Detection And Quantification of Eubacterium Saphenum and Porphyromonas Gingivalis In Periodontal Health and Disease (A Case-Control)

Detection And Quantification of Eubacterium Saphenum and Porphyromonas Gingivalis In Periodontal Health and Disease: A Case-Control Study

This case-control study will be conducted between January and June 2026 and will include 148 participants aged over 30 years to investigate differences between individuals with periodontitis (n = 111) and those with periodontal health (n = 37). Participants will be selected based on strict inclusion and exclusion criteria to minimize confounding factors, and ethical approval along with informed consent will be obtained prior to participation. Periodontitis will be diagnosed according to the 2017 classification criteria, requiring clinical attachment loss and periodontal pocketing, while periodontal health will be defined by minimal bleeding on probing, shallow pocket depths, and absence of attachment loss. The sample size will be determined using G*Power to ensure adequate statistical power.

Clinical periodontal parameters-including plaque index, gingival bleeding index, probing pocket depth, and clinical attachment loss-will be recorded using standardized methods at multiple sites per tooth by calibrated examiners to ensure reliability. Following clinical assessment, subgingival biofilm samples will be collected using sterile paper points under controlled conditions and stored at -80°C for subsequent real-time PCR analysis. Calibration procedures, including inter- and intra-examiner agreement assessed by intraclass correlation coefficients, will be implemented to enhance measurement consistency. Overall, the study will combine rigorous clinical evaluation with molecular techniques to enable a comprehensive assessment of periodontal status and associated microbiological factors.

Study Overview

• Status: Enrolling by invitation
• Conditions: Periodontitis; Periodontal Health
• Intervention / Treatment: Diagnostic test: Microbiological sampling After recording the clinical periodontal parameters, subgingival biofilm samples were collected. After selecting the sampling site, it was isolated with cotton rolls to keep i

Detailed Description

This case-control study will be conducted between January and June 2026 and will include a total of 148 participants aged over 30 years. The study population will consist of 111 subjects with periodontitis and 37 subjects with periodontal health. Ethical approval will be obtained from the Institutional Ethics Review Board (Certificate No. 2015-16/1118), and written informed consent will be secured from all participants prior to examination. Eligible individuals will be selected based on predefined inclusion and exclusion criteria and will be categorized into a healthy group and a periodontitis group according to clinical periodontal parameters. Inclusion criteria will require participants to be over 30 years old and to have at least 20 teeth. Exclusion criteria will include individuals with active caries, those undergoing orthodontic treatment, recent antibiotic use within three months, history of valve replacement requiring prophylactic antibiotics, recent use of non-steroidal anti-inflammatory drugs, pregnancy or lactation, alcohol use, and receipt of periodontal therapy within the last six months.

Participants in the periodontitis group will be diagnosed based on the 2017 classification, defined by interdental clinical attachment loss (CAL) at ≥2 non-adjacent teeth or buccal/oral CAL ≥3 mm with probing pocket depth (PPD) >3 mm at ≥2 teeth. Cases will be further characterized as generalized periodontitis affecting more than 30% of teeth, with unstable disease indicated by sites exhibiting PPD ≥5 mm or PPD ≥4 mm with bleeding on probing (BOP), and staged from I to IV. The control group will include individuals with periodontal health, defined by BOP <10%, PPD ≤3 mm, and absence of attachment loss. The sample size will be calculated using G*Power software (version 3.1.9.7) based on an expected prevalence of 25% in healthy individuals and 50% in periodontitis patients, with 80% power and a significance level of 0.05, resulting in a required total of 148 participants.

Clinical periodontal parameters will be assessed following participant selection. Plaque Index (PI) will be recorded at four sites per tooth using a disclosing agent and a dichotomous scoring system. Gingival Bleeding Index (GBI) will be evaluated at the same sites using a UNC-15 periodontal probe, with bleeding recorded 30 seconds after probing. Probing pocket depth (PPD) and clinical attachment loss (CAL) will be measured at six sites per tooth using a manual periodontal probe. To ensure measurement reliability, inter-examiner calibration will be performed between two examiners on selected quadrants and sites, while intra-examiner calibration will be conducted by repeating measurements in two sessions. Agreement will be analyzed using the intraclass correlation coefficient (ICC) in SPSS.

Following clinical examination, subgingival biofilm samples will be collected from selected sites. The sampling area will be isolated with cotton rolls to prevent contamination, and sterile paper points will be inserted into the base of the periodontal pocket for 30 seconds. Samples will then be transferred into sterile tubes containing preservative solution, properly labeled, and stored at -80°C to maintain DNA integrity. These samples will subsequently be analyzed using real-time polymerase chain reaction (RT-PCR) to identify and quantify bacterial presence.

• Study Type: Observational
• Enrollment (Estimated): 148

Contacts and Locations

Study Locations

• Iraq
  • Babel
    • Hillah, Babel, Iraq, 3001
      • University of babylon

Participation Criteria

Eligibility Criteria

• Ages Eligible for Study: Adult; Older Adult
• Accepts Healthy Volunteers: Yes

• Sampling Method: Probability Sample

Study Population

This study is an observational case-control study. Patients with generalized unstable periodontitis, at any grade, with (>4 mm) probing pocket depths (PPD), as well as periodontal health participants, will be recruited from the Department of Periodontics, College of Dentistry, University of Babylon. A written consent forms will be obtained from all participants. Ethical approval will be obtained.

Description

Inclusion Criteria:

• (1) Participants aged > 30 years old. (2) Participants who have at least 20 teeth

Exclusion Criteria:

• 1) Participants who have active caries. (2) Participants who are undergoing orthodontic treatment. (3) Participants with a history of antibiotic use within the last three months will not be eligible.

  (4) Participants with a history of valve replacement which requires prophylactic antibiotic coverage. (5) Participants who require systemic or topical non-steroidal anti-inflammatory drugs within the past three months. Participants who are pregnant or lactating women. (6) Participants with a history of alcohol use. (7) Participants who had periodontal therapy within the last six months

Study Plan

How is the study designed?

Number of groups / cohorts

2

Cohorts and Interventions

Group / Cohort	Intervention / Treatment
Peiordontal health; In the control group, the participants were diagnosed periodontal health characterized by BOP < 10%, PPD ≤ 3mm, and intact periodontium with no probable attachment loss (Chapple et al., 2018)	Diagnostic test: Microbiological sampling After recording the clinical periodontal parameters, subgingival biofilm samples were collected. After selecting the sampling site, it was isolated with cotton rolls to keep i; Microbiological sampling After recording the clinical periodontal parameters, subgingival biofilm samples were collected. After selecting the sampling site, it was isolated with cotton rolls to keep it dry and avoid saliva contamination. Then, a sterile paper point will be gently inserted into the selected sites reaching the base of the pocket and will be kept for 30 seconds. Then, the paper point will be carefully withdrawn and placed immediately into a sterile tube containing a preservative solution. The tube will be labeled with details, including the patient ID, site location, and date, and then sealed tightly to prevent contamination. The sample was stored at -80°C to preserve DNA quality for subsequent Real time-PCR analysis.
Periodontitis; Participants were diagnosed as periodontitis when they have interdental CAL detectable at ≥2 non-adjacent teeth, or facial and/or oral CAL ≥3 mm associated with pocketing >3mm detectable at ≥ 2 teeth. The diagnosis statement was: Generalized periodontitis (>30% of teeth having CAL), Unstable status (having at least one site with PPD ≥5 mm or PPD ≥4 mm with BOP), at stages I-IV (Tonetti, Greenwell and Kenneth S. Kornman, 2018)	Diagnostic test: Microbiological sampling After recording the clinical periodontal parameters, subgingival biofilm samples were collected. After selecting the sampling site, it was isolated with cotton rolls to keep i; Microbiological sampling After recording the clinical periodontal parameters, subgingival biofilm samples were collected. After selecting the sampling site, it was isolated with cotton rolls to keep it dry and avoid saliva contamination. Then, a sterile paper point will be gently inserted into the selected sites reaching the base of the pocket and will be kept for 30 seconds. Then, the paper point will be carefully withdrawn and placed immediately into a sterile tube containing a preservative solution. The tube will be labeled with details, including the patient ID, site location, and date, and then sealed tightly to prevent contamination. The sample was stored at -80°C to preserve DNA quality for subsequent Real time-PCR analysis.

What is the study measuring?

Primary Outcome Measures

Outcome Measure	Measure Description	Time Frame
The level of E. saphenum in periodontitis and periodontal health groups.	Measuring the level of E. saphenum and P. ginigvalis in study groups using Real-time PCR	8 MONTHS

Secondary Outcome Measures

Outcome Measure	Measure Description	Time Frame
Comparing the level of E. saphenum and P. gingivalis between study groups.	Comparing the level of E. saphenum and P. gingivalis between study groups.	8 months

Collaborators and Investigators

• Sponsor: Sarah Al-Rihaymee

Publications and helpful links

General Publications

• Castillo Y, Delgadillo NA, Neuta Y, Iniesta M, Sanz M, Herrera D, Pianeta R, Lafaurie GI, Castillo DM. Design and validation of a quantitative polymerase chain reaction test for the identification and quantification of uncultivable bacteria associated with periodontitis. Arch Oral Biol. 2023 Oct;154:105758. doi: 10.1016/j.archoralbio.2023.105758. Epub 2023 Jul 4.

Study record dates

Study Major Dates

• Study Start (Actual): January 31, 2026
• Primary Completion (Estimated): June 1, 2026
• Study Completion (Estimated): August 1, 2026

Study Registration Dates

• First Submitted: May 6, 2026
• First Submitted That Met QC Criteria: May 6, 2026
• First Posted (Actual): May 12, 2026

Study Record Updates

• Last Update Posted (Actual): May 12, 2026
• Last Update Submitted That Met QC Criteria: May 6, 2026
• Last Verified: May 1, 2026

More Information

Terms related to this study

• Keywords: periodontitis; Plaque; P. gingivalis; E.saphenum
• Additional Relevant MeSH Terms: Periodontal Diseases; Mouth Diseases; Stomatognathic Diseases; Pathological Conditions, Anatomical; Pathological Conditions, Signs and Symptoms; Periodontitis; Plaque, Amyloid
• Other Study ID Numbers: 2/5/2026; Sarah (Other Identifier: Sarah)

Plan for Individual participant data (IPD)

• Plan to Share Individual Participant Data (IPD)?: UNDECIDED
• IPD Plan Description: Undecided Yet.

Drug and device information, study documents

• Studies a U.S. FDA-regulated drug product: No
• Studies a U.S. FDA-regulated device product: No