Summary of Study ST002512

This data is available at the NIH Common Fund's National Metabolomics Data Repository (NMDR) website, the Metabolomics Workbench, https://www.metabolomicsworkbench.org, where it has been assigned Project ID PR001620. The data can be accessed directly via it's Project DOI: 10.21228/M89B04 This work is supported by NIH grant, U2C- DK119886.

See: https://www.metabolomicsworkbench.org/about/howtocite.php

This study contains a large results data set and is not available in the mwTab file. It is only available for download via FTP as data file(s) here.

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Study IDST002512
Study TitleGnotobiotic mice: Metabolites in intestinal contents of germ-free mice colonized with strains of gut bacterium Eggerthella lenta
Study TypeUntargeted LC-MS
Study SummaryThis dataset contains untargeted metabolomics analysis of intestinal contents of gnotobiotic mice either colonized with different strains of Eggerthella lenta for 2 weeks, or germ-free controls.
Institute
University of California, San Francisco
Last NameNoecker
First NameCecilia
Address513 Parnassus Ave HSW1501, San Francisco, CA 94143
Emailcecilia.noecker@ucsf.edu
Phone415-502-3264
Submit Date2023-03-21
Raw Data AvailableYes
Raw Data File Type(s)mzML, raw(Thermo)
Analysis Type DetailLC-MS
Release Date2023-04-10
Release Version1
Cecilia Noecker Cecilia Noecker
https://dx.doi.org/10.21228/M89B04
ftp://www.metabolomicsworkbench.org/Studies/ application/zip

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Project:

Project ID:PR001620
Project DOI:doi: 10.21228/M89B04
Project Title:Systems biology illuminates the alternative metabolic niche of the human gut bacterium Eggerthella lenta
Project Type:Untargeted LC-MS
Project Summary:Human gut bacteria perform diverse metabolic functions with consequences for host health. The prevalent and disease-linked Actinobacterium Eggerthella lenta performs several unusual chemical transformations, but it does not metabolize sugars and its core growth strategy remains unclear. To obtain a comprehensive view of the metabolic network of E. lenta, we generated several complementary resources: defined culture media, metabolomics profiles of strain isolates, and a curated genome-scale metabolic reconstruction. Stable isotope-resolved metabolomics revealed that E. lenta uses acetate as a key carbon source while catabolizing arginine to generate ATP, traits which could be recapitulated in silico by our updated metabolic model. We compared these in vitro findings with metabolite shifts observed in E. lenta-colonized gnotobiotic mice, identifying shared signatures across environments and highlighting catabolism of the host signaling metabolite agmatine as an alternative energy pathway. Together, our results elucidate a distinctive metabolic niche filled by E. lenta in the gut ecosystem.
Institute:University of California, San Francisco
Department:Microbiology and Immunology
Laboratory:Peter Turnbaugh
Last Name:Noecker
First Name:Cecilia
Address:513 Parnassus Ave HSW1501, San Francisco, CA 94143
Email:cecilia.noecker@ucsf.edu
Phone:415-502-3264
Funding Source:This work was supported by the National Institutes of Health (2R01HL122593; 1R01AT011117; 1R01DK114034 to P.J.T., F32GM140808 to C.N.). P.J.T. is a Chan Zuckerberg Biohub Investigator and held an Investigators in the Pathogenesis of Infectious Disease Award from the Burroughs Wellcome Fund.
Publications:https://doi.org/10.1101/2022.09.19.508335

Subject:

Subject ID:SU002612
Subject Type:Mammal
Subject Species:Mus musculus
Taxonomy ID:10090
Genotype Strain:C57BL/6J
Age Or Age Range:6-10 weeks
Gender:Male
Animal Animal Supplier:University of California, San Francisco Gnotobiotics core facility

Factors:

Subject type: Mammal; Subject species: Mus musculus (Factor headings shown in green)

mb_sample_id local_sample_id SampleType Group
SA252900Cecal_El15644_7Cecal 15644
SA252901Cecal_El15644_23Cecal 15644
SA252902Cecal_El15644_14Cecal 15644
SA252903Cecal_El15644_21Cecal 15644
SA252904Cecal_El15644_22Cecal 15644
SA252905Cecal_El15644_6Cecal 15644
SA252906Cecal_El2243_5Cecal 2243
SA252907Cecal_El2243_12Cecal 2243
SA252908Cecal_El2243_13Cecal 2243
SA252909Cecal_El2243_25Cecal 2243
SA252910Cecal_El2243_4Cecal 2243
SA252911Cecal_ElAB12n2_17Cecal AB12n2
SA252912Cecal_ElAB12n2_11Cecal AB12n2
SA252913Cecal_ElAB12n2_10Cecal AB12n2
SA252914Cecal_ElAB12n2_16Cecal AB12n2
SA252915Cecal_ElAB12n2_18Cecal AB12n2
SA252916Cecal_ElAB12n2_29Cecal AB12n2
SA252917Cecal_ElAB12n2_28Cecal AB12n2
SA252918Cecal_ElAB12n2_30Cecal AB12n2
SA252919Cecal_GF_3Cecal GF
SA252920Cecal_GF_27Cecal GF
SA252921Cecal_GF_2Cecal GF
SA252922Cecal_GF_26Cecal GF
SA252923Cecal_GF_1Cecal GF
SA252862pool_1- -
SA252863bk_3(1)- -
SA252864bk_3- -
SA252865SI_pool_0- -
SA252866pool_2- -
SA252867bk_2- -
SA252868pool_3- -
SA252869preBk- -
SA252870Bk_1- -
SA252871SI_pool_1- -
SA252872SI_pool_2- -
SA252873Cecal_pool_3- -
SA252874LI_pool_3- -
SA252875LI_pool_2- -
SA252876LI_pool_1- -
SA252877Cecal_pool_2- -
SA252878Cecal_pool_1- -
SA252879SI_pool_3- -
SA252880SI_El2243_24- -
SA252881Cecal_pool_0- -
SA252882bk_1- -
SA252883bk_1(1)- -
SA252884Bk_4- -
SA252885Bk_5- -
SA252886Bk_6- -
SA252887Bk_7- -
SA252888Bk_3(1)- -
SA252889Bk_3- -
SA252890Bk_1(1)- -
SA252891Bk_10- -
SA252892Bk_11- -
SA252893Bk_2(1)- -
SA252894Bk_8- -
SA252895Bk_9- -
SA252896LI_pool_0- -
SA252897bk_12- -
SA252898bk_11(1)- -
SA252899Cecal_El2243_24- -
SA252924LI_El15644_22LI 15644
SA252925LI_El15644_21LI 15644
SA252926LI_El15644_14LI 15644
SA252927LI_El15644_23LI 15644
SA252928LI_El15644_6LI 15644
SA252929LI_El15644_7LI 15644
SA252930LI_El2243_25LI 2243
SA252931LI_El2243_4LI 2243
SA252932LI_El2243_24LI 2243
SA252933LI_El2243_13LI 2243
SA252934LI_El2243_12LI 2243
SA252935LI_El2243_5LI 2243
SA252936LI_ElAB12n2_11LI AB12n2
SA252937LI_ElAB12n2_17LI AB12n2
SA252938LI_ElAB12n2_16LI AB12n2
SA252939LI_ElAB12n2_18LI AB12n2
SA252940LI_ElAB12n2_30LI AB12n2
SA252941LI_ElAB12n2_29LI AB12n2
SA252942LI_ElAB12n2_28LI AB12n2
SA252943LI_ElAB12n2_10LI AB12n2
SA252944LI_GF_3LI GF
SA252945LI_GF_26LI GF
SA252946LI_GF_27LI GF
SA252947LI_GF_2LI GF
SA252948LI_GF_1LI GF
SA252949SI_El15644_23SI 15644
SA252950SI_El15644_22SI 15644
SA252951SI_El15644_14SI 15644
SA252952SI_El15644_7SI 15644
SA252953SI_El15644_21SI 15644
SA252954SI_El15644_6SI 15644
SA252955SI_El2243_13SI 2243
SA252956SI_El2243_12SI 2243
SA252957SI_El2243_5SI 2243
SA252958SI_El2243_25SI 2243
SA252959SI_El2243_4SI 2243
SA252960SI_ElAB12n2_11SI AB12n2
SA252961SI_ElAB12n2_17SI AB12n2
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Collection:

Collection ID:CO002605
Collection Summary:Sample collection was performed as described in doi.org/10.1016/j.chom.2021.11.001 . Briefly, C57BL/6J mice (males, ages 4-8 weeks) were obtained from the University of California, San Francisco Gnotobiotics core facility (gnotobiotics.ucsf.edu) and housed in Iso positive cages (Tecniplast). Mice were colonized via oral gavage with E. lenta monocultures (109 CFU/mL, 200 μl gavage, n=6 E. lenta DSM 2243, n=5 E. lenta DSM 15644, n=4 E. lenta AB12n2) or control media (n=5 GF). Colonization was confirmed via anaerobic culturing and/or qPCR for an E. lenta specific marker (elnmrk1) [10,26]. Mice were colonized for 2 weeks prior to sacrifice and sample collection. Samples were collected of ileal, cecal, and colonic contents.
Sample Type:Intestine
Collection Frequency:single time point (72 hours)
Storage Conditions:-80℃

Treatment:

Treatment ID:TR002624
Treatment Summary:Mice were colonized via oral gavage with E. lenta monocultures (109 CFU/mL, 200 μl gavage, n=6 E. lenta DSM 2243, n=5 E. lenta DSM 15644, n=4 E. lenta AB12n2) or control media (n=5 GF). Colonization was confirmed via anaerobic culturing and/or qPCR for an E. lenta specific marker (elnmrk1). Mice were colonized for 2 weeks prior to sacrifice and sample collection. Samples were collected of ileal, cecal, and colonic contents, as well as serum.

Sample Preparation:

Sampleprep ID:SP002618
Sampleprep Summary:Intestinal samples (colon, cecum, ileum) were prepared individually using a single protocol as follows. Samples were kept frozen on dry ice and massed to at least 10 mg. Four microliters of -20oC extraction solvent (2:2:1 methanol:acetonitrile:water + stable isotope labeled internal standards) were added per milligram of intestinal sample. Six to eight 1mm zirconia silica beads were added to each sample followed by prompt bead beating (15 Hz, for 10 minutes). Following a 1 hour incubation in the -20oC freezer, samples were centrifuged at 4oC at 18,407 rcf for 5 minutes. Supernatant was collected and stored at -20oC prior to centrifugal plate filtration (0.2 micron polyvinylidene difluoride (PVDF) Agilent Technologies, Santa Clara CA) at 4oC at 4,122 rcf for 3 min. Collection plate was sealed and maintained at 4oC prior to prompt analysis. Serum samples were first thawed on wet ice. 20 μL of serum was extracted with 4 volumes of methanol, containing stable isotope labeled internal standards. Samples were homogenized by vortexing for 20 seconds and placed in a -20oC for 1 hour to maximize protein precipitation. After freezer incubation, samples were centrifuged at 4oC at 18,407 rcf for 5 minutes. Supernatant was removed and dried under vacuum via centrivap (Labconco Corp.). Dried samples were then resuspended in 30 μL of 80% acetonitrile in water containing exogenous standard CUDA at 60 ng/mL. Samples were maintained at 4oC prior to prompt analysis.

Combined analysis:

Analysis ID AN004136 AN004137
Analysis type MS MS
Chromatography type HILIC HILIC
Chromatography system Thermo Vanquish Thermo Vanquish
Column Waters ACQUITY UPLC BEH Amide (150 x 2.1mm,1.7um) Waters ACQUITY UPLC BEH Amide (150 x 2.1mm,1.7um)
MS Type ESI ESI
MS instrument type Orbitrap Orbitrap
MS instrument name Thermo Q Exactive HF hybrid Orbitrap Thermo Q Exactive HF hybrid Orbitrap
Ion Mode POSITIVE NEGATIVE
Units relative ion counts relative ion counts

Chromatography:

Chromatography ID:CH003064
Chromatography Summary:Samples, pools, and blanks were promptly added to a Thermo Vanquish Autosampler at 4°C in a Vanquish UHPLC (Thermo Fisher Scientific, Waltham, MA). Chromatographic separation was performed using an ACQUITY Bridged Ethylene Hybrid (BEH) Amide column 2.1 x 150 mm, 1.7-micron particle size, (Waters Corp. Milford, MA), using chromatographic conditions published elsewhere (HILIC method described in the Supplementary Methods of doi.org/10.1038/s41586-021-03707-9).
Instrument Name:Thermo Vanquish
Column Name:Waters ACQUITY UPLC BEH Amide (150 x 2.1mm,1.7um)
Column Temperature:40
Flow Gradient:The gradient profile was held at 100% B for 2 minutes, from 100% B to 70% B in 5 minutes, holding at 70% B for 0.7 minute, from 70% B to 40% B for 1.3 minutes, holding at 40% B for 0.5 minutes, from 40% B to 30% B for 0.75 minutes, before returning to 100% B for 2.5 minutes and holding at 100% B for 4 minutes.
Flow Rate:400 μL per minute
Solvent A:100% water; 0.125% formic acid; 10 mM ammonium formate, pH 3
Solvent B:95% acetonitrile/5% water; 0.125% formic acid; 10 mM ammonium formate
Chromatography Type:HILIC

MS:

MS ID:MS003883
Analysis ID:AN004136
Instrument Name:Thermo Q Exactive HF hybrid Orbitrap
Instrument Type:Orbitrap
MS Type:ESI
MS Comments:Full MS-ddMS2 data was collected, an inclusion list was used to prioritize MS2 selection of metabolites from our in-house ‘local’ library, when additional scan bandwidth was available MS2 was collected in a data-dependent manner. Mass range was 60-900 mz, resolution was 60k (MS1) and 15k (MS2), centroid data was collected, loop count was 4, isolation window was 1.5 Da. Metabolomics data was processed using MS-DIAL v4.60. Features were excluded from analysis if peak height was not at least 5-fold greater in one or more samples compared to the procedural blank average.
Ion Mode:POSITIVE
  
MS ID:MS003884
Analysis ID:AN004137
Instrument Name:Thermo Q Exactive HF hybrid Orbitrap
Instrument Type:Orbitrap
MS Type:ESI
MS Comments:Full MS-ddMS2 data was collected, an inclusion list was used to prioritize MS2 selection of metabolites from our in-house ‘local’ library, when additional scan bandwidth was available MS2 was collected in a data-dependent manner. Mass range was 60-900 mz, resolution was 60k (MS1) and 15k (MS2), centroid data was collected, loop count was 4, isolation window was 1.5 Da. Metabolomics data was processed using MS-DIAL v4.60. Features were excluded from analysis if peak height was not at least 5-fold greater in one or more samples compared to the procedural blank average.
Ion Mode:NEGATIVE
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