DOI: 10.5061/dryad.1jwstqk2w
title: Neglected patterns of variation in transgenerational plasticity: The importance of different sources of environmental variation differs across ages and sexes in a cyprinid fish

description: Adaptive transgenerational plasticity (TGP) requires individuals to
 integrate environmental experience across multiple sources. However, few
 empirical studies have considered that the relative relevance of certain
 sources might vary across ontogeny and sexes. Here, we address this
 knowledge gap by studying inducible antipredator defenses, one of the most
 convincing examples of TGP. We assessed individual and combined effects of
 perceived high predation risk in mothers, fathers, caring males and
 personal environments on the morphology of juvenile, adult male and adult
 female cyprinids Pimephales promelas. Parental rather than personal
 environmental experience determined morphological defense expression
 across ages and sexes, likely because parents had a longer sampling
 period. In juveniles and adult males, egg-mediated environmental
 experience outweighed sperm-mediated environmental experience in the
 induction of body shape differences, likely because eggs can transmit
 information beyond epigenomes. However, in adult females, where body shape
 responses can be interpreted as life-history plasticity, information from
 egg and sperm were equally important, likely resulting from different
 integration mechanisms between morphological and life-history plasticity.
 The importance of care-mediated relative to gamete-mediated variation
 changed between juveniles and adult males, likely because they represent
 short- and long-term environmental experience, respectively. Instead, in
 adult females, both sources were again equally important, potentially
 owing to lag-times of life-history plasticity. Parental care intensity
 only contributed marginally to defense formation. These results highlight
 age- and sex-specific prioritization of different environmental
 experiences so as to generate optimal phenotypes.
description: All methods are described in detail in the main manuscript and
 its supplementary information.
description: # Neglected patterns of variation in transgenerational plasticity: The
 importance of different sources of environmental variation differs across
 ages and sexes in a cyprinid fish
 [https://doi.org/10.5061/dryad.1jwstqk2w](https://doi.org/10.5061/dryad.1jwstqk2w) This data originates from a laboratory study on *Pimephales promelas*. In a factorial split-clutch design, mothers, fathers, caring parents and/or offspring were raised from hatching onwards under different levels of perceived predation risk (high-risk: conspecific alarm cues or low-risk: a water control). Morphological data of offspring was obtained at two developmental stages: in juveniles after completing larval development (18 days age) and in males and females at the onset of sexual maturity (180 days age). Whenever individuals were scheduled to receive parental care during the egg stage, caring parent behavioral data was additionally collected during the 4-day parental care period. Detailed information on how the data was obtained is contained in the corresponding manuscript. ## Description of the data and file structure **Data used in the analysis** Dataset.zip **Structure of Dataset.zip** \_juv - Juvenile data files \_mal - Adult male data files \_fem - Adult female data files \_landmarks files - Contain landmark data from the experimental specimens \_average_landmarks files - Contain landmarks from "average" individuals along principal components, necessary to visualize deformations along axes. \_average_outline files - Contain outlines from "average" individuals along principal components, necessary to visualize deformations along axes. \_addendum files - Contain associated non-landmark data. **Abbreviations in \_addendum** **data files** Family - Identifies the specific parental pair that sampled individuals were born from. Combo - The specific treatment combination that individuals have experienced. Definitions are given below: UNPCC = biparental low risk, personal low risk UNPCA = biparental low risk, personal high risk UPCCC = biparental low risk, low risk care, personal low risk UPCAC = biparental low risk, high risk care, personal low risk XNPCC = maternal high risk, paternal low risk, personal low risk XNPCA = maternal high risk, paternal low risk, personal high risk YNPCC = maternal low risk, paternal high risk, personal low risk YNPCA = maternal low risk, paternal high risk, personal high risk WNPCC = biparental high risk, personal low risk WNPCA = biparental high risk, personal high risk WPCCC = biparental high risk, low risk care, personal low risk WPCAC = biparental high risk, high risk care, personal low risk MatEnv - The maternal environment (A = high risk, C = low risk) PatEnv - The paternal environment (A = high risk, C = low risk) OffEnv - The personal environment (A = high risk, C = low risk) parcare - Identifies whether individuals received parental care (yes) or not (no). Carer.tank - ID of the tank that offspring received parental care in. NOTHING = not applicable. Carer.ID - ID of the male that offspring received parental care from. NOTHING = not applicable. Carer.family - Family identity of the male that offspring received parental care from. NOTHING = not applicable. meancare - The average proportion of care individuals received during their 4-day egg period (assessed daily in 5-minute intervals). NA = not applicable. care.type - Identifies whether parental care was provided to own or foreign (adopted, cross-fostered) eggs. NA = not applicable. tank - Identifies which tank an individual was raised in. ParTreat - Identifies the parental treatment combination. Definitions are given below: U = biparental low risk X = maternal high risk, paternal low risk Y = maternal low risk, paternal high risk W = biparental high risk Father.family - Identifies the specific parental pair that the father was born from. Father.SL..cm - The standard length (form the tip of the snout to the base of the tail fin) of the father (in cm), measured when individuals were paired up. Father.weight..g - The weight of the father (in g), measured when individuals were paired up. Father.Condition - The condition index of the father as calculated from standard length and weight according to Bolger, T. & Connolly, P.L. (1989). The selection of suitable indexes for the measurement and analysis of fish condition. J Fish Biol, 34, 171-182. Mother.family - Identifies the specific parental pair that the mother was born from. Mother.SL..cm - The standard length (form the tip of the snout to the base of the tail fin) of the mother (in cm), measured when individuals were paired up. Mother.weight..g - The weight of the mother (in g), measured when individuals were paired up. Mother.Condition - The condition index of the mother as calculated from standard length and weight according to Bolger, T. & Connolly, P.L. (1989). The selection of suitable indexes for the measurement and analysis of fish condition. J Fish Biol, 34, 171-182. CareEnv - Identifies the environment of the caring parent (PCA = high risk, PCC = low risk, NOTHING = not applicable). OffEnv.nc - Identifies personal environments in the treatments where no parental care was present (A = high risk, C = low risk, NOTHING = not applicable). MatEnvnc - Identifies maternal environments in the treatments where no parental care was present (A = high risk, C = low risk, NOTHING = not applicable). PatEnvnc - Identifies paternal environments in the treatments where no parental care was present (A = high risk, C = low risk, NOTHING = not applicable). ParEnv.c - Identifies parental environments in the treatments where parental care was present (U = biparental low risk, W = biparental high risk). MatEnvPatEnvCombo.nc - Identifies the combination of maternal and paternal environments in the treatments where parental care was absent (CC = both low risk, AA = both high risk, CA = maternal low risk, paternal high risk, AC= maternal high risk, paternal low risk, NOTHING = not applicable). PatEnvOffEnvCombo.nc - Identifies the combination of paternal and personal environments in the treatments where parental care was absent (CC = both low risk, AA = both high risk, CA = paternal low risk, personal high risk, AC= paternal high risk, personal low risk, NOTHING = not applicable). MatEnvOffEnvCombo.nc - Identifies the combination of maternal and personal environments in the treatments where parental care was absent (CC = both low risk, AA = both high risk, CA = maternal low risk, personal high risk, AC= maternal high risk, personal low risk, NOTHING = not applicable). **Photograph of the setup for juvenile images** 2019-01-12-juvenile-photo-setup.jpg This photo shows how the images underlying the juv_landmarks.tps file were obtained. To take a photograph, 18-day old juvenile *Pimephales promelas* were individually placed in a water-filled distortion-free and orthochromatic quartz glass cuvette (100-OS, outer dimensions 1.25x0.75x4.4 cm, inner dimensions 0.95x0.5x4.4 cm, Hellma, Germany) and the opening was immediately covered with Parafilm. The cuvette was then placed on its side in front of a matte black background and an associated ID number on folded paper was stationed next to the cuvette. The whole set-up was illuminated from the top by a 13W LED lamp (S9434, Long Neck Flood 40°, 5000 K, Satco, USA) from 3 cm above the cuvette (light incidence in a 90° angle with the fish). Pictures were then taken with a digital camera (Olympus E-3 with Zuiko Digital ED 50mm 1:2.0 macro objective). This photograph was taken on December 1st, 2019. **Photograph of the setup for adult images** 2019-01-12-adult-photo-setup.jpg This photo shows how the images underlying the mal_landmarks.tps and fem_landmarks.tps file were obtained. To take a photograph, 180-day adult *Pimephales promelas* were individually placed inside of a water-filled 34.5 x 27.0 x 18.5 cm tank where they were held into place within an 87 x 73 x 10 mm central area using a perforated plastic pane with attached sponge stripes. Illumination was provided by two 13W LED lamps (S9434, Long Neck Flood 40°, 5000 K, Satco, USA) from a distance of 15 cm each (light incidence in a 45° angle with the fish). White and size standards on a glass slide were placed next to the fish inside the tank, and an associated ID number on folded paper was stationed next to the fish. Pictures were then taken with a digital camera (Olympus E-3 with Zuiko Digital ED 50mm 1:2.0 macro objective). This photograph was taken on December 1st, 2019. **Sample juvenile images used for analysis** 2018-05-07-juvenile-photo-U1-NPC-A-2_0003.png This image is of a 18-day old *Pimephales promelas* with the ID number 0003. It originates from the family U1. Its parents received the low-risk treatment, it did not receive parental care, and it received the high-risk treatment. It originated from the first clutch of its parents and is part of the second replicate of the same family/treatment combination. We used tpsDig2 to obtain landmarks for our analysis from this picture (located in the juv_landmarks.tps file). This photograph was taken on May 7th, 2018. 2018-05-07-juvenile-photo-W2-NPC-A-1 Clutch 2_0739.png This image is of a 18-day old *Pimephales promelas* with the ID number 0739. It originates from the family W2. Its parents received the high-risk treatment, it did not receive parental care, and it received the high-risk treatment. It originated from the second clutch of its parents and is part of the first replicate of the same family/treatment combination. We used tpsDig2 to obtain landmarks for our analysis from this picture (located in the juv_landmarks.tps file). This photograph was taken on May 7th, 2018. 2018-05-15-juvenile-photo-X3-NPC-C-1_1237.png This image is of a 18-day old *Pimephales promelas* with the ID number 1237. It originates from the family X3. Its mother received the high-risk treatment and its father received the low-risk treatment, it did not receive parental care, and it received the low-risk treatment. It originated from the first clutch of its parents and is part of the first replicate of the same family/treatment combination. We used tpsDig2 to obtain landmarks for our analysis from this picture (located in the juv_landmarks.tps file). This photograph was taken on May 15th, 2018. 2018-09-17-juvenile-photo-Y10-NPC-A-1_1367.png This image is of a 18-day old *Pimephales promelas* with the ID number 1367. It originates from the family Y10. Its mother received the low-risk treatment and its father received the high-risk treatment, it did not receive parental care, and it received the high-risk treatment. It originated from the first clutch of its parents and is part of the first replicate of the same family/treatment combination. We used tpsDig2 to obtain landmarks for our analysis from this picture (located in the juv_landmarks.tps file). This photograph was taken on September 17th, 2018. **Sample adult images used for analysis** 2018-10-27-adult-photo-W1-PCC-C-2 Clutch 5_0545.png This image is of a 180-day old male *Pimephales promelas* with the ID number 0545. It originates from the family W1. Its parents received the high-risk treatment, it received parental care by a low-risk parent, and it received the low-risk treatment. It originated from the fifth clutch of its parents and is part of the second replicate of the same family/treatment combination. We used tpsDig2 to obtain landmarks for our analysis from this picture (located in the mal_landmarks.tps file). This photograph was taken on October 27th, 2018. 2018-10-13-adult-photo-X2-NPC-C-1 Clutch 3_1134.png This image is of a 180-day old female *Pimephales promelas* with the ID number 1134. It originates from the family X2. Its mother received the high-risk treatment and its father received the low-risk treatment, it did not receive parental care, and it received the low-risk treatment. It originated from the third clutch of its parents and is part of the first replicate of the same family/treatment combination. We used tpsDig2 to obtain landmarks for our analysis from this picture (located in the fem_landmarks.tps file). This photograph was taken on October 13th, 2018. 2018-10-21-adult-photo-Y2-NPC-A-2 Clutch 2_1286.png This image is of a 180-day old female *Pimephales promelas* with the ID number 1286. It originates from the family Y2. Its mother received the low-risk treatment and its father received the high-risk treatment, it did not receive parental care, and it received the high-risk treatment. It originated from the second clutch of its parents and is part of the second replicate of the same family/treatment combination. We used tpsDig2 to obtain landmarks for our analysis from this picture (located in the fem_landmarks.tps file). This photograph was taken on October 21st, 2018. **Sample parental care video files of one clutch over the four-day parental care period to derive parental care intensity (variable: "meancare") for analyses** 2018-07-13-D27-day-1-parental-care-video.mp4 This ~10 min long video was taken from an adult pair of *Pimephales promelas*. The male provides parental care for the clutch that contains the individuals that were later photographed. Both parents were from hatching onwards exposed to high-risk and cared for their own clutch here (family W7, high-risk care). The time spent within one standard length of the clutch within the last five minutes of this video was used as a datapoint to derive the data for the variable "meancare" (i.e., the average proportion of care individuals received during their 4-day egg period) in the \_addendum files. The tank ID is D27, and this video depicts the first day of parental care. This video was recorded on July 13th, 2018. 2018-07-14-D27-day-2-parental-care-video.mp4 This ~10 min long video was taken from an adult pair of *Pimephales promelas*. The male provides parental care for the clutch that contains the individuals that were later photographed. Both parents were from hatching onwards exposed to high-risk and cared for their own clutch here (family W7, high-risk care). The time spent within one standard length of the clutch within the last five minutes of this video was used as a datapoint to derive the data for the variable "meancare" (i.e., the average proportion of care individuals received during their 4-day egg period) in the \_addendum files. The tank ID is D27, and this video depicts the second day of parental care. This video was recorded on July 14th, 2018. 2018-07-15-D27-day-3-parental-care-video.mp4 This ~10 min long video was taken from an adult pair of *Pimephales promelas*. The male provides parental care for the clutch that contains the individuals that were later photographed. Both parents were from hatching onwards exposed to high-risk and cared for their own clutch here (family W7, high-risk care). The time spent within one standard length of the clutch within the last five minutes of this video was used as a datapoint to derive the data for the variable "meancare" (i.e., the average proportion of care individuals received during their 4-day egg period) in the \_addendum files. The tank ID is D27, and this video depicts the third day of parental care. This video was recorded on July 15th, 2018. 2018-07-16-D27-day-4-parental-care-video.mp4 This ~10 min long video was taken from an adult pair of *Pimephales promelas*. The male provides parental care for the clutch that contains the individuals that were later photographed. Both parents were from hatching onwards exposed to high-risk and cared for their own clutch here (family W7, high-risk care). The time spent within one standard length of the clutch within the last five minutes of this video was used as a datapoint to derive the data for the variable "meancare" (i.e., the average proportion of care individuals received during their 4-day egg period) in the \_addendum files. The tank ID is D27, and this video depicts the fourth day of parental care. This video was recorded on July 16th, 2018. ## Code/Software **Commented analysis code is contained in the following files** R-Code.html R-Code.Rmd **Software versions used** tpsDig2 '2.32' ( [https://www.sbmorphometrics.org/soft-dataacq.html )](https://www.sbmorphometrics.org/soft-dataacq.html) ``` ## R '4.2.1' ``` ``` ## ggforce ggpmisc ggpp showtext showtextdb sysfonts knitr ## "0.4.1" "0.5.1" "0.4.5" "0.9-5" "3.0" "0.8.8" "1.40" ## svglite scales forcats stringr dplyr purrr readr ## "2.1.0" "1.2.1" "0.5.2" "1.4.1" "1.0.10" "0.3.4" "2.1.3" ## tidyr tibble ggplot2 tidyverse rcompanion MASS geomorph ## "1.2.1" "3.1.8" "3.3.6" "1.3.2" "2.4.18" "7.3-57" "4.0.4" ## rgl RRPP emmeans nortest lmerTest lme4 Matrix ## "0.110.2" "1.3.1" "1.8.1-1" "1.0-4" "3.1-3" "1.1-30" "1.5-1" ## car carData ## "3.1-0" "3.0-5" ```