Presentation Title

Retrieving Novel Digestive Genes from an Uncultured Bacterial Phylum Found in the Human Oral Microbiome Found in Association with Periodontitis

Faculty Mentor

Cleber Ouverney

Start Date

23-11-2019 1:15 PM

End Date

23-11-2019 1:30 PM

Location

Markstein 205

Session

oral 3

Type of Presentation

Oral Talk

Subject Area

biological_agricultural_sciences

Abstract

This project aimed to understand the role that bacteria in the uncultured phylum, Candidatus Saccharibacteria (aka TM7), play in the human oral cavity. Some TM7 phylotypes have been associated with periodontitis, which affects nearly 18% of the adult population in developed nations,which when left untreated can lead to systemic complications. General understanding of these phylotypes could improve with axenic cultures as cultivation requires an understanding of the bacteria’s metabolism. It is unknown what proportion of the TM7 phylum contains a beta-galactosidase (β-gal) gene, but we’ve recently found one which matched to a TM7 genome. Our methodology took an approach to characterize this gene and homologs in the TM7 community of human oral plaque. We measured the diversity of TM7 β-gal genes in human oral plaque using polymerase chain reaction (PCR) followed by DNA sequencing. Sequences would then be tested for sequence similarity to sequenced TM7 genomes. We hypothesized that β-gal genes would be elusive in screened oral plaque samples. Genomic DNA was extracted from 144 oral plaque samples from about 70 healthy adult individuals and screened for β-gal gene using PCR-specific primers. Eighteen samples showed an amplicon from gel electrophoresis where sizes ranged from 500bp to 1,600 bp. Amplicons were cleaned (QIAprep kit protocol) and Sanger sequenced. Sequences were cleaned and contigs were analyzed for identity via BLAST and for similarity via DNA multi-sequence alignment. A phylogenetic tree was built to determine the evolutionary history of the sequences compared to original TM7 lactase and 40 reference sequences. Two of the PCR amplicons sequenced clustered with a TM7 genome from a human oral sample containing an alpha-glucosidase gene (accession number WP_146529164) at 40% similarity. These findings lead us to think that the TM7 phylum possibly has a diverse set of digestive enzymes to utilize saccharides within their environment.

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Nov 23rd, 1:15 PM Nov 23rd, 1:30 PM

Retrieving Novel Digestive Genes from an Uncultured Bacterial Phylum Found in the Human Oral Microbiome Found in Association with Periodontitis

Markstein 205

This project aimed to understand the role that bacteria in the uncultured phylum, Candidatus Saccharibacteria (aka TM7), play in the human oral cavity. Some TM7 phylotypes have been associated with periodontitis, which affects nearly 18% of the adult population in developed nations,which when left untreated can lead to systemic complications. General understanding of these phylotypes could improve with axenic cultures as cultivation requires an understanding of the bacteria’s metabolism. It is unknown what proportion of the TM7 phylum contains a beta-galactosidase (β-gal) gene, but we’ve recently found one which matched to a TM7 genome. Our methodology took an approach to characterize this gene and homologs in the TM7 community of human oral plaque. We measured the diversity of TM7 β-gal genes in human oral plaque using polymerase chain reaction (PCR) followed by DNA sequencing. Sequences would then be tested for sequence similarity to sequenced TM7 genomes. We hypothesized that β-gal genes would be elusive in screened oral plaque samples. Genomic DNA was extracted from 144 oral plaque samples from about 70 healthy adult individuals and screened for β-gal gene using PCR-specific primers. Eighteen samples showed an amplicon from gel electrophoresis where sizes ranged from 500bp to 1,600 bp. Amplicons were cleaned (QIAprep kit protocol) and Sanger sequenced. Sequences were cleaned and contigs were analyzed for identity via BLAST and for similarity via DNA multi-sequence alignment. A phylogenetic tree was built to determine the evolutionary history of the sequences compared to original TM7 lactase and 40 reference sequences. Two of the PCR amplicons sequenced clustered with a TM7 genome from a human oral sample containing an alpha-glucosidase gene (accession number WP_146529164) at 40% similarity. These findings lead us to think that the TM7 phylum possibly has a diverse set of digestive enzymes to utilize saccharides within their environment.