion to the extracellular matrix, human dental pulp cells, and human umbilical vein endothelial cells compared with the Cnm-negative S. mutans isolates. In conclusion, Cnm-positive bacteria colonize the apical lesion site using the extracellular matrix as a foothold and affect cerebral hemorrhage via the bloodstream.