Metabolite identification was carried out by comparing the processed peaks against a combination of an in-house database, publicly available metabolite databases (such as METLIN, HMDB), and prediction libraries. The identification process was based on accurate mass, retention time, and fragmentation patterns. The final list of identified metabolites was filtered based on a combined score above 0.5, and metabolites that passed the quality control (QC) threshold with a CV value of less than 0.3 in the QC samples were retained. The identified metabolites were then annotated using their chemical names and included in the dataset for further analysis.
"],"repository":["MetaboLights"],"study_status":["Public"],"ptm_modification":[""],"instrument_platform":["Liquid Chromatography MS - negative - reverse-phase","Liquid Chromatography MS - positive - reverse-phase"],"chromatography_protocol":["Chromatographic separation was performed using a Waters ACQUITY Premier HSS T3 column (1.8 µm, 2.1 mm × 100 mm). The gradient conditions were as follows:
0 min: 95% A, 5% B
2 min: 80% A, 20% B
5 min: 40% A, 60% B
6 min: 1% A, 99% B
7.5 min: 1% A, 99% B
7.6 min: 95% A, 5% B
10 min: 95% A, 5% B
Mobile phase A was 0.1% formic acid in water, and phase B was 0.1% formic acid in acetonitrile. The column temperature was maintained at 40°C, and the flow rate was set to 0.4 mL/min. The injection volume was 4 μL.
"],"publication":["Macrophage-mediated antibiotic evasion and competitive dominance of mcr-3-carrying Escherichia coli."],"submitter_name":["Shuyu Tan"],"submitter_affiliation":["China Agricultural University"],"organism_part":["RAW-264.7 cell"],"technology_type":["mass spectrometry assay"],"disease":[""],"extraction_protocol":["Samples were thawed on ice and 500 μL of 80% methanol-water internal standard extraction solution was added. The mixture was vortexed for 3 minutes to ensure complete suspension of the sample. The tube was then placed in liquid nitrogen and frozen for 5 minutes, followed by thawing on dry ice for 5 minutes, then on ice for 5 minutes. This freeze-thaw cycle was repeated 3 times.
After the final freeze-thaw cycle, the samples were centrifuged at 12,000 rpm for 10 minutes at 4°C. The supernatant (300 μL) was transferred to a new tube and stored at -20°C for 30 minutes. The sample was then centrifuged again at 12,000 rpm for 3 minutes at 4°C, and the resulting supernatant (200 μL) was transferred to a vial for analysis.
"],"organism":["Mus musculus"],"full_dataset_link":["https://www.ebi.ac.uk/metabolights/MTBLS13550"],"author":["Yanjun Dong. China Agricultural University. yanjund@cau.edu.cn.","Shuyu Tan. China Agricultural University. BS20233050521@cau.edu.cn.","Wang Yang. China Agricultural University. wangyang@cau.edu.cn.","Wenjuan Yin. Hebei University. ywj0501@hbu.edu.cn."],"data_transformation_protocol":["The raw data obtained from the mass spectrometry analysis were converted to mzML format using ProteoWizard software. The data were then processed using the XCMS pipeline for peak detection, alignment, and retention time correction. Peaks with a missing value rate greater than 50% were filtered out. For missing values, KNN imputation was applied if the missing value rate was below 50%, and a minimum value imputation (1/5 of the minimum value) was applied if the missing value rate was above 50%. The peak area was corrected using the Support Vector Regression (SVR) method. After correction, the processed peaks were identified using an in-house database, public databases, and prediction libraries. The final data, after filtering for QC samples with a coefficient of variation (CV) less than 0.3 and a combined score above 0.5, were used for further analysis. The data were then exported to an all_sample_data.xlsx file for statistical analysis.
"],"study_factor":["Group"],"submitter_email":["BS20233050521@cau.edu.cn"],"sample_collection_protocol":["RAW264.7 macrophage cells were cultured in DMEM medium supplemented with 10% fetal bovine serum (FBS) and 1% penicillin-streptomycin at 37°C in a 5% CO2 incubator. When infected, culture medium was changed to DMEM supplemented with 1% FBS without penicillin-streptomycin. Cells were infected with mcr-3-positive or -negative E. coli at a multiplicity of infection (MOI) of 1:20 for 1 hour, with equal volume of PBS treatment used as control, followed by gentamicin treatment to remove extracellular bacteria. Cells were collected at 12h post-infection and immediately frozen in liquid nitrogen for metabolite extraction.
"],"omics_type":["Metabolomics"],"study_design":["LC-MS data","Vanquish","Mus musculus","Immune Evasion","Q Exactive HF-X","untargeted analysis","competitive advantage","untargeted metabolites","mcr-3","Macrophage","RAW-264.7 cell"],"curator_keywords":["LC-MS data","Vanquish","Mus musculus","Immune Evasion","Q Exactive HF-X","untargeted analysis","competitive advantage","untargeted metabolites","mcr-3","Macrophage","RAW-264.7 cell"],"mass_spectrometry_protocol":["Mass spectrometric analysis was performed using a Q-Exactive HF Orbitrap mass spectrometer. The conditions for both ESI+ and ESI- modes were as follows:
Spray Voltage: 3500 V for ESI+, 3200 V for ESI-
Sheath gas (Arb): 30
Auxiliary gas (Arb): 5
Ion transfer tube temperature: 320°C
Vaporizer temperature: 300°C
Scan Range (MS1): 75–1000 Da
Resolution (MS1): 35,000
AGC Target (MS1): 1.00E+06
RF lens (% collision energy): 50
Scan Range (MS2): 75–1000 Da
Resolution (MS2): 17,500
AGC Target (MS2): 2.00E+05
Collision Energy (CE): 30–50 V for both ESI+ and ESI-
Signal Intensity Threshold: 1.00E+06 cps
Top N vs Top speed: 10
Exclusion Duration: 3 s (dynamic exclusion)
"],"metabolite_name":["4-Hydroxyproline","L-Phenylalanine","Dibutyl phthalate","3,4-Dihydroxymandelic acid","Pipecolic acid","Glu-Leu","Glucuronic Acid","Aniline","LPC(18:0/0:0)","L-Norleucine","Spermidine","Sphinganine","Epsilon-caprolactam","13-Docosenamide","Cytosine","1-Phenylethanol","Pyrrolidine","Piperidine","Flavin adenine dinucleotide","2-Picoline","2-Amino-1-phenylethanol","L-Glutamic acid","Cadaverine","Trp-glu","Niacinamide","Adenosine","S-(5-Adenosy)-L-Homocysteine","Phthalic acid","Dopamine","MG(18:2/0:0/0:0)","Choline Alfoscerate","DL-Leucine","L-Proline","Adenosine-5'-diphosphate","Choline","N1-Acetylspermidine","Inosine","3'-Adenylic acid","L-Valine"],"additional_accession":[]},"is_claimable":false,"name":"Macrophage-mediated antibiotic evasion and competitive dominance of mcr-3-carrying Escherichia coli","description":"Mobile colistin resistance (mcr) genes undermine the efficacy of last-line polymyxin antibiotics, and the global prevalence of mcr-3 continues to rise despite reduced colistin use. Here, we show that mcr-3-positive Escherichia coli (E. coli) confers a survival advantage by reprogramming macrophage immunity. MCR-3-mediated lipid A modification blunted TLR4-NF-kappaB signaling, suppressed macrophage reactive oxygen species (ROS) generation, and delayed phagosome-lysosome fusion, allowing mcr-3-positive strains to evade intracellular killing. Integrated transcriptomic and metabolomic analyses revealed extensive immunometabolic rewiring in infected macrophages, including altered glycerophospholipid metabolism and iron homeostasis. Consistently, mcr-3 enhanced bacterial tolerance to ferrous iron stress, likely mitigating host-induced ferroptotic damage. In a mouse co-infection model, mcr-3-positive strains outcompeted isogenic mcr-3-negative strains under antibiotic treatment without any difference in antibiotic susceptibility in vitro. These findings reveal a dual-action mechanism that mcr-3 endows E. coli with both antibiotic resistance and host immune suppression, enabling persistence under antibiotic pressure and highlighting the long-term threat of mcr-3 dissemination even in the absence of polymyxin use.
","dates":{"publication":"2026-05-30","submission":"2025-12-22"},"accession":"MTBLS13550","cross_references":{"HMDB":["HMDB0000162","HMDB0001406","HMDB0001065","HMDB0000159","HMDB0000086","HMDB0000050","HMDB0002107","HMDB0000097","HMDB0000195","HMDB0000725","HMDB0000070","HMDB0000883","HMDB0000073","HMDB0029082","HMDB0001866","HMDB0001645","HMDB0003540","HMDB0028823","HMDB0003012","HMDB0062769","HMDB0244507","HMDB0001276","HMDB0031641","HMDB0034301","HMDB0062203","HMDB0033244","HMDB0001248","HMDB0000630","HMDB0001341","HMDB0000939","HMDB0010384","HMDB0000269","HMDB0002322","HMDB0000148","HMDB0001257","HMDB0061888","HMDB0242115","HMDB0032619","HMDB0000127"]}}