Effects of Fire and Pollinator Visitation on the Reproductive Success of Asimina Reticulata (Annonaceae), the Netted Pawpaw

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Effects of Fire and Pollinator Visitation on the Reproductive Success of Asimina reticulata (Annonaceae), the Netted Pawpaw

Louise K. Barton1 and Eric S. Menges2 1Department of Ecology and Evolution, University of Colorado, Boulder, Colorado 80309 2Plant Ecology Program, Archbold Biological Station, Venus, Florida 33960

ABSTRACT The netted pawpaw (Annonaceae, Asimina reticulata) is a widespread Florida endemic plant that produces relatively few fruits. To understand its distribution and reproductive success, we posed three questions: (a) how do time-since-fire (TSF) and pollinator visitation influence reproductive success, (b) is it pollinator dependent, and (c) does outcrossing increase reproductive success? In 2016, we studied 157 plants in three flatwoods burn units (last burned, 2015, 2012, or 2001) and one grazed pasture at Archbold Biological Station in south-central Florida. Plants in pasture and recently burned flatwoods exhibited greater flower, fruit, and seed output, suggesting that habitat openness may promote competitive release or increased pollinator visitation. Pollinator visitation was greatest on plants within the 1-yr TSF, and the hairy flower scarab (Trichiotinus viridans) was the most prevalent visitor across all sites. Autogamously crossed flowers produced no fruit, confirming that seed production is pollinator dependent. Although fruit retention (10.0%) and seed germination (27.8%) were low after geitonogamous crossing, close and far outcrossing increased retention (30.8% and 39.6%) and germination success (41.6% and 46.4%). This research suggests that A. reticulata reaches its maximum reproductive potential in open areas after outcrossing events, and that low fruit set may be due to the lack of an effective pollinator, resulting in inbreeding depression. Additionally, an investigation into raccoons (Procyon lotor) as fruit consumers and potential seed dispersers suggests that seeds may benefit from gut scarification. Key words: Annonaceae, endemic, geitonogamy, pollination ecology, prescribed fire.

INTRODUCTION The southeastern Unit- beetle pollinator of the ancient Asimina flowers ed States is a global biodiversity hotspot (Noss may be long extinct, making the flowers of today et al. 2015), supporting enormous plant diversity a Pleistocene anachronism (Barlow 2002). and a vast array of regional endemics (Luna, Asimina reticulata most frequently establish- 2014). Within the genus Asimina, the only es in poorly drained flatwoods or disturbed temperate genus within the Annonaceae, seven habitats (Kral 1960, Abrahamson et al. 1984). A of the eight species are endemic to the southeast long-lived deciduous shrub, A. reticulata pro- and are primarily found in Florida (Kral 1960, duces an early spring display of showy, white, Wunderlin and Hansen 2011). One such species nodding flowers that emit a species-specific is Asimina reticulata Shuttlw. ex Chapm, which honey-like odor (Goodrich and Raguso 2009). is widespread throughout peninsular Florida and Flowers are typically 5–8 cm broad, peak in southern Georgia. Asimina reticulata exhibits abundance between February and April, and are low fruit set, possibly because of inbreeding produced before leaf emergence (Cox 1998, Kral 1960). The flowers are protogynous, with car- depression resulting from pollinator limitation, pellate-phase flowers developing into staminate- similar to its rarer congener, Asimina obovata phase flowers once the stigma is no longer (Willd.) Nash (Levitt et al. 2013). The effective receptive (Cox 1998). At a given time, stems will DOI: 10.2179/18-170 support flowers that are in a variety of stages: 323 324 CASTANEA VOL.83 bud, carpellate-phase, staminate-phase, senes- (AI) and show high levels of outcrossing (Pang cent, and fruiting. Fruits are berries that consist and Saunders 2014). A prior hand-pollination of several large (4–7-cm-long), clustered mono- study on A. reticulata determined incompatibil- carps, each of which contain two rows of 1–8 ity through AI, but not through geitonogamy, and seeds 1–2 cm long (Kral 1960). Seed dispersal found greater fruit set after cross-pollinations and germination success of this plant have not, than after geitonogamy (Cox 1998). Beyond fruit to our knowledge, been previously investigated. set, our study tracked potential inbreeding Along with the highest diversity of Asimina depression from hand-pollination treatments species, Florida supports numerous naturally through germination. Additionally, we incorpo- pyrogenic ecosystems and thus a variety of fire- rated two outcrossing treatments, close and far adapted species (Weekley and Menges 2003, crosses, to gain insight into the influence of Slapcinsky et al. 2010). After fire, many of these distance between parent plants and to assess species will resprout from belowground roots or potential for outbreeding depression. We also rhizomes (Menges and Kohfeldt 1995, Maguire used wildlife cameras to detect fruit predation and Menges 2011). Numerous studies in Florida and seed dispersal. This closer examination into have also shown that fire can also promote the reproductive biology of A. reticulata may seedling recruitment (Hartnett and Richardson provide valuable insight into why fruit set across 1989, Menges and Quintana-Ascencio 2004) and this species is notoriously low. stimulate flowering (Hartnett and Richardson METHODS 1989, McConnell and Menges 2002). Of the Study Sites Asimina species, Asimina longifolia Kral and We conducted this study in 2016 at Archbold Asimina pygmaea (W. Bartr.) Dunal are the Biological Station (ABS), a 2,101-ha preserve most fire dependent and will seldom flower in its located at the southern tip of the Lake Wales absence (Kral 1960). Postfire habitat conditions, Ridge in Highlands County, south-central Flor- including the addition of ash as a soil nutrient ida, (2781005000 N, 818210000 W). Known as a relic (Dean et al. 2015) and increases in light Pleistocene habitat, the Lake Wales Ridge and availability after the removal of high/dense associated Florida scrub support an array of vegetation (Abrahamson et al. 1984, Huffman Florida endemic plant species (Christman and and Werner 2000), may interact with effects of Judd 1990). Comprising naturally pyrogenic top removal from fire to create surges in flower ecosystems, the property is divided into pre- and fruit output. Although not fire dependent, it scribed burn units and is managed with varying is likely that A. reticulata is fire adapted and will ﬁre regimes (Menges et al. 2017b). show similar surges in reproductive output in To examine the effects of TSF on A. retic- more recently burned plots. ulata, we randomly selected three flatwoods In this study, the effects of time-since-fire sites with differing fire histories in which A. (TSF) on flower density, pollinator visitation, reticulata was present. These sites were domi- and the reproductive success of A. reticulata nated by either wiregrass (Aristida stricta were investigated within pine flatwoods with Michx.) or palmettos (Sabal etonia Swingle ex differing prescribed fire-return intervals. Plants Nash and Serenoa repens W. Bartram) and were within a grazed pasture habitat were also burned in 2015, 2012, or 2001 (1-yr, 4-yr, and 15- sampled to assess the influence of openness in yr TSF). Comprising 25% of the ABS area, the absence of fire. Although previous studies flatwoods are characterized by flat terrain with have concluded that A. reticulata responds to poorly drained soils and a scattered overstory of fire or mowing with rapid vegetative growth, Florida slash pines (Pinus elliottii Engelm. var. more long-term reproductive responses have not densa Little & Dorman) (Abrahamson et al. yet been examined (Kral 1960), and fire effects 1984). In addition to the flatwoods sites, we also on seed viability are largely unknown. selected a grazed pasture site within the Arch- To expand existing knowledge on the breeding bold Reserve to assess the influence of openness ecology of A. reticulata andtodetermine in the absence of fire. Although these pasture potential interactions between TSF and repro- sites are fire managed (last burned in 2010), duction, we also implemented a series of hand- actively grazed regions generally burn patchily, pollination treatments. Because of strong proto- suggesting that fire history within this site was gyny, flowers of Annonaceae avoid autogamy less relevant. In February 2016, we mapped and 2018 BARTON, MENGES: REPRODUCTIVE SUCCESS OF ASMINIA RETICULATA 325

Figure 1. (A) Flowering stems of Asimina reticulata Shuttlew. ex Chapm. with buds, carpellate-phase flowers, and staminate-phase flowers. (B) Pollination by a hairy flower scarab beetle (Trichiotinus rufobrunneus). (C) A cluster of fruit produced by a single flower. tagged reproductive individuals within the 1-yr to approximate the number of flowers each plant (n ¼ 52 plants), 4-yr (n ¼ 44), and 15 yr (n ¼ 35) produced throughout the season. postfire and grazed pasture (n ¼ 26) sites, for a Pollination surveys began in late March, when total of 157 study plants. In mid-June, when fruit flowering was at its peak. Initial pollinator began ripening, we counted the number of stems observations yielded no data; therefore, we and measured the height of the tallest branch, manually checked flowers by prying open outer canopy length from the longest horizontal petals to observe pollinators within (Norman and measurement, and canopy width from the widest Clayton 1986, Cox 1998). To survey all potential point perpendicular to the length. We estimated pollinators, flowers present on each study plant volume as the product of those three measures. were manually checked in either the morning (n The influence of TSF on flower and fruit output ¼ 6 d observation between 7:00 am and noon), was analyzed through a series of 1-way analyses afternoon (n¼12 between 1:00 PM and 6:00 PM), or of variance (ANOVAs), and pairwise differences evening (n¼3 between 7:00 PM and 11:00 PM) for a evaluated with a Tukey’s test. total of 21 d of observation. The order in which sites were visited was randomized, and exact Flower and Pollinator Surveys time of day was recorded for each pollinator To assess the influence of TSF on flower output observation. One specimen of each potential and pollinator visitation, we completed flower pollinator was collected, identified, and stored in and pollinator surveys on all study plants. the ABS flower-visitor collection. A visitor was Flower surveys began in mid-March, once plants deemed a potential pollinator if pollen covered had entered their budding stage and continued its body and if it was found in both staminate- every 5 d until early June, when all flowers had phase and carpellate-phase flowers. senesced. During that time, we recorded the presence of buds along with number of carpel- Hand-Pollination Experiment late-phase flowers, staminate-phase flowers, and To assess the breeding system of A. reticulata flowering stems for each plant (Figure 1). and the potential for inbreeding depression, we Staminate-phase flowers typically persist for 3– also conducted hand-pollination experiments. 6 d (Cox 1998); therefore, we used the total On each study plant, at least one unopened bud number of staminate flowers counted every 5 d was enclosed with a 25-cm by 30-cm, Delnet, 326 CASTANEA VOL.83

heat-sealed, nonwoven, pollinator-exclusion bag motion-sensor field cameras were trained to (Applied Extrusion Technologies, Inc., Middle- fruiting A. reticulata plants outside of the study ton, Delaware). Once stigmas were glistening populations. Camera traps were placed 2 m from and, therefore, receptive, we applied one of four ripe fruit and checked every 3 d during a 9-d randomly selected hand-pollination treatments span. to one flower on each plant and replaced the bag until the stigma had abscised. Because of Seed Germination protogyny, it was not necessary to emasculate Fruit from open-pollinated and hand-treated the flower (Cox 1998). Peroxtesmo esterase flowers were checked every 5 d and harvested indicator paper (Macherey-Nagel, Bathlehem, once fruit had become tender and ripe. We Pennsylvania) was used to confirm stigma measured fruit length and width, recorded the receptivity (Dafni and Maues´ 1998). number of seeds, and immediately sowed all We used four hand-pollination treatments. seeds into mini-petri dishes (35 mm by 20 mm). Autogamously treated flowers (n ¼ 46 flowers) Half of all collected seeds were scarified with a received no pollen and remained enclosed in razor before sowing. Open-pollinated seeds were pollinator exclusion bags to assess self-compat- collected from 90 fruits for a total of 308 seeds ibility (SI). Geitonogamously treated flowers (n (1-yr TSF ¼ 148 seeds, 4-yr TSF ¼ 56 seeds, 15-yr ¼ 34) received pollen from a different flower on TSF ¼ 7 seeds, and the pasture ¼ 97 seeds). the same plant, close outcross (n ¼ 39) from its Hand-pollinated seeds were collected from 44 closest flowering neighbor, and far outcross (n ¼ fruits for a total of 164 seeds (geitonogamy ¼ 18 38) from a study plant at least 50 m distant. We seeds, close outcross ¼ 77 seeds, and far collected pollen directly before application and outcross ¼ 69 seeds). All scarified seeds were applied it to receptive stigmas using a clean sown with the scarified side touching the sand. paintbrush. All other flowers on study plants Dishes were placed into terraria constructed were available for open pollination. from aluminum pans with perforated plastic lids. Indices of AI and SI were calculated using the Seeds were stored on an east-facing veranda at ABS, watered daily with deionized water, and following formulas: AI ¼ Sa/Ss and SI ¼ Ss/So, monitored daily for germination. To ensure equal where Sa, Ss, and So are the proportions of fruits resulting from autogamous, self, and outcross light and wind exposure, we rotated seed pollinators, respectively (Ramirez and Brito terraria regularly. One month passed after the 1990). If the AI was below the threshold of 0.3, last germination event before we concluded this then the species was nonautogamous. Similarly, study. if SI was below the 0.3 threshold, then the Field observations of A. reticulata whose seed species was not self-compatible. coats had been scraped or punctured by animals suggested that scarification might be an impor- Fruit Surveys tant component in germination. To examine We conducted fruit surveys every 5 d through- potential effects of scarification, we conducted out the fruiting season (late March to early an imbibition trial on 21 seeds. Initial seed July) until all fruit had either ripened or weights were recorded after drying for 12 hr; abscised. During surveys, we recorded the then, a small portion of the seed coat was number of flowers producing fruit, fruit per removed from one side of the seed by sanding or flower, and total fruit for each plant. As fruit scraping with a razor blade. Once treated, we approached ripeness, we enclosed fruiting submerged seeds in deionized water and branches in wire mesh cages to prevent weighed them after 3, 12, 18, and 24 hr of predation. We designed cages to fully enclose soaking. each branch and fastened fabric bags to the base of each cage to exclude small animals and RESULTS to capture aborted fruit. Fruit collection began Fire Management 12Juneandextendedto5July,whenthelast Although plant volume (mean – SE) did not fruit had ripened. differ significantly among sites (1-yr TSF, 0.13 – 0.02; 4-yr TSF, 0.19 – 0.04; 15-yr TSF, 0.25 – 0.08; Seed Dispersal and pasture, 0.22 – 0.04), mean stem numbers To detect potential seed dispersers and/or fruit within the 1-yr TSF and the pasture site were predators, two Bushnell (Overland Park, Kansas) significantly greater than those within the 15-yr 2018 BARTON, MENGES: REPRODUCTIVE SUCCESS OF ASMINIA RETICULATA 327

Figure 2. Time-since-fire effects on the production of flowers (A) and stems (B) in Asimina reticulata. Flower production is represented by the total flowers produced throughout the growing season and the maximum flowers present at a given time. Stem production is represented by total stem count and the maximum number of stems with flowers at given time. Results from 1-way analysis of variance models. Graphs depict means – SE, with lettering denoting significant differences among sites (Tukey’s honestly significant difference, p < 0.05).

TSF (1-way ANOVA, p ¼ 0.018 and p ¼ 0.028, TSF, 4.1 – 0.3; 4-yr TSF, 4.1 – 0.5; 15-yr TSF, 2.5 respectively). Study site also significantly pre- – 0.7; and pasture, 4.2 – 0.4). dicted total flower output, showiness (maximum flowers present on a plant at one time), and Pollinators maximum number of flowering stems (Figure 2). The most abundant pollinator in this study was Plants within 1-yr TSF or pastured sites pro- the hairy flower scarab, Trichiotinus rufobrun- duced significantly more flowers throughout the neus Casey (Scarabaeidae), which was recorded in 78 flowers across all sites (Table 1). Beetles season and had showier maximum floral displays were found in both staminate-phase and carpel- than did those within the 4-yr or 15-yr TSF sites. late-phase flowers. Other common visitors in- Differences in flowering between 1-yr TSF and cluded the love bug, Plecia nearctica Hardy pastured sites were not significant. (Pleciidae), and the small yellow cockroach, Similarly, differences in fruiting between 1-yr Cariblatta lutea Saussure & Zehntner (Ectobii- TSF and pastured sites were not significant (p > dae). Zebra swallowtail caterpillars, Eurytides 0.05). However, plants within those sites were marcellus Cramer (Papilionidae), were found on significantly more likely to initiate fruit, have 15 study plants, predominantly in the 1-yr TSF more flowers initiating fruits, and to initiate site. more fruits than those within 4-yr or 15-yr TSF The percentage of plants on which potential sites. Trends in fruit retention were comparable pollinators were observed was lowest in the but not significant (Figure 3). Length of retained 15-yr TSF site but did not differ significantly fruit and number of seeds varied among sites; among 1-yr (48.1% – 7.0%,), 4-yr (47.7% – however, differences were not significant (AN- 7.6%), 15-yr (23.1% – 8.4%), and pasture (40.0% OVA, p > 0.05): mean fruit length (1-yr TSF, 3.2 – 8.4%) sites (binary logistic regression, p ¼ cm – 0.2 cm; 4-yr TSF, 3.3 cm – 0.2 cm; 15-yr 0.154). Across sites, pollinators were signifi- TSF, 2.4 cm – 0.4 cm; and pasture, 3.5 cm – 0.2 cantly more likely to visit larger and more cm) and number of seeds within each fruit (1-yr showy plants, those with more flowers present 328 CASTANEA VOL.83

Figure 3. Time-since-fire effects on fruit initiation/retention and in Asimina reticulata Shuttlew. ex Chapm. (A) Results from binary logistic regression models with odds of presence or absence of initiated or retained fruit (%Initiated/ Retained) as dependent variables. (B) Results from 1-way nonparametric analysis of variance for number of flowers to initiate or retain fruit. Graphs depict means – SE, with lettering denoting significant differences among sites (p < 0.05). at one time, and those that produced more Breeding System total flowers throughout the season. Addition- Hand-pollination treatments signiﬁcantly inﬂu- ally, plants on which pollinators were observed enced fruit and seed production in A. reticulata initiated and retained significantly more fruits (Figure 4). Fruit initiation from autogamous (Table 2). treatments was miniscule and resulted entirely

Table 1. Number of flower visitors to Asimina reticulata in flatwood sites with varying time-since-fire (1, 4, and 15 yr TSF) and in an actively grazed pasture site. Plus (þ) sign indicates the specimen was collected in its larval state

Order Family, Genus species 1 yr 4 yr 15 yr Pasture

Blattodea Blattellidae, Cariblatta lutea Saussure & Zehntner 2 0 0 7 Coleoptera Cantharidae, Chauliognathus (black/grey) sp. Hentz 0 1 0 0 Cantharidae, Chauliognathus (orange/black) sp. Hentz 0 1 0 1 Cerambycidae, Typocerus zebra Olivier 4 14 1 1 Coccinellidae, Harmonia axyridis Pallas 0 0 0 2 Scarabaeidae, Trichiotinus rufobrunneus Casey 41 31 2 4 Scarabaeidae, Trichiotinus lunulatus Fabricius 3 1 0 0 Curculionidae Black vine weevil Otiorhynchus sulcatus Fabricius 0 1 0 0 Diptera Bibionidae, Plecia nearctica Hardy 5 1 1 13 Hymenoptera Apidae, Apis mellifera L. 0 0 0 2 Lepidoptera þ Papilionidae, Eurytides marcellus Cramer 8 3 1 3 Totals (excluding þ)5550430 2018 BARTON, MENGES: REPRODUCTIVE SUCCESS OF ASMINIA RETICULATA 329

Table 2. Pollinator visitation in relation to plant, flower, and fruit variables; p values represent results from binary logistic regression models of odds with presence or absence of pollinator on plant as the dependent variable. Values represent means – SE across all sites, and n ¼ 157 for all models.

Model Visited Not Visited P Value

Plant volume (m3) 0.24 – 0.04 0.15 – 0.03 0.037 Maximum flowers present at one time 23.39 – 2.76 13.00 – 1.55 0.002 Total flowers produced 50.29 – 5.66 25.49 – 3.03 <0.001 Max fruit initiated 12.21 – 2.03 4.32 – 0.80 <0.001 Fruits retained 1.05 – 0.20 0.30 – 0.86 0.001 in fruit abortion. Indices of AI and SI conﬁrmed and fruit retention was much less than initiation that A. reticulata was not autogamous (AI ¼ across all treatments (Figure 5). There were no 0.07), which was well below the threshold value significant interactive effects between TSF and of 0.3, but was self-compatible (SI ¼ 1.2). hand-pollination treatments on fruit initiation or Although fruit initiation was greatest after retention. However, pollen donated from plants geitonogamous pollination, few of those fruits within the 15-yr TSF site was significantly less were retained to maturity. Compared with likely to initiate fruit (21%) than pollen from the geitonogamy (mean percentage of fruits retained other sites (55–56%; binary logistic regression, p ¼ 9.1% – 5.1%), far-outcrossed flowers were ¼ 0.007) but had no significant effect on the significantly more likely to retain fruits (39.6% – number of fruits initiated or the fruits retained. 3.2%; binary logistic regression, p ¼ 0.003). Number of fruits retained per flower was also Length of retained fruit and numbers of seeds significantly greater after far outcross (0.87 – 0.2 varied slightly among pollination treatments; fruits) and close outcross (0.64 – 0.17) than it however, differences were not significant (AN- was after geitonogamy (0.12 – 0.07, p ¼ 0.002 OVA, p > 0.05) for the following variables: mean and p ¼ 0.054, respectively). There were no fruit length (geitonogamy, 3.5 cm – 0.4 cm; close significant differences in fruit initiation or outcross, 3.5 cm – 0.2 cm; and far outcross, 4.0 retention between close and far outcrosses, cm – 0.2 cm) and number of seeds within each

Figure 4. Effects of hand-pollination treatment on fruit initiation and retention. (A) Results from binary logistic regression models with odds of presence or absence of initiated or retained fruit as dependent variable. (B) Results from 1-way nonparametric analysis of variance for the number of fruit initiated or retained per flower. Graphs depict means – SE, with lettering denoting significant differences among treatments (Tukey’s honestly significant difference, p < 0.05). A: n ¼ 45 flowers, G: n ¼ 33 flowers, CC: n ¼ 39 flowers, and FC: n ¼ 38 flowers. 330 CASTANEA VOL.83

Across all TSF and pollination treatments, scariﬁcation marginally reduced days to germination but did not signiﬁcantly increase germination success (Table 3). Among the treatments with the greatest seed production (close and far outcross), close- outcrossed seeds germinated in significantly fewer days (p < 0.001). Although not significant, seeds from the 15-yr plot and from geitonogamous pollination treatments were slower to germinate and had less germination success than those from more recently burned plots (p ¼ 0.604 and p ¼ 0.465). No seeds were produced after autogamy and were, therefore, excluded from these analyses.

DISCUSSION Fire is a dominant ecological disturbance worldwide (Bond and Keeley 2005) Figure 5. Total water inundation (in grams) of seeds and may maintain species diversity and fecundity scarified with sandpaper, scarified with a razor, or not of various plant communities (Pickett and White scarified (controls) after being submerged for 3, 12, 18, 1985, Gardner and Engelhardt 2008). In Florida and 24 hr. Figure depicts means – SE, and n ¼ 7 for each treatment. scrub, TSF is especially important in affecting species composition, with many species, including Asimina obovata (Menges unpubl. data), fruit (geitonogamy, 4.2 – 0.9; close outcross, 3.8 exhibiting strong postfire resprouting (Menges – 0.4; and far outcross, 5.0 – 0.3). and Kohfeldt 1995, Schmalzer 2003). Fire also Seed Dispersal creates more and larger gaps within the shrub matrix of scrubby flatwoods (Dee and Menges Motion-sensor field cameras, trained to fruiting 2014) and rosemary scrub (Menges et al. 2017a). A. reticulata plants outside of the study popu- These gaps provide a competitive release from lations, captured three night-time visitors: a rat the roots and foliage of matrix shrubs and trees (Rattus sp. Fischer de Waldheim), a mouse (Mus (Petru and Menges 2003) and may also promote sp. L.), and a raccoon (Procyon lotor L). Footage greater soil water during drought (Weekley et al. clearly revealed the raccoon as it removed a fruit 2007). and proceeded to consume it whole. Seeds This study confirms that, although A. retic- within the fruit were not spit out during the ulata may persist in longer-unburned plots, its video but did not appear to be damaged during reproductive success may be reliant on either consumption. No seeds were visible under the frequent burning or mowing. These disturbances plant upon retrieval of footage. The rat was stimulated new growth (increased stem count), unable to retrieve the high dangling fruit without increased showiness (maximum number of toppling the flimsy stems, and the mouse flowers present at one time), and increased searched the plant after the raccoon had already flower output (total flowers produced). With removed all fruits. increased visibility to potential pollinators (e.g., the hairy flower scarab beetle) or increased Seed Germination resource availability, plants in more recently Water imbibition did not significantly differ burned or mowed sites were also able to initiate between our manual scarification treatments and retain more fruit than those in longer- (Figure 5). However, seeds that were scarified unburned sites. with either treatment were able to imbibe Our study confirms that A. reticulata is self- significantly more water over a 24-hr period than compatible only if pollen is able to move among unscarified controls (F ¼ 3.331, p ¼ 0.059). flowers, and we found that, in most cases, fruit Germination of all collected seeds extended was aborted prematurely and was found littering from 30 July 30 2016 to 13 February 2017 and the ground beneath the plants. Although the occurred between 35 d and 233 d after sowing. hairy flower scarab beetle is a frequent pollina- 2018 BARTON, MENGES: REPRODUCTIVE SUCCESS OF ASMINIA RETICULATA 331

Table 3. Germination percentage and time as a function of time-since-fire, hand-pollination treatment, and seed-scarification treatment. Results from binary logistic regression models with odds of presence or absence of germination as dependent variables and from 1-way nonparametric analyses of variance for days to germination. Table depicts means – SE. Significant differences within models (Tukey’s honestly significant difference, p < 0.05) are denoted with different letters

Variables % Germination No. of Seeds Time (d) No. of Seeds

Time-since-fire (yr) 1 36.5 – 4.0 148 132.4 – 7.4 54 4 35.7 – 6.4 56 116.8 – 12.5 20 15 42.9 – 20.2 26 153.3 – 2.9 3 Pasture 40.2 – 5.0 97 120.8 – 8.8 39 p value 0.604 0.465 Hand pollination Geitonogamy 27.8 – 10.9 18 139.8 – 23.3AB 5 Close cross 41.6 – 5.7 77 92.5 – 8.5B 32 Far cross 46.4 – 6.1 69 143.5 – 7.5A 32 p value 0.726 <0.001 Scarification Scarified 39.9 – 3.2 234 117.1 – 5.4 90 Control 38.5 – 3.2 238 130.9 – 5.7 95 p value 0.747 0.077 tor of A. reticulata, these data suggest that it Video footage of a raccoon consuming entire may be not be entirely effective in producing fruits suggests that seeds may benefit from gut high-quality pawpaw offspring. The percentage scarification, but future studies are needed to of fruit retention in open-pollinated flowers confirm that. Raccoons are known to disperse (6.2%) was most similar to that after geitonog- seeds of several species in North America, amy (9.1%), suggesting that these beetles are including other members of the genus Asimina, geitonogamously pollinating within a single plant and have been documented excreting intact and and that low levels of fruit initiation and germinable seeds (Willson 1993). Raccoons have retention may be due to the lack of an effective also been shown to increase germination per- pollinator. However, genetic data are necessary centages of some tree species in Illinois (but not to assess relatedness of plants within a popula- that of A. triloba; Cypher and Cypher 1999) and tion and are vital to determining whether low will often deposit large numbers of seeds in levels of fruit retention are a result of inbreeding ‘‘latrines,’’ which may not be exploited by seed predators during times of high food availability depression. (LoGiudice and Ostfeld 2002). Although more Despite low levels of fruit retention, seed observations are needed, it seems probable that viability was surprisingly high and may explain raccoons are successful dispersers of the hard why A. reticulata is less rare than its congener seeds from A. reticulata. A. obovata. Although not significant, likely Our findings for A. reticulata have strong because of low levels of fruit retention, germi- parallels with its co-occurring congener A. nation rates were lower after geitonogamous obovata (Levitt et al. 2013). Both species require crosses than they were after outcrossing treat- pollinators to produce seeds, have pollinator- ments. Although the potentially ancient breeding limited fruit set, and are pollinated by some of system of A. reticulata may not have evolved a the same beetles. In both species, geitonoga- mechanism to prevent geitonogamy, fruit abor- mous crosses result in high levels of fruit tion from inbreeding depression may be a abortion, likely because of inbreeding depres- mechanism to divert resources from seeds that sion. In A. reticulata, low fruit set in open- are less likely to germinate. Quicker germination pollinated plants was similar to that after times suggest that close outcrossing and manual geitonogamous crosses, likely because of the scarification may ensure the greatest reproduc- absence of an effective pollinator. Furthermore, tive success in A. reticulata; however, the our data suggest that A. reticulata reaches its effects of delayed germination have not been maximum reproductive potential in open areas, examined in this system. available shortly after fire, or in pastured areas 332 CASTANEA VOL.83 and has higher fecundity after outcrossing. Dean, S., E.C. Farrer, and E.S. Menges. 2015. Fire Frequent fires, which are often recommended effects on soil biogeochemistry in Florida for flatwood sites (Breininger et al. 2002), will, scrubby flatwoods. Amer. Midl. Naturalist therefore, favor seed production, recruitment, 174:49–64. and population growth in this Florida endemic Dee, J.F. and E.S. Menges. 2014. Gap ecology in shrub. the Florida scrubby flatwoods: effects of time- since-fire, gap area, gap aggregation and ACKNOWLEDGMENTS We thank mem- microhabitat on gap species diversity. J. Veg. bers of Archbold Biological Station’s Plant Sci. 25:1235–1246. Ecology Program, especially Stephanie Koontz, Lauren Maynard, Stacy Smith, Kathryn Tisshaw, Gardner, R.H. and K.A.M. Engelhardt. 2008. Brian Josey, Tabitha Petri, Evan Craine, and Spatial processes that maintain biodiversity Oliva Karas for support and advice throughout in plant communities. Perspect. Pl. Ecol. Evol. this project. 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