Summary of Study ST002933

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 PR001824. The data can be accessed directly via it's Project DOI: 10.21228/M8Z132 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.

Study ID	ST002933
Study Title	Coral endosymbiont growth is enhanced by metabolic interactions with bacteria
Study Summary	Bacteria are key contributors to microalgae resource acquisition, competitive performance, and functional diversity, but their potential metabolic interactions with coral microalgal endosymbionts (Symbiodiniaceae) have been largely overlooked. Here, we show that altering the bacterial composition of two widespread Symbiodiniaceae species, during their free-living stage, results in a significant shift in their cellular metabolism. Indeed, the abundance of monosaccharides and the key phytohormone indole-3-acetic acid (IAA) were correlated with the presence of specific bacteria, including members of the Labrenzia (Roseibium) and Marinobacter genera. Single-cell stable isotope tracking revealed that these two bacterial genera are involved in reciprocal exchanges of carbon and nitrogen with Symbiodiniaceae. We identified the provision of IAA by Labrenzia and Marinobacter, and this metabolite caused a significant growth enhancement of Symbiodiniaceae. By unravelling these interkingdom interactions, our work demonstrates how specific bacterial associates fundamentally govern Symbiodiniaceae fitness.
Institute	University of Technology Sydney
Last Name	Matthews
First Name	Jennifer
Address	15 Broadway, Sydney, NSW, 2007, Australia
Email	jennifer.matthews@uts.edu.au
Phone	0432404274
Submit Date	2023-10-14
Raw Data Available	Yes
Raw Data File Type(s)	qgd
Analysis Type Detail	GC-MS
Release Date	2023-11-10
Release Version	1

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Project:

Project ID:	PR001824
Project DOI:	doi: 10.21228/M8Z132
Project Title:	Coral endosymbiont growth is enhanced by metabolic interactions with bacteria
Project Summary:	Bacteria are key contributors to microalgae resource acquisition, competitive performance, and functional diversity, but their potential metabolic interactions with coral microalgal endosymbionts (Symbiodiniaceae) have been largely overlooked. Here, we show that altering the bacterial composition of two widespread Symbiodiniaceae species, during their free-living stage, results in a significant shift in their cellular metabolism. Indeed, the abundance of monosaccharides and the key phytohormone indole-3-acetic acid (IAA) were correlated with the presence of specific bacteria, including members of the Labrenzia (Roseibium) and Marinobacter genera. Single-cell stable isotope tracking revealed that these two bacterial genera are involved in reciprocal exchanges of carbon and nitrogen with Symbiodiniaceae. We identified the provision of IAA by Labrenzia and Marinobacter, and this metabolite caused a significant growth enhancement of Symbiodiniaceae. By unravelling these interkingdom interactions, our work demonstrates how specific bacterial associates fundamentally govern Symbiodiniaceae fitness.
Institute:	University of Technology Sydney
Last Name:	Matthews
First Name:	Jennifer
Address:	15 Broadway, Sydney, NSW, 2007, Australia
Email:	jennifer.matthews@uts.edu.au
Phone:	0432404274
Funding Source:	HFSP

Subject:

Subject ID:	SU003046
Subject Type:	Cultured cells
Subject Species:	Symbiodiniaceae

Factors:

Subject type: Cultured cells; Subject species: Symbiodiniaceae (Factor headings shown in green)

mb_sample_id	local_sample_id	Genotype	Treatment
SA318288	Sample 1	Breviolum minutum	Ab+Tx
SA318289	Sample 2	Breviolum minutum	Ab+Tx
SA318290	Sample 3	Breviolum minutum	Ab+Tx
SA318291	Sample 22	Breviolum minutum	Ab+Tx
SA318292	Sample 19	Breviolum minutum	Ab+Tx
SA318293	Sample 24	Breviolum minutum	Untreated
SA318294	Sample 10	Breviolum minutum	Untreated
SA318295	Sample 20	Breviolum minutum	Untreated
SA318296	Sample 6	Breviolum minutum	Untreated
SA318297	Sample 9	Breviolum minutum	Untreated
SA318298	Sample 26	Symbiodinium microadriaticum	Ab+Tx
SA318299	Sample 29	Symbiodinium microadriaticum	Ab+Tx
SA318300	Sample 23	Symbiodinium microadriaticum	Ab+Tx
SA318301	Sample 7	Symbiodinium microadriaticum	Ab+Tx
SA318302	Sample 17	Symbiodinium microadriaticum	Ab+Tx
SA318303	Sample 16	Symbiodinium microadriaticum	Untreated
SA318304	Sample 15	Symbiodinium microadriaticum	Untreated
SA318305	Sample 8	Symbiodinium microadriaticum	Untreated
SA318306	Sample 4	Symbiodinium microadriaticum	Untreated
SA318307	Sample 28	Symbiodinium microadriaticum	Untreated

Showing results 1 to 20 of 20

Showing results 1 to 20 of 20

Collection:

Collection ID:	CO003039
Collection Summary:	Sampled from Symbiodiniaceae cultures in F/2 media
Sample Type:	Cultured cells

Treatment:

Treatment ID:	TR003055
Treatment Summary:	Symbiodiniaceae cultures Two Symbiodiniaceae cultures were targeted from existing stocks at the University of Technology Sydney, Symbiodinium microadriaticum (ITS2: A1, culture ID: RT61), and Breviolum minutum (ITS2: B1, culture ID: RT2, CCMP2463) as preliminary trials allowed these cultures to be maintained for extended periods in an extracellular bacteria-free state. Each Symbiodiniaceae species was sub-cultured (n = 10 per Symbiodiniaceae species) by adding 10 mL of original cultures in 90 mL of autoclaved and filter sterilised (0.22 µm) artificial seawater (ASW) and F/2 media. Cultures were grown for one month (to achieve a minimum cell density of 106 cells/mL) at 26˚C with an irradiance of 85 ± 15 µmol photons m-2 s-1 (Philips TLD 18W/54 fluorescent tubes, 10 000 K on a 12h:12h light:dark cycle). Before use, cells were centrifuged at 700 × g for 10 mins at 26˚C and rinsed twice with ASW to remove residual media solution. Cells were resuspended in 100 mL ASW + F/2 media in sterile culture flasks. Untreated cultures Untreated Symbiodiniaceae cultures (n = 5) were maintained as above alongside the Ab+Tx treatments (below). Antibiotic treatment (AbTx) Each antibiotic treatment subculture (n = 5 per Symbiodiniaceae species) was provided with TritonX-100 detergent added to a final concentration of 20 µg/mL and placed on a shaker at mid speed for 30 s. All cultures (Ab+Tx treatment and untreated) were immediately centrifuged at 700 × g for 10 mins at 26˚C, and the supernatant discarded. Cells were rinsed in 20 mL ASW and centrifuged at 700 x g for 10 mins at 26˚C. Cells were transferred to sterile culture flasks and resuspended in 9 ml ASW+F/2. An aliquot of 1 mL custom antibiotic mix (Penicillin at 31.25 µg mL-1, Streptomycin and Kanamycin at 50 µg mL-1, and Neomycin, Ciprofloxacin and Ampicillin all at 100 µg mL-1; Ab+Tx) was added to each antibiotic treatment flask (and 1 mL ASW added to each untreated flask), and flasks were replaced in the incubator. After 48 hours, 90 mL ASW + F/2 was added to all cultures. Cultures were allowed to recover for 5 days prior to metabolism quenching.

Treatment ID:

TR003055

Treatment Summary:

Symbiodiniaceae cultures Two Symbiodiniaceae cultures were targeted from existing stocks at the University of Technology Sydney, Symbiodinium microadriaticum (ITS2: A1, culture ID: RT61), and Breviolum minutum (ITS2: B1, culture ID: RT2, CCMP2463) as preliminary trials allowed these cultures to be maintained for extended periods in an extracellular bacteria-free state. Each Symbiodiniaceae species was sub-cultured (n = 10 per Symbiodiniaceae species) by adding 10 mL of original cultures in 90 mL of autoclaved and filter sterilised (0.22 µm) artificial seawater (ASW) and F/2 media. Cultures were grown for one month (to achieve a minimum cell density of 106 cells/mL) at 26˚C with an irradiance of 85 ± 15 µmol photons m-2 s-1 (Philips TLD 18W/54 fluorescent tubes, 10 000 K on a 12h:12h light:dark cycle). Before use, cells were centrifuged at 700 × g for 10 mins at 26˚C and rinsed twice with ASW to remove residual media solution. Cells were resuspended in 100 mL ASW + F/2 media in sterile culture flasks. Untreated cultures Untreated Symbiodiniaceae cultures (n = 5) were maintained as above alongside the Ab+Tx treatments (below). Antibiotic treatment (AbTx) Each antibiotic treatment subculture (n = 5 per Symbiodiniaceae species) was provided with TritonX-100 detergent added to a final concentration of 20 µg/mL and placed on a shaker at mid speed for 30 s. All cultures (Ab+Tx treatment and untreated) were immediately centrifuged at 700 × g for 10 mins at 26˚C, and the supernatant discarded. Cells were rinsed in 20 mL ASW and centrifuged at 700 x g for 10 mins at 26˚C. Cells were transferred to sterile culture flasks and resuspended in 9 ml ASW+F/2. An aliquot of 1 mL custom antibiotic mix (Penicillin at 31.25 µg mL-1, Streptomycin and Kanamycin at 50 µg mL-1, and Neomycin, Ciprofloxacin and Ampicillin all at 100 µg mL-1; Ab+Tx) was added to each antibiotic treatment flask (and 1 mL ASW added to each untreated flask), and flasks were replaced in the incubator. After 48 hours, 90 mL ASW + F/2 was added to all cultures. Cultures were allowed to recover for 5 days prior to metabolism quenching.

Sample Preparation:

Sampleprep ID:	SP003052
Sampleprep Summary:	Seven days after the antibiotic treatment process, 3 × 1 mL aliquots were collected from each culture flask for flow cytometry cell counting of both bacteria and Symbiodiniaceae cells, bacteria community analysis via 16S rRNA amplicon sequencing analysis, and Symbiodiniaceae photophysiological measurements (methods described in more detail below). The remaining Symbiodiniaceae cells were concentrated by centrifugation at 700 × g for 10 mins at 26˚C, the media discarded, and Symbiodiniaceae pellets snap frozen in liquid nitrogen for metabolite profiling.

Sampleprep ID:

SP003052

Sampleprep Summary:

Seven days after the antibiotic treatment process, 3 × 1 mL aliquots were collected from each culture flask for flow cytometry cell counting of both bacteria and Symbiodiniaceae cells, bacteria community analysis via 16S rRNA amplicon sequencing analysis, and Symbiodiniaceae photophysiological measurements (methods described in more detail below). The remaining Symbiodiniaceae cells were concentrated by centrifugation at 700 × g for 10 mins at 26˚C, the media discarded, and Symbiodiniaceae pellets snap frozen in liquid nitrogen for metabolite profiling.

Combined analysis:

Analysis ID	AN004811
Analysis type	MS
Chromatography type	GC
Chromatography system	Shimadzu GC-2010
Column	Agilent DB5-MS (15m x 0.25mm, 0.25um)
MS Type	EI
MS instrument type	GC-TOF
MS instrument name	Shimazdu 2030
Ion Mode	POSITIVE
Units	relative abundance

Chromatography:

Chromatography ID:	CH003636
Instrument Name:	Shimadzu GC-2010
Column Name:	Agilent DB5-MS (15m x 0.25mm, 0.25um)
Column Temperature:	100-320
Flow Gradient:	10°C min-1
Flow Rate:	1 mL/min
Solvent A:	helium
Solvent B:	helium
Chromatography Type:	GC

MS:

MS ID:	MS004557
Analysis ID:	AN004811
Instrument Name:	Shimazdu 2030
Instrument Type:	GC-TOF
MS Type:	EI
MS Comments:	Approximately 520 quantifying MRM targets were collected using Shimadzu Smart Database along with qualifier for each target which covers about 350 endogenous metabolites and multiple 13C labelled internal standards. Both chromatograms and MRMs were evaluated using the Shimadzu GCMS browser and LabSolutions Insight software.
Ion Mode:	POSITIVE