Phylogenetic analyses of the 16S rRNA gene and Arg-gingipain A of Porphyromonas gingivalis clinical isolates

• Main Authors: Yi-Hsin Chan, 陳宜欣
• Other Authors: Keh-Liang Lin
• Format: Others
• Language: zh-TW
• Published: 2015
• Online Access: http://ndltd.ncl.edu.tw/handle/35377341251011792264

碩士 === 中山醫學大學 === 醫學檢驗暨生物技術學系碩士班 === 103 === Background and aim: Periodontitis has a high prevalent rate in Taiwan and Porphyromonas gingivalis is a major pathogen in destructive periodontal disease in humans. Porphyromonas gingivalis produces a unique class of cysteine proteinases termed gingipains that comprises Arg-gingipain A (RgpA). Gingipains have been implicated in the pathogenicity of Porphyromonas gingivalis, a major etiologic agent of chronic periodontitis. The purpose of our studies is to determine the phylogenies of 16S RNA gene and Arg-gingipain (RgpA) gene of clinical P. gingivalis strains among different years. Methods: Samples were collected from gingival pockets of periodontitis patients. All samples were treated with genomic DNA extraction, and PCR was used to amplify 16S RNA and RgpA genes for DNA sequencing. DNA sequences of all amplicons were subjected to phylogenetic analysis by using MEGA 6.01. In addition, the DNA sequences of 16S RNA genes were also deposited and submitted to National Center for Biotechnology Information Genbank database to acquire accession numbers. Results: Fifty four samples were collected from gingival pockets of periodontitis patients. The positive rates of P. gingivalis were 38.9% and 18.5% by PCR and by culture, respectively. 16S RNA and RgpA gene fragments from ten samples were amplified and sequencing. Phylogenetic analysis of 16S RNA genes indicated that P. gingivalis strains were divided into two clusters and our isolates from each other were similar to two clusters. Similarly, phylogenetic analysis of RgpA genes showed that compared with the other strains, the P.g_CSMU24、P.g_CSMU33、P.g_CSMU38、P.g_CSMU41 and P.g_CSMU51 of our clinical isolates were significantly different. Conclusion: P. gingivalis was steadily evolved on both 16S RNA gene and RgpA gene. The correlation between gene variations, fitness on human being and disease severity is requires further studies.