Assessment of the Impact of the Probiotic Limosilactobacillus Reuteri Prodentis (DSM 17938 and ATCC PTA 5289) on Clinical and Microbiological Parameters in Orthodontically Treated Patients With Concomitant Gingivitis
Phase: NA
Conditions: Gingivitis, Orthodontic Treatment
Sponsor: Medical University of Warsaw

2.01.2022 RESEARCH PROTOCOL 
 
 Title of the Study:  
Assessment of the Impact of the Probiotic Limosilactobacillus Reuteri Prodentis (DSM 
17938 and ATCC PTA 5289) on Clinical and Microbiological Parameters in Orthodontically 
Treated Patients with Concomitant Gingivitis 
 
 1. Current State of Knowledge Insufficient oral hygiene promotes the development of 
plaque-induced gingivitis. According to current knowledge, the development of gingivitis is 
associated with nonspecific bacterial plaque, and in the early stage of inflammation a 
disturbance in the balance between the physiological and pathogenic microbiota of the oral 
cavity plays a significant role. Untreated gingivitis may lead to the development of 
periodontitis, a chronic inflammatory disease that is one of the major risk factors for tooth 
loss and is associated with numerous systemic conditions, such as coronary heart disease, 
myocardial infarction, ischemic stroke, and diabetes mellitus. Biofilm-induced gingivitis 
represents a significant public health problem, affecting diverse patient populations. 
Moreover, its presence may lead to complications in conservative, orthodontic, or prosthetic 
dental treatment. Treatment with fixed orthodontic appliances promotes dental plaque 
accumulation and hampers effective oral hygiene procedures, thereby increasing the risk of 
plaque-induced gingivitis. Gingival inflammation during fixed orthodontic treatment may 
result in marginal gingival enlargement, loss of connective tissue attachment, and alveolar 
bone resorption, which may ultimately compromise orthodontic treatment outcomes. 
Professional periodontal therapy combined with strict adherence to meticulous oral hygiene 
procedures ensures a reduction in dental plaque and a significant decrease in pathogenic 
microorganisms responsible for the development of gingival and periodontal diseases. This is 
particularly important in orthodontic patients. In addition to professional plaque and calculus 
removal and rigorous hygiene instructions, shortterm use of chlorhexidine mouthrinses is 
commonly recommended as adjunctive therapy in the treatment of gingivitis. An alternative 
to antiseptic mouthrinses may be probiotic preparations for local application, which exhibit 
anti-inflammatory properties and modulate the oral microbiome. One such preparation is 
GUM PerioBalance, a commercially available natural dietary supplement. Each lozenge 
contains at least 200 million viable probiotic bacteria Lactobacillus reuteri Prodentis. 
According to the manufacturer’s information leaflet, the probiotic strains contained in the 
product enhance natural defense mechanisms, exert anti-inflammatory effects, and inhibit the 
growth of pathogenic bacteria responsible for gingival diseases. The tablets are gluten-free, 
suitable for vegetarians, do not cause tooth discoloration, and have a fresh mint flavor. The 
planned study aims to determine whether the use of probiotic preparations may represent a 
valuable adjunct to standard gingivitis therapy and whether probiotic supplementation during 
orthodontic treatment may be beneficial for maintaining gingival health. 
 

2.01.2022 RESEARCH PROTOCOL 
 
 Title of the Study:  
Assessment of the Impact of the Probiotic Limosilactobacillus Reuteri Prodentis (DSM 
17938 and ATCC PTA 5289) on Clinical and Microbiological Parameters in Orthodontically 
Treated Patients with Concomitant Gingivitis 
 
 
 References Laleman I et al. (2015). Probiotics in the dental practice: a review. Quintessence 
International, 46, 255–264. Teughels W et al. (2008). Probiotics and oral healthcare. 
Periodontology 2000, 48, 111–147. Alp S et al. (2018). Effects of probiotics on salivary 
Streptococcus mutans and Lactobacillus levels in orthodontic patients. American Journal of 
Orthodontics and Dentofacial Orthopedics, 154, 517–523. Lucchese A et al. (2018). Changes 
in oral microbiota due to orthodontic appliances: a systematic review. Journal of Oral 
Microbiology, 10, 1476645. 
 2. Aim of the Study  
The aim of this study is to evaluate the effect of a 12-week administration of a probiotic 
dietary supplement containing Lactobacillus reuteri Prodentis on: clinical periodontal 
parameters: Full-Mouth Plaque Index (FMPI), Bleeding on Probing (BOP), Probing Depth 
(PD), microbiological parameters of the dental biofilm: Fusobacterium nucleatum, Prevotella 
intermedia, Capnocytophaga gingivalis, Eubacterium nodatum, in adult patients undergoing 
fixed orthodontic treatment with concomitant gingivitis. 
 3. Methodology Study Design 
 A randomized interventional study with a control group. Study Population A total of 100 
adult patients aged 18–59 years undergoing fixed orthodontic treatment and presenting with 
clinical signs of gingivitis (BOP >30%, PD ≤4 mm) will be enrolled. Patients will be 
recruited from individuals routinely treated at the Department of Dental Hygiene, Medical 
University of Warsaw, within the framework of healthcare services reimbursed by the 
National Health Fund (NFZ). Clinical Procedures At the baseline visit (T1), a comprehensive 
periodontal examination will be performed using a graduated periodontal probe (1-mm 
increments), including: FMPI – dichotomous plaque index (presence/absence) assessed on 
four surfaces per tooth (distal, buccal, mesial, lingual), BOP – dichotomous bleeding index 
(presence/absence) assessed at six sites per tooth (distobuccal, mid-buccal, mesiobuccal, 
mesiolingual, midlingual, distolingual), PD – probing depth measured at the same six sites 
per tooth. All patients will undergo ultrasonic scaling and will receive standardized oral 
hygiene instructions. Subsequently, participants will be randomly assigned to one of two 
groups: Test group (A) – 50 patients instructed to use probiotic lozenges containing 

Lactobacillus reuteri Prodentis for 12 weeks, Control group (B) – 50 patients not receiving 
probiotic supplementation.  
2.01.2022 RESEARCH PROTOCOL 
 
 Title of the Study:  
Assessment of the Impact of the Probiotic Limosilactobacillus Reuteri Prodentis (DSM 
17938 and ATCC PTA 5289) on Clinical and Microbiological Parameters in Orthodontically 
Treated Patients with Concomitant Gingivitis 
 
Microbiological Assessment In the first 9 patients from the test group (A1) and 9 patients 
from the control group (B1), microbiological analysis will be performed. Prior to the 
periodontal examination and scaling, pooled dental biofilm samples will be collected from 
the mesial and distal approximal surfaces of teeth 15 and 35 using a sterile paper point 
inserted into the gingival sulcus for approximately 10 seconds with sterile tweezers. The 
collected samples will be transferred to transport tubes and sent to an analytical laboratory. 
Microbiological analysis will be performed using Real-Time PCR, ensuring high sensitivity 
and specificity, enabling accurate qualitative and quantitative determination of the selected 
bacterial species. Follow-up Visit After 12 weeks (T2), clinical and microbiological 
examinations will be repeated in the respective groups. 
 4. Eligibility Criteria 
 Inclusion Criteria: 
- generally healthy patient 
-age 18–59 years 
-treatment with fixed orthodontic appliances 
- plaque-induced gingivitis (BOP >10%, PD ≤4 mm).  
Exclusion Criteria: 
- hypersensitivity to any component of the probiotic preparation 
-smoking 
- use of probiotic supplements within the previous 30 days 
-antibiotic or other antimicrobial therapy within the previous month 
-use of chlorhexidine-containing mouthrinses within the previous 2 weeks 
- age <18 or >59 years. 
 

 
 
2.01.2022 RESEARCH PROTOCOL 
 
 Title of the Study:  
Assessment of the Impact of the Probiotic Limosilactobacillus Reuteri Prodentis (DSM 
17938 and ATCC PTA 5289) on Clinical and Microbiological Parameters in Orthodontically 
Treated Patients with Concomitant Gingivitis 
 
 
 5. Benefits, Risks, and Inconveniences Expected benefits: 
 Improvement in clinical periodontal parameters, including a reduction in BOP and PD, 
resolution of clinical signs of gingivitis (erythema, marginal gingival enlargement), and 
potential favorable changes in the qualitative and quantitative composition of the dental 
biofilm. Risks and inconveniences: Possible occurrence of hypersensitivity reactions to 
components of the dietary supplement. 
 6. Study Duration and Justification December 2021 – December 2025: clinical and 
microbiological data collection November 2025 – July 2026: data analysis and preparation of 
scientific publications and conference presentations Justification: The duration of the study is 
determined by the feasibility of recruiting an adequate number of eligible participants in each 
group in accordance with the adopted inclusion and exclusion criteria, as well as by the 
12-week intervention protocol for each participant.