Defensive symbiont genotype distributions are linked to parasitoid attack networks - Dataset and scripts

This R project includes data and scripts for paper - Defensive symbiont genotype distributions are linked to parasitoid attack networks

Script for analysis:

    Run all the scripts in order, to get all analyses and results in this paper. 

    Script_0_package_install_load.R: 
        Installs and loads the necessary R packages required for all subsequent scripts.
        This R project

    Script_1_Extract_from_Table_S2.R
        Extracts information from:
            Table_S2.csv (located in /Rawdata/)    Data_4_Aphid_sequences.fas
        Generates 10 files for further analyses:
            Aphid relatedness distance:
              Aphid_phylogenetic_relatedness_16species.csv
              Aphid_phylogenetic_relatedness_22species.csv
              Aphid_phylogenetic_relatedness_31species.csv
            Hamiltonella - Aphid Matrices:
              Hamiltonella_Aphid_matrix_16species.csv
              Hamiltonella_Aphid_matrix_22species.csv
              Hamiltonella_Aphid_matrix_31species.csv
            Parasitoid - Aphid Matrices:
              Para_Aphid_matrix_16species.csv
              Para_Aphid_matrix_22species.csv
            Plant - Aphid Matrices:
              Plant_Aphid_matrix_16species.csv
              Plant_Aphid_matrix_31species.csv

    Script_2_BarPlot_Fig1.R: 
        Generates Fig. 1 using: 
            Data_11_Parasitoid_genus_aphid_22_species_Fig.1.csv  Data_12_Plant_genus_aphid_31_species_Fig.1.csv
        The bar order in Fig. 1 is arranged by proportion, which is not directly supported in the ggplot2 package in R. Therefore, we need manually reordered if using the first plot code; or, manually colour it using the second plot code. 

    Script_3_MMRR_analysis.R
        Uses 10 distance/distribution matrices of Parasitoid, Plant, Hamiltonella, and Aphid relationships to compute matrix correlations with the MMRR (Multiple Matrix Regression with Randomization) model.

    Script_4_Species_linkage_Fig2_FigS4.R
        Uses 7 distribution matrices of Parasitoid, Plant, and Hamiltonella relationships with Aphids (excluding Aphid genetic distance matrices) to generate the species linkage diagrams for Fig. 2 and Fig. S4, For aesthetic purposes, unconnected sample points have been moved.

    Script_5_MMRR_Fig3_FigS5.R
        Plots the MMRR correlations using the 10 distance/distribution matrices for Parasitoid, Plant, Hamiltonella, and Aphid relationships (Fig. 3 & Fig. S5). 

    Script_6_parasitoid_specialization_Fig4.R
        Computes specialization levels using the H2 Index for:
            Parasitoid-Aphid
            Aphid-Hamiltonella
            Parasitoid-Hamiltonella relationships 
        Generates Figure 4.

    Script_7_Ecologicial_indices&plots_Table1_FigS3.R
        Uses 7 distribution matrices of Parasitoid, Plant, and Hamiltonella relationships with Aphids (excluding Aphid genetic distance matrices) to:
            Calculate ecological indices (Richness, Shannon Index, Simpson Index).
            Use linear models to analyze the relationships between Parasitoid/Plant-Aphid and Aphid-Hamiltonella communities.   
        Outputs results for Table 1 and Figure S3.

    Script_8_Bubble_plot_FigS2.R
        This script using 
            Hamiltonella_Aphid_matrix_31species.csv 
        and two phylogeny trees 
            Data_8_Aphid_species_phylogeny.txt & 
            Data_9_Hamiltonella_phylogeny.txt
        To create a Cophylogeny tree of Hamiltonella strains and aphid species that we identified in this study and previously known strains. (Fig. S2)

    Script_9_DADA2_pipeline.R
        This script is the DADA2 pipeline used to processing the raw COI sequencing data into ASV (Amplicon Sequence Variant) count files. After processing, the ASV files required manual curation to link each ASV with a distinct parasitoid and aphid species name based on BLAST results. In our updated version, we have provided both the raw data (BioProject PRJNA1139364) as well as the manually curated files (Data_1 and Data_2) which includes the ASV species names. All subsequent analyses can be reproduced using the curated datasets (Data_1 and Data_2), ensuring clarity and replicability for users.
        We have incorporated the SRA Toolkit to streamline the process. This allows users to download raw SRR sequencing files directly and convert them into FASTQ files, which can then be input into the DADA2 pipeline. The pipeline produces two ASV count files, one for each sequencing run.

Raw data and inofrmation:

    Table_S2.csv
        The Dataset from Supplementary Table S2: After manual pooling the Parasitoid/Aphid species based on the Data_1,2 and 6.
            No: the number of this sample
            City: The city that collected this sample
            Location: the location of sample collection
            Sample category: Aphid mummies or alive aphids
            Sample code: the name of sample (self defined)
            Collection year: when this sample was collected
            Aphid taxa / Wasp taxa / Plant species /Hamiltonella strain: The Aphid/Parasitoid/Plant/Hamiltonella species 
            Sequencing method: Illumina or Sanger sequencing   
            Sequencing date: the date of sending this sample to sequencing
            Data source: From this study or from Wu et al. 2022: Local adaptation to hosts and parasitoids shape Hamiltonella defensa genotypes across aphid species

    Data_1_Deep_sequencing_data_block_1.xlsx & Data_2_Deep_sequencing_data_block_2.xlsx 
        Sample: The sample name from Illumina sequencing, which can later be found on GenBank (BioProject PRJNA1139364).
        Aphid: The detected aphid species for each sample.
        Parasitoid: The detected parasitoid species.
        The remaining columns contain ASV (Amplicon Sequence Variant) data derived from high-throughput barcoding sequencing.
        Data1 and Data_2 are the manually curated versions of the BioProject PRJNA1139364 with added species names for each ASV. 

    Data_3_Parasitoid sequences.fas & 
    Data_4_Aphid sequences.fas & 
    Data_5_Hamiltonella sequences.fas,
        FASTA files for:
            Parasitoid species
            Aphid species
            Hamiltonella strains
        These sequences were used for phylogenetic reconstruction and subsequent analyses.

    Data_6_Parasitoid_Aphid_pooled_table.xlsx 
        Contains the OTU (Operational Taxonomic Unit) manual pooling results for Parasitoid and Aphid species based on 99% (4 base pair) sequence similarity:
            Parasitoid pooling together group & Aphid pooling together group: Original names from Illumina sequencing.
            Original name: Representative sequences for each group.
            Pooled species name: Final species names used in all analyses.
            Pooled sequences: Final representative sequences used in all analyses.
            The result of manual pooling see the Table_S2.csv

    Data_7_Parasitoid_species_phylogeny.txt & Data_8_Aphid_species_phylogeny.txt & Data_9_Hamiltonella_phylogeny.txt 
        Phylogenetic trees for:
            Parasitoid species
            Aphid species
            Hamiltonella strains
        These trees were generated using the PhyML tool on the ATGC Montpellier platform, original fasta file was Data_3, 4 & 5.

    Data_10_aphid_host_info.csv 
        Provides aphid host information for generating Figure 2 and Figure S4:
        Aphid: Names of aphid species included in this study.
        Host: Host categories:
            1: Herb aphids
            2: Grass aphids
            3: Tree aphids
        Host_category: Detailed descriptions of host categories.

    Data_11_Parasitoid_genus_aphid_22_species_Fig.1.csv & Data_12_Plant_genus_aphid_31_species_Fig.1.csv 
        Reduced matrices used for Figure 1, created by merging data from:
            OTUs associated with the same parasitoid species.
            Plant species belonging to the same genus.

    7 Distribution Matrices (from Script_1_Extract_from_Table_S2.R)
        Hamiltonella-Aphid Matrices:
           Hamiltonella_Aphid_matrix_16species.csv
            Hamiltonella_Aphid_matrix_22species.csv
            Hamiltonella_Aphid_matrix_31species.csv
        Parasitoid-Aphid Matrices:
            Para_Aphid_matrix_16species.csv
            Para_Aphid_matrix_22species.csv
        Plant-Aphid Matrices:
            Plant_Aphid_matrix_16species.csv
            Plant_Aphid_matrix_31species.csv
        Structure:
            Rows represent aphid species.
            Columns represent Hamiltonella strains, parasitoid species, or host plant species linked to each aphid species.
        Usage: These matrices were used to generate Figures 2, 3, 4, Figures S2, S3, S4, S5, and for MMRR tests, ecological indices, and H2 index calculations.

    3 Aphid phylogenetic relatedness matrices derived from Script_1_Extract_from_Table_S2.R
        Aphid_phylogenetic_relatedness_16species.csv
        Aphid_phylogenetic_relatedness_22species.csv
        Aphid_phylogenetic_relatedness_31species.csv
        These matrices provide phylogenetic distance information, showing pairwise genetic distances between the 31 aphid species included in this study.

    SraRunTable.csv
        The BioProject (PRJNA1139364) information downloaded directly from Genbank.