IADR Abstract Archives
Culture of Eight Saccharibacteria (TM7) Bacteria in Coculture With Actinobacteria
Objectives: The Candidate Phyla Radiation (CPR) group of bacteria comprises a large component of microbial dark matter on earth, bacterial phyla with no cultured representatives. The He laboratory (UCLA) recently cultured the first CPR bacterium, TM7x. They reported that TM7x grew as a parasitic bacterium on Actinomyces odontolyticus. The small genome sizes of known CPR bacteria suggest that they may share TM7x’s parasitic lifestyle. The goal of this research was to develop a reliable means for isolating other oral TM7s in coculture with their hosts.
Methods: The oral microbiomes from 9 oral niches in 35 subjects was examined to determine which oral sites and subjects possessed the highest abundance of TM7 bacteria. Samples from these subjects/sites were collected by either scaler or nylon brush for cultivation attempts. Because of the known small CPR organism size, samples were dispersed in saline and passed through a 0.2 μm nucleopore membrane to remove other bacteria. The filtrate containing TM7 bacteria was inoculated into broth cultures of several species from genera in the phylum Actinobacteria and passaged. Genomes were obtained by PacBio sequencing.
Results: Of six potential host species tested, eight independent strains representing several species of TM7 bacteria were found to grow on Pseudopropionibacterium propionicum. The eight binary cultures were found to passage readily at 37°C in TSBY/RPMI (50/50 v/v) medium under aerobic, microaerophilic or anaerobic conditions. Closed genomes for four TM7s were obtained and four are in progress. These genomes share significant synteny with other oral and environmental TM7s and show substantial loss of synthetic pathways leading to extreme auxotrophy.
Conclusions: Reliable methods have been identified for isolating TM7 bacteria in coculture with a suitable host. The genome sizes <0.9 MB shows that these TM7s have reduced genomes common to other parasitic bacteria.
Methods: The oral microbiomes from 9 oral niches in 35 subjects was examined to determine which oral sites and subjects possessed the highest abundance of TM7 bacteria. Samples from these subjects/sites were collected by either scaler or nylon brush for cultivation attempts. Because of the known small CPR organism size, samples were dispersed in saline and passed through a 0.2 μm nucleopore membrane to remove other bacteria. The filtrate containing TM7 bacteria was inoculated into broth cultures of several species from genera in the phylum Actinobacteria and passaged. Genomes were obtained by PacBio sequencing.
Results: Of six potential host species tested, eight independent strains representing several species of TM7 bacteria were found to grow on Pseudopropionibacterium propionicum. The eight binary cultures were found to passage readily at 37°C in TSBY/RPMI (50/50 v/v) medium under aerobic, microaerophilic or anaerobic conditions. Closed genomes for four TM7s were obtained and four are in progress. These genomes share significant synteny with other oral and environmental TM7s and show substantial loss of synthetic pathways leading to extreme auxotrophy.
Conclusions: Reliable methods have been identified for isolating TM7 bacteria in coculture with a suitable host. The genome sizes <0.9 MB shows that these TM7s have reduced genomes common to other parasitic bacteria.