Summary of study ST000570

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 PR000418. The data can be accessed directly via it's Project DOI: 10.21228/M81880 This work is supported by NIH grant, U2C- DK119886.

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Study IDST000570
Study TitleMetabolome analysis of the cecal contents of GF mice and GF mice colonized with dominant gut microbes present in the ceca of neonatal and adult mice
Study SummaryMetabolome profiles of GF or GF mice reconstituted with Esherichia coli (EC), Bacteroides acidifaciens (Bac), or Clostridia consortium (CL) were compared.
Institute
Keio University
DepartmentInstitute for Advanced Biosciences
Last NameFukuda
First NameShinji
AddressTsuruoka, Yamagata 997-0052, Japan
Emailsfukuda@sfc.keio.ac.jp
Phone+81-235-29-0528
Submit Date2017-03-09
Num Groups4
Total Subjects17
Raw Data AvailableYes
Raw Data File Type(s).bin, .stg, .xml, .xsd, .log
Analysis Type DetailLC-MS
Release Date2018-04-10
Release Version1
Shinji Fukuda Shinji Fukuda
https://dx.doi.org/10.21228/M81880
ftp://www.metabolomicsworkbench.org/Studies/ application/zip

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Project:

Project ID:PR000418
Project DOI:doi: 10.21228/M81880
Project Title:Metabolome analysis of the cecal contents of GF mice and GF mice colonized with dominant gut microbes present in the ceca of neonatal and adult mice
Project Summary:The high susceptibility of neonates to infections has been assumed to be due to immaturity of the immune system, but the mechanism remains unclear. By colonizing adult germ-free mice with the cecal contents of neonatal and adult mice, we show that the neonatal microbiota is impaired in mediating colonization resistance against two major pathogens causing mortality in neonates. The lack of colonization resistance was caused by the absence of Clostridiales in the neonatal microbiota. Administration of Clostridiales, but not Bacteroidales, restored colonization resistance and abrogated intestinal pathology upon pathogen challenge. Conversely, depletion of Clostridiales abolished colonization resistance in adult mice. Furthermore, intragastric administration of Clostridiales protected neonatal mice from pathogen infection. The neonatal bacteria enhanced the ability of these protective Clostridiales to colonize the gut. These results identify the gut microbiota as a critical determinant of increased susceptibility to enteric infection during the neonatal period.
Institute:Keio University
Department:Institute for Advanced Biosciences
Last Name:Fukuda
First Name:Shinji
Address:Tsuruoka, Yamagata 997-0052, Japan
Email:sfukuda@sfc.keio.ac.jp
Phone:+81-235-29-0528

Subject:

Subject ID:SU000592
Subject Type:Animal
Subject Species:Mus musculus
Taxonomy ID:10090
Genotype Strain:C57BL/6
Species Group:Mammal

Factors:

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

mb_sample_id local_sample_id colonized bacteria
SA0299251_B5Bacteroides acidifaciens
SA0299261_B1Bacteroides acidifaciens
SA0299271_B4Bacteroides acidifaciens
SA0299281_B3Bacteroides acidifaciens
SA0299291_B2Bacteroides acidifaciens
SA0299301_CL5Clostridia
SA0299311_CL4Clostridia
SA0299321_CL1Clostridia
SA0299331_CL2Clostridia
SA0299341_CL3Clostridia
SA0299352_EB3Esherichia coli
SA0299362_EB2Esherichia coli
SA0299372_EB1Esherichia coli
SA0299385_C4none (GF)
SA0299395_C2none (GF)
SA0299405_C1none (GF)
SA0299415_C3none (GF)
Showing results 1 to 17 of 17

Collection:

Collection ID:CO000586
Collection Summary:Feces of GF or GF mice reconstituted with Esherichia coli (EC), Bacteroides acidifaciens (Bac), or Clostridia consortium (CL)
Sample Type:Feces
Storage Conditions:-80℃

Treatment:

Treatment ID:TR000606
Treatment Summary:Animals Specific-pathogen-free (SPF) C57BL/6 mice were originally purchased from Jackson Laboratories. Wild-type mice on GF background were bred and maintained at the Germ-free Animal Core Facility of the University of Michigan. GF mice were maintained in flexible film isolators and were checked weekly for germ-free status by aerobic and anaerobic culture. The absence of microbiota was verified by microscopic analysis of stained cecal contents to detect unculturable contamination. All animal studies were performed according to approved protocols by the University of Michigan Committee on the Use and Care of Animals. Reconstitution of GF mice with cecal contents and defined bacteria Cecal contents from single neonatal and 7-week old SPF mice were diluted into 50 ml (per one adult mouse) or 5 ml (per one 4, 12, 16-day old mouse) of PBS, and passed through a 70 μm cell strainer to eliminate clumps and debris under sterile conditions) under an atmosphere of 5% H2, 5% CO2 and 90% N2 in an anaerobic growth chamber (Coy Manufacturing, Grass Lake, MI). Then 200 μl of the suspension was processed by intragastric gavage into each of the recipient GF mice. Bacteroides species were anaerobically grown at 37°C overnight in pre-reduced chopped-meat broth (Anaerobe systems, Morgan Hill, CA). E. coli were grown aerobically overnight in LB broth with shaking.
Treatment Protocol Filename:Treatment_Protocol.pdf
Treatment:Biotic
Treatment Compound:Suspension of diluted cecal contents
Treatment Route:Intragastric gavage
Treatment Dose:200 μl

Sample Preparation:

Sampleprep ID:SP000599
Sampleprep Summary:CE-TOFMS-based metabolome analysis(sample preparation) Fecal samples were freeze dried, and disrupted by vigorous shaking at 1,500 rpm for 10 min with four 3 mm zirconia beads by Shake Master NEO (Bio Medical Science Inc.). 10 mg (±0.5 mg) fecal samples were homogenized with 500 μl MeOH containing internal standards (20 μM each of methionine sulfone, and D-camphor-10-sulfonic acid (CSA)) and 100 mg of 0.1 mm and four of 3 mm zirconia/silica beads (BioSpec Products). After vigorous shaking (1,500 rpm for 5 min) by Shake Master NEO (Bio Medical Science Inc.), 200 μl of Milli-Q water and 500 μl of chloroform was added and then shaking in a same manner as before. After centrifugation at 4,600 × g for 15 min at 4°C, the supernatant was transferred to a 5kDa cutoff centrifugal filter tube. The filtrate was centrifugally concentrated at 40°C and reconstituted with 40 μl of Milli-Q water.
Sampleprep Protocol Filename:Sample_Prep_Protocol.pdf
Processing Method:Homogenized with 500 μl MeOH containing internal standards and 100 mg of 0.1 mm and four of 3 mm zirconia/silica beads
Processing Storage Conditions:On ice
Extraction Method:Chloroform
Extract Cleanup:5kDa cutoff centrifugal filter tube
Sample Resuspension:40 μl of Milli-Q water

Combined analysis:

Analysis ID AN000877 AN000878
Analysis type MS MS
Chromatography type CE CE
Chromatography system Agilent CE Agilent CE
Column COSMO(+), i.d.50 μm(anion), Fused-silica, i.d. 50 µm(cation) COSMO(+), i.d.50 μm(anion), Fused-silica, i.d. 50 µm(cation)
MS Type ESI ESI
MS instrument type TOF TOF
MS instrument name Agilent CE-TOFMS Agilent CE-TOFMS
Ion Mode POSITIVE NEGATIVE
Units nmol/g nmol/g

Chromatography:

Chromatography ID:CH000623
Instrument Name:Agilent CE
Column Name:COSMO(+), i.d.50 μm(anion), Fused-silica, i.d. 50 µm(cation)
Internal Standard:20 μM each of methionine sulfone, and D-camphor-10-sulfonic acid (CSA)
Chromatography Type:CE

MS:

MS ID:MS000778
Analysis ID:AN000877
Instrument Name:Agilent CE-TOFMS
Instrument Type:TOF
MS Type:ESI
Ion Mode:POSITIVE
  
MS ID:MS000779
Analysis ID:AN000878
Instrument Name:Agilent CE-TOFMS
Instrument Type:TOF
MS Type:ESI
Ion Mode:NEGATIVE
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