DOI: 10.5061/dryad.1ns1rn8x1
title: Impaired immune function accompanies social evolution in spiders

description: An efficient immune system is essential to the survival of many animals.
 Sociality increases risk of pathogen transmission, which should select for
 enhanced immune function. However, two hypotheses instead predict a
 weakened immune function: relaxed selection caused by social
 immunity/protection, and reduced efficacy of selection due to inbreeding,
 reproductive skew, and female bias in social species that reduces
 effective population size and accelerate genetic drift. We assessed the
 effect of social evolution on immune function in a comparative study of
 two social spider species and their closely related subsocial sister
 species (genus Stegodyphus). The haemolymph of social species was less
 efficient in inhibiting the growth of potentially pathogenic bacteria than
 that of subsocial species. Reduced efficacy of selection in social species
 was supported by comparative genomic analysis of immune genes showing
 substantially elevated non-synonymous substitutions in one of the social
 species. We propose that this impaired immune function is likely to be a
 result of reduced efficacy of selection because the evolution of sociality
 in spiders is accompanied by demographic processes that elevate genetic
 drift. There may also be positive feedback between pathogen-induced local
 extinctions and the resulting elevation of genetic drift which further
 weakens responses to selection by pathogens.
description: Two assays were conducted, one
 with the gram-positive <em>Bacillus</em>
 <em>subtilis</em> DSM10 and one with the gram-negative Escherichia
 coli OP50, both in NB medium (Scharlau, Spain). Samples of
 haemolymph were obtained
 from fully hydrated, sterilised and anaesthetized female spiders by piercing a book lung and
 collecting 2 μl from the emerging droplet. The extracted
 haemolymph was then randomly distributed between 100 μl samples of bacterial medium in 96-well
 plates. Plates were
 incubated at 27°C except during measurements at room temperature
 (~20°C). Each plate contained two treatment controls;
 one sample of NB medium without haemolymph and one sample of NB medium
 with Ringer solution, the latter to mimic the haemolymph salt composition.
 Sample sizes for inhibition assays were: <em>B</em>.
 <em>subtilis</em>:
 n<sub><em>africanus</em></sub> = 37,
 n<sub><em>mimosarum</em></sub> = 51,
 n<sub><em>tentoriicola</em></sub> = 14,
 n<sub><em>dumicola</em></sub> = 28,
 n<sub><em>NB</em></sub> = 27,
 n<sub><em>NB+Ringer</em></sub> = 27, and
 <em>E</em>. <em>coli</em>:
 n<sub><em>africanus</em></sub> = 35,
 n<em><sub>mimosarum</sub></em> = 52,
 n<sub><em>tentoriicola</em></sub> = 14,
 n<sub><em>dumicola</em></sub> = 24,
 n<sub><em>NB</em></sub> = 24,
 n<sub><em>NB+Ringer</em></sub> = 24. Before the
 addition of haemolymph, bacterial concentration in each well was measured
 as light absorbance using an Epoch microplate spectrophotometer to
 standardize start concentrations. After addition of haemolymph the light
 absorbance was measured every 2nd minute from 0 to 16 min, every
 5<sup>th</sup> minute from 20 to 50 min, and then at 60, 70,
 80, 100 and 120 min. To investigate if selection at the molecular
 level differs between social and subsocial species, we used 23 immune
 genes annotated in the <em>S</em>.
 <em>mimosarum</em> genome. Alignments consisting of ortholog sequences
 from <em>S</em>. <em>mimosarum</em>,
 <em>S.</em> <em>africanus</em> and
 <em>S</em>. <em>lineatus</em> were obtained from
 Bechsgaard et al. <em>Stegodyphus</em>
 <em>dumicola</em> sequences were extracted from the genome
 assembly published by Liu et al. and <em>S</em>.
 <em>tentoriicola</em> sequences were extracted from an
 unpublished genome assembly. We used PAML 4.6 to estimate dN/dS ratios for each branch
 separately, first for individual loci, and secondly for all loci
 concatenated.