DOI: 10.5061/dryad.kprr4xhbx
title: Data from: Temperature and carbon dioxide interactively drive shifts in the compositional and functional structure of peatland protist communities

description: 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.
description: # Data from: Temperature and carbon dioxide interactively drive shifts in
 the compositional and functional structure of peatland protist communities
 ## 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 &gt;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, &lt;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 1. **Protist sampling:** Superficial peat samples (3-5 cm depth) containing living Sphagnum moss tissue were collected and processed through washing with Carolina protist media. 2. **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. 3. **Density calibration:** Particle counts were calibrated to density estimates using known dilution series. 4. **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](mailto:science@christopher.eco)) * Jean P. Gibert ([jean.gibert@duke.edu](mailto:jean.gibert@duke.edu))