Data from: Temperature and carbon dioxide interactively drive shifts in the compositional and functional structure of peatland protist communities

Kilner, Christopher 1 ; Carrell, Alyssa2 ; Wieczynski, Daniel 1 ; Votzke, Samantha1 ; DeWitt, Katrina1 ; Yammine, Andrea1 ; Shaw, Jonathan 1 ; Pelletier, Dale2 ; Weston, David2 ; Gibert, Jean Philippe1

Published Aug 08, 2025 on Dryad. https://doi.org/10.5061/dryad.kprr4xhbx

Data files

Aug 08, 2025 version files 77.93 MB

Abstract

Microbes affect the global carbon cycle that influences climate change and are in turn influenced by environmental change. Here, we use data from a long-term whole-ecosystem warming experiment at a boreal peatland to answer how temperature and CO₂ jointly influence communities of abundant, diverse, yet poorly understood, non-fungi microbial Eukaryotes (protists). These microbes influence ecosystem function directly through photosynthesis and respiration, and indirectly, through predation on decomposers (bacteria, fungi). Using a combination of high-throughput fluid imaging and 18S amplicon sequencing, we report large climate-induced, community-wide shifts in the community functional composition of these microbes (size, shape, metabolism) that could alter overall function in peatlands. Importantly, we demonstrate a taxonomic convergence but a functional divergence in response to warming and elevated CO₂ with most environmental responses being contingent on organismal size: warming effects on functional composition are reversed by elevated CO₂ and amplified in larger microbes but not smaller ones. These findings show how the interactive effects of warming and rising CO₂ could alter the structure and function of peatland microbial food webs — a fragile ecosystem that stores 25% of terrestrial carbon and is increasingly threatened by human exploitation.

General Information

Date of data collection: June and September 2019

Geographic location of data collection: S1-Bog of the Marcell Experimental Forest (47° 30.4760′ N, 93° 27.1620′ W; 418 m above mean sea level), Minnesota, USA

Study Description

This dataset contains protist trait measurements, abundance data, and taxonomic composition data from the SPRUCE (Spruce and Peatland Responses Under Changing Environments) long-term whole-ecosystem warming experiment. Samples were collected from experimental enclosures with factorial temperature (+0, +2.25, +4.5, +6.75, +9°C above ambient) and CO2 treatments (ambient 450 ppm and elevated 900 ppm). Protist communities were analyzed using fluid imaging (FlowCAM) and 18S rRNA gene amplicon sequencing.

File Descriptions

1. SPRUCE_2019.csv

Description: Raw protist trait measurements from FlowCAM fluid imaging analysis. Each row represents an individual protist cell with associated morphological and optical measurements.

Dimensions: 211,039 rows × 48 columns

2. SPRUCE_Density_2019.csv

Description: Calibration data for converting FlowCAM particle counts to density estimates (particles per mL).

Dimensions: 20 rows × 6 columns

3. mapping.txt

Description: Sample metadata linking sample identifiers to experimental treatments and collection information for 18S rRNA gene amplicon sequencing analysis.

Dimensions: 50 rows × 6 columns

4. protist-dada2table.qza

Description: QIIME2 artifact containing the feature table (amplicon sequence variant abundance matrix) generated by DADA2 processing of 18S rRNA gene sequences. This file contains the abundance counts of each unique sequence variant (feature) across all samples after quality filtering, denoising, dereplication, chimera removal, and paired-end merging.

File format: QIIME2 artifact (.qza)

Content: Feature table with sequence variants as rows and samples as columns, containing integer abundance counts.

5. protist-taxonomy.qza

Description: QIIME2 artifact containing taxonomic classifications for each amplicon sequence variant (feature) identified in the protist-dada2table.qza file. Taxonomic assignments were made using the PR2 (Protist Ribosomal Reference) database for 18S rRNA gene sequences.

File format: QIIME2 artifact (.qza)

Content: Taxonomic classifications from Kingdom to Species level (where available) with confidence scores for each sequence variant.

Variable Definitions

SPRUCE_2019.csv Variables

Variable Name	Description	Units	Notes
Plot ID	Experimental plot identifier	categorical	Plot numbers corresponding to specific temperature/CO2 treatment combinations
Sample ID	Sample identifier within plot	categorical	North or South for location in Plot
Temperature (±C°)	Temperature treatment above ambient	±C°	Values: 0, 2.25, 4.5, 6.75, 9
Class	Type of taxa	categorical	Protists
Area (ABD)	Area based on equivalent circular diameter	μm²	Morphological measurement
Area (Filled)	Total filled area of particle	μm²	Morphological measurement
Aspect Ratio	Length to width ratio	dimensionless	Shape measurement; values >1 indicate elongated cells
Average Blue	Mean blue channel intensity	grayscale units (0-255)	Optical measurement
Average Green	Mean green channel intensity	grayscale units (0-255)	Optical measurement
Average Red	Mean red channel intensity	grayscale units (0-255)	Optical measurement
Circularity	Measure of how circular the particle is	dimensionless (0-1)	1 = perfect circle, <1 = more irregular
Circularity (Hu)	Hu's circularity measure	dimensionless	Alternative circularity calculation
Compactness	Measure of particle compactness	dimensionless	Higher values = more compact
Convex Perimeter	Perimeter of convex hull	μm	Morphological measurement
Convexity	Ratio of particle perimeter to convex perimeter	dimensionless (0-1)	1 = perfectly convex
Date	Date of sample analysis	date	Date when sample was processed through FlowCAM
Diameter (ABD)	Diameter based on area	μm	Calculated as 2×sqrt(Area/π)
Diameter (ESD)	Equivalent spherical diameter	μm	Diameter of sphere with same volume
Diameter (FD)	Feret diameter	μm	Maximum distance between two points on particle boundary
Elongation	Measure of particle elongation	dimensionless	Higher values = more elongated
Feret Angle Max	Maximum Feret angle	degrees	Orientation measurement
Feret Angle Min	Minimum Feret angle	degrees	Orientation measurement
Fiber Curl	Measure of fiber curvature	dimensionless	Higher values = more curved
Fiber Straightness	Measure of fiber straightness	dimensionless (0-1)	1 = perfectly straight
Filter Score	FlowCAM filter quality score	dimensionless (0-1)	Automated quality assessment score for particle detection
Geodesic Aspect Ratio	Length to width ratio using geodesic measurements	dimensionless	Morphological measurement
Geodesic Length	Maximum geodesic length	μm	Longest axis measurement
Geodesic Thickness	Geodesic thickness measurement	μm	Width measurement
Intensity	Mean grayscale intensity	grayscale units (0-255)	Overall optical density
Length	Maximum length	μm	Morphological measurement
Original Reference ID	Original particle reference identifier	categorical	Unique identifier assigned by FlowCAM software
Perimeter	Particle perimeter	μm	Morphological measurement
Ratio Blue/Green	Blue to green intensity ratio	dimensionless	Color ratio measurement
Ratio Red/Blue	Red to blue intensity ratio	dimensionless	Color ratio measurement
Ratio Red/Green	Red to green intensity ratio	dimensionless	Indicates resource acquisition mode; higher values suggest heterotrophy
Roughness	Measure of surface roughness	dimensionless	Higher values = rougher surface
Sigma Intensity	Standard deviation of grayscale intensity	grayscale units	Measure of cellular contents/heterogeneity
Sphere Complement	Sphericity complement measure	dimensionless	Measure of deviation from perfect spherical shape
Sphere Count	Count of sphere-like particles	count	Number of particles classified as sphere-like by FlowCAM
Sphere Unknown	Unknown sphere classification	categorical	Particles with uncertain spherical classification
Sphere Volume	Volume assuming spherical shape	μm³	Calculated volume assuming particle is perfectly spherical
Sum Intensity	Total grayscale intensity	grayscale units	Sum of all pixel intensities
Symmetry	Measure of particle symmetry	dimensionless (0-1)	1 = perfectly symmetric
Time	Time of particle detection	time	Time when individual particle was detected during analysis
Timestamp	Full timestamp of detection	datetime	Complete date and time stamp for particle detection
Transparency	Measure of particle transparency	dimensionless (0-1)	Higher values = more transparent
Volume (ABD)	Volume based on area	μm³	Calculated volume measurement
Volume (ESD)	Equivalent spherical volume	μm³	Volume of sphere with same diameter
Width	Maximum width	μm	Morphological measurement

SPRUCE_Density_2019.csv Variables

Variable Name	Description	Units	Notes
Plot ID	Experimental plot identifier	categorical	Plot numbers corresponding to specific temperature/CO2 treatment combinations
Sample ID	Sample identifier within plot	categorical	North or South for location in Plot
Temperature (±C°)	Temperature treatment above ambient	±C°	Values: 0, 2.25, 4.5, 6.75, 9
Class	Type of taxa	categorical	Protists
Count	Raw particle count	count	Number of particles detected by FlowCAM
Particles / ml	Calculated particle density	particles/mL	Particles per milliliter before dilution correction

mapping.txt Variables

Variable Name	Description	Units	Notes
SampleID	Sample identifier for sequencing	categorical	Unique identifier for each sequenced sample
plot	Experimental plot number	categorical	Links to Plot variable in SPRUCE_2019.csv
temp	Temperature treatment	°C	Temperature increase above ambient (0, 2.25, 4.5, 6.75, 9)
co2	CO2 treatment	categorical	"aCO2" = ambient CO2 (450 ppm), "eCO2" = elevated CO2 (900 ppm)
sampling	Sampling period	categorical	"sep" = September sampling period

Experimental Design

The data were collected from the SPRUCE experiment, which uses a factorial design of:

Temperature treatments: 5 levels (+0, +2.25, +4.5, +6.75, +9°C above ambient)
CO2 treatments: 2 levels (ambient 450 ppm, elevated 900 ppm)
Replication: Multiple subplots within each treatment combination

Data Collection Methods

Protist sampling: Superficial peat samples (3-5 cm depth) containing living Sphagnum moss tissue were collected and processed through washing with Carolina protist media.
Fluid imaging: Protist communities were analyzed using a FlowCAM (Yokagawa Fluid Imaging) at 10X magnification. Individual protist cells were imaged and measured for morphological and optical traits.
Density calibration: Particle counts were calibrated to density estimates using known dilution series.
18S rRNA gene sequencing: DNA was extracted from samples and the V4 region of 18S rRNA gene was amplified and sequenced on Illumina MiSeq for taxonomic identification.

Data Processing Notes

All protist images were manually curated to remove non-protist particles, cysts, and debris
Trait measurements represent individual cell-level data
Size classes were determined using Gaussian finite mixture modeling based on geodesic length
Raw trait data were transformed as appropriate for statistical analysis (see manuscript methods)

Missing Data Codes

NA: Data not available or not applicable
Empty cells: No data recorded

Quality Assurance

All samples were processed and imaged in a treatment-blind fashion
Images were manually curated by trained personnel
Calibration curves were used to convert raw counts to density estimates
Taxonomic assignments were made using the PR2 database

Related Publications

Kilner, C.L., Carrell, A.A., Wieczynski, D.J., Votzke, S., DeWitt, K., Yammine, A., Shaw, J., Pelletier, D.A., Weston, D.J., & Gibert, J.P. (2024). Temperature and CO2 interactively drive shifts in the compositional and functional structure of peatland protist communities. Global Change Biology, 30, e17203.

Contact Information

For questions about this dataset, please contact:

Christopher L. Kilner (science@christopher.eco)
Jean P. Gibert (jean.gibert@duke.edu)