#METABOLOMICS WORKBENCH estancliffe_20210128_125700 DATATRACK_ID:2432 STUDY_ID:ST001853 ANALYSIS_ID:AN003004 PROJECT_ID:000000
VERSION             	1
CREATED_ON             	June 29, 2021, 1:55 pm
#PROJECT
PR:PROJECT_TITLE                 	Longitudinal Metabolomics of Human Plasma Reveals Robust Prognostic Markers of
PR:PROJECT_TITLE                 	COVID-19 Disease Severity
PR:PROJECT_SUMMARY               	There is an urgent need to identify which COVID-19 patients will develop
PR:PROJECT_SUMMARY               	life-threatening illness so that medical resources can be optimally allocated
PR:PROJECT_SUMMARY               	and rapid treatment can be administered early in the disease course, when
PR:PROJECT_SUMMARY               	clinical management is most effective. To aid in the prognostic classification
PR:PROJECT_SUMMARY               	of disease severity, we perform untargeted metabolomics on plasma from 339
PR:PROJECT_SUMMARY               	patients, with samples collected at six longitudinal time points. Using the
PR:PROJECT_SUMMARY               	temporal metabolic profiles and machine learning, we build a predictive model of
PR:PROJECT_SUMMARY               	disease severity. We discover that a panel of metabolites measured at the time
PR:PROJECT_SUMMARY               	of study entry successfully determine disease severity. Through analysis of
PR:PROJECT_SUMMARY               	longitudinal samples, we confirm that the majority of these markers are directly
PR:PROJECT_SUMMARY               	related to disease progression and that their levels are restored to baseline
PR:PROJECT_SUMMARY               	upon disease recovery. Finally, we validate that these metabolites are also
PR:PROJECT_SUMMARY               	altered in a hamster model of COVID-19. Our results indicate that metabolic
PR:PROJECT_SUMMARY               	changes associated with COVID-19 severity can be effectively used to stratify
PR:PROJECT_SUMMARY               	patients and inform resource allocation during the pandemic.
PR:INSTITUTE                     	Washington University, St. Louis
PR:DEPARTMENT                    	Chemistry
PR:LABORATORY                    	Patti
PR:LAST_NAME                     	Patti
PR:FIRST_NAME                    	Gary
PR:ADDRESS                       	McMillen Chemistry Laboratory, Washington University 1 Brookings Dr @ Throop
PR:ADDRESS                       	Drive, Rm 102, St. Louis, MO 63130-4899
PR:EMAIL                         	gjpattij@wustl.edu
PR:PHONE                         	314-935-3512
#STUDY
ST:STUDY_TITLE                   	Longitudinal Metabolomics of Human Plasma Reveals Robust Prognostic Markers of
ST:STUDY_TITLE                   	COVID-19 Disease Severity
ST:STUDY_SUMMARY                 	There is an urgent need to identify which COVID-19 patients will develop
ST:STUDY_SUMMARY                 	life-threatening illness so that medical resources can be optimally allocated
ST:STUDY_SUMMARY                 	and rapid treatment can be administered early in the disease course, when
ST:STUDY_SUMMARY                 	clinical management is most effective. To aid in the prognostic classification
ST:STUDY_SUMMARY                 	of disease severity, we perform untargeted metabolomics on plasma from 339
ST:STUDY_SUMMARY                 	patients, with samples collected at six longitudinal time points. Using the
ST:STUDY_SUMMARY                 	temporal metabolic profiles and machine learning, we build a predictive model of
ST:STUDY_SUMMARY                 	disease severity. We discover that a panel of metabolites measured at the time
ST:STUDY_SUMMARY                 	of study entry successfully determine disease severity. Through analysis of
ST:STUDY_SUMMARY                 	longitudinal samples, we confirm that the majority of these markers are directly
ST:STUDY_SUMMARY                 	related to disease progression and that their levels are restored to baseline
ST:STUDY_SUMMARY                 	upon disease recovery. Finally, we validate that these metabolites are also
ST:STUDY_SUMMARY                 	altered in a hamster model of COVID-19. Our results indicate that metabolic
ST:STUDY_SUMMARY                 	changes associated with COVID-19 severity can be effectively used to stratify
ST:STUDY_SUMMARY                 	patients and inform resource allocation during the pandemic.
ST:INSTITUTE                     	Washington University, St. Louis
ST:DEPARTMENT                    	Chemistry
ST:LABORATORY                    	Patti
ST:LAST_NAME                     	Patti
ST:FIRST_NAME                    	Gary
ST:ADDRESS                       	McMillen Chemistry Laboratory, Washington University 1 Brookings Dr @ Throop
ST:ADDRESS                       	Drive, Rm 102, St. Louis, MO 63130-4899
ST:EMAIL                         	gjpattij@wustl.edu
ST:NUM_GROUPS                    	3
ST:TOTAL_SUBJECTS                	56
ST:NUM_FEMALES                   	56
ST:PHONE                         	314-935-3512
#SUBJECT
SU:SUBJECT_TYPE                  	Mammal
SU:SUBJECT_SPECIES               	Mesocricetus auratus
#SUBJECT_SAMPLE_FACTORS:         	SUBJECT(optional)[tab]SAMPLE[tab]FACTORS(NAME:VALUE pairs separated by |)[tab]Raw file names and additional sample data
SUBJECT_SAMPLE_FACTORS           	-	P H-433	Group:Influenza | Day post infection:4	RAW_FILE_NAME=P H-433_polar_pos.mzML,P H-433_polar_neg.mzML,P H-433_lipid_pos.mzML,P H-433_lipid_neg.mzML
SUBJECT_SAMPLE_FACTORS           	-	P H-457	Group:Influenza | Day post infection:4	RAW_FILE_NAME=P H-457_polar_pos.mzML,P H-457_polar_neg.mzML,P H-457_lipid_pos.mzML,P H-457_lipid_neg.mzML
SUBJECT_SAMPLE_FACTORS           	-	P H-458	Group:Influenza | Day post infection:4	RAW_FILE_NAME=P H-458_polar_pos.mzML,P H-458_polar_neg.mzML,P H-458_lipid_pos.mzML,P H-458_lipid_neg.mzML
SUBJECT_SAMPLE_FACTORS           	-	P H-459	Group:SARS-CoV-2 | Day post infection:14	RAW_FILE_NAME=P H-459_polar_pos.mzML,P H-459_polar_neg.mzML,P H-459_lipid_pos.mzML,P H-459_lipid_neg.mzML
SUBJECT_SAMPLE_FACTORS           	-	P H-460	Group:Influenza | Day post infection:4	RAW_FILE_NAME=P H-460_polar_pos.mzML,P H-460_polar_neg.mzML,P H-460_lipid_pos.mzML,P H-460_lipid_neg.mzML
SUBJECT_SAMPLE_FACTORS           	-	P H-461	Group:Influenza | Day post infection:6	RAW_FILE_NAME=P H-461_polar_pos.mzML,P H-461_polar_neg.mzML,P H-461_lipid_pos.mzML,P H-461_lipid_neg.mzML
SUBJECT_SAMPLE_FACTORS           	-	P H-462	Group:Influenza | Day post infection:6	RAW_FILE_NAME=P H-462_polar_pos.mzML,P H-462_polar_neg.mzML,P H-462_lipid_pos.mzML,P H-462_lipid_neg.mzML
SUBJECT_SAMPLE_FACTORS           	-	P H-463	Group:SARS-CoV-2 | Day post infection:6	RAW_FILE_NAME=P H-463_polar_pos.mzML,P H-463_polar_neg.mzML,P H-463_lipid_pos.mzML,P H-463_lipid_neg.mzML
SUBJECT_SAMPLE_FACTORS           	-	P H-464	Group:Mock | Day post infection:4	RAW_FILE_NAME=P H-464_polar_pos.mzML,P H-464_polar_neg.mzML,P H-464_lipid_pos.mzML,P H-464_lipid_neg.mzML
SUBJECT_SAMPLE_FACTORS           	-	P H-466	Group:Influenza | Day post infection:4	RAW_FILE_NAME=P H-466_polar_pos.mzML,P H-466_polar_neg.mzML,P H-466_lipid_pos.mzML,P H-466_lipid_neg.mzML
SUBJECT_SAMPLE_FACTORS           	-	P H-467	Group:Influenza | Day post infection:14	RAW_FILE_NAME=P H-467_polar_pos.mzML,P H-467_polar_neg.mzML,P H-467_lipid_pos.mzML,P H-467_lipid_neg.mzML
SUBJECT_SAMPLE_FACTORS           	-	P H-468	Group:SARS-CoV-2 | Day post infection:6	RAW_FILE_NAME=P H-468_polar_pos.mzML,P H-468_polar_neg.mzML,P H-468_lipid_pos.mzML,P H-468_lipid_neg.mzML
SUBJECT_SAMPLE_FACTORS           	-	P H-469 -1	Group:SARS-CoV-2 | Day post infection:2	RAW_FILE_NAME=P H-469 -1_polar_pos.mzML,P H-469 -1_polar_neg.mzML,P H-469 -1_lipid_pos.mzML,P H-469 -1_lipid_neg.mzML
SUBJECT_SAMPLE_FACTORS           	-	P H-469 -2	Group:SARS-CoV-2 | Day post infection:4	RAW_FILE_NAME=P H-469 -2 _polar_pos.mzML,P H-469 -2 _polar_neg.mzML,P H-469 -2 _lipid_pos.mzML,P H-469 -2 _lipid_neg.mzML
SUBJECT_SAMPLE_FACTORS           	-	P H-470	Group:Mock | Day post infection:14	RAW_FILE_NAME=P H-470_polar_pos.mzML,P H-470_polar_neg.mzML,P H-470_lipid_pos.mzML,P H-470_lipid_neg.mzML
SUBJECT_SAMPLE_FACTORS           	-	P H-471	Group:SARS-CoV-2 | Day post infection:4	RAW_FILE_NAME=P H-471_polar_pos.mzML,P H-471_polar_neg.mzML,P H-471_lipid_pos.mzML,P H-471_lipid_neg.mzML
SUBJECT_SAMPLE_FACTORS           	-	P H-472	Group:Mock | Day post infection:4	RAW_FILE_NAME=P H-472_polar_pos.mzML,P H-472_polar_neg.mzML,P H-472_lipid_pos.mzML,P H-472_lipid_neg.mzML
SUBJECT_SAMPLE_FACTORS           	-	P H-473	Group:Influenza | Day post infection:14	RAW_FILE_NAME=P H-473_polar_pos.mzML,P H-473_polar_neg.mzML,P H-473_lipid_pos.mzML,P H-473_lipid_neg.mzML
SUBJECT_SAMPLE_FACTORS           	-	P H-474	Group:Influenza | Day post infection:2	RAW_FILE_NAME=P H-474_polar_pos.mzML,P H-474_polar_neg.mzML,P H-474_lipid_pos.mzML,P H-474_lipid_neg.mzML
SUBJECT_SAMPLE_FACTORS           	-	P H-475	Group:Mock | Day post infection:14	RAW_FILE_NAME=P H-475_polar_pos.mzML,P H-475_polar_neg.mzML,P H-475_lipid_pos.mzML,P H-475_lipid_neg.mzML
SUBJECT_SAMPLE_FACTORS           	-	P H-476	Group:Mock | Day post infection:4	RAW_FILE_NAME=P H-476_polar_pos.mzML,P H-476_polar_neg.mzML,P H-476_lipid_pos.mzML,P H-476_lipid_neg.mzML
SUBJECT_SAMPLE_FACTORS           	-	P H-477	Group:Influenza | Day post infection:6	RAW_FILE_NAME=P H-477_polar_pos.mzML,P H-477_polar_neg.mzML,P H-477_lipid_pos.mzML,P H-477_lipid_neg.mzML
SUBJECT_SAMPLE_FACTORS           	-	P H-478	Group:Influenza | Day post infection:2	RAW_FILE_NAME=P H-478_polar_pos.mzML,P H-478_polar_neg.mzML,P H-478_lipid_pos.mzML,P H-478_lipid_neg.mzML
SUBJECT_SAMPLE_FACTORS           	-	P H-479	Group:SARS-CoV-2 | Day post infection:14	RAW_FILE_NAME=P H-479_polar_pos.mzML,P H-479_polar_neg.mzML,P H-479_lipid_pos.mzML,P H-479_lipid_neg.mzML
SUBJECT_SAMPLE_FACTORS           	-	P H-480	Group:Mock | Day post infection:4	RAW_FILE_NAME=P H-480_polar_pos.mzML,P H-480_polar_neg.mzML,P H-480_lipid_pos.mzML,P H-480_lipid_neg.mzML
SUBJECT_SAMPLE_FACTORS           	-	P H-481	Group:Mock | Day post infection:14	RAW_FILE_NAME=P H-481_polar_pos.mzML,P H-481_polar_neg.mzML,P H-481_lipid_pos.mzML,P H-481_lipid_neg.mzML
SUBJECT_SAMPLE_FACTORS           	-	P H-482	Group:Mock | Day post infection:4	RAW_FILE_NAME=P H-482_polar_pos.mzML,P H-482_polar_neg.mzML,P H-482_lipid_pos.mzML,P H-482_lipid_neg.mzML
SUBJECT_SAMPLE_FACTORS           	-	P H-483	Group:Influenza | Day post infection:14	RAW_FILE_NAME=P H-483_polar_pos.mzML,P H-483_polar_neg.mzML,P H-483_lipid_pos.mzML,P H-483_lipid_neg.mzML
SUBJECT_SAMPLE_FACTORS           	-	P H-484	Group:SARS-CoV-2 | Day post infection:14	RAW_FILE_NAME=P H-484_polar_pos.mzML,P H-484_polar_neg.mzML,P H-484_lipid_pos.mzML,P H-484_lipid_neg.mzML
SUBJECT_SAMPLE_FACTORS           	-	P H-485	Group:SARS-CoV-2 | Day post infection:14	RAW_FILE_NAME=P H-485_polar_pos.mzML,P H-485_polar_neg.mzML,P H-485_lipid_pos.mzML,P H-485_lipid_neg.mzML
SUBJECT_SAMPLE_FACTORS           	-	P H-486	Group:Influenza | Day post infection:6	RAW_FILE_NAME=P H-486_polar_pos.mzML,P H-486_polar_neg.mzML,P H-486_lipid_pos.mzML,P H-486_lipid_neg.mzML
SUBJECT_SAMPLE_FACTORS           	-	P H-487	Group:SARS-CoV-2 | Day post infection:4	RAW_FILE_NAME=P H-487_polar_pos.mzML,P H-487_polar_neg.mzML,P H-487_lipid_pos.mzML,P H-487_lipid_neg.mzML
SUBJECT_SAMPLE_FACTORS           	-	P H-488	Group:SARS-CoV-2 | Day post infection:4	RAW_FILE_NAME=P H-488_polar_pos.mzML,P H-488_polar_neg.mzML,P H-488_lipid_pos.mzML,P H-488_lipid_neg.mzML
SUBJECT_SAMPLE_FACTORS           	-	P H-489	Group:Mock | Day post infection:14	RAW_FILE_NAME=P H-489_polar_pos.mzML,P H-489_polar_neg.mzML,P H-489_lipid_pos.mzML,P H-489_lipid_neg.mzML
SUBJECT_SAMPLE_FACTORS           	-	P H-490	Group:Mock | Day post infection:14	RAW_FILE_NAME=P H-490_polar_pos.mzML,P H-490_polar_neg.mzML,P H-490_lipid_pos.mzML,P H-490_lipid_neg.mzML
SUBJECT_SAMPLE_FACTORS           	-	P H-491	Group:Influenza | Day post infection:6	RAW_FILE_NAME=P H-491_polar_pos.mzML,P H-491_polar_neg.mzML,P H-491_lipid_pos.mzML,P H-491_lipid_neg.mzML
SUBJECT_SAMPLE_FACTORS           	-	P H-492	Group:Influenza | Day post infection:2	RAW_FILE_NAME=P H-492_polar_pos.mzML,P H-492_polar_neg.mzML,P H-492_lipid_pos.mzML,P H-492_lipid_neg.mzML
SUBJECT_SAMPLE_FACTORS           	-	P H-495	Group:SARS-CoV-2 | Day post infection:6	RAW_FILE_NAME=P H-495_polar_pos.mzML,P H-495_polar_neg.mzML,P H-495_lipid_pos.mzML,P H-495_lipid_neg.mzML
SUBJECT_SAMPLE_FACTORS           	-	P H-496	Group:Influenza | Day post infection:14	RAW_FILE_NAME=P H-496_polar_pos.mzML,P H-496_polar_neg.mzML,P H-496_lipid_pos.mzML,P H-496_lipid_neg.mzML
SUBJECT_SAMPLE_FACTORS           	-	P H-497	Group:Influenza | Day post infection:14	RAW_FILE_NAME=P H-497_polar_pos.mzML,P H-497_polar_neg.mzML,P H-497_lipid_pos.mzML,P H-497_lipid_neg.mzML
SUBJECT_SAMPLE_FACTORS           	-	P H-498	Group:Influenza | Day post infection:4	RAW_FILE_NAME=P H-498_polar_pos.mzML,P H-498_polar_neg.mzML,P H-498_lipid_pos.mzML,P H-498_lipid_neg.mzML
SUBJECT_SAMPLE_FACTORS           	-	P H-499	Group:Influenza | Day post infection:14	RAW_FILE_NAME=P H-499_polar_pos.mzML,P H-499_polar_neg.mzML,P H-499_lipid_pos.mzML,P H-499_lipid_neg.mzML
SUBJECT_SAMPLE_FACTORS           	-	P H-501	Group:Mock | Day post infection:14	RAW_FILE_NAME=P H-501_polar_pos.mzML,P H-501_polar_neg.mzML,P H-501_lipid_pos.mzML,P H-501_lipid_neg.mzML
SUBJECT_SAMPLE_FACTORS           	-	P H-502	Group:Influenza | Day post infection:2	RAW_FILE_NAME=P H-502_polar_pos.mzML,P H-502_polar_neg.mzML,P H-502_lipid_pos.mzML,P H-502_lipid_neg.mzML
SUBJECT_SAMPLE_FACTORS           	-	P H-504	Group:SARS-CoV-2 | Day post infection:6	RAW_FILE_NAME=P H-504_polar_pos.mzML,P H-504_polar_neg.mzML,P H-504_lipid_pos.mzML,P H-504_lipid_neg.mzML
SUBJECT_SAMPLE_FACTORS           	-	P H-505	Group:Influenza | Day post infection:2	RAW_FILE_NAME=P H-505_polar_pos.mzML,P H-505_polar_neg.mzML,P H-505_lipid_pos.mzML,P H-505_lipid_neg.mzML
SUBJECT_SAMPLE_FACTORS           	-	P H-506	Group:Influenza | Day post infection:2	RAW_FILE_NAME=P H-506_polar_pos.mzML,P H-506_polar_neg.mzML,P H-506_lipid_pos.mzML,P H-506_lipid_neg.mzML
SUBJECT_SAMPLE_FACTORS           	-	P H-507	Group:Mock | Day post infection:4	RAW_FILE_NAME=P H-507_polar_pos.mzML,P H-507_polar_neg.mzML,P H-507_lipid_pos.mzML,P H-507_lipid_neg.mzML
SUBJECT_SAMPLE_FACTORS           	-	P H-509	Group:SARS-CoV-2 | Day post infection:6	RAW_FILE_NAME=P H-509_polar_pos.mzML,P H-509_polar_neg.mzML,P H-509_lipid_pos.mzML,P H-509_lipid_neg.mzML
SUBJECT_SAMPLE_FACTORS           	-	P H-510	Group:SARS-CoV-2 | Day post infection:14	RAW_FILE_NAME=P H-510_polar_pos.mzML,P H-510_polar_neg.mzML,P H-510_lipid_pos.mzML,P H-510_lipid_neg.mzML
SUBJECT_SAMPLE_FACTORS           	-	P H-511	Group:SARS-CoV-2 | Day post infection:14	RAW_FILE_NAME=P H-511_polar_pos.mzML,P H-511_polar_neg.mzML,P H-511_lipid_pos.mzML,P H-511_lipid_neg.mzML
SUBJECT_SAMPLE_FACTORS           	-	P H-512	Group:SARS-CoV-2 | Day post infection:6	RAW_FILE_NAME=P H-512_polar_pos.mzML,P H-512_polar_neg.mzML,P H-512_lipid_pos.mzML,P H-512_lipid_neg.mzML
SUBJECT_SAMPLE_FACTORS           	-	P H-513	Group:SARS-CoV-2 | Day post infection:2	RAW_FILE_NAME=P H-513_polar_pos.mzML,P H-513_polar_neg.mzML,P H-513_lipid_pos.mzML,P H-513_lipid_neg.mzML
SUBJECT_SAMPLE_FACTORS           	-	P H-514	Group:SARS-CoV-2 | Day post infection:4	RAW_FILE_NAME=P H-514_polar_pos.mzML,P H-514_polar_neg.mzML,P H-514_lipid_pos.mzML,P H-514_lipid_neg.mzML
SUBJECT_SAMPLE_FACTORS           	-	P H-515	Group:Influenza | Day post infection:6	RAW_FILE_NAME=P H-515_polar_pos.mzML,P H-515_polar_neg.mzML,P H-515_lipid_pos.mzML,P H-515_lipid_neg.mzML
SUBJECT_SAMPLE_FACTORS           	-	P H-516	Group:SARS-CoV-2 | Day post infection:2	RAW_FILE_NAME=P H-516_polar_pos.mzML,P H-516_polar_neg.mzML,P H-516_lipid_pos.mzML,P H-516_lipid_neg.mzML
#COLLECTION
CO:COLLECTION_SUMMARY            	Outbred female LVG golden Syrian hamsters (6-8 weeks of age) were obtained from
CO:COLLECTION_SUMMARY            	Charles River Laboratories (Kingston, NY). The hamsters were anesthetized by
CO:COLLECTION_SUMMARY            	intraperitoneal injection of a mixture of ketamine and xylazine prior to
CO:COLLECTION_SUMMARY            	intranasal inoculation with 0.1 mL of 1e5 plaque-forming units (PFU) of
CO:COLLECTION_SUMMARY            	SARS-CoV-2 (WA-1) or H1N1 influenza A virus (A/California/04/2009). On day 2, 4,
CO:COLLECTION_SUMMARY            	6, and 14 after infection, 3-6 anesthetized hamsters per infection group were
CO:COLLECTION_SUMMARY            	euthanized by exsanguination followed by intracardiac injection of veterinary
CO:COLLECTION_SUMMARY            	euthanasia solution (SleepAway; Fort Dodge). Plasma samples were treated by
CO:COLLECTION_SUMMARY            	exposure to germicidal UV-C light.
CO:SAMPLE_TYPE                   	Blood (plasma)
#TREATMENT
TR:TREATMENT_SUMMARY             	The hamsters were anesthetized by intraperitoneal injection of a mixture of
TR:TREATMENT_SUMMARY             	ketamine and xylazine prior to intranasal inoculation with 0.1 mL of 1e5
TR:TREATMENT_SUMMARY             	plaque-forming units (PFU) of SARS-CoV-2 (WA-1) or H1N1 influenza A virus
TR:TREATMENT_SUMMARY             	(A/California/04/2009).
#SAMPLEPREP
SP:SAMPLEPREP_SUMMARY            	Hamster plasma samples were diluted 1:4 with methanol (v/v), vortexed for 30
SP:SAMPLEPREP_SUMMARY            	seconds, and incubated at -20 C for 2 hours. Samples were centrifuged for 10
SP:SAMPLEPREP_SUMMARY            	minutes at 13,500 x g at 4°C and supernatant was transferred to a new
SP:SAMPLEPREP_SUMMARY            	centrifuge tube, concentrated, and stored at -80 C until reconstitution. Hamster
SP:SAMPLEPREP_SUMMARY            	plasma was thawed on ice. A 50 µL aliquot was transferred onto the
SP:SAMPLEPREP_SUMMARY            	solid-phase-extraction (SPE)-system CAPTIVA-EMR Lipid 96-wellplate (Agilent
SP:SAMPLEPREP_SUMMARY            	Technologies) before addition of 250 µL of acetonitrile containing 1% formic
SP:SAMPLEPREP_SUMMARY            	acid (v/v) and 10 µM internal standard (consisting of uniformly 13C and 15N
SP:SAMPLEPREP_SUMMARY            	labeled amino acids from Cambridge Isotope Laboratories, Inc). The samples were
SP:SAMPLEPREP_SUMMARY            	mixed for 1 min at 360 rpm on an orbital shaker at room temperature prior to a
SP:SAMPLEPREP_SUMMARY            	10 min incubation period at 4 C. Afterwards, 200 µL 80% acetonitrile in
SP:SAMPLEPREP_SUMMARY            	water (v/v) were added to the samples. The samples were mixed on an orbital
SP:SAMPLEPREP_SUMMARY            	shaker (360 rpm) for an additional 10 min at room temperature. The samples were
SP:SAMPLEPREP_SUMMARY            	then eluted into a 96-deepwell collection plate by centrifugation (10 min, 57 x
SP:SAMPLEPREP_SUMMARY            	g, 4 C followed by 2 min, 1000 x g, 4 C). Polar eluates were stored at -80 C
SP:SAMPLEPREP_SUMMARY            	until the day of LC/MS analysis. The SPE-plates were then washed twice with 500
SP:SAMPLEPREP_SUMMARY            	µL 80% acetonitrile in water (v/v). Lipids still bound to the SPE-material were
SP:SAMPLEPREP_SUMMARY            	then released into a second elution plate, in two elution steps applying 2x 500
SP:SAMPLEPREP_SUMMARY            	µL 1:1 methyl tert-butyl ether:methanol (v/v) onto the SPE cartridge and
SP:SAMPLEPREP_SUMMARY            	centrifuging for 2 min at 1000 g and 4 C. The combined eluates were dried under
SP:SAMPLEPREP_SUMMARY            	a stream of nitrogen (Biotage SPE Dry Evaporation System) at room temperature
SP:SAMPLEPREP_SUMMARY            	and reconstituted with 100 µL 1:1 2-propanol:methanol (v/v) prior to LC/MS
SP:SAMPLEPREP_SUMMARY            	analysis.
#CHROMATOGRAPHY
CH:CHROMATOGRAPHY_SUMMARY        	An aliquot of 2 µL of lipid extract was subjected to LC/MS analysis by using an
CH:CHROMATOGRAPHY_SUMMARY        	Agilent 1290 Infinity II LC-system coupled to an Agilent 6545 Q-TOF mass
CH:CHROMATOGRAPHY_SUMMARY        	spectrometer with a dual Agilent Jet Stream electrospray ionization source.
CH:CHROMATOGRAPHY_SUMMARY        	Lipids were separated on an Acquity UPLC® HSS T3 column (2.1 x 150 mm, 1.8 µm)
CH:CHROMATOGRAPHY_SUMMARY        	including an Acquity UPLC® HSS T3 VanGuard Pre-Column (2.1 x 5mm, 1.8 µm) at a
CH:CHROMATOGRAPHY_SUMMARY        	temperature of 60 C and a flow rate of 250 µLmin-1. The mobile phases
CH:CHROMATOGRAPHY_SUMMARY        	consisted of A: 60% acetonitrile, 40% water, 0.1% formic acid, 10 mM ammonium
CH:CHROMATOGRAPHY_SUMMARY        	formate, 2.5 µM medronic acid, and B: 90% 2-propanol, 10% acetonitrile, 0.1%
CH:CHROMATOGRAPHY_SUMMARY        	formic acid, 10 mM ammonium formate (dissolved in 1 mL water). The following
CH:CHROMATOGRAPHY_SUMMARY        	linear gradient was used: 0-2 min, 30% B; 17 min, 75% B; 20 min, 85%; 23-26 min,
CH:CHROMATOGRAPHY_SUMMARY        	100% B; 26, 30% B followed by a re-equilibration phase of 5 min.
CH:CHROMATOGRAPHY_TYPE           	Reversed phase
CH:INSTRUMENT_NAME               	Agilent 1290 Infinity II
CH:COLUMN_NAME                   	Acquity UPLC® HSS T3 column (2.1 x 150 mm, 1.8 µm) including an Acquity UPLC®
CH:COLUMN_NAME                   	HSS T3 VanGuard Pre-Column (2.1 x 5mm, 1.8 µm)
#ANALYSIS
AN:ANALYSIS_TYPE                 	MS
#MS
MS:INSTRUMENT_NAME               	Agilent 6545 QTOF
MS:INSTRUMENT_TYPE               	QTOF
MS:MS_TYPE                       	ESI
MS:ION_MODE                      	NEGATIVE
MS:MS_COMMENTS                   	gas temperature 250 C, drying gas flow 11 L*min-1, nebulizer pressure 35 psi,
MS:MS_COMMENTS                   	sheath gas temperature 300 C, sheath gas flow 12 Lmin-1, VCap 3000 V,
MS:MS_COMMENTS                   	nozzle voltage 500 V, Fragmentor 160 V, Skimmer 65 V, Oct 1 RF Vpp 750 V, and
MS:MS_COMMENTS                   	m/z range 50-1700. Data were acquired under continuous reference mass correction
MS:MS_COMMENTS                   	at m/z 121.0509 and 922.0890 in positive ion mode and m/z 119.0363 and 966.0007
MS:MS_COMMENTS                   	in negative ion mode. Polar metabolite identifications were supported by
MS:MS_COMMENTS                   	matching the retention time, accurate mass, and MS/MS fragmentation data to our
MS:MS_COMMENTS                   	in-house retention time and MS/MS library created from authentic reference
MS:MS_COMMENTS                   	standards (Mass Spectrometry Metabolite Library supplied by IROA Technologies,
MS:MS_COMMENTS                   	Millipore Sigma, St. Louis, MO, USA) and online MS/MS libraries (Human
MS:MS_COMMENTS                   	Metabolome Database (HMDB, https://hmdb.ca), Mass Bank of North America (MoNA,
MS:MS_COMMENTS                   	https://mona.fiehnlab.ucdavis.edu/), and mzCloud (https://mzcloud.org). Lipid
MS:MS_COMMENTS                   	iterative MS/MS data were annotated with the Agilent Lipid Annotator software.
MS:MS_COMMENTS                   	All data files were then analyzed in Skyline (Version 20.1.0.155) to obtain peak
MS:MS_COMMENTS                   	areas.
#MS_METABOLITE_DATA
MS_METABOLITE_DATA:UNITS	Relative Intensity
MS_METABOLITE_DATA_START
Samples	P H-433	P H-457	P H-458	P H-459	P H-460	P H-461	P H-462	P H-463	P H-464	P H-466	P H-467	P H-468	P H-469 -1	P H-469 -2	P H-470	P H-471	P H-472	P H-473	P H-474	P H-475	P H-476	P H-477	P H-478	P H-479	P H-480	P H-481	P H-482	P H-483	P H-484	P H-485	P H-486	P H-487	P H-488	P H-489	P H-490	P H-491	P H-492	P H-495	P H-496	P H-497	P H-498	P H-499	P H-501	P H-502	P H-504	P H-505	P H-506	P H-507	P H-509	P H-510	P H-511	P H-512	P H-513	P H-514	P H-515	P H-516
Factors	Group:Influenza | Day post infection:4	Group:Influenza | Day post infection:4	Group:Influenza | Day post infection:4	Group:SARS-CoV-2 | Day post infection:14	Group:Influenza | Day post infection:4	Group:Influenza | Day post infection:6	Group:Influenza | Day post infection:6	Group:SARS-CoV-2 | Day post infection:6	Group:Mock | Day post infection:4	Group:Influenza | Day post infection:4	Group:Influenza | Day post infection:14	Group:SARS-CoV-2 | Day post infection:6	Group:SARS-CoV-2 | Day post infection:2	Group:SARS-CoV-2 | Day post infection:4	Group:Mock | Day post infection:14	Group:SARS-CoV-2 | Day post infection:4	Group:Mock | Day post infection:4	Group:Influenza | Day post infection:14	Group:Influenza | Day post infection:2	Group:Mock | Day post infection:14	Group:Mock | Day post infection:4	Group:Influenza | Day post infection:6	Group:Influenza | Day post infection:2	Group:SARS-CoV-2 | Day post infection:14	Group:Mock | Day post infection:4	Group:Mock | Day post infection:14	Group:Mock | Day post infection:4	Group:Influenza | Day post infection:14	Group:SARS-CoV-2 | Day post infection:14	Group:SARS-CoV-2 | Day post infection:14	Group:Influenza | Day post infection:6	Group:SARS-CoV-2 | Day post infection:4	Group:SARS-CoV-2 | Day post infection:4	Group:Mock | Day post infection:14	Group:Mock | Day post infection:14	Group:Influenza | Day post infection:6	Group:Influenza | Day post infection:2	Group:SARS-CoV-2 | Day post infection:6	Group:Influenza | Day post infection:14	Group:Influenza | Day post infection:14	Group:Influenza | Day post infection:4	Group:Influenza | Day post infection:14	Group:Mock | Day post infection:14	Group:Influenza | Day post infection:2	Group:SARS-CoV-2 | Day post infection:6	Group:Influenza | Day post infection:2	Group:Influenza | Day post infection:2	Group:Mock | Day post infection:4	Group:SARS-CoV-2 | Day post infection:6	Group:SARS-CoV-2 | Day post infection:14	Group:SARS-CoV-2 | Day post infection:14	Group:SARS-CoV-2 | Day post infection:6	Group:SARS-CoV-2 | Day post infection:2	Group:SARS-CoV-2 | Day post infection:4	Group:Influenza | Day post infection:6	Group:SARS-CoV-2 | Day post infection:2
Cer_NS d18:2_16:0	68374	65549	61692	97078	52750	123486	100931	102544	70498	626204	55611	72375	194827	49265	83494	110670	299743	71989	209161	58616	254835	233257	113172	100782	415292	175207	87579	138990	57039	91384	288342	252931	115782	311563	76973	3905	237059	73758	116033	195411	73115	113357	68763	206502	167559	404090	132536	25281	194436	101028	72893	28425	130360	124234	112588	99824
PE 16:0_18:2	452595	287439	203791	973719	521200	770970	353824	773061	757250	373608	310846	742228	562208	710019	482139	1020264	1061455	704792	1065997	344578	371210	1038583	874134	1156423	676663	1011527	379839	674583	712173	381219	857317	220732	680996	1387477	453818	4805	958897	488504	786587	338232	310868	868044	643263	666904	664147	308608	573628	534865	450243	921397	538965	325012	968536	920223	878967	694608
PE 16:0_20:4	58843	31924	43207	110559	71229	72179	42209	102436	110001	44834	30237	91804	93777	119130	45673	129235	106268	90305	176930	46079	40360	111168	107893	149707	86312	102380	57093	49232	98023	48353	100656	134984	108039	125856	50422	89544	121642	73934	83754	52482	45329	108383	83040	85372	85638	59784	83235	57734	76565	113991	57839	51065	111927	107537	106450	96229
MS_METABOLITE_DATA_END
#METABOLITES
METABOLITES_START
metabolite_name
Cer_NS d18:2_16:0
PE 16:0_18:2
PE 16:0_20:4
METABOLITES_END
#END