Microbiota of complete removable dentures

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BACKGROUND

Edentulism, or complete tooth loss, is a prevalent condition. Demographic trends indicate that its incidence will continue to rise as life expectancy increases [1]. Tooth loss negatively impacts quality of life, leading to impaired speech, aged facial appearance, and, potentially, temporomandibular joint dysfunction, as well as dietary restrictions. Prosthetic rehabilitation is essential to restore masticatory function, diversify dietary options, and normalize speech. One widely used treatment option is the fabrication of complete removable acrylic dentures [2]. This type of prosthetic rehabilitation is one of the most common globally It effectively restores function, does not require high-end equipment, and is generally suitable for most edentulous patients [3]. However, treatment success depends not only on the quality of the prosthesis, but also on maintaining adequate denture hygiene.

Despite their advantages, complete removable acrylic dentures have limitations. These include the potential for allergic reactions and insufficient retention due to advanced bone and mucosal atrophy [4]. The denture base is made from acrylic resin, a porous material requiring meticulous daily cleaning and disinfection. Due to the low thermal conductivity of the acrylic base and the resulting “greenhouse effect,” the tissue surface of the denture creates favorable conditions for the proliferation of both pathogenic and opportunistic microorganisms [5], increasing the risk of inflammatory oral diseases. Denture stomatitis, frequently associated with microbial plaque on the denture surface, is among the most common conditions. Effective disinfection requires a thorough evaluation of oral hygiene and microbial contamination of dentures [6, 7]. The composition of the microbiota in the denture-bearing area depends on several factors: oral hygiene status, denture type and material, duration of denture use, the time interval between tooth extraction and prosthetic treatment, and the patient’s immune status [8]. Regardless of the type of removable dentures, their use leads to excessive microbial growth in the oral cavity and increases the risk of colonization by pathogenic species. As with natural dentition, a salivary glycoprotein-based acquired pellicle, containing salivary amylase, albumin, mucin, lysozyme, and immunoglobulins, forms on the denture surface once placed in the oral cavity [9]. The primary colonizers of dental enamel include gram-positive Streptococcus spp. (S. oralis, S. mutans, S. mitis, S. gordonii, S. sanguinis, and S. parasanguinis) as well as Veillonella spp., Neisseria spp., Rothia spp., Abiotrophia spp., Gemella spp., and Granulicatella spp. [10]. In 2019, Morse et al. investigated the cultivable flora on denture surfaces and reported predominant colonization by S. mutans, S. mitis, S. salivarius, and S. sanguis, along with gram-positive Actinomyces spp. (A. israelii, A. naeslundii, A. odontolyticus), Lactobacillus spp., and Veillonella spp. [11]. Understanding the qualitative and quantitative microbial composition is critical for developing effective denture disinfectants.

AIM: To assess the quantitative and qualitative composition of the microbiota on the surface of complete removable dentures after 1 and 4 years of use.

METHODS

Study Design

This was an observational, single-center, prospective, cross-sectional, controlled, blinded, randomized study.

Eligibility Criteria

Inclusion criteria: age 40 to 80 years, use of complete removable acrylic dentures for no longer than 5 years, and absence of allergic reactions.

Study Setting

The study was conducted at the Department of Propedeutics of Dental Diseases of the RUDN University, in the clinical and diagnostic center of the same institution (Moscow).

Study Duration

The study was carried out from October 2023 to July 2024.

Intervention

The quantitative and qualitative composition of the microbiota was determined by mass spectrometry of microbial markers. Swabs were collected from the inner surfaces of the maxillary and mandibular complete removable acrylic dentures without prior hygiene treatment. The samples were placed in labeled test tubes and transported in a cooled container to the laboratory. Upon arrival, the test tubes were sorted and checked for integrity. Prior to analysis, they were incubated in a thermostatic device to maintain optimal conditions for bacterial viability. The samples were then analyzed using the Maestro-αMS gas chromatograp–mass spectrometer (Interlab, Russia).

Main Study Outcome

Microbial contamination of all denture surfaces increased over time.

Additional Study Outcomes

In addition to microbial growth, denture hygiene deteriorated with prolonged use.

Subgroup Analysis

The study included 40 fully edentulous patients (К08.1) using acrylic complete removable dentures for no longer than 5 years. They were divided into 2 groups: Group 1 (n=20) used dentures for 1 year; Group 2 (n=20) used dentures for 4 years.

Outcomes Registration

Swabs were taken from denture surfaces, and the samples were analyzed using mass spectrometry to assess main and additional outcomes.

Statistical Analysis

Statistical analysis was performed using the Statistica 13 software. The data were normally distributed; therefore, descriptive statistics included calculation of the mean and standard deviation (mean ± SD). Intergroup differences were assessed using parametric tests, including the t-test with Bonferroni correction for multiple comparisons. A p value of 0.05 was considered significant.

RESULTS

Participants

Microbial contamination was assessed on the surfaces of complete removable acrylic dentures. In Group 1, biofilm was examined after 1 year of denture use; in Group 2, it was examined after 4 years.

Primary Results

The analysis of surface microbiota in patients who used removable dentures for 1 year and 4 years revealed the following (Table 1).

 

Table 1. Microbial concentration on denture surfaces (mean ± SD), ×10^⁵ CFU/g

Groups	Cocci and bacilli	Fungi and yeasts	Actinobacteria	Gram-negative rods	Anaerobes
Group 1 (1 year)	56±5	977±90	30±3	4±1	68±6
Group 2 (4 years)	107±8	1587±136	143±12	24±2	102±9
p-value	0.0	0.0003	0.0002	0.0005	0.0002

 

Both groups showed the presence of the following cocci and bacilli: Bacillus cereus, Bacillus megaterium, Enterococcus spp., Streptococcus spp., S. mutans (anaerobic), Staphylococcus aureus, and Staphylococcus epidermidis. These species were detected both in patients who used dentures for 1 year and those who used them for 4 years.

In Group 1, the concentration of cocci and bacilli was 56±5 (×10^⁵ CFU/g) (p=0.00). In Group 2, it increased to 107±8 (×10^⁵ CFU/g) (p=0.00) (Fig. 1).

 

Fig. 1. Concentration of cocci and bacilli on denture surfaces in Groups 1 and 2.

 

Fungal and yeast species (Aspergillus spp., Candida spp., campesterol) were identified in both groups. The concentration in Group 1 was 977±90 (×10^⁵ CFU/g) (p=0.0003). In Group 2, it increased to 1,587±136 (×10^⁵ CFU/g) (p=0.0003) (Fig. 2).

 

Fig. 2. Concentration of fungi and yeasts on denture surfaces in Groups 1 and 2.

 

The following actinobacteria were identified in both groups: Actinomyces spp., A. viscosus, Corynebacterium spp., Nocardia spp., N. asteroides, Mycobacterium spp., Pseudonocardia spp., Rhodococcus spp., Streptomyces spp., and S. farmamarensis. Their concentration in Group 1 was 30±3 (×10^⁵ CFU/g) (p=0.0002). In Group 2, it increased to 143±12 (×10^⁵ CFU/g) (p=0.0002) (Fig. 3).

 

Fig. 3. Concentration of actinobacteria on denture surfaces in Groups 1 and 2.

 

Gram-negative rods identified in both groups included the following: Alcaligenes spp./Klebsiella spp., Kingella spp., Flavobacterium spp., Moraxella spp./Acinetobacter spp., Porphyromonas spp., Pseudomonas aeruginosa, and Stenotrophomonas maltophilia. Their concentration was 4 ± 1 (×10^⁵ CFU/g) in Group 1 (p=0.0005) and 24±2 (×10^⁵ CFU/g) in Group 2 (p=0.0005) (Fig. 4).

 

Fig. 4. Concentration of gram-negative rods on denture surfaces in Groups 1 and 2.

 

Anaerobes found in both groups included: Bacteroides fragilis, Bifidobacterium spp., Blautia coccoides, Clostridium spp. (C. tetani group), C. difficile, C. histolyticum/Streptococcus pneumoniae, C. perfringens, C. propionicum, C. ramosum, Eubacterium spp., Eggerthella lenta, Fusobacterium spp./Haemophilus spp., Lactobacillus spp., Peptostreptococcus anaerobius, Prevotella spp., Propionibacterium spp. (P. acnes, P. freudenreichii, P. jensenii), Ruminococcus spp., and Veillonella spp. Their concentration in Group 1 was 68±6 (×10^⁵ CFU/g) (p=0.0002). In Group 2, it was 102±9 (×10^⁵ CFU/g) (p=0.0002) (Fig. 5).

 

Fig. 5. Concentration of anaerobes on denture surfaces in Groups 1 and 2.

 

Secondary Results

The observed increase in microbial counts over time suggests a deterioration in denture hygiene with prolonged use.

Adverse Events

No adverse events were reported.

DISCUSSION

Our findings demonstrate that fungi and yeasts dominate the microbial composition of denture surfaces. Long-term use (up to 4 years) leads to increased colonization not only by cocci and fungi but also by actinobacteria, gram-negative rods, and anaerobes. These findings are consistent with previous studies. For example, Vecherkina et al. reported a predominance of cocci and extensive Candida growth in cases of critically low pH (81% of cases) [12]. Bugorkov et al. identified Staphylococcus spp. and Candida spp. as the dominant pathogens [13]. An et al. concluded that Candida spp. play a key role in inflammatory oral diseases, with a significant increase in their counts after 5 years of denture use [14].

However, our data also indicate a significant rise in cocci, bacilli, actinobacteria, and anaerobes after 4 years of denture use.

CONCLUSION

The study findings indicate that microbial contamination of dentures correlates directly with duration of use. The longer a denture is worn, the more extensive the microbial colonization. Over a 4-year period, fungi and yeast counts increased by 91%, actinobacteria by 61%, gram-negative rods by 500%, and anaerobes by 50%.

ADDITIONAL INFORMATION

Funding sources: The authors declare no external funding for this study or its publication.

Disclosure of interests: The authors declare no explicit or potential conflicts of interests associated with the publication of this article.

Author contributions: All authors confirm that their authorship meets the international ICMJE criteria. The authors made the following contributions: S.N. Razumova: editing; A.S. Brago: selection of Russian-language sources; D.V. Serebrov: statistical analysis; K.D. Serebrov: writing; E.V. Adzhieva: selection of international sources; A.V. Rebriy: translation of international sources.

About the authors

Svetlana N. Razumova

Peoples’ Friendship University of Russia

Email: razumova-sn@rudn.ru
ORCID iD: 0000-0002-9533-9204
SPIN-code: 6771-8507

MD, Dr. Sci. (Medicine), Professor

Russian Federation, 6 Mikluho-Maklaja street, 117198 Moscow

Anzhela S. Brago

Peoples’ Friendship University of Russia

Email: brago-as@rudn.ru
ORCID iD: 0000-0001-8947-4357
SPIN-code: 2437-8867

MD, Cand. Sci. (Medicine)

Russian Federation, 6 Mikluho-Maklaja street, 117198 Moscow

Dmitrii V. Serebrov

Peoples’ Friendship University of Russia

Email: dserebrov@mail.ru
ORCID iD: 0000-0002-1030-1603
SPIN-code: 2161-9997

MD, Cand. Sci. (Medicine)

Russian Federation, 6 Mikluho-Maklaja street, 117198 Moscow

Elvira V. Adzhieva

Peoples’ Friendship University of Russia

Email: adzhieva-ev@rudn.ru
ORCID iD: 0000-0002-2735-4621
SPIN-code: 5667-4620
Russian Federation, 6 Mikluho-Maklaja street, 117198 Moscow

Astemir V. Rebriy

Peoples’ Friendship University of Russia

Email: rebriy-av@rudn.ru
ORCID iD: 0000-0002-5062-5979
Russian Federation, 6 Mikluho-Maklaja street, 117198 Moscow

Kirill D. Serebrov

Peoples’ Friendship University of Russia

Author for correspondence.
Email: k.serebrov@mail.ru
ORCID iD: 0000-0002-0353-1339
SPIN-code: 8649-7284
Russian Federation, 6 Mikluho-Maklaja street, 117198 Moscow

References

Supplementary files