analysis of cultivation techniques for the isolation of novel oral microorganisms; aerobic vs. anaerobic

• Online Access: http://hdl.handle.net/2047/d20002596

According to the American Society for Microbiology, the human body contains nearly ten times as many bacterial cells as somatic cells, comprised of thousands of different species, yet less than one percent has been cultivated in the laboratory setting. rRNA survey studies attribute over 700 different species to the oral microbiome, yet estimate less than a third have been isolated and characterized in vivo. This knowledge gap is dangerous to the study of human health, as many of the uncultivable organisms have deleterious effects on human health, from periodontitis to systemic cardiac issues. In order to increase the number of cultivable oral isolates, and build a complete microbiome library, novel approaches must be examined and optimized, including anaerobic cultivation which has demonstrated surprising preliminary success. This experiment seeks to quantitatively establish the high isolate yield potential of anaerobic cultivation, and serve as an academic basis and impetus for further research. After growing a single subgingival plaque sample on plates both aerobically and anaerobically, we sequenced and analyzed individual colonies to generate libraries of isolates. These data provided qualitative and quantitative evidence strongly reinforcing the potential of anaerobic growth techniques to decrease the quantity of uncultivable organisms within the oral microbiome. Species isolation consistency within a single condition remained above 50% for both growth states, yet the overlap of microorganisms was only 13.2%. These results clearly demonstrate the variation in cultivation results between aerobic and anaerobic laboratory environments, and strongly encourage future research to pursue anaerobic methods of isolation in order to ultimately complete the list of human microorganisms. To conclude, although this data does not suggest abandoning any technique, it provides numerical support to investigators seeking varied isolate yields in order to finally close the dangerous knowledge gap.