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This repository contains the data and code supporting the results presented in:
"Abrupt alkalinization alters microbial diversity and promotes the proliferation of marine parasites in coastal microcosm experiments."

Notes:

• The structure of this repository is designed to function as an R project. R markdown files and scripts are annotated internally.
• Figure images have been omitted from this repository as they appear in the associated manuscript, which is published under a CC BY license.
• "Mesocosm" was changed to "microcosm" for the published manuscript, but "mesocosm" is retained in all R markdown files and scripts to avoid code breaks.

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Overview of Repository Contents (oae_mesocosm_manuscript.zip)

oae_mesocosm_manuscript/
├── data/
│   ├── all_18S_seq_data/        # 18S rDNA amplicon sequencing data and processed outputs
│   ├── all_media_prep/          # Preparation of alkalinity stock solutions and spiking of filtered seawater for both experiments
│   ├── apr23_mesocosm/          # Spring 2023 mesocosm experiment data
│   ├── metadata/                # Sample metadata for both experiments
│   ├── nov23_mesocosm/          # Fall 2023 mesocosm experiment data
│   ├── sst_map/                 # Sea surface temperature (SST) and coastal map data
│   └── upwelling_indices/       # Biologically Effective Upwelling Transport Index (BEUTI) time series data files
├── src/
│   ├── dna_seq/                 # R Markdown files and scripts for 18S amplicon analysis and outputs
│   ├── dsi_experiment/          # R Markdown files for post hoc dissolved silicic acid (DSi) experiment
│   ├── mesocosm_combined/       # R Markdown files for producing physicochemical and FCM outputs
│   └── R/                       # R helper scripts sourced by R Markdown files

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Data Files

data/all_18S_seq_data/

This folder contains raw FASTQ files and processed outputs from 18S rDNA amplicon sequencing.

FASTQ files (fastq_files/spring_m2/ and fastq_files/fall_m3/)
Raw paired-end Illumina sequencing reads. Files follow the naming convention:

{EXPERIMENT}_{TREATMENT}[_BLK]_{TIMEPOINT}_{REPLICATE}_{SAMPLE_NUMBER}_L001_R{READ}_001.fastq

Example: M2_NAOH_T3_B_S7_L001_R1_001.fastq

OAE_all18s_M2M3_Sep2024_sample_data.xlsx / .csv
Sample sheet linking FASTQ file names to metadata fields from oae_mesocosm_2.xlsx/csv and oae_mesocosm_3.xlsx/csv.

seqtab.rds
R data file (RDS format). Amplicon sequence variant (ASV) count table output from DADA2.

seqtab_taxfiltered_20241015.rds
R data file. ASV count table after removal of non-target taxa.

eTax_w_troph_20241015.rds / .csv
Taxonomy table with trophic mode assigned to each ASV.

no_troph_20241015.csv
Subset of ASVs that could not be assigned a trophic mode.

meso_combined.csv
Table combining sample data for the spring and fall mesocosm experiments (e.g., carbonate chemistry, nutrients) to be used with ASVs.

mock_community_info_wManualBlast_20230124.csv
Mock community ASVs and relative abundances.

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data/all_media_prep/

oae_mesocosm_media.xlsx
Preparation of 1 M alkalinity stock solutions for both experiments. Seawater collection coordinates are also noted here. Metadata for this file is noted on the third excel tab.

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data/metadata/

oae_mesocosm_metadata.xlsx
Sample metadata table for both mesocosm experiment files (oae_mesocosm_2.csv/xlsx and oae_mesocosm_3.csv/xlsx). Columns include: Variable, Unit, and Description.

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data/apr23_mesocosm/ — Spring 2023 Experiment (M2)

oae_mesocosm_2.xlsx / .csv
Main data table for the spring mesocosm experiment. Each row is one sample. See /data/metadata/oae_mesocosm_metadata.xlsx for variable information. Units are also included in column labels (e.g., din_umol_l = dissolved inorganic nitrogen [µmol/L]).

dsi_experiment/oae_dsi_raw.xlsx / .csv
Raw data from the post hoc DSi addition experiment conducted after the spring mesocosm experiment (only conducted in the spring). Variables include sampling_date (date of collection), date label (tube label), bottle_type (type of bottle that solution was stored in; i.e., borosilicate, PP, or PC), treatment (alkalinity treatment), solution_starting_mass_g (starting solution mass), solute_concentration_mol_l (molar concentration of solution), hcl_umol_l (molar concentration of acid solution used to neutralize alkalinity stock solution prior to analysis), dilution_factor (dilution factor resulting from HCl addition), dsi_umol_l (DSi concentration), dsi_umol_l_corr (DSi concentrations adjusted for instrument method detection limits), dsi_umol_l_final (final DSi accounting for dilution factor), sample_collector (collecting researcher).

fcm_bigelow/apr23_fc_phyto.xlsx / .csv
Flow cytometry counts for phytoplankton populations (spring experiment). Variables include sampling_date (sample collection date), FCM file (file generated by FCM), tube number (cryovial tube number), sample_label (sample label), type (treatment), Flow rate (µl/min), Acq time (acquisition time; min), dilution (dilution factor), and cell abundance columns for distinct populations (e.g., Crypto counts, Nanoeuk1 counts ; units = cells mL⁻¹).

fcm_bigelow/apr23_fc_virbact.xlsx / .csv
Flow cytometry counts for virus and bacteria populations (spring experiment). Variables include sampling_date, FCM file, tube number, sample_label, type, Flow rate (µl/min), Acq time (min), dilution, cell abundance counts for bacteria (cells mL⁻¹), and virus-like particle counts (particles mL⁻¹).

fcm_bigelow/apr23_spring_bfc_sample_list_updated_FCM results_01072025.xlsx
Raw sample list and results file as received from Bigelow Laboratory flow cytometry facility.

icp_oes/oae_m2_icp_oes.xlsx / .csv
Concentrations of Na, Ca, Mg, Si, and P (all in ppm) as measured by ICP-OES. Metadata included on third excel tab (this metadata applies to icp_oes/oae_m3_icp_oes.xlsx as well).

irradiance/m2_irradiance.xlsx / .csv
Photosynthetically active radiation (PAR) measurements within incubation tank. Columns: Record, Date, Time_pt, INPUT1, INPUT2 (scalar LI-COR sensor 1 and 2; µmol photons m⁻² s⁻¹).

temp/oae_m2_temp.xlsx / .csv
Continuous temperature logger data for spring mesocosm incubation tank. Variables: Date Time, PT (date and time of measurement; Pacific Time), Temp, C (temperature; Celsius).

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data/nov23_mesocosm/ — Fall 2023 Experiment (Mesocosm M3)

Structure and variable names mirror the spring experiment (see above). Files are:

• oae_mesocosm_3.xlsx / .csv — Main data table (same variables as oae_mesocosm_2).
• fcm_bigelow/nov23_fc_phyto.xlsx / .csv — Flow cytometry phytoplankton counts.
• fcm_bigelow/nov23_fc_virbact.xlsx / .csv — Flow cytometry bacteria/virus-like particle counts.
• fcm_bigelow/nov23_fall_Bigelow_FC_RNA_sample_list_FCM analyses_01072025.xlsx — Raw Bigelow facility file.
• icp_oes/oae_m3_icp_oes.xlsx / .csv — ICP-OES data.
• irradiance/m3_irradiance.xlsm / .csv — PAR data.
• temp/oae_m3_temp.xlsx / .csv — Continuous temperature data.

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data/sst_map/

Contains SST data and a folder (gshhg_data) for the coastline file used to produce SST maps in oae_mesocosm_sbc_sst.Rmd. Users will need to download GSHHG coastlines, rivers, and borders in native binary format (gshhs_f.b file) from https://www.soest.hawaii.edu/pwessel/gshhg/index.html (last accessed 20260512) to reproduce the published maps.

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data/upwelling_indices/

Folder for monthly BEUTI csv files called within oae_mesocosm_pt1.Rmd. "BEUTI provides estimates of vertical nitrate flux near the coast (i.e., the amount of nitrate upwelled/downwelled), which may be more relevant than upwelling strength when considering some biological responses" (see Jacox MG, CA Edwards, EL Hazen, SJ Bograd (2018) Coastal upwelling revisited: Ekman, Bakun, and improved upwelling indices for the U.S. west coast, J. Geophysical Research, 123(10), 7332-7350).

The monthly BEUTI can be downloaded at https://mjacox.com/upwelling-indices/ (last accessed 20260512).

Two files are called within the R markdown file (not included in repository):

BEUTI_monthly.csv
Monthly BEUTI for the California Current System (2023).

BEUTI_monthly_1988-2022.csv
Extended historical BEUTI time series (1988–2022).

Note: Users will need to separate the downloaded monthly BEUTI data into individual files (i.e., 2023 and 1988-2022) or adjust the code to wrangle the data from a single file.

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Code (R Markdown Files and Scripts)

R Markdown files must be run in order within each subfolder (pt1 → pt2 → pt3...) because intermediate outputs are saved and read by subsequent files. The exception is oae_mesocosm_sbc_sst.Rmd, which is standalone.

src/dna_seq/ — 18S Amplicon Sequencing Analysis

Each R Markdown file is annotated internally. Run in this order:

figures/ — Output folder for figures created in R Markdown files (figures not included; see manuscript).

tables/ — Output folder for tables created in R Markdown files (tables not included; see manuscript).

taxonomy/dada2/ — DADA2 pipeline scripts:

taxonomy/eri_server_scripts/ — Scripts run on an external computing cluster for taxonomic assignment:

taxonomy/other_scripts/ — Scripts used for creating a taxonomic ensemble:

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src/mesocosm_combined/ — Physicochemical and Flow Cytometry Analysis

Each R Markdown file is annotated internally. Run in this order:

figures/ — Output folder for figures created in R Markdown files (figures not included; see manuscript).

tables/ — Output folder for tables created in R Markdown files (tables not included; see manuscript).

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src/dsi_experiment/ — Post Hoc DSi Experiment

figures/ — Output folder for figures created in R Markdown file (figures not included; see manuscript).

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src/R/ — Helper Scripts

These R scripts are sourced within the R Markdown files and do not need to be run independently (each is annotated internally):

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Acronyms

A reference list of acronyms used in this repository.

Approaches & Materials

Biological & Ecological

Chemical & Biogeochemical

Statistical

Physical & Oceanographic

Databases & Software

Instrumentation & Organizations