Microbial Profiling in Pockets Related to Chronic Periodontitis Patients Using 16s RNA Metagenomics Sequencing
- Conditions
- Periodontal Diseases
- Interventions
- Genetic: 16s RNA Metagenomics sequencing
- Registration Number
- NCT04425343
- Lead Sponsor
- Ajman University
- Brief Summary
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.
- 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
Recruitment & Eligibility
- Status
- COMPLETED
- Sex
- All
- Target Recruitment
- 80
- Patients recruited for the study were diagnosed as Stage II Generalized periodontitis
- Medically fit
- Patients having gingivitis or stage III periodontitis
- Medically unfit
Study & Design
- Study Type
- INTERVENTIONAL
- Study Design
- SINGLE_GROUP
- Arm && Interventions
Group Intervention Description Metgenomic analysis 16s RNA Metagenomics sequencing Analysis for whole bacterial count Periodontitis, Adult 16s RNA Metagenomics sequencing Plaque samples were taken from subgingival pocket and send to the lab for metagenomic analysis
- Primary Outcome Measures
Name Time Method Microbial Taxonomical Composition From Baseline to 3 months The composition was determined using subgingival plaque samples and performing 16s metagenomic sequencing
- Secondary Outcome Measures
Name Time Method
Trial Locations
- Locations (1)
Ajman University of Science and Technology
🇦🇪Ajman, United Arab Emirates