Dataset: microzoo_epi_abund
Deployment: MV1008

Cell abundance estimates of heterotrophic protists by size-class, based on epifluorescence microscopy.

Principal Investigator:

Michael R. Landry (University of California-San Diego Scripps, UCSD-SIO)

Contact:

Andrew G. Taylor (University of California-San Diego Scripps, UCSD-SIO)

BCO-DMO Data Manager:

Shannon Rauch (Woods Hole Oceanographic Institution, WHOI BCO-DMO)

Project:

Costa Rica Dome FLUx and Zinc Experiments (CRD FLUZiE)

Current State:

Final no updates expected

Version:

24 June 2014

Deployment Synonyms:

CRD FLUZiE

Version Date:

2014-06-24

Data URL:

https://www.bco-dmo.org/dataset-deployment/517422/data

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Description

Cell abundance estimates of heterotrophic protists by size-class, based on epifluorescence microscopy. Samples were collected on the MV1008 cruise in the Costa Rica Dome (CRD) region of the Eastern Tropical Pacific Ocean.

Methods & Sampling

Dataset acquisition description

Seawater samples (500 mL) for microscopical analysis were gently collected from the CTD and immediately preserved for slide preparation according to a modified protocol of Sherr and Sherr (1993). The samples were first preserved with 260 uL of alkaline Lugol’s solution, immediately followed by 10 mL of buffered formalin and 500 uL of sodium thiosulfate, with gentle mixing between each addition. Preserved samples were shielded from light and left to rest at room temperature for 1 h. After the rest period, 1 mL of proflavin (0.33% w/v) was added and the samples were stored in the dark for an additional hour. Just prior to filtration, the preserved samples were stained with 1 mL of DAPI (0.01 mg mL-1) and immediately transferred to the filtration manifold. A 50-mL aliquot (small volume, SV) of the sample was filtered through a 25-mm black polycarbonate filter with 0.8-um pore size, and the remaining 450 mL aliquot (large volume, LV) was filtered through a 25-mm black polycarbonate filter with 8.0-um pores. A 10-mm nylon backing filter was placed under all polycarbonate filters to promote even cell distribution, and filtered the samples under gentle vacuum pressure (<100 mm Hg). Each filter was then mounted onto glass slides with one drop of Type DF immersion oil and a No. 2 cover slip, and the prepared slides were frozen at -80°C for later analysis in the lab. NOTE: ciliated protists are poorly preserved by this method and are not included in these analyses.

Data Processing Description

Dataset Processing Description

Slides were digitally imaged using a Zeiss Axiovert 200 M inverted compound microscope equipped for high-throughput epifluorescence microscopy with a motorized focus drive, stage, objective and filters. Digital images were acquired with a Zeiss AxioCam MRc black and white 8-bit CCD camera. SV samples (50 mL aliquots) were viewed at 630X magnification, and LV samples (450 mL aliquots) were viewed at 200X magnification. A minimum of 20 random positions were imaged for each slide, with each position consisting of three to four fluorescent channels: Chl a, DAPI, FITC (SV and LV samples) and phycoerythrin (SV samples only).

Images were analyzed using ImagePro software to semi-automate the enumeration of eukaryotic cells larger than 1.5 um in length (Taylor et al., 2012). Whenever possible, 20 positions and >300 cells were counted for each slide. Each cell was manually identified and grouped into identifiable taxonomic groups. Heterotrophic cells were distinguished from autotrophs (eukaryotic phytoplankton) by very low red autofluorescence (low/absent chlorophyll and absence of defined chloroplasts). Cells with recently consumed autotrophic prey were counted as heterotrophs.

BCO-DMO Processing Notes:
- size_class and taxon columns were transposed from columns into rows.

More information about this dataset deployment

Funding

Award Number	Funding Source
OCE-0826626	NSF Division of Ocean Sciences

Instruments

Camera

Supplied Name: Zeiss AxioCam MRc

Supplied Description:

Digital images were acquired with a Zeiss AxioCam MRc black and white 8-bit CCD camera.

Instrument Type

Generic Name: Camera

Acronym: camera

Community Identifier: http://vocab.nerc.ac.uk/collection/L05/current/311/

Generic Description:

All types of photographic equipment including stills, video, film and digital systems.

Inverted Microscope

Supplied Name: Zeiss Axiovert 200 M inverted compound microscope

Supplied Description:

Instrument Type

Generic Name: Inverted Microscope

Community Identifier: http://vocab.nerc.ac.uk/collection/L05/current/LAB05/

Generic Description:

An inverted microscope is a microscope with its light source and condenser on the top, above the stage pointing down, while the objectives and turret are below the stage pointing up. It was invented in 1850 by J. Lawrence Smith, a faculty member of Tulane University (then named the Medical College of Louisiana).

Inverted microscopes are useful for observing living cells or organisms at the bottom of a large container (e.g. a tissue culture flask) under more natural conditions than on a glass slide, as is the case with a conventional microscope. Inverted microscopes are also used in micromanipulation applications where space above the specimen is required for manipulator mechanisms and the microtools they hold, and in metallurgical applications where polished samples can be placed on top of the stage and viewed from underneath using reflecting objectives.

The stage on an inverted microscope is usually fixed, and focus is adjusted by moving the objective lens along a vertical axis to bring it closer to or further from the specimen. The focus mechanism typically has a dual concentric knob for coarse and fine adjustment. Depending on the size of the microscope, four to six objective lenses of different magnifications may be fitted to a rotating turret known as a nosepiece. These microscopes may also be fitted with accessories for fitting still and video cameras, fluorescence illumination, confocal scanning and many other applications.

Parameters

Supplied Name	Supplied description	Supplied Units	Standard Name
event	Number referring to the particular activity (event) on the FluZiE cruise.	integer	event
cast	CTD Cast number from the FluZiE cruise.	integer	cast
cycle	Type and number of cruise sampling event. Either "Stn_n" or "Cycle_n". A transect of stations was sampled from 29 June to 03 July. Five quasi-Lagrangian experiments called "cycles" were conducted during the remainder of the cruise.	text	no_bcodmo_term
date_local	Date of CTD cast (local time zone of UTC -6). in the format mmddyyyy	unitless	date
lat	Latitude in degrees North.	decimal degrees	lat
lon	Longitude in degrees East.	decimal degrees	lon
depth	Sample depth.	meters	depth
niskin	Niskin bottle that the sample was taken from.	integer	bot_Nis
taxon	Taxon. heterotrophic_dinos = Heterotrophic Dinoflagellates; heterotrophic_flags = Heterotrophic Flagellates; all = all taxonomic groups (heterotrophic dinoflagellates + heterotrophic flagellates)	text	taxon
size_class	Size-Class Distribution (cell lengths are longest major axis): lt2 = cells < 2 um; 2_to_10 = cells 2-10 um; 10_to_20 = cells 10-20 um; gt20 = cells > 20 um; total = all sizes.	micrometers (um)	no_bcodmo_term
abundance	Abundance of the taxon size class.	cells per milliliter (cells/mL)	abundance

Database

Contribute Data

Dataset: microzoo_epi_abund
Deployment: MV1008

Dataset acquisition description

Dataset Processing Description

Database

Contribute Data

Dataset: microzoo_epi_abundDeployment: MV1008

Dataset acquisition description

Dataset Processing Description

Dataset: microzoo_epi_abund
Deployment: MV1008