DOI: 10.5061/dryad.c866t1gd1
title: Data from: Natural thermal stress-hardening of corals through cold temperature pulses in the Thai Andaman Sea

description: Stress-hardening by environmental priming could increase the odds for
 corals to resist ocean warming. Natural environmental fluctuations, such
 as those observed on offshore reefs in the Andaman Sea, provide an ideal
 natural environment to study these effects. Here, internal waves (IW)
 generate short cold-water pulses that peak from January to June and are
 absent from August to November. Additionally, only western shores of
 islands are exposed to this stress-hardening stimulus of IWs, while
 eastern shores remain sheltered. Therefore, this study examined (1)
 whether exposed corals were more heat stress resistant than their
 sheltered conspecifics and (2) whether this trait would persist during the
 season of stimulus absence. We exemplify that thermal regimes featuring
 cold-temperature pulses successfully induced thermal stress-hardening in
 corals. Corals from the IW-sheltered shore responded strongly to heat
 stress irrespective of the season, while stress responses of IW-exposed
 corals were either undetectable (during stimulus presence) or very weak
 (during stimulus absence). However, this demonstrates the relevance of
 stimulus re-occurrence in maintaining heat resistance. Furthermore,
 priming stimuli do not need to exceed certain upper thermal thresholds to
 be effective and we argue that cooling pulses represent a safer
 stress-hardening regimen potentially implemented in conservation
 strategies since it avoids warming-stress accumulation.
description: <strong>Study sites and coral
 collection</strong> Study sites were located at Racha
 Island in the Andaman Sea off the coast of Thailand, both at 15 m water
 depth (Figure 1 A-B). A reef on the western shore was chosen (7.595530°N,
 98.354320°E, Figure 1 B) where internal wave (IW) forcing as a potential
 stress-hardening stimulus induced environmental variability through
 frequent upwelling of deep, cool, and nutrient rich water onto the shelf
 (Schmidt
 et al., 2016; Wall et al., 2012). A reef on the eastern
 shore, sheltered from the stimulus of IWs, was chosen to represent a low
 variability reef (7.598910°N, 98.373100°E, Figure 1 B). Temperature
 fluctuations were monitored <em>in situ</em> as a proxy for IW
 impact and environmental variability. Temperature loggers (HOBO Pendant
 Temperature/Light 8K Data Logger, Onset, USA) were deployed at the study
 sites one month before heat stress assays were performed. At each study
 site, visually healthy coral colonies of <em>Pocillopora
 </em>sp. and <em>Porites </em>sp. were permanently
 tagged to assess their thermal resistance levels during the two seasons (n
 = 8 to n = 18, Figure 1 C, Table S1). These two coral species are
 cosmopolitan reef-builders in Thailand and within the entire Indo-Pacific
 region (Brown &
 Phongsuwan, 2012; Jain et al., 2023; Schmidt et al., 2012).
 Coral fragments were collected at the end of April 2018, during the season
 of highest IW intensity, and at the end of October
 (<em>Porites</em> sp.) and November
 (<em>Pocillopora</em> sp.), during the seasonal absence of the
 IW stimulus. Two fragments (<em>Porites</em> sp.: ø ~ 6 cm;
 <em>Pocillopora </em>sp.: length ~ 5 cm) per colony were
 collected using a chisel and a hammer (Table S1). <strong>Short-term heat stress
 assays</strong> Collected fragments were
 instantly transported to the Phuket Marine Biological Center (Phuket,
 Thailand) where they were maintained in two 500 L flow-through tanks with
 a flow rate of 2.8 ± 1.31 L/min until the start of each heat stress assay.
 Another 500 L source tank constantly supplied both flow-through tanks with
 5 μm-filtered seawater from the reef adjacent to the research center. Its
 temperature was held at constant 29.43 ± 0.32 °C using a
 temperature-controlling device including a chiller and a heater (Titanium
 Heater 100 W, Schego, Germany; Temperature Switch TS 125, HTRONIC,
 Germany; Aqua Medic Titan 1500 Chiller, Germany). LED lights (135 W, Hydra
 Fiftytwo HD LED, Aqua Illumination, USA) mimicked the average light
 conditions of the sampling sites (Text S1). For
 each heat stress assay (Figure 1 D), two 40 L experimental tanks were set
 up inside each of the 500 L flow-through tanks that were used as
 temperature-controlling water baths (Table S2). The seawater of all four
 experimental tanks was supplied by daily, manual 50% water changes from
 the source tank. Each experimental tank was equipped with a
 temperature-controlling device, one heater, air supply, a small current
 pump and a temperature logger (Temperature Switch TS 125, HTRONIC,
 Germany; Titanium Heater 100 W, Schego, Germany; Koralia nano 900 L/h,
 Hydor, Italy; HOBO Pendant Temperature/Light 8K Data Logger, Onset, USA).
 Two coral fragments per coral colony were randomly distributed among the
 four tanks “34 °C” (<em>n</em> = 2) and “29 °C”
 (<em>n</em> = 2), resulting in one fragment per colony per
 treatment. The 34 °C-treatment was established over the course of one day
 by ramping temperatures from 29 °C to 34 °C for 4 h, holding at 34 °C for
 5 h or 6 h (<em>Pocillopora </em>sp.) or for 6 h or 7 h
 (<em>Porites</em> sp.), and decreasing temperatures to 29 °C
 within 4 h. After the heat exposure, corals were maintained at ambient
 temperatures of 29 °C for 10 h until the next day. While<em>
 Pocillopora </em>sp. fragments were subjected to the short-term heat
 exposure once, resulting in a 24 h experiment,
 <em>Porites</em> sp. corals were exposed to the treatment over
 two consecutive days resulting in a duration of 72 h (Figure 1
 D)<em>.</em> <strong>Coral stress response
 variables</strong> We measured two variables that
 assessed the thermal stress response of each fragment before and after
 each heat stress assay (timepoints (1) and (2) in Figure 1 D). Tissue
 coloration, a proxy for microalgal symbiont cell density in coral tissues
 and therefore an indicator of holobiont health and coral bleaching
 severity, was assessed using a “bleaching score”. The coloration of each
 individual fragment was visually categorized on the scale from 1
 (bleached, pale tissues) to 6 (healthy, dark tissues) using a coral
 bleaching chart (Siebeck et al.,
 2006). A minimum and maximum score was recorded per fragment
 and averaged. Photosynthetic efficiency of microalgal symbionts was
 determined by measuring effective quantum efficiency (yield Φ PSII = (Fm’
 – F) / Fm’ = ΔF / Fm’, (Genty et al.,
 1989) of electron transport using a pulse amplitude-modulated
 fluorometer (Diving-PAM, Walz, Germany). <strong>Statistical
 analyses</strong> ∆-values of each measured thermal
 stress response variable (end – start of each experimental part) were
 calculated to reflect their change over time. Based on these ∆-values,
 effect sizes were estimated using <em>dabestR </em>v0.2.3 6
 (Ho et
 al., 2019). Effects of the high temperature treatment (“Heat”
 vs. “Ambient”) were compared between the sites of origin (“IW exposed
 shore” and “IW sheltered shore”) and between the seasons (“Season of IW
 stimulus presence” and “Season of IW stimulus absence”). Statistical
 significance was tested in<em> R</em> (R Core Team,
 2013) using linear mixed effect models
 (<em>nlme</em> v4 3.1-148 and <em>lme4
 </em>v1.1-23 package). Where applicable, coral colony genotype was
 used as a random factor. References Brown, B.,
 & Phongsuwan, N. (2012). Delayed mortality in bleached massive
 corals on intertidal reef flats around Phuket, Andaman Sea, Thailand.
 <em>Phuket Marine
 Biological Center Research Bulletin</em>,
 <em>48</em>(April 2010), 43–48. Wall, M., Schmidt, G. M., Janjang, P., Khokiattiwong, S., & Richter, C. (2012). Differential Impact of Monsoon and Large Amplitude Internal Waves on Coral Reef Development in the Andaman Sea. <em>PloS One</em>, <em>7</em>(11), e50207. https://doi.org/10.1371/journal.pone.0050207 Schmidt, G. M., Wall, M., Taylor, M., Jantzen, C., & Richter, C. (2016). Large-amplitude internal waves sustain coral health during thermal stress. <em>Coral Reefs </em>, 1–13. https://doi.org/10.1007/s00338-016-1450-z Siebeck, U. E., Marshall, N. J., Klüter, A., & Hoegh-Guldberg, O. (2006). Monitoring coral bleaching using a colour reference card. <em>Coral Reefs </em>, <em>25</em>(3), 453–460. https://doi.org/10.1007/s00338-006-0123-8 Ho, J., Tumkaya, T., Aryal, S., Choi, H., & Claridge-Chang, A. (2019). Moving beyond P values: data analysis with estimation graphics. <em>Nature Methods</em>, <em>16</em>(7), 565–566. https://doi.org/10.1038/s41592-019-0470-3 Genty, B., Briantais, J.-M., & Baker, N. R. (1989). The relationship between the quantum yield of photosynthetic electron transport and quenching of chlorophyll fluorescence. <em>Biochimica et Biophysica Acta (BBA) - General Subjects</em>, <em>990</em>(1), 87–92. https://doi.org/10.1016/S0304-4165(89)80016-9 Jain, T., Buapet, P., Ying, L., & Yucharoen, M. (2023). Differing Responses of Three Scleractinian Corals from Phuket Coast in the Andaman Sea to Experimental Warming and Hypoxia. <em>Journal of Marine Science and Engineering</em>, <em>11</em>(2), 403. https://doi.org/10.3390/jmse11020403
description: # Natural thermal stress-hardening of corals through cold temperature
 pulses in the Thai Andaman Sea
 ## Description of the data
 and file structure This dataset includes all data for the manuscript
 "Natural thermal stress-hardening in corals: Cold-water pulses boost
 heat resistance" by Wall M./Roik A. et al. which investigated the
 phenomenon of stress-hardening through environmental priming in corals
 during two distinct seasons. Data were collected at the Andaman Sea coast
 of Thailand. This data collection contains environmental data (temperature
 °C, HOBO Pendant Temperature/Light 8K Data Logger, Onset, USA), coral
 bleaching score data (decline of coral tissue coloration, assessed as
 established in Siebeck et al., 2006), and the effective quantum efficiency
 (yield Φ PSII, pulse amplitude-modulated fluorometer Diving-PAM, Walz,
 Germany). The two latter variables are metrics that capture the coral
 stress response and were assessed during short-term heat stress assays
 (the mothofology is also described in details in Doering et al. 2021). The
 stress response data were the delta values of pre-heat stess and post-heat
 stress measurements. The stress response data were analyzed comparing
 between reef sites of origin and seasons using effect sizes (following the
 instruction to the R package dabestR v0.2.3 6 (Ho et al., 2019)).
 Additionally, linear-Model based analyses were conducted with the response
 data (following instructions in the package nlme v4 3.1-148 and lme4
 v1.1-23, R). All analyses were performed in R studio. Visualizations were
 generated via ggplot2 and assembled in Adobe Illustrator. These figures
 are also shared herein. ### DATA FILE STRUCTURE EnvironPrim_Data.tar.gz
 EnvPriming_ReadMe.md \---EnvPriming_data +---Data \|
 Input_Data_delta_BSC.xlsx # Input data for downstream analyses. Contains
 the delta values of bleaching score averages calculated from the max. and
 min. values recorded at each asesssement time point. Delta values are
 mostly negative, i.e. = indicating coral bleaching and delcining health =
 stress response; the data is organzsed into 4 tabs by coral species and
 season; ; All acronyms in the headers and tabs are explained in the LEGEND
 SECTION below \| Input_Data_delta_PAM.xlsx # Input data for downstream
 analyses. Contains the delta values of the effective quantum efficiency
 (yield Φ PSII) measured by a diving-PAM (pulse amplitude-modulated
 fluorometer); the data is organized into 4 tabs by coral species and
 season; All acronyms in the headers and tabs are explained in the LEGEND
 SECTION below \| Rawdata_BSC.xlsx # Original collected data (transcript
 from underwater proof field sheets). Contains bleaching scores recorded
 during the heat stress assays, see score scala by Siebeck et al., 2006 (we
 use internsity scores 1-6). At each assessment the minimum and the maximum
 score of a fragment were recorded; All acronyms in the headers and tabs
 are explained in the LEGEND SECTION below \| Rawdata_PAM.xlsx # Original
 collected data (transcript from underwater proof field sheets). Contains
 effective quantum efficiencies (yield Φ PSII) measured by a diving-PAM
 (pulse amplitude-modulated fluorometer); At each assessment the 3
 measurements per coral fragment were recorded; All acronyms in the headers
 and tabs are explained in the LEGEND SECTION below +---Effectsizes_output
 \| Fig_3_CB.tif # calculated effect sizes from dabestR are represented
 along a color gradient in this heatmap \| Fig_4_CB.tif # this figure is a
 composition of all "Cumming estimation plot" from dabestR sorted
 by coral species and season \| Fig_3-4_Tables.xlsx # the tabs contain:
 TAB1) raw output from the dabestR analyses (bleaching score data); TAB2)
 raw output from the dabestR analyses (diving-PAM data); TBA3) the output
 from Tab1) and Tab2) organized in a summary table; TAB4)summary table form
 Tab3) rearranged for plotting the values in heatmaps \| Fig3_Rcode.html #
 R code produces the elements for the heatmap in Fig_3, R code, versions
 and results \| Fig4_Rcode_bsc.html # R code for dabestR conducted with
 bleaching score data, R code, versions and results \| Fig4_Rcode_pam.html
 # R code for dabestR conducted with diving-PAM data, R code, versions and
 results +---LMs_output \|
 Rcode_MixedModels_bsc_poc_apr_Sep2021FinalV1.html # Statistical analysis
 of delta-bleaching score , R code, versions and results \|
 Rcode_MixedModels_bsc_poc_nov_Sep2021FinalV1.html # Statistical analysis
 of delta-bleaching score , R code, versions and results \|
 Rcode_MixedModels_bsc_por_apr_Sep2021FinalV1.html # Statistical analysis
 of delta-bleaching score , R code, versions and results \|
 Rcode_MixedModels_bsc_por_nov_Sep2021FinalV1.html # Statistical analysis
 of delta-bleaching score , R code, versions and results \|
 Rcode_MixedModels_pam_poc_apr_Sep2021FinalV1.html # Statistical analysis
 of Delta-Y (effective quantum yield), R code, versions and results \|
 Rcode_MixedModels_pam_poc_nov_Sep2021FinalV1.html # Statistical analysis
 of Delta-Y (effective quantum yield), R code, versions and results \|
 Rcode_MixedModels_pam_por_apr_Sep2021FinalV1.html # Statistical analysis
 of Delta-Y (effective quantum yield), R code, versions and results \|
 Rcode_MixedModels_pam_por_nov_Sep2021FinalV1.html # Statistical analysis
 of Delta-Y (effective quantum yield), R code, versions and results
 +---Temperature \| Fig_2_CB.tif # Timeseries plot of temperature records
 from the study sites; created in R environment \|
 Input_Data_Temperature.xlsx # In situ temperatures in the study sites RE
 and RW (one site presented per tab) recorded in Celsius using HOBO Pendant
 loggers \| Fig_2_Rcode_Temperature.R # R Code that generates Fig_2, R
 code, versions and results ### LEGEND POC = Pocillopora sp. (coral
 species) POR = Porites sp. (coral species) season 1 = April 2018 (season
 of internal waves peak) season 2 = November 2018 (season of internal waves
 absence) apr = April nov = November site = study site / reef location
 where corals were collected RW = Ko Racha West Shore 7.595530°N,
 98.354320°E RE = Ko Racha East Shore 7.598910°N, 98.373100°E treatment =
 temperature exposure treatment of the short term heat stress assays (also
 see Doering et al. 2021) A = ambient temperature treatment of 29 °C
 (control treatment) H = heat exposure of 34 °C colony = indicated the
 coral colony number N = Replicate number tank = Experimental tanks
 "A1", "A2", "H1", "H2"
 deltaBSCboth = delta values of pre-heat and post-heat bleaching score
 measurements deltaY = delta values of pre-heat and post-heat measurements
 by PAM (i.e., effective quantum efficiency (yield Φ PSII) measured by a
 diving-PAM, pulse amplitude-modulated fluorometer) Y 1-4 = technical
 replicates of effective quantum yield measurements Y1, Y2, Y3, Y4
 (diving-PAM) NA = value not available HM = heatmap IW = internal waves LMs
 = Linear Models PAM = pulse amplitude-modulated fluorometer Timestamp =
 date and time of temperature record from the HOBO Pendant loggers dd/mm/yy
 hh:mm Temperature = recorded in situ temperatures in Celsius ###
 REFERENCES Doering, T., Wall, M., Putchim, L., Rattanawongwan, T.,
 Schroeder, R., Hentschel, U., & Roik, A. (2021). Towards enhancing
 coral heat tolerance: a “microbiome transplantation” treatment using
 inoculations of homogenized coral tissues. Microbiome, 9(1), 102.
 Ho, J., Tumkaya, T.,
 Aryal, S., Choi, H., & Claridge-Chang, A. (2019). Moving beyond P
 values: data analysis with estimation graphics. Nature Methods, 16(7),
 565–566. Siebeck, U. E.,
 Marshall, N. J., Klüter, A., & Hoegh-Guldberg, O. (2006). Monitoring
 coral bleaching using a colour reference card. Coral Reefs, 25(3),
 453–460. ## Authors:data
 preparation and data curation Dr. Marlene Wall,
 Dr. Anna Roik,
 ## Sharing/Access
 information The original preprint can be accessed from here:
 
 Supplementary material can be downloaded here: