DOI: 10.5061/dryad.rjdfn2zdz
title: Swida amomum seed germination

description: Premise: Effective seed dispersal is essential in population dynamics of
 plant species. Swida amomum (Silky dogwood) exhibits a dispersal syndrome
 characteristic of autumn-ripening shrubs with fleshy fruits, where
 attached fruits are ingested and defecated by birds while fallen fruits
 are consumed by ground-foraging birds and mammals.  Methods: We
 documented that fallen fruits of this shrub were consumed by two aquatic
 turtle species (Eastern painted turtle, Chrysemys picta and Red-eared
 slider, Trachemys scripta) and that their seeds were defecated.
  We compared germination success (percentage of seeds germinated)
 of defecated seeds, seeds collected from pond surface, and seeds removed
 from shrubs. Results: While four seed taxa were identified in fecal
 samples, seeds of S. amomum were the most frequent (93%) among samples and
 the most numerous (106 seeds) in any sample.  Average proportion
 of fecal seeds germinated (85.99%) exceeded that of seeds from pond
 surface (82.76%) and from shrubs (60.24%), albeit the difference in
 germination success was insignificant. When analyzed using fecal samples
 from Painted turtles only, the difference in germination success between
 fecal seeds and those collected from pond or shrub became significant.
 Conclusions: Our findings represent the first report of S. amomum seeds
 being dispersed by turtle gut passage and suggest aquatic turtles could be
 an important part of a secondary seed dispersal process influencing woody
 plant community composition in temperate wetland ecosystems.
description: Study site: We conducted this study at a small
 (surface area 0.03 km2) freshwater impoundment (Sturtevant Pond; UTM
 Easting: 433438.26, UTM Northing: 46170104684.94, UTM zone: 24Z) of a
 semi-perennial stream located in Bridgewater, Massachusetts, USA.
 Pine-mixed eastern temperate deciduous forests and woodlands comprised the
 dominant vegetation of the upland habitats. The watershed is largely
 forested with a substantial extent of suburban and residential
 development.    Field sampling and seed
 collection­: We conducted this study for three
 consecutive years (2017–2019), from late August to early October in each
 year. This timeframe coincided with both the turtle activity, autumn bird
 migration, and fruit production in silky dogwood.  For capturing turtles,
 we targeted two species (Chrysemys picta, Eastern painted turtles, and
 Trachemys scripta, Red-eared sliders) and deployed a single hoop-net trap
 (1 m diam., 2.5 cm mesh; Memphis Net & Twine Co., Memphis,
 Tennessee, USA) baited with canned, oil-emersed sardines.  For a given
 year, we deployed the trap overnight for 24 hours, 1–2 times per week. We
 recovered the trap after each trap night to check for captured turtles.
 Our methods are standard field techniques for surveying turtles and have
 been employed for capturing a representative sample of freshwater turtles
 elsewhere (Dodd, 2016). We individually housed turtles
 in a 38 L aquarium filled with tap water to a depth of 2–3 cm for five
 consecutive days without feeding to collect fecal matter.  Afterward, we
 returned all animals to the site of capture.  For each turtle captured,
 seeds were manually isolated from fecal matter, rinsed, sorted, and
 counted.  Two sources of Swida amomum control seeds were collected from
 the study site in each year we trapped for turtles: 1) fruits floating on
 the pond surface (a random sample of 11–26 fruits) and 2) ripe fruits
 harvested from nearby shrubs (donor shrubs were 1–2 m apart, 11–15 fruits
 collected per plant, one fruit per inflorescence).  Prior to storage, all
 pulp was removed by agitation after soaking in water for 24–48 h.  In the
 first year of study (2017), control fruits were limited to only those
 floating on pond surface.  All seeds were surface sterilized with a 30 s
 0.05% hypochlorite solution wash, followed by two sterile water rinses,
 and stored in sterile water for 4 months at 5°C to break dormancy (Allen
 and Farmer, 1972).  To measure germination, we placed
 entire seed samples (1–64 seeds per fecal sample; 12–26 seeds per “pond”
 control; 12–15 seeds per “shrub” control) on moist filter paper, with
 individual seeds physically apart from other seeds, in petri dishes. 
 Dishes were positioned in a growth chamber (Percival CU30L2C8; Percival
 Scientific, Perry, Iowa, USA) under a 12h light/dark, 25/15°C cycle for 50
 consecutive days. Seeds were observed once every two days for germination
 and filter paper wetted as needed with sterile water. Seeds were removed
 from the dishes as they germinated. For each seed sample, both controls
 and fecal, percentage seed germination was calculated ((number
 germinated/total seeds in sample)*100). Statistical
 analyses: To test for significant differences in
 percent seed germination between fecal, pond, and shrub samples, we ran a
 one-way Analyses of Variance (1-ANOVA) permutational (20,000 permutations)
 test (Legendre and Legendre 2012). For post hoc pairwise comparisons, we
 performed unpaired, Yuen’s trimmed means robust test for unequal variance
 (Welch approximation), and applied Benjamini and Hochberg (1995) method to
 correct for false discovery rate and adjust probability estimations. The
 graphics were generated following Patil (2021). We
 opted for permutation tests since our sample sizes were low, non-normal,
 and unbalanced. In permutation tests, instead of comparing test statistics
 to a standard statistical distribution, the reference distribution is
 generated by randomly permuting the dataset under the study (Legendre and
 Legendre 2012). Likewise, the robust test we applied for pairwise
 comparisons is distribution independent and performs well despite
 heteroscedasticity and small sample sizes. Since just one fecal sample was
 collected from Trachemys scripta, we ran tests both including and
 excluding the fecal samples collected from this species. Statistical
 inferences were made at 95% confident intervals. We used R Statistical
 Programming Environment (R Core Team 2022) within RStudio integrated
 Developmental Environment for all the analyses (RStudio 2022.02.3 Build
 492).