CORE: A Phylogenetically-Curated 16S rDNA Database of the Core Oral Microbiome

Comparing bacterial 16S rDNA sequences to GenBank and other large public databases via BLAST often provides results of little use for identification and taxonomic assignment of the organisms of interest. The human microbiome, and in particular the oral microbiome, includes many taxa, and accurate identification of sequence data is essential for studies of these communities. For this purpose, a phylogenetically curated 16S rDNA database of the core oral microbiome, CORE, was developed. The goal was to include a comprehensive and minimally redundant representation of the bacteria that regularly reside in the human oral cavity with computationally robust classification at the level of species and genus. Clades of cultivated and uncultivated taxa were formed based on sequence analyses using multiple criteria, including maximum-likelihood-based topology and bootstrap support, genetic distance, and previous naming. A number of classification inconsistencies for previously named species, especially at the level of genus, were resolved. The performance of the CORE database for identifying clinical sequences was compared to that of three publicly available databases, GenBank nr/nt, RDP and HOMD, using a set of sequencing reads that had not been used in creation of the database. CORE offered improved performance compared to other public databases for identification of human oral bacterial 16S sequences by a number of criteria. In addition, the CORE database and phylogenetic tree provide a framework for measures of community divergence, and the focused size of the database offers advantages of efficiency for BLAST searching of large datasets. The CORE database is available as a searchable interface and for download at http://microbiome.osu.edu

Beyond Streptococcus mutans: dental caries onset linked to multiple species by 16S rRNA community analysis.

Dental caries in very young children may be severe, result in serious infection, and require general anesthesia for treatment. Dental caries results from a shift within the biofilm community specific to the tooth surface, and acidogenic species are responsible for caries. Streptococcus mutans, the most common acid producer in caries, is not always present and occurs as part of a complex microbial community. Understanding the degree to which multiple acidogenic species provide functional redundancy and resilience to caries-associated communities will be important for developing biologic interventions. In addition, microbial community interactions in health and caries pathogenesis are not well understood. The purpose of this study was to investigate bacterial community profiles associated with the onset of caries in the primary dentition. In a combination cross-sectional and longitudinal design, bacterial community profiles at progressive stages of caries and over time were examined and compared to those of health. 16S rRNA gene sequencing was used for bacterial community analysis. Streptococcus mutans was the dominant species in many, but not all, subjects with caries. Elevated levels of S. salivarius, S. sobrinus, and S. parasanguinis were also associated with caries, especially in subjects with no or low levels of S. mutans, suggesting these species are alternative pathogens, and that multiple species may need to be targeted for interventions. Veillonella, which metabolizes lactate, was associated with caries and was highly correlated with total acid producing species. Among children without previous history of caries, Veillonella, but not S. mutans or other acid-producing species, predicted future caries. Bacterial community diversity was reduced in caries as compared to health, as many species appeared to occur at lower levels or be lost as caries advanced, including the Streptococcus mitis group, Neisseria, and Streptococcus sanguinis. This may have implications for bacterial community resilience and the restoration of oral health

Bacterial 16S Sequence Analysis of Severe Caries in Young Permanent Teeth ▿

Previous studies have confirmed the association of the acid producers Streptococcus mutans and Lactobacillus spp. with childhood caries, but they also suggested these microorganisms are not sufficient to explain all cases of caries. In addition, health-associated bacterial community profiles are not well understood, including the importance of base production and acid catabolism in pH homeostasis. The bacterial community composition in health and in severe caries of the young permanent dentition was compared using Sanger sequencing of the ribosomal 16S rRNA genes. Lactobacillus species were dominant in severe caries, and levels rose significantly as caries progressed from initial to deep lesions. S. mutans was often observed at high levels in the early stages of caries but also in some healthy subjects and was not statistically significantly associated with caries progression in the overall model. Lactobacillus or S. mutans was found either at low levels or not present in several samples. Other potential acid producers observed at high levels in these subjects included strains of Selenomonas, Neisseria, and Streptococcus mitis. Propionibacterium FMA5 was significantly associated with caries progression but was not found at high levels. An overall loss of community diversity occurred as caries progressed, and species that significantly decreased included the Streptococcus mitis-S. pneumoniae-S. infantis group, Corynebacterium matruchotii, Streptococcus gordonii, Streptococcus cristatus, Capnocytophaga gingivalis, Eubacterium IR009, Campylobacter rectus, and Lachnospiraceae sp. C1. The relationship of acid-base metabolism to 16S rRNA gene-based species assignments appears to be complex, and metagenomic approaches that would allow functional profiling of entire genomes will be helpful in elucidating the microbial pathogenesis of caries

Correlation between relative levels of Veillonella and acidogenic streptococci in white spot lesions.

<p>The total % abundance of <i>S. mutans</i>, <i>S. sobrinus</i>, and <i>S. vestibularis/salivarius</i> combined is plotted against the abundance of the <i>Veillonella atypica/dispar/parvula</i> group expressed as a fraction of the remaining community. The result of a linear regression is shown as a line with the indicated parameters.</p

Candidate microbial risk and protective factors.

<p>Candidate microbial risk and protective factors are listed for the onset of caries in subjects that were caries free at baseline and the progression of caries in subjects that had caries at baseline.</p

Relative levels of bacterial taxa by advancing stage of caries.

<p>Graphs at the level of phylum, genus and species are shown. Taxa are sorted by magnitude of change with stage of caries (linear mixed effects model estimates), so that taxa associated with health sort at the bottom and taxa associated with caries are shown at the top. “*” indicates taxa with <i>p</i><0.05 and “**” indicates taxa with <i>p</i><0.01 after the false discovery rate correction was applied. Only genera found at greater than 0.1% of total clones and species found at greater than 0.2% of total clones are shown, and only those taxa significantly associated with caries or health are shown in the species-level graph.</p

Sample sizes at baseline and outcomes at longitudinal follow-up.

<p>Sample sizes at baseline and outcomes at longitudinal follow-up.</p

Among-subject differences visualized by non-metric multidimensional scaling (NMDS) ordination based on Bray-Curtis Dissimilarity.

<p>Among-subject differences by longitudinal caries activity were observed. The plots represent baseline community profiles for healthy subjects and subjects with caries. The baseline patient class and subsequent disease activity are color-coded. Samples from healthy enamel were the only stage available from all subjects and so were used here. The top panel shows the NMDS plot, with the centroid for each clinical group marked. The <i>p</i>-value is for the overall ANOSIM model. The points in lower panels are sized by abundance for the most common species significantly associated with caries and health, and <i>p</i>-values are for the linear mixed effects model estimates.</p

Decreasing species diversity was observed with increasing caries severity both within and among subjects.

<p>Mean Shannon Diversity Indices with 95% confidence intervals are shown. The upper panel shows diversity <i>within</i> subjects for stage of caries at baseline. Diversity was modeled using a linear mixed effects model (SAS PROC MIXED), and is shown as a dashed line (estimate =  −0.26). <i>Post hoc</i> comparisons between sample types were significant, except between white spot and cavitated lesions. The lower panel shows species diversity comparisons <i>among</i> subjects by their baseline and longitudinal caries status for samples collected from noncarious enamel (the only type of sample available from all groups) using ANOVA. Significant <i>post hoc</i> comparisons are indicated by blue lines.</p

Heatmap and cluster analysis of baseline samples from white spot lesions.

<p>Abundances of those bacterial species significantly associated with caries are shown, except for Veillonella which was ubiquitous and therefore omitted. The samples (one from each of 36 subjects) are arranged by hierarchical clustering using the average method and Bray-Curtis dissimilarity. Abundance as percentage of the total community is indicated by the color scale. The bar along the left side indicates longitudinal caries activity.</p