DOI: 10.5061/dryad.ncjsxkt57
title: Effects of benign and heat-stress conditions on parent clones and their selfed offspring of the pea aphid

description: In the face of global climate change, understanding the interaction
 between heat stress and inbreeding depression is a critical issue. We
 evaluated the magnitude of inbreeding depression under benign and
 heat-stress conditions using parent clones and their selfed offspring of
 Acyrthosiphon pisum.    The number of nymphs produced by
 an adult during the first seven days was counted, and the total area of
 all nymphs was measured as fitness components of a clone. Inbreeding
 depression was observed during the development of the fundatrix and was
 exemplified by the occurrence of sterile fundatrices, which accounted for
 28.6% of the adults.    Once clonal lines were
 established, no significant overall differences were detected in these
 fitness components between a parent clone and its 30 selfed-offspring
 clones at a constant 20 °C. However, pairwise comparisons revealed a
 significant reduction in the fitness components in a few selfed clones,
 whereas one selfed clone significantly outperformed the parent in terms of
 the total area of the nymphs. These results demonstrate substantial
 variation in the fitness components among sibling clones derived from
 selfing.    However, when clones were exposed to either
 a single heat stress (35 °C for 6 h) or repetitive heat stresses (35 °C
 for 6 h over three consecutive days), inbreeding depression was
 exacerbated under the more intense stress conditions. These results
 suggest that inbred clones could spread in the population in regions with
 mild climates, but are likely to be largely eliminated when exposed to
 high summer temperatures.
description: In the first experiment, a single clone of the pea aphid
 (Acyrthosiphon pisum, clone 05E) and its selfed offspring clones were
 prepared. We evaluated the fecundity and growth rate of each clone at
 20 °C under a 16L:8D photoperiod. From each clone, a single first-instar
 nymph was selected as the test aphid and transferred to a new seedling,
 where it was reared to adulthood. The first day of larviposition was
 recorded as day 1, and on day 7, all offspring produced were collected and
 preserved in 80% ethanol. For each of the 30 selfed clones, we prepared 10
 to 12 replicates (mean = 11.4), and for the parental clone 05E, we
 prepared 13 replicates. The number of nymphs produced over the first seven
 days (7-day fecundity) was counted. All nymphs were placed in a petri dish
 containing a small amount of 80% ethanol, and their image was captured
 using a digital microscope (Dino-Lite Edge, AnMo Electronics) under
 standardized lighting conditions. Images were binarized using a consistent
 threshold in ImageJ, and the total area of all nymphs was calculated based
 on the processed images (https://imagej.net/ij/). The second experiment involved two families: the parental clone 05E and its selfed offspring (05E-self), and the parental clone 07C and its selfed offspring (07C-self). In each of the four clones, five first-instar nymphs were randomly selected within 6 hours of birth and transferred onto a new seedling inside a plastic cage. Seedlings with nymphs were randomly assigned to one of two heat stress treatments or a control. In the single heat stress treatment (SH), cages containing first instars were transferred from 20 °C to 35 °C and maintained at this temperature for 6 hours. After treatment, they were returned to 20 °C under a 16L:8D photoperiod. In the repetitive heat stress treatment (RH), the SH treatment was repeated for three consecutive days, with intervals at 20 °C between heat exposures. In the control (CT), first instars were reared on seedlings continuously at 20 °C. For the SH, RH, and CT treatments, 20–51 aphids (mean = 35.5), 35–40 aphids (mean = 37.8), and 25–35 aphids (mean = 29.3) were used, respectively. After completion of the SH and RH treatments, surviving nymphs were individually transferred to new seedlings and maintained at 20 °C. This transfer was performed because heat stress may have adversely affected the broad bean seedlings. When the nymphs reached the fourth-instar stage, all test aphids—including those in the control group—were transferred individually to new seedlings and allowed to reproduce. Larviposition and mortality were monitored twice daily. On day 7, all nymphs produced by each test aphid were collected and preserved in 80% ethanol. The 7-day fecundity was thus recorded for each aphid. To evaluate the effects of heat stress on each clone, the 7-day fecundity of the selfed offspring was compared with that of the parental clone. The coefficient of inbreeding depression (δ) was calculated for each treatment and family as follows (Lynch & Walsh, 1998): δ = 1- XI/XO where XO is the mean 7-day fecundity of the parental clone and XI is that of its selfed offspring. Parental clones are assumed to be derived from outbreeding.
description: # Effects of benign and heat-stress conditions on parent clones and their
 selfed offspring of the pea aphid
 [https://doi.org/10.5061/dryad.ncjsxkt57](https://doi.org/10.5061/dryad.ncjsxkt57) ## Description of the data and file structure Here, we provide three data files containing: (1) the number of nymphs produced by test aphids over the first seven days (7-day fecundity) and the total area occupied by the nymphs, (2) mortality data of aphids exposed to different temperature conditions, and (3) fecundity data of aphids exposed to different temperature conditions. The aphid clones used in the experiments are identified, and temperature treatments are coded as CT, SH, and RH. CT (control) indicates a constant temperature of 20 °C; SH (single heat stress) refers to 35 °C for 6 hours followed by 20 °C for the remaining time; and RH (repetitive heat stress) consists of 35 °C for 6 hours per day for three consecutive days, with 20 °C for the rest of the time. ### Files and variables #### File: Figs.\_1\_and\_2-dataset.csv **Description:**  This csv file is used for creating Figs. 1 and 2 and includes the data of the number of nymphs produced by aphids for the first seven days and the total area of the nymphs. Test aphids were randomly selected from a parent clone (05E) and its selfed offspring clones. ##### Variables * Clone type: parent (P) or selfed offspring (Off1 or Off2). Off1 and 2 derived from different females of the same clone (05E). * Clone: parent clone (P) or each clone of its selfed offspring, which was discriminated by offspring type (Off1 or 2) and clone ID following the underscore. * Generation: the number of generations from the fundatrix generation (1) for test aphids. * Sample no: replicates (different test aphids) in each clone. * No. offspring: the number of nymphs produced by test aphids for the first 7 days. * Area in mm2: the total area (mm2) of nymphs produced by test aphids for the first 7 days. * Treatment: temperature treatments including a constant 20 °C only. #### File: Fig.\_3-dataset.csv **Description:**  This file is used for creating Fig. 3 and includes the data of survival (1) or death (0) to adulthood of test aphids under different temperature conditions. Test aphids were selected from four clones: 05E-parent, 05E-offspring, 07C-parent, and 07C-offspring, and subjected to one of three temperature conditions: a constant 20  °C (CT), 35  °C for 6 h (SH), and the repetition of 35  °C for 6 h over 3 days (RH). ##### Variables * ID: test aphids used for the experiment. * Family: 05E family or 07C family. * Clone: clone names consisting of family name (05E or 07C) followed by \_p (parent) or \_self (selfed offspring). * Status: parent (par) or selfed offspring (off). * Treatment: temperature treatments. CT (control), a constant 20 °C; SH (single heat stress), 35 °C for 6h and 20 °C for the remaining time; RH (repetitive heat stress), repetition of 35 °C for 6h over 3 days and 20 °C for the remaining time. * Comparison: combination between clone names and treatments. * Fate: LS, larviposited successfully; LD, larviposited but died; and DB, died before larviposition. * Fate2: 1, survived to adulthood; 0, died before adulthood. #### File: Fig.\_4\_Table\_1-dataset.csv **Description:**  This file is used for creating Fig. 4 and Table 1 and includes the data of the number of nymphs produced by test aphids for the first seven days. Test aphids were selected from four clones: 05E-parent, 05E-offspring, 07C-parent and 07C-offspring, and subjected to one of three temperature conditions: a constant 20  °C (CT), 35  °C for 6 h (SH), and the repetition of 35  °C for 6 h over 3 days (RH). ##### Variables * ID: test aphids used for the experiment. * Family: 05E family or 07C family. * Status: parent (par) or selfed offspring (off). * Clone: clone names consisting of family name (05E or 07C) followed by \_p (parent) or \_self (selfed offspring). * Treatment: temperature treatments. CT (control), a constant 20 °C; SH (single heat stress), 35 °C for 6h and 20 °C for the remaining time; RH (repetitive heat stress), repetition of 35 °C for 6h over 3 days and 20 °C for the remaining time. * Group: groups of aphids that were simultaneously subjected to each temperature treatment in a cage. * Fecundity: the number of nymphs produced for the first 7 days. * Comparison: combination between clone names and treatments. * Comments: comments