The Microflora Diversity And Profiles In Dental Plaque Biofilms From The Isolated Population With And Without Caries

Dental caries is the most common disease,which do harm to the health of oral cavity. Dental plaque biofilms is the main etiological factor. The ecological plaque hypothesis suggests under the dynamic balance the complex bacteria are normal flora, and caries is a result of a shift in the balance of the resident microflora driven by changes in local environmental conditions.There are about 700 oral bacterial species detected in oral biofilms, of which about 50% has not yet been cultivated.The research on these microflora experienced some stages as follows:several cariogenic pathogens,dental plaque,biofilm,the microbial complexity in biofilms.Study on diversity with biofilm with oral diseases,specially with dental caries has become as an international focus.There some limited reports on this field in isolated population.Their living enviroment,life styles and diets habits are relatively closed and chang less.However different individual suscep--- to caries is discrepance.Therefore many play an much important role in canogenicity.Object:The present study used Human Oral Microbe Identification Microarray (HOMIM) to define the bacterial species and phylotypes of microflora in dental plaque from the children with sever caries and the healthy controls in isolated population, and compare the diversity alteration of microbial distribution and their quantitative composition in dental plaque biofilms between sever caries and healthy controls in isolated population, also between isolated population and opened population,in order to understand the phylogenetic relationship between bacteria and caries. For the purpse of to detecting some known pathogenic in these plaque samples,real-time PCR was used to evaluante the method and result. Methods:Plaque samples from 30 children with sever caries and 20 healthy controls were pooled from the intact enamel of the first permanent tooth. Bacterial DNA was extracted from each pooled plaque sample, and from which 16S rRNA genes were amplified by PCR with 16S rRNA universal primers and were labeled via incorporation of Cy3-dCTP in a second nested PCR. Hybridization of labeled amplicons on HOMIM, the data acquired from professional software were analyzed.The Streptococcus mutans, Streptococcus oralis, Streptococcus sobrinus, Lactobacillus, Actinomyces naeslundii in these plaque sample by real-time fluorescent quantitative PCR, and were Statistical analyzed with cluster analyses, Chi-Square x2 Test, and Wilcoxon rank sum test et.al.Results:(1) Bacterial profile of 30 children with sever caries and 20 healthy controls showed 97 bacterial species or clusters representing 6 bacterial phyla and 35 genus. The species/clusters detected in the plaque with sever caries(median is 30) is lower than the healthy controls (median is 39). (2)Subjects with healthy controls were distinguished from sever caries by a significantly higher frequency or quantum of putative pathogens, such as Bacteroidetes[G-2] sp clone AU126_ot274, Capnocytophaga sputigena_ot775, Porphyromonas catoniae and sp clone CW034_ot279, Haemophilus sp clone BJ095_ot036, Campylobacter gracilis_ot623, Campylobacter showae_ot763, Neisseria flavescens_ot610, Streptococcus gordonii_ot622, Streptococcus sanguinis and sp clone C3MLM097_ot058₇58, Actinomyces gerencseriae_ot618, Rothia mucilaginosa_ot681.(P<0.05) (3) Streptococcus mutans,Streptococcus sobrinus were not detectable in the two groups. Actinomyces naeslundii was detectable in only 1 case in the caries-free group, and just 1 smple from caries-active group presented detectable Lactobacillus. (4)There are 3 other bacterial phyla and 2 genus are detected in the opened population:Deferribacteres:Synergistetes[G-3]; Spirochaetes:Treponema; SRI.(5)Regarding the caries-active group in the both population:Prevotella, Eubacterium, Kingella oralis_ot706, Kingella denitrificans_ot582, Streptococcus gordonii_ot622, Streptococcus australis,Actinomyces gerencseriae_ot618, Actinomyces naeslundii_ot176, Veillonella parvula_ot161 and Selenomonas sputigena in opened population respectively present the detection rate of 100%,100%,60%,70%,90%, 90%,60%,60%,100%,70%, which are higher than that in the isolated population 53.3%,26.7%,13.3%,13.3%,33.3%,40%,0,0,40%,20%(P<0.05). The higher quantum of Streptococcus australis,Streptococcus infantis FN042_ot065₆38, Capnocytophaga sputigena_ot775, Prevotella, Campylobacter,Veillonella,Haemophilus were found in opened population(P<0.05).(6)Regarding the caries-free group:Eubacterium, Capnocytophaga ochracea, Prevotella multisaccharivorax_ot794, Kingella oralis_ot706),Neisseria sp clone BM052_ot009,Eubacterium[14][G-1] saburreum_ot494, Selenomonas noxia and sp clone EL028 and Gemella sanguinis_ot757 were identified at significantly higher level than those.(p<0.05) (7)Species/clusters that were more quantum in the caries-active than in the caries-free group in the opened population included Prevotella melaninogenica and sp clone BE073_ot298₄69, Haemophilus sp clone BJ095_ot036, Streptococcus australis and sp clone FN042_ot065₀73, however the results are reverse in the isolated population,there are more quantum in the health controls.(8) Streptococcus mutans, Streptococcus oralis, Streptococcus sobrinus, Lactobacillus, Actinomyces naeslundii are detectable in all samples by using real-time fluorescent quantitative PCR.The detective level of Streptococcus mutans, Lactobacillus and Actinomyces naeslundii in caries-free group were higher level in caries-active group.Nevertheless, these diffirences have not statistical significance.Conclusion:A lower bacterial diversity was observed in the isolated population compared to the opened population.The distribution diffirence of Species/clusters between the two population is significant difference. Some species such as Prevotella melaninogenica, Haemophilus sp clone BJ095_ot036 and Streptococcus australis are pathogenic in opened group, however these are beneficial bacteria in isolated population. Streptococcus mutans, Lactobacillus, Streptococcus sobrinus maybe not necessary for cariogenicity.Possibility caries is the outcome of the overall activity of the total plaque microflora, and as a result of a shift in the balance of the resident microflora,which support the ecological plaque hypothesis.Real-time quantitative PCR for quantitative detection of microbial composition in dental plaque,issensitive,but its reliability and accuracy still need to to be improved.