Heredity 86 (2001) 243±250 Received 13 November 2000, accepted 13 November 2000

Lack of genetic differentiation among widely spaced subpopulations of a butter¯y with home range behaviour

MARCUS R. KRONFORST* & THEODORE H. FLEMING Department of Biology, University of Miami, Coral Gables, , U.S.A.

We examine seven geographically separate subpopulations of charithonia, a butter¯y with well-documented home range behaviour, in Miami±Dade County, Florida, for genetic di€erentiation using cellulose acetate electrophoresis. These subpopulations exhibit little genetic variation (percent polymorphic loci ˆ 27, average heterozygosity ˆ 0.103) especially in comparison to populations of the same and related from mainland South America. Allele frequencies do not di€er among

the subpopulations in south Florida and estimates of Wright's ®xation index (FST) support that there is no detectable genetic di€erentiation among them. This result supports an earlier ®nding that the dispersal ability of Heliconius butter¯ies may be underestimated. However, it is unlikely that increased dispersal ability alone could account for the lack of genetic di€erentiation observed among subpopulations separated by almost 80 km. Given the likely e€ective population size of these

subpopulations (Ne ˆ 205) and the average generation time of this species in the subtropics (in the range of 30±90 days), this lack of genetic di€erentiation is best explained by current or very recent gene ¯ow following a stepping-stone model. Furthermore, this result provides evidence that the current extensive degree of habitat fragmentation surrounding the city of Miami does not limit gene ¯ow among urban subpopulations of Heliconius charithonia.

Keywords: allozyme electrophoresis, gene ¯ow, genetic di€erentiation, Heliconius charithonia, stepping-stone model, urban fragmentation.

Introduction ¯ies use speci®c foraging routes by which they return daily to sites of adult host (for and ) Butter¯ies of the genus Heliconius (: Nymp- and larval host plants (for egg laying and mating) halidae) are common organisms in New World tropical (Turner, 1971; Ehrlich & Gilbert, 1973; Cook et al., and subtropical forests. This genus is well represented 1976; Mallet & Jackson, 1980; Quintero, 1988). Addi- throughout South and Central America, into , tionally, Heliconius butter¯ies exhibit marked ®delity to and the southern United States. Heliconius butter¯ies night roosts (Turner, 1971; Brown, 1981; Waller & are a major focus of research due to their longevity, Gilbert, 1982; Mallet, 1986a). These behaviours suggest Mullerian , use of pollen as a nutritional that Heliconius butter¯ies travel within a limited range supplement, unpalatability to predators, aposematic throughout their lifetimes. Limited range, in turn, coloration, pupal-mating system, gregarious roosting, suggests that gene ¯ow between geographically separ- striking coevolution with plants of the genus Passi¯ora, ated populations is likely to be low. inter- and intraspeci®c wing colour-pattern variation, Mallet (1986b), however, found that dispersal distances and home-range behaviour (Turner, 1971; Ehrlich & of Heliconius butter¯ies may be substantially underesti- Gilbert, 1973; Gilbert, 1975; Gilbert, 1991). mated. Prereproductive dispersal is common in , so Results of mark±recapture studies suggest that dis- Mallet investigated movement of marked Heliconius erato persal rates in Heliconius butter¯ies are low, apparently individuals from the site of pupal eclosion. He found that as a result of home-range behaviour. Heliconius butter- most dispersal occurs before the ®rst capture of an individual in a typical mark±recapture study. Thus, individuals may disperse some distance (an average of *Correspondence and present address: School of Biological Sciences, 296 m in H. erato; Mallet, 1986b) before they exhibit Section of Integrative Biology, University of at Austin, Austin, home-range behaviour. Additionally, one individual TX, 78712, U.S.A. E-mail: [email protected]

Ó 2001 The Genetics Society of Great Britain. 243 244 M. R. KRONFORST & T. H. FLEMING moved 1000 m between the sites of eclosion and ®rst capture, and 820 m between sites of ®rst and second capture. This example shows that extended dispersal is possible for Heliconius butter¯ies. Mallet (1986b) used his dispersal data to estimate rates of gene ¯ow between populations of H. erato. Using estimates of neighbour- hood deme size, Mallet predicted Wright's ®xation index values (FST values) would range between 0.02 and 0.04, relative to 100 local neighbourhoods. Values this low indicate little genetic di€erentiation between populations. In this study we analyse the e€ect of butter¯y dispersal on gene ¯ow by measuring allele frequencies in several widely spaced subpopulations of H. charitho- nia in south Florida. From these allele frequencies we calculate Wright's (1951) F-statistics, as well as a number of standard statistics to determine whether subpopulations separated by distan- ces much greater than the proposed dispersal distance exhibit signi®cant genetic di€erentiation. In addition, we compare our results with other studies to determine (1) whether our measure of genetic variability (hetero- zygosity) for H. charithonia is typical of the genus Heliconius and butter¯ies in general, (2) what factors might be responsible for any observed patterns of Fig. 1 Map of the seven subpopulations of Heliconius chari- heterozygosity within the subfamily , and thonia included in this study (each indicated by an arrow). (3) whether our estimated pattern of gene ¯ow corres- Math, Matheson Hammock; Deer, Deering Estate; Cast, ponds to that of other species in which both Castellow Hammock; Grey, West Greynolds Park; Trop, dispersal and gene ¯ow have been estimated. Finally, we Tropical Park; Ever, Everglades National Park; New, consider our results in the context of the extensive residential backyard. human population growth, which has led to a highly fragmented natural landscape in south Florida, in order mately 40 min at 200 V using a Tris±glycine bu€er. to determine what e€ect this has had, and potentially After electrophoresis, the plates were stained for the will have, on the pattern of gene ¯ow in this area. appropriate enzyme (Table 1) following standard reci- pes (Harris & Hopkins, 1978; Richardson et al., 1986) scaled to approximately 2 mL total volume, and applied Materials and methods to plates as overlays mixed 1 : 1 with 1.5% agar solution Between April 1997 and January 1998, we collected 20 following Hebert & Beaton (1993). adult H. charithonia for allozyme electrophoresis from After an initial survey for variation at 15 loci, 11 were each of seven subpopulations in Miami±Dade County, found to be consistently scorable. Of these, three were Florida (Fig. 1). Five of the sites are remnant patches of found to be polymorphic in a sample of 24 individuals, native forest (either pineland or tropical hardwood 12 of which were captured from areas outside the study hammock), one site was Gumbo Limbo Hammock in sites and 12 were randomly selected study individuals. Everglades National Park, and the ®nal site was a The remaining eight loci were monomorphic. Enzyme suburban backyard. Butter¯ies were transported live to names, abbreviations, and E.C. numbers for the 11 loci the laboratory, where they were frozen and stored at scored are given in Table 1. The allelic state of each )80°C until processing for electrophoresis. individual was determined for the three polymorphic Tissue extracts were prepared by grinding one-quarter loci, and percentage polymorphic loci (%P), average to one-half of the abdomen of each butter¯y in 250 lL alleles per locus (APL), and Nei's (1978) unbiased of extraction bu€er (Hagen & Scriber, 1991), and the estimate of average heterozygosity (H) were calculated extracts were then centrifuged for 6 min at 9000 ´ g. for each of the seven subpopulations. Supernatant from the centrifuged extracts was applied Since larger areas of habitat are expected to support to thin layer cellulose acetate plates (`Titan III' [94 larger Heliconius populations, and larger populations by 76 mm], Helena Laboratories, Beaumont, TX) for may exhibit greater genetic variation, average hetero- electrophoresis and these were then run for approxi- zygosity was compared to the size of each of the

Ó The Genetics Society of Great Britain, Heredity, 86, 243±250. HELICONIUS POPULATION GENETICS 245

Table 1 Enzymes, abbreviations, and E.C. numbers for the reduction in heterozygosity of an individual due to 11 loci scored in Heliconius charithonia. Note, the eight nonrandom mating within its subpopulation, and monomorphic loci were determined by screening a random Wright's overall inbreeding coecient (FIT), which sample of 24 individuals measures the reduction in heterozygosity of an individ- Poly/ ual relative to the total population, were calculated for E.C. Mono each of the three polymorphic loci. A chi-squared test Enzyme Locus number morphic* was used to determine if the allelic frequencies di€ered signi®cantly among subpopulations. Allelic frequencies Phosphoglucomutase Pgm 5.4.2.2 Poly at each locus for each subpopulation were also tested Phosphoglucose Isomerase Pgi 5.3.1.9 Poly against Hardy±Weinberg expectations. Calculation of Hydroxybutyrate Hbdh 1.1.1.30 Poly heterozygosities, F-statistics, and tests for deviation Dehydrogenase Malate Dehydrogenase Mdh 1.1.1.37 Mono from Hardy±Weinberg equilibrium were performed Fumarate Hydratase Fum 4.2.1.2 Mono using the program TFPGA (Miller, 1997). Peptidase Pep 3.4.11 Mono Mannose-6-Phosphate Mpi 5.3.1.8 Mono Results Isomerase Isocitrate Dehydrogenase Idh 1.1.1.42 Mono Allele frequencies at the three polymorphic loci for each 6-Phosphogluconate 6Pgdh 1.1.1.44 Mono subpopulation are reported in Table 2. There were no Dehydrogenase signi®cant deviations from Hardy±Weinberg expecta- Aspartate Aminotransferase Aat 2.6.1.1 Mono tions at these loci in any subpopulation. Average alleles Arginine Phosphokinase Apk 2.7.3.3 Mono per locus (APL) and average heterozygosity (H) for each * Polymorphic locus determined as frequency of most common subpopulation are given in Table 3. Since the sub- allele <95%. populations shared the same three polymorphic loci as well as the same eight monomorphic loci, the percentage localities from which the individuals were taken using polymorphic loci for each subpopulation was invariant Spearman's rank correlation test. The localities, which at 27.3%. Average alleles per locus varied slightly are primarily urban parks, were ranked in size by among subpopulations, primarily due to two rare alleles. visually comparing their boundaries on a city map. An S- allele at the Pgm locus was present in one The degree of genetic subdivision was estimated by individual in each of the Deer and Trop subpopulations. calculating Wright's ®xation index (FST) for each of the Additionally, an F+ allele at the Pgi locus was present three polymorphic loci using the method of Weir & in one individual in the Ever subpopulation (Table 2). Cockerham (1984). The three estimates were then Average heterozygosity varied slightly among subpop- combined into one jackknife mean FST. FST, which ulations, however, there was no correlation between ranges from 0 to 1, indicates the amount of genetic average heterozygosity and size of each habitat patch subdivision between subpopulations (values near 0 (rs ˆ 0.571, P > 0.05). indicate little genetic subdivision and values near 1 Across all subpopulations, the loci produced similar indicate substantial genetic subdivision). Additionally, values of FST, ranging from )0.0121 to 0.0070 with a Wright's inbreeding coecient (FIS), which measures the jackknife mean of 0.0029, which is not signi®cantly

Table 2 Allele frequencies at the three polymorphic loci scored for each Subpopulation subpopulation of Heliconius Locus Allele Math Deer Cast Grey Trop Ever New v2 P charithonia. Alleles are designated as F+ (fastest migrating allele), F (fast Hbdh M 0.850 0.875 0.875 0.900 0.950 0.925 0.875 2.99 0.811 migrating allele), M (moderate S 0.150 0.125 0.125 0.100 0.050 0.075 0.125 migrating allele), S (slow migrating Pgm F 0.150 0.175 0.200 0.200 0.200 0.225 0.050 14.98 0.243 allele), and S± (slowest migrating M 0.800 0.800 0.700 0.800 0.750 0.650 0.875 allele) with respect to the other alleles. S 0.050 Ð 0.100 Ð 0.025 0.125 0.075 N = 20 for each locus at each site S± Ð 0.025 Ð Ð 0.025 Ð Ð (number of alleles = 2N) Pgi F+ Ð Ð Ð Ð Ð 0.025 Ð 13.84 0.311 F 0.150 0.250 0.125 0.175 0.050 0.150 0.075 M 0.600 0.500 0.575 0.500 0.775 0.600 0.600 S 0.250 0.250 0.300 0.325 0.175 0.225 0.325

Ó The Genetics Society of Great Britain, Heredity, 86, 243±250. 246 M. R. KRONFORST & T. H. FLEMING

Table 3 Average alleles per locus (APL) and Nei's (1978) 27% of the loci were polymorphic. Although this result unbiased estimate of average heterozygosity (H) for each of could be due to the smaller number of loci studied the seven subpopulations of Heliconius charithonia (11 vs. 17), it could also indicate an overall reduction _ in genetic variation in this species in south Florida Subpopulation APL H compared with other species in Central and South Math 1.45 0.107 America. Furthermore, the mean average hetero- Deer 1.45 0.109 zygosity within subpopulations was 10.3% in this study Cast 1.45 0.116 whereas Turner et al. (1979) found that average hetero- Grey 1.36 0.104 zygosity ranged from 9% to 24% for the eight Central Trop 1.55 0.080 and South American species (Fig. 2). Packer et al. Ever 1.55 0.113 (1998) found that average heterozygosity of 56 lepid- New 1.45 0.091 opteran species (not including any Heliconius species) was 10.5%, indicating an increased level of genetic variability in the Central and South American species Table 4 The inbreeding coecient (F ), overall inbreeding IS as compared to south Florida subpopulations of coecient (FIT) and ®xation index (FST) for each of the three polymorphic loci, the jackknife mean of each, and the H. charithonia. 95% con®dence interval as determined by bootstrapping The low heterozygosity of H. charithonia in this study over loci. These F-statistics were estimated using the is probably due to the geographical location of these method of Weir & Cockerham (1984) subpopulations on the peninsula of Florida, where the in¯ux of genetic variation from dispersing individuals is Locus FIS FIT FST presumably less than in the inland areas of South and Hbdh )0.0310 )0.0435 )0.0121 Central America. This explanation is strongly supported Pgm )0.0608 )0.0534 0.0070 by comparing the average heterozygosity for these Pgi )0.0087 )0.0049 0.0038 subpopulations to that of other H. charithonia popula- Mean )0.0265 )0.0235 0.0029 tions and to that of its sibling species, H. peruvianus 95% C. I. )0.0087 to )0.0049 to 0.0070 to (Jiggins & Davies, 1998). Populations of H. charithonia )0.0608 )0.0534 )0.0121 from the Caribbean islands of Jamaica and have heterozygosities identical to that of the south Florida subpopulations (Fig. 2) and the extent of geographical di€erent from 0 (Table 4). Therefore, at most only a isolation on these Caribbean islands is probably similar fraction of a percentage of the overall genetic diversity to the isolation experienced by the subpopulations on of these subpopulations results from variation among the Florida peninsula. In contrast to these isolated subpopulations (Table 4). In support of this conclusion, populations, H. charithonia from the South American a chi-squared analysis revealed that the allele frequen- mainland country of Ecuador exhibit considerably cies for the three polymorphic loci were not signi®cantly greater heterozygosity (Fig. 2). As expected, popula- di€erent among the seven subpopulations (Table 2). FIS tions of H. peruvianus have a heterozygosity inter- values were small but consistently negative (Table 4), mediate to the mainland and island populations of indicating that at each of the three polymorphic loci H. charithonia (Fig. 2). The genetic variation of H. peru- there was an excess of heterozygotes. Given the negative vianus is less than that of parapatric populations of values of FIS and the very small values of FST, FIT is also H. charithonia probably as a result of its limited negative for all three polymorphic loci (Table 4). As geographical distribution, yet the heterozygosity of with FIS, these negative values indicate that there was an H. peruvianus is greater than that of the island and excess of heterozygotes at each of the three polymorphic Florida populations as a result of occasional genetic loci for the entire population. input due to H. peruvianus/H. charithonia hybridization (as documented by Jiggins & Davies, 1998). Notice that Discussion this scenario is partially mirrored by that of H. himera,a recently derived taxon in the same region of Ecuador Our results suggest that subpopulations of H. charitho- and Peru. Like H. peruvianus, H. himera exhibits a nia in south Florida contain relatively little genetic slightly decreased (although not signi®cantly so) hetero- variation and are not genetically subdivided. Our results zygosity in comparison to its widespread parapatric di€er in some respects from a previous allozyme study of sister species H. erato (Jiggins et al., 1997). Heliconius butter¯ies. Turner et al. (1979) reported that Our results indicate that there is no genetic di€eren- 47% to 76% of the 17 loci they studied in eight species tiation among the seven subpopulations of H. charitho- of Heliconiine butter¯ies were polymorphic. Here, only nia. This is most likely due to the homogenizing e€ect of

Ó The Genetics Society of Great Britain, Heredity, 86, 243±250. HELICONIUS POPULATION GENETICS 247

Fig. 2 Average heterozygosity for the eight Heliconiine species studied by Turner et al. (1979) (white), H. erato and H. himera studied by Jiggins et al. (1997) (diagonal lines), H. charithonia and H. peruvianus studied by Jiggins & Davies (1998) (grey), and H. charithonia from this study (black). gene ¯ow. Support for this conclusion comes from the on neighbourhood deme size, was 0.02±0.04. Our negative values of inbreeding coecients, indicating an estimate of FST for H. charithonia falls well below this excess of heterozygotes. If subpopulations were genetic- range. Although this ®nding supports the hypothesis ally isolated, we would expect to ®nd evidence for that Heliconius butter¯ies disperse a greater distance inbreeding (e.g. a de®ciency of heterozygotes indicated than was once believed based on their home range by positive inbreeding coecients). behaviour, Mallet found that even after taking early In their review of literature on genetic variation in dispersal into account, the average distance travelled natural insect populations, Peterson & Denno (1998) remained relatively low. Yet, we have shown that rates identify a number of factors which could potentially of gene ¯ow remain high at distances of up to almost in¯uence gene ¯ow among insect populations. A major 80 km between subpopulations. Although it is likely factor, obviously, is the dispersal behaviour of insects. that H. charithonia individuals disperse even greater Thus, the dispersal-gene ¯ow hypothesis holds that there distances than those determined for H. erato, it remains should be a positive correlation between the extent of doubtful that increased movement alone can account for dispersal and levels of gene ¯ow. Indeed, such a this ®nding. correlation was found (Peterson & Denno, 1998). Given In other insect systems, in which both dispersal and this, our results suggest that, at a spatial scale of gene ¯ow distances are known, gene ¯ow typically 5±80 km, dispersal of individuals is a potential explan- occurs over greater distances than those exhibited by ation for the low genetic di€erentiation observed among dispersing individuals (e.g. McCauley & Eanes, 1987; the subpopulations of H. charithonia. Rosenberg, 1989; Michalakis et al., 1993; Peterson, This ®nding supports that of Mallet (1986b), who 11995, 1996). In many instances, these results are not showed that dispersal distances in Heliconius butter¯ies explained by direct gene ¯ow between populations, but probably have been substantially underestimated by rather by genes being dispersed in a `stepping-stone' mark±recapture studies. Mallet's estimate of FST, based fashion, whereby continued gene ¯ow between neigh-

Ó The Genetics Society of Great Britain, Heredity, 86, 243±250. 248 M. R. KRONFORST & T. H. FLEMING bouring populations spreads genes greater distances Considering the e€ect of alone, we can than the dispersal ability of the individuals might predict predict how long it will take to reach various degrees (Kimura, 1953). Therefore, rather than individuals of genetic di€erentiation using the relationship FST ˆ t carrying genes directly between two distant populations, 1 1 1 (Hartl & Clark, 1989). Although these genes are indirectly transported through geo- 2 Ne‡1=2† graphically intermediate populations. FST has a theoretical maximum of 1, observed values Although this pattern of gene ¯ow occurring over a are usually much less than this. Wright (1978) suggested larger distance than the dispersal of individuals is that FST values which exceed 0.25 indicate very great common and supports a stepping-stone model of gene genetic di€erentiation. Given complete isolation, two ¯ow, other explanations are possible. With few excep- subpopulations should exhibit genetic divergence of the tions, allozymes are considered to be selectively neutral, order of FST ˆ 0.3 in the span of 12 to 36.1 years, but if this were not the case for depending on the estimate of generation time. Likewise, particular allelic states could have a homogenizing e€ect two subpopulations should exhibit a moderate level on subpopulations. Additionally, if these subpopula- of di€erentiation (FST ˆ 0.1) in the span of 3.6 to tions represent fragments of a recently continuous 10.7 years or a low degree of genetic di€erentiation population, the allele frequencies might still remain (FST ˆ 0.05) in the span of 1.7 to 5.2 years. In the case similar among them. Pashley (1989) and Bossart & of H. charithonia in south Florida, the FST among all Pashley Prowell (1998) warn against making inferences subpopulations is essentially zero and all pairwise FST of extensive gene ¯ow based solely on allozyme data, on comparisons are below 0.05. Therefore, if gene ¯ow the grounds that if no di€erences between groups are had ceased among any of these subpopulations one detected, high levels of gene ¯ow cannot be assumed thousand, one hundred, or even 20 years ago there because markers may not be re®ned enough to distin- would be some detectable level of di€erentiation today. guish current gene ¯ow from gene ¯ow that ceased Interestingly, the history of urbanization in south perhaps hundreds or thousands of years ago. Although Florida suggests that if urban fragmentation could this is a fair warning, we believe that we can attribute the limit gene ¯ow in this species, it should have already lack of genetic di€erentiation observed here to current happened and we should see the resulting genetic or very recent gene ¯ow given the relatively low density di€erentiation by now. of individuals in local H. charithonia populations and This study was not carried out in a typically natural the potentially rapid generation time for this species in area, but rather in the suburban areas surrounding the the subtropics. city of Miami. In these areas, the population of H. Based on Quintero's (1988) data for a population of charithonia is subject to urban fragmentation. Therefore, the seven subpopulations in this study are not geo- H. charithonia in Puerto Rico, we estimate Ne to be approximately 205, after accounting for population size graphically separated solely by a region of suitable ¯uctuations and a consistent male-biased sex ratio in habitat, but rather, by an expanse of developed land and the adult stage. Estimating generation time is a bit more major roadways in many cases. Heavy drainage and complex due to the potentially extended life span of development of the areas around the city of Miami Heliconius butter¯ies (Gilbert, 1972 and Ehrlich & began in the 1920s, and by 1975 only 4% of Miami Gilbert, 1973.) and the life-long reproduction exhibited Rockland forest remained outside of Everglades by females exposed to sucient pollen resources National Park (Shaw, 1975). Thus, fragmentation of (Dunlap-Pianka et al., 1977). However, the minimum this habitat occurred relatively recently but was swift generation time, measured as the mean time for a and extensive. In light of this fragmentation, it is female to hatch, grow, pupate, mate and lay an egg of interesting to note that gene ¯ow remains high among her own is approximately 30 days in this genus. subpopulations. Apparently fragmentation of the nat- Measured in the same way, the maximum generation ural habitat has not reached such a level as to restrict time can potentially exceed ®ve months if females are gene ¯ow among many local neighbourhoods. Given the exposed to unlimited pollen. In the subtropical area of stepping-stone model by which gene ¯ow is presumably south Florida the primary pollen resources used by occurring, it is likely that subpopulations that occur in Heliconius species in South and Central America open urban lots, small parks, and residential backyards (Psiguria and Gurania vines of the family Cucurbita- provide the means for gene ¯ow among the larger ceae) are absent and therefore pollen resources are natural areas studied here such as Matheson Hammock, likely to be limited. Therefore, we estimate the maxi- the Deering Estate, and Everglades National Park. mum generation time to be in the neighbourhood of 90 Although current levels of habitat fragmentation may days and the average is probably on the lower end of not a€ect gene ¯ow, continued human growth and this range (approximately 45 days). expansion in Miami±Dade County could have an

Ó The Genetics Society of Great Britain, Heredity, 86, 243±250. HELICONIUS POPULATION GENETICS 249 increasing impact on these subpopulations. More devel- HARRIS, H.AND HOPKINSON , D. A. 1978. Handbook of Enzyme opment could potentially cut certain paths of gene ¯ow Electrophoresis in Human Genetics. American Elsevier, New and e€ectively separate the now continuous population York. , D. L.AND CLARK , A. G. 1989. Principles of Population into two or more gene-exchanging units. HARTL Genetics, 2nd Edn. Sinauer, Sunderland, MA. HEBERT, P. D. N.AND BEATON , M. J. 1993. Methodologies for Acknowledgements Allozyme Analysis Using Cellulose Acetate Electrophoresis. Helena Laboratories, University of Guelph, Ontario. We thank K. Spitze for his generous support of this JIGGINS, C. D.AND DAVIES , N. 1998. Genetic evidence for a project in both time and resources, D. Serrano for sibling species of Heliconius charithonia (Lepidoptera; collecting butter¯ies from the New subpopulation, and ). Biol. J. Linn. Soc., 64, 57±67. B. Wee for assistance in preparing the ®gures. We also JIGGINS, C. D., MCMILLAN, W. O., KING, P.AND MALLET , J. 1997. thank D. Altshuler, M. Cardoso, L.E. Gilbert, D. Kapan, The maintenance of species di€erences across a Heliconius P. Schappert, J.R.G. Turner, and an anonymous hybrid zone. Heredity, 79, 495±505. reviewer for discussion and comments on the manuscript KIMURA, M. 1953. `Stepping-stone' model of population. Ann. Report Nat. Inst. Genet., Japan, 3, 62±63. and C. Jiggins for sharing his genotype frequency data MALLET, J. 1986a. Gregarious roosting and home range in for the Ecuadorian and Caribbean H. charithonia and Heliconius butter¯ies. Nat. Geo. Res., 2, 198±215. H. peruvianus. Butter¯ies were collected under permits MALLET, J. 1986b. Dispersal and gene ¯ow in a butter¯y granted by the Miami-Dade County Park and Recrea- with home range behavior: Heliconius erato (Lepidoptera: tion Department and the National Park Service. Fund- Nymphalidae). 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