Aim: Apical periodontitis(AP) caused by root canal infection is the most frequent pathological lesion in the jaws. Bacterial products, host immune cells and biologically active factors called locally produced cytokines(such as IL-1, TNF-α) have been reported to play an important role in the pathogenesis of AP. Metalloproteinases(MMP), is a measurable biomarker that plays an important role in the degradation and regeneration of collagen and is an indicator of collagen. This study aimed to determine the main bacterial species in the microbiota as Gr(+) and Gr(-) and to compare the relationship between MMP-9 and TNF-α with controlled patient groups. Methodology:60 patients with AP and extraction indication were included in the study. 30 systemically and orally healthy volunteers were selected as the control group. After access cavity preparation, an initial microbiologic sample(S1) was taken from the root canal. After atraumatic extraction of the tooth, second microbial sample(S2) was taken from the external root surface. After bacterial DNA extraction, 16S rRNA gene primer was designed for sequence analysis. Bacterial community profiling was made by Sanger sequencing of the PCR products. Besides, blood samples were collected from all of the patients. Enzyme-linked immunosorbent assay was used to measure levels of MMP-9 and TNF-α.

60 patients with AP and extraction indication were included in the study. 30 systemically and orally healthy volunteers were selected as the control group. After access cavity preparation, an initial microbiologic sample(S1) was taken from the root canal. After atraumatic extraction of the tooth, second microbial sample(S2) was taken from the external root surface. After bacterial DNA extraction, 16S rRNA gene primer was designed for sequence analysis. Bacterial community profiling was made by Sanger sequencing of the PCR products. Besides, blood samples were collected from all of the patients. Enzyme-linked immunosorbent assay was used to measure levels of MMP-9 and TNF-α Approximately 450 patients who applied to xxxx University, Faculty of Dentistry, Endodontics Department for routine examination due to apical periodontitis complaints were examined and their demographic characteristics were recorded. Patients who had any acute or chronic inflammatory disease, known infection status, history of trauma or surgical operation within 6 months that may affect MMP-9 levels, pregnant and/or lactating, malignity, morbid obesity, arthritis, immunologic diseases, cardiovascular diseases or use of vitamins with high biotin were not included in the study. Additionally, patients who used antibiotics or anti-inflammatory drugs within the last 6 months, and patients with periodontal disease were excluded from the study. Of these patients, 60 patients with AP and extraction indication (due to restorative causes and lesion size) were included in the study. A total of 30 volunteers who were determined to be systemically and orally healthy (not having AP indication), were included in the study as the control group. The study was conducted between March 2022 and February 2023.

Demographic Characteristics & Clinical and Radiographic Evaluation Laboratory Analysis ELISA measurement procedure of MMP-9 and TNF-α levels Bacteriological Sampling from the Root Canal System and Outer Root Surface Genomic DNA Isolation and Measurement of DNA Concentration PCR (16S Universal primer) Purification and sequencing of the 16S rRNA gene Power Analysis and Sample Size Calculations

Inclusion Criteria:

• AP and extraction indication (due to restorative causes and lesion size) were included in the study

Exclusion Criteria:

• Patients who had any acute or chronic inflammatory disease, known infection status, history of trauma or surgical operation within 6 months that may affect MMP-9 levels, pregnant and/or lactating, malignity, morbid obesity, arthritis, immunologic diseases, cardiovascular diseases or use of vitamins with high biotin were not included in the study. Additionally, patients who used antibiotics or anti-inflammatory drugs within the last 6 months, and patients with periodontal disease were excluded from the study.