Supplementary materials for "Evolution of immune genes in island birds: reduction in population sizes can explain island syndrome"

Here is a brief description of the files present in each folder:<br> <br> <strong>List of genes: </strong> <strong>List Immunity genes</strong>: Description and references for the immunity genes. <strong>list_Sma3s:</strong> List of genes identified by Sma3s program <strong>list_database:</strong> List of genes identified Immunome Knowledge Base, InnateDB and gene annotation. <strong>Polymorphism analysis :</strong> <strong>README: </strong>help to run seq_stat_coding <strong>seq_stat_coding</strong>: Executable C++ file and source file (.cpp) to estimate synonymous nucleotide genetic diversity (Ps) and non-synonymous nucleotide diversity (Pn) <strong>removeStopCodon:</strong> Executable C++ file and source file (.cpp) to remove the stop codon from the alignments <strong>Clean_Alignment : </strong>Executable C++ fileand source file (.cpp) to exclude the site with less than a define number of individuals <strong>TLR7_Pmajor_XP_015492521.1</strong>: Input file example <strong>Scripts and dataset : </strong> <strong>Simulations with SliM:</strong> <strong>SliM__immunity.slim</strong>: Script used to simulate sequences under balancing selection using codon format and exon-intron format <strong>Formating dataset:</strong> <strong>out_Seq_stat_*</strong>: Output of Seq_stat_coding according to our differents gene categories and selection pressure <strong>Formating_dataframe_script</strong>: Script to calculate Pn/Ps ratio and make a dataframe ready to plot. <strong>Plots and models :</strong> <strong>Dataframe_ready_to_plot.csv</strong>: Output from Formating_dataframe_script. Table containing : Species, PNPS, PN, PS, D_Taj, GC, S, category, Origin, family and selection regime. <strong>Plot_and_models_script</strong>: Script to plot results, and make differents models <strong>data_from_Leroy_etal_2021.csv : </strong>Informations and statistics about dataset from Leroy et al., 2021. <strong>Mitochondrial_phylogeny.treefile:</strong> Phylogeny based on mitochondrial genes of species from the dataset reconstructed by maximum likelihood method (IQTREE model GTR+Gamma and ultrafast bootstrap). <strong>Simulation analysis</strong> <strong>Tab_h_overdominance </strong>: Effect of parameters h (dominance coefficient) on Pn/Ps for sequences simulated under overdominace. <strong>Tab_m3_freq_dep </strong>: Effect of parameters S (selection coefficient) on Pn/Ps for sequences simulated under frequency dependent. <strong>Tab_Ne_overdominance </strong>: Effect of population size on Pn/Ps for sequences simulated under overdominance <strong>Tab_Ne_freq_dep </strong>: Effect of population size on Pn/Ps for sequences simulated under frequency dependent. <strong>Plot_simulated_results: </strong>Script to plot the effect of parameters on Pn/Ps from simulations. <strong>Supplementary table and figures :</strong> <strong>Figure S1 : </strong>Distribution of the percentage of contaminating contigs. The red line represents the 80% quantile. <strong>Figure S2: </strong>PCA of <em>Cyanistes species</em> <strong>Figure S3: </strong>PCA of <em>Cyanomitra </em>and <em>Turdus </em>species <strong>Figure S4: </strong>PCA of <em>Ploceus </em>species <strong>Figure S5: </strong>Fis ~Nucleotide diversity <strong>Figure S6: </strong>Correlation between Pn/Ps (a) and Ps (b) calculated on the control genes in this study's dataset and those calculated by Leroy et al. (2021). <strong>Figure S7 - Missing_data_ps_Phylloscopus.pdf</strong> : Relationship between the maximum number of missing individuals allowed and synonymous nucleotide diversity (Ps) <em>Phylloscopus trochilus</em> and <em>Fringilla coelebs</em>. <strong>Figure S8: </strong>Effect of sub-sampling size on PN/PS <strong>Figure S9: </strong>Pn/PS according to Ps for sub-sampling control, TLR and BD genes. <strong>Figure S10:</strong> Boxplot of Pn/Ps according to population size for simulated sequences under overdomiance via SLiM <strong>Figure S11:</strong> Boxplot of Pn/Ps according to population size for simulated sequences under frequency dependence via SLiM <strong>Figure S12:</strong> Boxplot of Pn/Ps according to a) initial selection coefficient of the mutation under frequency dependence b) dominance coefficient (h) for simulated sequences under overdominance via SLiM <strong>Table S1 - Model comparison using reduce number of families :</strong> Model selection of all genes categories using reduce number of families (we grouped Turdidae within Muscicapidae, Nectariniidae, and Estrildidae within Ploceidae and Fringillidae within Thraupidae). <strong>Table S2 - Lm &amp; PGLS on dPn/Ps </strong>: Alternative models (Lm for linear models and PGLS for Phylogenetic Generalized Least Squares) using the difference between Pn/Ps of immunity genes and control genes (Pn/Ps) as dependent variable, and species origin as explanatory variable. <strong>Table S3 - Samples &amp; sequencing information : </strong>Table with sampling and sequencing information regarding the samples newly-sequenced in this study, and those obtained from Leroy et al. 2021 <strong>Table S4 - Quality of sequences per individual : </strong>Table with sequencing quality information ( number of genes analysed, proportion of available positions, depth coverage) regarding the samples newly-sequenced in this study, and those obtained from Leroy et al. 2021 <strong>Table S5 to S14 : </strong>Model selection by AICc criterion and ANOVA test. Summary of the best models. <strong>Mitochondrial phylogeny:</strong> <strong>AllSp_ultrafastaboot.treefile:</strong> The species phylogeny was estimated using mitochondrial genes and a maximum likelihood method implemented in IQTREE (model GTR+Gamma and ultrafast bootstrap <strong>AllSp_mito.fst</strong> : Alignment of the mitochondrial genes in fasta format. <strong>Table and figure of the main text:</strong> <strong>Figure 1: </strong>Phylogeny based on mitochondrial genes of species from the dataset <strong>Figure 2 :</strong> Conceptual diagram showing the expected results <strong>Figure 3 : </strong>Boxplot of Pn/Ps according to species origin for different gene categories under purifying selection. <strong>Figure 4 :</strong> Boxplot of Pn/Ps according to species origin for different gene categories under purifying selection. <strong>Figure 5 : </strong>Effect of mutation type on Pn/Ps acording to Ne <strong>Figure 6 : </strong>Boxplot of Pn/Ps according to species origin (mainland in green and insular in orange) for different gene categories under balancing selection. <strong>Table 1: </strong>List of species and sampling localities, along with the type of data obtained and the number of individuals (N). <strong>Table 2 : </strong>Statistical model explaining Pn/Ps variation of Toll-Like Receptors, Beta-Defensins genes, and control genes. The p-values of ANOVA test between simpler models are not reported if a more complex model explains a larger proportion of the variance. <strong>Table 3 : </strong>Summary of the best statistical model selected using AICc explaining variation in Pn/Ps in control genes, Toll-Like receptors and Beta-Defensins genes under purifying selection with origin, gene category parameters.* indicates significances : * &lt; 0.05; ** &lt; 0.01; *** &lt; 0.001. <strong>Table 4 : </strong>Statistical model explaining Pn/Ps variation of genes under balancing selection (i.e MHC class I and II), and simulated sequences under frequency dependence.