Skip to main content
U.S. Department of Energy
Office of Scientific and Technical Information

Targeted isolation and cultivation of uncultivated bacteria by reverse genomics

Journal Article · · Nature Biotechnology
 [1];  [2];  [3];  [2];  [1];  [4];  [5];  [3];  [3];  [4];  [3];  [1];  [1]
  1. Oak Ridge National Lab. (ORNL), Oak Ridge, TN (United States); Univ. of Tennessee, Knoxville, TN (United States)
  2. Oak Ridge National Lab. (ORNL), Oak Ridge, TN (United States); Northwest Missouri State Univ., Maryville, MO (United States)
  3. Oak Ridge National Lab. (ORNL), Oak Ridge, TN (United States)
  4. The Ohio State Univ., Columbus, OH (United States)
  5. Knoxville Periodontics, Knoxville, TN (United States)

The majority of microorganisms from all taxonomic levels are uncultured. Single-cell genomes and metagenomes continue to increase the known diversity of Bacteria and Archaea; however, while ’omics can be used to infer physiological or ecological roles for species in a community, most of these hypothetical roles remain unvalidated. Here, we report a technique to capture specific microorganisms from complex communities into pure cultures using genome-informed antibody engineering. We apply our reverse genomics approach to isolate and sequence single cells and to cultivate three different species-level lineages of human oral Saccharibacteria (TM7). Using our pure cultures, we show that all three Saccharibacteria species are epibionts of diverse Actinobacteria. We also isolate and cultivate human oral SR1 bacteria, which are members of a lineage of previously uncultured bacteria. Reverse-genomics-enabled cultivation of microorganisms can be applied to any species from any environment and has the potential to unlock the isolation, cultivation and characterization of species from as-yet-uncultured branches of the microbial tree of life.

Research Organization:
Oak Ridge National Laboratory (ORNL), Oak Ridge, TN (United States)
Sponsoring Organization:
USDOE; National Institute of Health (NIH); National Science Foundation (NSF)
Grant/Contract Number:
AC05-00OR22725
OSTI ID:
1570133
Journal Information:
Nature Biotechnology, Journal Name: Nature Biotechnology Journal Issue: 11 Vol. 37; ISSN 1087-0156
Publisher:
Springer NatureCopyright Statement
Country of Publication:
United States
Language:
English

References (94)

Improved recognition of native-like protein structures using a combination of sequence-dependent and sequence-independent features of proteins journal January 1999
Prediction of continuous B-cell epitopes in an antigen using recurrent neural network journal August 2006
Learning generative models for protein fold families
  • Balakrishnan, Sivaraman; Kamisetty, Hetunandan; Carbonell, Jaime G.
  • Proteins: Structure, Function, and Bioinformatics, Vol. 79, Issue 4 https://doi.org/10.1002/prot.22934
journal January 2011
Predicting transmembrane protein topology with a hidden markov model: application to complete genomes11Edited by F. Cohen journal January 2001
Protein Data Bank (PDB): The Single Global Macromolecular Structure Archive book January 2017
Complex Evolution of the Mismatch Repair System in Eukaryotes is Illuminated by Novel Archaeal Genomes journal January 2021
Phenotypic and Physiological Characterization of the Epibiotic Interaction Between TM7x and Its Basibiont Actinomyces journal November 2015
A comparison of human dental plaque microcosm biofilms grown in an undefined medium and a chemically defined artificial saliva journal June 2001
Siderophores from Neighboring Organisms Promote the Growth of Uncultured Bacteria journal March 2010
Novel Microbial Diversity and Functional Potential in the Marine Mammal Oral Microbiome journal December 2017
The phenomenon of microbial uncultivability journal October 2013
A novel recombinant vaccine candidate comprising PBP2a and autolysin against Methicillin Resistant Staphylococcus aureus confers protection in the experimental mice journal November 2017
A new symbiotic nanoarchaeote (Candidatus Nanoclepta minutus) and its host (Zestosphaera tikiterensis gen. nov., sp. nov.) from a New Zealand hot spring journal January 2019
Sequence-structure relationships in polysaccharide co-polymerase (PCP) proteins journal February 2009
The feline oral microbiome: A provisional 16S rRNA gene based taxonomy with full-length reference sequences journal February 2015
Distinctive Attributes of β-Lactam Target Proteins in Acinetobacter baumannii Relevant to Development of New Antibiotics journal December 2011
A new phylum of Archaea represented by a nanosized hyperthermophilic symbiont journal May 2002
Insights into the phylogeny and coding potential of microbial dark matter journal July 2013
Unusual biology across a group comprising more than 15% of domain Bacteria journal June 2015
Culturing of ‘unculturable’ human microbiota reveals novel taxa and extensive sporulation journal May 2016
Asgard archaea illuminate the origin of eukaryotic cellular complexity journal January 2017
Genome sequences of rare, uncultured bacteria obtained by differential coverage binning of multiple metagenomes journal May 2013
Genomics-informed isolation and characterization of a symbiotic Nanoarchaeota system from a terrestrial geothermal environment journal July 2016
Thousands of microbial genomes shed light on interconnected biogeochemical processes in an aquifer system journal October 2016
Harnessing the landscape of microbial culture media to predict new organism–media pairings journal October 2015
HHblits: lightning-fast iterative protein sequence searching by HMM-HMM alignment journal December 2011
Practical innovations for high-throughput amplicon sequencing journal September 2013
QIIME allows analysis of high-throughput community sequencing data journal April 2010
Culture of previously uncultured members of the human gut microbiota by culturomics journal November 2016
A new view of the tree of life journal April 2016
Obtaining genomes from uncultivated environmental microorganisms using FACS–based single-cell genomics journal April 2014
In situ cultivation of previously uncultivable microorganisms using the ichip journal September 2017
Recovery of nearly 8,000 metagenome-assembled genomes substantially expands the tree of life journal September 2017
GABA-modulating bacteria of the human gut microbiota journal December 2018
Biosynthetic capacity, metabolic variety and unusual biology in the CPR and DPANN radiations journal September 2018
Crystal structure of a peptidoglycan glycosyltransferase suggests a model for processive glycan chain synthesis journal March 2007
Dissecting biological "dark matter" with single-cell genetic analysis of rare and uncultivated TM7 microbes from the human mouth journal July 2007
Crystal structure of the membrane-bound bifunctional transglycosylase PBP1b from Escherichia coli journal May 2009
Crystal structure of Staphylococcus aureus transglycosylase in complex with a lipid II analog and elucidation of peptidoglycan synthesis mechanism journal April 2012
UGA is an additional glycine codon in uncultured SR1 bacteria from the human microbiota journal March 2013
Assessing the utility of coevolution-based residue-residue contact predictions in a sequence- and structure-rich era journal September 2013
Cultivation of a human-associated TM7 phylotype reveals a reduced genome and epibiotic parasitic lifestyle journal December 2014
Biogeography of a human oral microbiome at the micron scale journal January 2016
Microfluidic droplet platform for ultrahigh-throughput single-cell screening of biodiversity journal February 2017
Nonlinear partial differential equations and applications: Cultivating the uncultured journal November 2002
Streptococcus agalactiae Capsule Polymer Length and Attachment Is Determined by the Proteins CpsABCD journal February 2015
The multi-omics promise in context: from sequence to microbial isolate journal May 2017
Environmental Diversity of Bacteria and Archaea journal August 2001
Environmental Diversity of Bacteria and Archaea journal August 2001
SPAdes: A New Genome Assembly Algorithm and Its Applications to Single-Cell Sequencing journal May 2012
Search and clustering orders of magnitude faster than BLAST journal August 2010
Geneious Basic: An integrated and extendable desktop software platform for the organization and analysis of sequence data journal April 2012
InterProScan 5: genome-scale protein function classification journal January 2014
Prokka: rapid prokaryotic genome annotation journal March 2014
Immunogenicity of meningococcal PBP2 during natural infection and protective activity of anti-PBP2 antibodies against meningococcal bacteraemia in mice journal March 2006
The integrated microbial genomes (IMG) system journal January 2006
Pfam: the protein families database journal November 2013
eggNOG 4.5: a hierarchical orthology framework with improved functional annotations for eukaryotic, prokaryotic and viral sequences journal November 2015
Intestinal TM7 bacterial phylogenies in active inflammatory bowel disease journal December 2008
CheckM: assessing the quality of microbial genomes recovered from isolates, single cells, and metagenomes journal May 2015
Diversity and genomic insights into the uncultured C hloroflexi from the human microbiota : Uncultured human-associated journal April 2014
Critical biogeochemical functions in the subsurface are associated with bacteria from new phyla and little studied lineages: N- and C-cycling organisms in the subsurface journal July 2015
The penicillin-binding proteins: structure and role in peptidoglycan biosynthesis journal March 2008
The penicillin-binding proteins: structure and role in peptidoglycan biosynthesis journal May 2008
Recent findings on the viable but nonculturable state in pathogenic bacteria journal July 2010
Structural Insight into the Transglycosylation Step of Bacterial Cell-Wall Biosynthesis journal March 2007
Protein structure determination using metagenome sequence data journal January 2017
Interbacterial Adhesion Networks within Early Oral Biofilms of Single Human Hosts journal March 2017
Axenic Culture of a Candidate Division TM7 Bacterium from the Human Oral Cavity and Biofilm Interactions with Other Oral Bacteria journal August 2014
Targeted Access to the Genomes of Low-Abundance Organisms in Complex Microbial Communities journal March 2007
New Approaches for Isolation of Previously Uncultivated Oral Bacteria journal November 2011
Prevalence of Bacteria of Division TM7 in Human Subgingival Plaque and Their Association with Disease journal March 2003
Microcolony Cultivation on a Soil Substrate Membrane System Selects for Previously Uncultured Soil Bacteria journal December 2005
Identification of Streptococcus pneumoniae Cps2C Residues That Affect Capsular Polysaccharide Polymerization, Cell Wall Ligation, and Cps2D Phosphorylation journal March 2011
The Human Oral Microbiome journal July 2010
Impact of Culture-Independent Studies on the Emerging Phylogenetic View of Bacterial Diversity journal September 1998
Molecular Detection of Cellulosimicrobium cellulans as the Etiological Agent of a Chronic Tongue Ulcer in a Human Immunodeficiency Virus-Positive Patient journal August 2005
Insights into the Evolution of Host Association through the Isolation and Characterization of a Novel Human Periodontal Pathobiont, Desulfobulbus oralis journal March 2018
Draft Genome Sequence of Comamonas aquatilis Strain LK (= CSUR P6418 = CECT 9772), Isolated from the Planarian Schmidtea mediterranea journal February 2021
Present and Future of Culturing Bacteria journal September 2017
Hidden Markov model speed heuristic and iterative HMM search procedure journal August 2010
Establishment and metabolic analysis of a model microbial community for understanding trophic and electron accepting interactions of subsurface anaerobic environments journal January 2010
Insights into archaeal evolution and symbiosis from the genomes of a nanoarchaeon and its inferred crenarchaeal host from Obsidian Pool, Yellowstone National Park journal April 2013
Fine-scale population structure and evidence for local adaptation in Australian giant black tiger shrimp (Penaeus monodon) using SNP analysis journal September 2020
Generalized Fragment Picking in Rosetta: Design, Protocols and Applications journal August 2011
The Canine Oral Microbiome journal April 2012
Multiple Single-Cell Genomes Provide Insight into Functions of Uncultured Deltaproteobacteria in the Human Oral Cavity journal March 2013
In Vitro Cultivation of ‘Unculturable’ Oral Bacteria, Facilitated by Community Culture and Media Supplementation with Siderophores journal January 2016
Cutadapt removes adapter sequences from high-throughput sequencing reads journal May 2011
A Complex Endomembrane System in the Archaeon Ignicoccus hospitalis Tapped by Nanoarchaeum equitans journal June 2017
Isolation of Uncultured Bacteria from Antarctica Using Long Incubation Periods and Low Nutritional Media journal July 2017
eggNOG 4.5: a hierarchical orthology framework with improved functional annotations for eukaryotic, prokaryotic and viral sequences text January 2016
Anvi’o: an advanced analysis and visualization platform for ‘omics data journal January 2015
Linking pangenomes and metagenomes: the Prochlorococcus metapangenome journal January 2018

Cited By (10)

Molecules Produced by Probiotics and Intestinal Microorganisms with Immunomodulatory Activity journal February 2020
Additional file 1 of Cultivation of common bacterial species and strains from human skin, oral, and gut microbiota dataset January 2021
Additional file 3 of Exploring Co-occurrence patterns and microbial diversity in the lung microbiome of patients with non-small cell lung cancer image January 2023
Additional file 4 of Exploring Co-occurrence patterns and microbial diversity in the lung microbiome of patients with non-small cell lung cancer dataset January 2023
Additional file 1 of Genomic representativeness and chimerism in large collections of SAGs and MAGs of marine prokaryoplankton image January 2024
Next-generation physiology approaches to study microbiome function at single cell level journal February 2020
Culturing the uncultured journal October 2019
Genome‐inferred spatio‐temporal resolution of an uncultivated Roizmanbacterium reveals its ecological preferences in groundwater journal December 2019
Methane oxidation and methylotroph population dynamics in groundwater mesocosms journal April 2020
Insights Obtained by Culturing Saccharibacteria With Their Bacterial Hosts journal February 2020

Similar Records

Single-Cell Genomics and the Oral Microbiome
Journal Article · Sun Feb 23 23:00:00 EST 2020 · Journal of Dental Research · OSTI ID:1606754

Dissecting Biological Dark Matter: Single Cell Genetic Analysis of TM7, a Rare and Uncultivated Microbe from the Human Mouth
Journal Article · Sun Jul 01 00:00:00 EDT 2007 · PNAS · OSTI ID:934712

Related Subjects