- Top of page
- Materials and Methods
- Supporting Information
Pollinators are often thought to be driving floral evolution (Fenster et al., 2004). Indeed, pollinator specialization seems to have driven rapid evolution in some systems (e.g. Kay et al., 2005) and is the main hypothesis put forth to explain the diversity of flowering plants (e.g. Fenster et al., 2004). Moreover, key innovations that allow greater pollinator specialization seem to lead to diversification, as is the case for nectar spurs in Aquilegia (Hodges, 1997). Pollination syndromes (a collection of floral traits associated with attracting a particular group of pollinators) can also explain floral-trait variation (Wilson et al., 2004), suggesting that pollinator specialization has been an important force in floral evolution. Although pollinators seem to be significant drivers of floral evolution on a macro-evolutionary scale, and natural selection on floral traits is common (although not consistent) on a micro-evolutionary scale (Harder & Johnson, 2009), a major gap in our knowledge is whether pollinators are the agents of natural selection within populations (Ashman & Morgan, 2004).
Because flower size and display size are likely to be attractive to pollinators, these floral phenotypes are often assumed to be the result of selection by pollinators (Barrett & Harder, 1996; Ashman & Morgan, 2004). Furthermore, a recent review of phenotypic selection on floral traits shows that selection for larger flowers is common, and selection for flower production even more so (Harder & Johnson, 2009). However, it is also clear that floral signals used for pollinator attraction can be perceived and used by other organisms (Raguso, 2009), making them particularly vulnerable to conflicting selection (Strauss & Irwin, 2004). For example, antagonists such as herbivores and predispersal seed predators, as well as abiotic factors, can drive natural selection on floral traits (Strauss & Whittall, 2006). To conclusively determine the agents of selection, the selective environment must be manipulated (Wade & Kalisz, 1990; Conner & Hartl, 2004). However, only six studies have employed this approach to test the role of pollinators in natural selection.
We explicitly set out to test whether pollinators were acting as agents of selection on floral traits of Penstemon digitalis. Pollinators are thought to have played an important role in the diversification of the genus Penstemon (Wilson et al., 2004) and following the traditional pollination framework this could suggest that pollinators are acting as selective agents in particular Penstemon species. Therefore, we compared natural selection in open- and hand-pollinated P. digitalis to assess whether pollinators were exerting selection on floral traits. To gauge the generality of our findings, we evaluated the role of pollinators as selective agents for multiple species by reviewing studies that specifically manipulated pollination and measured natural selection. We then compared pollinators with two other potential agents of selection (herbivores and co-flowering species).
- Top of page
- Materials and Methods
- Supporting Information
We found pollinator-mediated selection in P. digitalis on two traits thought to be important for pollinator attraction: flower size and display size (Table 1). Selection for larger flowers was present in open-pollinated plants, but not in hand-pollinated plants. Natural selection on flower morphology selected for larger values (Fig. 2; Harder & Johnson, 2009) and fits with the general hypothesis that pollinators select for larger flowers because they are more conspicuous and/or may be associated with larger rewards (e.g. Blarer et al., 2002). Natural selection for larger displays is also common (Fig. 2; Harder & Johnson, 2009) and we found stronger directional selection on flower number in open-pollinated P. digitalis (Table 1). Because flower number sets the upper limit for fruit number, in many systems such as P. digitalis, there is a direct, positive relationship between flower and fruit number. Thus, positive selection would be expected on flower number independently of pollinator-mediated selection. However, we found stronger directional selection in the open-pollinated plants, suggesting that the difference in strength of selection was the result of an added benefit of larger displays attracting pollinators. We also found stabilizing selection on flower number in the open-pollinated plants, but not in the hand-pollinated plants, suggesting a cost of having too many flowers. Large displays can have increased geitonogamy, which could be costly through reduced fitness of selfed seeds (Harder & Barrett, 1995). Because hand-pollinations probably reduced the number of selfed seeds, we would expect the cost of large displays to be likewise reduced, suggesting that the difference in stabilizing selection between our treatments was a result of stabilizing selection by pollinators.
We found natural selection by pollinators despite the lack of pollen limitation. Although stronger selection via female fitness can be correlated with a greater degree of pollen limitation (Ashman & Morgan, 2004), selection by pollinators is not always associated with pollen limitation. Galen (1996) found selection by pollinators without pollen limitation, and two studies found pollen limitation, but no selection by pollinators (Andersson, 1996; Totland et al., 1998). Therefore, there may be an increased likelihood of finding selection by pollinators in pollen-limited populations because selection is probably stronger. However, this cannot be assumed to be true for all populations.
Field estimates of phenotypic selection can be biased as a result of environmental covariance between traits and fitness (Rausher, 1992). We attempted to control for this bias by physically pairing our treatments and using a blocked design. There were phenotypic differences between our two blocks which suggest that pollination, competition and/or resources may have differed in these parts of the population. However, when we included block as a random factor in our selection models, the significant patterns remained, suggesting that an environmental gradient was not responsible for the overall selection we found.
If biotic agents frequently exert natural selection on plants, then we would expect stronger selection when the agents are interacting with the plant, compared to when they are experimentally removed. For pollinators specifically, we expect selection on floral traits to be stronger when pollinators are selecting plants than when experimenters hand-pollinate them. Indeed, we found that natural selection on floral traits was stronger in the presence than in the absence of pollinators (Fig. 2b). Furthermore, we do not necessarily expect herbivores to be strong selective agents on flowers, although they could influence floral evolution in a number of ways (Strauss & Irwin, 2004). We found that selection on floral traits was equivalent regardless of whether herbivores were present or absent (Fig. 2c). However, there were few selection coefficients with which to test this hypothesis, so these findings should be interpreted with caution. Co-flowering species could either have competitive or facilitative effects on the focus species, which could lead to divergent or convergent evolution of floral traits (Caruso, 2001). Therefore, it is difficult to predict across systems whether selection should be stronger with or without a co-flowering species. When co-flowering species were manipulated, selection was stronger in the absence of the co-flowering plant (Fig. 2d). However, there were also many more reversals when co-flowering species were removed (i.e. positive selection became negative, or vice versa), suggesting that community context could alter selection on floral traits (Caruso, 2000). Surprisingly, we found significant trends for manipulations of both pollinator and co-flowering species, despite our small sample size and the fact that selection is frequently weak in natural populations (Kingsolver et al., 2001; Knapczyk & Conner, 2007). Indeed, the majority of the selection coefficients included were close to zero and not significant. However, our ability to detect trends suggests that these patterns could be strong in nature.
Our survey data could be biased by three major limitations. First, although our complete data set included plants spanning multiple functional pollinator groups, pollinator manipulations have been limited to hymenopteran and dipteran pollinators. Pollinator shifts have been proposed as a major driver of speciation (e.g. Fenster et al., 2004) and some shifts seem more common than others. For example, bee-to-bird pollination is more common than the reverse (Thomson & Wilson, 2008). However, it is uncertain whether one functional group would exert stronger selection on floral traits than another. Thus, the addition of plants pollinated by other functional groups, such as birds, beetles, moths, etc., may alter the pattern we detected. Second, although our data set spans almost as many families and species as studies, it only includes herbaceous plants. Studying selection in herbaceous plants is a bias common to the plant literature; however, it is important to note that it may affect our ability to generalize the results to all flowering plants (e.g. Geber & Griffen, 2003). Because natural selection by pollinators is of general interest in floral evolution, measuring selection by pollinators in nonherbaceous plants and/or nonbee/fly-pollinated plants will provide further insights into their role as agents of natural selection. Lastly, our survey was necessarily limited to studies that measured the selection coefficients described by Lande & Arnold (1983) and directly manipulated an agent of selection. This allowed direct comparisons to be made across studies but could also introduce bias.
We were unable to directly compare the impacts of pollinators with selection by other floral visitors, such as florivores (McCall & Irwin, 2006) or predispersal seed herbivores (these data have simply not been collected, but see Wise & Cummins, 2007). In particular, predispersal seed predators are expected to conflict with pollinator selection on floral traits because they should visit fertilized flowers (or those that will be fertilized). Moreover, they can exert selection on floral traits in multiple systems (Pilson, 2000; Cariveau et al., 2004; Rey et al., 2006; Parachnowitsch & Caruso, 2008), making a direct comparison between seed predator- and pollinator-mediated selection a relevant question. For our population of P. digitalis, we found no evidence that predispersal seed predators influenced selection on flower size or number, or affected selection on aborted fruits. Only five of the 281 plants in this experiment showed evidence of any fruit damage, and our results were robust to excluding these plants (data not shown). Predispersal seed herbivores can be much more frequent in P. digitalis (Mitchell & Ankeny, 2001; A. L. Parachnowitsch, unpublished) and it is possible that they do exert natural selection in other populations and/or years.
An important limitation of our studies and those reviewed here is that they generally only measure female fitness in hermaphroditic plants in one season. Natural selection can vary over time (Siepielski et al., 2009). Therefore, to fully understand the evolutionary trajectory for a population, selection should be measured in multiple years. Moreover, if most of the natural selection on flowers by pollinators or other agents occurs via male function, then these types of experiments may fail to detect their impact (see Conner (1997) for a theoretical discussion and counter example). Although few natural-selection studies examine both male and female fitness estimates, the general pattern from the literature does not support the male-function hypothesis. That is, selection on attractive floral traits was not always stronger via male fitness, but rather selection through male and female fitness was context-dependent (Ashman & Morgan, 2004). Therefore, hand-pollinations can provide valuable information about selection by pollinators but are still only half of the story. Manipulations of the agents of selection via both male and female fitness are necessary to give a more complete picture and reveal functional differences.
Flowers offer a conundrum to evolutionary ecologists. On the one hand it seems fairly obvious that flowers are adapted to their pollinators (reviewed in Harder & Johnson, 2009) and that shifts in pollination systems have led to diversification in plants (e.g. Kay et al., 2005). Moreover, there are a limited number of examples of pollinator-mediated natural selection, such as those found in P. digitalis. However, the majority of selection estimates on floral traits from manipulations of pollination do not support selection by pollinators (Fig. 2b), and studies that have used path analyses often find that pollinators weakly effect fitness (Conner et al., 1996; Gómez, 2000; Rey et al., 2006; Ashman & Penet, 2007). Harder & Johnson (2009) argue that natural-selection studies may be measuring the effects of pollinators during periods of relative stasis and that we should expect selection by pollinators on novel traits or in new environments. In other words, the lion’s share of pollinator-mediated natural selection shaping flowers may have occurred during and shortly after speciation. While this view needs further testing, we are left to explain what the agents of selection are in cases when there is significant selection on floral traits, but not by pollinators (e.g. Andersson, 1996; Totland et al., 1998; Fishman & Willis, 2008; Parachnowitsch & Caruso, 2008; Sandring & Ågren, 2009). If pollinators truly exert most of their selection on flowers during cladogenesis, then an interesting question follows. During a period of stasis, how far from these adaptive peaks can nonpollinator agents push floral traits within a particular population or species?