{
"METABOLOMICS WORKBENCH":{"STUDY_ID":"ST002868","ANALYSIS_ID":"AN004701","VERSION":"1","CREATED_ON":"September 13, 2023, 3:02 am"},

"PROJECT":{"PROJECT_TITLE":"Pathogenic Staphylococcus epidermidis ICE25 response to skin and blood pH","PROJECT_TYPE":"Proteomic and metabolomic study","PROJECT_SUMMARY":"Staphylococcus epidermidis (SE) is one of the most common bacteria of the human skin microbiota. Despite its role as a commensal, SE has emerged as an opportunistic pathogen, associated with 80% of medical devices related infections. Moreover, these bacteria are extremely difficult to treat due to their ability to form biofilms and accumulate resistance to almost all classes of antimicrobials developed so far. Thus new preventive and therapeutic strategies are urgently needed. In spite of its clinical importance, the molecular mechanisms associated with SE colonisation and disease are still poorly understood. A deeper understanding of the metabolic and cellular processes associated with response to environmental factors characteristic of SE ecological niches in health and disease might provide new clues on colonisation and disease processes. Here we studied the impact of pH conditions, mimicking the skin pH (5.5) and blood pH (7.4), in a S. epidermidis pathogenic strain, belonging to the A/C clonal lineage, by means of next-generation proteomics and 1H NMR-based metabolomics. Moreover, we evaluated the metabolic changes occurring when a sudden pH change arise, simulating the skin barrier break produced by a catheter.","INSTITUTE":"ITQB NOVA","LAST_NAME":"Gonçalves","FIRST_NAME":"Luís","ADDRESS":"Avenida Republica, Oeiras, Not USCanada, 2780-157 Oeiras, Portugal","EMAIL":"lgafeira@itqb.unl.pt","PHONE":"214469464"},

"STUDY":{"STUDY_TITLE":"Pathogenic Staphylococcus epidermidis ICE25 response to skin and blood pH","STUDY_SUMMARY":"Staphylococcus epidermidis (SE) is one of the most common bacteria of the human skin microbiota. Despite its role as a commensal, SE has emerged as an opportunistic pathogen, associated with 80% of medical devices related infections. Moreover, these bacteria are extremely difficult to treat due to their ability to form biofilms and accumulate resistance to almost all classes of antimicrobials developed so far. Thus new preventive and therapeutic strategies are urgently needed. In spite of its clinical importance, the molecular mechanisms associated with SE colonisation and disease are still poorly understood. A deeper understanding of the metabolic and cellular processes associated with response to environmental factors characteristic of SE ecological niches in health and disease might provide new clues on colonisation and disease processes. Here we studied the impact of pH conditions, mimicking the skin pH (5.5) and blood pH (7.4), in a S. epidermidis pathogenic strain, belonging to the A/C clonal lineage, by means of next-generation proteomics and 1H NMR-based metabolomics. Moreover, we evaluated the metabolic changes occurring when a sudden pH change arise, simulating the skin barrier break produced by a catheter.","INSTITUTE":"ITQB NOVA","LAST_NAME":"Gonçalves","FIRST_NAME":"Luís G.","ADDRESS":"Avenida Republica","EMAIL":"lgafeira@itqb.unl.pt","PHONE":"214469464"},

"SUBJECT":{"SUBJECT_TYPE":"Bacteria","SUBJECT_SPECIES":"Staphylococcus epidermidis","TAXONOMY_ID":"1282","GENOTYPE_STRAIN":"Staphylococcus epidermidis ICE25"},
"SUBJECT_SAMPLE_FACTORS":[
{
"Subject ID":"-",
"Sample ID":"SE_1_190530",
"Factors":{"Class":"I77"},
"Additional sample data":{"RAW_FILE_NAME":"SE_1_190530"}
},
{
"Subject ID":"-",
"Sample ID":"SE_10_190530",
"Factors":{"Class":"I57"},
"Additional sample data":{"RAW_FILE_NAME":"SE_10_190530"}
},
{
"Subject ID":"-",
"Sample ID":"SE_11_190530",
"Factors":{"Class":"I57"},
"Additional sample data":{"RAW_FILE_NAME":"SE_11_190530"}
},
{
"Subject ID":"-",
"Sample ID":"SE_18_190530",
"Factors":{"Class":"I55"},
"Additional sample data":{"RAW_FILE_NAME":"SE_18_190530"}
},
{
"Subject ID":"-",
"Sample ID":"SE_23_190530",
"Factors":{"Class":"I55"},
"Additional sample data":{"RAW_FILE_NAME":"SE_23_190530"}
},
{
"Subject ID":"-",
"Sample ID":"SE_24_190530",
"Factors":{"Class":"I55"},
"Additional sample data":{"RAW_FILE_NAME":"SE_24_190530"}
},
{
"Subject ID":"-",
"Sample ID":"SE_4_190530",
"Factors":{"Class":"I57"},
"Additional sample data":{"RAW_FILE_NAME":"SE_4_190530"}
},
{
"Subject ID":"-",
"Sample ID":"SE_5_190530",
"Factors":{"Class":"I77"},
"Additional sample data":{"RAW_FILE_NAME":"SE_5_190530"}
},
{
"Subject ID":"-",
"Sample ID":"SE_6_190530",
"Factors":{"Class":"I77"},
"Additional sample data":{"RAW_FILE_NAME":"SE_6_190530"}
},
{
"Subject ID":"-",
"Sample ID":"SE_7_190530",
"Factors":{"Class":"I57"},
"Additional sample data":{"RAW_FILE_NAME":"SE_7_190530"}
},
{
"Subject ID":"-",
"Sample ID":"SE_9_190530",
"Factors":{"Class":"I77"},
"Additional sample data":{"RAW_FILE_NAME":"SE_9_190530"}
},
{
"Subject ID":"-",
"Sample ID":"SE_a03_190712",
"Factors":{"Class":"I57"},
"Additional sample data":{"RAW_FILE_NAME":"SE_a03_190712"}
},
{
"Subject ID":"-",
"Sample ID":"SE_a04_190712",
"Factors":{"Class":"I55"},
"Additional sample data":{"RAW_FILE_NAME":"SE_a04_190712"}
},
{
"Subject ID":"-",
"Sample ID":"SE_a06_190712",
"Factors":{"Class":"I77"},
"Additional sample data":{"RAW_FILE_NAME":"SE_a06_190712"}
},
{
"Subject ID":"-",
"Sample ID":"SE_a07_190712",
"Factors":{"Class":"I55"},
"Additional sample data":{"RAW_FILE_NAME":"SE_a07_190712"}
},
{
"Subject ID":"-",
"Sample ID":"SE_a09_190712",
"Factors":{"Class":"I57"},
"Additional sample data":{"RAW_FILE_NAME":"SE_a09_190712"}
},
{
"Subject ID":"-",
"Sample ID":"SE_a11_190712",
"Factors":{"Class":"I55"},
"Additional sample data":{"RAW_FILE_NAME":"SE_a11_190712"}
},
{
"Subject ID":"-",
"Sample ID":"SE_a12_190712",
"Factors":{"Class":"I77"},
"Additional sample data":{"RAW_FILE_NAME":"SE_a12_190712"}
},
{
"Subject ID":"-",
"Sample ID":"SE_a13_190712_rep",
"Factors":{"Class":"I57"},
"Additional sample data":{"RAW_FILE_NAME":"SE_a13_190712_rep"}
},
{
"Subject ID":"-",
"Sample ID":"SE_a16_190712",
"Factors":{"Class":"I77"},
"Additional sample data":{"RAW_FILE_NAME":"SE_a16_190712"}
},
{
"Subject ID":"-",
"Sample ID":"SE_a17_190712",
"Factors":{"Class":"I57"},
"Additional sample data":{"RAW_FILE_NAME":"SE_a17_190712"}
},
{
"Subject ID":"-",
"Sample ID":"SE_a18_190712",
"Factors":{"Class":"I55"},
"Additional sample data":{"RAW_FILE_NAME":"SE_a18_190712"}
},
{
"Subject ID":"-",
"Sample ID":"SE_a20_190712",
"Factors":{"Class":"I77"},
"Additional sample data":{"RAW_FILE_NAME":"SE_a20_190712"}
}
],
"COLLECTION":{"COLLECTION_SUMMARY":"The Staphylococcus epidermidis 19N strain was collected from the anterior nares of a healthy person in Portugal in 2001. This strain was previously characterised by whole genome sequencing and belongs to clonal lineage B. A single colony from a S. epidermidis 19N strain culture grown O/N at 37ºC (TSA, BactoTM), was used to pre-inoculate Tryptic Soy Broth (TSB) medium with two different pH (5.5 and 7.4) that was incubated overnight at 37ºC under agitation. Pre-inoculums were adjusted either to pH 5.5 or pH 7.4, with hydrochloric acid. In this work, three pH transitions from pre-inoculum to inoculum were assayed. S. epidermidis pre-inoculums and the growth were performed at medium with pH 7.4, to mimic the blood pH; and pH 5.5, to mimic the skin pH. The pre-inoculum cellular density was adjusted to 0.06 (OD600) (aprox.1.5x108 CFU/mL) and used to inoculate fresh medium in the three conditions depicted in Figure 1, simulating S. epidermidis at skin and blood and a pH shock endured by S. epidermidis during the infection process from skin to blood transition. The cell cultures incubated at 37ºC with 225 rpm were followed by OD600 and recovered at mid-exponential phase for further analysis.","SAMPLE_TYPE":"Staphylococcus epidermidis intracellular","STORAGE_CONDITIONS":"-80℃"},

"TREATMENT":{"TREATMENT_SUMMARY":"n this work, three pH transitions from pre-inoculum to inoculum were assayed. S. epidermidis pre-inoculums and the growth were performed at medium with pH 7.4, to mimic the blood pH; and pH 5.5, to mimic the skin pH. The pre-inoculum cellular density was adjusted to 0.06 (OD600) (aprox.1.5x108 CFU/mL) and used to inoculate fresh medium in the three conditions depicted in Figure 1, simulating S. epidermidis at skin and blood and a pH shock endured by S. epidermidis during the infection process from skin to blood transition. The cell cultures incubated at 37ºC with 225 rpm were followed by OD600 and recovered at mid-exponential phase for further analysis. Pre-inocula were prepared in TSB medium at pH 5.5 or 7.4. Inocula at pH 5.5 was used for the cultures grown at 5.5 (N55) and 7.4 (N57), and the inoculum at pH 7.4 for the culture at the same pH (N77). The cells were harvested at mid-exponential phase."},

"SAMPLEPREP":{"SAMPLEPREP_SUMMARY":"Cells were recovered at mid-exponential phase from 100 mL cultures following a protocol adapted from Somerville & Powers (Somerville and Powers 2014). Eight biological replicates of each independent growth condition were obtained. Cells were harvested by centrifugation at 5000 x g for 5 min at 4ºC. Cells were washed with 20 mM phosphate buffer pH 7.2-7.4 and centrifuged for 1 min at 13,000 rpm. Cell pellet was suspended in the same buffer with a final OD600 of 20 and stored at -80ºC for further metabolite extraction. Cells were thawed in a water bath at room temperature and 750 µL of 60% methanol were added and subjected to three freeze-thaw cycles using liquid nitrogen. Extracted samples were centrifuged at 21,000 g for 5 min at 4ºC. The extraction process on the pellets was repeated twice. The supernatants were kept and stored together at -20ºC overnight and dried in a SpeedVac. Dried samples were dissolved in: 750 µL phosphate buffer (33 mM, pH 7.0 in D2O with 2 mM of sodium azide) with 0.21 mM of 3-(trimethylsilyl)propionic-2,2,3,3-d4 (TSP). The suspensions were centrifuged at 21,000 g for 5 min at 4ºC and the resulting supernatants were then transferred to 5 mm NMR tubes."},

"ANALYSIS":{"ANALYSIS_TYPE":"NMR"},

"NM":{"INSTRUMENT_NAME":"Bruker Avance II+ 800 MHz","INSTRUMENT_TYPE":"FT-NMR","NMR_EXPERIMENT_TYPE":"1D-1H","SPECTROMETER_FREQUENCY":"800","NMR_PROBE":"5 mm TXI-Z H/C/N/-D","NMR_SOLVENT":"D2O","NMR_TUBE_SIZE":"5 mm","SHIMMING_METHOD":"Topshim","CHEMICAL_SHIFT_REF_CPD":"TSP","TEMPERATURE":"25"},

"NMR_METABOLITE_DATA":{
"Units":"nanomoles",

"Data":[{"Metabolite":"2-Hydroxyisobutyrate","SE_18_190530":"2.625","SE_23_190530":"3.9","SE_24_190530":"4.125","SE_a04_190712":"4.275","SE_a07_190712":"2.625","SE_a11_190712":"1.5","SE_a18_190712":"1.425","SE_10_190530":"0.9","SE_11_190530":"0.675","SE_4_190530":"0.825","SE_7_190530":"0.6","SE_a03_190712":"0.675","SE_a09_190712":"3.15","SE_a13_190712_rep":"3.975","SE_a17_190712":"2.7","SE_1_190530":"1.425","SE_5_190530":"1.2","SE_6_190530":"1.2","SE_9_190530":"1.2","SE_a06_190712":"1.35","SE_a12_190712":"2.475","SE_a16_190712":"2.25","SE_a20_190712":"1.425"},{"Metabolite":"3-Hydroxyisovalerate","SE_18_190530":"3.675","SE_23_190530":"3.525","SE_24_190530":"3.525","SE_a04_190712":"9.675","SE_a07_190712":"10.2","SE_a11_190712":"2.85","SE_a18_190712":"2.1","SE_10_190530":"4.95","SE_11_190530":"5.325","SE_4_190530":"4.575","SE_7_190530":"4.8","SE_a03_190712":"10.5","SE_a09_190712":"9.375","SE_a13_190712_rep":"2.1","SE_a17_190712":"10.575","SE_1_190530":"3.3","SE_5_190530":"3.225","SE_6_190530":"4.425","SE_9_190530":"4.35","SE_a06_190712":"10.425","SE_a12_190712":"2.325","SE_a16_190712":"10.875","SE_a20_190712":"2.925"},{"Metabolite":"AMP","SE_18_190530":"4.2","SE_23_190530":"6.375","SE_24_190530":"8.625","SE_a04_190712":"17.1","SE_a07_190712":"14.85","SE_a11_190712":"17.175","SE_a18_190712":"15.45","SE_10_190530":"15.225","SE_11_190530":"11.55","SE_4_190530":"14.25","SE_7_190530":"16.95","SE_a03_190712":"18.375","SE_a09_190712":"13.425","SE_a13_190712_rep":"12.6","SE_a17_190712":"12.825","SE_1_190530":"22.95","SE_5_190530":"21.9","SE_6_190530":"11.775","SE_9_190530":"17.85","SE_a06_190712":"20.025","SE_a12_190712":"18.225","SE_a16_190712":"19.8","SE_a20_190712":"21.075"},{"Metabolite":"Acetate","SE_18_190530":"43.875","SE_23_190530":"35.175","SE_24_190530":"67.425","SE_a04_190712":"65.775","SE_a07_190712":"58.65","SE_a11_190712":"53.175","SE_a18_190712":"59.775","SE_10_190530":"30.3","SE_11_190530":"36.525","SE_4_190530":"43.8","SE_7_190530":"32.775","SE_a03_190712":"39.825","SE_a09_190712":"51.9","SE_a13_190712_rep":"39.75","SE_a17_190712":"37.95","SE_1_190530":"61.125","SE_5_190530":"48.9","SE_6_190530":"54.6","SE_9_190530":"52.275","SE_a06_190712":"71.4","SE_a12_190712":"125.025","SE_a16_190712":"116.025","SE_a20_190712":"75.75"},{"Metabolite":"Adenine","SE_18_190530":"3.15","SE_23_190530":"4.275","SE_24_190530":"4.35","SE_a04_190712":"7.725","SE_a07_190712":"4.425","SE_a11_190712":"4.5","SE_a18_190712":"10.575","SE_10_190530":"3.75","SE_11_190530":"1.875","SE_4_190530":"4.05","SE_7_190530":"6.225","SE_a03_190712":"6.6","SE_a09_190712":"5.7","SE_a13_190712_rep":"5.55","SE_a17_190712":"4.35","SE_1_190530":"7.05","SE_5_190530":"4.575","SE_6_190530":"9.075","SE_9_190530":"5.025","SE_a06_190712":"14.25","SE_a12_190712":"17.4","SE_a16_190712":"18.075","SE_a20_190712":"13.725"},{"Metabolite":"Adenosine","SE_18_190530":"3.15","SE_23_190530":"6.75","SE_24_190530":"11.25","SE_a04_190712":"21.3","SE_a07_190712":"15.225","SE_a11_190712":"16.2","SE_a18_190712":"17.55","SE_10_190530":"14.175","SE_11_190530":"13.2","SE_4_190530":"16.8","SE_7_190530":"14.4","SE_a03_190712":"14.85","SE_a09_190712":"12.9","SE_a13_190712_rep":"11.625","SE_a17_190712":"16.575","SE_1_190530":"22.8","SE_5_190530":"21.075","SE_6_190530":"14.175","SE_9_190530":"21.45","SE_a06_190712":"22.425","SE_a12_190712":"25.275","SE_a16_190712":"20.1","SE_a20_190712":"23.025"},{"Metabolite":"Alanine","SE_18_190530":"10.275","SE_23_190530":"20.1","SE_24_190530":"27.15","SE_a04_190712":"50.7","SE_a07_190712":"47.475","SE_a11_190712":"40.2","SE_a18_190712":"54.675","SE_10_190530":"33.675","SE_11_190530":"23.475","SE_4_190530":"22.275","SE_7_190530":"36.9","SE_a03_190712":"39.9","SE_a09_190712":"24.825","SE_a13_190712_rep":"24","SE_a17_190712":"35.025","SE_1_190530":"111.825","SE_5_190530":"42.9","SE_6_190530":"36.3","SE_9_190530":"51","SE_a06_190712":"73.65","SE_a12_190712":"101.925","SE_a16_190712":"87.975","SE_a20_190712":"83.325"},{"Metabolite":"Arabinose","SE_18_190530":"1.65","SE_23_190530":"4.35","SE_24_190530":"3.6","SE_a04_190712":"9.525","SE_a07_190712":"5.625","SE_a11_190712":"7.275","SE_a18_190712":"8.175","SE_10_190530":"0.75","SE_11_190530":"2.25","SE_4_190530":"3.6","SE_7_190530":"2.85","SE_a03_190712":"3.975","SE_a09_190712":"4.125","SE_a13_190712_rep":"1.65","SE_a17_190712":"3.525","SE_1_190530":"11.175","SE_5_190530":"14.625","SE_6_190530":"19.125","SE_9_190530":"11.25","SE_a06_190712":"13.65","SE_a12_190712":"12.3","SE_a16_190712":"24.225","SE_a20_190712":"13.95"},{"Metabolite":"Asparagine","SE_18_190530":"13.425","SE_23_190530":"19.5","SE_24_190530":"24.6","SE_a04_190712":"19.95","SE_a07_190712":"17.625","SE_a11_190712":"14.1","SE_a18_190712":"19.725","SE_10_190530":"11.175","SE_11_190530":"8.1","SE_4_190530":"14.1","SE_7_190530":"11.85","SE_a03_190712":"18.075","SE_a09_190712":"8.7","SE_a13_190712_rep":"12","SE_a17_190712":"10.05","SE_1_190530":"12","SE_5_190530":"13.95","SE_6_190530":"13.725","SE_9_190530":"18.75","SE_a06_190712":"18.675","S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Id":"C00387"},{"Metabolite":"Histidine","RefMet Name/Standardized Name*":"Histidine","WorkBench":"ME545945","PubChem":"6274","Kegg Id":"C00135"},{"Metabolite":"Isoleucine","RefMet Name/Standardized Name*":"Isoleucine","WorkBench":"ME545946","PubChem":"6306","Kegg Id":"C00407"},{"Metabolite":"Isovalerate","RefMet Name/Standardized Name*":"Isovaleric acid","WorkBench":"ME545947","PubChem":"10430","Kegg Id":"C08262"},{"Metabolite":"Lactate","RefMet Name/Standardized Name*":"Lactic acid","WorkBench":"ME545948","PubChem":"107689","Kegg Id":"C00186"},{"Metabolite":"Leucine","RefMet Name/Standardized Name*":"Leucine","WorkBench":"ME545949","PubChem":"6106","Kegg Id":"C00123"},{"Metabolite":"Lysine","RefMet Name/Standardized Name*":"Lysine","WorkBench":"ME545950","PubChem":"5962","Kegg Id":"C00047"},{"Metabolite":"NAD+","RefMet Name/Standardized Name*":"NAD+","WorkBench":"ME545951","PubChem":"5893","Kegg Id":"C00003"},{"Metabolite":"NADP+","RefMet Name/Standardized Name*":"NADP+","WorkBench":"ME545952","PubChem":"5886","Kegg Id":"C00006"},{"Metabolite":"Nicotinate","RefMet Name/Standardized Name*":"Nicotinic acid","WorkBench":"ME545953","PubChem":"938","Kegg Id":"C00253"},{"Metabolite":"Phenylalanine","RefMet Name/Standardized Name*":"Phenylalanine","WorkBench":"ME545954","PubChem":"6140","Kegg Id":"C00079"},{"Metabolite":"Phosphoenolpyruvic acid","RefMet Name/Standardized Name*":"Phosphoenolpyruvic acid","WorkBench":"ME545955","PubChem":"1005","Kegg Id":"C00074"},{"Metabolite":"sn-Glycero-3-phosphocholine","RefMet Name/Standardized Name*":"sn-Glycero-3-phosphocholine","WorkBench":"ME545965","PubChem":"71920","Kegg Id":"C00670"},{"Metabolite":"Succinate","RefMet Name/Standardized Name*":"Succinic acid","WorkBench":"ME545956","PubChem":"1110","Kegg Id":"C00042"},{"Metabolite":"Sucrose","RefMet Name/Standardized Name*":"Sucrose","WorkBench":"ME545957","PubChem":"5988","Kegg Id":"C00089"},{"Metabolite":"Threonine","RefMet Name/Standardized Name*":"Threonine","WorkBench":"ME545958","PubChem":"6288","Kegg Id":"C00188"},{"Metabolite":"Tryptophan","RefMet Name/Standardized Name*":"Tryptophan","WorkBench":"ME545959","PubChem":"6305","Kegg Id":"C00078"},{"Metabolite":"Tyrosine","RefMet Name/Standardized Name*":"Tyrosine","WorkBench":"ME545960","PubChem":"6057","Kegg Id":"C00082"},{"Metabolite":"UMP","RefMet Name/Standardized Name*":"UMP","WorkBench":"ME545961","PubChem":"6030","Kegg Id":"C00105"},{"Metabolite":"Uracil","RefMet Name/Standardized Name*":"Uracil","WorkBench":"ME545962","PubChem":"1174","Kegg Id":"C00106"},{"Metabolite":"Uridine","RefMet Name/Standardized Name*":"Uridine","WorkBench":"ME545963","PubChem":"6029","Kegg Id":"C00299"},{"Metabolite":"Valine","RefMet Name/Standardized Name*":"Valine","WorkBench":"ME545964","PubChem":"6287","Kegg Id":"C00183"}]
}

}