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Keywords:

  • coadaptation;
  • coevolution;
  • conflict;
  • Ficus;
  • fig wasp;
  • mutualism;
  • pollination

Abstract

  1. Top of page
  2. Abstract
  3. Introduction
  4. Methods
  5. Ficus sampling and floral sex ratio data
  6. Direct observations of pollination behaviour
  7. Phylogenies
  8. Mapping changes in pollination behaviour
  9. Correlated evolution between wasp behaviour and FSR
  10. Results
  11. Direct observations of behaviour
  12. Phylogeny and changes in pollination behaviour
  13. Correlated evolution of pollinator behaviour and FSR
  14. Discussion
  15. Evolution of pollination behaviour in Pleistodontes
  16. Correlated evolution of pollination behaviour and FSR
  17. Why is wasp pollination behaviour and fig FSR coevolving continuously?
  18. Acknowledgments
  19. References

Only three insect lineages have evolved complex active pollination behaviour and only fig wasps (Agaonidae) have also reverted from active to passive pollination. Previously, it was assumed that there was a single origin of active pollination in fig wasps, followed by one independent loss in each of five genera. We show here that there have been three to six changes in pollination behaviour within just one genus (Pleistodontes). The results suggest multiple gains of active pollination in fig wasps, but are sensitive to assumptions about the relative costs of gaining and losing this complex behaviour. In addition, previous comparative studies at higher taxonomic levels have reported correlated evolution between active pollination in wasps and low anther/ovule ratios in figs. We report that changes in pollination behaviour between congeneric species correlate perfectly with changes in anther/ovule ratios in the host figs, showing no phylogenetic inertia in coadaptation at the species level.


Introduction

  1. Top of page
  2. Abstract
  3. Introduction
  4. Methods
  5. Ficus sampling and floral sex ratio data
  6. Direct observations of pollination behaviour
  7. Phylogenies
  8. Mapping changes in pollination behaviour
  9. Correlated evolution between wasp behaviour and FSR
  10. Results
  11. Direct observations of behaviour
  12. Phylogeny and changes in pollination behaviour
  13. Correlated evolution of pollinator behaviour and FSR
  14. Discussion
  15. Evolution of pollination behaviour in Pleistodontes
  16. Correlated evolution of pollination behaviour and FSR
  17. Why is wasp pollination behaviour and fig FSR coevolving continuously?
  18. Acknowledgments
  19. References

Plants and their insect pollinators provide some classic examples of mutualism. Some insect-pollinated plant species have many pollinators, whereas others are associated with just a few, or even one, specialist pollinator species. There are many examples of plant characteristics (e.g. colour, smell, mimicry, nectar rewards) that attract insect pollinators, both specialist and/or generalist. However, there are very few examples of insect adaptations that act to improve pollination of the plants. The most striking case is active, or ethodynamic, pollination, in which insects have evolved specific behaviours for collecting and/or dispensing pollen. Active pollination has been reported to have evolved once in each of only three insect lineages –Yucca moths (Pellmyr, 1997), Senita moths (Fleming & Holland, 1998) and fig (Ficus) wasps (Ramirez, 1969; Frank, 1984; Kjellberg et al., 2001). Each case involves an intimate and highly species-specific mutualism between a host plant and an insect pollinator that is also its seed predator.

The fig/fig wasp interaction stands out from these examples as the only system in which some pollinator species are active, whereas others are passive. Active pollinators use a combination of specialized morphological (Ramirez, 1969) and behavioural adaptations (Frank, 1984) to transfer pollen between fig trees. Modified combs of hairs (corbiculae) on their fore coxae are used to gather pollen from within their natal fig inflorescence (syconium) and store it in thoracic pockets (Frank, 1984). On arrival at a new syconium they then use their forelegs to distribute pollen onto the stigmatic surfaces of receptive female flowers (Frank, 1984; Kjellberg et al., 2001). Wasps that pollinate passively lack coxal combs and do not load or deposit pollen actively (Kjellberg et al., 2001). Instead, they transmit pollen, which has adhered to their bodies prior to emergence from their natal syconia. Once outside the syconium the wasps groom themselves, but some pollen is trapped in folds in the integument when the gaster contracts on contact with drier external conditions (Galil & Neeman, 1977). When the wasps arrive at a new syconium, the pollen is either rubbed off, or falls off as the body expands again, amongst the receptive female flowers.

Intuitively, the efficacy of active pollination should allow figs to produce fewer male flowers and re-allocate resources to other functions (Galil & Eisikowitch, 1971). This prediction is supported by a wide survey across most genera of fig wasps showing that Ficus sp. with active pollinators have much lower anther/ovule ratios than those with passive pollinators (Kjellberg et al., 2001). The study also suggests that the presence of coxal combs always indicates active pollination behaviour, whereas pollen pockets provide a good, but imperfect, index. Kjellberg et al. (2001) did not control for phylogeny when testing for evolutionary correlations, but a subsequent study by Jousselin et al. (2003b), using a smaller species set and incorporating a Ficus molecular phylogeny, showed a strong association between active pollination and low anther/ovule ratios across wasps from many genera.

Active pollination may benefit wasps, because there is some evidence that pollination of a flower improves the nutrition or survival of the larva that it contains (Jousselin & Kjellberg, 2001), possibly by increasing the success of gall initiation (Jousselin et al., 2003a). However, there is presumably also a significant cost to active pollination, in terms of both the time and energy expended by ovipositing wasps (Cook & Rasplus, 2003). Fig wasps are probably generally time-limited as they live for only 1 or 2 days as adults (Compton et al., 1994) and, during this short time period, they must mate, collect pollen, emerge from their natal syconium, locate a receptive syconium and oviposit.

There are 20 genera of fig-pollinating wasps, of which 13 contain only active and two only passive pollinators, whereas the remaining five genera contain both active and passive species (Kjellberg et al., 2001). A wealth of evidence supports both the monophyly of fig-pollinating wasps (family Agaonidae) and a basal position for the exclusively passive pollinators in the genus Tetrapus (Wiebes, 1982; Boucek, 1988; Rasplus et al., 1998; Machado et al., 2001; Weiblen, 2001; Cook & Rasplus, 2003). These observations have been used to argue that active pollination evolved once, early in the radiation of fig wasps, and has been lost once (see Fig. 1.) in each of several different lineages (Kjellberg et al., 2001; Machado et al., 2001; Weiblen, 2001; Jousselin et al., 2003b). However, this hypothesis has not been tested by either statistical analysis of phylogenies, or by detailed investigation of the phylogenetic relationships between active and passive species in genera that include both types of pollinator.

image

Figure 1. A genus level reconstruction of pollinator phylogeny by Machado et al. (2001), using COI sequence data. Pollination mode at the genus level is shown as active (white), passive (black), or polymorphic (grey). Several Ceratosolen sp. were included to clarify the position of the passive ‘cuckoo’ species C. galili, which coexists with active pollinators.

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As active pollination is generally rare in nature, an initial hypothesis implying the minimum number of origins necessary to explain the distribution of this complex adaptive trait across fig wasp genera seems reasonable (Fig. 1). However, this hypothesis also requires explicit testing, especially in light of the coevolutionary selection pressures involved. The timescale of coadaptation is also worthy of consideration, as trait matching observed at higher taxonomic levels may not always occur at the species level. In this study, we focus on a single genus of wasps, Pleistodontes Saunders, that contains both active and passive pollinators (Wiebes, 1990, 1991, 1994; Ramirez & Malavasi, 1997; Lopez-Vaamonde et al., 2002) and address the following questions:

  • 1
    Is the general assumption of one change – a loss of active pollination – in behaviour in the genus Pleistodontes supported by existing evidence?
  • 2
    Do we observe coadapted trait matching of pollinator behaviour and host fig anther/ovule ratios even when changes occur between species within the same genus?

Ficus sampling and floral sex ratio data

  1. Top of page
  2. Abstract
  3. Introduction
  4. Methods
  5. Ficus sampling and floral sex ratio data
  6. Direct observations of pollination behaviour
  7. Phylogenies
  8. Mapping changes in pollination behaviour
  9. Correlated evolution between wasp behaviour and FSR
  10. Results
  11. Direct observations of behaviour
  12. Phylogeny and changes in pollination behaviour
  13. Correlated evolution of pollinator behaviour and FSR
  14. Discussion
  15. Evolution of pollination behaviour in Pleistodontes
  16. Correlated evolution of pollination behaviour and FSR
  17. Why is wasp pollination behaviour and fig FSR coevolving continuously?
  18. Acknowledgments
  19. References

Ficus is a large genus containing >750 species and divided into 20 intra-generic sections (Berg, 1989). Section Malvanthera is endemic to Australasia and contains 22 species (Dixon, 2002), each of which is pollinated by one (or, in a few cases, two) wasp species in the genus Pleistodontes (Dixon, 2002; Lopez-Vaamonde et al., 2002). All malvantheran fig species are monoecious, with growth forms ranging from rainforest hemi-epiphytes to lithophytic deciduous shrubs (Dixon, 2002). Monoecious figs have many unisexual flowers of each sex in each syconium (fig inflorescence) and we estimated floral sex ratios (FSR), defined here as the proportion of flowers that are male (i.e. anthers/anthers + ovules), by counting and sexing all flowers in three to six syconia for each of 14 malvantheran species (Table 1). All syconia were collected from trees growing within their natural ranges in different parts of Australia and Papua New Guinea.

Table 1.  Flower sex ratios (FSR) of Ficus sp. and correlated morphology of their pollinators.
Ficus sp.Syconia (trees)FSRPleistodontes sp.Coxal combs*Pollen pockets*Mode
  1. *Wasp morphological characters are from Lopez-Vaamonde et al. (2002) and Wiebes (1994).

  2. Ficus obliqua is pollinated by different wasp species in northern (P. xanthocephalus) and southern (P. greenwoodi) Australia (Lopez-Vaamonde et al., 2002).

  3. ‡Data for outgroup species from Kjellberg et al. (2001).

brachypoda4 (1)0.077macrocainusPresentLargeActive
crassipes5 (3)0.453addicottiAbsentSmallPassive
destruens6 (3)0.246rigisamosAbsentSmallPassive
hesperidiformis3 (1)0.256plebejusAbsentAbsentPassive
lilliputiana6 (2)0.088proximusPresentLargeActive
macrophylla6 (3)0.248froggattiAbsentLargePassive
obliqua5 (1)0.109greenwoodiPresentLargeActive
obliqua6 (3)0.134xanthocephalusPresentLargeActive
platypoda4 (2)0.153cuneatusPresentLargeActive
pleurocarpa6 (2)0.533regalisAbsentSmallPassive
rubiginosa6 (3)0.125imperialisPresentLargeActive
triradiata3 (1)0.315schizodontusAbsentSmallPassive
watkinsiana6 (4)0.399nigriventrisAbsentSmallPassive
xylosycia5 (1)0.332rieckiAbsentAbsentPassive
ingens0.040Platyscapa sorariaPresentPresentActive

Although Kjellberg et al. (2001) showed, for three distantly related fig species from other Ficus sections, that sampling just two syconia gives a >95% chance of placing the species into the correct (high or low) FSR category, where possible we sampled six syconia per species from different trees and different sites. We also conducted our own investigation of within species variability in FSR by collecting data from a much larger sample (approximately 30 syconia), sourced from several trees and localities, for one actively pollinated and one passively pollinated malvantheran fig species.

Direct observations of pollination behaviour

  1. Top of page
  2. Abstract
  3. Introduction
  4. Methods
  5. Ficus sampling and floral sex ratio data
  6. Direct observations of pollination behaviour
  7. Phylogenies
  8. Mapping changes in pollination behaviour
  9. Correlated evolution between wasp behaviour and FSR
  10. Results
  11. Direct observations of behaviour
  12. Phylogeny and changes in pollination behaviour
  13. Correlated evolution of pollinator behaviour and FSR
  14. Discussion
  15. Evolution of pollination behaviour in Pleistodontes
  16. Correlated evolution of pollination behaviour and FSR
  17. Why is wasp pollination behaviour and fig FSR coevolving continuously?
  18. Acknowledgments
  19. References

Direct observations are very difficult for many species of pollinating wasps, due to the problems inherent in obtaining receptive syconia (short time window) containing live foundress wasps (24–48 h lifespan), especially from rainforest canopy Ficus sp. In addition, foundresses of some species have been found to leave syconia when the latter are opened (J.M. Cook, personal observation). In other species, foundresses remain and continue to oviposit and pollinate in an apparently normal manner. In such cases, individual females may continue oviposition and pollination for up to 12 hours (J.M. Cook & S.A. Power, personal observation), as long as the syconium is not allowed to dessicate.

Kjellberg et al. (2001) observed pollen deposition behaviour for several species from various fig wasp genera and identified a morphological trait (presence of coxal combs) that always indicated active pollination. This trait is easy to observe and allows inference of behaviour for a much wider set of species than can be observed in the act of pollination. The presence of pollen pockets also generally indicates active pollination, but there are exceptions involving passive species in a genus that is primarily active (Kjellberg et al., 2001). The genus Pleistodontes contains the only species highlighted as a possible exception to the striking general pattern of fig-pollinator coadaptation (Kjellberg et al., 2001; Jousselin et al., 2003b). Ficus macrophylla Desf. ex Pers. has a high anther/ovule ratio, typical of a passively pollinated fig. However, its pollinator, Pleistodontes froggatti Mayr, although lacking coxal combs (suggesting passive pollination), has pollen pockets that can contain concentrated pollen (suggesting active pollination). On balance, this species has been considered an active pollinator in two previous studies (Kjellberg et al., 2001; Jousselin et al., 2003b).

Where possible, and especially for the controversial P. froggatti, we made direct observations of the pollen deposition behaviour of live foundresses. Receptive figs were collected, sliced open along the long axis, and enclosed between the two halves of a snap-top Petri dish. We then observed the wasps under a binocular microscope at 20× magnification. For the other species, we inferred active/passive behaviour from the presence/absence of coxal combs (Table 1). Assigning Pleistodontes sp. (except P. froggatti) as active or passive is very easy as each species has either two diagnostic features (large pollen pockets plus coxal combs) of active pollinators or two diagnostic features (small or absent pollen pockets with no coxal combs) of passive pollinators (see Table 1, Lopez-Vaamonde et al., 2002).

Phylogenies

  1. Top of page
  2. Abstract
  3. Introduction
  4. Methods
  5. Ficus sampling and floral sex ratio data
  6. Direct observations of pollination behaviour
  7. Phylogenies
  8. Mapping changes in pollination behaviour
  9. Correlated evolution between wasp behaviour and FSR
  10. Results
  11. Direct observations of behaviour
  12. Phylogeny and changes in pollination behaviour
  13. Correlated evolution of pollinator behaviour and FSR
  14. Discussion
  15. Evolution of pollination behaviour in Pleistodontes
  16. Correlated evolution of pollination behaviour and FSR
  17. Why is wasp pollination behaviour and fig FSR coevolving continuously?
  18. Acknowledgments
  19. References

We estimated phylogenies using sequence data from the nuclear 28S rDNA (28S) and internal transcribed spacer rDNA (ITS2) regions. These data were generated and aligned by Lopez-Vaamonde et al. (2001) for a study of cospeciation in pollinating and nonpollinating fig wasps and the alignment is available from TreeBASE, http://www.herbaria.harvard.edu/treebase/console.html).

Pleistodontes is one of several genera of wasps that pollinate monoecious figs in the Ficus subgenus Urostigma and which appear to form a monophyletic group (Machado et al., 2001; Weiblen, 2001), separated from the pollinators of dioecious figs. Following Lopez-Vaamonde et al. (2001), we used Platyscapa soraria Wiebes, 1980 (pollinator of another fig from subgenus Urostigma) as the outgroup.

We generated a Bayesian estimate of phylogeny, using MrBayes 2.01 (Huelsenbeck & Ronquist, 2001) and specifying the general time reversible (GTR) model of sequence evolution, with gamma rate variation between sites, which is the recommended substitution model for noncoding DNA (Hall, 2001; Huelsenbeck & Ronquist, 2001). We ran the program for 1 million generations, sampling a tree every 1000 generations and then calculated a 50% majority rule consensus tree in paup4.0b10 (Swofford, 2002). We excluded the first 200 trees, which represented the ‘burn-in’ period before the likelihood value stabilized. The proportion of sampled trees that contain a given node provides the posterior probability of that node, which is a measure of its support, given the model and data supplied.

We also generated a maximum parsimony (MP) phylogeny, using paup in a heuristic search, involving 1000 random additions and TBR branch-swapping and treating gaps as a fifth character state. We then carried out nonparametric bootstrapping (1000 replicates) to assess support for nodes in the MP analysis.

Mapping changes in pollination behaviour

  1. Top of page
  2. Abstract
  3. Introduction
  4. Methods
  5. Ficus sampling and floral sex ratio data
  6. Direct observations of pollination behaviour
  7. Phylogenies
  8. Mapping changes in pollination behaviour
  9. Correlated evolution between wasp behaviour and FSR
  10. Results
  11. Direct observations of behaviour
  12. Phylogeny and changes in pollination behaviour
  13. Correlated evolution of pollinator behaviour and FSR
  14. Discussion
  15. Evolution of pollination behaviour in Pleistodontes
  16. Correlated evolution of pollination behaviour and FSR
  17. Why is wasp pollination behaviour and fig FSR coevolving continuously?
  18. Acknowledgments
  19. References

We mapped pollination behaviour onto phylogenies and analysed the pattern of changes using parsimony techniques in MacClade 4 (Maddison & Maddison, 2000). We first treated behaviour as an unordered, binary (active/passive) character and weighted gains and losses of active pollination equally in seeking the most parsimonious reconstruction (MPR) of changes. This first analysis implied at least one gain of active pollination within the genus, which contrasts with the existing view that active pollination has arisen only once in fig wasps overall (Kjellberg et al., 2001; Machado et al., 2001; Jousselin et al., 2003b), so we conducted a further analysis. In the second analysis we set the weight of losses to one and then progressively increased the weight of gains until the MPR involved only losses, as implied by the conventional view of the evolution of pollination behaviour (Kjellberg et al., 2001; Machado et al., 2001; Jousselin et al., 2003b).

We also took a second approach to investigate support for a gain of active pollination within Pleistodontes. This involved generating phylogenies (as described above), but subject to the constraint that all the active species (including the outgroup) formed a single, monophyletic clade. In other words, the constrained tree represents the conventional hypothesis of a single loss of active pollination in Pleistodontes. We then compared the length or likelihood of this constrained tree with that of the corresponding unconstrained tree, using a Kishino & Hasegawa (1989) test of length differences for MP trees and a log likelihood comparison for Bayesian trees.

Correlated evolution between wasp behaviour and FSR

  1. Top of page
  2. Abstract
  3. Introduction
  4. Methods
  5. Ficus sampling and floral sex ratio data
  6. Direct observations of pollination behaviour
  7. Phylogenies
  8. Mapping changes in pollination behaviour
  9. Correlated evolution between wasp behaviour and FSR
  10. Results
  11. Direct observations of behaviour
  12. Phylogeny and changes in pollination behaviour
  13. Correlated evolution of pollinator behaviour and FSR
  14. Discussion
  15. Evolution of pollination behaviour in Pleistodontes
  16. Correlated evolution of pollination behaviour and FSR
  17. Why is wasp pollination behaviour and fig FSR coevolving continuously?
  18. Acknowledgments
  19. References

We tested whether active pollination results in a more female-biased FSR both with and without taking phylogeny into account. We treated pollination as a binary (active/passive) character and FSR as a continuous variable (see Table 1 for data). For the phylogeny-free analysis, we compared the FSRs of active and passive species using a nonparametric Wilcoxon signed ranks test. For the phylogenetic comparative analysis, we used the ‘brunch’ option in caic 2.0.0 (Purvis & Rambaut, 1995) to calculate four independent contrasts for FSR. If FSR is independent of pollination mode, we would expect equal numbers of contrasts to be positive and negative with a mean of zero. In order to test this hypothesis we thus compared the mean of the contrasts to zero using a t-test.

Direct observations of behaviour

  1. Top of page
  2. Abstract
  3. Introduction
  4. Methods
  5. Ficus sampling and floral sex ratio data
  6. Direct observations of pollination behaviour
  7. Phylogenies
  8. Mapping changes in pollination behaviour
  9. Correlated evolution between wasp behaviour and FSR
  10. Results
  11. Direct observations of behaviour
  12. Phylogeny and changes in pollination behaviour
  13. Correlated evolution of pollinator behaviour and FSR
  14. Discussion
  15. Evolution of pollination behaviour in Pleistodontes
  16. Correlated evolution of pollination behaviour and FSR
  17. Why is wasp pollination behaviour and fig FSR coevolving continuously?
  18. Acknowledgments
  19. References

We were able to make direct observations of pollination behaviour, which was clearly bimodal, for four species. Each species was observed in at least two localities. If there was a single wasp present, it was observed for at least 20 oviposition events, whereas multiple foundresses were observed as a group for 30 min. Pleistodontes imperialis Saunders (n = 8 wasps) and P. xanthocephalus Lopez-Vaamonde, Dixon, Cook & Rasplus (n = 3) foundresses pollinate all, or nearly all, flowers in which they oviposit by actively transferring pollen from their pollen pockets using their forelegs, as described in some Pegoscapus sp. (Frank, 1984). The movements are very obvious to the observer and occur at the end of an oviposition sequence for a single flower. In contrast, P. addicotti Wiebes (n = 2) and P. froggatti (n = 21) do not deposit pollen actively. Given that P. froggatti was classified previously as an active pollinator, we observed 21 different individuals (in both single and multiple foundress figs) from two different localities (Melbourne and Brisbane) for a total of 10 h, but did not observe any active pollination. In addition, we did not observe any aggressive interference between multiple foundresses in the same syconium (see Bronstein et al., 1998).

Phylogeny and changes in pollination behaviour

  1. Top of page
  2. Abstract
  3. Introduction
  4. Methods
  5. Ficus sampling and floral sex ratio data
  6. Direct observations of pollination behaviour
  7. Phylogenies
  8. Mapping changes in pollination behaviour
  9. Correlated evolution between wasp behaviour and FSR
  10. Results
  11. Direct observations of behaviour
  12. Phylogeny and changes in pollination behaviour
  13. Correlated evolution of pollinator behaviour and FSR
  14. Discussion
  15. Evolution of pollination behaviour in Pleistodontes
  16. Correlated evolution of pollination behaviour and FSR
  17. Why is wasp pollination behaviour and fig FSR coevolving continuously?
  18. Acknowledgments
  19. References

Bayesian and MP analyses led to very similar estimates of phylogeny. In fact, the topology of the strict consensus of the two MPTs was identical to that of the 50% majority-rule consensus Bayesian tree and this tree (used for all mappings) is shown in Fig. 2. Many nodes received strong support with both methods, whereas those with weak support from one method also had weak support from the other. In the first reconstruction, with gains and losses weighted equally, the minimum number of changes is four. There is more than one MPR (Fig. 2a), but each involves at least one gain of active pollination. In addition, the ancestral state for Pleistodontes is implied to be passive, not active, pollination. In the second analysis, we progressively increased the weights of gains relative to losses until the MPR involved only gains. This was not achieved until gains were more than twice as costly as losses and increased the implied number of changes to six losses (Fig. 2b).

image

Figure 2. Parsimony reconstruction of changes (*) between active (white) and passive (black) pollination behaviour. In (a) gains and losses of active pollination are assigned equal costs, whereas in (b) the cost of one gain is more than twice the cost of one loss. Numbers indicate support (when >50%) for nodes in the phylogeny in Bayesian (above line = posterior probability) and maximum parsimony analyses (below line = bootstrap value).

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Our alternative approach was to constrain phylogenies such that they were consistent with just a single loss of active pollination in Pleistodontes. In both MP and Bayesian approaches the phylogenies so produced were significantly worse than the unconstrained phylogenies that imply multiple changes in behaviour. The constrained MP tree (1589 steps) was significantly longer (t1 = 6.44, P < 0.0001) than the unconstrained MP tree (1504 steps), whereas the constrained Bayesian tree (log likelihood = 6825.31) was significantly (χ2 = 26.54, d.f = 1, P < 0.001) less likely than the unconstrained one (log likelihood = −6798.76).

Correlated evolution of pollinator behaviour and FSR

  1. Top of page
  2. Abstract
  3. Introduction
  4. Methods
  5. Ficus sampling and floral sex ratio data
  6. Direct observations of pollination behaviour
  7. Phylogenies
  8. Mapping changes in pollination behaviour
  9. Correlated evolution between wasp behaviour and FSR
  10. Results
  11. Direct observations of behaviour
  12. Phylogeny and changes in pollination behaviour
  13. Correlated evolution of pollinator behaviour and FSR
  14. Discussion
  15. Evolution of pollination behaviour in Pleistodontes
  16. Correlated evolution of pollination behaviour and FSR
  17. Why is wasp pollination behaviour and fig FSR coevolving continuously?
  18. Acknowledgments
  19. References

In the phylogeny-free analysis, FSR was significantly lower (less anthers) in actively pollinated figs (Wilcoxon test, one-tailed: W1 = 0, P < 0.0005) (Fig. 3a). This difference is further emphasized by the bimodal pattern found when the distribution of species means for FSR is plotted for active and passive species (Fig. 3b). When taking phylogeny into account, we were able to make four independent contrasts, which again showed that FSR was significantly lower in actively pollinated figs (t-test, one-tailed: t4 = 3.1949, P < 0.05).

image

Figure 3. Correlated evolution of pollination behaviour and flower sex ratios. (a) Mean floral sex ratio (FSR) for active (white) and passive (black) species (bars show 1 SE). (b) Distribution of mean FSR for active (white) and passive (black) species.

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Finally, our assessment of within species variation revealed that the one actively pollinated (F. obliqua) and one passively pollinated (F. macrophylla) fig species have abutting, but nonoverlapping, FSR distributions (Fig. 4). Although it would be possible, but highly unlikely, for a very small sample of syconia to yield a mean in the tail of one of the distributions, this would not lie in the ‘wrong’ category, and would give a borderline value, demanding further investigation (see Kjellberg et al., 2001).

image

Figure 4. Within-species variability in floral sex ratios (proportion male flowers). White bars show values for 36 Ficus obliqua syconia (active pollinator) and black bars for 26 F. macrophylla syconia (passive pollinator).

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Evolution of pollination behaviour in Pleistodontes

  1. Top of page
  2. Abstract
  3. Introduction
  4. Methods
  5. Ficus sampling and floral sex ratio data
  6. Direct observations of pollination behaviour
  7. Phylogenies
  8. Mapping changes in pollination behaviour
  9. Correlated evolution between wasp behaviour and FSR
  10. Results
  11. Direct observations of behaviour
  12. Phylogeny and changes in pollination behaviour
  13. Correlated evolution of pollinator behaviour and FSR
  14. Discussion
  15. Evolution of pollination behaviour in Pleistodontes
  16. Correlated evolution of pollination behaviour and FSR
  17. Why is wasp pollination behaviour and fig FSR coevolving continuously?
  18. Acknowledgments
  19. References

This is the first study to combine a molecular phylogeny of many species from a single fig wasp genus with information on their (variable) pollination behaviour. Two recent studies at higher taxonomic levels have either not explicitly involved phylogeny (Kjellberg et al., 2001), or mapped traits onto a Ficus phylogeny (Jousselin et al., 2003b). Consequently, our study extends previous work both by investigating a lower taxonomic level and by utilizing a wasp rather than a fig phylogeny. Previous studies hypothesized a single origin of active pollination, followed by at least five independent losses, with only one of these in Pleistodontes (Fig. 1). A major result of our study is evidence for multiple changes in pollination behaviour within just one genus (Pleistodontes) of fig wasps. Furthermore, character mapping suggests an ancestral state of passive pollination, with at least one gain of active pollination during the radiation of the genus. These results have major implications for our view of the evolution of pollination behaviour across fig wasps as a whole (see Fig. 1) and suggest two possible scenarios:

  • 1
    Active pollination did not arise until after the origin of Pleistodontes, and then arose independently in Pleistodontes and in other fig wasps. Under this scenario there are multiple independent origins of active pollination.
  • 2
    Active pollination arose after the divergence of the basal genus Tetrapus, but before the origin of Pleistodontes. Active pollination was then lost in the Pleistodontes lineage and subsequently regained (and possibly lost and/or regained again). Under this scenario there was a single origin of active pollination, but it was lost and regained in Pleistodontes.

Distinguishing between these two scenarios requires well-resolved and densely sampled phylogenies and pollination data for fig wasp species from many genera. Inclusion of further basal Pleistodontes sp. from New Guinea is likely to provide further support for passive pollination as the ancestral behaviour in this genus because all species known from New Guinea lack coxal combs and have small or no pollen pockets (Wiebes, 1991, 1994; Lopez-Vaamonde et al., 2002; J.-Y. Rasplus, personal communication). Unfortunately, the species delimitations and host records of these New Guinean wasps (some known only from light traps) are not yet complete. However, we stress that either scenario above implies that pollination behaviour has changed from passive to active on at least two occasions in fig wasps. This is noteworthy as it suggests that the loss of an unusual key trait (active pollination) is reversible, a possibility that has been largely overlooked in previous studies (Machado et al., 2001; Jousselin et al., 2003b), although Kjellberg et al. (2001) did mention that the genera Wiebesia and Pleistodontes were potential candidates for recent acquisition of active pollination. Furthermore, investigation of the behaviour and phylogeny of other fig wasp genera that contain both active and passive pollinators may reveal evidence for further changes in one or both directions.

Ancestral character states and patterns of change are difficult to estimate for algorithmic reasons (Cunningham et al., 1998; Webster & Purvis, 2001). Nevertheless, exploring different reconstruction options, such as weighting of changes, can be highly instructive (e.g. Omland, 1997; Belshaw & Quicke, 2002). A major issue here is the weighting of gains and losses of active pollination. Assigning equal weights suggests that changes in both directions occur within Pleistodontes; however, if gains are more than twice as costly as losses, the MPR involves only losses (but six independent ones). The origin of a complex trait with several components – coxal corbiculae, pollen pockets, pollen-loading and pollen deposition – seems less likely than its loss, which requires loss of only one component. Consequently, we might decide to weight gains more heavily, but it is impossible to know the appropriate difference in cost. Nonetheless, it should also be noted that pollen pockets are sometimes retained in passively pollinating members of predominantly actively pollinating clades (Kjellberg et al., 2001), so re-gain of active pollination may be easier than its de novo origination. Previous studies that have mapped pollination mode onto wasp or fig phylogenies (Machado et al., 2001; Jousselin et al., 2003b) have not considered differential weighting of gains and losses, but this has a major impact on conclusions and deserves greater consideration.

Our reconstruction of changes in pollination behaviour also depends on the accuracy of the estimate of phylogeny. For this reason, it would be valuable to revisit this issue in the future using wasp phylogenies generated from sequences of other DNA regions. In addition, it would be particularly interesting to compare results with the changes suggested by mapping characters onto the corresponding Ficus phylogeny, should this become available. However, the current data clearly reject the hypothesis of a single loss of active pollination in Pleistodontes.

Correlated evolution of pollination behaviour and FSR

  1. Top of page
  2. Abstract
  3. Introduction
  4. Methods
  5. Ficus sampling and floral sex ratio data
  6. Direct observations of pollination behaviour
  7. Phylogenies
  8. Mapping changes in pollination behaviour
  9. Correlated evolution between wasp behaviour and FSR
  10. Results
  11. Direct observations of behaviour
  12. Phylogeny and changes in pollination behaviour
  13. Correlated evolution of pollinator behaviour and FSR
  14. Discussion
  15. Evolution of pollination behaviour in Pleistodontes
  16. Correlated evolution of pollination behaviour and FSR
  17. Why is wasp pollination behaviour and fig FSR coevolving continuously?
  18. Acknowledgments
  19. References

The correlation between low anther/ovule ratio and active pollination observed between distantly related fig wasp species (Kjellberg et al., 2001) is also observed within a single genus of wasps (Pleistodontes), in which there have been multiple changes in behaviour. This strengthens the case for a very strong co-evolutionary link between the two characters, as well as the absence of major phylogenetic inertia which might disrupt correlated evolution in the face of one partner changing state frequently. The rapid turnover of FSR within a section of figs and correlated turnover in pollination behaviour in the associated genus of wasps further emphasizes the belief that figs and wasps impose key reciprocal selection pressures upon each other and evolve in response to these pressures. Although parasite virulence and host resistance provide a general example of coevolving traits, there are few other pairs of traits for which there is a clear basis for reciprocal selection (Clayton et al., 1999). Figs and their pollinators provide an excellent model for coevolutionary studies, because there are several such pairs of traits (others include fig breeding system and wasp ovipositor length –Jousselin et al., 2003b).

Kjellberg et al. (2001) found an almost perfect match between active pollination by wasps and low FSRs in figs and considered P. froggatti/F. macrophylla to be the only clear exception. However, we found that this wasp species does not pollinate actively and so the high FSR of the host fig is appropriate for its passive pollination. Pleistodontes froggatti has no coxal combs, but does have large pollen pockets. In this respect, it is like members of certain other fig wasp genera, in which some passive pollinators retain pollen pockets, despite lacking pollination behaviour (Kjellberg et al., 2001). These might contain pollen because of the dramatic dehiscence of anthers in some passively pollinated figs like F. macrophylla.

The case of P. froggatti is instructive, because it also emphasizes that active pollination is a syndrome with several identifiable components. Wasps must: (i) collect pollen, (ii) place it into pockets and (iii) deposit it onto receptive flowers, and to do this they require coxal combs and pollen repositories. Loss of any one of the behaviours (i)–(iii) removes active pollination. Given the previous report of pollen in P. froggatti pollen pockets (Kjellberg et al., 2001), it is possible that this species (or some individuals) retain pollen collection behaviour, although we have not observed this to be the case. Whereas the discrete division between active and passive pollinators (mirrored by the striking bimodal FSR distribution) is clearly the major variation in pollination behaviour, there may be further minor variation within the ‘passive’ and ‘active’ categories (e.g. see Ramirez & Malavasi, 1997). In this study, the range of mean species FSR values was low (0.08–0.15) in figs that are pollinated actively, but higher (0.25–0.53) in those which are pollinated passively. With more detailed knowledge of wasp behaviour and morphology, we may be able to provide adaptive explanations not only for the bimodality of FSR, but also for some of the variation within each category. A further possibility is that foundress behaviour may change with time in the syconium, such that pollination is active early on and/or with few competitors, but passive late in life due to changes in the opportunity costs of pollination, or even exhaustion of pollen in the pollen pockets. Finally, it would also be instructive to determine whether F. macrophylla (and other malvantheran species) has a synstigmatic platform in its receptive syconia. This structure is generally associated with active pollination, whereas passively pollinated figs have individualized female flowers (Jousselin et al., 2003b). However, it is possible that changes in stigma structure, which mostly affect seed set, might respond to a change in pollination mode more, or less, quickly than changes in FSR, which mostly affect pollen export.

Why is wasp pollination behaviour and fig FSR coevolving continuously?

  1. Top of page
  2. Abstract
  3. Introduction
  4. Methods
  5. Ficus sampling and floral sex ratio data
  6. Direct observations of pollination behaviour
  7. Phylogenies
  8. Mapping changes in pollination behaviour
  9. Correlated evolution between wasp behaviour and FSR
  10. Results
  11. Direct observations of behaviour
  12. Phylogeny and changes in pollination behaviour
  13. Correlated evolution of pollinator behaviour and FSR
  14. Discussion
  15. Evolution of pollination behaviour in Pleistodontes
  16. Correlated evolution of pollination behaviour and FSR
  17. Why is wasp pollination behaviour and fig FSR coevolving continuously?
  18. Acknowledgments
  19. References

Previous studies suggested at least six changes in pollination behaviour, with only one of these in Pleistodontes. Our study raises this estimate to at least nine and investigation of other polymorphic genera may well raise this estimate further. This continued turnover of coadapted traits emphasizes that coevolution does not reach an ‘endpoint’ and that selection pressures on mutualists are not constant. Further understanding will come from closer investigation of the costs and benefits. Existing evidence suggests that active pollinators tend to pollinate flowers in which they lay their eggs, whereas the eggs of passive pollinators are often in unpollinated flowers (Jousselin et al., 2001, 2003a; Jousselin & Kjellberg, 2001). There is also evidence that the offspring of active pollinators gain fitness benefits from developing in pollinated flowers (Jousselin et al., 2003a). We need to identify factors that influence the relative costs and benefits of active and passive pollination for wasps and for figs. For example, Jousselin et al. (2003b) suggested that loss of active pollination in wasps may be selected for if figs reroute pollen tube growth so that wasp larvae do not gain the benefit of developing in actively pollinated flowers. Intuitively, active pollination appears beneficial to figs, but this may not be true if too many of the actively pollinated flowers also receive a wasp egg (Jousselin et al., 2003b). The repeated correlated evolution of wasp behaviour and fig anther/ovule ratios is clear and probably stems from underlying reproductive tensions and changes in the costs and benefits for both wasps and figs. Our view of this mutualism and its diversity must increasingly take account of the reciprocal selection pressures caused by these reproductive tensions (Herre, 1989; Anstett et al., 1997; Herre & West, 1997; Cook & Rasplus, 2003).

References

  1. Top of page
  2. Abstract
  3. Introduction
  4. Methods
  5. Ficus sampling and floral sex ratio data
  6. Direct observations of pollination behaviour
  7. Phylogenies
  8. Mapping changes in pollination behaviour
  9. Correlated evolution between wasp behaviour and FSR
  10. Results
  11. Direct observations of behaviour
  12. Phylogeny and changes in pollination behaviour
  13. Correlated evolution of pollinator behaviour and FSR
  14. Discussion
  15. Evolution of pollination behaviour in Pleistodontes
  16. Correlated evolution of pollination behaviour and FSR
  17. Why is wasp pollination behaviour and fig FSR coevolving continuously?
  18. Acknowledgments
  19. References
  • Anstett, M.C., Hossaert McKey, M. & Kjellberg, F. 1997. Figs and fig pollinators: evolutionary conflicts in a coevolved mutualism. Trends Ecol. Evol. 0: 9499.
  • Belshaw, R. & Quicke, D.L.J. 2002. Robustness of ancestral state estimates: evolution of life history strategy in ichneumonoid parasitoids. Syst. Biol. 51: 450477.
  • Berg, C.C. 1989. Classification and distribution of Ficus. Experientia 45: 605611.
  • Boucek, Z. 1988. Australasian Chalcidoidea (Hymenoptera). CAB International, Wallingford, UK.
  • Bronstein, J.L., Vernet, D. & Hossaert-McKey, M. 1998. Do fig wasps interfere with each other during oviposition? Entomol. Exp. Appl. 87: 321324.
  • Clayton, D.H., Lee, P.L.M., Tompkins, D.M. & Brodie, E.D. 1999. Reciprocal natural selection on host–parasite phenotypes. Am. Nat. 154: 261270.
  • Compton, S.G., Rasplus, J.Y. & Ware, A.B. 1994. African fig wasp parasitoid communities. In: Parasitoid Community Ecology (B.Hawkins & W.Sheehan, eds), pp. 323348. Oxford University Press, Oxford.
  • Cook, J.M. & Rasplus, J.Y. 2003. Mutualists with attitude: coevolving fig wasps and figs. Trends Ecol. Evol. 18: 241248.
  • Cunningham, C., Omland, K.E. & Oakley, T.H. 1998. Reconstructing ancestral character states: a critical re-appraisal. Trends Ecol. Evol. 13: 361366.
  • Dixon, D.J. 2002. A taxonomic revision of the Australian Ficus species in the section Malvanthera Ficus subg. Urostigma: Moraceae). Telopea 10: 125153.
  • Fleming, T.H. & Holland, J.N. 1998. The evolution of obligate pollination mutualisms: senita cactus and senita moth. Oecologia 114: 368375.
  • Frank, S.A. 1984. The behaviour and morphology of the fig wasps Pegoscapus assuetus and P. jimenezi: descriptions and suggested behavioural characters for phylogenetic studies. Psyche 91: 289308.
  • Galil, J. & Eisikowitch, D. 1971. Studies on the mutualistic symbiosis between syconia and sycophilous wasps in monoecious figs. New Phytol. 70: 773787.
  • Galil, J. & Neeman, G. 1977. Pollen transfer and pollination in the common fig (Ficus carica L.). New Phytol. 79: 163171.
  • Hall, B.G. 2001. Phylogenetic Trees Made Easy. Sinauer, Sunderland, MA, USA.
  • Herre, E.A. 1989. Coevolution of reproductive characteristics in 12 species of New World figs and their pollinator wasps. Experientia 45: 637647.
  • Herre, E.A. & West, S.A. 1997. Conflict of interest in a mutualism: documenting the elusive fig wasp seed trade-off. Proc. R. Soc. Lond. B. 264: 15011507.
  • Huelsenbeck, J.P. & Ronquist, F. 2001. MRBAYES: bayesian inference of phylogenetic trees. Biometrics 17: 754755.
  • Jousselin, E. & Kjellberg, F. 2001. The functional implications of active and passive pollination in dioecious figs. Ecol. Lett. 4: 151158.
  • Jousselin, E., Hossaert-McKey, M., Vernet, D. & Kjellberg, F. 2001. Egg deposition patterns of fig pollinating wasps: implications for studies on the stability of the mutualism. Ecol. Entomol. 26: 602608.
  • Jousselin, E., Hossaert-McKey, M., Herre, E.A. & Kjellberg, F. 2003a. Why do fig wasps actively pollinate monoecious figs? Oecologia 134: 381387.
  • Jousselin, E., Rasplus, J.Y. & Kjellberg, F. 2003b. Convergence and coevolution in a mutualism: evidence from a molecular phylogeny of Ficus. Evolution 57: 12551269.
  • Kishino, H. & Hasegawa, M. 1989. Evaluation of the maximum likelihood estimate of the evolutionary tree topologies from DNA sequence data and the branching order in Hominoidea. J. Mol. Evol. 29: 170179.
  • Kjellberg, F., Jousselin, E., Bronstein, J.L., Patel, A., Yokoyama, J. & Rasplus, J.Y. 2001. Pollination mode in fig wasps: the predictive power of correlated traits. Proc. R. Soc. Lond. B. 268: 11131121.
  • Lopez-Vaamonde, C., Rasplus, J.Y., Weiblen, G.D. & Cook, J.M. 2001. Molecular phylogenies of fig wasps: partial co-cladogenesis of pollinators and parasites. Mol. Phylogenet. Evol. 21: 5571.
  • Lopez-Vaamonde, C., Dixon, D., Cook, J.M. & Rasplus, J.Y. 2002. Revision of the Australian species of Pleistodontes (Hymenoptera: Agaonidae) fig-pollinating wasps and their host plant affiliations. Zool. J. Linn. Soc. 136: 637683.
  • Machado, C.A., Jousselin, E., Kjellberg, F., Compton, S.G. & Herre, E.A. 2001. Phylogenetic relationships, historical biogeography and character evolution of fig-pollinating wasps. Proc. R. Soc. Lond. B. 268: 685694.
  • Maddison, D.R. & Maddison, W.P. 2000. MacClade. Sinauer, Sunderland, MA, USA.
  • Omland, K.E. 1997. Examining two standard assumptions of ancestral reconstructions: repeated loss of dichromatism in dabbling ducks (Anatini). Evolution 51: 16361646.
  • Pellmyr, O. 1997. Pollinating seed eaters: why is active pollination so rare? Ecology 78: 16551660.
  • Purvis, A. & Rambaut, A. 1995. Comparative analysis by independent contrasts (CAIC) – an Apple-Macintosh application for analyzing comparative data. Comp. Applic. Biosci. 11: 247251.
  • Ramirez, B.W. 1969. Fig wasps: mechanism of pollen transfer. Science 163: 580581.
  • Ramirez, W. & Malavasi, J. 1997. Fig wasps: mechanisms of pollen transfer in Malvanthera and Pharmacosycea figs (Moraceae). Revista De Biologia Tropical 45: 16351640.
  • Rasplus, J.Y., Kerdelhue, C., Le Clainche, I. & Mondor, G. 1998. Molecular phylogeny of fig wasps Agaonidae are not monophyletic. Comptes Rendus Acad. Sci. Ser. III-Sci. Vie-Life Sci. 321: 517527.
  • Swofford, D.L. 2002. PAUP*: Phylogenetic Analysis using Parsimony (*and Other Methods). Sinauer, Sunderland, MA, USA.
  • Webster, A. & Purvis, A. 2001. Testing the accuracy of methods for reconstructing the ancestral states of continuous characters. Proc. R. Soc. Lond. B. 269: 143149.
  • Weiblen, G.D. 2001. Phylogenetic relationships of fig wasps pollinating functionally dioecious Ficus based on mitochondrial DNA sequences and morphology. Syst. Biol. 50: 243267.
  • Wiebes, J.T. 1982. The phylogeny of the Agaonidae (Hymenoptera, Chalcidoidea). Neth. J. Zool. 32: 395411.
  • Wiebes, J.T. 1990. Species of Pleistodontes from the Australian continent (Hymenoptera, Agaonidae). Beaufortia 41: 219225.
  • Wiebes, J.T. 1991. Agaonidae (Hymenoptera: Chalcidoidea) and Ficus (Moraceae): fig wasps and their figs, vii. (Pleistodontes). Proc. Kon. Ned. Akad. Wet. Ser. C. 94: 137152.
  • Wiebes, J.T. 1994. The Indo-Australian Agaoninae (pollinators of figs). Proc. Kon. Ned. Akad. Wet. Ser. C. 92: 1208.