Microbial Profiling in Pockets Related to Chronic Periodontitis Patients Using 16s RNA Metagenomics Sequencing

Microbial Profiling in Pockets Related to Chronic Periodontitis Patients in UAE Population Using 16s RNA Metagenomics Sequencing

The study was developed in compliance with the Helsinki Declaration on medical research involving the Ethics Committee at Ajman University (2017-A-DN-04). Informed consent from all the participants was obtained before participation in the study. The participants were systemically healthy with no history of antibiotics for the past three months. Patients recruited for the study were diagnosed as Stage II Generalized periodontitis according to the classification from the 2017 world workshop on the "classification of periodontal and peri-implant disease and conditions". Periodontal status indicating the severity of interdental clinical attachment loss of 3-4mm, radiographic bone loss between 15%-33% and with no tooth loss were included. The complexity of periodontitis with a maximum probing depth of ≤ 5mm with horizontal bone loss and having an extent and distribution with >30% teeth involved were included in the study. A total of 80 plaque samples were collected with the clinical characteristics of the patient comprising of age between 25-39 years, 36 females and 44 males. The subgingival plaque was collected using a sterile curette from the buccal aspect of maxillary molars and lingual aspect of mandibular incisors, in a vial containing 200µl of Buffer CL.

Study Overview

• Status: Completed
• Conditions: Periodontal Diseases
• Intervention / Treatment: Genetic: 16s RNA Metagenomics sequencing

Detailed Description

The collected plaque samples were then freshly prepared for the DNA isolation using ABIOpure TM Total DNA (version 2.0) (Cat No: M501DP100) according to the manufacturer's instruction. All the samples were assessed for DNA quantification using spectrophotometry and further quantified using the fluorometric method. This was performed using DeNovix DS-11 FX (DeNovix). Further to the DNA quantification, assessment of DNA integrity was resolved on a 0.8% agarose gel with ethidium bromide. For NGS library preparation, all the samples underwent PCR amplification of the 16S rRNA gene in the isolated bacterial DNA. The primers used had targeted in the V3-V4 region of the 16S rRNA gene. The full length primer sequences, using standard IUPAC nucleotide nomenclature, to follow the protocol targeting this region are:

16SAmpliconPCRForwardPrimer=5' TCGTCGGCAGCGTCAGATGTGTATAAGAGACAGCCTACGGGNGGCWGCAG 16SAmpliconPCRReversePrimer=5' GTCTCGTGGGCTCGGAGATGTGTATAAGAGACAGGACTACHVGGGTATCTAATCC Gel electrophoresis method was used for the size selection and the bioinformatics pipeline used for processing microbiome 16S sequence data was QIIME

• Study Type: Interventional
• Enrollment (Actual): 80
• Phase: Not Applicable

Contacts and Locations

Study Locations

• United Arab Emirates
  • Ajman, United Arab Emirates, 009716
    • Ajman University of Science and Technology

Participation Criteria

Eligibility Criteria

• Ages Eligible for Study: 16 years to 58 years (Adult)
• Accepts Healthy Volunteers: No
• Genders Eligible for Study: All

Description

Inclusion Criteria:

• Patients recruited for the study were diagnosed as Stage II Generalized periodontitis
• Medically fit

Exclusion Criteria:

• Patients having gingivitis or stage III periodontitis
• Medically unfit

Study Plan

How is the study designed?

Design Details

• Primary Purpose: Health Services Research
• Allocation: Randomized
• Interventional Model: Single Group Assignment
• Masking: Single

Number of Arms

2

Arms and Interventions

Participant Group / Arm	Intervention / Treatment
Experimental: Periodontitis, Adult; Plaque samples were taken from subgingival pocket and send to the lab for metagenomic analysis	Genetic: 16s RNA Metagenomics sequencing; The subgingival plaque was collected using a sterile curette from the buccal aspect of maxillary molars and lingual aspect of mandibular incisors, in a vial containing 200µl of Buffer CL.; Other Names: DNA isolation
Experimental: Metgenomic analysis; Analysis for whole bacterial count	Genetic: 16s RNA Metagenomics sequencing; The subgingival plaque was collected using a sterile curette from the buccal aspect of maxillary molars and lingual aspect of mandibular incisors, in a vial containing 200µl of Buffer CL.; Other Names: DNA isolation

What is the study measuring?

Primary Outcome Measures

Outcome Measure	Measure Description	Time Frame
Microbial Taxonomical Composition	The composition was determined using subgingival plaque samples and performing 16s metagenomic sequencing	From Baseline to 3 months

Collaborators and Investigators

• Sponsor: Ajman University
• Investigators: Principal Investigator: Sudhir Varma, MDS, Assistant Professor

Study record dates

Study Major Dates

• Study Start (Actual): November 10, 2017
• Primary Completion (Actual): September 15, 2018
• Study Completion (Actual): October 7, 2019

Study Registration Dates

• First Submitted: June 7, 2020
• First Submitted That Met QC Criteria: June 9, 2020
• First Posted (Actual): June 11, 2020

Study Record Updates

• Last Update Posted (Actual): June 11, 2020
• Last Update Submitted That Met QC Criteria: June 9, 2020
• Last Verified: June 1, 2020

More Information

Terms related to this study

• Keywords: metagenomics; 16S; Periodontitis, Adult
• Additional Relevant MeSH Terms: Stomatognathic Diseases; Mouth Diseases; Periodontitis; Periodontal Diseases; Chronic Periodontitis
• Other Study ID Numbers: F-H-17-11-03

Plan for Individual participant data (IPD)

• Plan to Share Individual Participant Data (IPD)?: No
• IPD Plan Description: Microbial Taxonomical Composition

Drug and device information, study documents

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