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Isolation of previously uncultured rumen bacteria by dilution to extinction using a new liquid culture medium
Abstract
A new anaerobic medium that mimics the salts composition of rumen fluid was used in conjunction with a dilution method of liquid culture to isolate fermentative bacteria from the rumen of a grass-fed sheep. The aim was to inoculate a large number of culture tubes each with a mean of <1 culturable cell, which should maximize the number of cultures that develop from a single bacterium. This minimizes the effort that has to be put into purifying the resultant cultures. Of 1000 tubes, 139 were growth positive. Of the 93 that were able to be subcultured, 54 (58%) appeared to be pure cultures. The phylogenetic placements of these 54 cultures, together with another 6 cultures obtained from a preliminary study, were determined. Using a criterion of <93% 16S rRNA gene sequence identity to a previously named bacterium as a proxy for defining a new genus, 27 (45%) of the 60 cultures belonged to 14 potentially novel genera. Many of these had 16S rRNA genes that shared >97% sequence identity to genes of uncultured bacteria detected in various gastrointestinal environments. This strategy has therefore allowed us to cultivate many novel rumen bacteria, opening the way to overcoming the lack of cultures of many of the groups detected using cultivation-independent methods.
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“Our ability to culture rumen bacteria has improved in recent years through the development of culture media (Kenters et al., 2011).”
From Addressing global ruminant agricultural challenges through understanding the rumen microbiome: past, present, and future (2018) · cites this paper as Kenters et al., 2011
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“Techniques such as dilution to extinction have also improved our ability to culture bacteria in many ecosystems, including the rumen (Kenters et al., 2011).”
From Addressing global ruminant agricultural challenges through understanding the rumen microbiome: past, present, and future (2018) · cites this paper as Kenters et al., 2011
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Metadata
- Authors
- Kenters, N., Henderson, G., Jeyanathan, J., Kittelmann, S., & Janssen, P. H.
- Affiliations
- AgResearch (NZ)
- Corresponding author
- Janssen, Peter H.
- Venue
- Journal of Microbiological Methods
- Publisher
- Elsevier BV
- Published
- 2011-01
- Volume
- 84
- Issue
- 1
- Pages
- 52-60
- ISSN
- 0167-7012 (ISSN-L)
- OpenAlex
- W1965203437
- Cited by
- 108
- References
- 57
- Citations by year
- 2026: 3 · 2025: 6 · 2024: 8 · 2023: 9 · 2022: 5 · 2021: 9 · 2020: 8 · 2019: 8 · 2018: 6 · 2017: 11 · 2016: 5 · 2015: 8 · 2014: 10 · 2013: 5 · 2012: 7
- FWCI
- 8.12
- Type
- journal article
- Language
- en
- Open access
- closed
- Locations
- Journal of Microbiological Methods (publishedVersion)
- PubMed (publishedVersion)
- Ghent University Academic Bibliography (Ghent University) (submittedVersion)
- Funders
- Ministry of Agriculture and Forestry, New Zealand
- New Zealand Foundation for Research Science and Technology's
- Topics
- Life Sciences › Agricultural and Biological Sciences › Animal Science and Zoology › Ruminant Nutrition and Digestive Physiology
- Physical Sciences › Environmental Science › Ecology › Microbial Community Ecology and Physiology
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Gut microbiota and health
- Keywords
- rumen bacteria, uncultured bacteria, dilution-to-extinction, rumen fluid, sheep, novel bacterial genus, 16S rRNA phylogeny, bacterial isolation, rumen fermentation
- MeSH terms
- Anaerobiosis, Animals, Bacteria, Bacteriological Techniques, Culture Media, DNA, Bacterial, DNA, Ribosomal, Molecular Sequence Data, Phylogeny, RNA, Ribosomal, 16S, Rumen, Sheep, Cluster Analysis, Sequence Analysis, DNA
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