Home Physical Sciences A study on the bacterial adhesion of Streptococcus mutans in various dental ceramics: In vitro study
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A study on the bacterial adhesion of Streptococcus mutans in various dental ceramics: In vitro study

  • Fuat Bislimi EMAIL logo , Jagoda Bajevska , Mrinmoy Garai and Arianit A. Reka
Published/Copyright: November 4, 2020
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Open Chemistry
From the journal Open Chemistry Volume 18 Issue 1

Abstract

Streptococcus mutans (S. mutans) has been identified as a major etiologic agent of human dental caries and forms a significant proportion of oral streptococci in carious lesions. This study investigates the correlation of surface properties (effect of contact angle [CA] and free surface energy) on three restorative materials (zirconia, nickel–chromium–molybdenum alloy and composites) used in dental prosthetics with bacterial adhesion to S. mutans. Ten samples of each material (zirconia, nickel–chromium–molybdenum alloy and composites) of 8 mm diameter and 2.5 mm thickness were used. Aqueous CA measurements, free surface energy and bacterial adhesion to the sample surfaces were performed. Bacterial adhesion is determined by planting samples in the blood agar cultures and using an electron microscope (scanning electron microscopy [SEM]). The highest values of bacterial adhesion are found in composites, followed by the metal alloy, while the lowest values are observed in zirconia. Measurements show that zirconia has 17 bacteria; Ni–Cr–Mo alloy has 65, while the composite has 80 bacteria. The composites showed the highest degree of bacterial adhesion, compared to the other investigated materials, which correlates with the free surface energy of the samples (24.31 mJ/m2 for zirconia, 31.78 mJ/m2 for Ni–Cr–Mo alloy and 48.82 mJ/m2 for the composite).

Keywords: Streptococcus mutans; bacterial adhesion; hydrophobicity; free surface energy; SEM

1 Introduction

Dental implants and prosthodontic restorations are one of the most routinely used treatment options for the replacement of missing teeth [1,2,3]. The oral microflora and its interactions with the implant substrata seem to have a crucially impact on the long-term success or failure of dental implants. Once the implant surfaces are exposed to the human oral cavity, they are promptly colonized by microorganisms [4,5,6,7].

The biological response to the dental implants or prosthodontic restorations is determined by a number of physical and chemical features. These features include mechanical properties and physiochemical properties such as chemical composition, surface energy, surface wettability and surface topography [8].

Surface topography plays a very important role in bacterial adhesion, such as hydrophobicity, free surface energy, chemical composition and smoothness or roughness of surfaces. Microscopic studies of early dental plaque formations have shown adhesion of the initial colonized bacteria along the cracks and pits in the enamel, indicating the influence of surface structure on bacterial adhesion [9].

The success of esthetic restorations on a long-term basis depends on the quality and quantity of the attached biofilm. Initial attachment and subsequent colonization of bacteria on the surface of restorative materials is key to the pathogenesis of secondary caries promoted in particular by S. mutans and Streptococcus sobrinus (S. sobrinus). S. mutans has been identified as the major etiologic agent of human dental caries and forms a significant proportion of oral streptococci in carious lesions [10,11].

The various types of materials and their specific textures and physicochemical surface properties influence the quantity and quality of microbial colonization [12,13,14,15]. Therefore, knowledge of bacterial adhesion and subsequent biofilm formation (multicellular structures with microbial cells embedded in the extracellular matrix) is important for the study of bacterial pathogenicity. The human mouth provides a unique environment for the formation of complex biofilms; for example, it hosts more than 1,000 species of microorganisms [16]. Among several microorganisms, bacteria are a major group and are capable of forming biofilms. Thus, it is important to know the mechanisms involved in bacterial adhesion in various materials used.

Today, in dental prosthetics, different materials are used, which have different physicochemical properties and act differently on the bacterial adhesion of S. mutans. In nickel–chromium alloys, despite the good physical chemical properties of the material, biofilm formation on its surface is crucial. The same problem is present in composite resins used in dentistry; as in some complex resins, bacterial adhesion is stimulated by the released residual monomer [17].

This study investigates the correlation of surface characteristics (effect of contact angle [CA] and free surface energy) on three restorative materials (zirconia, nickel–chromium–molybdenum alloy and composites) used in dental prosthetics with bacterial adhesion (by means of the test specie S. mutans).

2 Materials and methods

For the purpose of studying the bacterial adhesion of S. mutans in various dental ceramics, three various starting materials were used: zirconium (Ceramill, Zolid FX White, Germany), metal alloy (Kera N, Eisenbacher Dentalwaren ED-GMBH, Germany) and composite (Sr ADORO – Ivoclar Vivadent, Liechtenstein). Ten specimens in cylindrical shape (dimensions: 8 mm in diameter and 2.5 mm in thickness) from all three types of starting are used during this research. The surfaces of the specimens were treated with standard polishing methods as specified by the material manufacturer.

To form biofilm formations for further analysis, we used S. mutans bacteria, reference strain ATCC 35668, which according to the manufacturer’s instructions was developed in laboratory conditions in Petri dishes with 5% blood agar incubation of the plates at 37°C in an incubator with microaerophilic conditions at 10% CO2 for 24 h. Bacteria obtained in culture are used to make a bacterial suspension at a concentration of 0.5 Mc Farland. The concentration of bacteria in the suspension is measured with the DensiCHEK Plus-bio Merieux apparatus. The samples will then be cleaned and sterilized in an autoclave for 30 min at 121°C. Each set of samples (3 sets of 10 samples) will be covered with the bacterial suspension for 15 min at 37°C; the samples were removed from the suspension and washed with physiological solution, then incubated in Petri dishes containing the blood agar culture at 37°C for 48 h under microaerophilic conditions with 10% CO2. Then, after 48 h, specimens are prepared for scanning electron microscopy (SEM).

The CA measurements were performed on the basis of an average of three consecutive measurements of each sample according to a standard procedure with distilled water droplets, ethylene glycol and glycerol. CA measurements were made using the See System E instrument with Software 7.0, Advex Instrument. Measurements were performed on three types of samples, and the results were obtained by averaging three consecutive measurements of each sample according to the standard distilled water drop procedure.

Surface-free energy (SFE) calculations are performed according to the Owens–Wendt Regression model. For this purpose, CA measurements were performed on the basis of an average of three consecutive measurements of each sample according to a standard procedure with distilled water droplets, ethylene glycol and glycerol.

To observe the morphology of bacterial adhesion, specimens are prepared for SEM. The samples were fixed in 2% glutaraldehyde for 24 h at room temperature, washed three times with buffer phosphate solution (pH 7.4) and dehydrated through a series of sorted ethanol solutions (20%, 40%, 60%, 80% and 100%). The samples are then dried, coated with gold spray and scanned using an electron microscope VEGA3 LMU coupled with energy-dispersive X-ray spectroscopy (INCA Energy 250 Microanalysis System). The accelerating voltage of the SE detector was set to 20 kV.

XRPD analysis was performed on Rigaku Ultima IV X-ray diffractometer equipped with D/teX high-speed 1-dimensional detector using CuKα radiation (λ = 1.54178 Å) in 2θ range from 5° to 60°. The accelerating voltage and the current power were set to 40 kV and 40 mA, respectively.

The chemical compositions of the starting materials as provided by the producer for all three starting materials are shown below (Table 1).

Table 1

Chemical composition of starting materials

MaterialsManufacturerComposition
ZirconiumCeramill* Zolid fx white, GermanyY2O3 (9.15–9.55%), HfO2 (<5%), Al2O3 (<0.5), others (0.1%)
Metal alloyKera N, Eisenbacher-Dentalwaren ED-GMBH, GermanyNi (61.4%), Cr (25.9%), Mo (11%), Si (1.5%), others (<0.1%)
CompositesSr ADORO, Ivoclare Vivadent, LiechtensteinUretan dimetakrilat (UDMA), (SiO2, 70%)

  1. Ethical approval: The conducted research is not related to either human or animal use.

3 Results and discussion

3.1 XRPD analysis of the starting materials

The XRPD analysis (Figure 1a) shows the presence of amorphous phase present in the sample, manifested with a wide bump in the region 15–30° 2θ. This bump is due to the amorphous silica [18,19]. Evident is the presence of crystalline phases: quartz in the following 2θ 26.58° (d 3.35 Å), 50.10° (d 1.81 Å) and 59.70° (d 1.54 Å) [20,21]; peaks at 30.10° (d 2.96 Å) and 34.82° (d 2.57 Å) due to the presence of cubic zirconia. Figure 1b shows the presence of two peaks at 43.86° (d 2.06 Å) and 50.86° (d 1.79 Å) as a result of the Ni–Cr–Mo alloy present in the sample. Figure 1c depicts crystalline behavior of the sample manifested by the appearance of peaks characteristic for cubic zirconia as well as quartz peaks. The peaks at 2θ 20.92° (d 4.24 Å) and 26.70° (d 3.34 Å) are due to the presence of quartz, while 30.20° (d 2.95 Å), 35.06° (d 2.55 Å), 50.26° (d 1.81 Å), 59.82° (d 1.54 Å) and 62.72° (d 1.47 Å) are as a result of cubic zirconia [20,21,22].

Figure 1 XRPD of a – composite (Q – quartz, Z – zirconia), b – alloy (A – Ni–Cr–Mo alloy) and c – zirconium (Q – quartz, Z – zirconia).
Figure 1

XRPD of a – composite (Q – quartz, Z – zirconia), b – alloy (A – Ni–Cr–Mo alloy) and c – zirconium (Q – quartz, Z – zirconia).

3.2 CAs and free surface energy

Table 2 shows the results of the CA for the three tested samples (zirconium, alloy and composite). Based on the CA measurements, the following is observed: the highest CA is observed with zirconium sample. The CA of the metal alloy is characterized by a CA lower by 4.35% than the zirconium sample, while the CA of the composite material is characterized by an even lower angle by 3.04%. This observed difference is not statistically significant. Therefore, based on the measurements made, the following can be concluded: based on the CA measurements, it can be concluded that the most hydrophobic characteristics are the samples with ordinal numbers: 1, 3 and 2.

Table 2

CA of zirconium, metal alloy and composite

SpecimenCA (°)
Zirconium90.34
Metal alloy (Ni–Cr–Mo alloy)86.41
Composite87.59

Table 3 shows the results of the SFE of three tested samples subject of this in vitro study. The data obtained from the SFE according to the OWR model show highly significant differences among the different materials. From the measurements of the SFE, it can be concluded that the highest values show the samples in ordinal numbers: 3, 2 and 1. These results, especially the CA for the composite, are most likely due to the amorphous matter present in the sample (Figure 1a).

Table 3

SFE of zirconium, metal alloy and composite

SpecimenSFE according to OWR model [mJ/m2]
Zirconium24.31
Metal alloy (Ni–Cr–Mo alloy)31.78
Composite48.82

3.3 SEM analysis

The SEM micrographs of the surfaces of zirconium, metal alloy and the composite are shown in Figure 2. Based on the provided results, it is observed that the surfaces of all three materials are remarkably different. The composite seems to be with the smoothest surface, while zirconia has a rigid surface topography.

Figure 2 SEM micrographs of a – zirconia composite, b – metal alloy, c – composite.
Figure 2

SEM micrographs of a – zirconia composite, b – metal alloy, c – composite.

While surface morphology plays a crucial role in the process of bacterial adhesions; in this study, the opposite is observed. Although the rigidness of the surface of zirconia is clearly observable compared to the other two, yet the average number of bacteria is observed to be significantly higher in the composite material. Table 4 shows the results obtained based on the number of bacteria developed in all three specimens. It is evident that the number of bacteria in metal alloys is around 3.8 times higher than zirconia, while this number is around 4.5 times higher in the composite material.

Table 4

Number of bacteria in different specimens (zirconium, metal alloy and composite)

SpecimenNumber of bacteria
Zirconium17
Metal alloy (Ni–Cr–Mo alloy)65
Composite80

4 Conclusions

This in vitro study evaluated the bacterial adhesion of S. mutans to zirconia, nickel–chromium–molybdenum alloys and composite materials. The obtained results show different values of CA, free surface energy and bacterial adhesion for all three materials. No correlation was found between CA and bacterial adhesion, whereas SFE correlated with bacterial adhesion, so that with increasing free surface energy, the bacterial adhesion of S. mutans increased. The analysis of the susceptibility of the selected dental materials to the adhesion of microorganisms as part of this study showed that the composite material is more susceptible to the adhesion of microorganisms. The hydrophobicity of the surfaces shows no association with bacterial adhesion in the test materials, whereas the free surface energy is correlated with the bacterial adhesion of S. mutans. Differences in the adhesion of microorganisms to the surface zirconia vs composite were statistically significant. Based on the above-mentioned results, it can be concluded that zirconia as dental ceramics has the lowest values for the presence of bacteria, followed by nickel–chromium–molybdenum alloy, while the highest values are found in the composites.

Acknowledgment

The authors thank the Ministry of Education and Science of North Macedonia and the Chemists Society of Turkey and for providing the financial support. The authors also thank Professor Blazo Boev and Mr. sc. Maja Lazarova for their support during the process of characterization of the samples with SEM, and Professor Metodija Najdovski for performing the XRPD analysis.

  1. Conflict of interest: The authors declare no conflict of interest.

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Received: 2020-04-08
Revised: 2020-08-20
Accepted: 2020-08-21
Published Online: 2020-11-04

© 2020 Fuat Bislimi et al., published by De Gruyter

This work is licensed under the Creative Commons Attribution 4.0 International License.

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  98. Adsorption and sugarcane-bagasse-derived activated carbon-based mitigation of 1-[2-(2-chloroethoxy)phenyl]sulfonyl-3-(4-methoxy-6-methyl-1,3,5-triazin-2-yl) urea-contaminated soils
  99. Antimicrobial and antifungal activities of bifunctional cooper(ii) complexes with non-steroidal anti-inflammatory drugs, flufenamic, mefenamic and tolfenamic acids and 1,10-phenanthroline
  100. Application of selenium and silicon to alleviate short-term drought stress in French marigold (Tagetes patula L.) as a model plant species
  101. Screening and analysis of xanthine oxidase inhibitors in jute leaves and their protective effects against hydrogen peroxide-induced oxidative stress in cells
  102. Synthesis and physicochemical studies of a series of mixed-ligand transition metal complexes and their molecular docking investigations against Coronavirus main protease
  103. A study of in vitro metabolism and cytotoxicity of mephedrone and methoxetamine in human and pig liver models using GC/MS and LC/MS analyses
  104. A new phenyl alkyl ester and a new combretin triterpene derivative from Combretum fragrans F. Hoffm (Combretaceae) and antiproliferative activity
  105. Erratum
  106. Erratum to: A one-step incubation ELISA kit for rapid determination of dibutyl phthalate in water, beverage and liquor
  107. Review Articles
  108. Sinoporphyrin sodium, a novel sensitizer for photodynamic and sonodynamic therapy
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  110. Plant description, phytochemical constituents and bioactivities of Syzygium genus: A review
  111. Evaluation of elastomeric heat shielding materials as insulators for solid propellant rocket motors: A short review
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  114. Study on online soft sensor method of total sugar content in chlorotetracycline fermentation tank
  115. Studies on the Anti-Gouty Arthritis and Anti-hyperuricemia Properties of Astilbin in Animal Models
  116. Effects of organic fertilizer on water use, photosynthetic characteristics, and fruit quality of pear jujube in northern Shaanxi
  117. Characteristics of the root exudate release system of typical plants in plateau lakeside wetland under phosphorus stress conditions
  118. Characterization of soil water by the means of hydrogen and oxygen isotope ratio at dry-wet season under different soil layers in the dry-hot valley of Jinsha River
  119. Composition and diurnal variation of floral scent emission in Rosa rugosa Thunb. and Tulipa gesneriana L.
  120. Preparation of a novel ginkgolide B niosomal composite drug
  121. The degradation, biodegradability and toxicity evaluation of sulfamethazine antibiotics by gamma radiation
  122. Special issue on Monitoring, Risk Assessment and Sustainable Management for the Exposure to Environmental Toxins
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  124. Source apportionment of soil contamination based on multivariate receptor and robust geostatistics in a typical rural–urban area, Wuhan city, middle China
  125. Special Issue on 13th JCC 2018
  126. The Role of H2C2O4 and Na2CO3 as Precipitating Agents on The Physichochemical Properties and Photocatalytic Activity of Bismuth Oxide
  127. Preparation of magnetite-silica–cetyltrimethylammonium for phenol removal based on adsolubilization
  128. Topical Issue on Agriculture
  129. Size-dependent growth kinetics of struvite crystals in wastewater with calcium ions
  130. The effect of silica-calcite sedimentary rock contained in the chicken broiler diet on the overall quality of chicken muscles
  131. Physicochemical properties of selected herbicidal products containing nicosulfuron as an active ingredient
  132. Lycopene in tomatoes and tomato products
  133. Fluorescence in the assessment of the share of a key component in the mixing of feed
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  135. Effectiveness of removal of sulphur compounds from the air after 3 years of biofiltration with a mixture of compost soil, peat, coconut fibre and oak bark
  136. Special Issue on the 4th Green Chemistry 2018
  137. Study and fire test of banana fibre reinforced composites with flame retardance properties
  138. Special Issue on the International conference CosCI 2018
  139. Disintegration, In vitro Dissolution, and Drug Release Kinetics Profiles of k-Carrageenan-based Nutraceutical Hard-shell Capsules Containing Salicylamide
  140. Synthesis of amorphous aluminosilicate from impure Indonesian kaolin
  141. Special Issue on the International Conf on Science, Applied Science, Teaching and Education 2019
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  143. The effect of nitrite food preservatives added to se’i meat on the expression of wild-type p53 protein
  144. Biocompatibility and osteoconductivity of scaffold porous composite collagen–hydroxyapatite based coral for bone regeneration
  145. Special Issue on the Joint Science Congress of Materials and Polymers (ISCMP 2019)
  146. Effect of natural boron mineral use on the essential oil ratio and components of Musk Sage (Salvia sclarea L.)
  147. A theoretical and experimental study of the adsorptive removal of hexavalent chromium ions using graphene oxide as an adsorbent
  148. A study on the bacterial adhesion of Streptococcus mutans in various dental ceramics: In vitro study
  149. Corrosion study of copper in aqueous sulfuric acid solution in the presence of (2E,5E)-2,5-dibenzylidenecyclopentanone and (2E,5E)-bis[(4-dimethylamino)benzylidene]cyclopentanone: Experimental and theoretical study
  150. Special Issue on Chemistry Today for Tomorrow 2019
  151. Diabetes mellitus type 2: Exploratory data analysis based on clinical reading
  152. Multivariate analysis for the classification of copper–lead and copper–zinc glasses
  153. Special Issue on Advances in Chemistry and Polymers
  154. The spatial and temporal distribution of cationic and anionic radicals in early embryo implantation
  155. Special Issue on 3rd IC3PE 2020
  156. Magnetic iron oxide/clay nanocomposites for adsorption and catalytic oxidation in water treatment applications
  157. Special Issue on IC3PE 2018/2019 Conference
  158. Exergy analysis of conventional and hydrothermal liquefaction–esterification processes of microalgae for biodiesel production
  159. Advancing biodiesel production from microalgae Spirulina sp. by a simultaneous extraction–transesterification process using palm oil as a co-solvent of methanol
  160. Topical Issue on Applications of Mathematics in Chemistry
  161. Omega and the related counting polynomials of some chemical structures
  162. M-polynomial and topological indices of zigzag edge coronoid fused by starphene
https://doi.org/10.1515/chem-2020-0070
Keywords for this article
Streptococcus mutans; bacterial adhesion; hydrophobicity; free surface energy; SEM
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  107. Review Articles
  108. Sinoporphyrin sodium, a novel sensitizer for photodynamic and sonodynamic therapy
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  118. Characterization of soil water by the means of hydrogen and oxygen isotope ratio at dry-wet season under different soil layers in the dry-hot valley of Jinsha River
  119. Composition and diurnal variation of floral scent emission in Rosa rugosa Thunb. and Tulipa gesneriana L.
  120. Preparation of a novel ginkgolide B niosomal composite drug
  121. The degradation, biodegradability and toxicity evaluation of sulfamethazine antibiotics by gamma radiation
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  123. Insight into the cadmium and zinc binding potential of humic acids derived from composts by EEM spectra combined with PARAFAC analysis
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  129. Size-dependent growth kinetics of struvite crystals in wastewater with calcium ions
  130. The effect of silica-calcite sedimentary rock contained in the chicken broiler diet on the overall quality of chicken muscles
  131. Physicochemical properties of selected herbicidal products containing nicosulfuron as an active ingredient
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  133. Fluorescence in the assessment of the share of a key component in the mixing of feed
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  136. Special Issue on the 4th Green Chemistry 2018
  137. Study and fire test of banana fibre reinforced composites with flame retardance properties
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  145. Special Issue on the Joint Science Congress of Materials and Polymers (ISCMP 2019)
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  147. A theoretical and experimental study of the adsorptive removal of hexavalent chromium ions using graphene oxide as an adsorbent
  148. A study on the bacterial adhesion of Streptococcus mutans in various dental ceramics: In vitro study
  149. Corrosion study of copper in aqueous sulfuric acid solution in the presence of (2E,5E)-2,5-dibenzylidenecyclopentanone and (2E,5E)-bis[(4-dimethylamino)benzylidene]cyclopentanone: Experimental and theoretical study
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  159. Advancing biodiesel production from microalgae Spirulina sp. by a simultaneous extraction–transesterification process using palm oil as a co-solvent of methanol
  160. Topical Issue on Applications of Mathematics in Chemistry
  161. Omega and the related counting polynomials of some chemical structures
  162. M-polynomial and topological indices of zigzag edge coronoid fused by starphene
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