DOI: 10.5061/dryad.s4mw6m9cm
title: Data for: Does parasitoid species diversity promote protective symbiont diversity?

description: How does diversity in nature come about? One factor contributing to this
 diversity are species interactions; diversity on one trophic level can
 shape diversity on lower or higher trophic levels. For example, parasite
 diversity enhances host immune diversity. Protective symbionts mediate
 host resistance and are therefore also engaged in reciprocal selection
 with their host’s parasites. Here, we applied experimental evolution in a
 well-known symbiont-aphid-parasitoid system to study whether parasitoid
 diversity contributes to maintaining symbiont diversity. We used caged
 populations of black bean aphids (Aphis fabae), containing uninfected
 individuals and individuals infected with different strains of the
 bacterial endosymbiont Hamiltonella defensa, which protects aphids against
 parasitoids. Over multiple generations, these populations were exposed to
 three different species of parasitoid wasps (Aphidius colemani, Binodoxys
 acalephae, or Lysiphlebus fabarum), simultaneous or sequential mixtures of
 these species, or no wasps. Surprisingly, we observed little selection for
 H. defensa in most treatments, even when it clearly provided protection
 against a fatal parasitoid infection. This seemed to be caused by high
 induced costs of resistance: aphids surviving parasitoid attacks suffered
 an extreme reduction in fitness. In marked contrast to previous studies
 looking at the effect of different genotypes of a single parasitoid
 species, we found little evidence for a diversifying effect of multiple
 parasitoid species on symbiont diversity in hosts.
description: Data on aphid demograph from the experimental evolution
 experiments ("basic") was collected by counting aphids (roughly estimated by counting aphids in groups
 of ca. 10 individuals) and mummies (exact counts) and measuring plant size
 (i.e. total stem length of all plants) for each pot in each cage. Prior to
 data analysis, data for each cage was combined to obtain a single value
 for each cage in each generation (see associated paper for
 details). To analyse <em>H. Defensa
 </em>composition ("mol") we first collected aphids at the
 end of generations 3 and 6, extracted their DNA, used diagnostic PCRs to
 amplify part of the bacterial 16S rRNA and part of <em>H.
 defensa</em>'s murE gene for sequencing to distinguish
 <em>H. defensa</em> haplotypes. Data was recorded as either
 <em>H. defensa</em> absent or present and in the later case
 the identity of the haplotype. From this we calculate the proportion of
 <em>H. defensa </em>negative aphids and each <em>H.
 defensa </em>haplotype. This proportion was used in subsequent
 analysis (see associated
 paper for details). Data on the experimental infection test
 ("test") was collected by counting aphids and mummies on each
 plant at different time points (see associated paper for
 details).
description: # **Data for Does parasitoid species diversity promote protective symbiont
 diversity?** ### Experimental procedures We conducted an experimental
 evolution experiment with 30 replicate populations (cages) over six
 generations each with the same initial composition of different haplotypes
 of the protective symbiont Hamiltonella defensa or no H. defensa. In each
 generation our artificial populations were exposed to different wasp
 species either individually or in combination. In each generation we
 collected data on aphid demographics and in generation three and six. In
 addition we conducted infection tests using each possible symbiont
 haplotype – parasitoid species combination. ### Results Contrary to our
 expectations we found little effect of parasitoid treatment on *H.
 defensa* composition. ## **Description of the data and file structure**
 ### basic.txt: Data on aphid demographics collected during transfer in
 each generation. * id1: Unique identifier for each pot-generation
 combination * Generation: Generation * Date: Day when sampling took place
 (DD/MM) * ID: Identity of each cage * ID-orig: in two cases replicates
 were lost very early in the experiments and replaced from another cage of
 the same treatments (see associated paper) * ID_orig states the IDs of
 these cages. In all other cases ID and ID-orig are identical * Pot:
 Identifier of each pot in each cage. In each cage we had three pots each
 containing a single plant * Treatment: Experimental treatment; levels are:
 Baca: Cages exposed to *Binodoxys acalephae*; Lfab: Cages exposed to
 *Lysiphlebus fabarum*; Acol: Cages exposed *Aphidius colemani*; Sim: Cages
 simultaneously exposed to all three wasp species; Seq: Cages sequentially
 exposed to all three wasp species, i.e. t a single species in each
 generation. But alternating between generations. * WaspLine: Wasp line
 used for this cage in this generation. Except for the Seq treatment this
 always corresponds to treatmen * Block: In order to manage the work load,
 work was split over two consecutive days. Block indicates on which of
 these days each cage was handled * N_aphids: Number of aphids * N_mummies:
 Number of mummies * L_stem: Plant length (cm) * N_transferred: Number of
 aphids transferred to the next generation. We aimed for 10 aphids from
 each pot (30/ cage). Where this was not possible due to a lack of
 surviving aphids this number derivates from 10. NA indicates that no data
 could be obtained for this replicate and column. In the remark column it
 indicates that nothing noteworthy happened in this replicate. ### mol.txt:
 Data on the presence/ absence and identity of *H. Defensa* collected in
 generation 3 and 6. Data for generation 0 presents the initial aphid
 composition. * id1: Unique identifier for each pot-generation combination
 * Generation: Generation * ID: Identity of each cage * pot: Identifier of
 each pot in each cage. In each cage we had three pots each containing a
 single plant * Treatment: Experimental treatment; levels are: Baca: Cages
 exposed to *Binodoxys acalephae*; Lfab: Cages exposed to *Lysiphlebus
 fabarum*; Acol: Cages exposed *Aphidius colemani*; Sim: Cages
 simultaneously exposed to all three wasp species; Seq: Cages sequentially
 exposed to all three wasp species, i.e. t a single species in each
 generation. But alternating between generations. * WaspLine: Wasp species
 used for this cage in this generation. Except for the Seq treatment this
 always corresponds to treatment * plate: 96 well plate on which this
 sample was collected and processed * col & row: Location on this place
 (column and row respectively) * H0: *H. defensa* present or absent (*H.
 defensa* present=0, *H.defensa* absent=1) * Hpos: *H. defensa* present or
 absent (*H. defensa* present=1, *H.defensa* absent=0) * weakHam: Band for
 *H. defensa* in the diagnostic PCR weak * H15: *H. defensa* haplotype 15?
 (yes=1, no=0) * H76: *H. defensa* haplotype 76? (yes=1, no=0) * Done:
 Molecular analysis of this sample complete? (yes=1, no=0) * Use_pres: Data
 can be used to analyse the presence/ absence of *H. defensa* * Use_haplo:
 Data can be used to analyse the haplotype identity of *H. defensa*. E.g.
 data in which sequencing failed had to be excluded. ### test.txt: Data for
 the experimental infection experiments * Id: Unique identifier for each
 plant/ pot * Time: Round of the experiment (pre vs. post experimental
 evolution experiment). Some experimental details differed between rounds
 (see accompanying paper) * Symb: Haplotype of *H. defensa* present in
 aphids used in this replicate (H-: *H. defensa* free aphids, H15: *H.
 defensa* haplotyie H15, H76: H. defensa haplotype H76) * Wasp: Wasp used
 for experimental exposure (Baca: *Binodoxys acalephae*; Lfab: *Lysiphlebus
 fabarum*; Acol *Aphidius colemani*) * N_aphids1: Number of aphids prior to
 the addition of parasitoid wasps * N_aphids2: Number of aphids 10 to 11
 days after the addition of parasitoid wasps * N_mummies: Number of mummies
 10 to 11 days after the addition of parasitoid wasps NA indicates that no
 data could be obtained for this replicate and column. In the remark column
 it indicates that nothing noteworthy happened in this replicate. ##
 **Code/Software** We used R, version 4.0.0 (2020-04-24) to analyse data.
 Scripts were written using R markdown (version 2.2) and run in R studio
 (version 2023.06.1). To run the code make sure that the files (basic.txt,
 mol.txt, & test.txt) are in your working directory. The following
 packages were attached or loaded: ### attached base packages: stats,
 graphics, grDevices, utils, datasets, methods, base ### other attached
 packages: stringr_1.4.0, entropart_1.6-4, metacom_1.5.3,
 data.table_1.12.8,SpadeR_0.1.1, gmodels_2.18.1, DHARMa_0.3.2.0,
 factoextra_1.0.7,rcompanion_2.3.25, GLMMadaptive_0.7-0, magrittr_1.5,
 flextable_0.5.10,glmmTMB_1.0.1, vegan_2.5-6, permute_0.9-5,
 tidyr_1.1.0,taRifx_1.0.6.2, optimx_2020-4.2, lawstat_3.4,
 psyphy_0.1-9,blmeco_1.4, emmeans_1.4.7, dplyr_1.0.0,
 purrr_0.3.4,lmerTest_3.1-2, car_3.0-8, carData_3.0-4,
 fitdistrplus_1.1-1,officer_0.3.11, cowplot_1.0.0, gridExtra_2.3,
 reshape_0.8.8,plyr_1.8.6, Hmisc_4.4-0, Formula_1.2-3,
 lattice_0.20-41,multcomp_1.4-13, TH.data_1.0-10, MASS_7.3-51.5,
 survival_3.1-12,mvtnorm_1.1-1, lme4_1.1-23, Matrix_1.2-18, ggplot2_3.3.1
 ### loaded via a namespace (and not attached): readxl_1.3.1, uuid_0.1-4,
 backports_1.1.7,systemfonts_0.2.3, TMB_1.7.16,
 splines_4.0.0,digest_0.6.25, foreach_1.5.0, htmltools_0.4.0,gdata_2.18.0,
 checkmate_2.0.0, cluster_2.1.0,openxlsx_4.1.5, matrixStats_0.56.0,
 Kendall_2.2,sandwich_2.5-1, jpeg_0.1-8.1, colorspace_1.4-1,ggrepel_0.8.2,
 haven_2.3.1, xfun_0.14,crayon_1.3.4, libcoin_1.0-5,
 zoo_1.8-8,iterators_1.0.12, glue_1.4.1, gtable_0.3.0,abind_1.4-5,
 scales_1.1.1, bibtex_0.4.2.2,Rcpp_1.0.4.6, xtable_1.8-4,
 htmlTable_2.0.0,foreign_0.8-78, stats4_4.0.0,
 htmlwidgets_1.5.1,RColorBrewer_1.1-2, acepack_1.4.1,
 ellipsis_0.3.1,modeltools_0.2-23, farver_2.0.3,
 pkgconfig_2.0.3,nnet_7.3-13, multcompView_0.1-8,
 labeling_0.3,tidyselect_1.1.0, reshape2_1.4.4,
 munsell_0.5.0,cellranger_1.1.0, tools_4.0.0, generics_0.0.2,evaluate_0.14,
 EMT_1.1, arm_1.11-1,yaml_2.2.1, knitr_1.28, zip_2.0.4,coin_1.3-1,
 nlme_3.1-147, xml2_1.3.2,pbkrtest_0.4-8.6, compiler_4.0.0,
 rstudioapi_0.11,curl_4.3, png_0.1-7, tibble_3.0.1,statmod_1.4.34,
 DescTools_0.99.36, stringi_1.4.6,forcats_0.5.0, gdtools_0.2.2,
 nloptr_1.2.2.1,vctrs_0.3.0, pillar_1.4.4, lifecycle_0.2.0,Rdpack_1.0.0,
 lmtest_0.9-37, estimability_1.3,gbRd_0.4-11, R6_2.4.1,
 latticeExtra_0.6-29,rio_0.5.16, codetools_0.2-16,
 boot_1.3-24,gtools_3.8.2, withr_2.2.0, nortest_1.0-4,mgcv_1.8-31,
 expm_0.999-4, parallel_4.0.0,hms_0.5.3, grid_4.0.0,
 rpart_4.1-15,coda_0.19-3, minqa_1.2.4, rmarkdown_2.2,numDeriv_2016.8-1.1,
 base64enc_0.1-3