Inbreeding depression in a sexually selected weapon and the homologue in females
Data files
Nov 08, 2023 version files 71.28 KB
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inb560_-_for_submission_JEB.xlsx
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README.md
Nov 20, 2023 version files 1.26 GB
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DATA_inb560.csv
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Pictures_of_all_the_measured_insectsX.zip
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README_DATA_inb560.txt
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README_pictures_of_all_the_measured_insects.txt
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README.md
Abstract
Theory predicts that traits with heightened condition-dependence, such as sexually selected traits, should be affected by inbreeding to a greater degree than other traits. The presence of environmental stress may compound the negative consequences of inbreeding depression. In this study we examined inbreeding depression across multiple traits and whether it increased with a known form of environmental stress. We conducted our experiment using both sexes of the sexually dimorphic leaf-footed cactus bug, Narnia femorata (Hemiptera: Coreidae). Adult male cactus bugs have enlarged hind legs used as weapons in male-male contests; these traits, and their homologue in females, have been previously found to exhibit high condition-dependence. In this study, we employed small developmental group size as an environmental stress challenge. Nymph N. femorata aggregate throughout their juvenile stages, and previous work has shown the negative effects of small group size on survivorship and body size. We found evidence of inbreeding depression for survival and seven out of the eight morphological traits measured, in both sexes. Inbreeding depression was higher for the size of the male weapon and the female homolog. Additionally, small developmental group size negatively affected survival to adulthood. However, small group size did not magnify the effects of inbreeding on morphological traits. These findings support the hypothesis that traits with heightened condition-dependence exhibit higher levels of inbreeding depression.
README: Inbreeding depression in a sexually selected weapon and the homologue in females
https://doi.org/10.5061/dryad.2ngf1vhvw
In this study, we aimed to determine the effects of inbreeding and environmental stress (in the form of reduced group size during development) on traits with heightened condition-dependence versus other morphological traits. For this purpose, we used a true bug (Hemiptera) from the family Coreidae, Narnia femorata. As all hemipterans do, this insect has a nymphal juvenile stage in which small instars (tiny versions of adult) hatch out of the egg and then growth through molting processes between instars until they reach the winged adult stage.
The data presented here are the morphological measurements for all the adult insects used in this study, all 560 insects for which we collected all 8 morphological measurements, all measurements are in millimeters.
The experimental treatments represent the environment that the nymphs experienced from the 2nd instar until they became adults, and their exoskeleton became fully sclerotized (hardened).
DATA_inb560
Here are the column descriptions, all the hind leg traits* are part of the male weapon or the female homologue, and they all display higher levels of condition-dependence that the other morphological trait.
Here are the column descriptions:
Data column title | Description |
---|---|
Rep | treatment repetition within the same treatment combination in the same block (e.g., inbred/. Goes from 1-5 in 2-nymph treatments, and from 1-2 in the 10-nymph treatments. Each individual Rep represents a unique rearing cup. |
Num | Unique bug number assigned to each adult insect used in the experiment |
Sex | F – female or M – male |
Genetic | inbred or outbred, genetic treatment |
Block | Unique block designation, from A through P. Missing blocks E and K due to small sample sizes. |
Family | family number within each block, from 1 to 4, 1 and 2 are the inbred families, and 3 and 4 are the outbred families. |
Den | 2 or 10 nymphs, group size during juvenile development, density treatment. Low density is considered a more stressful environment as survival is lower in this treatment across different experiments, including this one. |
HFL* | hind femur length |
HFW* | hind femur width |
HTA* | hind tibia area |
FFL | front femur length |
HL | head length |
BL | beak length |
HFA* | hind femur area |
PW | pronotum width – proxy for overall body size |
The 8 treatment combinations used in this experiment are as follow:
1. Female inbred/2-nymph
2. Female inbred/10-nymph
3. Female outbred/2-nymph
4. Female outbred/10-nymph
5. Male inbred/2-nymph
6. Male inbred/10-nymph
7. male outbred/2-nymph
8. Male outbred/10-nymph
Pictures_of_all_the_measured_insectsX
Within the Pictures of all the measured insects folder, you will find 18 folders. Each folder corresponds to a single picture-taking bout by the technician. The name of the folder is the date in 2015 when the pictures were taken, e.g., APRIL_10(8) = pictures taken April 10th 2015, and this folder includes the pictures for (8) individual insects. Some days include 2 folders, that just means pictures were taken in two different bouts on the same day. In each folder, you will find 4 pictures per insect (A-D; see key below), and an additional picture of the ruler (one per folder) used to calibrate the measuring software, ImageJ. This is the reason why you must use this folder structure to measure the insects, as the ruler only works to measure the insects contained in the same folder.
The pictures are labelled as follows:
Example: 621_M_O_N_4_2_2_A
Formula: Num_Sex_Genetic_Block_Family_Den_Rep_picture type
- Num = Unique bug number
- Sex = F or M
- Genetic = inbred or outbred
- Block = Unique block designation
- Family = family number within each block
- Den = 2 or 10 nymphs
- Rep = treatment repetition, same family
- Picture type = A – hind legs, B – front legs, C – insect body top side, D – insect body underside
In the DATA_inb560.csv file, you will easily find the corresponding measurements by using the Unique bug number (Num). All the pictures taken are included here but we only used those for 560 insects, those for which we could obtain all 8 measurements.
Methods
Parental generation (Fp): Fifth instar nymphs were collected from the Ordway-Swisher Biological Station (Melrose, FL) in September 2014. Individual nymphs were kept in plastic deli containers, with topsoil, and a cactus pad (Opuntia mesacantha) with ripe fruit attached to it. This rearing protocol works for all life stages in this species (Allen et al., 2018). After nymphs molted into adults and reached sexual maturity (two weeks after molting), we randomly paired males and females.
Parental pairs (Fp): Male-female pairs were allowed to mate freely, and ripe fruit was provided (one per cup). After eggs were laid, we removed them and placed them in a new container with cactus and no fruit (1st instar nymphs do not feed). We selected the first 15 eggs from each of 32 females over a two-week period.
Founding generation nymphs (F0): Once the eggs hatched and nymphs reached the 2nd instar, each female’s nymphs (15) were split equally into three cups. We supplied two ripe fruit per cup. After nymphs molted into adults and reached sexual maturity (two weeks after molting) we paired them as described below.
Founding generation adults (F0): To create inbred and outbred lines we used the ‘block’ design proposed by Roff (Roff,1998; Fox & Reed, 2011; Vega-Trejo et al., 2015; Joseph et al., 2016; Marsh et al., 2017; Hooper & Bonduriansky 2022). A block consisted of two (A + B) randomly paired families from the 32 Fp pairs we had available, therefore creating 16 blocks. From each family we haphazardly chose two females and two males; we crossed offspring from the two families to create two outbred lines (♀A/♂B & ♀B/♂A) and crossed offspring within each family to create two inbred lines (♀A/♂A & ♀B/♂B). This design ensured that inbred and outbred families were created from the same set of alleles providing an advantage over other experimental designs (Fox, 2005). Male-female pairs were allowed to mate freely, and ripe fruit was provided as needed. Eggs from all the pairs were extracted and set up separately, using the same methodology as in the Parental pairs section (see also Allen et al., 2018 for more details). Within each block, we obtained 30 eggs per cross (new F1 families).
The inbred/outbred generation (F1): From each block (N=16) we obtained 60 inbred nymphs (30/cross) and 60 outbred nymphs (30/cross). After molting to the 2nd instar, nymphs from each cross were separated equally into 2-nymph (5 cups) and 10-nymph groups (2 cups). Two-nymph groups got one fruit/cup and 10-nymph groups five fruits/cup. After individuals molted into adults and reached sexual maturity (two weeks after molting) they were frozen.
Insect measurements: The measuring protocol follows the same methodology used by Procter et al. (2012). A digital camera (Canon EOS 50D) attached to a dissecting microscope (Leica M165 C) was used to photograph all the extremities and body; the software ImageJ (Rasband, 2011) was used for the linear and area measurements. The morphological traits chosen were beak length (=rostrum [mouthparts]), head length, pronotum width, front femur length, hind femur length, and hind femur width. Legs and body were separated to facilitate the measuring procedure. Area measurements were taken of the hind tibia and femur, as these are the most enlarged traits part of the male weapon, and the most condition-dependent traits (Allen & Miller, 2017). We used pronotum width as a proxy for body size; in this species, it is highly correlated with overall body size (Gillespie et al., 2014; Miller et al., 2016; Allen & Miller, 2017). Where applicable, means of paired body parts were used for analyses. For the purposes of this study, all male hind leg traits were considered to be sexually selected traits because of their involvement in signaling and fighting (Procter et al., 2012; Nolen et al., 2017); in contrast, female hind leg traits were referred to as the female homologous traits.