Diurnal versus nocturnal pollination of Brunsvigia gregaria ... - CiteSeerX

South African Journal of Botany 72 (2006) 291 – 294 www.elsevier.com/locate/sajb

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Diurnal versus nocturnal pollination of Brunsvigia gregaria R.A. Dyer (Amaryllidaceae) at a coastal site B. Balmford a, J. Balmford a, A. Balmford b,c, S. Blakeman a, A. Manica c, R.M. Cowling d,* a

Cape St Francis Travelling Academy, c/o Department of Botany, Nelson Mandela Metropolitan University, P.O. Box 77000, Port Elizabeth 6031, South Africa b Percy Fitzpatrick Institute of African Ornithology, University of Cape Town, Private Bag, Rondebosch 7701, South Africa c Department of Zoology, University of Cambridge, Cambridge CB2 3EJ, UK d Department of Botany, Nelson Mandela Metropolitan University, P.O. Box 77000, Port Elizabeth 6031, South Africa Received 2 June 2005; accepted 25 July 2005

Abstract Members of the amaryllid genus Brunsvigia appear to show considerable variation in pollination syndromes, but experimental studies have been limited to two species, one pollinated by sunbirds and one by moths. Here we present the results of a simple exclusion experiment designed to shed light on the pollinators of the narrowly distributed species B. gregaria. Experimental flowers were left unbagged, or were bagged during night-time only, day-time only, or both day-time and night-time. Treatments started when flowers first matured and continued until senescence, after which we counted the number of fertile seeds per capsule. Flowers in treatment groups left open during the daytime produced significantly more seeds than flowers bagged during the daytime, whereas nocturnal bagging made little difference to seed number, indicating near-complete dependence on diurnal pollinators. Based on anecdotal observations we speculate that sunbirds — specifically Nectarinia famosa and possibly Cinnyris afer — are the most likely pollinators of B. gregaria. D 2005 SAAB. Published by Elsevier B.V. All rights reserved.

Plants of the Greater Cape Floristic Region (the combined succulent karoo and fynbos regions; Ju¨rgens, 1997; Born et al., in press) — most notably bulbs (or geophytes) — are renowned for their attractive blooms, and many are well-established horticultural plants (Manning et al., 2002). This region — hereafter referred to as the Cape — has the largest bulb flora in the world; some 2100 species in 20 families have been recorded from this area, 84% of them endemic (Proches¸ et al., in press). The showiness of their flowers is thought to be a response of a largely obligate out-crossing flora (Johnson and Bond, 1997) that flowers mainly in the cool-season (winter and spring; Johnson, 1992), when pollinator activity is constrained by inclement weather (Struck, 1994; Johnson and Bond, 1997). Consequently, many Cape plants are pollinator-limited (Johnson and Bond, 1997) and so invest heavily in what we perceive as attractive flowers as a means of advertising for the services of scarce insect and vertebrate pollinators (Cowling et al., 1999).

* Corresponding author. Tel.: +27 42 2980259. E-mail address: [email protected] (R.M. Cowling).

Numerous highly specialised plant – pollinator systems are a hallmark of the Cape flora (e.g. Goldblatt et al., 1995; Manning and Goldblatt, 1996; Johnson, 1997; Steiner, 1998; Goldblatt and Manning, 2000a; Johnson and Steiner, 2000; Pauw, 2004). Pollinators are thought to be major drivers of speciation in the flora, especially among geophytes (Johnson, 1996a,b; Johnson and Steiner, 1997, 2000). Brunsvigia (Amaryllidaceae: Amaryllideae) is a bulbous genus of 17 species, all endemic to southern Africa (Snijman and Archer, 2003). Of these, 13 species are Cape plants, confined to dry fynbos and succulent karoo vegetation, from southern Namibia to the vicinity of Port Elizabeth (Goldblatt and Manning, 2000b). They are summer-deciduous and hysteranthous, with flowers appearing before the leaves (Snijman and Linder, 1996). The striking pink, red or white flowers, which appear in late summer to autumn (Feb –May), are arranged in a spherical head comprising several helicoid cymes, which are borne on a stout, leafless stem or scape (Mu¨ller-Doblies, 1977). After pollination, the dry inflorescence abscises at ground level, and is carried by the wind — tumbleweed like — thus enabling dispersal of the fleshy, ovoid

0254-6299/$ - see front matter D 2005 SAAB. Published by Elsevier B.V. All rights reserved. doi:10.1016/j.sajb.2005.07.008

B. Balmford et al. / South African Journal of Botany 72 (2006) 291 – 294

seeds that are loosely held in large, inflated capsules that form the outer periphery of the inflorescence (Snijman and Linder, 1996). The few studies conducted to date indicate considerable variation in pollination syndromes within Brunsvigia. Detailed experimental and observational evidence indicates that B. orientalis (L.) Aiton ex Eckl. is pollinated diurnally, very largely by malachite and lesser double-collared sunbirds, Nectarinia famosa and Cinnyris chalybea (Pauw, 2004). On the other hand similar studies of B. bosmaniae F.M. Leight reveal that it is pollinated at night, by sphingid and noctuid moths (Raimondo, 1998). For other species the available evidence is more anecdotal, although observations suggest that B. grandiflora Lindl. and B. striata (Jacq.) Aiton are pollinated by the diurnal long-tongued fly Prosoeca ganglbaurii (Goldblatt and Manning, 2000a), and B. marginata is pollinated by the butterfly Meneris tulbaghia (Johnson and Bond, 1994). Very little is known about the pollination biology of the species examined here, B. gregaria R.A. Dyer, which has so far only been recorded from coastal sites between Cape St Francis and Port Elizabeth. Observational evidence has been used to infer that it too is pollinated by P. ganglbaurii (Goldblatt and Manning, 2000a). B. gregaria inflorescences lack the features which facilitate bird pollination in B. orientalis (Pauw, 2004) and, putatively, B. litoralis and B. josephinae (D. Snijman pers. comm.). The latter species have flowers borne on stout, upwardly curving pedicels — thereby providing easy access for visiting birds — whereas the 20 –40, dark pink to crimson flowers of B. gregaria are borne on relatively narrow and straight pedicels: consequently, the flowers are not easily accessed by birds. Moreover, in the bird-pollinated B. orientalis, the curvature of the pedicel ensures that the stamens have an erect orientation, which facilitates contact with the head of nectar-probing sunbirds (Pauw, 2004). In B. gregaria, the stamens are deflexed downwards, thereby reducing the potential for such contact. On the other hand, unlike the mothpollinated B. bosmaniae (Raimondo, 1998), the flowers of B. gregaria are not strongly scented at night, and are more brightly coloured. In this paper, we report the results of an experimental study of pollination and seed set in B. gregaria at a coastal site in Cape St Francis, in the eastern part of the Cape. Specifically, we placed fine-mesh bags on flowers either by day or night (or both) following a matched-flowers design and then quantified subsequent seed number to examine the relative roles of nocturnal (moths) versus diurnal (birds or other insects) vectors in pollinating this plant. Our study area was located immediately west of the small village of Cape St Francis (24-50VE, 34-13VN) on a rise 30 – 50 m above the high water mark that is exposed to the full brunt of the prevailing south-westerly winds. The plant community consisted of a dense species-rich turf of Stenotaphrum secundatum growing on a shelly, calcareous sand of marine origin. B. gregaria occurred in a patch approximately 70 m long and 30 m across, at a density of up to 3 plants m 2. The nearest patches of similar size were ¨1 km away. We have lodged a voucher specimen from the study area in NBG.

We set up the experiment on 29 March 2005. Working from the centre of the patch, where B. gregaria grew most densely, we selected 20 inflorescences, the only criterion being that each had to have at least four mature but unopened buds. We then randomly assigned one bud per inflorescence to each of four treatments: Treatment A (Control 1) — left continually exposed, with no bag; Treatment B — bagged during night-time only, to exclude nocturnal but not diurnal vectors; Treatment C — bagged during day-time only, to exclude diurnal but not nocturnal vectors; Treatment D (Control 2) — bagged during both day and night, but with the bag removed and immediately replaced once daily (so that the flower experienced a similar degree of manipulation to flowers in treatments B and C). The exclusion bags were made of 15 denier nylon tights supported on a 1 mm-gauge wire frame 50 mm in diameter and 50 mm deep and secured to the pedicel using fine copper wire. To reduce the risk of pollen contamination between flowers, each flower in treatments B – D had its own bag. The bags were placed on buds in treatments B and D at dusk on 29 March. After that, plants were visited twice daily: once around dawn (between 0625 and 0715), when we removed bags from Treatment B flowers, placed bags on Treatment C flowers, and removed and immediately replace bags on Treatment D flowers; and once around dusk (between 1805 and 1850) to replace Treatment B bags, and remove Treatment C bags. These visits continued until the flowers senesced, which occurred between 7 and 10 April. Upon senescence we bagged all the study flowers, to prevent seed predation, until the capsules had ripened. We then harvested the capsules and on 23 April scored the number of fertile seeds per capsule. Before this could be done we lost 11 flowers (one each from 12 10

No. seeds/ capsule

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8 6 4 2 0

A

B

C

D

Treatment group Fig. 1. Seed set in Brunsvigia gregaria at Cape St Francis during April 2005. Treatment groups depicted in box and whisker plots are: A = unbagged; B = bagged during night-time only; C = bagged during day-time only; D = bagged during both day and night. Horizontal bars within the boxes are medians, the boxes shows the inter-quartile ranges, and the upper bars are maxima; minima are zero in all treatments. For each treatment the sample size is 17, except Treatment C where n = 18.


seven plants, and all four from one plant) as a result of damage by wind or dogs, so the results reported here are from a reduced sample of 69 flowers. To provide context on levels of pollination, on 3 April we also dissected an additional 20 freshly opened flowers (not included in the present sample), and counted the numbers of ovules contained per capsule. We found that seed number per capsule in our unmanipulated flowers (Treatment A) was variable and generally low. In our dissections, the ovaries of freshly opened flowers contained a median of 13 ovules (range = 12– 20, n = 20 flowers). In contrast, the median number of seeds in the ripe capsules of flowers in Treatment A, which had been continually exposed to pollinators, was only 3 (range = 0 –10, n = 17 flowers). The median (and range in) number of seeds per capsule in each of the four treatment groups is plotted in Fig. 1. To take account of our matched-flowers design and the non-parametric distribution of our response scores we initially analysed these data using a Friedman test. This was highly significant (S [adjusted for ties] = 21.86, n = 12 plants with complete set of treatments, P < 0.001), showing there was a strong overall effect of treatment on seed number. However, the post hoc pairwise tests associated with the Friedman test are weak, and by using only inflorescences with complete data, this analysis made no use of results from seven of our study plants. For a more powerful and detailed assessment of differences in seed number between treatment groups, we therefore fitted a Generalised Linear Model (GLM, in R version 2.0.1) with a quasi-Poisson error structure to data for all 69 flowers. Entering treatment as a factor and plant identity as a block, this was again highly significant ( F 21, 47 = 4.71, P < 0.001). To look at between-treatment differences we then performed Tukey post hoc tests on the GLM (using the package Multcomp v0.4 – 8). These showed that flowers from both Treatments A and B (both exposed during the day) had significantly higher seed number than flowers from either Treatments C or D (both bagged during the day; Tukey post hoc results: A vs C: P < 0.01; A vs. D: P < 0.01; B vs. C: P < 0.05; B vs. D: P < 0.05). Hence bagging during daytime significantly reduces seed number per capsule. In contrast, we found only a very weak difference in seed number between flowers in Treatments A and B (unbagged throughout vs bagged during the night only; P < 0.1) and no difference between flowers in Treatments C and D (bagged during daytime only vs bagged throughout; NS). It seems likely that the non-significant difference between groups A and B reflects the effects of daily bagging, and we are therefore left with no evidence that bagging during the night reduces seed number. Our results suggest that B. gregaria is an obligate outcrosser. Only one of the 17 flowers in Treatment D (bagged throughout) and four out of 18 in Treatment C (bagged during daytime) set seed. These seeds could conceivably have been produced by cross-fertilisation, through a few buds having slightly opened before the experiment began, or via extremely early daytime or occasional night-time pollination (in Treatment C), or accidental pollen transfer by us. However, the mean seed number in these groups (just 0.06 and 0.33 seeds per

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capsule, respectively) are similar to those recorded for permanently bagged or manually self-pollinated flowers in B. orientalis (means of 0.26 and 0.38 seeds per capsule; Pauw, 2004). Very limited self-pollination therefore seems a more likely explanation, confirming the importance of pollinators in Brunsvigia breeding systems. For B. gregaria in Cape St Francis, the consistent effect of daytime but not night-time bagging in substantially lowering seed number indicates that these pollinators are diurnal. Moth pollination is rare among amaryllids that grow in the fynbos region of the Greater Cape Floristic Region, possibly owing to the absence of suitable larval food plants for the sphingid pollinators (Manning and Snijman, 2002). While our experiment was not designed to help us further identify the pollinators, several lines of evidence suggest that they are most likely to be sunbirds. Two species of sunbird — the malachite and the greater double-collared (Cinnyris afer) — are abundant at Cape St Francis during March and April. We frequently observed an apparently territorial male malachite in the study area, feeding on B. gregaria (including treated inflorescences) and chasing away conspecifics. Last, by the end of the flowering season many inflorescences had pedicels bearing apparent clawmarks along their length. It is possible that long-tongued flies play an additional role in pollinating B. gregaria, but although Prosoeca is present in the area, we have not seen it there during March or April. Further fieldwork on this issue is clearly warranted. Despite the abundance of sunbirds, it seems probable that B. gregaria is pollinator-limited at Cape St Francis. Even in our permanently exposed flowers (Treatment A), median seed number was less than one quarter of the median ovule count per capsule. Frequent visits by a territorial male malachite to treated inflorescences suggest it is unlikely that low seed number arose because of any deterrent effect of the exclusion bags on other flowers on the inflorescence. Maternal control may provide an alternative explanation that requires direct testing (Roach and Wulff, 1987). However, the fact that in B. orientalis similar levels of natural seed production were substantially boosted by manual cross-pollination (Pauw, 2004) suggests that pollinator-limitation may be likely in B. gregaria too. Acknowledgements Comments by Anton Pauw and two anonymous referees greatly improved this manuscript. In addition we thank Dee Snijman for advice and encouragement, Shirley Pierce for floral expertise and tights, Liesel von der Marwitz for help with fieldwork, Anton Pauw and Tilla Raimondo for permission to cite their unpublished work, and the owners of Rocky Coast Farm for permission to work on their property. References Born J., Linder H.P., Desmet P., in press. The Greater Cape Floristic Region. Journal of Biogeography. Cowling, R.M., Esler, K.J., Rundel, P.W., 1999. Namaqualand, South Africa — an overview of a unique winter – rainfall desert ecosystem. Plant Ecology 142, 3 – 21.

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