Comparative Analysis Of Microbial Diversity With Discordant Caries Phenotype Twins And Their Mothers In Supragigival Plaque

Dental caries is one of the most common diseases which is harmful to children's oral health.Early childhood caries(ECC)is defined as the presence of one or more decayed(non cavitated or cavitated lesions),missing teeth(due to caries),or filled tooth surfaces in any primary tooth in a child 71 months age or younger.The incidence of dental caries among 5-year-old Chinese children has reached to 66%and remained much higher than other countries in the world,according to the Third National Epidemiological Survey on Oral Health which conducted in 2005.It suggested that ECC is still one of the most frequently occurring diseases in oral cavity among preschool children,and it has become an important public health problem in China,furthermore it has harmful influence on children's health and growth.The four factors theory thinks ECC is the consequence of complex interplay of host,oral microbial,the food and time,among which the oral microbial as the most important factor received a great deal of attention from oral clinical doctors and researchers.Later,there are many researches pay close attention to the influence of overall health,diet custom and environmental factors for ECC.However,much of these studies were focused on the microorganisms which can be cultured in the laboratory such as Streptococcus mutans genetic,Lactobacillus and Actinomyces in oral cavity,and the influence of family environment and host genetics for ECC are still not clearly.Objective:To investigate the role of the oral microbiome and the impact of host genetics and environmental factors on microbiota variation,supragigival plaque were pooled and then analyzed from 14 twin pairs,a set of triplets with discordant caries phenotype and 15 mothers,applying the technique of Human Oral Microbe Identification using Next Generation Sequencing(HOMINGS).Thus can lay the foundation of identifying risk factors of ECC,and develop effective preventive strategies.Methods:31 children aged from 3-6 years old contains 14 pairs of twins,a set of triplets and 15 mothers were selected and accepted oral health check,according to the forth guidelines of World Health Organization(WHO)for dental caries diagnose.All sebjects were not taking antibiotics over the past three monthes,and had no drug or food allergy and other systemic diseases.Moreover,none of these selected subjects had moderate or severe periodontitis and significant oral mucosal disease.Twins should fulfill the condition that one had carious teeth and the other was healthy in oral cavity.Supragingival plaque were collected and pooled,and then the whole DNA of bacteria was extracted.HOMINGS was performed to identify the bacteria composition of supragingival plaque,and then bioinformatics analysis were used to compare the community structure and species diversity.Results:A total of 2,293,650 reads and 20,431 OTUs had been captured,aligned with the reference of Greengene database and clustered OTUs at the similarity level of 97%,revealed 11 phyla,116 genera and 139 species of microflora with the platform of Miseq in HOMINGS.At the phylum level,the abundance of Bacteroidetes and Firmicutes are much higher in Mother group compared with Caries and Caries-free group(P<0.05),while at the genus level,the relative abundances of these strains varied dramatically among the children.Comparative analysis between Caries group and CariesFree group revealed that the frequency of Actinomyces,Lactobacillus,Haemophilus and Streptococcus's detection are much higher in caries group(P<0.05).The sample from the mother group has a relative higher abundance of Prevotella,Tannerella,Parvimonas and Bacteroidales_[G-2]than Caries group and CariesFree group(P<0.05).Analyzing alpha diversity and it suggested that the species richness and evenness of supragingival plaque exhibited much higher level in mother group compared with caries and caries-free group(P<0.05).Furthermore,monozygotic twins exhibited much higher similarities than dizygotic twins.Conclusion:The results demonstrated that Streptococcus,Actinomyces,Lactobacillus and Haemophilus are strongly associated with ECC and can be potential biomarkers of dental caries in the primary dentition.Moreover,the species richness and evenness of supragingival plaque in adults has a significant difference with children,suggesting that the composition of oral microbial was changeable at different stage of life.In addition,the oral microbiota of adults tends to be susceptible to periodontal diseases.Furthermore,the weighted unifrac distance between monozygotic twins was much shorter than dizygotic twins,indicating that the host genetic factors have an influence on the composition of oral mirobiota.