Project abstract for group rudneyjd

Metaproteomic Analysis of Severe Early Childhood Caries

Severe caries in children under the age of six (S-ECC) remains a major problem in dentally underserved communities. S-ECC etiology has been linked to frequent sucrose exposure, but S-ECC also occurs in children without that dietary pattern. This may indicate that S-ECC children harbor a distinct cariogenic microbiota. 16S rRNA studies indicate which taxa are present in the microbiota, but do not provide information on its collective biochemical activity. Quantitative metaproteomics can provide expression-based information on the metabolic activity of microbial biofilms. However, it is difficult to obtain sufficient protein for deep proteomic analysis from dental plaque. This project will address that problem by using a published validated biofilm reactor model to grow complex oral microcosm biofilms representative of the supragingival plaque microbiota from individual subjects. 

Preliminary analysis of microcosms grown in the presence and absence of sucrose revealed major, subject-specific, differences in taxonomic diversity and protein expression patterns. This demonstrates that that the microcosm approach to metaproteomics is both feasible and informative, making possible deep quantitative metaproteomic comparisons of oral microcosms grown from individual S-ECC and caries-free (CF) children. State-of-the-art proteomic and bioinformatic workflows will be used to test three alternative (but not mutually exclusive) hypotheses. The sucrose selection hypothesis predicts that sucrose-induced changes in community composition and protein expression patterns will be fundamentally similar between the S-ECC and CF groups. That would support a primary role for sucrose in the etiology of S-ECC. The phenotypic variation hypothesis predicts that differences in sucrose response patterns between the S-ECC and CF groups will be greater for protein expression than for community composition. This would suggest that S-ECC children harbor highly acidogenic and aciduric strains of common species. The keystone pathogen hypothesis predicts that differences in sucrose response patterns between the S-ECC and CF groups will reflect differences in the relative abundance of virulence factors produced by Streptococcus mutans, and other putative pathogens. This would suggest that the S-ECC children are infected by a distinct cariogenic microbiota. Each outcome will have different implications for risk assessment, prevention, and treatment. 

This project has three main aims: refine current MS methods for deep metaproteomic analysis of oral microcosm biofilms to further increase the already large number of distinct microbial proteins/peptides that can be identified; work with MSI to refine the informatics workflow for analysis of large-scale metaproteomic data to further enhance identification/quantification of microbial peptides/proteins; hypothesis testing via metaproteomic comparison of microcosms from 35 S-ECC and 35 CF children. Microcosms will be grown in the presence and absence of sucrose, to determine whether metaproteomic results best fit the sucrose selection, phenotypic variation, or keystone pathogen hypotheses. 

A bibliography of this group’s publications acknowledging MSI is attached.