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MB Sample ID: SA328818

Local Sample ID:M_Cefto_3g_95h_E
Subject ID:SU003150
Subject Type:Bacteria
Subject Species:Pseudomonas aeruginosa
Taxonomy ID:287
Genotype Strain:CW41
Age Or Age Range:NA
Weight Or Weight Range:NA
Gender:Not applicable

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Subject:

Subject ID:SU003150
Subject Type:Bacteria
Subject Species:Pseudomonas aeruginosa
Taxonomy ID:287
Genotype Strain:CW41
Age Or Age Range:NA
Weight Or Weight Range:NA
Gender:Not applicable

Factors:

Local Sample IDMB Sample IDFactor Level IDLevel ValueFactor Name
M_Cefto_3g_95h_ESA328818FL039050M_Cefto_3g_95htreatment

Collection:

Collection ID:CO003143
Collection Summary:A hypermutable P. aeruginosa clinical isolate, CW41, was challenged with ceftolozane-tazobactam (Zerbaxa®, MSD, Australia) in the HFIM (C3008-1 cartridges; FiberCell Systems Inc., Frederick, MD, USA), in five biological replicates performed across two studies. The first study, with replicates 1 and 2, was conducted over 167 h and the second study, with replicates 3, 4 and 5, over 215 h. Briefly, the studied isolate was characterized as susceptible to ceftolozane-tazobactam (MIC 4 mg/L), and MDR (i.e. resistant to at least 1 antibiotic from each of ≥3 antibiotic classes) (17-20). The HFIM studies used cation-adjusted Mueller Hinton broth (CAMHB) and agar (CAMHA) [Becton Dickinson & Co., Sparks, MD, USA, with 25.0 mg/L Ca2+ and 12.5 mg/L Mg2+]. Ceftolozane-tazobactam was administered to simulate steady-state concentrations of ceftolozane predicted to occur in the epithelial lining fluid of the lung in patients with CF, following daily doses of 3 g/1.5 g and 6 g/3 g via continuous infusion (10.6 and 21.3 mg/L, respectively) (21-23). Total bacterial populations were quantified on antibiotic-free CAMHA, and resistant subpopulations on CAMHA containing ceftolozane-tazobactam (12 and 20 mg/L).
Sample Type:Bacterial cells

Treatment:

Treatment ID:TR003159
Treatment Summary:Ceftolozane-tazobactam was administered to simulate steady-state concentrations of ceftolozane predicted to occur in the epithelial lining fluid of the lung in patients with CF, following daily doses of 3 g/1.5 g and 6 g/3 g via continuous infusion (10.6 and 21.3 mg/L, respectively) (21-23). Total bacterial populations were quantified on antibiotic-free CAMHA, and resistant subpopulations on CAMHA containing ceftolozane-tazobactam (12 and 20 mg/L).
Treatment Compound:Ceftolozane-tazobactam
Treatment Dose:3 g and 6 g
Treatment Dosevolume:10.6 and 21.3 mg/L

Sample Preparation:

Sampleprep ID:SP003156
Sampleprep Summary:Each sample (25 µL) was added to 100 µL of pre-chilled methanol containing the internal standards (CHAPS, CAPS, TRIS and PIPES) at 1 µM. This mixture was vortexed, and subsequently centrifuged at 14800 x g and 4°C for 10 min .The final supernatant samples containing the extracted extracellular metabolites were stored at -80°C until LC-MS analysis was performed (Figure S2B).
Processing Method:IDEOM
Processing Storage Conditions:-80℃
Extract Storage:-80℃

Combined analysis:

Analysis ID AN004977 AN004978
Analysis type MS MS
Chromatography type HILIC HILIC
Chromatography system Thermo Dionex Ultimate 3000 Thermo Dionex Ultimate 3000
Column Merck SeQuant ZIC-pHILIC (150 x 4.6mm,5um) Merck SeQuant ZIC-pHILIC (150 x 4.6mm,5um)
MS Type ESI ESI
MS instrument type Orbitrap Orbitrap
MS instrument name Thermo Q Exactive Orbitrap Thermo Q Exactive Orbitrap
Ion Mode POSITIVE NEGATIVE
Units peak height peak height

Chromatography:

Chromatography ID:CH003758
Chromatography Summary:ZIC-pHILIC chromatography at pH 9 using pos neg switching
Methods Filename:Metabolomics_pHILIC_Parkville_v1.pdf
Instrument Name:Thermo Dionex Ultimate 3000
Column Name:Merck SeQuant ZIC-pHILIC (150 x 4.6mm,5um)
Column Temperature:25 C
Flow Gradient:0 min - 80%B, 15 min - 50%B, 18 min - 5%B, 21 min - 5%B, 24 min - 80%B, 32 min - 80%B
Flow Rate:0.3 ml/min
Solvent A:20 mM ammonium carbonate
Solvent B:acetonitrile
Washing Buffer:syringe wash 50% IPA
Chromatography Type:HILIC
  
Chromatography ID:CH003759
Chromatography Summary:ZIC-pHILIC chromatography at pH 9 using pos neg switching
Methods Filename:Metabolomics_pHILIC_Parkville_v1.pdf
Instrument Name:Thermo Dionex Ultimate 3000
Column Name:Merck SeQuant ZIC-pHILIC (150 x 4.6mm,5um)
Column Temperature:25 C
Flow Gradient:0 min - 80%B, 15 min - 50%B, 18 min - 5%B, 21 min - 5%B, 24 min - 80%B, 32 min - 80%B
Flow Rate:0.3 ml/min
Solvent A:20 mM ammonium carbonate
Solvent B:acetonitrile
Washing Buffer:syringe wash 50% IPA
Chromatography Type:HILIC

MS:

MS ID:MS004717
Analysis ID:AN004977
Instrument Name:Thermo Q Exactive Orbitrap
Instrument Type:Orbitrap
MS Type:ESI
MS Comments:polarity switching used, resolution 35 k, full scan
Ion Mode:POSITIVE
Capillary Temperature:300 C
Capillary Voltage:3.5 kV
Collision Energy:NA
Dry Gas Flow:50
Dry Gas Temp:120
Ion Source Temperature:120 C
Mass Accuracy:3 ppm
Precursor Type:[M+H]+
Acquisition Parameters File:Metabolomics_pHILIC_Parkville_v1.pdf
Analysis Protocol File:PQMS3-MPMF-WIN-0501_LCMS_data_acquisition_for_untargeted_metabolomics_analysis.pdf
  
MS ID:MS004718
Analysis ID:AN004978
Instrument Name:Thermo Q Exactive Orbitrap
Instrument Type:Orbitrap
MS Type:ESI
MS Comments:polarity switching used, resolution 35 k, full scan
Ion Mode:NEGATIVE
Capillary Temperature:300 C
Capillary Voltage:4 kV
Collision Energy:NA
Dry Gas Flow:50
Dry Gas Temp:120
Ion Source Temperature:120 C
Mass Accuracy:3 ppm
Precursor Type:[M-H]-
Acquisition Parameters File:Metabolomics_pHILIC_Parkville_v1.pdf
Analysis Protocol File:PQMS3-MPMF-WIN-0501_LCMS_data_acquisition_for_untargeted_metabolomics_analysis.pdf
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