Flow cytometry measurements from HHQ experiments conducted during the MesoHux mesocosm experiment, May 2017, Bergen, Norway

Website: https://www.bco-dmo.org/dataset/753431
Data Type: experimental
Version: 1
Version Date: 2019-01-23

Project
» Collaborative Research: Building a framework for the role of bacterial-derived chemical signals in mediating phytoplankton population dynamics (HHQSignals)
ContributorsAffiliationRole
Harvey, ElizabethSkidaway Institute of Oceanography (SkIO)Principal Investigator
Rowley, DavidUniversity of Rhode Island (URI)Co-Principal Investigator
Whalen, Kristen E.Haverford College (Haveford)Co-Principal Investigator
Copley, NancyWoods Hole Oceanographic Institution (WHOI BCO-DMO)BCO-DMO Data Manager

Abstract
This dataset includes flow cytometry measurements from HHQ experiments conducted during the MesoHux mesocosm experiment, May 2017, Bergen, Norway. Microbial mesocosms were spiked with 2-heptyl-4-quinolone (HHQ).


Coverage

Spatial Extent: Lat:60.221 Lon:5.281
Temporal Extent: 2017-05-16 - 2017-05-31

Dataset Description

This dataset includes flow cytometry measurements from HHQ experiments conducted during the MesoHux mesocosm experiment, May 2017, Bergen, Norway. Microbial mesocosms were spiked with 2-heptyl-4-quinolone (HHQ).


Acquisition Description

Triplicate 5 mL samples were preserved for flow cytometry with 0.5% glutaraldehyde (final concentration), incubated at 4°C for 10 min and frozen (-80°C) until analysis (within 2-3 weeks; Kemp et al. 1993). To calculate phytoplankton group abundances, 200 µl aliquots of fixed sample were added to a 96-well plate and run on a Guava flow cytometer (Millipore). Filtered seawater (0.45 µm) was run as a blank and instrument-specific beads were used to calibrate the cytometer. Samples were analyzed at low flow rate (0.24 µl s-1) for 3 min. Three major phytoplankton groups were distinguishable based on plots of forward scatter vs. orange (phycoerythrin-containing, Synechococcus spp.) or red (pico- and nanoeukaryotes) fluorescence signals (Worden and Binder 2003). 

Samples for enumerating bacteria were stained prior to running on the Guava in 0.5% v/v SybrGreen I DNA stain for 1 hour at room temperature in the dark.

Mesocosm treatment for all HHQ experiments was as follows:
Redfield: N:P added in a 16:1 ratio during the first 3 days of the experiment, no shading

HHQ treatments here are as follows:
High HHQ - 100 ng mL-1 (410 uM) added to triplicate 5L bottles.
DMSO control - equivalent (v:v) DMSO added to triplicate 5L bottles.

 All bottles were incubated for 24h in a flow-through tank, that was shaded to mimic in situ conditions. Chlorophyll samples were taken at T0 and T24 for all experiments.

Data were processed in Excel with statistics run in Excel, R, or Matlab.


Processing Description

BCO-DMO Processing Notes:
- added conventional header with dataset name, PI name, version date
- modified parameter names to conform with BCO-DMO naming conventions


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Related Publications

Kemp, P., B. F. Sherr, E. B. Sherr, and J. J. Cole (Eds.). (1994). Handbook of methods in aquatic microbial ecology. Lewis Publishing, Boca Raton, FL 33431, 777 pp.
Worden, A., & Binder, B. (2003). Application of dilution experiments for measuring growth and mortality rates among Prochlorococcus and Synechococcus populations in oligotrophic environments. Aquatic Microbial Ecology, 30, 159–174. doi:10.3354/ame030159

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Parameters

ParameterDescriptionUnits
Datesampling date formatted as Mon dd yyyy unitless
Samplesample identifier unitless
Experiment_numexperiment number unitless
Timetime since start of experiment hours
Replicationreplicate number unitless
Bacterianumber of bacterial cells cells/milliliter
Synechococcusnumber of Synechococcus cells cells/milliliter
Picoeukaryotesnumber of Picoeukaryotes cells cells/milliliter
Nanoeukaryotesnumber of Nanoeukaryotes cells cells/milliliter
Total_Phytoplankton_lt_15umtotal number of phytoplankton cells less than 15 microns in diameter cells/milliliter


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Instruments

Dataset-specific Instrument Name
5 L Niskin
Generic Instrument Name
Niskin bottle
Dataset-specific Description
Used to collect water samples.
Generic Instrument Description
A Niskin bottle (a next generation water sampler based on the Nansen bottle) is a cylindrical, non-metallic water collection device with stoppers at both ends. The bottles can be attached individually on a hydrowire or deployed in 12, 24 or 36 bottle Rosette systems mounted on a frame and combined with a CTD. Niskin bottles are used to collect discrete water samples for a range of measurements including pigments, nutrients, plankton, etc.

Dataset-specific Instrument Name
Millipore Guava inCyte BG HT flow cytometer
Generic Instrument Name
Flow Cytometer
Dataset-specific Description
Used for cell counts
Generic Instrument Description
Flow cytometers (FC or FCM) are automated instruments that quantitate properties of single cells, one cell at a time. They can measure cell size, cell granularity, the amounts of cell components such as total DNA, newly synthesized DNA, gene expression as the amount messenger RNA for a particular gene, amounts of specific surface receptors, amounts of intracellular proteins, or transient signalling events in living cells. (from: http://www.bio.umass.edu/micro/immunology/facs542/facswhat.htm)


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Project Information

Collaborative Research: Building a framework for the role of bacterial-derived chemical signals in mediating phytoplankton population dynamics (HHQSignals)

Coverage: Bergen, Norway


NSF Abstract: Bacteria and phytoplankton play a central role in the modification and flow of materials and nutrients through the marine environment. While it has been established that interactions between these two domains are complex, the mechanisms that underpin these interactions remain largely unknown. There is increasing recognition, however, that dissolved chemical cues govern these microbial interactions. This project focuses on establishing a mechanistic framework for how bacterially derived signaling molecules influence interactions between phytoplankton and bacteria. The quorum-sensing (QS) molecule, 2-heptyl-4-quinolone (HHQ) will be used as a model compound for these investigations. Previously published work suggests that exposure to very low levels of HHQ results in phytoplankton mortality. Gaining a mechanistic understanding of these ecologically important interactions will help to inform mathematical models for the accurate prediction of the cycling of material through the marine microbial loop. This work initiates a new, hybrid workshop-internship undergraduate research program in chemical ecology, with a focus Bacteria and phytoplankton play a central role in the modification and flow of materials and nutrients through the marine environment. While it has been established that interactions between these two domains are complex, the mechanisms that underpin these interactions remain largely unknown. There is increasing recognition, however, that dissolved chemical cues govern these microbial interactions. This project focuses on establishing a mechanistic framework for how bacterially derived signaling molecules influence interactions between phytoplankton and bacteria. The quorum-sensing (QS) molecule, 2-heptyl-4-quinolone (HHQ) will be used as a model compound for these investigations. Previously published work suggests that exposure to very low levels of HHQ results in phytoplankton mortality. Gaining a mechanistic understanding of these ecologically important interactions will help to inform mathematical models for the accurate prediction of the cycling of material through the marine microbial loop. This work initiates a new, hybrid workshop-internship undergraduate research program in chemical ecology, with a focus into bacteria-phytoplankton interactions. Undergraduate students participate in an intense summer learning experience where research and field-based exercises are supplemented with short-lecture based modules. Students return to their home institutions and work closely with the PIs to conduct interdisciplinary research relating to the aims and scope of the summer research. This research also provides training and career development to two graduate students and a postdoctoral scientist. Interactions between phytoplankton and bacteria play a central role in mediating biogeochemical cycling and microbial trophic structure in the ocean. The intricate relationships between these two domains of life are mediated via excreted molecules that facilitate communication and determine competitive outcomes. Despite their predicted importance, identifying these released compounds has remained a challenge. The PIs recently identified a bacterial QS molecule, HHQ, produced by globally distributed marine gamma-proteobacteria, which induces phytoplankton mortality. The PIs therefore hypothesize that bacteria QS signals are critical drivers of phytoplankton population dynamics and, ultimately, biogeochemical fluxes. This project investigates the timing and magnitude of HHQ production, and the physiological and transcriptomic responses of susceptible phytoplankton species to HHQ exposure, and quantifies the influence of HHQ on natural algal and bacterial assemblages. The work connects laboratory and field-based experiments to understand the governance of chemical signaling on marine microbial interactions, and has the potential to yield broadly applicable insights into how microbial interactions influence biogeochemical fluxes in the marine environment.


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Funding

Funding SourceAward
NSF Division of Ocean Sciences (NSF OCE)

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