Specificity, Rank Preference, and the Colonization of a Non-Native Host

Oecologia (2013) 172:177–188 DOI 10.1007/s00442-012-2476-8

PLANT-ANIMAL INTERACTIONS - ORIGINAL RESEARCH

Speciﬁcity, rank preference, and the colonization of a non-native host plant by the Melissa blue butterﬂy

M. L. Forister • C. F. Scholl • J. P. Jahner • J. S. Wilson • J. A. Fordyce • Z. Gompert • D. R. Narala • C. Alex Buerkle • C. C. Nice

Received: 24 April 2012 / Accepted: 7 September 2012 / Published online: 4 October 2012 Ó Springer-Verlag 2012

Abstract Animals often express behavioral preferences individuals from alfalfa-associated populations, there for different types of food or other resources, and these appears to have been a decrease in specificity: females preferences can evolve or shift following association with from these populations lay fewer eggs on the native host novel food types. Shifts in preference can involve at least and more eggs on the exotic relative to females from two phenomena: a change in rank preference or a change in native-host populations. However, females from alfalfa- specificity. The former corresponds to a change in the order associated populations did not lay more eggs on a third in which hosts are preferred, while a shift in specificity can plant species, which suggests that preferences for specific be an increase in the tendency to utilize multiple hosts. hosts in this system can potentially be gained and lost These possibilities have been examined in relatively few independently. Geographic variation in oviposition pref- systems that include extensive population-level replication. erence in L. melissa highlights the value of surveying a The Melissa blue butterfly, Lycaeides melissa, has colo- large number of populations when studying the evolution nized exotic alfalfa, Medicago sativa, throughout western of a complex behavioral trait. North America. We assayed the host preferences of 229 females from ten populations associated with novel and Keywords Exotic species Hierarchical Bayesian Á native hosts. In four out of five native-associated popula- model Host range Lycaeides Oviposition Á Á Á Á tions, a native host was preferred over the exotic host, Specialization while preference for a native host characterized only two out of five of the alfalfa-associated populations. Across all Introduction

Communicated by Konrad Fiedler. Closely related insects commonly exhibit behavioral pref- M. L. Forister (&) C. F. Scholl J. P. Jahner erences for different plant species (Jaenike 1990; Thomp- J. S. Wilson D. R.Á Narala Á Á son and Pellmyr 1991; Gripenberg et al. 2010), and shifts Á Department of Biology, University of Nevada, in host use associated with the colonization of novel plants Reno, NV 89557, USA are thought to underlie major instances of diversiﬁcation e-mail: [email protected] (Fordyce 2010; Janz 2011). Thus, it is of interest to ask J. A. Fordyce how behavioral changes in oviposition preference come Department of Ecology and Evolutionary Biology, about, and at least two modes or routes of change can be University of Tennessee, Knoxville, TN 37996, USA identiﬁed (Singer 1982). One mode is a shift in rank Z. Gompert C. C. Nice preference, which can be a transition from a state in which Department ofÁ Biology, Texas State University, an ancestral host is preferred to a state in which a novel San Marcos, TX 78666, USA host is preferred. ‘‘Preferred’’ in this context refers to the amount of reproductive output a given female devotes to a C. Alex Buerkle Department of Botany, University of Wyoming, particular plant given a choice between species: a preferred Laramie, WY 82071, USA host receives more eggs. A second mode of change

123 178 Oecologia (2013) 172:177–188 involves a change in specificity, where ‘‘specificity’’ refers (Singer et al. 1994) in combination with the presence of both to the number of hosts that are utilized and more heritable variation in oviposition preference, and a similar generally to the distribution of eggs within a given pref- process seems to be happening with another butterfly erence hierarchy (Singer 1982; Thompson and Pellmyr (Pieris oleracea) on another exotic host (Keeler and Chew 1991). A highly specific female will lay all of her eggs on a 2008). Similarly, in Rhagoletis flies associated with apple single host, whereas a female with lower specificity might, trees as the exotic host, there has been an evolution of for example, lay most of her eggs on a preferred host and preference such that apple-associated flies respond most a smaller number of eggs on a second-ranked host. A readily to odors of the novel host (Linn et al. 2003). conceptual model for changes in rank preference and In contrast to the evolution in preference exhibited by specificity is presented in Fig. 1, which includes a change E. editha, P. oleracea, and Rhagoletis, rank preference in rank preference but not specificity (transition ‘‘a’’), a hierarchies in swallowtail butterflies tend to be more change in rank preference and specificity (‘‘b’’), and a broadly conserved, with some widespread species exhibit- change in specificity but not rank preference (‘‘c’’). Spec- ing similar preference hierarchies in different geographic ificity and rank preference are both useful descriptors of regions despite association with different local host species oviposition behavior (Singer 1982, Courtney 1989, Singer (Thompson 1998). A recently described hybrid swallowtail et al. 1992a), and a change in oviposition behavior might species (between Papilio glaucus and P. canadensis) involve a shift in either preference or specificity. retains the oviposition preferences of one of the parental Some of the systems that have most influenced thinking species, even though the preferred host is locally absent about the evolution of oviposition behavior involve insects (Mercader et al. 2009). In Colias philodice butterflies, the and exotic or invasive plants (Carroll and Dingle 1996). utilization of a novel host was not accompanied by a The checkerspot butterfly Euphydryas editha has under- change in preference, though in this case native-feeding gone repeated bouts of diet breadth evolution, with one of populations seem to have been pre-adapted with preference the most well known involving a shift in rank preference for the new host (Tabashnik 1983). The diversity of ecol- favoring the exotic host Plantago lanceolata (Singer et al. ogies and life histories of herbivorous insects would sug- 1993). Selection in this example appears to have acted as a gest that additional studies are needed before we can draw simple response to high larval survival on the exotic generalities regarding modes of evolution of oviposition behavior. Here we examine the Melissa blue butterfly, Lycaeides a melissa, which has been the focus of studies in the ecology of host use (Nice et al. 2002; Fordyce and Nice 2003; Change in Forister et al. 2011; Scholl et al. 2012), and which is part of rank preference but not the Lycaeides species complex that has provided examples specificity of interspecific hybridization and hybrid speciation Preference (Gompert et al. 2006; Lucas et al. 2008; Gompert et al. Plant Plant 2010). L. melissa can be found throughout much of western b North America, where it widely utilizes exotic alfalfa, Change in Medicago sativa, which was brought to the continent Change c specificity in rank within the last 200 years (Michaud et al. 1988). Although but not rank preference cultivated alfalfa is used in some locations, L. melissa is preference and more often found with roadside or feral alfalfa. In at least specificity some areas where L. melissa populations have been studied, alfalfa is an inferior larval resource relative to a common native host, Astragalus canadensis (Forister et al. Preference Preference Preference 2009). However, populations persist on the novel host, which is due at least in part to the abundance of mutualistic Plant Plant ants on alfalfa (Forister et al. 2011). Ants tend caterpillars and protect them from natural enemies in exchange for Fig. 1 Conceptual model for the evolution of oviposition preference through changes in rank preference and specificity. Each panel sugary excretions produced by specialized organs of the illustrates the fraction of eggs laid by a single female on three plants caterpillars. (indicated by different symbols). The female in the upper left is from a Previous studies of preference involving L. melissa hypothetical ancestral population characterized by a strong preference populations have reported contrasting results: Nice et al. for the first host (circle). The three paths (labeled a, b, and c) indicate three ways in which the ancestral preference might evolve through (2002) found weak preference or an absence of preference different combinations of change in specificity or rank preference in a single population depending on the plants used in

123 Oecologia (2013) 172:177–188 179 choice tests, while Forister et al. (2009) described rela- GLA tively strong preferences in two populations. These studies SUV encompassed a small number of populations, and used LAC different plants and experimental designs. In the present BWP SLA study, we use a single design and a larger number of VCP populations to compare preferences of alfalfa-associated REW WLA populations to preferences of native-associated popula- GAV tions. If preferences differ between these groups, we ask to what extent changes involve a shift in rank preference or a BHP shift in specificity (Fig. 1). One prediction is that individuals from alfalfa-associated and native-associated populations will express similar preferences. Perhaps, for Native, A. canadensis example, native hosts are preferred, as was found in two Native, L.L. argenteusargenteus populations by Forister et al. (2009), and preference for Native, G. lepidota A. canadensis && alfalfaalfalfa alfalfa has not increased because it is an inferior larval Alfalfa resource: females that express a preference for the novel host would have lower fitness. Such a scenario would be most likely where native and novel hosts occur in some 0 100 200 km geographic proximity, giving females with a preference for native hosts a relative fitness advantage. However, in our Fig. 2 Map of study populations and host associations in the states of study areas on the western edge of the Great Basin, pop- California and Nevada in the western United States. Symbol locations ulations only rarely have more than one available host are approximate, and are shifted slightly so that overlapping symbols can be seen; exact locations are given in Table 1 species and are separated by inhospitable desert. In geographic isolation on the novel host, preference for the native host could be lost through drift, leading to the that we have used elsewhere for characterizing variation in alternative prediction that alfalfa-associated populations preference among Lycaeides populations (Fordyce et al. might have a higher preference for alfalfa. Of course, this 2011). In some cases the target sample was obtained in a second prediction assumes an independent genetic basis for single visit, and in some cases more than one visit was preference associated with different hosts, which is not necessary. All collections were made during July and known in Lycaeides but has been suggested for other her- August, when L. melissa populations were in their second bivorous insects (Singer et al. 1992b; Hawthorne and Via and third generations. After capture in the field, females 2001; Forister et al. 2007). These contrasting scenarios were kept cool and transferred in glassine envelopes to the illustrate some of the possibilities for the evolution of University of Nevada, Reno, for preference tests. preference in L. melissa, which we investigated by sur- We focus here solely on preferences of wild-caught as veying preferences of females collected from ten popula- opposed to lab-reared females. Lab-reared females can be tions: five associated with alfalfa, four associated with used in preference experiments to standardize female native hosts, and one population that utilizes both a native condition and age, and also to control for the possibility of host and alfalfa. early-adult experiences influencing preference (Davis 2008). Lab-reared females of L. melissa have been studied for preference elsewhere (Forister et al. 2009), but this Materials and methods approach was not logistically compatible with the goals of the present study, which included wide geographic sam- The location of the ten populations sampled is shown in pling and a large number of populations. Moreover, lab- Fig. 2. These populations were chosen to encompass both reared L. melissa females tend to lay fewer eggs relative to native host and alfalfa associations, and are generally wild-caught females (MLF, unpubl. data), which compro- characteristic of low-elevation, Great Basin L. melissa mises statistical power in preference tests. habitat. With respect to the habitat of alfalfa-associated populations, some of these (VCP and BWP) are associated Preference tests with feral plants not in immediate proximity to cultivated stands, while the other alfalfa locations include the margins Three-way choice arrays were used to test the preferences of cultivated fields or roadsides near managed alfalfa fields. of female butterflies. The three hosts in all tests consisted Our goal was to assay the preferences of between 20 and of (1) a native host, (2) the novel host, alfalfa, and (3) a 30 females from each population, which is a sample size negative control, Lotus nevadensis, the host of a closely

123 180 Oecologia (2013) 172:177–188 related Lycaeides species (L. anna; formerly L. idas anna). within 72 h of the start of preference experiments and The identities of the latter two plants, alfalfa and the stored in a refrigerator at 5 °C. In collecting plants from a negative control, were constant across tests, while the restricted number of geographic locations, we intentionally native host was specific to the populations involved. When minimized potential variation associated with interpopu- testing females from the native-associated populations, the lation differences in, for example, phytochemistry. Such native host in test arrays was simply the host from the variation could be quite important, and could be involved population of origin (either Astragalus canadensis, Lupinus in the local adaptation of insects preferring local plants; argenteus, or Glycyrrhiza lepidota). For choice tests however, this possibility is beyond the scope of the present involving females from alfalfa-associated populations, we study. The collection locality (VCP) for the alfalfa used in used Astragalus canadensis as the representative native. preference cages is a feral stand not associated with culti- We used this native host in tests of alfalfa-associated vation, and is not managed or harvested (the identity of the females both because it is a common native host in the variety or cultivar of alfalfa growing at VCP is not known western Great Basin, and also because we have studied it to us). previously, which allows for continuity with previously Preference arrays consisted of large, opaque plastic published studies. Throughout the course of experiments, drinking cups containing sprigs of each of the three plant Astraglus canadensis was collected from Silver Lake species, with the ends of the sprigs pushed through holes in (SLA), just north of Reno, Nevada, and alfalfa was col- the bottom of the cup into water in a second, smaller cup. lected from Verdi (VCP), west of Reno, Nevada (Table 1). The top of the larger cup was covered in bridal veil that had Lupinus argenteus and Glycyrrhiza lepidota were both been soaked in fruit punch flavor GatoradeÒ, which was collected from the same locations where butterflies were available to the butterflies throughout the course of the collected in association with these hosts (LAC and BHP, experiment (GatoradeÒ on the mesh was refreshed with respectively; Table 1). In all cases, plants were collected water from a spray bottle three times each day).

Table 1 Collection details and related information, including host associations Population Lat., Long. Host Sample sizes and collection dates Eggs per female (±SE)

Silver Lake (SLA) 39.6497, Astragalus canadensis 25 (17 July 2011) 18.20 (3.4) -119.9263 Washoe Lake (WLA) 39.2346, Astragalus canadensis 6 (19 July 2011), 12.87 (2.0) -119.7831 25 (22 August 2011) Beckwourth Pass (BWP) 39.7797, Astragalus canadensis, 23 (28 July 2011) 19.09 (3.29) -120.0734 Medicago sativa Lamoille Canyon (LAC) 40.6847, Lupinus argenteus 12 (29 August 2011) 11.33 (2.64) -115.4734 Bishop (BHP) 37.3585, Glycyrrhiza lepidota 20 (20 July 2011), 8.33 (1.58) -118.3906 7 (17 August 2011) Goose Lake (GLA) 41.9860, Medicago sativa 33 (21 July 2011) 19.24 (3.09) -120.2925 Red Earth Way (REW) 38.9808, Medicago sativa 20 (20 July 2011) 11.45 (2.35) -118.8371 Gardnerville (GAV) 38.9608, Medicago sativa 21 (11 August 2011), 12.84 (2.07) -119.7788 4 (29 August 2011) Surprise Valley (SUV) 41.2821, Medicago sativa 10 (21 July 2011) 5.6 (2.02) -120.0987 Verdi Crystal Peak (VCP) 39.5102, Medicago sativa 2 (8 July 2011), 10.87 (2.09) -119.9952 12 (9 August 2011), 9 (12 August 2011) Preference data were collected from a total of 229 females; 118 from native-associated populations (including BWP, where two hosts are present, native and exotic), and 111 from populations associated only with alfalfa, Medicago sativa. The last column has the mean (±standard error) number of eggs per female per population Sample sizes refer to the number of females that laid eggs in preference assays (in some cases, females were collected on more than one day)

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Experiments were set up in the late afternoon of the day that The model is implemented in the R package bayespref females were collected, and taken down 48 h later, at which in a Markov chain Monte Carlo (MCMC) framework, and a time eggs on foliage were counted (eggs that had fallen off number of options are available for hypothesis testing. or were laid on the cup or mesh were not counted). Pref- Within individual populations, the model can be run with erence tests were conducted in an outdoor lath house, with individuals constrained to have equal preference among filtered shade and at ambient temperature. host plants (i.e., lay an equal fraction of eggs, on average, The negative control (L. nevadensis) is the primary host of on each of three plants). The DIC (deviance information a closely related species, Lycaeides anna, at Yuba Gap, CA, criterion) value from the constrained model is then com- USA, on the west slope of the Sierra Nevada mountains. pared to the DIC value from an unconstrained model (in Previous experiments found that L. melissa will lay a small which preferences among host plant species are permitted fraction of eggs on L. nevadensis in choice experiments to vary). DIC is the appropriate model selection tool in a involving A. canadensis, alfalfa, and L. nevadensis (Forister Bayesian framework when the posterior distributions are et al. 2009), and L. melissa caterpillars can be readily reared approximated using MCMC (Spiegelhalter et al. 2002). on L. nevadensis to adult weights that equal or exceed adult DIC is interpreted similar to AIC values (Burnham and weights of individuals reared on the native host A. canadensis Anderson 2002), where the lowest value indicates the best- (Scholl et al. 2012). This plant was chosen for the experiments fit model. Thus, smaller DIC values obtained from the described here to gain insight into potentially generalist unconstrained model, compared to the constrained model, oviposition behavior. For example, one possible route to would suggest that substantial preference variation exists preference evolution could be an increase in preference for for the different plants (not all plants receive an equal the exotic host specifically, while a second route of change fraction of eggs). could be a simple decrease in specificity (path ‘‘c’’ in Fig. 1), For tests among populations, groups of populations can which could be seen as eggs more evenly distributed across be constrained to have the same preferences. This was all three hosts, including the negative control. used, for example, to ask whether native and alfalfa-asso- Finally, we note that only choice tests were used, as ciated populations differ in their preferences for the three opposed to no-choice tests. In some cases, and with certain hosts. The constraint approach was also used within the insects, the two types of experimental design can produce native and alfalfa-associated populations in separate con- different results. However, we have found qualitatively strained analyses to ask to what extent significant among- similar results using the two designs with Lycaeides population variance exists in preference within these (Forister et al. 2009; Fordyce et al. 2011) and with other groups. Finally, a comparison of constrained and uncon- lycaenid butterflies (Forister 2008). strained models was used to address variation in preference among sampling dates. For populations that were visited Analyses more than once (Table 1), females from the two dates were scored as belonging to two ‘‘populations’’ and bayespref Preference tests have traditionally been analyzed with was run with and without females from the two dates being nonparametric analyses, such as the Friedman or Quade constrained to have the same preferences (the two females tests, the goal of which is to evaluate the null hypothesis of from the first date at VCP were too few and were not no difference in preference for the items being chosen included in these tests). (Conover 1999). Instead, we utilized a hierarchical In addition to analyses within populations, and among Bayesian model designed for ecological count data (Ford- groups of populations (e.g., native and alfalfa-associated yce et al. 2011). This approach offers a number of populations considered separately), an analysis was also advantages, in particular the estimation of parameters that performed in which we treated all native-associated indi- explicitly describe the strength of preference for specific viduals as belonging to a single population, and likewise plants, which is more informative than simply rejecting a for all alfalfa-associated females (i.e., ignoring population null hypothesis of no preference. The hierarchical model structure within those two host-associated groups). uses the observed count data (the numbers of eggs on Assuming that individual females were representative different plants) to estimate preferences of individual samples of these two groups allowed us to directly estimate females as parameters of a multinomial distribution; pref- preferences and ask whether, for example, alfalfa-associ- erence at the population level is in turn modeled as a ated females have a lower or higher preference for the Dirichlet distribution from which individual preferences native host relative to females from native-associated are sampled. By estimating the strength of preference, and populations. This approach (pooling all individuals within comparing these estimates among populations, we can the two groups of populations) is complementary to the address both changes in rank preference and changes in analyses described above involving model comparison and specificity, as illustrated in Fig. 1. constraints in preference within groups of populations.

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In all cases, MCMC runs were conducted for 5,000 in the better fit of the first, unconstrained model reported in generations with the first 1,000 generations discarded as a Table 2, and in visual inspection of the posterior proba- burn-in. Significant differences between estimates of bility distributions for population-level preferences illus- preference for different host plants within populations were trated in Figs. 3 and 4. Variation in preference among detected through pairwise comparisons of sampled values populations is also reflected in the better fit (lower DIC during post-burn-in MCMC steps. In this procedure, if scores) for the unconstrained models within groups of both preference for a particular plant is greater than preference native and alfalfa-associated populations (Table 2). for another plant in more than 95 % of the sampled MCMC Between sampling dates within populations, model com- iterations, then the two preference estimates are considered parisons suggested that preference was consistent in time: to be different (Fordyce et al. 2011). For simplicity, single all DDIC values comparing models that were or were not MCMC search chains were used for all analyses, after constrained to have similar preferences across sampling verifying the stability of results with multiple search chains dates had values of \2. for a representative sample of populations and model A clear hierarchy of preference was observed in many comparisons. Diagnostic plots of MCMC samples were populations, specifically a hierarchy in which the native examined in all cases to ensure mixing of search chains. host was preferred over both alfalfa and the negative control, as was true at four out of the five native-associated populations (SLA, WLA, BWP, and LAC), and at two of Results the alfalfa-associated populations (GLA and REW), as can be seen in Figs. 3 and 4 as well as in the model compari- A total of 3,771 eggs were laid by 229 females in the sons reported in Table 3 (the constrained model reflecting preference experiments (an average of 23 females were no preference was a better fit at only BHP and SUV). In no tested per population, laying an average of 15 eggs per case did either alfalfa or the negative control rank first as female). Table 1 includes the number of females assayed the most preferred host (alfalfa was ranked first at SUV, but from each site as well as the mean number of eggs laid per the difference between alfalfa and the second-ranked native female within populations. Considerable variation in pref- was not significant; Fig. 4). However, there was variation erence was observed among populations: this can be seen in rank preference, specifically in the extent to which alfalfa was preferred above the negative control. In the four native-associated populations where females expressed a Table 2 Model comparisons investigating consistency of preference preference for the native host, they did not distinguish among all populations, and within groups of native- and alfalfa- between the other two plants (Fig. 3). In contrast, prefer- associated populations considered separately ence was exhibited for alfalfa over the negative control at Model DIC DDIC two of the alfalfa-associated populations, GLA and GAV (Fig. 4). At the latter population (GAV), preference was All populations equivalent for the native host and alfalfa, which was also Unconstrained model: interpopulation variance in 257.10 true at VCP (Fig. 4). preference allowed Across populations, the greatest disparity in behavior Partly constrained model: populations assigned to 293.20 36.10 two groups (native- and alfalfa-associated), with involved preferences for the native host (Fig. 5), which preferences constrained to be similar within the ranged from nearly 80 % at WLA and SLA to below 30 % two groups at BHP, while preferences for alfalfa were generally below Fully constrained model: all populations (native- 382.80 125.7 50 %. Variance in preference among individuals (reflected and alfalfa-associated) constrained to have in the parameter) within populations was similar across similar preferences x most populations (Fig. 5d), with the exception of two Native populations alfalfa-associated populations, SUV and GAV, which Unconstrained model: interpopulation variance in -87.26 preference allowed showed elevated values of this parameter, indicating less Constrained model: preferences constrained -70.00 17.26 variation among individuals (i.e., higher ‘‘precision’’ of Alfalfa populations estimates from the Bayesian model). When females were Unconstrained model: interpopulation variance in 329.52 pooled within host-associated groups, an overall decrease preference allowed in preference for native hosts and an increase in preference Constrained model: preferences constrained 373.58 44.06 for alfalfa was estimated for alfalfa-associated females compared to native-associated females (Fig. 6). Alfalfa The first three models refer to analyses involving all populations (DDIC values reported in rows two and three are with reference to the and native-associated females showed a similar reluc- first, unconstrained model), while the lower four models are from tance to lay eggs on L. nevadensis, the negative control native- and alfalfa-associated populations considered separately (Fig. 6).

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Fig. 3 Posterior probability distributions for population- a SLA a WLA level (solid lines) and a a individual-level (dotted lines) b

10 15 b 10 15 preferences estimated from hierarchical Bayesian models of host preference for populations associated with native host Probability plants. The three colors correspond to the different plant species as indicated in the 05 05 legend, and lower-case letters 0.0 0.2 0.4 0.6 0.8 1.0 0.0 0.2 0.4 0.6 0.8 1.0 denote posterior probabilities of [0.95 for differences in preference. Three-letter codes BWP LAC designate populations; see also Fig. 2 and Table 1 a 10 15 a 10 15 a b a b Probability 05 05 0.0 0.2 0.4 0.6 0.8 1.0 0.0 0.2 0.4 0.6 0.8 1.0 Preference BHP

Population level Individual level

Native Probability Alfalfa Neg. control 0 5 10 15 0.0 0.2 0.4 0.6 0.8 1.0 Preference

Discussion GLA females (that comparison also involves a change in rank, as GLA females distinguished between alfalfa and the Oviposition preference can evolve in a number of ways negative control while REW females did not). Across all (Fig. 1), and previous research on herbivorous insects has populations, there was a general preference for native revealed shifts in rank preference (e.g., Singer et al. 2008) hosts, but alfalfa-associated females had a lower preference as well as instances of conserved preference hierarchies for native hosts and a higher preference for alfalfa than (Thompson 1998). Variation in specificity is likely com- native-associated females did (Fig. 6). mon among populations and species of herbivorous insects These results are suggestive of evolutionary changes in (e.g., Singer et al. 1994, Mercader and Scriber 2007), oviposition preference associated with the colonization of a though changes in specificity are not as frequently novel host plant, which raises the issue of plasticity (or addressed in the literature as are simple changes in rank induction) versus genetically fixed preferences. Plasticity, preference. In comparisons among L. melissa populations, through imprinting of oviposition preferences associated we observed differences in rank preference as well as with juvenile experience in herbivorous insects, was once differences in specificity. For example, GLA and GAV considered likely (as Hopkins’ host-selection principle), females differed in rank preference, with the native being then quite unlikely (Barron 2001; Janz et al. 2009). preferred in the former and the native and exotic hosts Developmental plasticity in oviposition is now considered being equally preferred in the latter (Fig. 4). A difference possible under certain circumstances, such as preference in specificity can be seen in the comparison between GLA induction through early adult experience in some species and REW (Fig. 4), where the native host received pro- (Zhang et al. 2007; Davis 2008). Ideally, all of the exper- portionally fewer eggs from REW females as compared to iments described here would have been carried out with

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Fig. 4 Posterior probability a distributions for population- GLA REW level (solid lines) and b individual-level (dotted lines) c preferences estimated from a a hierarchical Bayesian models of b host preference for populations associated with the exotic host Probability alfalfa M. sativa. The three colors correspond to the different plant species as 0 5 10 15 0 5 10 15 indicated in the legend, and 0.0 0.2 0.4 0.6 0.8 1.0 0.0 0.2 0.4 0.6 0.8 1.0 lower-case letters denote posterior probabilities of [0.95 for differences in preference. GAV SUV Three-letter codes designate a populations; see also Fig. 2 and Table 1 bb a b ab Probability 0 5 10 15 0 5 10 15

0.0 0.2 0.4 0.6 0.8 1.0 0.0 0.2 0.4 0.6 0.8 1.0

Preference VCP

a Population level Individual level ab b Native Probability Alfalfa Neg. control 051015 0.0 0.2 0.4 0.6 0.8 1.0 Preference

Table 3 Model comparisons investigating host preference within lab-reared females. This is impractical in Lycaeides, lar- populations gely because most lab-reared females will only lay a Population Full DIC Constrained DIC DDIC small number of eggs, insufficient for preference tests (MLF, unpubl. data). However, lab-reared females from SLA (N) -67.82 -33.56 34.26 one population have been previously tested, and were WLA (N) -104.61 -36.92 67.69 found to express behavioral preferences identical to wild- BWP (AN) 76.55 99.03 22.48 caught individuals from the same population (Forister LAC (N) -56.13 -3.39 52.74 et al. 2009). A genetic basis for fixed preferences, rather BHP (N) 63.20 -28.83 -92.03 than plasticity, can also be inferred from the results GLA (A) 21.00 60.30 39.30 reported here. Females from GLA and REW expressed REW (A) 75.69 82.11 6.42 preferences that were qualitatively similar to those GAV (A) 116.39 141.22 24.83 expressed by native-associated females (Figs. 3, 4). SUV (A) 24.97 -29.83 -54.8 Females from GLA in particular laid a high fraction of VCP (A) 92.47 99.43 6.96 eggs on the native host, with which they presumably had Full DIC values correspond to models in which preference for hosts is no previous contact in the wild (L. melissa butterflies are allowed to vary. Constrained values are from models in which pref- fairly sedentary, and we know of no A. canadensis plants erence for the three hosts is constrained to be the same (i.e., preference anywhere near the GLA population). Lacking previous for each host = 0.33). Delta (D) values correspond to the difference experience with the native host, the preferences of GLA between the full and constrained DIC values; larger, positive DDIC values indicate greater strength of host preference. Association with and REW females would not have been induced by larval alfalfa (A), native hosts (N), or both (AN) is indicated in parentheses or adult experiences.

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a 1.0 a ab e bcd d b 1.0 abc bcde f ab cde a bc a cd e bcd ef a ab de

0.8 0.8

0.6 0.6

0.4 0.4

0.2 0.2 Preference for alfalfa for Preference Preference for native for Preference

0.0 0.0 SLA SLA LAC LAC GLA GLA GAV GAV SUV VCP BHP BHP VCP SUV WLA WLA BWP BWP REW REW

c 1.0 a cd d bc cd d bc c abc ab abc cd cd ab bc d c c a bc c 50 0.8 40

0.6 30 ω 0.4 20

0.2 10 Preference for neg. control neg. for Preference 0.0 0 SLA SLA LAC LAC GLA GLA GAV GAV VCP BHP BHP SUV SUV VCP WLA WLA BWP BWP REW REW

Fig. 5 Summaries of preference (a–c) across populations for the Figs. 3, 4). Lower case letters at the top of each panel indicate three host species, and x, which is a measure of the variance in posterior probabilities [0.95 for differences in preference. Symbols oviposition behavior among individuals within populations (d). The reﬂect host association, as in Fig. 2. For each panel, native-associated preference values in a–c are medians and 95 % credible intervals populations (solid symbols) and alfalfa-associated populations (open from posterior probability distributions estimating preference at circles) are ordered with decreasing values (on the y axis) from left to the population level (full posterior distributions are illustrated in right

Another methodological issue involves potential varia- generation can be highly synchronized. Later in the season tion in host plants used in preference arrays. Intraspecific (when experiments were conducted), emergence is less variation in host quality and chemistry can of course be synchronized and generations overlap, making it more substantial and can even be greater (from the perspective of likely that samples from each population were composed of ovipositing females) than variation among plant species a mix of relatively young and relatively old females, (Singer et al. 2002). To minimize such variation, we col- though this was not quantified. With respect to these lected our primary three plants (alfalfa, the negative con- methodological issues, a comparison involving GLA is trol, and the most common native host, A. canadensis) from again informative. Females from GLA and SUV were the same three locations throughout the course of the collected on the same day, and preference experiments experiments. Furthermore, all assays were conducted in the were conducted at the same time using the same batches of second half of summer, and model comparisons addressing plants. However, these two populations are thoroughly variation among sampling dates within sites did not detect divergent in their preferences (Fig. 4), which is consistent any significant differences. Although there is no particular with both a genetic basis for preference (females from both reason to expect female age to be a source of bias in this populations were associated with roadside alfalfa) and an species, the time window in the latter half of the summer absence of experimental artifacts (such as variation in plant also reduced the possibility of temporal biases in the vigor quality) affecting behavior in cages. or health of females. Early in the season (May or June), it is Assuming factors such as developmental plasticity or possible on a given day to collect mostly fresh or mostly variation in plant quality did not substantially affect the worn (old) females, as the emergence of adults in the first behavior of females, our results raise some interesting

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more deﬁnitive test of these issues, and a quantitative genetics approach would be needed to address the potentially independent genetic basis of preference for different hosts. Beyond questions that can be raised about the process of host range expansion, the observed variation in preference 0.4 0.6 among populations also highlights much that we do not know about the L. melissa system. For example, what

Preference explains the great variation in oviposition behavior observed among alfalfa-associated populations (Fig. 4)? One possibility, that we cannot test at this time, is that

0.0 0.2 populations are of different ages: preference for the native Native Alfalfa Neg. control plant might be lost given sufficient geographic and tem- Fig. 6 Preference for the three kinds of hosts used in preference poral isolation. In this context, the GLA location (where experiments, comparing between native (solid circles) and alfalfa- females exhibited the strongest preference for the native associated (open circles) individuals in analyses pooled across host among the alfalfa populations) might have been col- populations within those two host-associated groups. Values shown onized most recently from a native source. are medians and credible intervals from posterior probability distributions. For both native hosts and alfalfa (the left and middle panels, Perhaps even more intriguing than the variation in respectively), the estimated preferences are different: the probability preference among alfalfa-associated populations is the lack that native-associated females have a greater preference for native of preference exhibited by the native-associated population hosts is 96.7 % (left panel); the probability that alfalfa-associated BHP. Females from BHP were sampled twice, one month individuals have a greater preference for alfalfa is 99.8 % (middle panel). Preferences for the negative control do not differ apart, and showed the same lack of preference on both occasions. Unlike all of the other native populations, the BHP location is a weedy lot in a semi-urban setting questions about the evolution of host preference associated (Bishop, CA, USA). Other L. melissa populations are with the colonization of a novel plant. Most of the alfalfa- known from the immediate vicinity associated with alfalfa associated females exhibited a weaker preference for a (MLF, pers. obs.), thus raising the possibility that BHP is native host (Fig. 6), and in some cases females did not the converse of GLA, a native-associated location that has distinguish between alfalfa and the native plant used in been recently colonized from an alfalfa-associated source. arrays (e.g., at SUV and VCP). These preferences contrast An understanding of the historical and landscape context in with those of native-associated females, as discussed above which focal populations exist will have to be gained using (Fig. 6). Thus, our results generally suggest route ‘‘c’’ from population genetic data in future studies. Fig. 1, a decrease in specificity, as a prominent mode of In summary, the results reported here include interpop- preference evolution for L. melissa. However, the fact that ulation variation in oviposition preference in a system that the native- and alfalfa-associated females did not differ in includes the recent colonization of a novel host plant. Some preference for the negative control suggests the intriguing of the interpopulation variation in oviposition preference possibility that preferences for different hosts can evolve can be understood through host association: females asso- independently (McBride 2007). In other words, preference ciated with the novel host have reduced preference for a for the native host can be lost and preference for the novel native plant and increased preference for a novel host host can be gained without an increase in the tendency to (Fig. 6). These results contribute to our understanding of use a lower-ranked host. Independent evolution of prefer- host range evolution by reinforcing the idea that oviposi- ences for different hosts has been suggested in Euphyrdr- tion behavior is evolutionarily labile and can change in yas editha, where acceptance of a novel host increased with complex ways, including changes in both specificity and no associated change in behavior with respect to discrim- rank order of preference. At the same time, the conclusions ination among phenotypes of an ancestral host (Singer that we present also point toward much that is unknown et al. 1992b). An alternate possibility, suggested by the with respect to the evolution of host range. For example, oscillation hypothesis of Janz and Nylin (2008), would why might natural selection favor an increase in preference have been an increase in generalized behavior (acceptance for any particular host? After all, the novel host is at least of multiple hosts) associated with an expansion of host an acceptable host to females associated with native plants: range. Generalism in diet has been shown to be associated preference for alfalfa was estimated to be around 20 % for with the tendency to colonize exotic hosts in a number of these females. other butterflies (Jahner et al. 2011). A wider array of In locations where multiple host species occur, selection lower-ranked hosts could be included in future assays for a could certainly act on preference, assuming heritable

123 Oecologia (2013) 172:177–188 187 variation in larval performance. However, the novel host is Forister ML, Nice CC, Fordyce JA, Gompert Z (2009) Host range generally an inferior larval resource, and the one location evolution is not driven by the optimization of larval performance: the case of Lycaeides melissa (Lepidoptera: Lycaenidae) studied where both native and novel hosts are used (BWP) and the colonization of alfalfa. Oecologia 160:551–561 is characterized by preference for the native (Fig. 3). Most Forister ML, Gompert A, Nice CC, Forister ML, Fordyce JA (2011) of our populations are geographically isolated on a single Ant association facilitates the evolution of diet breadth in a host, which raises the possibility that selection has favored lycaenid butterfly. Proc R Soc B 278:1539–1547 Gompert Z, Fordyce JA, Forister ML, Shapiro AM, Nice CC (2006) preference for the relevant host through the mechanism of Homoploid hybrid speciation in an extreme habitat. Science increased efficiency in host finding and egg laying. Perhaps 314:1923–1925 preference for alfalfa has slowly increased in alfalfa-asso- Gompert Z, Lucas LK, Fordyce JA, Forister ML, Nice CC (2010) ciated populations because of selection for sensory systems Secondary contact between Lycaeides idas and L. melissa in the Rocky Mountains: extensive admixture and a patchy hybrid that cue in on that host quickly: even if it is a relatively zone. Mol Ecol 19:3171–3192 inferior larval resource, it might still pay for a short-lived Gripenberg S, Mayhew PJ, Parnell M, Roslin T (2010) A meta- butterfly to make efficient decisions with respect to the analysis of preference-performance relationships in phytopha- only host present. Increased efficiency associated with gous insects. Ecol Lett 13:383–393 Hawthorne DJ, Via S (2001) Genetic linkage of ecological special- specialized behavior has been suggested in other species ization and reproductive isolation in pea aphids. 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