Methanobrevibacter olleyae YLM1: Difference between revisions
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[[File:Puromycin MIC determination in M. olleyae YLM1. Source- Dr. Nana Shao.jpg|thumb|Puromycin MIC determination in ''M. olleyae'' YLM1. Source: Dr. Nana Shao]]'''Solid Medium''' | [[File:Puromycin MIC determination in M. olleyae YLM1. Source- Dr. Nana Shao.jpg|thumb|Puromycin MIC determination in ''M. olleyae'' YLM1. Source: Dr. Nana Shao]]'''Solid Medium''' | ||
Puromycin susceptibility of ''M.'' ''olleyae'' YLM1 was assessed on solid medium under anaerobic conditions, using the absence of visible colonies as the endpoint. The apparent MIC was 10 µg/mL at an inoculum of | Puromycin susceptibility of ''M.'' ''olleyae'' YLM1 was assessed on solid medium under anaerobic conditions, using the absence of visible colonies as the endpoint. The apparent MIC was 10 µg/mL at an inoculum of 10<sup>5</sup> cells/plate and 15 µg/mL at 10<sup>7</sup> cells/plate. | ||
'''Liquid Medium''' | '''Liquid Medium''' | ||
Puromycin and nourseothricin susceptibility was assessed in liquid MMSM at initial inoculum densities of | Puromycin and nourseothricin susceptibility was assessed in liquid MMSM at initial inoculum densities of 8×10<sup>5</sup> and 2×10<sup>7</sup> cells/mL, using the absence of a detectable increase in OD600 throughout the observation period as the endpoint. | ||
{| class="wikitable" | {| class="wikitable" | ||
|+ | |+ | ||
!Antibiotics | !Antibiotics | ||
|MIC at | |MIC at 8×10<sup>5</sup> cells/mL | ||
|MIC at | |MIC at 2×10<sup>7</sup> cells/mL | ||
|Observation period | |Observation period | ||
|- | |- | ||
Revision as of 19:53, 23 October 2026
Introduction
Methanobrevibacter olleyae YLM1 was a hydrogenotrophic methanogen isolated from the rumen of a lamb[1]. Unlike Methanobrevibacter ruminantium M1[2], YLM1 carries comADE genes supporting its predicted ability to synthesize coenzyme M. It also possesses the mrtBDGA operon encoding methyl-coenzyme M reductase II, which is absent from M1 and may contribute to differences in growth under varying hydrogen concentrations[3]. These genomic features make YLM1 a useful strain for comparative studies of rumen methanogen physiology and the development of methane mitigation strategies. Its robust growth and high plating efficiency, observed in the Whitman Lab, make it particularly valuable for research supported by the Global Methane Hub.
Global Methane Hub Metadata
| Origin: | Lucidome Bio, NZ |
| Sequenced by: | Plasmidsaurus, ONT + Illumina |
| Assembled by: | Nana Shao |
| Methylome called by: | William G. Alexander |
| Methyltransferases cloned by: | Roy Rodriguez Carrero |
| MIC and Culturing conditions by: | Nana Shao |
| Transformation by: |
Genome
The draft genome of Methanobrevibacter olleyae YLM1 is 2,201,192 bp, with a GC content of 26.9% and 1,834 predicted protein-coding genes[3]. It contains one approximately 40-kb prophage, two CRISPR regions, and no identified plasmids. Notable features include the hdrABC gene encoding CoB-CoM heterodisulfide reductase subunits, the comADE genes involved in coenzyme M biosynthesis, the mrtBDGA operon encoding methyl-coenzyme M reductase II, and 64 large adhesin-like protein genes. The GenBank accession number is CP014265.
DefenseFinder Results
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Methylome
Pending Roy's input.
- Searching dataset for following motifs: GSNGA, GNSNGA
- Motifs explain the following about the (6mA) modified dataset:
- Low-quality methylated sites: 7170/6856 explained (104.57992998833139%)
- Mid-quality methylated sites: 7158/6524 explained (109.71796443899449%)
- High-quality methylated sites: 300/244 explained (122.95081967213115%)
- No 5mC motifs present in output file, skipping...
- No 4mC motifs present in output file, skipping...
- Only one replicon present, skipping differential genomic coverage
Methylating E. coli
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Culturing Conditions
M. olleyae YLM1 is cultivated in Modified Methanobrevibacter smithii and Methanosphaera stadtmanae medium (MMSM) at 37°C. To ensure good gas transfer to the media, tubes are incubated on their sides (or at a shallow angle so that the media does not contact the stoppers) with gentle shaking at 50 rpm and 207 kPa of H2/CO2 (80:20, v/v) as an energy substrate for 2-4 days. This methanogen can also be cultivated on MMSM plates using 1.5% agar as a gelling agent with the same pressure of H2/CO2 as an energy substrate for 3-7 days.
Antimicrobial MIC Tests
Solid Medium
Puromycin susceptibility of M. olleyae YLM1 was assessed on solid medium under anaerobic conditions, using the absence of visible colonies as the endpoint. The apparent MIC was 10 µg/mL at an inoculum of 105 cells/plate and 15 µg/mL at 107 cells/plate.
Liquid Medium
Puromycin and nourseothricin susceptibility was assessed in liquid MMSM at initial inoculum densities of 8×105 and 2×107 cells/mL, using the absence of a detectable increase in OD600 throughout the observation period as the endpoint.
| Antibiotics | MIC at 8×105 cells/mL | MIC at 2×107 cells/mL | Observation period |
|---|---|---|---|
| Puromycin | 10 µg/mL | 20 µg/mL | ~700 h |
| nourseothricin | 40 µg/mL | 70 µg/mL | ~630 h |
Overall, the results demonstrated inoculum-dependent inhibition under the tested conditions, with higher apparent MICs at higher inoculum densities in both solid and liquid media.
| Antibiotics | Concentration | |
|---|---|---|
| Liquid | Solid | |
| Puromycin | ||
| nourseothricin | μM | |
Protocols
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Transformation Results
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External Links
- ↑ Skillman LC, Evans PN, Naylor GE, Morvan B, Jarvis GN, Joblin KN. 16S ribosomal DNA-directed PCR primers for ruminal methanogens and identification of methanogens colonising young lambs. Anaerobe. 2004 Oct;10(5):277-85. doi: 10.1016/j.anaerobe.2004.05.003.
- ↑ Leahy SC, Kelly WJ, Altermann E, Ronimus RS, Yeoman CJ, Pacheco DM, Li D, Kong Z, McTavish S, Sang C, Lambie SC, Janssen PH, Dey D, Attwood GT. The genome sequence of the rumen methanogen Methanobrevibacter ruminantium reveals new possibilities for controlling ruminant methane emissions. PLoS One. 2010 Jan 28;5(1):e8926. doi: 10.1371/journal.pone.0008926.
- ↑ 3.0 3.1 Kelly WJ, Li D, Lambie SC, Cox F, Attwood GT, Altermann E, Leahy SC. Draft Genome Sequence of the Rumen Methanogen Methanobrevibacter olleyae YLM1. Genome Announc. 2016 Apr 7;4(2):e00232-16. doi: 10.1128/genomeA.00232-16.
