View source for Ligilactobaillus ruminis WC1T17

== Introduction ==

=== Strain characteristics and function in the rumen ===
[[File:Lruminis WC1T17 microscopy.png|thumb|Microscopy of ''L. ruminis'' WC1T17 at 100X magnification.]]
''Ligilactobacillus ruminis'' WC1T17 is a Gram-positive, rod-shaped, aerotolerant anaerobic bacterium that typically grows in a single bacillus or diplo-bacillus cellular arrangement. ''Lactobacillus'' species are of increasing interest for methane mitigation research in ruminant animals as their metabolic network in the rumen can function as a hydrogen sink. ''Ligilactobacillus'' are lactic acid bacteria which ferment glucose in the rumen and produce lactate, this lactate can be utilized by syntrophic lactate utilizing bacteria to produce propionate, an important metabolite for the host animal, and this process also requires absorbs hydrogen thus reducing available substrate for methanogenesis.  ''L. ruminis'' is of particular interest to the Global Methane Hub as it is the only known member of the ''Lactobacillus'' genus to be motile making it an attractive prospect for ruminant methane mitigation research as it is theoretically better suited to translocate to multiple niches within the rumen biome.

=== Strain history and information ===
''L. ruminis'' WC1T17 was isolated from the rumen of a healthy cow in Palmerston North, New Zealand by the Institute for Bioeconomy Science, formerly known as AgResearch. The strain contains a circular 1.9 Mb genome with no extrachromosomal elements such as plasmids. This strain can be readily cultured in MRS broth and on MRS agar plates under anaerobic conditions with an average doubling time of 40 minutes.

Minimum inhibitory concentration (MIC) screening of antibiotics and counter-selective agents in liquid media shows that the strain is sensitive to the following tested antibiotics: chloramphenicol, ampicillin, tetracycline, and streptomycin and the following counter-selective agents: 8-azehypoxanthine, 2-deoxy-d-galactose, 5-fluordeoxyuridine, and sucrose.

''L. ruminis'' WC1T17 is genetically tractable and has been successfully transformed via electroporation with the replicative shuttle vector [https://plasmidvectors.com/general-cloning/ pMTL89151]. This vector confers resistance to chloramphenicol and contains the pAMB1 Gram-positive origin of replication machinery in addition to a multiple cloning sight for integration of reporter genes or biological payloads of interest.

== Global Methane Hub Metadata ==
{| class="wikitable"
| valign="top" |Strain origin:
| valign="top" |New Zealand<sup>1</sup>
|-
| valign="top" |Isolated from:
| valign="top" |Cow rumen
|-
| valign="top" |Sequenced by:
| valign="top" |Rhiannon Cecil<sup>2</sup>
|-
| valign="top" |Assembled by:
| valign="top" |William G. Alexander<sup>3</sup>
|-
| valign="top" |Genome and methylome analysis by:
| valign="top" |William G. Alexander<sup>3</sup> & Ashley Hall<sup>2</sup>
|-
| valign="top" |MIC and culturing conditions by:
| valign="top" |Rhiannon Cecil & Ashley Howes<sup>2</sup>
|-
| valign="top" |Transformed by:
| valign="top" |Ashley Howes
|}
<sup>1</sup>Institute for Bioeconomy Science, Palmerston North, New Zealand. <sup>2</sup>BiomEdit, Indianapolis, Indiana, United States. <sup>3</sup>Oak Ridge National Lab, Oak Ridge, Tennessee, United States.

== Genome ==
{| class="wikitable"
| colspan="2" valign="top" |'''''L. ruminis'' WC1T17 Genome Summary'''
|-
| valign="top" |'''Sequencing method'''
| valign="top" |'''Oxford Nanopore'''
|-
| valign="top" |'''Contigs'''
| valign="top" |'''1'''
|-
| valign="top" |'''Base pairs'''
| valign="top" |'''1.9 Mb'''
|-
| valign="top" |'''CDSs'''
| valign="top" |'''1,830'''
|-
| valign="top" |'''CRISPR'''
| valign="top" |'''1'''
|-
| valign="top" |'''Genes'''
| valign="top" |'''1,952'''
|-
| valign="top" |'''rRNA'''
| valign="top" |'''19'''
|-
| valign="top" |'''tRNA'''
| valign="top" |'''73'''
|-
| valign="top" |'''Miscellaneous RNA'''
| valign="top" |'''29'''
|-
| valign="top" |'''Antimicrobial resistance genes'''
| valign="top" |'''None'''
|}

== Methylome ==

=== Methylome analysis summary results ===
{| class="wikitable"
| colspan="2" |''L. ruminis'' WC1T17 Methylome Summary
|-
| valign="top" |Isolation Motif
| valign="top" |% Motif Methylation
|-
| valign="top" |GTTGN(6mA)C
| valign="top" |99.13
|-
| valign="top" |CAC(6mA)G
| valign="top" |92.58
|-
| valign="top" |CAGR(6mA)G
| valign="top" |99.54
|}

=== DefenseFinder Results ===
{| class="wikitable"
| colspan="2" valign="top" |''L. ruminis'' WC1T17 DefenseFinder Results
|-
| valign="top" |DefenseFinder System
| valign="top" |# Found
|-
| valign="top" |Cas (Class I, Type I-E)
| valign="top" |1
|-
| valign="top" |RM Type III
| valign="top" |1
|-
| valign="top" |PD-T7-2
| valign="top" |1
|-
| valign="top" |RM Type IIG
| valign="top" |3
|}

=== Methylating ''Escherichia coli'' strains ===
''E. coli'' strain AGXXXXX engineered to carry methytransferases X, X, X for methylating plasmid DNA for ''L. ruminis'' WC1T17. 

== Culturing Conditions ==
''L. ruminis'' WC1T17 can be readily cultured in MRS broth or agar plates under anaerobic conditions (75% N<sub>2</sub>, 5% H<sub>2</sub>, and 20% CO<sub>2</sub>) at 39°C for 12 – 48 hours of incubation. ''L. ruminis'' WC1T17 colonies typically form on MRS agar plates overnight and colonies inoculated from an agar plate or transferred from an active liquid culture will also grow overnight; however, when reviving from cryostock the strain takes ~48 hours to exhibit growth in broth. The strain will remain viable in liquid or solid medium for ~1 week at room temperature under anaerobic conditions. For long-tern storage 1mL of active culture with an OD<sub>600</sub> >0.8 should be combined with anaerobic 50% glycerol to a final glycerol concentration of 25% in anaerobic screw-cap cryovials then stored at -80°C. Revival of cryostocked cells is achieved by transferring a small amount of the cryostock to either an MRS agar plate or to MRS broth followed by incubation under anaerobic conditions.

== Antimicrobial Susceptibility ==

== Transformation Results ==

== External Links ==