Dataset: BALO Metabolites 15T and 21T MSDial

Name: Ultrahigh reolution mass spectrometry data on lysis products following predation by micropredators from samples generated in 2022-07 by laboratory microcosm experiments and data collected in 2022-07 and 2023-04
Published: 2024-03-18
Keywords: oceans

View Data: Data not available yet

Cite This Dataset

Temporal Extent: 2022-07-07 - 2023-04-01

Project:

Excellence in Research: Assessing the Control by Multiple Micropredators on Bacterial Communities in Estuarine Environments and Characterization of Prey Lysis Products Resulting from Each Predator (Predators of bacteria)

Principal Investigator:

Henry Neal Williams (Florida A&M University, FAMU)

Co-Principal Investigator:

Huan Chen (Florida State University - National High Magnetic Field Lab, FSU - NHMFL)

Ahkinyala Cobb-Abdullah (Virginia Union University, VUU)

Sven Kranz (Florida State University, FSU)

Michael Stukel (Florida State University, FSU)

BCO-DMO Data Manager:

Amber D. York (Woods Hole Oceanographic Institution, WHOI BCO-DMO)

Version:

Version Date:

2024-03-18

Restricted:

Yes

Release Date:

2024-05-31

Validated:

Current State:

Data not available

Ultrahigh reolution mass spectrometry data on lysis products following predation by micropredators from samples generated in 2022-07 by laboratory microcosm experiments and data collected in 2022-07 and 2023-04

Abstract:

This dataset contains potential metabolites identified by 14.5T and 21T FT-ICR MS analyzed by MSDial. Heterotrophic protists and bacteriophages are considered to be major contributors to bacterial mortality in the microbial loop. Although evidence shows a similar role for the predatory bacterium, Halobacteriovorax (HBx), it has been largely ignored. Since the predation mechanism and utilization of prey cellular compounds differ for each predator, we examine by ultra-high resolution mass spectrometry the molecular-level composition of dissolved organic matter released at lyses of the prey. Equal volumes of prey bacteria suspension in artificial seawater were dispensed into three flasks and tested against a single strain of protist, bacteriophages, or HBx. Following incubation and prey lysis, each predator-prey culture was filtered sequentially to remove cell debris, leaving extracellular metabolites in the filtrate. Filtrates were acidified and extracted by solid phase extraction, followed by reverse-phase liquid chromatography coupled with positive and negative electrospray ionization (ESI) orbitrap and FT-ICR mass spectrometry. Results show that predation by protists, HBx, and phages, resulted in separate, distinguishable products. Prey lysis products unique to each micropredator were identified by comparing metabolite profiles. The structural information of these predator-specific signatures was further validated by tandem MS analysis. The chemical composition assessment of the lysis products by micropredators provides insights into how each predator may contribute in different ways to nutrient cycling in the ocean.

Expand/Collapse All

Description

This dataset includes two forms of the data, one as plain-text .csv (See "Data Files" section, and one in Microsoft Excel (.xlsx) format from MSDial software (See "Data Files" and "Supplemental Files" sections).

Methods & Sampling

The first experiment was done using one strain of Vibrio vulnificus as prey. Equal volumes of the prey bacterial suspensions in artificial seawater were dispensed and tested against a single strain of protist, bacteriophages, and HBx. Predator-free control was also included in the analysis. Predator-prey dual cultures were filtered sequentially through 0.8µm, 0.45µm, 0.2 µm, and 0.01µm membrane filters to obtain extracellular metabolites. Filtrates were acidified and extracted by solid phase extraction (Dittmar et al. 2008) using the C18 cartridges, followed by reverse-phase liquid chromatography coupled with positive and negative electrospray ionization (ESI) FT-ICR MS.

Instrument summary:

Liquid Chromatography
Reverse phase LC separations were performed online with a C18 custom packed (ReprosSil_Pur 120 Å C18-AQ, 5 μm., 360 μm o.d. X 75 μm i.d., 7.5 cm) outfitted on our nano-HPLC system (ACQUITY M-class, Waters, Milford, MA). An analytical gradient was modified from published gradients to fit our LC-MS/MS set-up (Soule et al. 2015). Analytical gradient going from 5% B to 100% B was carried out (A: 0.1% formic acid in water B: 0.1% formic acid in methanol). Samples were run in triplicates with 4 µL injections at a flow rate of 0.3 uL/min for the analytical separation.

Tandem Mass Spectrometry
Electrospray ionization (ESI) voltage was set to positive 3.5 kV with a source temperature of 300 ºC. MS analysis was performed on the NHMFL 14.5 T and 21 T FT-ICR MS mass spectrometer outfitted with a Velos Pro linear ion trap (Thermo Fisher Scientific) at the front end (Hendrickson et al. 2015; Schaub et al. 2008). Precursor spectra were obtained in the ICR cell with a 1E6 automatic gain control (AGC), sum 4 transient acquisition, and resolving power of 600k at 400 m/z. CID fragmentation spectra were collected from the Velos Pro Ion Trap on the top 3 most abundant ions in the precursor scan with sum 3 transient acquisition and collision energy of 40. Precursor and fragment spectra were collected in the 150-1500 m/z dynamic range.

Location Summary:

Samples were generated in 2022-July by laboratory microcosm experiments, and the data were collected in 2022-07 and 2023-04.

In vitro microcosm experiments to isolate and characterize the prey cellular organic matter released as lysis products as a result of micropredators were conducted at PI Williams lab at Florida A&M University.

FT-ICR MS data collection and analysis was conducted by co-PI Chen in the ICR user facility at the National High Magnetic Field Laboratory.

Orbitrap MS data were collected at the Systems Mass Spectrometry Core at Georgia Institute of Technology in collaboration with Dr. Facundo Fernandez's lab.

Data Processing Description

First, MSDial 4.90 (RIKEN), a freely available analysis software, was used for metabolite analysis loaded with a public library of tandem mass spectrometry experiments (13,303 compounds: positive mode) (Tsugawa et al. 2015). MSDial was used due to the ability to identify unique metabolites as well as compare relative abundance of metabolites within each sample. Data was organized by predator types for comparisons. Precursor parameters were set for accurate mass tolerance for MS and MS/MS (0.001 Da and 0.025 Da) and a minimum peak detection of 10k amp. Eight analyte adducts were selected, ([M+NH4]+, [M+Na]+ , [M+Na]+ , [2M+NH4]+ , [M+2H]2+, [M+H+NH4]2+, [M+H+Na]2+, [M+3H]3+) based on previous studies.

The identification score was set to 80% in MSDial for identifications and to limit false positives. A standard mixture was made with B12, ferrichrome, and ferrioxamine E, each at 5 mM. The standard mix runs were used for the alignment of all spectra for comparison. MSDial output includes metabolite retention times, m/z, compound name, and relative abundance within each sample set.

The dataset was also analyzed using Compound Discoverer (ThermoFisher Scientific, version 3.3 SP1).

After analysis, the data were exported from MSDial and saved as .xlsx. The pdf files (*_CompoundDiscovererReport.pdf) were exported from Compound Discoverer. See the "Supplemental Files" section for access to the .xslx and .pdf files.