DOI: 10.5061/dryad.34tmpg4q1
title: Data from: Dispersal and establishment traits provide a colonization advantage for a polyploid apomictic plant

description: Premise: Apomictic plants (reproducing asexually through seed) often have
 larger ranges and occur at higher latitudes than closely related sexuals,
 a pattern known as geographical parthenogenesis (GP). Explanations for GP
 include differences in colonizing ability due to reproductive assurance
 and direct/indirect effects of polyploidy (most apomicts are polyploid) on
 ecological tolerances. While life history traits associated with dispersal
 and establishment also contribute to the potential for range expansion,
 few studies compare these traits in related apomicts and
 sexuals.  Methods: We investigated differences in early life
 history traits between diploid-sexual and
 polyploid-apomictic Townsendia hookeri (Asteraceae),
 which displays a classic pattern of GP. Using lab and greenhouse
 experiments, we measured seed dispersal traits, germination success, and
 seedling size and survival in sexual and apomictic populations from across
 the range.  Key Results: While theory predicts that trade-offs
 between dispersal and establishment traits should be common, this was
 largely not the case in T. hookeri. Apomictic seeds had both
 lower terminal velocity (staying aloft longer when dropped) and higher
 germination success than sexual seeds. While there were no differences in
 seedling size between reproductive types, apomicts did, however, have
 slightly lower seedling survival than sexuals.  Conclusions:
 These differences in early life history traits, combined with reproductive
 assurance conferred by apomixis, suggest that apomicts achieve a greater
 range through advantages in their ability to both spread and
 establish. 
description: <strong>Study
 system</strong> <em>Townsendia</em>
 <em>hookeri</em> is a diminutive perennial member of the
 sunflower family (Asteraceae) with two forms: diploids that reproduce
 sexually and autopolyploids (mainly triploids) that reproduce via
 gametophytic apomixis (Beaman1957; Thompson and Whitton 2006). Sexual
 individuals are self-incompatible, while apomicts set seed autonomously,
 without the need for pollen to fertilize the endosperm.
 <em>Townsendia</em>
 <em>hookeri</em> produces single-seeded diaspores with
 persistent feathery pappus bristles (Strother 1993) usually associated
 with wind dispersal (Andersen 1992). Despite differing in ploidy and
 reproductive mode, the two forms are macro-morphologically
 indistinguishable in the field but can be distinguished by pollen traits.
 Polyploids produce larger viable pollen grains than diploids and have much
 lower pollen viability. To date, our lab group has characterized more than
 90 populations throughout the species’ range and found a tight association
 between low pollen viability and larger genome size (Thompson &
 Whitton 2006; Garani 2014; Lee 2015), supporting Beaman’s (1957)
 contention that diploids are sexual and polyploids are apomictic (Thompson
 & Whitton 2006; Thompson et al. 2008).
 Sexual populations have a much
 smaller range than the apomicts and primarily occur between Boulder, CO
 and Laramie, WY (Lee 2015). Apomictic populations extend from southern WY
 along the eastern side of the Rocky Mountains as far north as BC, Canada.
 In addition, a small number of diploid-sexual and polyploid-apomict populations occurs in a
 disjunct distribution in the Yukon territory (Thompson & Whitton
 2006; Garani 2014). <strong>Seed dispersal traits and terminal velocity
 measurements and analysis</strong>
 We compared diaspore traits of apomicts and sexuals
 using field-collected diaspores (technically cypselae, but from now on
 referred to as “seeds”) obtained between 2008 and 2013 from five diploid
 sexual and seven polyploidapomictic populations from
 across <em>T</em>.
 <em>hookeri</em>’s range, including northern populations in
 British Columbia and the Yukon territory (Appendix S1, Table
 S1; see the Supplementary
 Data with this article).  We selected two seeds from each of up to ten
 maternal plants per population, using only filled and darkly coloured
 seeds (traits that indicate viability; Garani 2014) with an intact pappus.
 In order to restore the pappus to a comparably open state (relative to the
 somewhat flattened state that resulted from storage in the collection
 envelopes), we placed seeds on wet filter paper in a sealed petri dish
 for ∼24 hours, then removed them and allowed them to
 air-dry for another 24 hours. At this point, the pappus had achieved a
 more regular form consistent with what is seen at the time of field
 collection. For each seed, we estimated the mass using an analytical
 balance, recorded the number of pappus bristles, and measured the length
 of two bristles from the center-apex using an ocular micrometer. We
 measured the angle of attack, the maximum angle across the open pappus
 bristles centered on their point of attachment to the seed proper, using a
 protractor (see Figure 2 in O’Connell and Eckert (2001) for a diagram
 depicting diaspore traits). We estimated terminal velocity by dropping
 individual seeds down a clear 120 cm
 long acrylic glass tube. Seeds were dropped by holding
 the pappus with tweezers and releasing. After conducting preliminary
 trials, we determined that the seeds would reach terminal velocity over
 the first 70 cm of the drop. We then recorded the bottom 50 cm of the fall
 with a video camera (shooting at 30 frames per second). We calculated
 the terminal velocity of each seed by dividing the drop time by drop
 distance (50 cm), taking great care throughout not to damage or disturb
 the structure of the pappus. We analyzed differences in seed traits (terminal
 velocity, angle of attack, bristle length, number of bristles, and seed
 mass) between reproductive modes using mixed effects models. Terminal
 velocity, angle of attack, and bristle length were analyzed using linear
 mixed effect models (LMMs), and number of bristles was analyzed used
 generalized linear mixed effects models (GLMMs) with a log link function
 for Poisson data. Reproductive mode was a fixed factor; population and mom
 (nested within population) were included as random factors. In order to
 test for differences in variance in terminal velocity between reproductive
 modes, we used Fligner-Killeen’s Test of Homogeneity of Variances
 (fligner.test() in R) which is robust to departures from
 normality.  We used linear models to test the effects of
 each diaspore trait on terminal velocity separately. Traits were centered
 and scaled using the scale() function in R with default options, which
 subtracts the mean from each value and divides by the standard
 deviation.  We explored the relationship between diaspore traits and
 terminal velocity using simple models that included each trait as a fixed
 effect (without the random effects of population or
 mom).  <strong>Germination in the
 lab</strong> For the germination and greenhouse experiments,
 we collected seeds from six diploid sexual populations and six polyploid
 apomict populations in the spring of 2013 and 2014 (Appendix S1, Table
 S1). Seeds were stored at room temperature in paper coin envelopes. The
 ploidy of each population used in this study was assessed previously by
 flow cytometry (Lee 2015).  We chose 11 maternal plants (hereafter known as
 “moms”) from each population, and 32 dark, filled seeds from each mom.
 We assessed germination in agar-filled petri dishes (hereafter referred to
 as “plates”). Each plate was divided into four quadrants, with four seeds
 from a single mom randomly assigned to each quadrant. Within quadrants,
 seeds were spaced to avoid contact. Due to the large number of seeds, we
 filled the plates over the course of three days (beginning on June 18,
 2014). We stacked the plates on the lab bench at room temperature
 (21°C) away from direct
 sunlight and re-ordered the stacks every 3–4 days. We checked the plates
 every 1–2 days until no new seeds had germinated for 10 days, and scored
 germination as successful based on the emergence of both a radicle and a
 pair of cotyledons. <strong>Seedling
 traits</strong> We transplanted seeds from agar into racks of
 cone-tainer pots (Proptek - Watsonville, California) 2–3 days after
 germination over a period of two weeks. We filled the pots with a
 well-draining soil mixture (4 potting soil: 2 sand: 1 perlite) and placed
 the seedling racks in growth chambers (Conviron, various models -
 Winnipeg, Canada). Growth chambers were set for 12h light
 (20°C) / 12h dark (10°C) cycles. We hand-watered seedlings daily in
 the week following transplanting and every two days after that. We rotated
 the racks between chambers once per week to account for potential
 differences among growth chamber models. After most seedlings reached a
 height of at least 2 cm (early July 2013), we hardened the seedlings in a
 greenhouse at the University of British Columbia Farm (Vancouver, Canada).
 The greenhouse environment was regulated by Argus Controls (Surrey,
 Canada) (set to 12h light at 20°C / 12h dark at 12°C), and the seedlings were bottom-watered on
 flood tables. Because the seedlings grown here were subsequently
 transplanted in a common garden experiment (the results of which are in
 Hersh 2020), we moved the plants outside to harden them to UV light (3–5
 hours per day). After two weeks of hardening, we took our final
 measurements, censusing all plants for survival and estimating plant size
 using total number of leaves, and the length of one fully expanded leaf
 per seedling. Thus, our measurement of seedling size reflects the size
 achieved from the day of germination until the census approximately two
 months later. While not every seed germinated on the same day (and
 therefore did not have equal time to grow until the census), 75% of seeds
 that germinated did so within a five-day period, so we expect the
 variation in total growth time had minimal effects on our growth
 estimates. 
 <strong>Analysis of
 germination and seedling traits</strong>
 We used GLMMs to analyze
 differences in germination success (logit link - binomial) and germination
 speed (log link - poisson, due to speed being rep- resented by day counts)
 between reproductive modes. Reproductive mode was a fixedfactor; plate,
 population, and mom (nested within population) were included as random
 factors. Similar to terminal velocity above, we tested for differences in
 variance in germination success and germination speed using
 Fligner-Killeen’s Test. We used GLMMs to analyze differences between
 reproductive modes for seedling survival (logit link - binomial) and leaf
 number (log link - poisson), and LMMs for leaf length. Reproductive mode
 was a fixed factor; rack, population, and mom (nested within population)
 were included as random factors. Including rack as a random effect
 resulted in model singularities for the leaf length and leaf number
 models, so we removed it from those analyses. Because populations were the
 main level of replication, we used subsequent mixed models to determine
 whether there were differences among populations, with population as a
 fixed effect and the same random factors as above. 
 We performed the mixed-effect
 analyses using the lmer and glmer functions implemented
 in the lme4 package (Bates et al. 2015) and fixed effect
 analyses using the lm function in R version 3.6.1 (R Core team 2019). We
 evaluated the strength of effects by using a combination of likelihood
 ratio tests (LRTs; comparing fully fitted model to a model with the tested
 term removed) and model predictions using
 the <em>ggeffects</em> package (Lüdecke 2018). We considered that we had strong evidence for
 differences between groups when both the LRT P-values were below 0.05
 and confidence intervals from model predictions were largely
 non-overlapping. In cases where only one of these criteria was met (i.e.,
 LRT P-value below 0.05 but substantial overlap in predicted confidence
 intervals), we considered that the evidence for differences was
 moderate.