DOI: 10.6086/d1m97d
title: Data from: Correlated evolution between orb weaver glue droplets and supporting fibers maintains their distinct biomechanical roles in adhesion

description: Orb weaving spiders employ a “silken toolkit” to accomplish a range of
 tasks, including retaining prey that strike their webs. This is
 accomplished by a viscous capture thread spiral thread that features tiny
 glue droplets, supported by a pair of elastic flagelliform fibers. Each
 droplet contains a glycoprotein core responsible for adhesion. However,
 prey retention relies on the integrated performance of multiple glue
 droplets and their supporting fibers, with previous studies demonstrating
 that a suspension bridge forms, whose biomechanics sum the adhesive forces
 of multiple droplets while dissipating the energy of the struggling
 insect. While the interdependence of the droplet’s glycoprotein and
 flagelliform fibers for functional adhesion is acknowledged, there has
 been no direct test of this hypothesized linkage between the material
 properties of each component. Spider mass, which differs greatly across
 orb weaving species, also has the potential to affect flagelliform fiber
 and glycoprotein material properties. Previous studies have linked spider
 mass to capture thread performance but have not examined the relationship
 between spider mass and thread material properties. We extend earlier
 studies to examine these relationships in 16 orb weaving species using
 phylogenetic generalized least squares. This analysis revealed that
 glycoprotein stiffness (elastic modulus) was correlated with flagelliform
 fiber stiffness, and that spider mass was related to the glycoprotein
 volume, flagelliform fiber cross-sectional area and droplets per unit
 thread length. By shaping the elastic moduli of glycoprotein adhesive and
 flagelliform fibers, natural selection has maintained the biomechanical
 integration of this adhesive system.
description: Thread measurements:  Web samples were
 collected from fresh orb webs in nature. A metal frame with double sided
 tape (3M #9086K29550360) on its rim was pressed from behind a web,
 ensuring that a web’s native tensions were preserved. Once adhered to the
 taped rim, the remaining web was separated by hand along the outer edge of
 the frame, isolating the sample. Samples were brought back to lab as soon
 as possible for testing. To prepare individual droplets for testing, we
 collected a thread on carbon tape covered forceps to ensure natural thread
 tension was maintained. After contacting a thread strand with the forceps,
 we cut the connecting threads with a pair of iris scissors. This sample
 then spanned the 4.8mm space between the supports of a microscope slide
 sampler (Opell et al., 2011). We ensured that these threads were
 perpendicular to the supports, guaranteeing consistency in the length of
 the tested thread and the angle of droplet extension. To ensure that only
 a single droplet contacted our probe, we isolated the central droplet of
 the suspended strand. Droplets on either side were slid away from the
 central droplet using a wooden applicator stick that was whittled to a
 fine tip, exposing a few xylem fibers. Droplets were tested
 in a sealed chamber to maintain humidity at 55% relative humidity. With
 the desired humidity achieved, droplets were extended using a probe.
 Before each test, the 413µm tip of this polished steel probe was cleaned
 with 100% ethanol on a Kimwipe® or Whatman® filter paper. After inserting
 the probe into a port in the side of the chamber, the probe was locked
 into a support resting beside the microscope to prevent its movement
 (Opell et al., 2018). The isolated droplet was then brought into contact
 with the probe tip using the microscope’s mechanical stage. The adhered
 droplet was extended until it pulled off from the thread. Extending these
 droplets provided the angle of extension and stress strain curves, thus
 providing the elastic modulus and toughness. Seperate droplets were
 flattened with a device the dropped a coverslip onto the threads, allowing
 us to measure droplet volume and glycoprotein from surface area
 measurements.  Flagelliform fiber
 properties of A. pegnia, A. argentata, M. sagitatta, and N. oaxacensis,
 and T. elongata capture threads were newly measured. Additional threads
 from each individual’s web were secured to cardboard samplers and sent to
 the American Museum of Natural History for characterization of their
 elastic modulus and toughness using a Nano Bionix instrument. Similar
 methodology and instrumentation was used by Sensenig et al., 2010 to
 determine these properties for eleven of the study species present in the
 literature. The flagelliform fiber features of C. conica were used for the
 similarly sized species C. turbinata included in this study (Sensenig et
 al., 2010). PGLS:
 We
 used phylogenetic generalized-least squares (PGLS) to examine
 relationships among the capture thread material properties under Brownian
 Motion, with Pagel’s lambda to detect phylogenetic signal (Pagel, 1999).
 The phylogeny used in this analysis is based on a time calibrated tree
 produced from BEAST (Dimitrov et al., 2017). Details on how this tree was
 edited can be found in glycoPGLS.R. Having a complete 16 species tree, we
 used PGLS to examine relationships among traits and plot phylomorphospace
 plots, carried out using the ape, caper, geiger, and phytools packages in
 R (Revell, 2012; Pennell et al., 2014; Orme et al., 2018; Paradis
 & Schliep, 2019; R Core Team, 2019). Sensitivity
 analyses: Our tree editing
 introduced considerable uncertainty in our results, as the placement of
 additional taxa stems from taxonomic knowledge instead of phylogenetic.
 Four species: A. pegnia, N. crucifera, N. oaxacensis, and M. sagitatta
 were added in this manner. The placement of A. aurantia and V. arenata
 were based on other phylogenies, but these were derived from different
 datasets that feature different sampling of spider diversity (Garrison et
 al., 2016; Scharff et al., 2019). What may compound this uncertainty is
 the varying nodal support across the branches of our input tree, with
 broad confidence intervals around the divergence times among these
 relationships as well. Therefore, it’s necessary to examine alternate
 topologies and branch lengths to assess the influence of constructing our
 16 species phylogeny in its present form. To assess the impact of adding
 species to our phylogeny, we created seven alternate phylogenies. Six of
 these alternate phylogenies featured a random placement of a single “new”
 species (A. pegnia, A. aurantia, N. crucifera, N. oaxacensis, M. sagitatta
 or V. arenata). The randomized trees were the basis of our sensitivity
 analysis (prepped in glycoPGLS.R). The actual sensitivty analyses (using
 SensiPhy package in R) were carried out in Sensitivity.R.