Summary of Study ST002051

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 PR001297. The data can be accessed directly via it's Project DOI: 10.21228/M82H7B 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.

Perform statistical analysis  |  Show all samples  |  Show named metabolites  |  Download named metabolite data  
Download mwTab file (text)   |  Download mwTab file(JSON)   |  Download data files (Contains raw data)
Study IDST002051
Study TitleThe apicomplexan parasite Toxoplasma gondii forms bradyzoite-containing tissue cysts that cause chronic and drug-tolerant infections.
Study SummaryThe apicomplexan parasite Toxoplasma gondii forms bradyzoite-containing tissue cysts that cause chronic and drug-tolerant infections. Here, we developed a human myotube-based in vitro culture model of functionally mature tissue cysts. Metabolomic characterization of purified cysts reveals global changes that comprise increased levels of amino acids and decreased abundance of nucleobase- and tricarboxylic acid cycle-associated metabolites. In contrast to fast replicating tachyzoite forms of T. gondii these tissue cysts tolerate exposure to the aconitase inhibitor sodium fluoroacetate.
Institute
Robert Koch-Institute
DepartmentNG2
LaboratoryNG2
Last NameBlume
First NameMartin
AddressSeestraße 10
Emailblumem@rki.de
Phone+49 30 18754 2572
Submit Date2022-01-04
Raw Data AvailableYes
Raw Data File Type(s)mzML
Analysis Type DetailLC-MS
Release Date2022-02-28
Release Version1
Martin Blume Martin Blume
https://dx.doi.org/10.21228/M82H7B
ftp://www.metabolomicsworkbench.org/Studies/ application/zip

Select appropriate tab below to view additional metadata details:


Project:

Project ID:PR001297
Project DOI:doi: 10.21228/M82H7B
Project Title:In vitro maturation of Toxoplasma gondii bradyzoites in human myotubes and their metabolomic characterization
Project Type:characterization of in vitro T. gondii stages
Project Summary:The apicomplexan parasite Toxoplasma gondii forms bradyzoite-containing tissue cysts that cause chronic and drug-tolerant infections. Here, we developed a human myotube-based in vitro culture model of functionally mature tissue cysts. Metabolomic characterization of purified cysts reveals global changes that comprise increased levels of amino acids and decreased abundance of nucleobase- and tricarboxylic acid cycle-associated metabolites. In contrast to fast replicating tachyzoite forms of T. gondii these tissue cysts tolerate exposure to the aconitase inhibitor sodium fluoroacetate.
Institute:Robert Koch-Institut
Last Name:Blume
First Name:Martin
Address:Seestr. 10, Berlin, Berlin, 13353, Germany
Email:blumem@rki.de
Phone:+49 30 18754 2572

Subject:

Subject ID:SU002133
Subject Type:Other organism
Subject Species:Toxoplasma gondii
Taxonomy ID:5811

Factors:

Subject type: Other organism; Subject species: Toxoplasma gondii (Factor headings shown in green)

mb_sample_id local_sample_id Factor
SA1935100424_Blank_4Blank
SA1935110424_Blank_1Blank
SA1935120424_Blank_2Blank
SA1935130427_Blank_1Blank
SA1935140427_Blank_2Blank
SA1935150427_Blank_4Blank
SA1935160427_Blank_3Blank
SA1935170424_Blank_3Blank
SA1935580424_NP_T1NP_bead
SA1935590427_NP_T1NP_bead
SA1935180427_NP_Cysten_T1_2NP_Cysten_bead
SA1935190427_NP_Cysten_T1_4NP_Cysten_bead
SA1935200424_NP_Cysten_T1_1NP_Cysten_bead
SA1935210427_NP_Cysten_T1_3NP_Cysten_bead
SA1935220427_NP_Cysten_T1_1NP_Cysten_bead
SA1935230424_NP_Cysten_T1_4NP_Cysten_bead
SA1935240424_NP_Cysten_T1_3NP_Cysten_bead
SA1935250424_NP_Cysten_T1_2NP_Cysten_bead
SA1935260427_NP_Tachy_HFF_2NP_Tachy_HFF
SA1935270427_NP_Tachy_HFF_4NP_Tachy_HFF
SA1935280427_NP_Tachy_HFF_3NP_Tachy_HFF
SA1935290424_NP_Tachy_HFF_1NP_Tachy_HFF
SA1935300427_NP_Tachy_HFF_1NP_Tachy_HFF
SA1935310424_NP_Tachy_HFF_3NP_Tachy_HFF
SA1935320424_NP_Tachy_HFF_2NP_Tachy_HFF
SA1935330424_NP_Tachy_HFF_4NP_Tachy_HFF
SA1935340427_NP_Tachy_HFF_T1_2NP_Tachy_HFF_bead
SA1935350427_NP_Tachy_HFF_T1_4NP_Tachy_HFF_bead
SA1935360427_NP_Tachy_HFF_T1_3NP_Tachy_HFF_bead
SA1935370427_NP_Tachy_HFF_T1_1NP_Tachy_HFF_bead
SA1935380424_NP_Tachy_HFF_T1_3NP_Tachy_HFF_bead
SA1935390424_NP_Tachy_HFF_T1_1NP_Tachy_HFF_bead
SA1935400424_NP_Tachy_HFF_T1_4NP_Tachy_HFF_bead
SA1935410424_NP_Tachy_HFF_T1_2NP_Tachy_HFF_bead
SA1935420427_NP_Tachy_Tubes_3NP_Tachy_Tubes
SA1935430427_NP_Tachy_Tubes_1NP_Tachy_Tubes
SA1935440427_NP_Tachy_Tubes_2NP_Tachy_Tubes
SA1935450424_NP_Tachy_Tubes_3NP_Tachy_Tubes
SA1935460424_NP_Tachy_Tubes_1NP_Tachy_Tubes
SA1935470427_NP_Tachy_Tubes_4NP_Tachy_Tubes
SA1935480424_NP_Tachy_Tubes_4NP_Tachy_Tubes
SA1935490424_NP_Tachy_Tubes_2NP_Tachy_Tubes
SA1935500424_NP_Tachy_Tubes_T1_4NP_Tachy_Tubes_bead
SA1935510427_NP_Tachy_Tubes_T1_1NP_Tachy_Tubes_bead
SA1935520427_NP_Tachy_Tubes_T1_2NP_Tachy_Tubes_bead
SA1935530427_NP_Tachy_Tubes_T1_4NP_Tachy_Tubes_bead
SA1935540424_NP_Tachy_Tubes_T1_1NP_Tachy_Tubes_bead
SA1935550424_NP_Tachy_Tubes_T1_3NP_Tachy_Tubes_bead
SA1935560427_NP_Tachy_Tubes_T1_3NP_Tachy_Tubes_bead
SA1935570424_NP_Tachy_Tubes_T1_2NP_Tachy_Tubes_bead
SA1935600427_NP_uninf_T1_3NP_uninf_bead
SA1935610427_NP_uninf_T1_2NP_uninf_bead
SA1935620427_NP_uninf_T1_4NP_uninf_bead
SA1935630424_NP_uninf_T1_3NP_uninf_bead
SA1935640424_NP_uninf_T1_1NP_uninf_bead
SA1935650427_NP_uninf_T1_1NP_uninf_bead
SA1935660424_NP_uninf_T1_2NP_uninf_bead
SA1935670424_NP_uninf_T1_4NP_uninf_bead
SA1936080424_PP_T1PP_bead
SA1936090427_PP_T1PP_bead
SA1935680424_PP_Cysten_T1_2PP_Cysten_bead
SA1935690424_PP_Cysten_T1_1PP_Cysten_bead
SA1935700427_PP_Cysten_T1_4PP_Cysten_bead
SA1935710427_PP_Cysten_T1_3PP_Cysten_bead
SA1935720424_PP_Cysten_T1_4PP_Cysten_bead
SA1935730424_PP_Cysten_T1_3PP_Cysten_bead
SA1935740427_PP_Cysten_T1_1PP_Cysten_bead
SA1935750427_PP_Cysten_T1_2PP_Cysten_bead
SA1935760427_PP_Tachy_HFF_2PP_Tachy_HFF
SA1935770427_PP_Tachy_HFF_4PP_Tachy_HFF
SA1935780427_PP_Tachy_HFF_1PP_Tachy_HFF
SA1935790427_PP_Tachy_HFF_3PP_Tachy_HFF
SA1935800424_PP_Tachy_HFF_2PP_Tachy_HFF
SA1935810424_PP_Tachy_HFF_1PP_Tachy_HFF
SA1935820424_PP_Tachy_HFF_3PP_Tachy_HFF
SA1935830424_PP_Tachy_HFF_4PP_Tachy_HFF
SA1935840427_PP_Tachy_HFF_T1_2PP_Tachy_HFF_bead
SA1935850427_PP_Tachy_HFF_T1_4PP_Tachy_HFF_bead
SA1935860427_PP_Tachy_HFF_T1_1PP_Tachy_HFF_bead
SA1935870427_PP_Tachy_HFF_T1_3PP_Tachy_HFF_bead
SA1935880424_PP_Tachy_HFF_T1_3PP_Tachy_HFF_bead
SA1935890424_PP_Tachy_HFF_T1_1PP_Tachy_HFF_bead
SA1935900424_PP_Tachy_HFF_T1_2PP_Tachy_HFF_bead
SA1935910424_PP_Tachy_HFF_T1_4PP_Tachy_HFF_bead
SA1935920427_PP_Tachy_Tubes_3PP_Tachy_Tubes
SA1935930427_PP_Tachy_Tubes_2PP_Tachy_Tubes
SA1935940424_PP_Tachy_Tubes_1PP_Tachy_Tubes
SA1935950427_PP_Tachy_Tubes_1PP_Tachy_Tubes
SA1935960427_PP_Tachy_Tubes_4PP_Tachy_Tubes
SA1935970424_PP_Tachy_Tubes_4PP_Tachy_Tubes
SA1935980424_PP_Tachy_Tubes_2PP_Tachy_Tubes
SA1935990424_PP_Tachy_Tubes_3PP_Tachy_Tubes
SA1936000427_PP_Tachy_Tubes_T1_2PP_Tachy_Tubes_bead
SA1936010427_PP_Tachy_Tubes_T1_4PP_Tachy_Tubes_bead
SA1936020427_PP_Tachy_Tubes_T1_1PP_Tachy_Tubes_bead
SA1936030427_PP_Tachy_Tubes_T1_3PP_Tachy_Tubes_bead
SA1936040424_PP_Tachy_Tubes_T1_2PP_Tachy_Tubes_bead
SA1936050424_PP_Tachy_Tubes_T1_3PP_Tachy_Tubes_bead
SA1936060424_PP_Tachy_Tubes_T1_1PP_Tachy_Tubes_bead
SA1936070424_PP_Tachy_Tubes_T1_4PP_Tachy_Tubes_bead
Showing page 1 of 2     Results:    1  2  Next     Showing results 1 to 100 of 108

Collection:

Collection ID:CO002126
Collection Summary:For metabolome measurements of tissue cysts and tachyzoites, tachyzoite isolation, cyst maturation and isolation were performed as described above. Metabolites were extracted in 80 % acetonitrile (Carl Roth) and 20 % water (Carl Roth) containing internal standards (phenolphthalein, CAPS, PIPES (Sigma-Aldrich)). Cell pellets were sonicated for 5 min and after centrifugation (21,500 x g, 5 min, 0 °C), the supernatants were transferred to MS vials for immediate LC/MS analysis. 5 µl of each sample were collected to generate a pooled biological quality control (PBQC). 20 µl of the in vitro cysts, bead control and host cell background samples, and 5 µl of the tachyzoite samples were injected. The injection order of the samples was randomized, blanks and PBQCs were injected periodically. The samples were analyzed on a Q-Exactive Plus mass spectrometer (Thermo Fisher Scientific) via 70k MS1 scans, with intermittent 35k data-dependent 35k MS2 scans in positive and negative mode separately. Chromatographic separation was achieved on a Vanquish Flex fitted with an ACQUITY UPLC BEH Amide column (Waters). Running a 25 min linear gradient starting with 90 % eluent A (10 mM ammonium carbonate in acetonitrile) / 10 % eluent B (10 mM ammonium carbonate in water) and ending with 40 % eluent A / 60 % eluent B, followed by washing and equilibration steps. XCalibur 4.2.47 software and Compound Discoverer 3.1 software (Thermo Fisher Scientific) was used for recording and used for peak detection, combination of adducts and compound annotation, respectively. Metabolite identifications were based on either retention time and accurate mass match to an in-house library of 160 authentic standards, or by matching accurate mass and MS2 fragments to m/z cloud database (mirrored offline in m/z vault v 2.1.22.15 in May 2020) (Thermo Fisher Scientific).
Sample Type:Cultured human myotubes with Toxoplasma gondii

Treatment:

Treatment ID:TR002145
Treatment Summary:Cells were either uninfected, tachyzoites, tachyzoites with beads, bradyzoites (cysts), beads only

Sample Preparation:

Sampleprep ID:SP002139
Sampleprep Summary:Uninfected and infected myotubes were prepared. In both cases, two T150 dishes were pooled into one sample. For bradyzoite samples, myotubes were infected with 3.2*106 Pru-tdTomato tachyzoites corresponding to a MOI of 0.3 and cyst formation was induced for indicated time. On the day of harvest, infected samples and uninfected host cell controls were placed on ice, medium was removed and monolayers were washed three times with ice-cold PBS. Cells were then harvested by scraping into 10 ml ice-cold 0.05 % BSA in PBS per T150 dish. Cysts were released from the monolayer via forcing through a 23G needle (Sterican®) 25 times with a syringe and collected via centrifugation (1,200 x g, 10 min, 0 °C). The supernatant was removed, the pellet was resuspended carefully in ice-cold 2 % BSA in PBS containing 200 µl DBA-coupled beads (preparation described below) and samples were incubated for 1 h at 4 °C with gentle shaking. Subsequently, the samples were placed in a magnetic stand on ice, washed five times with 0.1 % BSA in PBS to remove cell debris, followed by two washing steps with PBS to remove residual BSA. Cysts and beads were then collected via centrifugation (1,200 x g, 10 min, 0 °C), shock frozen in liquid nitrogen and stored at -80 °C until extraction. Tachyzoite samples were generated in T150 dishes by infecting myotubes and HFF cells with 3.2*107 tachyzoites corresponding to an MOI of 3 for 48 h. Medium was replaced by ice-cold PBS and monolayers were scraped and passaged through a 27G needle. Tachyzoites were filter-purified through a 3 µm filter (Whatman) and PBS-washed by centrifugation (1,200 x g, 10 min, 0 °C) three times. All samples were extracted simultaneously in 80 % acetonitrile for LC/MS analysis as described below. Bead-only controls were processed equally. Bead-supplemented tachyzoite controls were processed equally to cyst samples, replacing washing steps via magnetic stand by centrifugation (1,200 x g, 10 min, 0 °C). Preparation of beads Coupling of DynabeadsTM MyONETM Streptavdin T1 (Thermo Fisher Scientific) to DBA was done as described in the manufacturer's protocol. Briefly, 200 µl beads/sample were resuspended in 1 ml PBS by vortexing, washed three times with PBS in a magnetic stand and resuspended in 1 ml PBS containing 50 µg DBA / sample. The tube containing the DBA-magnetic bead mixture was incubated on a rotary mixer for 45 min at RT. Uncoupled DBA was removed by washing the coated beads three times with PBS. After washing, the DBA-coated beads were resuspended in 2 ml PBS containing 2 % BSA.

Combined analysis:

Analysis ID AN003338
Analysis type MS
Chromatography type HILIC
Chromatography system Thermo Vanquish
Column Waters Acquity BEH Amide (150 x 2.1mm, 1.7um)
MS Type ESI
MS instrument type Orbitrap
MS instrument name Thermo Exactive Plus Orbitrap
Ion Mode UNSPECIFIED
Units counts

Chromatography:

Chromatography ID:CH002472
Chromatography Summary:Chromatographic separation was achieved on a Vanquish Flex fitted with an ACQUITY UPLC BEH Amide column (Waters). Running a 25 min linear gradient starting with 90 % eluent A (10 mM ammonium carbonate in acetonitrile) / 10 % eluent B (10 mM ammonium carbonate in water) and ending with 40 % eluent A / 60 % eluent B, followed by washing and equilibration steps.
Instrument Name:Thermo Vanquish
Column Name:Waters Acquity BEH Amide (150 x 2.1mm, 1.7um)
Chromatography Type:HILIC

MS:

MS ID:MS003107
Analysis ID:AN003338
Instrument Name:Thermo Exactive Plus Orbitrap
Instrument Type:Orbitrap
MS Type:ESI
MS Comments:The injection order of the samples was randomized, blanks and PBQCs were injected periodically. The samples were analyzed on a Q-Exactive Plus mass spectrometer (Thermo Fisher Scientific) via 70k MS1 scans, with intermittent 35k data-dependent 35k MS2 scans in positive and negative mode separately.XCalibur 4.2.47 software and Compound Discoverer 3.1 software (Thermo Fisher Scientific) was used for recording and used for peak detection, combination of adducts and compound annotation, respectively. Metabolite identifications were based on either retention time and accurate mass match to an in-house library of 160 authentic standards, or by matching accurate mass and MS2 fragments to m/z cloud database (mirrored offline in m/z vault v 2.1.22.15 in May 2020) (Thermo Fisher Scientific). Data were exported to Excel for grouping, combination of datasets from positive and negative ionization runs and blank subtraction.
Ion Mode:UNSPECIFIED
  logo