Microbial culturomics: paradigm shift in the human gut microbiome study. Clin Microbiol Infect

Aix Marseille Université, URMITE, UM63, CNRS 7278, IRD 198, INSERM 1095  Service de Nutrition, Maladies Métaboliques et Endocrinologie, UMR-INRA U1260, CHU de la Timone, Marseille, France  IRD, UMR CNRS 7278-IRD 198, Route des Pères Maristes, Dakar, Sénégal  National Centre for Biotechnology Information, National Library of Medicine, National Institutes of Health, Bethesda, MD, USA.
Clinical Microbiology and Infection (Impact Factor: 5.77). 09/2012; 449(12). DOI: 10.1111/1469-0691.12023
Source: PubMed

ABSTRACT Clin Microbiol Infect 2012; 18: 1185–1193
Comprehensive determination of the microbial composition of the gut microbiota and the relationships with health and disease are major challenges in the 21st century. Metagenomic analysis of the human gut microbiota detects mostly uncultured bacteria. We studied stools from two lean Africans and one obese European, using 212 different culture conditions (microbial culturomics), and tested the colonies by using mass spectrometry and 16S rRNA amplification and sequencing. In parallel, we analysed the same three samples by pyrosequencing 16S rRNA amplicons targeting the V6 region. The 32 500 colonies obtained by culturomics have yielded 340 species of bacteria from seven phyla and 117 genera, including two species from rare phyla (Deinococcus-Thermus and Synergistetes, five fungi, and a giant virus (Senegalvirus). The microbiome identified by culturomics included 174 species never described previously in the human gut, including 31 new species and genera for which the genomes were sequenced, generating c. 10 000 new unknown genes (ORFans), which will help in future molecular studies. Among these, the new species Microvirga massiliensis has the largest bacterial genome so far obtained from a human, and Senegalvirus is the largest virus reported in the human gut. Concurrent metagenomic analysis of the same samples produced 698 phylotypes, including 282 known species, 51 of which overlapped with the microbiome identified by culturomics. Thus, culturomics complements metagenomics by overcoming the depth bias inherent in metagenomic approaches.

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    • "The biggest difference among the microorganisms with regard to rigidity is the grampositive or gram-negative structure of the cell wall. At least half of the gut-residing bacteria are gram-positive (Gossling and Slack 1974; Lagier, Million et al. 2012), and the commonly hardto-lyse domain of the Archaea is represented predominantly by methanogenic Methanobrevibacter spp. with a variable and overall low prevalence (Hoffmann et al. 2013). Moreover, the microbiota comprises taxa containing aerobic and anaerobic bacteria belonging to the genera of Bacillus, Clostridium, and others within the Firmicutes, potentially forming endospores, which are probably the most robust cellular structures known. "
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    ABSTRACT: Analyzing human as well as animal microbiota composition has gained growing interest because structural components and metabolites of microorganisms fundamentally influence all aspects of host physiology. Originally dominated by culture-dependent methods for exploring these ecosystems, the development of molecular techniques such as high throughput sequencing has dramatically increased our knowledge. Because many studies of the microbiota are based on the bacterial 16S ribosomal RNA (rRNA) gene targets, they can, at least in principle, be compared to determine the role of the microbiome composition for developmental processes, host metabolism, and physiology as well as different diseases. In our review, we will summarize differences and pitfalls in current experimental protocols, including all steps from nucleic acid extraction to bioinformatical analysis which may produce variation that outweighs subtle biological differences. Future developments, such as integration of metabolomic, transcriptomic, and metagenomic data sets and standardization of the procedures, will be discussed. © The Author 2015. Published by Oxford University Press on behalf of the Institute for Laboratory Animal Research. All rights reserved. For permissions, please email:
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    • "The aerobic bacteria tested were Escherichia coli CSUR P729, Staphylococcus aureus CSUR P749, Staphylococcus epidermidis CSUR P748, Klebsiella pneumoniae CSUR P931 and Pseudomonas aeruginosa CSUR P1003. All of the strains tested were isolated in the laboratory as part of a culturomics study as previously described [7] [8]. Aerobic and anaerobic bacterial strains were routinely subcultured using 5% sheep blood agar medium (bioMérieux, Marcy-l'Étoile, France). "
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    • "Comment étudier le résistome ? Si la culture des bactéries du microbiote intestinal demeure possible mais difficile [10], les méthodes s'affranchissant de la culture et particulièrement celles basées sur la métagénomique ont réellement permis de démarrer l'exploration du résistome intestinal. La métagénomique dite « classique » (ou shotgun metagenomics) consiste à séquencer directement , sans spécificité, un extrait d'ADN d'un échantillon de selles. "
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