Andromonoecy and buzz pollination in Solanum species ( Solanaceae ) endemic to the Canary Islands by

We investigated the pollination and reproductive biology of two Canary Island endemics, Solanum vespertilio and S. lidii (Solanaceae). We measured male function (pollen development), female function (fruit initiation) and spatial arrangement of reproductive parts within flowers and inflorescences. Furthermore, we observed flower visitors and monitored visitation rates. Both species of Solanum display andromonoecy: Longstyled flowers are functionally hermaphrodite and borne proximally on the inflorescences, while short-styled flowers are functionally male and borne distally on inflorescences. Large bees capable of buzzing were the main flower visitors. In particular, the endemic Canarian bumblebee, Bombus terrestris canariensis, was a frequent visitor and pollen vector of S. vespertilio.


Introduction
In the endemic flora of the Canary Islands (514 spp.), relatively few species show sexual dimorphism: heterostyly (one sp.) (Olesen & al., 2003); dioecy (16 spp.); monoecy, gynomonoecy or gynodioecy (46 spp.) (Helfgott & al., 2000).Andromonoecy has not been reported.However, two endemic Canarian species of Solanum, S. vespertilio and S. lidii, bear short-styled and long-styled flowers on the same plant individual.Functionally they may be andromonoecious (Whalen & Costrich, 1986;Diggle, 1991;Diggle, 1994).Pollen is contained in sac-like poricidal anthers characteristic of buzz-pollination.This implies that pollen can only be removed from the anthers by thoracic muscle vibration of bees.However, to date no studies have investigated the pollination and re-productive systems of these rare endemic species.
Here we address the questions: Does female function (fruit initiation) and male function (pollen development) differ between short and long styled flowers?What is the spatial arrangement of reproductive parts within and between flowers?Which insects visit the flowers, and which are potential pollinators?

Materials and methods
The two Solanum species of this study belong to the subgenus Leptostemonum section Nycterium.Solanum lidii Sunding is confined to Gran Canaria, while S. vespertilio Aiton is found on Tenerife and Gran Canaria.Both species are classified as vulnerable (Gómez Campo, 1996).The plants are small woody shrubs.Inflorescences are non-branching cymes in S. lidii, while secondary branching occurs in S. vespertilio.The blue-purple corolla is weakly zygomorphic.Pollen is contained in yellow poricidal anthers borne on short filaments.The stamens are dimorphic, each flower having 3-4 stamens with short anthers on the dorsal side and one stamen with a long, slender anther on the ventral side of the flower.The style is filamentous and dimorphic (short or long), and deflected to the opposite side of the long stamen (enantiostyly) (unpubl.).
A small population of S. lidii consisting of four flowering plants was studied in Temisa, Gran Canaria (27°55'N, 15°30'W).Vegetation was a low xerophytic shrub.A population of S. vespertilio consisting of 20 plants was studied in the Anaga peninsula of Tenerife (28°32'N, 16°09'W).Vegetation was a shrub dominated by Erica arborea L., located at the borders of the laurel forest.
Pollen grains were analysed by SEM.Diameter of the tricolpate pollen grains was defined as the distance from one aperture to the opposite side through the centre of the pollen grain.Pollen diameter was measured in 10 pollen grains in each of the eight categories: short and long anthers of short-styled and long-styled flowers in each of the two species (Table 1).We tested female function of flowers by marking 49 short-styled flowers (six individuals) and 47 longstyled flowers (five individuals) of S. vespertilio.Fruit initiation was monitored two weeks later.Furthermore, we recorded numbers of long-styled and shortstyled flowers, flower buds and fruits and their spatial arrangement within inflorescences in 60 inflorescences of S. lidii and in 11 inflorescences of S. vespertilio.
Flower-visitors of S. lidii, were observed on February 10 th 2003, while S. vespertilio was observed on February 10 th , March 11-12 th  (Bombus terrestris canariensis Perez, two specimens) were analysed by SEM.

Reproductive system
Pollen grains were developed in both long-styled and short-styled flowers.We found no differences in structure or size of pollen grains of short-styled and longstyled flowers within species (Table 1).Furthermore, no differences were found between pollen from short and long anthers within flowers (Table 1).Whether pollen viability differs remains to be investigated.Of the 49 marked short-styled flowers of S. vespertilio, 45 had withered and showed no fruit initiation, while four could not be found two weeks after flowering.Of the 47 marked long-styled flowers, 40 (85%) showed fruit initiation, while seven had withered.Thus, both species were functionally andromonoecious, short-styled flowers being functionally male and long-styled flowers being functionally hermaphodite.Andromonoecy is common among species of the subgenus Leptostemonum (Symon, 1979;Whalen & Costrich, 1986).
In both Canarian Solanum species, flowers open in sequence, one or two at a time, from the proximal to the distal end of an inflorescence.Fruits were borne proximally on the inflorescences, followed by longstyled flowers, long-styled flower buds, short-styled flowers, short-styled flower buds and flower buds of indeterminable style length in sequence towards the distal end.This floral arrangement is typical of andromonoecious Solanum species (Whalen & Costrich, 1986).Andromonoecy combined with temporal segregation in flowering of male and hermaphrodite flowers may promote out-crossing (Coleman & Coleman, 1982;Anderson & Symon, 1989).Cross pollination may be further enhanced by enantiostyly, which facilitates pollen transfer between mirror-image flowers (Bowers, 1975;Dulberger, 1981) Anales del Jardín Botánico de Madrid 63( 1

Pollination
As in most other species of Solanum, only large insects capable of buzzing can remove pollen from the anthers and function as pollen vectors (Symon, 1979).For S. lidii, only one female individual of Amegilla Friese sp.(Apidae) was observed buzzing the flowers during the observation period.Small bees of the genus Lasioglossum Curtis (Halictidae) and small beetles (Meligethes Stephens sp.) were observed on the anthers, possibly searching for pollen.However, the latter two species did not buzz, nor did they touch the stigma in long-styled flowers, and thus they were not effective pollinators.
In S. vespertilio, the Canarian bumblebee, Bombus t. canariensis, was a regular visitor.The bumblebees were observed buzzing the flowers from early morning to late afternoon.Bombus t. canariensis was the only insect, which regularly visited the flowers of S. vespertilio (0.15-0.24 visits/flower/h).During a visit, bumblebees grasped the short stamens firmly with the mandibles and legs, while making a high-pitched sound.Bite marks were regularly seen on the short stamens of old flowers.Although bumblebees did not collect pollen from the long stamen, the apical pore of this anther and the stigma of long-styled flowers often touched the dorsal side of the bee during a visit.SEM analysis revealed that many Solanum pollen grains were deposited on both the upper and lower parts of the abdomen, while few adhered to the head.
Other less frequent visitors were Apis mellifera L. (0-0.04 visits/flower/h), Anthophora alluaudi Perez (Apidae) (0-0.01 visits/flower/h) and Lasioglossum chalcodes Brulle (Halictidae).The behaviour of Anthophora alluaudi on Solanum flowers was similar to that of B. t. canariensis.In contrast, Apis mellifera was unable to buzz the anthers, and did not touch the stigma of long-styled flowers.Lasioglossum chalcodes buzzed the short anthers, but was not sufficiently large to contact the tip of the long anther and stigma during a visit.Thus, the latter two species are poor pollen vectors.
The pollinator fauna of oceanic islands is usually less diverse compared to mainland areas.In particular, large-bodied, specialised flower-visitors are rare (Barrett, 1996).The small potential pollinator pool of the Canarian Solanum species may be due to the limited number of large bees capable of buzzing, which inhabit the islands.In comparison, Solanum rostratum Dunal and S. elaeagnifolium L. growing in a continental desert region (Arizona, USA) are visited frequently by several species of pollen collecting bees (eight and nine spp., respectively) (Linsley & Cazier, 1963).Solanum elaeagnifolium is a close relative of S. vespertilio (Bohs & Olmstead, 2001).
Although B. t. canariensis was the only frequent visitor of S. vespertilio, it is a reliable pollinator under various climatic conditions.On many occasions during fieldwork in the Canary Islands, we have observed active B. t. canariensis individuals.Weather conditions at which B. t. canariensis was foraging ranged from 9°C in rainy weather to 29°C in sunny and windy weather (Y.L. Dupont & C. Skov, unpubl. data).Furthermore, B. t. canariensis visits at least 88 different plant species (30 families) in the Canary Islands, and most likely is an important pollinator of many endemic plants (Hohmann & al., 1993).It may thus be regarded as a keystone pollinator species.
and May 31 st 2001, and March 15 th and 28 th 2003.Rates of visitation were monitored during three 1-h trials during the peak activity period of flower-visitors (10-15h) on March 15 th 2003.Pollen load of the most frequent flower-visitors 64 Y.L. Dupont & J.M. Olesen
): 63-66.January-June 2006.ISSN: 0211-1322 a Mann-Whitney U-test of difference in pollen diameter between long and short anthers within flowers.P > 0.05 = n.s.b Mann-Whitney U-test of difference in pollen diameter between flowers of different style length.P > 0.05 = n.s.