Metagenomic Analysis Of The Oral Microbiome Reveals The Different Signatures Of Aggressive Periodontitis Before And After Treatment

Objective:Oral cavity is the channel between human body and the outside environment. It’s the portal through which microorganisms intrude into human’s digestive and respiratory tract. Therefore microorganisms are easy to enter the oral cavity. Oral cavity is also one of the most complex environments in human body with the complex microbial community. In general, there are more than seven hundred of bacteria, fungus,virus and so on. Those microorganism are associated with parasitifer’s caries and periodontitis. Normally, the alien microbes in the oral cavity exists complex symbiotic relationships, to keep the relative balance between microbial community, while maintaining the dynamic balance between microorganism and the host. Moreover, the balance is very important to maintain the oral health. This balance is not invariable, microbial species and quantity change along with the age, diet, oral hygiene status, local and systemic conditions. Thus, the balance is relative, conditional and variable. The host can maintain oral health, when it keeps the dynamic balance not only between bacteria in the flora but also between bacteria within the host. Otherwise, when the balance is destroyed, it will cause a host of oral diseases, and even affect the health of the whole body.Nowadays, the relationships between metagenome and human healthy and disease state have drawn more and more attentions. Microbiomes at different body sites and functional metagenomics must be considered as parts of systemic biology. The profound differences between microbiomes in various body sites reveal how metagenomes contribute to tissue and organ function. Differences in microbial composition and function are being linked to health and disease states in different organs and tissues. More detailed knowledge of human microbiomes will yield next-generation diagnostics and therapeutics for various human diseases.This study focused on the microbial composition and diversity of aggressive periodontitis before and after periondontal initial therapy. Our results revealed the microbial composition and diversity in the level of microbial community before and after treatment. Moreover, our study explored the relationship between the variation of microflora structure and aggressive periodontitis.Methods:The medical history of two patients with aggressive periodontitis were required and recorded. The oral cavity was examined, and pocket probing was performed and panoramic radiography was taken. The periodontal parameters of bleeding on probing (BOP) and probing depth (PD) were recorded. Two patients received mechanical debridement (including supragingival and subgingival scaling and root planning). Periodontal clinical examination and subgingival samples were taken at baseline and one month after periodontal therapy. We cleaned the surface of teeth with cotton swab, collected the subgingival plaque with the curratage, and transfered the dental plaque into EP tube with 0.5ml PBS in it, and then centrifuged the EP tube 5min at 3000r/min. Next the supernate was discarded and the EP tubes with precipatates were put into refrigerator at -80℃. Total genome DNA from samples was extracted using CTAB method. V4 hypervariable regions of 16s rDNA were picked up for PCR amplification. Finally, the PCR products of V4 hypervariable region were sequenced with Illumina MiSeq platform and 250 bp.Results:1 One month after initial periodontal therapy, both the clinical features and periodontal parameters (BOP, PD) improved dramatically.2 PCR products 16 samples of V4 hypervariable region were sequenced with Illumina MiSeq platform and 250 bp. Those samples could be annotation to the level of genus. With Illumina Miseq, we obtained 952272 raw resds. After filtration, we obtained 915626 nochime reads. After comparing and analyzing with gene bank, we got And 887803 reads for classification.3 The composition and structure of subgingival bacterial community changed after periodontal initial therapy. The percentage of Actinobacteria and Proteobacteria increased obviously. The percentage of Bacteroidetes、Spirochaetes and Synergistetes decreased dramatically. The difference of bacterial composition and diversity at two time points was remarkably. Before treatment, the genus of microbiomes was more diverse and the structure of bacterial community was more complicated than those after therapy. The structure of subgingival plaque was similar between different individuals before treatment, while it was quite different between different individuals after therapy.Conclusion:In conclusion, this study examined the change of microbial composition and diversity of by high-throughput metagenomics analysis for two generalized aggressive periodontitis patients before and after periodontal initial therapy. Our results indicated that Bacteroidetes, spirochaetes and Synergistetes were related to pathogenicity of aggressive periodontitis while Actinobacteria and Proteobacteria seemed associated with clinical symptoms resolution. This study demonstrated the richness and unevenness of the subgingival bacterial community, as well as the change of composition and diversity of community following periodontal treatment. Further study may collect a larger set of subjects to examine the causal relationship between the subgingival microbiota and periodontal health and disease.