https://doi.org/10.5061/dryad.m0cfxpp99

Data derived from experimental measurements during multiple stages of the experiment (F1 adult through F2 early life) analyzed in the categories outlined below. Each category has its own R notebook and corresponding data:

• F1 adult performance (growth in terms of length and weight)
• F1 adult fecundity (spawning success; gonad tissue thickness measured via ultrasound)
• F2 performance (survival at 10, 28, and 97 DPF; size at 10 and 97 DPF; Inorganic/Shell mass at 97 DPF)
• Data documenting seawater chemistry (temperature and pH/pCO2) throughout these phases of the experiment also included.

Description of the data and file structure

F1 Adult performance
Length and Weight measurements analyzed for F1 adult abalone. Data taken in Fall of 2019, Fall of 2020, and Spring of 2021. Difference in length and weight from Fall 2019 to Fall 2020 used as growth metric for analyses.

Files: F1_Weights_Lengths_12.19.23.csv

Code: ORA_F1_performance_8.17.23.Rmd

1. Number of variables: 26

2. Number of cases/rows: 120

   1. Variable list (*denotes used or described in analyses presented in manuscript):

      *Family - Maternal identity of the F1 abalone (individuals C, D, E as outlined in Swezey et al. 2020)

      *Juv_Treatment - (Juvenile Treatment) Treatment the F1 abalone experienced during the first three months of life.

      *Adult_Treatment - Treatment the F1 abalone experienced as adults during the second exposure.

      Code - A short hand way to indicate the combination of juvenile and adult treatment each abalone experienced (W = High-High, X = High-Low, Y = Low-High, Z = Low-Low).

      Juv_Adult_Code - An abbreviated indication of the combined juvenile and adult treatment each abalone experienced (Hi = High pCO2, Lo = Low pCO2).

      *Tag - The individual identifying tag number demarcating each individual abalone throughout the experiment.

      *Bucket_2019 - The bucket the adult F1 abalone were housed in during the 11 month adult exposure portion of the experiment (Nov 2019 - Oct 2020). Each bucket contained 5-7 adult abalone.

      ID - A unique numerical identification for each individual abalone independent of their tag number.

      Length_11_19 - The length of the abalone as measured along the longest axis of the shell in November of 2019 (mm).

      Weight_11_19 - The weight of the abalone measured via balance in November of 2019 (g).

      Notes_11_19 - Notes taken during the measuring of adult abalone in November of 2019.

      Length_10_20 - The length of the abalone as measured along the longest axis of the shell in October 2020 (mm).

      Weight_10_20 - The weight of the abalone measured via balance in October of 2020 (g).

      Notes_10_20 - Notes taken during the measuring of adult abalone in October of 2020.

      Length_4_21 - The length of the abalone as measured along the longest axis of the shell in April of 2021 (mm).

      Weight_4_21 - The weight of the abalone measured via balance in April of 2021 (g).

      4_21_Notes - Notes taken during the measuring of adult abalone in April of 2021.

      *Length_Change_2019_2020 - The length of each abalone measured in November of 2019 subtracted from their length as measured in October 2020 (mm).

      *Weight_Change_2019_2020 - The weight of each abalone measured in November of 2019 subtracted from their weight as measured in October 2020 (g).

      Length_Change_2020_2021 - The length of each abalone measured in October 2020 subtracted from their length as measured in April 2021 (mm).

      Weight_Change_2020_2021 - The weight of each abalone measured in October 2020 subtracted from their weight as measured in April 2021 (g).

      Length_Change_2019_2021 - The length of each abalone measured in November 2019 subtracted from their length as measured in April 2021 (mm).

      Weight_Change_2019_2021 - The weight of each abalone measured in November 2019 subtracted from their weight as measured in April 2021 (g).

      Weight_Length_2020 - The ratio of Weight to Length (Weight(g)/Length(mm)) as measured in October of 2020.

      Weight_Length_2019 - The ratio of Weight to Length (Weight(g)/Length(mm)) as measured in November 2019.

      WL_Change - Change in Weight/Length Ratio from 2019 to 2020 (Weight/Legth Ratio 2020 - Weight/Length Ratio 2019)

3. Missing data codes: None

4. Specialized formats or abbreviations used:

   High = Stressful high pCO2 (lower pH) treatment

   Low = Control or ambient low pCO2 (higher pH) treatment

F1 Fecundity\
Spawning success (binary response) along with gonadal tissue measurements via ultrasound documented during spawning event in Spring of 2021.

Files: GonadSexSpawn_12.18.23.csv

Code: ORA_F1_fecundity_8.17.23.Rmd

1. Number of variables: 14
2. Number of cases/rows: 143

3. Variable list (*denotes used or described in analyses presented in manuscript):

   *Family - Maternal identity of the F1 abalone (individuals C, D, E as outlined in Swezey et al. 2020)

   *Juv_Treatment - (Juvenile Treatment) Treatment the F1 abalone experienced during the first three months of life.

   *Adult_Treatment - Treatment the F1 abalone experienced as adults during the second exposure.

   Treatment_Code - A short hand way to indicate the combination of juvenile and adult treatment each abalone experienced (W = High-High, X = High-Low, Y = Low-High, Z = Low-Low).

   *Tag_ID - The individual identifying tag number demarcating each individual abalone throughout the experiment.

   *Bucket - The bucket the adult F1 abalone were housed in during the 11 month adult exposure portion of the experiment (Nov 2019 - Oct 2020). Each bucket contained 5-7 adult abalone.

   Spawned - Denotes whether the abalone successfully spawned (produced sperm or eggs) during the spawning event in April of 2021 (responses are "yes" or "no").

   *Spawned_Binom - Denotes whether the abalone successfully spawned (produced sperm or eggs) during the spawning event in April of 2021 (responses are binary 0 or 1).

   *Sex - The sex of the abalone as determined by their production of either sperm or eggs across spawning attempts (October 2020 and April 2021). Blank cells denote animals where sex was not able to be determined as they did not successfully spawn.

   Gonad_Score - A qualitative score given to abalone during ultrasound that captures the thickness/ripeness of the gonadal tissue (see Boles et al. 2022 for full details).

   Gonad_mean_Winter - The thickness of the gonadal tissue as measured from still ultrasound images and Fiji software. Values are calculated from averaging three measurements across the thickest, medial portion of the gonad tissue (cm). Ultrasound images captured in January of 2021.

   *Gonad_mean_Spring - The thickness of the gonadal tissue as measured from still ultrasound images and Fiji software. Values are calculated from averaging three measurements across the thickest, medial portion of the gonad tissue (cm). Ultrasound images captured on the day of spawning, prior to spawning, in April of 2021.

   Length_4_21 - The length of the abalone as measured along the longest axis of the shell in April of 2021 (mm).

   Gonad_Spring_Length - The gonad thickness measured in April 2021 divided by the length of the animal in April of 2021 to see if animal size impacted gonadal thickness.

4. Missing data codes:

   Sex - Blank cells denote animals where sex was not able to be determined as they did not successfully spawn.

   Gonad Score - Blank cells denote animals where we were unable to get a clear ultrasound image and therefore did not get to accurately score.

   Gonad Mean (Winter and Spring) - We were not able to ultrasound all abalone prior to there spawning. Missing values indicate individuals where we were unable to capture still images to analyze.

   Length - Blank cells are missing data from that timepoint.

5. Specialized formats or abbreviations used:

   High = Stressful high pCO2 (lower pH) treatment

   Low = Control or ambient low pCO2 (higher pH) treatment

F2 performance
Survival was calculated from animal counts per settlement cup at 10, 28, and 97 DPF (days post-fertilization) and survival from 10-28 and 28-97 DPF periods were analyzed. Growth (area of abalone shell) was measured via microscope photos at 10 DPF and 97 DPF and calculated in Fiji image processing software. Final dry weights were obtained using a microbalance. Foil boats were placed in muffle furnace and weighed individually empty (starting weight of foil boat), then weighed with dried animals (taken at 97 DPF) (total mass), then after boats placed into muffle furnace and all organic matter burned off (shell/inorganic mass), this final weight subtracted from the dried weight gives organic mass and finally organic/inorganic mass ratio can be calculated from there.

Files: ORA_finalCounts_6.16.22.csv; Ash_weight_abalone_12.19.23.csv; 10dpf_Size.csv; F2_GrowthData_Density_8.23.22.csv

Code: ORA_F2_performance_8.17.23.Rmd

ORA_finalCounts_6.16.22.csv

1. Number of variables: 15
2. Number of cases/rows: 192

3. Variable list (*denotes used in analyses presented in manuscript):

   Treatment_Code - A short hand way to indicate the combination of juvenile and adult treatment each F1 parent abalone experienced (W = High-High, X = High-Low, Y = Low-High, Z = Low-Low).

   Rep - The replicate number of each cup used to house the baby F2 abalone during the exposure period. Each treatment combination (F1 early life, F1 adult, F2 early life) had 24 replicate cups (8 treatment combinations, 192 cups total).

   *ID - The unique identifier for each cup combining treatment code and replicate.

   *Current_Treatment - The treatment applied to each cup housing F2 offspring abalone for the three month exposure period.

   *Juv_Treatment - (Juvenile Treatment) Treatment the F1 parental abalone experienced during the first three months of life.

   *Adult_Treatment - Treatment the F1 parentl abalone experienced as adults during the second exposure.

   DPF_10_Total - The number of abalone observed in each cup via dissecting microscope at 10 days post fertilization.

   DPF_28_Total - The number of abalone observed in each cup via dissecting microscope at 28 days post fertilization.

   DPF_97_Total - The number of abalone observed in each cup via dissecting microscope at 97 days post fertilization.

   *Surv_10_28 - The proportion of abalone that survived in each cup from day 10 to day 28 post fertilization.

   *Surv_28_97 - The proportion of abalone that survived in each cup from day 28 to day 97 post fertilization.

   Surv_10_97 - The proportion of abalone that survived in each cup from day 10 to day 97 post fertilization.

   DPF10_minus_28 - The number of surviving abalone in each cup from day 10 to day 28 post fertilization (subtract counts from day 10 from counts from day 28).

   DPF10_minus_97 - The number of surviving abalone in each cup from day 10 to day 97 post fertilization (subtract counts from day 10 from counts from day 97).

   DPF28_minus_97 - The number of surviving abalone in each cup from day 28 to day 97 post fertilization (subtract counts from day 28 from counts from day 97).

4. Missing data codes: Empty cells in Survival and columns subtracting counts from each other (e.g., DPF10_minus_28) indicate that all animals had died (count = zero) at the earliest time point so there was no calculation to conduct (for example, if all animals were dead at 28 DPF there is no value for the proportion of survivors between 28 DPF and 97 DPF, cannot divide by zero). In some cases, there was an error in the counts and more animals were counted at a later time point than there had been previously (this happened between 10 and 28 DPF as abalone are hardest to count when they are young and very small and some still swimming and yet to settle, these were removed as outliers as survival rate cannot be above 100%).

5. Specialized formats or abbreviations used:

   High = Stressful high pCO2 (lower pH) treatment

   Low = Control or ambient low pCO2 (higher pH) treatment

   DPF = days post fertilization

Ash_weight_abalone_12.19.23.csv

1. Number of variables: 21
2. Number of cases/rows: 96

3. Variable list (*denotes used in analyses presented in manuscript):

   Weigh_boat_num - The number assigned to the foil weigh boat used to measure dry and ash weight of abalone.

   Treatment - A shorthand that encompasses all treatments into one code, current treatment (Hi = High, Lo = Low), parental treatment (W = High-High, X = High-Low, Y = Low-High, Z = Low-Low) and the cup replicate (1-24).

   *Current_Trt - The treatment applied to each cup housing F2 offspring abalone for the three month exposure period.

   Trt_Code - A short hand way to indicate the combination of juvenile and adult treatment each F1 parent abalone experienced (W = High-High, X = High-Low, Y = Low-High, Z = Low-Low).

   *F1_juv_Trt - (Juvenile Treatment) Treatment the F1 parental abalone experienced during the first three months of life.

   *F1_adult_Trt - Treatment the F1 parental abalone experienced as adults during the second exposure.

   Cup_ID_Code - The replicate number of each cup used to house the baby F2 abalone during the exposure period.

   *Number_abalone - The number of abalone left in each cup after the three month experiment that were used in these measurements. Each weigh boat contains abalone originating from a single cup.

   Weigh_boat - The weight of the foil weigh boat used to measure the abalone after being ashed at 500°C for 3h and weighed using a microbalance (µg).

   Dry_abs_boat - The weight of the dried abalone (paced drying oven at 30°C for 24 h) plus the weight of the foil boat µg.

   Ash_abs_boat - The weight of the ashed abalone (boat and dried abalone placed in a muffle furnace heated to 460°C for 4 h) and the weigh boat (µg).

   *Dry_abs - The weight of the dried abalone and the weigh boat minus the weight of the foil boat on its own (µg).

   *Ash_abs - The weight of the ashed abalone and the weigh boat minus the weight of the foil boat on its own (µg). This represents the shell or inorganic mass of the abalone.

   *Dry_minus_ash - The weight of the dried abalone minus the weight of the ashed abalone (µg). This represents the tissue or organic mass of the abalone.

   *Area_Total - The total are of shell as measured in Fiji added across all abalone left in a given cup (mm^2).

   Area_Avg - The total are of shell divided by the number of abalone in a cup to get the average shell area per abalone.

   *Shell_div_AreaTotal - The weight of the ashed abalone (Ash_abs) divided by the total area of shell in the cup (Area_Total) to get the shell mass per unit area (g/mm^2).

   *Tissue_div_AreaTotal - The weight of the dried organic mass or tissue (Dry_minus_ash) divided by the total area of shell in the cup (Area_Total) to get the tissue mass per unit area (g/mm^2).

   *Shell_div_Tissue - The ratio of shell mass to tissue mass as calculated by dividing Shell_div_AreaTotal by Tissue_div_AreaTotal.

   Dry_div_AreaTotal - The weight of the whole animal dried (Dry_abs; Shell and Tissue) divided by the total area of shell in the cup (Area_Total) to get mass per unit area (g/mm^2).

   Notes - Notes relevant to data collection or analysis.

4. Missing data codes: Some extreme outliers removed (see notes column).

5. Specialized formats or abbreviations used: None

10dpf_Size.csv

1. Number of variables: 6
2. Number of cases/rows: 94

3. Variable list (*denotes used in analyses presented in manuscript):

   *F2_Treatment - The treatment applied to each cup housing F2 offspring abalone for the three month exposure period.

   *Adult_Treatment - Treatment the F1 parental abalone experienced as adults during the second exposure.

   *Juv_Treatment - (Juvenile Treatment) Treatment the F1 parental abalone experienced during the first three months of life.

   Code_Treatment - A short hand way to indicate the combination of juvenile and adult treatment each F1 parent abalone experienced (W = High-High, X = High-Low, Y = Low-High, Z = Low-Low).

   Cup - A shorthand that encompasses all treatments into one code, current treatment (Hi = High, Lo = Low), parental treatment (W = High-High, X = High-Low, Y = Low-High, Z = Low-Low) and the cup replicate (1-24).

   *Area - The shell area of a subset of individuals at 10 DPF measured by photographing them under a microscope and using Fiji software tracing the outline of each individual abalone to calculate their area (mm^2).

4. Missing data codes: None.

5. Specialized formats or abbreviations used:

   Hi = High = Stressful high pCO2 (lower pH) treatment

   Lo = Low = Control or ambient low pCO2 (higher pH) treatment

   DPF = days post fertilization

F2_GrowthData_Density_8.23.22.csv

1. Number of variables: 10
2. Number of cases/rows: 101

3. Variable list (*denotes used in analyses presented in manuscript):

   *Current_Trt - The treatment applied to each cup housing F2 offspring abalone for the three month exposure period.

   Code - A short hand way to indicate the combination of juvenile and adult treatment each F1 parent abalone experienced (W = High-High, X = High-Low, Y = Low-High, Z = Low-Low).

   *Juv_Trt - (Juvenile Treatment) Treatment the F1 parental abalone experienced during the first three months of life.

   *Adult_Trt - Treatment the F1 parental abalone experienced as adults during the second exposure.

   Cup - A shorthand that encompasses the parental treatments into one code, parental treatment (W = High-High, X = High-Low, Y = Low-High, Z = Low-Low) and the cup replicate (1-24).

   Cup_ID - A shorthand that encompasses all treatments into one code, current treatment (High, Low), parental treatment (W = High-High, X = High-Low, Y = Low-High, Z = Low-Low) and the cup replicate (1-24).

   Avg_Area - The estimated area per individual abalone (mm^2) calculated from the total area summed across each abalone in the image and divided by the number of abalone in each image (Ab_Num_97dpf). The area for each individual was measured by photographing them under a microscope and using Fiji software tracing the outline of each individual abalone to calculate their area (mm^2) at 97 DPF.

   Ab_Num_97dpf - The number of abalone in each cup and photographed at 97 DPF.

   Ab_Num_28dpf - The number of abalone in each cup and counted at 28 DPF.

   Notes - Notes relevant to data collection or analysis.

4. Missing data codes: None

5. Specialized formats or abbreviations used:

   High = Stressful high pCO2 (lower pH) treatment

   Low = Control or ambient low pCO2 (higher pH) treatment

   DPF = days post fertilization

Water chemistry:

File: ORA_Chemistry Data_12.18.23.xlsx

Tab 1: ORA_F1_Phase 1

Water chemistry measured weekly throughout the adult F1 exposure period (November 2019 - October 2020).

1. Number of variables: 8
2. Number of cases/rows: 1018

3. Variable list:

   Date - The date a measurement was collected.

   Slope_percent - A value that verifies the probe is properly calibrated. Values between 85-100 are considered in acceptable range (see additional notes below).

   Slope_mV - A value that verifies the probe is properly calibrated. These values represent the offset from the slope.

   Source - Which treatment the water was collected from, high or low.

   Bucket_Sump - Where the water was collected from (a bucket housing abalone in the experimental system identified by number, incoming water from the flow-through seawater system prior to entering the experimental system, and sumps that held the treated water for both the high and low treatments).

   pH - The pH of the water sample.

   Temperature - The temperature of the water sample (°C).

   mV - Millivolts of the sample (mV).

4. Missing data codes: Empty cells represent NA or that data were not collected from that source/date. (See additional notes for the Slope_percent and Slope_mV columns).

5. Specialized formats or abbreviations used:

   mV = millivolt; 0 mV = 7 on the pH scale

   Slope percent and Slope mV = These values are recorded after calibrating the pH probe as a way to compare measurements across time and ensure proper functioning of the pH probe. These two values are recorded once and apply to all values collected from the pH probe in a given session/date. They are independent from the observations recorded in that row, they are mapped to a given date.

   High = Stressful high pCO2 (lower pH) treatment

   Low = Control or ambient low pCO2 (higher pH) treatment

Tab 2: ORA_F1_Phase 2

Water chemistry measured weekly throughout the adult F1 exposure period (February 2021 - April 2021). Water samples were collected from their source and placed into falcon tubes for measurements in the lab.

1. Number of variables: 8
2. Number of cases/rows: 333

3. Variable list:

   Date - The date a measurement was collected.

   Slope_percent - A value that verifies the probe is properly calibrated. Values between 85-100 are considered in acceptable range (see additional notes below).

   Slope_mV - A value that verifies the probe is properly calibrated. These values represent the offset from the slope.

   Source - Which treatment the water was collected from, high or low.

   Bucket_Sump - Where the water was collected from (a bucket housing abalone in the experimental system identified by number, incoming water from the flow-through seawater system prior to entering the experimental system, and sumps that held the treated water for both the high and low treatments).

   pH - The pH of the water sample.

   Temperature - The temperature of the water sample (°C).

   mV - Millivolts of the sample (mV).

4. Missing data codes: Empty cells represent NA or that data were not collected from that source/date. (See additional notes for the Slope_percent and Slope_mV columns).

5. Specialized formats or abbreviations used:

   mV = millivolt; 0 mV = 7 on the pH scale

   Slope percent and Slope mV = These values are recorded after calibrating the pH probe as a way to compare measurements across time and ensure proper functioning of the pH probe. These two values are recorded once and apply to all values collected from the pH probe in a given session/date. They are independent from the observations recorded in that row, they are mapped to a given date.

   High = Stressful high pCO2 (lower pH) treatment

   Low = Control or ambient low pCO2 (higher pH) treatment

Tab 3: ORA_F2_Larval Bucket_probe

Water chemistry measured in all larval buckets twice throughout the larval exposure period (0-7 DPF). Water samples were collected from their source and placed into falcon tubes for measurements in the lab.

1. Number of variables: 8
2. Number of cases/rows: 50

3. Variable list:

   Date - The date a measurement was collected.

   Slope_percent - A value that verifies the probe is properly calibrated. Values between 85-100 are considered in acceptable range (see additional notes below).

   Slope_mV - A value that verifies the probe is properly calibrated. These values represent the offset from the slope.

   Source - Which treatment the water was collected from, high, low, or incoming (water from the flow-through seawater system prior to entering our experimental system).

   Bucket_Sump - Where the water was collected from (a bucket housing larval abalone in the experimental system identified by letter or sumps that held the treated water for both the high and low treatments).

   pH - The pH of the water sample.

   Temperature - The temperature of the water sample (°C).

   mV - Millivolts of the sample (mV).

4. Missing data codes: Empty cells represent NA or that data were not collected from that source/date. (See additional notes for the Slope_percent and Slope_mV columns).

5. Specialized formats or abbreviations used:

   mV = millivolt; 0 mV = 7 on the pH scale

   Slope percent and Slope mV = These values are recorded after calibrating the pH probe as a way to compare measurements across time and ensure proper functioning of the pH probe. These two values are recorded once and apply to all values collected from the pH probe in a given session/date. They are independent from the observations recorded in that row, they are mapped to a given date.

   High = Stressful high pCO2 (lower pH) treatment

   Low = Control or ambient low pCO2 (higher pH) treatment

Tab 4: ORA_F2_in situ

Water chemistry measured in all larval buckets twice throughout the larval exposure period (0-7 DPF). Water samples were collected from their source and placed into falcon tubes for measurements in the lab (pH) as well as measured in situ within the system (Insitu pH) and the pH reading from our sensors built into the experimental system were also recorded.

1. Number of variables: 9
2. Number of cases/rows: 81

3. Variable list:

   Date - The date a measurement was collected.

   Salinity - Salinity of the seawater (ppt) measured once per observation session on incoming seawater prior to entering the experimental system.

   Source - Which treatment the water was collected from, high, low, or incoming water prior to entering the experimental system.

   Bucket_Sump - Where the water was collected from (sumps that held the treated water for both the high and low treatments, empty cells signify that the measurements were collected from incoming seawater, see above).

   Temp - The temperature of the water measured within the sump or in the flow of incoming water (°C).

   pH - The pH of the water as measured by pH probe (see Tabs 1-3).

   Insitu pH - The pH of the water as measured within the experimental system.

   Wall pH - The pH reading from our sensors built into the experimental system.

4. Missing data codes: Empty cells represent NA or that data were not collected from that source/date.

   In the Salinity column, these values are recorded once on incoming water once and apply to all values collected in a given session/date. They are independent from the observations recorded in that row, they are mapped to a given date).

5. Specialized formats or abbreviations used:

   High = Stressful high pCO2 (lower pH) treatment

   Low = Control or ambient low pCO2 (higher pH) treatment

Code/Software

We performed all analyses using R version 3.5.1 and the lme4 package for model construction (Bates et al. 2015). We assessed the significance of fixed effects in our models by analysis of deviance, Type II Wald chi-square tests using the ANOVA function from the car package (Fox & Weisberg 2019). For models with scaled parameters, we used a Kenward-Roger approximation of degrees of freedom and used these to assess the significance of fixed effects using F scores (Venables & Ripley 2002, Kenward & Roger 2009). We explored any potential pair-wise differences using a Bonferroni correction and the “contrasts” function in the emmeans package (Searle et al. 1980).