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3 Department of Evolution, Ecology, and Organismal Biology, The Ohio State University, 1735 Neil Avenue, Columbus, Ohio 43210-1293 USA; and 4 Department of Biological Sciences, Clemson University, Clemson, South Carolina 29634-1903 USA
Received for publication August 10, 1999. Accepted for publication January 18, 2000.
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ABSTRACT |
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 TOP ABSTRACT INTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONLITERATURE CITED |
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Key Words: allozyme marker • Hibiscus • male fitness • Malvaceae • pollen tube competition • pollination • timing of pollen deposition
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INTRODUCTION |
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TOPABSTRACTINTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONLITERATURE CITED |
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; Snow, 1986
; Snow and Roubik, 1987
; Levin, 1990
; Spira et al., 1992
). When this occurs, plants with the greatest success as pollen donors may be those whose pollen arrives earliest on the stigmas of newly open flowers and/or those whose pollen is able to sire seeds most quickly after pollen mixtures are deposited. These two types of competition often take place during the first few hours after anthesis and are influenced by the complex dynamics of pollinator movements, availability of ovules, and variable rates of pollen tube growth. Despite a large number of studies on nonrandom fertilization in plants, we know little about whether this process influences male fitness in natural populations (Stephenson and Bertin, 1983
; Lyons et al., 1988
; Marshall and Folsom, 1991
; Walsh and Charlesworth, 1992
; Snow, 1994
).
Hibiscus moscheutos, commonly known as wild rose mallow, is a well-studied species in which pollen competition may affect male fitness (Snow and Spira, 1996
). At our study sites in Edgewater, Maryland, USA, bumble bees (Bombus spp.) and a specialist anthophorid bee (Ptilothrix bombiformis) make frequent visits to the large, 1-d flowers. More than half of the consecutive pollinator visits we observed occurred within 15 min of each other, and pollinators typically deposited surplus pollen (more than 
350 pollen grains) within 2–3 h of anthesis (Snow and Spira, 1991a
; Spira, Snow, and Puterbaugh, 1996
). Pollen carryover, frequent stigma contacts by pollinators, and the close proximity of neighboring plants suggest that stigmas often receive pollen from multiple donor plants, either simultaneously or following brief delays between visits (Spira et al., 1992
).
When excessive amounts of pollen from several donors are deposited on stigmas simultaneously, the speed of pollen tube growth down the 5–6 cm style of H. moscheutos can determine the siring success of competing pollen donors. Using two-donor hand-pollination experiments, we found that individual pollen donors often differed in the proportion of seeds sired, in some cases by ratios exceeding 2:1 (Snow and Spira, 1996
). Furthermore, these differences were consistent across a range of recipient plants. Competitive hierarchies were also consistent among donors, allowing us to identify "fast" and "slow" pollen donors relative to the population as a whole. Therefore, we concluded that pollen competitive ability may be an important component of male reproductive success in these populations (Snow and Spira, 1991b, 1996
). Differential siring success is likely due to corresponding differences in pollen tube growth rates (Snow and Spira, 1991a
), although this was not examined in all of our studies.
A complicating factor in evaluating the fitness consequences of pollen tube competition in H. moscheutos is the fact that several pollinator visits were often required to foster strong competition for ovules, giving a head-start advantage to pollen that arrived early (Spira et al., 1992
). For pollen donors with similar competitive abilities, we found that delays of only 15–30 min between visits ensured that 60–75% of a flower's ovules were fertilized by the first of two donors, as compared to 50% from each donor following simultaneous deposition (Spira, Snow, and Puterbaugh, 1996
). However, these time-dependent inequalities would not necessarily offset differences in pollen competitive ability if the order of arrival is random with respect to pollen tube growth rates, as is likely to occur in nature. The penalty of arriving late should be greater for slow donors than for fast ones. Therefore, we undertook the present study to test the assumption that the average siring success of fast pollen donors is greater than that of slow donors when each is subjected to a 15 or 30 min delay relative to the other. We hypothesized that when influenced by similar time delays, fast pollen donors would sire significantly more seeds than slow donors.
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MATERIALS AND METHODS |
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TOPABSTRACTINTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONLITERATURE CITED |
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). Individual plants consist of multiple 1–2 m tall stems that emerge each spring from a woody rootstock. Genets can be propagated vegetatively, but they are not clonal under natural conditions. Flowering occurs in mid- to late summer. Automatic self-pollination is prevented by spatial separation of the anthers and stigmas, and outcrossing rates are estimated to be 64% (Snow and Spira, 1991b
; Snow et al., 1995
). The showy white or pink flowers are visited frequently by bees and typically produce 120 seeds per fruit. The round, hard-coated seeds are often destroyed by insect seed predators; undamaged seeds are bouyant and appear to be dispersed by water (Spira, 1989
).
We collected plants from the Mill Swamp population in Edgewater, Maryland, as described in our previous studies, and planted them in 20-L pots kept outdoors at the South Carolina Botanical Garden at Clemson University, Clemson, South Carolina, USA. Although this site is distant from the source population in Maryland, the climate is similar and the plants became dormant and resprouted each year on a schedule similar to their counterparts in Maryland (but plants grown in South Carolina flowered 1 mo earlier than those in Maryland). Each genotype was cultivated in the same manner from 1993 to 1996, and in many cases these genotypes were propagated as several potted ramets to increase the numbers of available flowers per genotype. These genotypes (hereafter referred to as pollen donors or pollen recipients) had three Mendelian alleles for GPI (glucose phosphate isomerase), based on starch gel electrophoresis with Gottlieb's (1981)
extraction buffer and procedures described in Soltis et al. (1983)
. Plants used as competing pollen donors differed in allozyme markers, such that the siring success of each donor could be determined by scoring seeds from experimental crosses (Table 1).
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. Pollen donors were designated as "fast" or "slow" based on their competitive ability in previous experiments (Snow and Spira, 1996
; unpublished data). We carried out mixed-donor hand-pollinations using each of four pairs of pollen donors in the following treatments: (1) no delay—fast and slow pollen applied simultaneously, (2) 15-min delay with either fast pollen first (F-15) or slow pollen first (S-15), or (3) 30-min delay with either fast pollen first (F-30) or slow pollen first (S-30).
Flowers on recipient plants were bagged just prior to anthesis and rebagged after hand-pollination to exclude pollinators. On newly opened flowers, two of the five stigmas received pollen from each donor in a pair, with the fifth stigma left unpollinated. Each stigma was coated with pollen by gently brushing it with pollen-laden anthers held in forceps. This resulted in more than twice the amount of pollen from each donor required to fertilize all available ovules (800 grains/donor, 1600 grains per flower; Snow and Spira, 1991a
). We did not attempt to mix pollen from the two sources prior to application, as advocated by Mitchell and Marshall (1995)
, because we wanted both early- and late-arriving pollen to be in contact with clean stigmatic surfaces. Thus, pollen from the two donors did not have the opportunity to interact on the surface of the stigma, although their pollen tubes converged 1–2 cm below the stigma, at the base of the stylar branches (Snow and Spira, 1991a
). In similar studies by other investigators, later pollen loads were placed directly over earlier pollen to test for stigma "clogging" (e.g., Galen, Gregory, and Galloway, 1989
). We opted not to use this approach because the five large stigmas of each Hibiscus flower can accommodate a total of 2000 pollen grains (Snow and Spira, 1991a
), so stigma clogging may be rare, and in any case we wanted to focus on pollen tube competition as a possible mechanism for nonrandom fertilization.
For each pair of competing donors, we pollinated eight flowers per treatment on each of three recipient plants, taking care to alternate which of the five treatments was started first on different days (4 donor pairs x 5 treatments x 3 recipients x 8 flowers = 480 flowers). Pollinations were carried out between 0830 and 1100 on days that were not rainy or unusually cool. About 4 wk later, mature but undehisced fruit capsules were collected and stored in labeled envelopes. To induce germination, seeds were nicked and placed on moist filter paper in petri dishes at room temperature. We then scored 150 seedlings for each treatment for electrophoretic markers to assign paternity (3 recipients x 5 fruits x 10 seedlings per fruit). This measure of paternal success likely reflects fertilization success rather than differences in seed abortion, germination, or seedling survivorship (Snow and Spira, 1991a, b
). Aborted embryos were extremely rare, seed germination exceeded 90%, and very few seedlings died prior to sampling, so we assumed that survivorship at these life stages did not affect paternal reproductive success.
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RESULTS |
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TOPABSTRACTINTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONLITERATURE CITED |
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62:38 (Table 1, Fig. 1). In two of the four pairs of donors, fast donors sired significantly more seeds than slow donors when each was subjected to delays of 15 or 30 min, as expected (donor pairs I and III; Fig. 1). However, the other two donor pairs did not show differences in siring ability under these conditions (donor pairs II and IV; Fig. 1). Pooling data from the four delay treatments (S-15, F-15, S-30, F-30) showed that the advantage of fast donors seen with simultaneous pollinations diminished and was not significant at P < 0.05 in two of the four pairs of pollen donors (Table 2).
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DISCUSSION |
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TOPABSTRACTINTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONLITERATURE CITED |
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Flower age could also contribute to the variation we observed in the effects of delayed pollination. In a previous paper, we reported that the penalty of arriving late decreased when pollinations were started at 1130 as opposed to 0930 (Spira et al., 1996
). In the present study, however, treatments were assigned at random throughout the morning and generally resulted in lower siring success after a time delay, suggesting that flower age was not responsible for the inconsistencies in our results.
Other factors that could affect pollen competitive ability are the genotypes of the three pollen recipients used with each pair of donors, or possible donor-by-recipient interactions (Table 1). Snow and Spira (1996)
found that differences in siring ability were significant across samples of 11–15 recipients, but no attempt was made to test for variation in siring success due to possible interactions between pollen donors and recipients. Here, with different groups of recipients for each donor pair, it is possible that such interactions could have had unanticipated effects on the dynamics of competing pollen tubes, although G tests (not shown) revealed few differences among recipients within treatments. Also, donor-by-recipient interactions would have to account for differences in relative siring success between delayed pollinations and simultaneous applications.
Regardless of the underlying reasons for our unexpected findings, this research shows that competition between pollen grains from different pollinator visits is more complicated than we initially assumed. Staggered pollen arrival times are common in natural populations, so earlier arriving pollen often has a substantial head start over later arriving pollen in sending pollen tubes to the ovary (e.g., Mulcahy, Curtis, and Snow, 1983
; Spira et al., 1992
). In H. moscheutos this head start weakens the intensity of competition among pollen genotypes and sometimes changes the outcome of competition based on what is expected following simultaneous pollinations (Fig. 1; note, however that all significant differences between pairs of donors were in the predicted direction). Here, pollen from relatively fast pollen donors did not always sire more seeds than pollen from slow donors with the same delay in arrival. Delays of only 15 min sometimes masked differences in siring success that were seen following simultaneous pollinations. Perhaps a larger sample of competing pollen donors (and more seeds sampled per donor) would have identified plants that exhibit greater or more consistent differences in siring success. In conclusion, results from this study indicate that fitness consequences of pollen competition in natural populations of H. moscheutos may be negligible or difficult to detect.
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FOOTNOTES |
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2 Author for reprint requests.
3 Current address: Department of Biology, Florida International University, Miami, Florida 33199 USA.
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LITERATURE CITED |
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TOPABSTRACTINTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONLITERATURE CITED |
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Gottlieb, L. D. 1981 Gene number in species of Asteraceae that have different chromosome numbers. Proceedings of the National Academy of Sciences, USA 78: 3726–3729.
Levin, D. A. 1990 Sizes of natural microgametophyte populations in pistils of Phlox drummondii. American Journal of Botany 77: 356–363.
Lyons, E. E., N. M. Waser, M. V. Price, J. Antonovics, and A. F. Motten. 1988 Sources of variation in plant reproductive success and implications for concepts of female choice, male–male competition, and sexual selection. American Naturalist 134: 409–433.[CrossRef][ISI]
Marshall, D. L., and M. W. Folsom. 1991 Mate choice in plants: an anatomical to population perspective. Annual Review of Ecology and Systematics 22: 37–63.
Mitchell, R. J., and D. L. Marshall. 1995 Effects of pollination method on paternal success in Lesquerella fendleri (Brassicaceae). American Journal of Botany 82: 462–467.[CrossRef][ISI]
Mulcahy, D. L., P. S. Curtis, and A. A. Snow. 1983 Pollen competition in a natural population. In C. E. Jones and R. J. Little [eds.], Handbook of experimental pollination biology, 330–337. Van Nostrand Reinhold, New York, New York, USA.
SAS. 1994 SAS Institute, Cary, North Carolina, USA.
Snow, A. A. 1986 Pollination dynamics of Epilobium canum (Onagraceae): consequences for gametophytic selection. American Journal of Botany 73: 139–151.[CrossRef][ISI]
———. 1994 Postpollination selection and male fitness in plants. American Naturalist 144: S69–S83.[CrossRef][ISI]
———, and D. W. Roubik. 1987 Pollen deposition and removal by bees visiting two tree species in Panama. Biotropica 19: 57–63.
———, and T. P. Spira. 1991a Differential pollen-tube growth rates and nonrandom fertilization in Hibiscus moscheutos (Malvaceae). American Journal of Botany 78: 1419–1426.[CrossRef][ISI]
———, and ———. 1991b Pollen vigor and the potential for sexual selection in plants. Nature 352: 796–797.[CrossRef]
———, and ———. 1996 Pollen-tube competition and male fitness in Hibiscus moscheutos. Evolution 50: 1866–1870.
———, ———, R. Simpson, and R. A. Klips. 1995 The ecology of geitonogamous pollination. In D. L. Lloyd and S. C. H. Barrett [eds.], Floral biology, 191–216. Chapman and Hall, New York, New York, USA.
Soltis, D. E., C. H. Haufler, D. C. Darrow, and G. J. Gastony. 1983 Starch gel electrophoresis of ferns: a compilation of grinding buffers, gel and electrode buffers, and staining schedules. American Fern Journal 73: 9–27.[CrossRef][ISI]
Spira, T. P. 1989 Reproductive biology of Hibiscus moscheutos. In J. H. Bock and Y. B. Linhart [eds.], The evolutionary ecology of plants, 247–255. Westview, Boulder, Colorado, USA.
———, A. A. Snow, and M. N. Puterbaugh. 1996 The timing and effectiveness of sequential pollination in Hibiscus moscheutos. Oecologia 105: 230–235.
———, ———, D. F. Whigham, and J. L. Leak. 1992 Flower visitation, pollen deposition, and pollen-tube competition in Hibiscus moscheutos (Malvaceae). American Journal of Botany 79: 428–433.[CrossRef][ISI]
Stephenson, A. J., and R. I. Bertin. 1983 Male competition, female choice, and sexual selection in plants. In L. Real [ed.], Pollination biology, 110–149. Academic Press, New York, New York, USA.
Walsh, N. E., and D. Charlesworth. 1992 Evolutionary interpretation of differences in pollen tube growth rates. Quarterly Review of Biology 67: 19–37.[CrossRef]
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| HOME | HELP | FEEDBACK | SUBSCRIPTIONS | ARCHIVE | SEARCH | TABLE OF CONTENTS |
