DOI: 10.5061/dryad.wpzgmsbqr
title: Sustained mangrove reproduction despite major turnover in pollinator community composition at expanding range edge

description: Background and Aims How well plants reproduce near their geographic range
 edge can determine whether distributions will shift in response to
 changing climate. Reproduction at the range edge can be limiting if
 pollinator scarcity leads to pollen limitation, or if abiotic stressors
 affect allocation to reproduction. For many animal-pollinated plants with
 expanding ranges, the mechanisms by which they have overcome these
 barriers are poorly understood. Methods In this study, we examined
 plant-pollinator interactions hypothesized to impact reproduction of the
 black mangrove, Avicennia germinans, which is expanding northward in
 coastal Florida, USA. We monitored insects visiting A. germinans
 populations varying in proximity to the geographic range edge, measured
 the pollen loads of the most common insect taxa and pollen receipt by A.
 germinans stigmas, and quantified flower and propagule production. Key
 Results We found that despite an 84% decline in median floral visits by
 insects at northernmost vs. southernmost sites, range-edge pollen receipt
 remained high. Notably, local floral visitor assemblages exhibited
 substantial turnover along the study’s latitudinal gradient, with
 large-bodied bees and hover flies increasingly common at northern sites.
 We also observed elevated flower production in northern populations and
 higher per capita reproductive output at the range edge. Furthermore, mean
 propagule mass in northern populations was 18% larger than propagules from
 the southernmost populations. Conclusions These findings reveal no erosion
 of fecundity in A. germinans populations at range limits,
 allowing rapid expansion of mangrove cover in the region. These results
 also illustrate that substantial turnover in the assemblage of
 flower-visiting insects can occur at an expanding range edge without
 altering pollen receipt.
description: Data were collected over the course of three years at eleven
 sites containing <em>A. germinans</em> populations in eastern
 Florida. The southernmost sites (latitudes 27.1–27.9°N) contain mixed
 stands of all three Floridian mangrove species (with <em>Rhizophora
 mangle</em> L. [Rhizophoraceae] and <em>Laguncularia
 racemosa</em> [L.] C.F. Gaertn. [Combretaceae]), in varying stages
 of regeneration following impoundment for mosquito control in the first
 half of the 20th century. Farther north, in the southern reaches of the
 mangrove-marsh ecotone (latitudes 28.5–29.1°N), many sites are also
 recovering from impoundment or other human alteration, but here are
 characterized by mixed-mangrove stands edging waterways, with salt marsh
 vegetation (primarily <em>Distichlis spicata</em> [L.] Greene
 [Poaceae], <em>Spartina alterniflora</em> Loisel. [Poaceae],
 <em>Batis maritima</em> L. [Bataceae], and
 <em>Salicornia</em> sp. L. [Amaranthaceae]) landward. At the
 northern end of the study region (latitudes 29.6–29.9°N), sites contain
 some of Florida’s northernmost mangroves (Cavanaugh et al. 2019). Here,
 clusters of short (generally &lt;3m tall) mangroves – almost
 exclusively <em>A. germinans</em> – are embedded in a salt
 marsh matrix consisting primarily of <em>S. alterniflora</em>,
 <em>B. maritima</em>, and <em>Salicornia</em> sp.
 Most of these northern sites fall within the Guana-Tolomato-Matanzas
 National Estuarine Research Reserve (GTM). The identity
 and frequency of <em>A. germinans</em>’ floral visitors were
 assessed during the 2013, 2014, and 2015 flowering seasons. All 11 sites
 were monitored at least once over these three years, though only seven
 were monitored in any given year, with each site visited repeatedly within
 a flowering season. Across all sites in a given year, visitation data were
 collected over 105.5hrs, 67hrs, and 33.5hrs in 2013, 2014, and 2015,
 respectively. <em>A. germinans</em> flowers
 were monitored for floral visitors during 15-minute observation periods,
 which we conducted in fair weather from mid-morning to mid-day. Focal
 <em>A. germinans </em>were selected haphazardly; trees were at
 least 10m apart and contained at least 10 open flowers. We conducted a
 single observation period per day at each focal tree (i.e. no tree was
 monitored twice on the same date). During each observation period, the
 observer selected 4–10 open flowers in close proximity to one another and
 recorded the identity of each floral visitor and the number of focal
 flowers visited by each individual insect. Insects were typically
 identified to family, with the exception of the Apidae (Hymenoptera) which
 were identified to genus or species. During each observation period, the
 observer recorded the focal tree’s height (to the nearest 0.5m) and
 estimated the total number of open flowers on the tree.
 To approximate the relative importance of individual taxa as
 pollinators of <em>A. germinans</em>, we captured floral
 visitors and measured the size of their pollen loads. Using hand nets, we
 collected individual insects representing 11 of the 12 most frequently
 observed floral visitor taxa, directly from <em>A.
 germinans</em> flowers (one frequent taxon – Pompilidae – was not
 encountered during this time). We collected as close to 10 individuals per
 taxon as possible. We quickly immobilized the insects in coolers with ice
 packs and later transferred them to a freezer to kill them. They were
 subsequently pinned and inspected using a dissecting microscope. In the
 lab, each insect was swabbed with a ~2mm<sup>3</sup> cube of
 fuchsin jelly for up to 10 minutes to sample its pollen load. In the case
 of <em>Apis mellifera</em> L. (Hymenoptera:Apidae) and
 <em>Bombus</em> (Hymenoptera:Apidae), hydrated corbicular
 pollen was avoided, as it is thought to be unlikely to contribute to
 pollination (Thorp 2000). The fuchsin jelly was then transferred to a
 microscope slide, where the number of <em>Avicennia</em> and
 non-<em>Avicennia</em> pollen grains could be counted using a
 compound microscope at 100X and 400X magnification. To
 assess pollen deposition, we collected <em>A. germinans</em>
 stigmas from six sites in 2015. Each site was visited twice – once in the
 early flowering season and once in the mid-flowering season. During each
 site visit, up to 12 trees were haphazardly selected, and we collected up
 to eight flowers with mature stigmas from throughout the tree canopy.
 Mature stigmas could be identified by their spread lobes, which open on
 approximately day 3 of anthesis (personal observation); flowers are
 syncarpous, with a single stigma per flower. In total, we collected 768
 stigmas from all six sites. In the lab, each stigma was removed from its
 flower and mounted in fuchsin jelly on a microscope slide, which we
 inspected at 100X and 400X magnification on a compound microscope. Given
 the sometimes-large number of pollen grains present, we counted the number
 of <em>A. germinans</em> pollen grains three times on each
 stigma and used the average of these three counts for analysis.
 Flower production per tree was measured during each observation
 period for floral visitors, as described above. In late October-early
 November of 2014 and 2015, when propagules were maturing on parent trees,
 we established transects at multiple sites to measure the densities of
 reproductive <em>A. germinans</em> trees and fecundity
 (propagule production per tree). In each year, transects were established
 at six sites (though not the same set of sites in both years). At each
 site, we identified three areas of high <em>A. germinans
 </em>density and laid out one 20m transect in each area (oriented to
 maximize the number of trees intersected), resulting in three transects
 per site. We recorded the number of reproductive <em>A.
 germinans</em> over 0.5m tall that occurred within 1m of the
 transect tape, on either side. To measure propagule
 production per tree, or fecundity, we randomly selected five reproductive
 trees along each transect (or in close proximity to the transect, if fewer
 than five were immediately adjacent) and estimated the number of
 propagules present by counting the number on a representative portion of
 the tree and extrapolating to the entire canopy. Because different sites
 were in different stages of propagule drop, we then corrected these
 estimates to account for propagules that had already fallen. We did this
 using infructescences at each site that had been covered in mesh bags
 prior to the onset of propagule drop (three bags per tree on up to 30
 haphazardly selected <em>A. germinans</em> per site); these
 bags collected falling propagules and allowed us to calculate the fraction
 of total propagules that remained attached to the pedicel. By multiplying
 our original propagule counts by the inverse of this fraction, we
 estimated the total number of propagules originally produced by the
 trees. Fruit set rates, defined as the number of mature
 propagules produced per floral bud, were also measured using these same
 bags. Once each bag was collected, we counted the number of propagules
 (both abscised and still attached to the pedicel) as well as the total
 number of floral bud scars. Fruit set rate was calculated as the total
 number of propagules produced by a given inflorescence, divided by the
 number of floral bud scars. In 2015, propagules were
 collected from seven sites and weighed. To avoid bias resulting from
 earlier phenology at the northern sites, we collected only mature
 propagules that had dropped from their pedicels. To do this, we
 haphazardly selected 13–22 <em>A. germinans</em> at each site
 in late summer, once flowering had ended; we installed large-mesh (~1.0cm)
 bags over three inflorescences, each on a separate branch. In late
 October/early November, we returned to the sites, collected a total of 326
 mature propagules that had dropped from the pedicels into the bags, and
 measured their fresh weight to the nearest 0.001g.
description: Data files are CSVs and can be opened with any software
 compatible with this file type. The programming script is in R, originally
 written in R version 3.5.0.