Summary of Study ST003308

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 PR002058. The data can be accessed directly via it's Project DOI: 10.21228/M8KZ5Q 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.

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Study ID	ST003308
Study Title	An untargeted metabolic profiling strategy for the dissection of the oat (Avena sativa L.) plant innate immune response to various Pseudomonas syringae pathovars
Study Type	Plant Metabolomics
Study Summary	One of the most important characteristics that plants utilise to successfully defend themselves is the ability to rapidly identify potential threats in the surrounding environment. Plants rely on the perception of microbe-derived molecular pattern chemicals for this recognition, which initiates a number of induced defence reactions that ultimately increase plant resistance. The metabolome acts as a metabolic fingerprint of the biochemical activities that take place in a biological system under particular conditions and therefore provides a functional readout of the cellular mechanisms involved in a biological system. In this study, an untargeted metabolomics approach was applied to decipher the biochemical processes involved in oat plant defence responses to inoculation with various pathovars of Pseudomonas syringae (pathogenic and non-pathogenic on oat) such as P. syringae pv. coronafaciens (Ps-c), -pv. tabaci (Ps-t), -pv. tomato DC3000 (DC3000) and -pv. tomato DC3000 hrcC mutant (hrcC−) and thereby identify signatory markers that are involved in host or nonhost defence responses. At the seedling growth stage, metabolic alterations in the Dunnart oat cultivar (tolerant to Ps-c) in response to inoculation with the respective Pseudomonas syringae pathovars were examined. Following inoculation, plants were monitored for symptom development and harvested at 2-, 4- and 6 d.p.i.. Methanolic metabolite extracts were prepared, and ultra-high-performance liquid chromatography (UHPLC) connected to a qTOF high-definition mass spectrometer was used to analyse the extracts. Chemometric modelling and multivariate statistical analysis revealed host- and time-related metabolic alterations that point to host and nonhost interactions in response to bacterial inoculation/infection. Metabolic profiles from further multivariate data analyses revealed a range of metabolite classes involved in the respective defence responses, including phenolic amides, saponins, phenolic acids, flavonoids, fatty acids, amino acids and alkaloids. The findings in this study allowed the elucidation of metabolic changes involved in oat defence responses to a range of pathovars of Pseudomonas syringae and ultimately contributed to a more comprehensive view of the oat plant metabolism under biotic stress during host vs nonhost interactions. Note: Initial optimisation tests revealed that the majority of extractable metabolites ionised better in the ESI (–) mode; thus, only these data sets are provided.
Institute	University of Johannesburg
Department	Biochemistry
Laboratory	Plant Metabolomics
Last Name	Pretorius
First Name	Chanel
Address	Corner Kingsway and University Road, Auckland Park, Johannesburg, 2092
Email	chanelpretorius5@outlook.com
Phone	0660328667
Submit Date	2024-07-04
Raw Data Available	Yes
Raw Data File Type(s)	raw(Waters)
Analysis Type Detail	LC-MS
Release Date	2024-07-30
Release Version	1

Select appropriate tab below to view additional metadata details:

Project:

Project ID:	PR002058
Project DOI:	doi: 10.21228/M8KZ5Q
Project Title:	An untargeted metabolic profiling strategy for the dissection of the oat (Avena sativa L.) plant innate immune response to various Pseudomonas syringae pathovars
Project Type:	Untargeted MS
Project Summary:	One of the most important characteristics that plants utilise to successfully defend themselves is the ability to rapidly identify potential threats in the surrounding environment. Plants rely on the perception of microbe-derived molecular pattern chemicals for this recognition, which initiates a number of induced defence reactions that ultimately increase plant resistance. The metabolome acts as a metabolic fingerprint of the biochemical activities that take place in a biological system under particular conditions and therefore provides a functional readout of the cellular mechanisms involved in a biological system. In this study, an untargeted metabolomics approach was applied to decipher the biochemical processes involved in oat plant defence responses to inoculation with various pathovars of Pseudomonas syringae (pathogenic and non-pathogenic on oat) such as P. syringae pv. coronafaciens (Ps-c), -pv. tabaci (Ps-t), -pv. tomato DC3000 (DC3000) and -pv. tomato DC3000 hrcC mutant (hrcC−) and thereby identify signatory markers that are involved in host or nonhost defence responses. At the seedling growth stage, metabolic alterations in the Dunnart oat cultivar (tolerant to Ps-c) in response to inoculation with the respective Pseudomonas syringae pathovars were examined. Following inoculation, plants were monitored for symptom development and harvested at 2-, 4- and 6 d.p.i.. Methanolic metabolite extracts were prepared, and ultra-high-performance liquid chromatography (UHPLC) connected to a qTOF high-definition mass spectrometer was used to analyse the extracts. Chemometric modelling and multivariate statistical analysis revealed host- and time-related metabolic alterations that point to host and nonhost interactions in response to bacterial inoculation/infection. Metabolic profiles from further multivariate data analyses revealed a range of metabolite classes involved in the respective defence responses, including phenolic amides, saponins, phenolic acids, flavonoids, fatty acids, amino acids and alkaloids. The findings in this study allowed the elucidation of metabolic changes involved in oat defence responses to a range of pathovars of Pseudomonas syringae and ultimately contributed to a more comprehensive view of the oat plant metabolism under biotic stress during host vs nonhost interactions.
Institute:	University of Johannesburg
Department:	Biochemistry
Laboratory:	Plant Metabolomics
Last Name:	Pretorius
First Name:	Chanel
Address:	Cnr Barry Hertzog and Napier Road, Richmond, Johannesburg
Email:	chanelpretorius5@outlook.com
Phone:	0660328667

Subject:

Subject ID:	SU003429
Subject Type:	Plant
Subject Species:	Avena sativa L.
Taxonomy ID:	4498

Factors:

Subject type: Plant; Subject species: Avena sativa L. (Factor headings shown in green)

mb_sample_id	local_sample_id	Sample source	Treatment	Days post inoculation
SA358699	DUNHCDAY2B1R3b	Dunnart oat cultivar	Control	2 dpi
SA358700	DUNHCDAY2B1R1b	Dunnart oat cultivar	Control	2 dpi
SA358701	DUNHCDAY2B2R1b	Dunnart oat cultivar	Control	2 dpi
SA358702	DUNHCDAY2B2R2b	Dunnart oat cultivar	Control	2 dpi
SA358703	DUNHCDAY2B2R3b	Dunnart oat cultivar	Control	2 dpi
SA358704	DUNHCDAY2B3R1b	Dunnart oat cultivar	Control	2 dpi
SA358705	DUNHCDAY2B3R2b	Dunnart oat cultivar	Control	2 dpi
SA358706	DUNHCDAY2B3R3b	Dunnart oat cultivar	Control	2 dpi
SA358707	DUNHCDAY2B1R2b	Dunnart oat cultivar	Control	2 dpi
SA358708	DUNHCDAY4B3R1b	Dunnart oat cultivar	Control	4 dpi
SA358709	DUNHCDAY4B2R2b	Dunnart oat cultivar	Control	4 dpi
SA358710	DUNHCDAY4B1R1b	Dunnart oat cultivar	Control	4 dpi
SA358711	DUNHCDAY4B1R3b	Dunnart oat cultivar	Control	4 dpi
SA358712	DUNHCDAY4B2R1b	Dunnart oat cultivar	Control	4 dpi
SA358713	DUNHCDAY4B1R2b	Dunnart oat cultivar	Control	4 dpi
SA358714	DUNHCDAY4B2R3b	Dunnart oat cultivar	Control	4 dpi
SA358715	DUNHCDAY4B3R2b	Dunnart oat cultivar	Control	4 dpi
SA358716	DUNHCDAY4B3R3b	Dunnart oat cultivar	Control	4 dpi
SA358717	DUNHCDAY6B1R3b	Dunnart oat cultivar	Control	6 dpi
SA358718	DUNHCDAY6B1R1b	Dunnart oat cultivar	Control	6 dpi
SA358719	DUNHCDAY6B2R1b	Dunnart oat cultivar	Control	6 dpi
SA358720	DUNHCDAY6B2R2b	Dunnart oat cultivar	Control	6 dpi
SA358721	DUNHCDAY6B2R3b	Dunnart oat cultivar	Control	6 dpi
SA358722	DUNHCDAY6B3R1b	Dunnart oat cultivar	Control	6 dpi
SA358723	DUNHCDAY6B3R2b	Dunnart oat cultivar	Control	6 dpi
SA358724	DUNHCDAY6B3R3b	Dunnart oat cultivar	Control	6 dpi
SA358725	DUNHCDAY6B1R2b	Dunnart oat cultivar	Control	6 dpi
SA358726	DUNDC3000DAY2B1R2b	Dunnart oat cultivar	DC3000	2 dpi
SA358727	DUNDC3000DAY2B1R1b	Dunnart oat cultivar	DC3000	2 dpi
SA358728	DUNDC3000DAY2B3R3b	Dunnart oat cultivar	DC3000	2 dpi
SA358729	DUNDC3000DAY2B2R3b	Dunnart oat cultivar	DC3000	2 dpi
SA358730	DUNDC3000DAY2B3R2b	Dunnart oat cultivar	DC3000	2 dpi
SA358731	DUNDC3000DAY2B3R1b	Dunnart oat cultivar	DC3000	2 dpi
SA358732	DUNDC3000DAY2B1R3b	Dunnart oat cultivar	DC3000	2 dpi
SA358733	DUNDC3000DAY2B2R2b	Dunnart oat cultivar	DC3000	2 dpi
SA358734	DUNDC3000DAY2B2R1b	Dunnart oat cultivar	DC3000	2 dpi
SA358735	DUNDC3000DAY4B1R1b	Dunnart oat cultivar	DC3000	4 dpi
SA358736	DUNDC3000DAY4B3R3b	Dunnart oat cultivar	DC3000	4 dpi
SA358737	DUNDC3000DAY4B3R2b	Dunnart oat cultivar	DC3000	4 dpi
SA358738	DUNDC3000DAY4B3R1b	Dunnart oat cultivar	DC3000	4 dpi
SA358739	DUNDC3000DAY4B2R3b	Dunnart oat cultivar	DC3000	4 dpi
SA358740	DUNDC3000DAY4B2R2b	Dunnart oat cultivar	DC3000	4 dpi
SA358741	DUNDC3000DAY4B2R1b	Dunnart oat cultivar	DC3000	4 dpi
SA358742	DUNDC3000DAY4B1R3b	Dunnart oat cultivar	DC3000	4 dpi
SA358743	DUNDC3000DAY4B1R2b	Dunnart oat cultivar	DC3000	4 dpi
SA358744	DUNDC3000DAY6B3R2b	Dunnart oat cultivar	DC3000	6 dpi
SA358745	DUNDC3000DAY6B3R3b	Dunnart oat cultivar	DC3000	6 dpi
SA358746	DUNDC3000DAY6B1R1b	Dunnart oat cultivar	DC3000	6 dpi
SA358747	DUNDC3000DAY6B1R3b	Dunnart oat cultivar	DC3000	6 dpi
SA358748	DUNDC3000DAY6B2R1b	Dunnart oat cultivar	DC3000	6 dpi
SA358749	DUNDC3000DAY6B2R2b	Dunnart oat cultivar	DC3000	6 dpi
SA358750	DUNDC3000DAY6B2R3b	Dunnart oat cultivar	DC3000	6 dpi
SA358751	DUNDC3000DAY6B3R1b	Dunnart oat cultivar	DC3000	6 dpi
SA358752	DUNDC3000DAY6B1R2b	Dunnart oat cultivar	DC3000	6 dpi
SA358753	DUNHRCCDAY2B3R1b	Dunnart oat cultivar	HrcC mutant	2 dpi
SA358754	DUNHRCCDAY2B3R3b	Dunnart oat cultivar	HrcC mutant	2 dpi
SA358755	DUNHRCCDAY2B3R2b	Dunnart oat cultivar	HrcC mutant	2 dpi
SA358756	DUNHRCCDAY2B2R1b	Dunnart oat cultivar	HrcC mutant	2 dpi
SA358757	DUNHRCCDAY2B2R3b	Dunnart oat cultivar	HrcC mutant	2 dpi
SA358758	DUNHRCCDAY2B2R2b	Dunnart oat cultivar	HrcC mutant	2 dpi
SA358759	DUNHRCCDAY2B1R3b	Dunnart oat cultivar	HrcC mutant	2 dpi
SA358760	DUNHRCCDAY2B1R2b	Dunnart oat cultivar	HrcC mutant	2 dpi
SA358761	DUNHRCCDAY2B1R1b	Dunnart oat cultivar	HrcC mutant	2 dpi
SA358762	DUNHRCCDAY4B3R3b	Dunnart oat cultivar	HrcC mutant	4 dpi
SA358763	DUNHRCCDAY4B3R2b	Dunnart oat cultivar	HrcC mutant	4 dpi
SA358764	DUNHRCCDAY4B3R1b	Dunnart oat cultivar	HrcC mutant	4 dpi
SA358765	DUNHRCCDAY4B2R3b	Dunnart oat cultivar	HrcC mutant	4 dpi
SA358766	DUNHRCCDAY4B2R2b	Dunnart oat cultivar	HrcC mutant	4 dpi
SA358767	DUNHRCCDAY4B1R3b	Dunnart oat cultivar	HrcC mutant	4 dpi
SA358768	DUNHRCCDAY4B1R2b	Dunnart oat cultivar	HrcC mutant	4 dpi
SA358769	DUNHRCCDAY4B1R1b	Dunnart oat cultivar	HrcC mutant	4 dpi
SA358770	DUNHRCCDAY4B2R1b	Dunnart oat cultivar	HrcC mutant	4 dpi
SA358771	DUNHRCCDAY6B3R1b	Dunnart oat cultivar	HrcC mutant	6 dpi
SA358772	DUNHRCCDAY6B3R3b	Dunnart oat cultivar	HrcC mutant	6 dpi
SA358773	DUNHRCCDAY6B3R2b	Dunnart oat cultivar	HrcC mutant	6 dpi
SA358774	DUNHRCCDAY6B2R2b	Dunnart oat cultivar	HrcC mutant	6 dpi
SA358775	DUNHRCCDAY6B2R3b	Dunnart oat cultivar	HrcC mutant	6 dpi
SA358776	DUNHRCCDAY6B2R1b	Dunnart oat cultivar	HrcC mutant	6 dpi
SA358777	DUNHRCCDAY6B1R3b	Dunnart oat cultivar	HrcC mutant	6 dpi
SA358778	DUNHRCCDAY6B1R2b	Dunnart oat cultivar	HrcC mutant	6 dpi
SA358779	DUNHRCCDAY6B1R1b	Dunnart oat cultivar	HrcC mutant	6 dpi
SA358780	DUNPSCDAY2B3R1b	Dunnart oat cultivar	Pseudomonas coronafaciens	2 dpi
SA358781	DUNPSCDAY2B3R3b	Dunnart oat cultivar	Pseudomonas coronafaciens	2 dpi
SA358782	DUNPSCDAY2B3R2b	Dunnart oat cultivar	Pseudomonas coronafaciens	2 dpi
SA358783	DUNPSCDAY2B1R3b	Dunnart oat cultivar	Pseudomonas coronafaciens	2 dpi
SA358784	DUNPSCDAY2B2R3b	Dunnart oat cultivar	Pseudomonas coronafaciens	2 dpi
SA358785	DUNPSCDAY2B2R2b	Dunnart oat cultivar	Pseudomonas coronafaciens	2 dpi
SA358786	DUNPSCDAY2B1R1b	Dunnart oat cultivar	Pseudomonas coronafaciens	2 dpi
SA358787	DUNPSCDAY2B2R1b	Dunnart oat cultivar	Pseudomonas coronafaciens	2 dpi
SA358788	DUNPSCDAY2B1R2b	Dunnart oat cultivar	Pseudomonas coronafaciens	2 dpi
SA358789	DUNPSCDAY4B1R1b	Dunnart oat cultivar	Pseudomonas coronafaciens	4 dpi
SA358790	DUNPSCDAY4B1R2b	Dunnart oat cultivar	Pseudomonas coronafaciens	4 dpi
SA358791	DUNPSCDAY4B1R3b	Dunnart oat cultivar	Pseudomonas coronafaciens	4 dpi
SA358792	DUNPSCDAY4B2R1b	Dunnart oat cultivar	Pseudomonas coronafaciens	4 dpi
SA358793	DUNPSCDAY4B2R3b	Dunnart oat cultivar	Pseudomonas coronafaciens	4 dpi
SA358794	DUNPSCDAY4B3R1b	Dunnart oat cultivar	Pseudomonas coronafaciens	4 dpi
SA358795	DUNPSCDAY4B3R2b	Dunnart oat cultivar	Pseudomonas coronafaciens	4 dpi
SA358796	DUNPSCDAY4B3R3b	Dunnart oat cultivar	Pseudomonas coronafaciens	4 dpi
SA358797	DUNPSCDAY4B2R2b	Dunnart oat cultivar	Pseudomonas coronafaciens	4 dpi
SA358798	DUNPSCDAY6B3R1b	Dunnart oat cultivar	Pseudomonas coronafaciens	6 dpi

Showing page 1 of 2 Results: 1 2 Next Showing results 1 to 100 of 135

Collection:

Collection ID:	CO003422
Collection Summary:	At the three-leaf growth stage the leaves were treated by spraying with the Ps-c, Ps-t, DC3000 and hrcC− bacterial suspensions (prepared in PBS with 0.1 % Tween 20), diluted to OD600 ≈0.3. The leaves were then harvested at 2, 4 and 6 days post inoculation (d.p.i.). The leaf material was quenched with liquid nitrogen before being crushed into powder with a mortar and pestle. The samples were weighed (1 g) suspended in 80% cold (4 °C) aqueous analytical grade methanol at a 1:10 m/v ratio.
Sample Type:	Leaves

Treatment:

Treatment ID:	TR003438
Treatment Summary:	The leaves were treated by spraying with the Ps-c, Ps-t, DC3000 and hrcC− bacterial suspensions (prepared in PBS with 0.1 % Tween 20), diluted to OD600 ≈0.3. The vehicle control (VC) plants were sprayed with a solution free of the bacteria and the healthy control (HC) groups were untreated (i.e., not sprayed with either solution), all grown under normal growth conditions.

Sample Preparation:

Sampleprep ID:	SP003436
Sampleprep Summary:	The harvested leaf material was quenched with liquid nitrogen before being crushed into powder with a mortar and pestle. The samples were weighed (1 g) suspended in 80% cold (4 °C) aqueous analytical grade methanol (Romil's Chemistry, Cambridge, UK).at a 1:10 m/v ratio. The mixture was then homogenised with a probe sonicator (Bandelin Sonopuls, Berlin, Germany) at 55% power for 10 seconds per sample. To avoid cross-contamination, equipment was cleansed between each sample. The homogenates were centrifuged at 5100 x g for 20 min at 4 °C in a benchtop centrifuge after which the supernatants were kept and concentrated by evaporating the methanol under vacuum to approximately 1 mL using a rotary evaporator set to 55 °C. The concentrated samples were transferred to 2 mL microcentrifuge tubes and dried in a centrifugal evaporator under vacuum. The dried extracts were then reconstituted by dissolving in 500 μL of 50% aqueous methanol (MilliQ deionised water and LC-grade methanol (Romil, Cambridge, UK). The samples were subsequently filtered through nylon syringe filters (0.22 μm) into chromatography vials fitted with 500 μL inserts, capped, and kept at 4 °C until analysis.

Sampleprep ID:

SP003436

Sampleprep Summary:

The harvested leaf material was quenched with liquid nitrogen before being crushed into powder with a mortar and pestle. The samples were weighed (1 g) suspended in 80% cold (4 °C) aqueous analytical grade methanol (Romil's Chemistry, Cambridge, UK).at a 1:10 m/v ratio. The mixture was then homogenised with a probe sonicator (Bandelin Sonopuls, Berlin, Germany) at 55% power for 10 seconds per sample. To avoid cross-contamination, equipment was cleansed between each sample. The homogenates were centrifuged at 5100 x g for 20 min at 4 °C in a benchtop centrifuge after which the supernatants were kept and concentrated by evaporating the methanol under vacuum to approximately 1 mL using a rotary evaporator set to 55 °C. The concentrated samples were transferred to 2 mL microcentrifuge tubes and dried in a centrifugal evaporator under vacuum. The dried extracts were then reconstituted by dissolving in 500 μL of 50% aqueous methanol (MilliQ deionised water and LC-grade methanol (Romil, Cambridge, UK). The samples were subsequently filtered through nylon syringe filters (0.22 μm) into chromatography vials fitted with 500 μL inserts, capped, and kept at 4 °C until analysis.

Combined analysis:

Analysis ID	AN005420
Analysis type	MS
Chromatography type	Reversed phase
Chromatography system	Waters Acquity
Column	Waters ACQUITY UPLC HSS T3 (150 x 2.1mm,1.8um)
MS Type	ESI
MS instrument type	QTOF
MS instrument name	Waters Synapt G1
Ion Mode	NEGATIVE
Units	m/z

Chromatography:

Chromatography ID:	CH004109
Chromatography Summary:	The run was set to 30 min per injection with an elution gradient carried out via a binary solvent system consisting of 0.1% aqueous formic acid with 2.5% isopropanol (solvent A) and 0.1% formic acid in acetonitrile with isopropanol (Romil, Cambridge, UK; solvent B) at a flow rate of 0.4 mL/min. Concave chromatography gradient carried out via binary solvent system.
Instrument Name:	Waters Acquity
Column Name:	Waters ACQUITY UPLC HSS T3 (150 x 2.1mm,1.8um)
Column Temperature:	60
Flow Gradient:	The initial conditions were 95% A and 5% B and held for 1 min. A gradient was applied to change the chromatographic conditions to 10% A and 90% B at 25 min; and changed to 5% A and 95% B at 25.10 min. These conditions were held for 2 min and then changed to the initial conditions at 28 min. The analytical column was allowed to equilibrate for 2 min before each subsequent injection.
Flow Rate:	0.4 mL/min
Solvent A:	97.5% MilliQ water/2.5% isopropanol; 0.1% formic acid
Solvent B:	97,5% acetonitrile/2.5% isopropanol; 0.1% formic acid
Chromatography Type:	Reversed phase

MS:

MS ID:	MS005146
Analysis ID:	AN005420
Instrument Name:	Waters Synapt G1
Instrument Type:	QTOF
MS Type:	ESI
MS Comments:	A high definition SYNAPT G1 quadrupole time-of-flight (qTOF) mass spectrometry system (Waters Corporation, Manchester, UK) was coupled to the UHPLC chromatography system to detect metabolites and acquire data in both positive and negative electrospray ionisation (ESI) operation modes. The controlling software was MassLynx XSTM (Waters, Manchester, UK). A reference calibrant, leucine encephalin (554.2615 Da) was used as the ‘lockmass’ calibrant and allowed for typical mass accuracies between 1 to 3 mDa. The respective capillary and sampling cone voltages were set as 2.5 kV and 30 V. The desolvation temperature used was 450 °C, with the source temperature set to 120 °C, cone gas flow was set to 50 L/h, and the desolvation gas flow set to 550 L/h. An m/z range of 50–1200 was set with a scan time of 0.1 s. The desolvation-, collision- and cone gas used at a flow rate of 700 L/h was high-purity nitrogen. Data was acquired using five different collision energies (MSE), ramping from 0-50 eV to cause fragmentation of the initial ions to ensure that information regarding the fragmentation of the respective compounds could be obtained for downstream structural elucidation and metabolite annotation.
Ion Mode:	NEGATIVE