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20 pages, 12695 KiB

Open AccessArticle

Necrotizing Gingivitis: Microbial Diversity and Quantification of Protein Secretion in Necrotizing Gingivitis

by Nicolas Gerhard, Thomas Thurnheer, Susanne Kreutzer, Rudolf Dominik Gmür, Thomas Attin, Giancarlo Russo and Lamprini Karygianni

Antibiotics 2021, 10(10), 1197; https://doi.org/10.3390/antibiotics10101197 - 1 Oct 2021

Cited by 1 | Viewed by 2957

Abstract

Necrotizing gingivitis (NG) is a necrotizing periodontal disease that differs from chronic gingivitis (CG). To date, both the microbiological causes and the involved host cytokine response of NG still remain unclear. Here, we investigated corresponding interdental plaque and serum samples from two groups [...] Read more.

Necrotizing gingivitis (NG) is a necrotizing periodontal disease that differs from chronic gingivitis (CG). To date, both the microbiological causes and the involved host cytokine response of NG still remain unclear. Here, we investigated corresponding interdental plaque and serum samples from two groups of Chinese patients with CG (n = 21) or NG (n = 21). The microbiota were studied by 16S rRNA Illumina MiSeq sequencing of the microbial metagenome and by assessing quantitatively the abundance of the phylum Bacteroidetes, the genus Prevotella and the species T. forsythia, P. endodontalis, and P. gingivalis using fluorescence in situ hybridization (FISH). With respect to the associated host response, the levels of 30 inflammatory mediators were quantified by multiplex immunoassay analysis. Differential microbial abundance analysis of the two disease groups revealed at the phylum level that Proteobacteria accounted for 67% of the differentially abundant organisms, followed by organisms of Firmicutes (21%) and Actinobacteria (9%). At the species level, significant differences in abundance were seen for 75 species of which 58 species were significantly more abundant in CG patients. Notably, the FISH analysis revealed that Bacteroidetes was the most prevalent phylum in NG. The multiplex cytokine assay showed significant quantitative differences between the disease groups for eight analytes (GM–CSF, G–CSF, IFN–α, IL–4, IL–13, TNF–α, MIG, and HGF). The G–CSF was found to be the most significantly increased inflammatory protein marker in NG. The next-generation sequencing (NGS) data supported the understanding of NG as a multi-microbial infection with distinct differences to CG in regard to the microbial composition. Full article

(This article belongs to the Special Issue Oral Microorganisms and Inactivation of Oral Biofilms)

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11 pages, 1546 KiB

Open AccessArticle

Antibacterial Effect of Sodium Hypochlorite and EDTA in Combination with High-Purity Nisin on an Endodontic-like Biofilm Model

by Ericka T. Pinheiro, Lamprini Karygianni, Thomas Attin and Thomas Thurnheer

Antibiotics 2021, 10(9), 1141; https://doi.org/10.3390/antibiotics10091141 - 21 Sep 2021

Cited by 11 | Viewed by 4092

Abstract

Antimicrobial peptides have been proposed as antibiofilm agents. Therefore, we evaluated the effect of endodontic irrigants combined or not with the antimicrobial peptide nisin against an endodontic biofilm model composed of eleven bacterial species. Biofilms were grown on hydroxyapatite discs for 3, 15 [...] Read more.

Antimicrobial peptides have been proposed as antibiofilm agents. Therefore, we evaluated the effect of endodontic irrigants combined or not with the antimicrobial peptide nisin against an endodontic biofilm model composed of eleven bacterial species. Biofilms were grown on hydroxyapatite discs for 3, 15 and 21 days and treated with 1.5% sodium hypochlorite (NaOCl) or 17% EDTA followed by high-purity nisin (nisin ZP) or saline for 5 min each. Differences between groups were tested by two-way ANOVA and Tukey’s multiple comparisons test (p < 0.05). Treatment with 1.5% NaOCl completely eliminated 3-d and 15-d biofilms but did not eradicate 21-d biofilms. Treatment with 1.5% NaOCl and 17% EDTA was equally effective against 21-d biofilms, showing 5-log and 4-log cell reduction, respectively, compared to the untreated control (9 log₁₀, p < 0.05). No significant difference was found between 1.5% NaOCl + nisin ZP and 1.5% NaOCl in 21-d biofilms (p > 0.05). Likewise, no significant difference was found between 17% EDTA + nisin ZP and 17% EDTA treatments (p > 0.05). In conclusion, 1.5% NaOCl or 17% EDTA were effective strategies to combat mature biofilms. The additional use of nisin did not improve the activity of conventional irrigants against multispecies biofilms. Full article

(This article belongs to the Special Issue Oral Microorganisms and Inactivation of Oral Biofilms)

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18 pages, 1689 KiB

Open AccessArticle

Subinhibitory Antibiotic Concentrations Enhance Biofilm Formation of Clinical Enterococcus faecalis Isolates

by Sara Bernardi, Annette Anderson, Guido Macchiarelli, Elmar Hellwig, Fabian Cieplik, Kirstin Vach and Ali Al-Ahmad

Antibiotics 2021, 10(7), 874; https://doi.org/10.3390/antibiotics10070874 - 19 Jul 2021

Cited by 44 | Viewed by 4774

Abstract

Enterococcus faecalis is a microorganism that can be found in the oral cavity, especially in secondary endodontic infections, with a prevalence ranging from 24–70%. The increase in the ability to form biofilms in the presence of subinhibitory antibiotic concentrations is a phenomenon that [...] Read more.

Enterococcus faecalis is a microorganism that can be found in the oral cavity, especially in secondary endodontic infections, with a prevalence ranging from 24–70%. The increase in the ability to form biofilms in the presence of subinhibitory antibiotic concentrations is a phenomenon that is observed for a wide variety of bacterial pathogens and is associated with increased resistance. In this study, therefore, six E. faecalis isolates from an endodontic environment and two control strains were exposed to subinhibitory concentrations of Penicillin G, Amoxicillin, Doxycycline, Fosfomycin, Tetracycline and Vancomycin and examined for their biofilm formation abilities. The minimum inhibitory concentration (MIC) was determined for all E. faecalis isolates. A culture of the isolate was mixed with a serial dilution series of the respective antibiotic, incubated overnight and the biofilm formation was analyzed using a microtiter plate assay. All isolates were able to form biofilms in the absence of an antibiotic. A significant increase in biofilm formation of up to more than 50% was found in the isolates exposed to subinhibitory concentrations of various antibiotics. Most isolates showed a significant increase in Fosfomycin (7/8), Doxycycline (6/8) and Tetracycline (6/8). Three endodontic isolates showed a significant increase in five of the antibiotics examined at the same time. On exposure to Vancomycin, three endodontic isolates and the two control strains showed an increase. The increase in the ability to form biofilms extended over a concentration range from 1/2 to 1/64 of the MIC concentration. Antibiotics may reach certain niches in the oral cavity at subinhibitory concentrations only. This can increase the biofilm formation by enterococci, and in turn lead to decreased susceptibility of these taxa to antibiotics. Full article

(This article belongs to the Special Issue Oral Microorganisms and Inactivation of Oral Biofilms)

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12 pages, 1902 KiB

Open AccessArticle

Antimicrobial and Antibiofilm Activity of the Probiotic Strain Streptococcus salivarius K12 against Oral Potential Pathogens

by Andrea Stašková, Miriam Sondorová, Radomíra Nemcová, Jana Kačírová and Marián Maďar

Antibiotics 2021, 10(7), 793; https://doi.org/10.3390/antibiotics10070793 - 29 Jun 2021

Cited by 13 | Viewed by 3777

Abstract

Oral probiotics are increasingly used in the harmonization of the oral microbiota in the prevention or therapy of various oral diseases. Investigation of the antimicrobial activity of the bacteriocinogenic strain Streptococcus salivarius K12 against oral pathogens shows promising results, not only in suppressing [...] Read more.

Oral probiotics are increasingly used in the harmonization of the oral microbiota in the prevention or therapy of various oral diseases. Investigation of the antimicrobial activity of the bacteriocinogenic strain Streptococcus salivarius K12 against oral pathogens shows promising results, not only in suppressing growth, but also in eliminating biofilm formation. Based on these findings, we decided to investigate the antimicrobial and antibiofilm activity of the neutralized cell-free supernatant (nCFS) of S. salivarius K12 at various concentrations against selected potential oral pathogens under in vitro conditions on polystyrene microtiter plates. The nCFS of S. salivarius K12 significantly reduced growth (p < 0.01) in Streptococcus mutans Clarke with increasing concentration from 15 to 60 mg/mL and also in Staphylococcus hominis 41/6 at a concentration of 60 mg/mL (p < 0.001). Biofilm formation significantly decreased (p < 0.001) in Schaalia odontolytica P10 at nCFS concentrations of 60 and 30 mg/mL. Biofilm inhibition (p < 0.001) was also observed in Enterobacter cloacae 4/2 at a concentration of 60 mg/mL. In Schaalia odontolytica P10 and Enterobacter cloacae 4/2, the nCFS had no effect on their growth. Full article

(This article belongs to the Special Issue Oral Microorganisms and Inactivation of Oral Biofilms)

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10 pages, 1613 KiB

Open AccessArticle

Efficacy of 0.05% Chlorhexidine and 0.05% Cetylpyridinium Chloride Mouthwash to Eliminate Living Bacteria on In Situ Collected Biofilms: An In Vitro Study

by Kathrin Becker, Giulia Brunello, Luisa Scotti, Dieter Drescher and Gordon John

Antibiotics 2021, 10(6), 730; https://doi.org/10.3390/antibiotics10060730 - 17 Jun 2021

Cited by 12 | Viewed by 4858

Abstract

Chlorhexidine (CHX) mouthwashes are frequently used as an adjunctive measure for the treatment of periodontitis and peri-implantitis, as well as in patients on maintenance therapy. However, their prolonged use is associated with several side effects. This study aimed at evaluating if a mouthwash [...] Read more.

Chlorhexidine (CHX) mouthwashes are frequently used as an adjunctive measure for the treatment of periodontitis and peri-implantitis, as well as in patients on maintenance therapy. However, their prolonged use is associated with several side effects. This study aimed at evaluating if a mouthwash with a reduced concentration of CHX combined with cetylpyridnium chloride (CPC) was as effective as a conventional CHX mouthwash in the reduction in living cells in oral biofilms attached to hydroxyapatite (HA) and micro-rough titanium (Ti) surfaces. Four healthy volunteers wore a customized acrylic appliance containing HA and Ti discs for in situ plaque accumulation. Biofilms were grown on the discs for 24 or 48 h and then randomly exposed for 60 s to: 0.05% CHX + 0.05% CPC, 0.1% CHX (positive control) or sterile saline (negative control). Viability assay and live-dead staining were performed to quantify bacterial viability and to distinguish live and dead cells, respectively. At both time points, contrary to saline, CHX, both alone and in combination with CPC, exhibited high antibacterial properties and induced a significant reduction in biofilm viability. This study demonstrates the potential of mouthwashes containing a low concentration of CHX combined with CPC as effective antibacterial agents for long-term applications with reduced undesired side effects. Full article

(This article belongs to the Special Issue Oral Microorganisms and Inactivation of Oral Biofilms)

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15 pages, 1332 KiB

Open AccessArticle

Trans-Cinnamaldehyde Attenuates Enterococcus faecalis Virulence and Inhibits Biofilm Formation

by Islam A. A. Ali, Jukka P. Matinlinna, Celine M. Lévesque and Prasanna Neelakantan

Antibiotics 2021, 10(6), 702; https://doi.org/10.3390/antibiotics10060702 - 11 Jun 2021

Cited by 30 | Viewed by 3687

Abstract

Enterococcus faecalis as an important nosocomial pathogen is critically implicated in the pathogenesis of endocarditis, urinary tract, and persistent root canal infections. Its major virulence attributes (biofilm formation, production of proteases, and hemolytic toxins) enable it to cause extensive host tissue damage. With [...] Read more.

Enterococcus faecalis as an important nosocomial pathogen is critically implicated in the pathogenesis of endocarditis, urinary tract, and persistent root canal infections. Its major virulence attributes (biofilm formation, production of proteases, and hemolytic toxins) enable it to cause extensive host tissue damage. With the alarming increase in enterococcal resistance to antibiotics, novel therapeutics are required to inhibit E. faecalis biofilm formation and virulence. Trans-cinnamaldehyde (TC), the main phytochemical in cinnamon essential oils, has demonstrated promising activity against a wide range of pathogens. Here, we comprehensively investigated the effect of TC on planktonic growth, biofilm formation, proteolytic and hemolytic activities, as well as gene regulation in E. faecalis. Our findings revealed that sub-inhibitory concentrations of TC reduced biofilm formation, biofilm exopolysaccharides, as well as its proteolytic and hemolytic activities. Mechanistic studies revealed significant downregulation of the quorum sensing fsr locus and downstream gelE, which are major virulence regulators in E. faecalis. Taken together, our study highlights the potential of TC to inhibit E. faecalis biofilm formation and its virulence. Full article

(This article belongs to the Special Issue Oral Microorganisms and Inactivation of Oral Biofilms)

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13 pages, 1418 KiB

Open AccessArticle

Long-Term Use of Oral Hygiene Products Containing Stannous and Fluoride Ions: Effect on Viable Salivary Bacteria

by Anne Brigitte Kruse, Nadine Schlueter, Viktoria Konstanze Kortmann, Cornelia Frese, Annette Anderson, Annette Wittmer, Elmar Hellwig, Kirstin Vach and Ali Al-Ahmad

Antibiotics 2021, 10(5), 481; https://doi.org/10.3390/antibiotics10050481 - 22 Apr 2021

Cited by 10 | Viewed by 3857

Abstract

The aim of this randomized, controlled clinical trial was to isolate and identify viable microorganisms in the saliva of study participants that continuously used a stannous and fluoride ion (F/Sn)-containing toothpaste and mouth rinse over a period of three years in comparison to [...] Read more.

The aim of this randomized, controlled clinical trial was to isolate and identify viable microorganisms in the saliva of study participants that continuously used a stannous and fluoride ion (F/Sn)-containing toothpaste and mouth rinse over a period of three years in comparison to a control group that used stannous ion free preparations (noF/Sn) over the same time period. Each group (F/Sn and noF/Sn) included 16 participants that used the respective oral hygiene products over a 36-month period. Stimulated saliva samples were collected at baseline (T0) and after 36 months (T1) from all participants for microbiological examination. The microbial composition of the samples was analyzed using culture technique, matrix-assisted laser desorption ionization–time of flight (MALDI–TOF) mass spectrometry, and 16S rDNA Polymerase Chain Reaction (PCR). There were only minor differences between both groups when comparing the absolute values of viable microbiota and bacterial composition. The treatment with F/Sn led to a slight decrease in disease-associated and a slight increase in health-associated bacteria. It was shown that the use of stannous ions had no negative effects on physiological oral microbiota even after prolonged use. In fact, a stabilizing effect of the oral hygiene products containing stannous ions on the health-associated oral microbiota could be expected. Full article

(This article belongs to the Special Issue Oral Microorganisms and Inactivation of Oral Biofilms)

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13 pages, 6284 KiB

Open AccessArticle

Single DNase or Proteinase Treatment Induces Change in Composition and Structural Integrity of Multispecies Oral Biofilms

by Lamprini Karygianni, Pune N. Paqué, Thomas Attin and Thomas Thurnheer

Antibiotics 2021, 10(4), 400; https://doi.org/10.3390/antibiotics10040400 - 7 Apr 2021

Cited by 6 | Viewed by 2690

Abstract

Biofilm virulence is mainly based on its bacterial cell surrounding biofilm matrix, which contains a scaffold of exopolysaccharides, carbohydrates, proteins, lipids, and nucleic acids. Targeting these nucleid acids or proteins could enable an efficient biofilm control. Therefore, the study aimed to test the [...] Read more.

Biofilm virulence is mainly based on its bacterial cell surrounding biofilm matrix, which contains a scaffold of exopolysaccharides, carbohydrates, proteins, lipids, and nucleic acids. Targeting these nucleid acids or proteins could enable an efficient biofilm control. Therefore, the study aimed to test the effect of deoxyribonuclease I (DNase I) and proteinase K on oral biofilms. Six-species biofilms (Streptococcus mutans, Streptococcus oralis, Actinomyces oris, Fusobacterium nucleatum, Veillonella dispar, and Candida albicans) were exposed to DNase I (0.001 mg/mL, 0.002 mg/mL) or proteinase K (0.05 mg/mL, 0.1 mg/mL) for 1 h during biofilm formation. After 64 h, biofilms were harvested, quantified by culture analysis and visualized by image analysis using CLSM (confocal laser scanning microscopy). Statistical analysis was performed by ANOVA, followed by the Tukey test at a 5% significance level. The biofilm treatment with proteinase K induced a significant increase of Logs₁₀ counts in S. mutans and a decrease in C. albicans, while biofilm thickness was reduced from 28.5 μm (control) to 9.07 μm (0.05 mg/mL) and 7.4 μm (0.1 mg/mL). Treatment with DNase I had no effect on the total bacterial growth within the biofilm. Targeting proteins of biofilms by proteinase K are promising adjunctive tool for biofilm control. Full article

(This article belongs to the Special Issue Oral Microorganisms and Inactivation of Oral Biofilms)

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12 pages, 10739 KiB

Open AccessArticle

Adjuvant Transgingival Therapy with Visible Light Plus Water-Filtered Infrared-A (VIS + wIRA) in Periodontal Therapy—A Randomized, Controlled, Stratified, Double-Blinded Clinical Trial

by Stefanie Anna Peikert, Anil Fischer, Anne Brigitte Kruse, Ali Al-Ahmad, Johan Peter Woelber, Kirstin Vach, Andreas Braun and Petra Ratka-Krüger

Antibiotics 2021, 10(3), 251; https://doi.org/10.3390/antibiotics10030251 - 3 Mar 2021

Cited by 2 | Viewed by 2261

Abstract

The aim of this randomized, controlled, double-blinded clinical trial was to examine the additional healing effect of transgingival visible light and water-filtered infrared-A (VIS + wIRA) in the treatment of periodontitis patients compared with the standard therapy by subgingival instrumentation (SI). Therefore, forty [...] Read more.

The aim of this randomized, controlled, double-blinded clinical trial was to examine the additional healing effect of transgingival visible light and water-filtered infrared-A (VIS + wIRA) in the treatment of periodontitis patients compared with the standard therapy by subgingival instrumentation (SI). Therefore, forty patients with untreated periodontitis received a non-surgical periodontal treatment. Using a split-mouth study design, one quadrant of the upper jaw was randomly either exposed to VIS + wIRA four times for 20 min within two weeks in addition to SI or received only SI. Three and 6 months after intervention, clinical parameters (probing depths (PDs), clinical attachment level, bleeding on probing (BOP), furcation, tooth mobility, plaque control record, and papilla bleeding index) were re-evaluated. In the presence of PD of 4 mm and positive BOP or PD > 4 mm, SI was performed again. Moreover, the patients were asked about their discomfort using a visual analog scale from 1 to 10 for each side of the maxilla. Statistical analysis demonstrated no differences between quadrants at re-evaluation for clinical parameters (p > 0.05) after 3 and 6 months. Concerning pain perception, patients described less pain on the irradiated side (p = 0.016). In the treatment of patients with periodontitis, VIS + wIRA did not show an additional effect on the clinical outcome after 3 and 6 months. Patients described less pain on the irradiated quadrant after treatment. Full article

(This article belongs to the Special Issue Oral Microorganisms and Inactivation of Oral Biofilms)

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9 pages, 6088 KiB

Open AccessArticle

Antibacterial Effect of High-Purity Nisin Alone and in Combination with D-Amino Acids or Chlorhexidine in an Endodontic-Like Biofilm Model

by Ericka T. Pinheiro, Lamprini Karygianni, Thomas Attin and Thomas Thurnheer

Antibiotics 2021, 10(2), 149; https://doi.org/10.3390/antibiotics10020149 - 2 Feb 2021

Cited by 4 | Viewed by 2223

Abstract

New strategies to eradicate endodontic biofilms are needed. Therefore, we evaluated the effect of high-purity nisin alone and in combination with D-amino acids (D-AAs) or chlorhexidine (CHX) against an “endodontic-like” biofilm model. Biofilms were grown on hydroxyapatite discs for 64 h and treated [...] Read more.

New strategies to eradicate endodontic biofilms are needed. Therefore, we evaluated the effect of high-purity nisin alone and in combination with D-amino acids (D-AAs) or chlorhexidine (CHX) against an “endodontic-like” biofilm model. Biofilms were grown on hydroxyapatite discs for 64 h and treated with nisin, eight D-AAs mixture, nisin + eight D-AAs, 2% CHX, and nisin + 2% CHX. After the 5 min and 24 h treatments, biofilm cells were harvested and total colony-forming units were counted. Differences between groups were tested by two-way ANOVA followed by Tukey’s multiple comparisons test (p < 0.05). Nisin and D-AAs, alone or in combination, were not effective in reducing bacteria after short or long exposure times. After 5 min, treatment with 2% CHX and nisin + 2% CHX resulted in 2 and 2.4-log cell reduction, respectively, compared with the no treatment control (p < 0.001). After 24 h, 2% CHX and nisin + 2% CHX drastically reduced bacterial counts. In conclusion, high-purity nisin alone or in combination with D-AAs did not show antibacterial activity against multispecies biofilms. Moreover, combined treatment using nisin and CHX showed similar antibiofilm activity compared with the use of CHX alone. Full article

(This article belongs to the Special Issue Oral Microorganisms and Inactivation of Oral Biofilms)

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19 pages, 2179 KiB

Open AccessArticle

Staphylococcus aureus Interferes with Streptococci Spatial Distribution and with Protein Expression of Species within a Polymicrobial Oral Biofilm

by Etyene Schnurr, Pune N. Paqué, Thomas Attin, Paolo Nanni, Jonas Grossmann, Silva Holtfreter, Barbara M. Bröker, Christian Kohler, Binh An Diep, Apoena de Aguiar Ribeiro and Thomas Thurnheer

Antibiotics 2021, 10(2), 116; https://doi.org/10.3390/antibiotics10020116 - 26 Jan 2021

Cited by 9 | Viewed by 3346

Abstract

We asked whether transient Staphylococcus aureus in the oral environment synergistically interacts with orally associated bacterial species such as Actinomyces oris, Candida albicans, Fusobacterium nucleatum, Streptococcus oralis, Streptococcus mutans, and Veillonella dispar (six-species control biofilm 6S). For this [...] Read more.

We asked whether transient Staphylococcus aureus in the oral environment synergistically interacts with orally associated bacterial species such as Actinomyces oris, Candida albicans, Fusobacterium nucleatum, Streptococcus oralis, Streptococcus mutans, and Veillonella dispar (six-species control biofilm 6S). For this purpose, four modified biofilms with seven species that contain either the wild type strain of the S. aureus genotype (USA300-MRSA WT), its isogenic mutant with MSCRAMM deficiency (USA300-MRSA ΔMSCRAMM), a methicillin-sensitive S. aureus (ST72-MSSA-) or a methicillin-resistant S. aureus (USA800-MRSA) grown on hydroxyapatite disks were examined. Culture analyses, confocal-laser-scanning microscopy and proteome analyses were performed. S. aureus strains affected the amount of supragingival biofilm-associated species differently. The deletion of MSCRAMM genes disrupted the growth of S. aureus and the distribution of S. mutans and S. oralis within the biofilms. In addition, S. aureus caused shifts in the number of detectable proteins of other species in the 6S biofilm. S. aureus (USA300-MRSA WT), aggregated together with early colonizers such as Actinomyces and streptococci, influenced the number of secondary colonizers such as Fusobacterium nucleatum and was involved in structuring the biofilm architecture that triggered the change from a homeostatic biofilm to a dysbiotic biofilm to the development of oral diseases. Full article

(This article belongs to the Special Issue Oral Microorganisms and Inactivation of Oral Biofilms)

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11 pages, 699 KiB

Open AccessArticle

Initial Bacterial Adhesion and Biofilm Formation on Aligner Materials

by Sibel Tektas, Thomas Thurnheer, Theodore Eliades, Thomas Attin and Lamprini Karygianni

Antibiotics 2020, 9(12), 908; https://doi.org/10.3390/antibiotics9120908 - 15 Dec 2020

Cited by 16 | Viewed by 3762

Abstract

The present study aims to assess the initial bacterial adhesion and biofilm formation on different aligner materials. A total of four different aligner materials, CA-medium (CAM), copolyester (COP), Duran (DUR), Erkodur (ERK), were tested. Stimulated human saliva was obtained from six healthy volunteers. [...] Read more.

The present study aims to assess the initial bacterial adhesion and biofilm formation on different aligner materials. A total of four different aligner materials, CA-medium (CAM), copolyester (COP), Duran (DUR), Erkodur (ERK), were tested. Stimulated human saliva was obtained from six healthy volunteers. Salivary bacteria were harvested by centrifugation, and 1 mL of the salivary suspension was injected onto each sample surface for 2 h and 3 days, respectively. The samples were then washed twice with 5 mL 0.9% NaCl solution, and non-adherent bacteria were removed. The adherent microorganisms were dislodged from the sample surfaces after ultrasonication for 4 min in 1 mL 0.9% NaCl on ice. After the incubation of the adherent salivary bacteria under both aerobic and anaerobic conditions on Columbia blood agar plates at 37 °C and 5% CO₂ and in anaerobic jars overnight, several dilutions thereof were used for the determination of CFUs. This protocol was applied three times, obtaining an average of nine independent measurements for each material group. Overall, the differences between the tested aligner materials as well as between the materials and controls were not of statistical significance (p > 0.05). Regarding initial bacterial attachment and biofilm formation, the tested aligner materials are comparable to enamel and metal orthodontic brackets and can be therefore considered for clinical use. The four tested aligner materials CAM, COP, DUR, ERK showed no significant differences in initial microbial attachment and biofilm formation of aerobic and anaerobic species compared to enamel and conventional brackets. Full article

(This article belongs to the Special Issue Oral Microorganisms and Inactivation of Oral Biofilms)

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16 pages, 2425 KiB

Open AccessArticle

Influence of Probiotics on the Salivary Microflora Oral Streptococci and Their Integration into Oral Biofilm

by Nicole B. Arweiler, Thorsten M. Auschill, Christian Heumann, Elmar Hellwig and Ali Al-Ahmad

Antibiotics 2020, 9(11), 803; https://doi.org/10.3390/antibiotics9110803 - 13 Nov 2020

Cited by 8 | Viewed by 3343

Abstract

Probiotics’ ability to integrate into dental biofilms is not yet clarified. The aim of this trial was to detect probiotic bacteria from probiotic products in dental biofilm and saliva during and after intake. In this parallel, randomized clinical trial, 39 subjects wore customized [...] Read more.

Probiotics’ ability to integrate into dental biofilms is not yet clarified. The aim of this trial was to detect probiotic bacteria from probiotic products in dental biofilm and saliva during and after intake. In this parallel, randomized clinical trial, 39 subjects wore customized appliances to build up intra-oral biofilms (72-h periods). The trial was divided into screening (S) to determine baseline biofilm flora, intervention (I), and wash out (WO). During I (28 days), subjects consumed a product containing (a) Enterococcus faecalis (b) Lactobacilluscasei, or (c) Lactobacillus rhamnosus GG. Probiotic bacteria and Streptococci spp. were detected in the biofilms and saliva of the 35 subjects that were included in the analysis. During I and WO, the ratio of probiotics in the biofilm was very low compared to total bacterial load, while saliva had slightly but not significantly higher values. No significant changes of probiotic bacteria (p > 0.05) were found at any visit during I or WO. The proportion of streptococci was significantly reduced (p < 0.05) during I and even lower in WO, compared to S. Probiotic bacteria could neither integrate nor persist in dental biofilm and saliva but did influence the growth of streptococci in biofilm and saliva. Full article

(This article belongs to the Special Issue Oral Microorganisms and Inactivation of Oral Biofilms)

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12 pages, 1627 KiB

Open AccessArticle

Comparison of Three Endodontic Irrigant Regimens against Dual-Species Interkingdom Biofilms: Considerations for Maintaining the Status Quo

by Om Alkhir Alshanta, Saeed Alqahtani, Suror Shaban, Khawlah Albashaireh, William McLean and Gordon Ramage

Antibiotics 2020, 9(9), 634; https://doi.org/10.3390/antibiotics9090634 - 22 Sep 2020

Cited by 8 | Viewed by 3961

Abstract

Endodontic infections are often interkingdom biofilms, though current clinical management rarely considers this phenomenon. This study aimed to evaluate new and standard endodontic antimicrobial regimens against simple and complex Candida albicans and Enterococcus faecalis mono- and dual-species biofilms. C. albicans and E. faecalis [...] Read more.

Endodontic infections are often interkingdom biofilms, though current clinical management rarely considers this phenomenon. This study aimed to evaluate new and standard endodontic antimicrobial regimens against simple and complex Candida albicans and Enterococcus faecalis mono- and dual-species biofilms. C. albicans and E. faecalis mono- and dual-species biofilms were grown upon Thermanox™ coverslips and treated for 5 min with 3% NaOCl, 3% NaOCl followed by 17% EDTA, or 9% HEDP dissolved in 3% NaOCl. The number of cells remaining immediately after treatment at 0 h and after 72 h of regrowth were assessed using real-time quantitative PCR. All three treatment arms showed a similar positive antimicrobial effect on C. albicans and E. faecalis in both mono- and dual-species biofilms following initial treatment, resulting in ≥98% reduction in colony forming equivalent (CFE). Regardless of species or biofilm type (mono- or dual- species), the antimicrobial effect of NaOCl:HEDP mixture was comparable to that of NaOCl alone, with both showing significant regrowth after 72 h, whereas sequential treatment with NaOCl and EDTA consistently prevented significant regrowth. Our data suggest that sequential irrigation with NaOCl and EDTA remains the antimicrobial strategy of choice as it significantly reduces biofilm persistence and regrowth in our experimental dual-species biofilm conditions. Full article

(This article belongs to the Special Issue Oral Microorganisms and Inactivation of Oral Biofilms)

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13 pages, 2068 KiB

Open AccessArticle

Improving the Efficiency of Photodynamic Chemotherapy in Root Canals against Enterococcus faecalis In Vitro

by Christian Tennert, Yoana Zinovieva, Kalin Shishkov, Lamprini Karygianni, Makus Jörg Altenburger, Richard J Wierichs and Ali Al-Ahmad

Antibiotics 2020, 9(9), 543; https://doi.org/10.3390/antibiotics9090543 - 26 Aug 2020

Cited by 6 | Viewed by 2988

Abstract

The aim of this study was to evaluate the effect of photoactivated chemotherapy (PACT) on Enterococcus faecalis (E. faecalis) biofilms in root canals using an 90% isopropanol (IPA)-based photosensitizer and removing excess photosensitizer before light incubation. Three hundred and seven extracted [...] Read more.

The aim of this study was to evaluate the effect of photoactivated chemotherapy (PACT) on Enterococcus faecalis (E. faecalis) biofilms in root canals using an 90% isopropanol (IPA)-based photosensitizer and removing excess photosensitizer before light incubation. Three hundred and seven extracted human teeth with one root canal were infected with E. faecalis for 72 h and treated in groups: IPA irrigation; PACT; PACT and final rinse with IPA; PACT with photosensitizer removal using either 0.9% NaCl solution or sterile paper points or both; PACT using IPA-based photosensitizer with and without a final rinse of IPA. Root canals were sampled using sterile paper points and dentin chips collected from the root canal walls. Additionally, SEM (Scanning Electron Microscopy) images of the specimens were taken to evaluate the root canal walls for residue bacterial contamination. In all antimicrobial treatment groups treatments E. faecalis counts were significantly reduced in the root canals. Using IPA-based photosensitizer the antimicrobial effect of PACT was significantly enhanced. Irrigation with IPA alone or after PACT significantly increased the antimicrobial effect compared to PACT alone. The collected dentin chips revealed the highest amount of culture negative root canals (10%) after PACT using IPA-based photosensitizer. In the other groups, the culture negative samples ranged from only 0–2 specimens of 30 specimens. REM images show remaining E. faecalis cells on the root canal wall and inside dentin tubules. Using IPA-based photosensitizer significantly enhanced the antimicrobial effect of PACT against E. faecalis in the root canals. Full article

(This article belongs to the Special Issue Oral Microorganisms and Inactivation of Oral Biofilms)

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19 pages, 1957 KiB

Open AccessFeature PaperArticle

Microbial Composition of Oral Biofilms after Visible Light and Water-Filtered Infrared a Radiation (VIS+wIRA) in Combination with Indocyanine Green (ICG) as Photosensitizer

by Thomas Burchard, Lamprini Karygianni, Elmar Hellwig, Annette Wittmer and Ali Al-Ahmad

Antibiotics 2020, 9(9), 532; https://doi.org/10.3390/antibiotics9090532 - 23 Aug 2020

Cited by 8 | Viewed by 3342

Abstract

In view of increasing antibiotic resistance, antimicrobial photodynamic therapy (aPDT) is an alternative treatment method used to eradicate the microbial community of oral biofilms that can be responsible for different oral infections. In order to investigate changes in the microbial composition after application [...] Read more.

In view of increasing antibiotic resistance, antimicrobial photodynamic therapy (aPDT) is an alternative treatment method used to eradicate the microbial community of oral biofilms that can be responsible for different oral infections. In order to investigate changes in the microbial composition after application of aPDT with visible light and water-filtered infrared A (VIS+wIRA) in combination with indocyanine green (ICG), oral microorganisms of the initial and mature biofilm were evaluated by mass spectrometry (MALDI-TOF-MS). To determine surviving microorganisms using MALDI-TOF-MS, an in situ biofilm was irradiated with VIS+wIRA for five minutes in the presence of ICG (300 and 450 µg/mL, respectively). Treatment with chlorhexidine (0.2%) served as positive control. Identified microorganisms of the initial biofilm treated with ICG showed a clear reduction in diversity. The microbial composition of the mature oral biofilm also showed changes after the implementation of aPDT, which mainly resulted in a shift in the percentage of bacterial species. The resulting destruction of the microbial balance within the oral biofilm by aPDT using VIS+wIRA and ICG can be seen as an advantageous supplementary approach in the adjunctive treatment of periodontitis and peri-implantitis. Full article

(This article belongs to the Special Issue Oral Microorganisms and Inactivation of Oral Biofilms)

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20 pages, 650 KiB

Open AccessArticle

Analysing the Relationship between Nutrition and the Microbial Composition of the Oral Biofilm—Insights from the Analysis of Individual Variability

by Kirstin Vach, Ali Al-Ahmad, Annette Anderson, Johan Peter Woelber, Lamprini Karygianni, Annette Wittmer and Elmar Hellwig

Antibiotics 2020, 9(8), 479; https://doi.org/10.3390/antibiotics9080479 - 4 Aug 2020

Cited by 6 | Viewed by 3032

Abstract

The influence of a change in nutrition on the oral microbiota are discussed in literature, but usually only changes of population mean values are reported. This paper introduces simple methods to also analyse and report the variability of patients’ reactions considering data from [...] Read more.

The influence of a change in nutrition on the oral microbiota are discussed in literature, but usually only changes of population mean values are reported. This paper introduces simple methods to also analyse and report the variability of patients’ reactions considering data from the culture analysis of oral biofilm. The framework was illustrated by an experimental study exposing eleven participants to different nutrition schemes in five consecutive phases. Substantial inter-individual variations in the individual reactions were observed. A new coherence index made it possible to identify 14 instances where the direction of individual changes tended to coincide with the direction of the mean change with more than 95% probability. The heterogeneity in variability across different bacteria species was limited. This allowed us to develop recommendations for sample sizes in future studies. For studies measuring the concentration change of bacteria as a reaction to nutrition change, the use of replications and analysis of the variability is recommended. In order to detect moderate effects of a change in nutrition on the concentration of single bacterial taxa, 30 participants with three repetitions are often adequate. Insights into the relationship between nutrition and the microbial composition can be helpful for the development of dietary habits that promote the establishment of a healthy microbial flora and can therefore prevent the initiation of oral diseases such as caries and periodontitis. Full article

(This article belongs to the Special Issue Oral Microorganisms and Inactivation of Oral Biofilms)

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Review

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13 pages, 775 KiB

Open AccessReview

A Qualitative and Comprehensive Analysis of Caries Susceptibility for Dental Fluorosis Patients

by Qianrui Li, Jiaqi Shen, Tao Qin, Ge Zhou, Yifeng Li, Zhu Chen and Mingyun Li

Antibiotics 2021, 10(9), 1047; https://doi.org/10.3390/antibiotics10091047 - 27 Aug 2021

Cited by 7 | Viewed by 3453

Abstract

Dental fluorosis (DF) is an endemic disease caused by excessive fluoride exposure during childhood. Previous studies mainly focused on the acid resistance of fluorotic enamel and failed to reach a consensus on the topic of the caries susceptibility of DF patients. In this [...] Read more.

Dental fluorosis (DF) is an endemic disease caused by excessive fluoride exposure during childhood. Previous studies mainly focused on the acid resistance of fluorotic enamel and failed to reach a consensus on the topic of the caries susceptibility of DF patients. In this review, we discuss the role of DF classification in assessing this susceptibility and follow the “four factors theory” in weighing the pros and cons of DF classification in terms of host factor (dental enamel and saliva), food factor, bacteria factor, and DF treatment factor. From our analysis, we find that susceptibility is possibly determined by various factors such as the extent of structural and chemical changes in fluorotic enamel, eating habits, fluoride levels in diets and in the oral cavity, changes in quantity and quality of saliva, and/or oral hygiene. Thus, a universal conclusion regarding caries susceptibility might not exist, instead depending on each individual’s situation. Full article

(This article belongs to the Special Issue Oral Microorganisms and Inactivation of Oral Biofilms)

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12 pages, 282 KiB

Open AccessReview

Clindamycin as an Alternative Option in Optimizing Periodontal Therapy

by Ionut Luchian, Ancuta Goriuc, Maria Alexandra Martu and Mihai Covasa

Antibiotics 2021, 10(7), 814; https://doi.org/10.3390/antibiotics10070814 - 4 Jul 2021

Cited by 37 | Viewed by 6220

Abstract

Periodontal disease is an oral infectious and inflammatory disease caused by microorganisms that determine the host-mediated destruction of soft and hard periodontal tissues, which ultimately leads to tooth loss. Periodontitis affects a large part of the population, with various degrees of severity. Treatment [...] Read more.

Periodontal disease is an oral infectious and inflammatory disease caused by microorganisms that determine the host-mediated destruction of soft and hard periodontal tissues, which ultimately leads to tooth loss. Periodontitis affects a large part of the population, with various degrees of severity. Treatment consists of etiologic therapy: the removal of biofilm through mechanical debridement plus microbial elimination by supplementary measures. Antibiotic administration, either systemically or through local delivery, has been shown to improve clinical outcomes after mechanical periodontal treatment. Clindamycin is a lincosamide with a broad spectrum, being active against aerobic, anaerobic, and β-lactamase-producing bacteria. This antibiotic offers several advantages and some disadvantages and has been used in periodontal treatment both systemically and locally with various degrees of success. Among the properties that recommend it for periodontal treatment is the bacteriostatic effect, the inhibition of bacterial proteins synthesis, the enhancement of neutrophil chemotaxis, phagocytosis and the oxidative burst–oxidative stress storm. Furthermore, it is easily absorbed at the level of oral tissues in a considerable amount. This substantial tissue penetration, especially inside the bone, is synergistic with a stimulating effect on the host immune system. The aim of this review is to explore the applicability of this antibiotic agent and to evaluate its antimicrobial potential and limitations at the level of the oral biofilm associated with periodontal disease. Full article

(This article belongs to the Special Issue Oral Microorganisms and Inactivation of Oral Biofilms)

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