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Basic and Applied Ecology 10 (2009) 737–744 www.elsevier.de/baae

Contrasting effects of plant population size on florivory and pollination

Virve Sobera,�, Tiit Tederb, Mari Mooraa

aDepartment of Botany, Institute of Ecology and Earth Sciences, University of Tartu, Lai 40, 51005 Tartu, EstoniabDepartment of Zoology, Institute of Ecology and Earth Sciences, University of Tartu, Vanemuise 46, 51014 Tartu, Estonia

Received 4 December 2008; accepted 19 June 2009

Abstract

Changes in plant population size, induced by various forms of habitat degradation, can affect the performance ofplants by altering their interactions with other organisms such as pollinators and herbivores. However, studies on plantreproductive response to variation in population size that simultaneously consider different interactions are rare. Inthis study, we examined (1) how levels of pollinator visitation and florivory vary with population size of a self-incompatible herb, Verbascum nigrum (Scrophulariaceae), (2) the relative effect of these two interactions on host seedset, and (3) whether the intensity of florivory influences pollinator visitation rate. The intensity of florivory increased,whereas pollinator visitation rate decreased with host population size. Although average seed production wasnegatively affected by the intensity of florivory, seed production was independent of population size. The directnegative effect of florivores on plant seed set was indirectly enforced by their negative effect on pollinator visitationrate. Our results emphasize the complexity of responses of different plant–animal interactions to plant population size.However, interactions involving specialized organisms are likely to disrupt first as plant population size decreases.r 2009 Gesellschaft fur Okologie. Published by Elsevier GmbH. All rights reserved.

Zusammenfassung

Veranderungen in der Populationsgroße, die durch verschiedene Formen der Habitatverschlechterung verursachtwerden, konnen die Performanz der Pflanzen andern, indem sie die Interaktionen mit anderen Organismen wieBestaubern oder Herbivoren verandern. Dennoch gibt es wenige Untersuchungen uber die reproduktive Reaktion derPflanzen auf eine Variation der Populationsgroße, die gleichzeitig verschiedene Interaktionen berucksichtigen. In dieserStudie untersuchten wir bei einem selbstinkompatiblen Kraut, Verbascum nigrum (Scrophulariaceae), (1) wie die Levelder Blutenbesuche durch Bestauber und der Florivorie mit der Populationsgroße variierten, (2) den relativen Effektdieser zwei Interaktionen auf den Samenansatz und (3) ob die Intensitat der Florivorie die Besuchsrate der Bestauberbeeinflusste. Die Intensitat der Florivorie stieg mit der Große der Wirtspopulation, wahrend die Besuchsrate durchBestauber abnahm. Obwohl die durchschnittliche Samenproduktion durch die Intensitat der Florivorie beeinflusstwurde, war die Samenproduktion unabhangig von der Populationsgroße. Der direkte negative Effekt der Florivorenauf den Samenansatz der Pflanzen wurde indirekt durch ihren negativen Effekt auf die Besuchsrate durch Bestauberverstarkt. Unsere Ergebnisse betonen die Komplexitat der Reaktion von verschiedenen Pflanzen-Tier-Interaktionen

e front matter r 2009 Gesellschaft fur Okologie. Published by Elsevier GmbH. All rights reserved.

ae.2009.06.003

ing author. Tel.: +372 7376221; fax: +3727376222.

ess: virve.sober@ut.ee (V. Sober).

ARTICLE IN PRESSV. Sober et al. / Basic and Applied Ecology 10 (2009) 737–744738

auf die Populationsgroße der Pflanzen. Dennoch ist es wahrscheinlich, dass die Interaktionen, die spezialisierteOrganismen einschließen, zuerst zusammenbrechen, wenn die Populationsgroße der Pflanzen abnimmt.r 2009 Gesellschaft fur Okologie. Published by Elsevier GmbH. All rights reserved.

Keywords: Floral herbivory; Habitat fragmentation; Verbascum nigrum; Pollinator visitation; Seed set

Introduction

Widespread habitat degradation and concomitantreduction in plant population size may strongly impact(e.g. Cunningham 2000; Tylianakis, Didham, Bas-compte, & Wardle 2008) or even terminate (e.g.Lennartsson 2002) plant interactions with animals suchas herbivores and pollinators. Indeed, the quantitativeeffect of pairwise plant–animal interactions on plantreproductive output has frequently been shown todepend on plant population size (e.g. (Agren 1996;Aguilar, Ashworth, Galetto, & Aizen 2006; Severns2003). Changes in plant population size, however, arenot likely to influence different plant–animal interac-tions uniformly. The response of an interacting speciesto plant population size could depend on variousfactors, such as the observed spatial scale, nature ofthe interaction with the plant (mutualistic or antagonis-tic) and/or the degree of the specialization betweeninteracting species (Tscharntke, Steffan-Dewenter,Kruess, & Thies 2002).

The simultaneous action of different biotic interac-tions and their potential interference has gained moreattention recently (Strauss & Irwin 2004). As a result,the number of studies focusing on the combined effect ofdifferent interactions on plant reproductive output israpidly increasing (e.g. Gomez 2005; Herrera et al. 2002;McCall 2008; Sanchez-Lafuente 2007). However, the neteffect of different plant–animal interactions on plantperformance as dependent on plant population size hasrarely been addressed. The few known studies examiningthe net effect of different interactions on seed set inresponse to plant population size have found nouniform pattern ( (Agren, Ehrlen, & Solbreck 2008;Colling & Matthies, 2004; Kolb 2008; Steffan-Dewenter,Munzenberg, & Tscharntke 2001). Thus, the effect ofchange in population size on plant performance throughchanges in co-acting biotic interactions is not straight-forward. More studies on species with different char-acteristics and with different combinations of bioticinteractions are therefore needed to draw meaningfulgeneralizations. Moreover, most research on this topichas been conducted using simplified experimental de-signs at relatively small spatial scales (e.g. Kolb 2008).Although manipulative studies are necessary to revealcausal connections, correlative analyses assessing thestrength of interactions in natural populations are noless important.

Here, we used an obligate outcrossing perennial,black mullein (Verbascum nigrum), as a model species toestimate the combined effect of two different plant–in-sect interactions, florivory (i.e. floral herbivory) andpollination, on seed set in different-sized populations.Black mullein is pollinated by generalist pollinators(mostly bumblebees), and its flowers are consumed by aspecialist florivore. In general, plants in small or sparsepopulations are often less attractive to plant consumers.In case of pollination, for example, plants may receivefewer pollinator visits and less pollen than thoseoccurring in larger or denser populations, resulting inlower reproductive success ( (Agren 1996; Ashman et al.2004; Klinkhamer & de Jong 1990). Furthermore,because generalist pollinators can visit multiple speciesduring one foraging bout, plants in small populations orat low density may receive fewer conspecific and moreheterospecific pollen than in large populations or at highdensity (e.g. Groom 1998; Jakobsson, Padron, &Traveset 2008; Murphy & Aarssen 1995). Additionally,pollinators tend to visit more flowers per plant in smallor sparse populations, which may decrease both seedquantity and quality because of increased selfing rates(Willi, Van Buskirk, & Fischer 2005). The latter togetherwith reduced number of compatible mates (Campbell &Husband 2007) can result in higher inbreeding depres-sion and lower reproductive success in small and sparsepopulations (Ashman et al. 2004; Fischer, Hock, &Paschke 2003). Alternatively, reproductive output ofindividual plants may also decrease in very large plantpopulations, if pollinators are satiated by overabundantresources (Goulson, Stout, Hawson, & Allen 1998;Rathcke 1983; Steven, Rooney, Boyle, & Waller 2003).The influence of specialist florivores is also expected todepend on host population size. Larger plant popula-tions can support larger populations of plant consumersjust because they provide more resource, resulting inlower probability of stochastic extinction (e.g. Piessens,Adriaens, Jacquemyn, & Honnay 2009). Moreover,density of specialist herbivores in larger plant popula-tions may further increase through their density-dependent aggregation (Root 1973). Floral herbivoresare thus expected to decrease seed production more inlarger host populations than in small populations(Tscharntke et al., 2002).

In addition to the direct gamete consumption,florivores may reduce plant reproduction also indirectlyby decreasing floral attractiveness to pollinators (Hishii,

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Hirabayashi, & Kudo 2008; McCall 2008). Althoughsimilar indirect effects across the cascade of plant–ani-mal interactions are probably common in ecosystems,studies distinguishing between direct and indirect effectsare relatively rare. For example, indirect interactionsmay link frugivores to pollinators, when reduced pollina-tion (due to habitat loss or plant population reduction)leads to reduced fruit set and hence to reduced fruitavailability for frugivores (e.g. Garcıa & Chacoff 2007).Similarly, indirect interactions may emerge betweenmutualistic and antagonistic animals using the same fruitresources (e.g. Santos & Tellerıa 1994).

In this study, we asked (a) how the intensity offlorivory and pollinator visitation rate vary with hostplant population size, (b) what the relative effects ofthese two interactions are on host seed set and (c)whether the intensity of florivory influences pollinatorvisitation rate. We hypothesized that (1) plant popula-tion size would correlate positively with the intensity offlorivory and pollinator visitation rate, (2) plant seed setwould correlate positively with pollinator visitation rateand the inversely with the intensity of florivory, and (3)the intensity of florivory and pollinator visitation ratewere negatively correlated.

Material and methods

Model system

V. nigrum L. (Scrophulariaceae) is a mono- tooligocarpic, perennial herb occupying open disturbedhabitats such as gravel pits and roadsides (Reier, 2007). Itis an insect-pollinated self-incompatible species that doesnot reproduce vegetatively. In northern Europe, it flowersfrom early July until September, with peak flowering inmid-July. A typical individual has a single floweringshoot with one inflorescence consisting of up to 400 smallyellow flowers. The inflorescences rarely branch and theirmean length at our study sites was 33 cm (range:10–97 cm). Mean plant height was 90 cm (range:27–152 cm). After obligatory pollination, the flowerdevelops into a capsule containing several hundreds verysmall seeds (one seed weighs about 0.1mg). The flowersof V. nigrum are consumed by a specialist florivore, theweevil Cionus nigritarsis (Coleoptera, Curculionidae).Females of this species lay their eggs into inflorescences,from which larvae start to emerge shortly after thebeginning of flowering. Typically, larvae pupate in thesame inflorescence by the end of flowering.

Study sites

The study was carried out in summer 2006 in 21natural V. nigrum populations that were located within aradius of 70 km in eastern Estonia, northern Europe.

Minimum distance between populations was 11 km andmaximum 95 km. All chosen populations were located ingravel pits, and formed, therefore, relatively distinctpatches. All study sites had the same type of soil andvegetation and shared similar numbers of co-floweringplant species with similarly low abundances (ten plantspecies in average per site). Weather conditions (Esto-nian Institute of Meteorology and Hydrology 2006) andthe surrounding landscape were also relatively similaramong the study sites. Eleven of the 21 gravel pits wereactive. Due to excavation and/or weather conditions,some parameters in some populations could not bemeasured as is indicated by different degrees of freedomin the statistical analyses (see below).

Data collection and measurement

Plant population size was determined by counting thenumber of flowering plants during the peak floweringperiod between July 8 and 16. All flowering individualswere counted in populations of up to 300 individuals. Inlarger populations, flowering plants were counted in adefined area, and this density index was extrapolated toestimate the total number of individuals. V. nigrum

population sizes varied between 2 and 1500 individuals.The intensity of florivory was estimated by examining

flowering plants for the presence or absence of C.

nigritarsis larvae. Fifty randomly chosen floweringplants in all populations larger than 50 and allindividuals in smaller populations were examined duringthe peak of florivore abundance on July 11–13. Theintensity of florivory was then calculated as thepercentage of plants with larvae among all observedplants.

Observations to quantify pollinator visitation rate

were performed once during peak flowering (8–16 July).A patch with 5–15 flowering plants was chosenhaphazardly, avoiding population edges, and its areawas measured (1–6m2 depending on the density offlowering individuals). The patch was defined as a groupof individuals that were located at least 2m from anyother patch. In each patch, pollinator activity wasobserved for 30min during the peak visiting hours(between 5.30 and 8.30 a.m.), which coincided withmaximum floral receptivity. Pollinator visitation rateper patch was measured as the average number of visitsper plant per unit time. Multiple visits by one pollinatorto the same plant were treated as separate visits.Pollinators were classified into coarse taxonomic cate-gories. The density of flowering plants was thencalculated for each patch, and the height of each plant(from ground to the top) and inflorescence (the portionof plant with flowers) in the patch was measured. Theabundance (estimated visually: 1 – low abundance, 2 –medium abundance, and 3 – high abundance) of other

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b = 0.29F1;19 = 14.14R2 = 0.43 p = 0.0013

0

1

0 0.5 1 1.5 2 2.5 3 3.5

Inte

nsity

of f

loriv

ory

2.5

0.8

0.6

0.4

0.2

Log (population size)

V. Sober et al. / Basic and Applied Ecology 10 (2009) 737–744740

flowering species in the populations and weatherparameters (air temperature, cloudiness and windstrength) were also recorded immediately after theobservation period. As weather parameters had noeffect on visitation rates (p40.05 in all cases), they wereexcluded from the subsequent analyses. Because of theinterest in natural pollination levels and not in thestochasticity created by bee-keeping, the visits of Apis

mellifera were excluded from the measures of visitationrate in the analyses, except when examining the influenceof visitation rate on seed set.

To estimate seed set, individuals with mature seedswere collected on August 18. Twenty-five plants werecut in each population consisting of more than 25individuals; in smaller populations, all plants werecollected. Plant height and inflorescence height of thecollected individuals were measured. The total numberof fruits was counted for each individual plant, dry seedswere then separated from fruits, cleaned and weighed.Since counting all seeds of every collected individual wasnot feasible (one plant can produce up to 70,000 tinyseeds; Eichwald et al. 1969), standardized seed set wascalculated as a measure of seed production for eachindividual. To do this, the total seed-set weight and theweight of 100 randomly chosen seeds were recorded foreach individual plant. This enabled calculating anestimate of the total seed number per individual: [totalseed-set weight� 100]/[weight of 100 seeds]. Standar-dised seed set was obtained by dividing the estimatednumber of seeds by inflorescence height of the corre-sponding individual. This method allowed us toeliminate the influence of plant height. Standardizedseed set helped to account for the decrease in fruitnumber caused by florivores (flowers damaged by weevillarvae dehisce and disappear before seeds mature).Moreover, such standardization was necessary becausethe mean plant height (incl. inflorescence height) variedamong populations, possibly due to variation in theabiotic environment. Importantly, however, plant heightwas independent of population size (b ¼ 0.43;F1;14 ¼ 0.06; R2

¼ 0.0014; p ¼ 0.89). For the statisticalanalyses, mean standardized seed set was calculated foreach population.

b = -0.49F1;16 = 5.51 R2 = 0.26 p = 0.032

0

0.5

1

1.5

2

1 1.5 2 2.5 3 3.5

Log

(vis

itatio

n ra

te)

Log (population size)

Fig. 1. The intensity of florivory (A) and pollinator visitation

rate (B) in relation to population size of V. nigrum.

Statistical analyses

All statistical analyses were conducted at the popula-tion level, i.e. each population was regarded as a singleobservation. Two variables, population size and polli-nator visitation rate, were log10-transformed to obtainapproximate normal distribution of residuals.

Most analyses were performed with simple andmultiple regressions using type III sums of squares testsof significance (PROC GLM; SAS Institute Inc. 2003).We first examined the factors affecting pollinator

visitation rate and the intensity of florivory. Populationsize and density of flowering V. nigrum individuals, andthe abundance of other flowering species were used asexplanatory variables in these analyses. We thenanalysed plant seed production with regard to popula-tion size, the intensity of florivory and pollinatorvisitation rate. Finally, we examined the associationbetween the intensity of florivory and pollinatorvisitation rate.

To visualize qualitatively all direct and indirect effectsof population size on variation in seed set, we performedpath analysis using structural equation modelling (SEM;PROC CALIS; Mitchell 1992; SAS Institute Inc. 2003).We based the path diagram on the assumptions thathost plant population size influences seed set directly,but also indirectly through population-size-dependentlevels of pollination and florivory. Additionally, flor-ivory was assumed to affect pollinator visitation ratenegatively. These assumptions were based on the resultsof simple and multiple regressions.

Results

Florivores were present in all populations except one,but the proportion of plants attacked by the larvae of C.

nigritarsis varied greatly, ranging from 10% to almost100% (Fig. 1A). The intensity of florivory increasedsignificantly with V. nigrum population size (Fig. 1A).

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050

100150200250300350400450

Stan

dard

ized

see

d se

t

0.5 1 1.5 2Log (population size)

2.5 3 3.5

Fig. 3. Population mean standardized seed set in relation to

population size of V. nigrum.

V. Sober et al. / Basic and Applied Ecology 10 (2009) 737–744 741

Less than one-third of the plants were attacked byflorivores in the smallest populations, whereas almost100% were damaged in the largest populations.

Large variation was also observed in pollinatorvisitation rates, which ranged from 0 to 145 visits perplant (Fig. 1B). Most of the 4898 recorded pollinatorvisits were made by Bombus ssp. (76% of total visits).The visits by A. mellifera (11%), Syrphidae (7%) andother insects (6%) were relatively infrequent. Pollinatorvisitation rate decreased significantly with increasingpopulation size (Fig. 1B). Pollinator visitation rate didnot depend either on the density of flowering V. nigrum

individuals (b ¼ 0.61; F1;16 ¼ 2.73; R2¼ 0.22; p ¼ 0.12)

or on the abundance of other flowering species(F1;16 ¼ 0.29; R2

¼ 0.02; p ¼ 0.59).The intensity of florivory was negatively correlated

with the visitation rate of pollinators (Fig. 2). Thenegative correlation of florivory with visitation rateremained marginally significant after accounting for theeffect of plant population size (GLM, type I SS:F2;15 ¼ 6.4; R2

¼ 0.4; p ¼ 0.076).Standardized seed set varied nearly 10-fold among

populations. As expected, standardized seed set wasconsiderably lower in populations where the intensity offlorivory was higher (Table 1). In the population withthe highest intensity of florivory, the average seed setwas nearly 10 times lower than in the population withthe lowest florivore abundance (population meanstandardized seed set 43 and 425, respectively). Contraryto expectations, the effect of pollinator visitation rate onseed set remained insignificant (Table 1). Finally, there

r = -0.59N = 18p = 0.01

0

0.5

1

1.5

2

2.5

0 0.2 0.4 0.6 0.8

Log

(vis

itatio

n ra

te)

1Intensity of florivory

Fig. 2. The correlation between pollinator visitation rate and

the intensity of florivory.

Table 1. Results of a GLM analysis examining the influence

of the intensity of florivory and pollinator visitation rate on

population mean standardized seed set.

Source of variation b d.f. F P

Intensity of florivory �267.83 2;11 8.08 0.016

Visitation rate �18.78 2;11 0.13 0.72

R2¼ 0.51.

was no effect of population size on plant seed set(b ¼ �34.9; F1;14 ¼ 0.63; R2

¼ 0.05; p ¼ 0.44; Fig. 3).Path analysis visualized all direct and indirect effects

of population size on variation in seed set. According tothe path diagram (Fig. 4), population size directlyenhanced the intensity of florivory, but decreasedpollinator visitation rate. A relatively strong directnegative effect of florivory on pollination was alsorevealed. The only significant indirect effect of popula-tion size on seed set is through florivory, and, asexpected, it is negative. The direct effect of populationsize on seed set was relatively weak and not significant.The path model as a whole explained a marginallysignificant amount of the total variation.

Discussion

The levels of florivory and pollination responded tothe increase in host plant population size in a contrast-ing manner. The intensity of florivory clearly increased,whereas pollinator visitation rate declined with hostpopulation size. Seed set was strongly negativelyaffected by the intensity of florivory. In contrast, theeffect of pollinator visitation rate on average seed setwas negligible. Nevertheless, average seed set did notdepend on plant population size.

Only a minor proportion of plants were attacked byflorivores in the smallest populations of V. nigrum,whereas most plants were damaged in the largest ones.The positive effect of plant population size on florivoreabundance is consistent with the respective patternsfound in other studies. The abundance of strictlyspecialized herbivores such as C. nigritarsis typicallyincreases with host plant population size (the resourceconcentration hypothesis; Colling & Matthies 2004;Munzbergova 2006; Root 1973; Tscharntke et al. 2002).

Several potentially complementary mechanisms mayaccount for the decrease in pollinator visitation rate

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Populationsize

-0.48

-0.034

0.49

-0.41

-0.77

-0.11

R2=0.5-1; p=0.06Pollinator visitation

rate

Seed set

Intensity of florivory

Fig. 4. Path diagram depicting potential causal relationships between population size, intensity of florivory, pollinator visitation

rate and seed set of V. nigrum. Numbers above the arrows refer to the standardized path coefficients.

V. Sober et al. / Basic and Applied Ecology 10 (2009) 737–744742

with plant population size. A decrease in visitation rateis usually attributed to pollinator saturation by anoverabundance of floral resources (Goulson et al. 1998;Rathcke 1983; Steven et al. 2003). Even if the number ofpollinators is the same in all populations, their visits arestretched over more flowers in larger populations, andthus visitation rate per flower decreases. In our study,however, the decline of pollinator visits with plantpopulation size could also be associated with the otherfocal interaction, florivory. Even after correcting forplant population size, the effect of florivory onpollinator visitation rate remained marginally signifi-cant. Path analysis also revealed a negative effect offlorivory on pollinator visitation rates. Indeed, bydestroying plant reproductive organs, florivores reducethe actual number of rewarding flowers to pollinators,making these inflorescences potentially less attractive(Hishii et al. 2008). Moreover, McCall (2008) showed inan annual plant that even damaging flowers (andthereby reducing floral display) without harming repro-ductive organs can reduce pollinator visits. The ob-served negative correlation between pollinator visitationrate and the intensity of florivory implies that the directnegative impact of florivores on plant seed set may beeven further enforced through changed behaviour ofpollinators. Still, the results of the path analysis suggestthat the direct effect of population size on pollinatorvisitation is larger than the indirect effect mediated byflorivory. An experimental approach is required toevaluate the indirect effects of florivory more rigorously(Sober, Moora & Teder in preparation). Additionally,the reduced attractiveness of smaller V. nigrum popula-tions may have been compensated for by co-floweringspecies within the habitat (Ghazoul 2006). The majorpollinators of V. nigrum bumblebees – are broadgeneralists, and can therefore rely on all floral resourcessupported by a habitat fragment (Carvell 2002). Thepresent results, however, provide no support for this

rationale: pollinator visitation rate was not related tothe abundance of co-flowering species.

Average seed production differed greatly amongpopulations, varying by an order of magnitude. Thestrong negative effect of the intensity of florivory onseed set has a clear mechanistic basis, since florivoresdirectly destroy generative organs of V. nigrum. In someplant individuals, no seeds matured at all. As aconsequence, florivores in some heavily attacked popu-lations reduced the average seed set to a tenth comparedto populations with low florivore abundance. Incontrast, the effect of pollinator visitation rate onaverage seed set was negligible. Because V. nigrum isself-incompatible, we expected that higher pollinatorvisitation rates would lead to higher seed sets. The lackof a positive effect is most likely attributable to therelatively high abundance of pollinators: except for acouple of populations, most plant individuals receivednumerous pollinator visits during the short observationperiod.

Surprisingly, average seed production per plant didnot depend on plant population size, even though boththe lower intensity of florivory and higher pollinatorvisitation rate in smaller populations could havefavoured larger seed set. Various mechanisms mayaccount for this pattern. For example, in self-incompa-tible plants such as V. nigrum, seed production maydecrease because of limited availability of appropriatemating partners resulting from reduced genetic variationin smaller populations (Fischer et al., 2003; Willi et al.,2005). Previously, a positive association between plantfitness and the mean level of genetic variation has beenfound for self-incompatible – but not for self-compatible– species (Leimu, Mutikainen, Koricheva & Fischer2006). Furthermore, pollinators may become lessefficient from the plant perspective at small populationsizes, because most cross-flower movements do notoccur between conspecifics (Ashman et al. 2004).

ARTICLE IN PRESSV. Sober et al. / Basic and Applied Ecology 10 (2009) 737–744 743

In conclusion, our results emphasize the complexity ofresponses of different biotic interactions to host plantpopulation size: in our study system, the intensity offlorivory increased, whereas pollinator visitation ratedecreased with host population size. Due to thesevarying effects, predicting the response of plant repro-ductive performance in rapidly changing landscapesrequires detailed analyses of major biotic interactions.However, more specialized taxa – and the interactionsthey are involved in – are more sensitive to habitatfragmentation compared to more generalized taxa: adecrease in plant population size is likely to disruptinteractions with specialist taxa first.

Acknowledgements

We thank Lars Gotzenberger and Ants Kaasik forstatistical advice and Robert Szava-Kovats, MartinZobel and three anonymous reviewers for their valuablecomments on the manuscript. This study was supportedby the Estonian Science Foundation (Grant nos. 7371and 6619), the Estonian Ministry of Education andScience (targeted financing projects SF0180098s08,SF0180122s08), the FP 6 Integrated Project ‘‘ALARM’’(Assessing LArge scale environmental Risks for biodi-versity with tested Methods: GOCE-CT-2003-506675;www.alarmproject.net) and by the European Unionthrough the European Regional Development Fund(Centre of Excellence FIBIR).

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