The diapause response of Rhagoletis pomonella to varying environmental conditions and its significance for geographic and host plant‐related adaptation

The recent shift of Rhagoletis pomonella Walsh (Diptera: Tephritidae) from its ancestral host hawthorn to apple is a model for incipient sympatric speciation in action. Previous studies have shown that changes in the over‐wintering pupal diapause are critical for differentially adapting R. pomonella flies to a difference in the fruiting times of apples vs. hawthorns, generating ecologically based reproductive isolation. Here, we exposed pupae of the hawthorn race to various combinations of pre‐ and over‐wintering rearing conditions and analyzed their effects on eclosion time and genetics. We report certain unexpected results in regards to a combination of brief pre‐winter and over‐wintering periods indicative of gene*environment interactions requiring a reassessment of our current understanding of R. pomonella diapause. We present a hypothesis that involves physiological factors related to stored energy reserves in pupae that influences the depth and duration of Rhagoletis diapause. This ‘pupal energy reserve’ hypothesis can account for our findings and help clarify the role host plant‐related life history adaptation plays in phytophage biodiversity.     

Influence of protein hydrolysate baits on oviposition in the apple maggot, Rhagoletis pomonella

Ovipositional responses of apple maggot (AM), Rhagoletis pomonella (Walsh), females were studied in the laboratory on apples (var: Golden Delicious) treated with different rates of four protein hydrolysate baits in choice and no-choice tests. Protein hydrolysate baits at rates of 0.5 and 1% had no significant effect, but oviposition was greatly reduced at higher rates of 5 and 10%. Apple maggot females exposed to apples treated with protein hydrolysate baits at a rate of 10% made 41–71% fewer punctures and laid 41–73% fewer eggs than in untreated control. No oviposition activity was shown on apples treated with 25 and 100% Nulure®. In no-choice tests the AM females laid 75–96% fewer eggs in apples treated with 10 and 25% Nulure compared to controls and no oviposition occurred in apples treated with 100% Nulure. Apple maggot females arrived in similar numbers on apples treated with 10% Nulure and untreated apples, but only 5% of those arriving on Nulure-treated apples showed ovipositor boring with no egg deposition while 60% of females arriving on untreated apples showed ovipositor boring activity and laid an average of 2.5 eggs per apple. In another experiment, individual AM females displayed similar behavioral responses to 10% Nulure-treated apples; none of the 56 females tested on treated apples displayed ovipositor boring activity, but 59% of the females (N=56) tested on untreated apples displayed ovipositor boring within 5 min of their arrival. Ninetyeight percent of AM females stayed and fed on fruit surfaces for 5 min on Nulure-treated apples without ovipositor boring compared to only 2% on untreated apples. Of the females that arrived on untreated apples, 39% flew away within 5 min without ovipositor boring compared to only 2% of those that arrived on Nulure-treated apples. Results of these two behavioral experiments suggest that upon arrival on a protein bait-treated apple, an apparent change of behavior occurs in AM females and instead of attempting to oviposit, they attempt to feed on fruit surfaces resulting in reduced oviposition activity. These results indicate that the feeding and oviposition-related activities of AM females are probably mutually exclusive and that the feeding behavior preempts oviposition activities on host fruits treated with higher rates of protein hydrolysate baits.     

Geographic variation in fruit volatiles emitted by the hawthorn Crataegus mollis and its consequences for host race formation in the apple maggot fly,Rhagoletis pomonella

The shift of the apple (AP) maggot fly, Rhagoletis pomonella (Walsh) (Diptera: Tephritidae), from its ancestral host downy hawthorn, Crataegus mollis (DH) (Torr. & A. Gray) Scheele, to introduced domesticated AP, Malus domestica Borkh. (both Rosaceae), is a model for ecological divergence and incipient sympatric speciation with gene flow. However, a portion of the variation contributing to the sympatric host shift from DH to AP appears to have a different biogeographic history, pre‐dating the shift. One potential source of standing variation may trace to a number of different native hawthorn species infested by R. pomonella in the southern USA, where the AP‐attacking race is absent. Herein, we investigate this possibility for the southern red hawthorn (SR) endemic to Texas, Crataegus mollis var. texana (Buckl.), which has been described as a member of the Molles series that includes the more northern distributed DH. We report results from chemical analyses of host fruit volatiles, fly behavioural responses to synthetic fruit blends, and microsatellite surveys of fly populations, implying that R. pomonella infesting SR may behaviourally and genetically represent a native host race differing from the DH‐infesting fly. No fly reared from SR responded to AP fruit volatiles in flight tunnel assays. However, coupled gas chromatographic‐electroantennographic detection (GC‐EAD) profiles for SR fruit contain all five of the component esters that comprise the standard AP volatile blend inducing behavioural orientation for AP‐infesting flies, compounds that appear to be largely missing from volatile profiles for DH fruit. Thus, SR‐infesting flies do not represent a source for a preassembled AP‐accepting phenotype. However, they may help explain why the ancestral DH race that shifted to AP in the northeastern USA had the ability to recognize AP fruit esters, potentially enabling the shift to AP. Our results highlight how categorizing speciation into different geographic modes may not adequately describe the evolutionary origins of important genetic variation fuelling adaptive radiation and the genesis of new biodiversity.     

Specificity of olfactory responses in the tephritid fruit fly, Rhagoletis pomonella

Electroantennograms (EAG) were recorded from, and behavior observed of female apple maggot flies, Rhagoletis pomonella (Walsh) (Diptera: Tephritidae), in response to over 60 individual esters. For acetates through decanoates, 2 methylbutyrates, and isobutyrates, we tested homologous series of systematically altered chain lengths. Most of the compounds had been isolated from behaviorally active fractions derived from extracts of volatiles produced by host fruits of R. pomonella. For the acetates through hexanoates, maximum EAG amplitudes were elicited by esters with chain lengths of 9 carbons and for the heptanoates through nonanoates, by 10 carbon esters. Recovery time, or the rate at which the EAG trace returned to the baseline following maximum depolarization, was slowest for straight chain esters that were 9–11 carbons long. Branching of the chain by addition of a methyl group to either side of the ester resulted in a decline in amplitude and a faster recovery time. Compared to EAG results, only 5 esters (butyl and pentyl hexanoate; propyl and butyl heptanoate; propyl octanoate) were highly active in wind tunnel bioassays, demonstrating (1) the hazard of assigning significance to EAG-active compounds without accompanying behavioral data, but more importantly; (2) a high degree of olfactory specificity. Maximum behavioral response was contingent upon the following rules regarding size and structure of the molecule. The ester must (1) be a straight chain; (2) be 10–11 carbons in length; (3) have an acid portion of 6–8 carbons and an alcohol portion of 3–5 carbons. One of the active esters, butyl hexanoate, appears in significant concentrations in the headspace of host fruit and, because this ester elicits such a pronounced behavioral response, our results suggest that R. pomonella is adapted to perception of a compound that is typical of its hosts.

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Résumé Les réactions de R. pomonella à plus de 60 esters différents ont été enregistreées par électroantennogrammes (EAG) et par ovservation du comportement. Pour les acétates, avec les décanoates, les 2-méthylbutyrates et les iso butyrates, nous avons examiné des séries homologues de chaînes aux longueurs systématiquement altérées. La plupart de ces composés ont été isolés à partir des fractions actives sur le comportement, issues des extraits des substances volatiles des fruits des plantes hôtes de cette téphritidae. Pour les acétates, avec les hexanoates, les EAG aux plus grandes amplitudes ont été obtenus avec des esters dont la longueur des chaînes est de 9 carbones, et avec les heptanoates et les nonanoates pour les esters à 10 carbones. Le temps de récupération ou temps mis par l'EAG pour revenir à l'ordonnée de départ après dépolarisation maximum, a été plus lent pour les esters à chaînes droites avec 9 à 11 carbones. La ramification de la chaîne par addition d'un groupe méthyl de chaque côté de l'ester a provoqué une réduction de l'amplitude et une accélération de la récupération. 5 esters seuls provoquent une forte réaction dans le tunnel à vent (hexanoates butilique et pentylique, heptanoates propylique et butylique, octanoate propylique). Ceci montre (1) le risque qu'il y a à attribuer une signification aux substances provoquant une réaction en EAG, lorsqu'il n'y a pas d'observations comportementales parallèles, et, surtout; (2) le degré élevé de spécificité olfactive.La résponse comportementale optimale obéit aux règles suivantes concernant la taille et la structure de la molécule. L'ester doit avoir: (1) une chaîne linéaire; (2) 10–11 carbone de longueur; (3) une portion acide de 6 à 8 C et une portion alcool de 3 à 5 C. Le butyl hexanoate, l'un des esters actifs, présent en concentration suffisante près de fruits utilisés, provoque pour cette raison une réponse comportementale tellement marquée, que nos résultats suggèrent que R. pomonella est adapté à la perception des substances caractéristiques de leurs hôtes.

     

It''s about time: the evidence for host plant-mediated selection in the apple maggot fly, Rhagoletis pomonella, and its implications for fitness trade-offs in phytophagous insects

The apple maggot fly, Rhagoletis pomonella, Walsh (Diptera: Tephritidae), provides a unique opportunity to address the issue of host-related fitness trade-offs for phytophagous insects. Rhagoletis pomonella has been controversial since the 1860's when Benjamin Walsh cited the fly's shift from hawthorn (Crataegus spp.) to apple (Malus pumila) as an example of an incipient sympatric speciation event. Allozyme and mark-release-recapture studies have subsequently confirmed the status of apple and hawthorn flies as partially reproductively isolated and genetically differentiated host races, the hypothesized initial stage in sympatric divergence. Here, we review the ecological and genetic evidence for host-plant mediated selection in R. pomonella. We reach the following three major conclusions: First, although developmental timing is not everything, it is a good deal of the story. Differences in the fruiting phenologies of apple and hawthorn trees exert different selection pressures on the diapause and eclosion time characteristics of the host races. In particular, the 3-week earlier mean fruiting phenology of apples in eastern North America appears to select for a slower rate of metabolism or deeper pupal diapause in apple than hawthorn flies. Second, host-related fitness trade-offs for R. pomonella may not be due to disruptive selection affecting any one specific life-history stage. Rather, it is the sum total of directional selection pressures acting across different life-stages that generates divergent selection on apple and hawthorn flies. For example, selection favors the alleles Me 100, Acon-2 95 and Mpi 37 (or linked genes) in the larval stage in both host races. However, these same alleles are disfavored in the pupal stage to follow, where they correlate with early adult eclosion, and by inference premature diapause termination. Because apple trees fruit an average of 3 weeks earlier than hawthorn trees, this counter-balancing selection is stronger on apple-fly pupae. The net result is that the balance of selective forces is different between apple and hawthorn flies, helping to maintain the genetic integrity of the host races in sympatry in the face of gene flow. Finally, natural R. pomonella populations harbor a good deal of genetic variation for development-related traits. This variation allows fly populations to rapidly respond to temporal vagaries in local environmental conditions across years, as well as to broad-scale geographic differences that exist across the range of the species. Perhaps most importantly, this variation gives R. pomonella the flexibility to explore and adapt to novel plants. Taken together, our results underscore how difficult it can be to document host plant-related fitness trade-offs for phytophagous insects due to the need to consider details of the entire life-cycle of a phytophagous insect. Our findings also show how reproductive isolation can arise as a by-product of host-associated adaptation in insects, a central theme for models of sympatric speciation via host shifts.     

Genetic variation among apple and hawthorn host races of Rhagoletis pomonella across an ecological transition zone in the Mid-Western United States

In previous studies, we have shown that apple and hawthorn populations of Rhagoletis pomonella (Diptera: Tephritidae) represent partially reproductively isolated and genetically differentiated host races; a result consistent with predictions of sympatric speciation models. The geographic pattern of allozyme variation for these flies is complex, however, as inter-host differences are superimposed on latitudinal allele frequency clines within the races. In addition, pronounced allele frequency shifts exist among R. pomonella populations across three major ecological transition zones in the mid-western United States. This suggests that selection related to environmental heterogeneity is responsible for the allele frequency shifts, but does not rule out secondary contact as an alternative possibility. Resolution of this issue is important, because if secondary contact is involved, then we would have to reassess the relationship host race formation has with speciation in the R. pomonella group.Here, we present results from a detailed genetic analysis of fly populations spanning the deciduous/prairie transition zone near the border between the states of Wisconsin and Illinois. Allele frequencies for hawthorn populations within the zone formed spikes, rather than the expected steps, and these frequency peaks correlated with variation in local ambient temperature conditions. Ambient temperature, and not secondary contact, therefore appears to be an important determinant of the shape of R. pomonella allele frequency clines. Allele frequency heterogeneity was also observed among apple populations, but was less pronounced compared to that for hawthorn flies. This suggests that ambient temperature differentially affects the host races, possibly through differences in the fruiting phenologies of apple and hawthorn trees. Several pairs of linked loci displayed concordant allele frequency changes and were in disequilibrium among both apple and hawthorn populations along the Wisconsin/Illinois transect. Although we do not know the reason for the observed pattern of disequilibrium, site to site variation in levels of inter-host migration, coupled with selection, seem the most likely explanations. We conclude by discussing how host specific adaptations, such as those associated with ambient temperature, may interact with host recognition traits to drive the sympatric speciation process for R. pomonella group flies.     

Radiation and divergence in the Rhagoletis pomonella species group: inferences from allozymes

The Rhagoletis pomonella species group has for decades been a focal point for debate over the possibility of sympatric speciation via host shift. Here I present the first extensive analysis of genetic (allozyme) divergence in the pomonella group, including all known taxa/populations except the allopatric Mexican population of R. pomonella. The phylogeny is estimated for all four described species (pomonella, mendax, zephyria, and cornivora) plus two undescribed species (the "flowering dogwood fly" and "sparkleberry fly"). Allozyme data for two additional populations of uncertain status (the "plum fly" and "mayhaw fly") are presented for the first time. Two data sets were analyzed, one for 17 loci from 77 populations and one for an additional 12 loci for a subset of 12 of these populations, with more than 4000 flies analyzed in total. Interspecific Nei unbiased genetic distances were generally small, being as low as 0.040. No fixed autapomorphic alleles beyond those already known for R. cornivora and R. zephyria were revealed in the new data, but several loci displaying frequency patterns useful in discriminating the species were discovered. The phylogenetic placement of the flowering dogwood fly differed depending on whether a molecular clock was assumed (UPGMA of Nei distance) or not assumed (frequency parsimony) for tree building. Other than this, however, trees under either assumption were essentially identical. The best tree was used to test the prediction of the sympatric speciation hypothesis that sister taxa should be broadly sympatric. This prediction was not rejected, but the best tree was weakly supported by bootstrap analysis. An unexpected finding was that R. pomonella populations representing ends of its strong latitudinal clines did not cluster together. One possible explanation is that the current R. pomonella is the result of a genetic fusion of two previously isolated, genetically differentiated populations. Such a fusion prior to the origin of the other species in the group could contribute to the poor resolution of the phylogeny.     

A second case of genetic host races in Rhagoletis? A population genetic comparison of sympatric host populations in the European cherry fruit fly,Rhagoletis cerasi

Despite an increasing acceptance in the biological community for sympatric speciation as a mode of species formation, well documented examples of sympatrically evolved ‘incipient species’ remain rare. The sympatric host races of apple maggot, Rhagoletis pomonella (Walsh), represent one of the most prominent case studies for sympatric speciation via a host shift. The European cherry fruit fly, R. cerasi (L.), shows strong ecological similarities to R. pomonella: (1) infestation of two different host plants, Lonicera xylosteum L. and Prunus avium L., and (2) divergent phenological and behavioral adaptations of flies on different hosts. The population genetic study presented here addresses whether the host associated populations of R. cerasi also represent genetically differentiated true host races. Out of a total of 29 allozyme loci examined, six were polymorphic and used to analyze six sympatric pairs of R. cerasi populations on Lonicera and Prunus from Switzerland and Germany. A direct comparison of allele frequencies between sympatric sites showed no pattern indicative of host races in R. cerasi. However, the hierarchical F‐statistic for one locus, mannose 6‐phosphate isomerase (Mpi), showed significant population differentiation that was in accordance with host race differentiation. Mpi is one of several loci that are also diagnostic for host race differentiation in R. pomonella. Results from Mpi suggest the formation of sympatric host races in R. cerasi, but additional polymorphic markers are necessary.     

A field test for host fruit odour discrimination and avoidance behaviour for Rhagoletis pomonella flies in the western United States

Prezygotic isolation due to habitat choice is important to many models of speciation-with-gene-flow. Habitat choice is usually thought to occur through positive preferences of organisms for particular environments. However, avoidance of non-natal environments may also play a role in choice and have repercussions for post-zygotic isolation that preference does not. The recent host shift of Rhagoletis pomonella (Diptera: Tephritidae) from downy hawthorn, Crataegus mollis, to introduced apple, Malus domestica, in the eastern United States is a model for speciation-with-gene-flow. However, the fly is also present in the western United States where it was likely introduced via infested apples ≤ 60 years ago. R. pomonella now attacks two additional hawthorns in the west, the native C. douglasii (black hawthorn) and the introduced C. monogyna (English ornamental hawthorn). Flight tunnel tests have shown that western apple-, C. douglasii- and C. monogyna-origin flies all positively orient to fruit volatile blends of their respective natal hosts in flight tunnel assays. Here, we show that these laboratory differences translate to nature through field-trapping studies of flies in the state of Washington. Moreover, western R. pomonella display both positive orientation to their respective natal fruit volatiles and avoidance behaviour (negative orientation) to non-natal volatiles. Our results are consistent with the existence of behaviourally differentiated host races of R. pomonella in the west. In addition, the rapid evolution of avoidance behaviour appears to be a general phenomenon for R. pomonella during host shifts, as the eastern apple and downy hawthorn host races also are antagonized by non-natal fruit volatiles.     

Hawthorn-infesting populations of Rhagoletis pomonella in Mexico and speciation mode plurality

Categorizing speciation into dichotomous allopatric versus nonallopatric modes may not always adequately describe the geographic context of divergence for taxa. If some of the genetic changes generating inherent barriers to gene flow between populations evolved in geographic isolation, whereas others arose in sympatry, then the mode of divergence would be mixed. The apple maggot fly, Rhagoletis pomonella, has contributed to this emerging concept of a mixed speciation mode "plurality." Genetic studies have implied that a source of diapause life-history variation associated with inversions and contributing to sympatric host race formation and speciation for R. pomonella in the United States may have introgressed from the Eje Volcanico Trans Mexicano (EVTM; a.k.a. the Altiplano) in the past. A critical unresolved issue concerning the introgression hypothesis is how past gene flow occurred given the current 1200-km disjunction in the ranges of hawthorn-infesting flies in the EVTM region of Mexico and the southern extreme of the U.S. population in Texas. Here, we report the discovery of a hawthorn-infesting population of R. pomonella in the Sierra Madre Oriental Mountains (SMO) of Mexico. Sequence data from 15 nuclear loci and mitochondrial DNA imply that the SMO flies are related to, but still different from, U.S. and EVTM flies. The host affiliations, diapause characteristics, and phylogeography of the SMO population are consistent with it having served as a conduit for gene flow between Mexico and the United States. We also present evidence suggesting greater permeability of collinear versus rearranged regions of the genome to introgression, in accord with recent models of chromosomal speciation. We discuss the implications of the results in the context of speciation mode plurality. We do not argue for abandoning the terms sympatry or allopatry, but caution that categorizing divergence into either/or geographic modes may not describe the genetic origins of all species. For R. pomonella in the United States, the proximate selection pressures triggering race formation and speciation stem from sympatric host shifts. However, some of the phenological variation contributing to host-related ecological adaptation and reproductive isolation in sympatry at the present time appears to have an older history, having originated and become packaged into inversion polymorphism in allopatry.     

Potential geographical distribution of Rhagoletis pomonella (Diptera: Tephritidae) in China

Abstract Apple maggot fly, Rhagoletis pomonella (Walsh) is a major pest causing considerable economic losses of fruits in North America. During the development of international trade, apple maggot fly has become a threat to Chinese agriculture. In this study, CLIMEX and ArcGIS were used to predict the potential geographical distribution of apple maggot fly in China. The parameters used in CLIMEX for apple maggot fly were derived from ecological data and the present geographical distribution of apple maggot fly in North America. Then the potential distribution map in China was presented based on the adjusted values of these parameters. The results showed that apple maggot fly has a wide potential distribution area in China; 47.5% of 748 meteorological stations presented high or medium suitability of pest establishment. These high suitable stations are mainly located in northeast, southwest and northwest of China, such as Liaoning, Shandong, Gansu and Shaanxi Provinces. These areas are also the central regions of apple, pear and peach production in China. Two hundred and twenty‐five stations (30.1%) in western and southern China, such as Tibet, Qinghai, Guangdong, Guangxi, Hainan and Taiwan, were unsuitable for establishment of apple maggot fly populations. In order to prevent the introduction of apple maggot fly in China, the present plant quarantine measures should be enhanced, especially in the areas with high suitability for the presence of apple maggot fly.     

Isolation and characterization of microsatellite loci from the apple maggot fly Rhagoletis pomonella (Diptera: Tephritidae)

We report the isolation and development of 81 novel primers for amplifying microsatellite loci in the Rhagoletis pomonella sibling species complex, and the sequencing, characterization and analysis of basic population genetic parameters for nine of these genes. We also report the successful cross‐species amplification of several of these loci. The R. pomonella sibling species complex is a textbook example of genetic differentiation in sympatry via host‐plant shifting. Microsatellite markers can be useful for mapping host‐plant‐associated adaptations in Rhagoletis that generate reproductive isolation and facilitate speciation, as well as for resolving the genetic structure and evolutionary history of fly populations.     

Rapid and repeatable host plant shifts drive reproductive isolation following a recent human-mediated introduction of the apple maggot fly,Rhagoletis pomonella

Ecological speciation via host-shifting is often invoked as a mechanism for insect diversification, but the relative importance of this process is poorly understood. The shift of Rhagoletis pomonella in the 1850s from the native downy hawthorn, Crataegus mollis, to introduced apple, Malus pumila, is a classic example of sympatric host race formation, a hypothesized early stage of ecological speciation. The accidental human-mediated introduction of R. pomonella into the Pacific Northwest (PNW) in the late 1970s allows us to investigate how novel ecological opportunities may trigger divergent adaptation and host race formation on a rapid timescale. Since the introduction, the fly has spread in the PNW, where in addition to apple, it now infests native black hawthorn, Crataegus douglasii, and introduced ornamental hawthorn, Crataegus monogyna. We use this “natural experiment” to test for genetic differentiation among apple, black, and ornamental hawthorn flies co-occurring at three sympatric sites. We report evidence that populations of all three host-associations are genetically differentiated at the local level, indicating that partial reproductive isolation has evolved in this novel habitat. Our results suggest that conditions suitable for initiating host-associated divergence may be common in nature, allowing for the rapid evolution of new host races when ecological opportunity arises.     

Life history phenology differences between southern and northern populations of the apple maggot fly, Rhagoletis pomonella

The textbook version of sympatric host race formation in Rhagoletis pomonella (Walsh) (Diptera: Tephritidae) features an ancestral hawthorn-infesting race and a derived apple race differing in life-history timing and response to host fruit odor. However, previous research has focused largely on northeastern North America. To investigate life-history timing in poorly studied southeastern North American populations with very diverse hosts, we performed common garden experiments on emergence time on six southern R. pomonella populations and four other R. pomonella species-group populations. Findings were: (1) the diverse adult emergence times of southern populations resulted largely from genetic differences; (2) some southern populations had bimodal emergence curves; (3) correlations of allozyme markers and emergence times were uncommon in southern populations; and (4) mean emergence times from common garden rearings and dates of field collection of the populations were strongly correlated.     

Variability in response specificity of apple, hawthorn, and flowering dogwood-infesting Rhagoletis flies to host fruit volatile blends: implications for sympatric host shifts

Rhagoletis pomonella Walsh (Diptera: Tephritidae) originating from domesticated apple (Malus pumila), hawthorn (Crataegus mollis) (Rosaceae), and flowering dogwood (Cornus florida) (Cornaceae) were tested sequentially in flight‐tunnel assays to volatile blends previously identified from the three fruit types. The majority of flies flew to odor sources containing their natal blend (68–83%). Some flies from each fruit type also flew to non‐natal fruit blends (11–39%), but of these non‐natal responders the vast majority were flies that responded to their natal blend as well. The results indicate that individual flies within R. pomonella populations infesting different host types have different degrees of specificity with respect to discriminating among fruit volatile blends, and that a moderate proportion of apple, hawthorn, and dogwood flies (10–30%) are broad responders, with the capacity to recognize and orient to more than one blend. The observed variability in response specificity could facilitate sympatric shifts to new host plants.     

Radiation and divergence in the Rhagoletis pomonella species complex: inferences from DNA sequence data

Here, we investigate the evolutionary history and pattern of genetic divergence in the Rhagoletis pomonella (Diptera: Tephritidae) sibling species complex, a model for sympatric speciation via host plant shifting, using 11 anonymous nuclear genes and mtDNA. We report that DNA sequence results largely coincide with those of previous allozyme studies. Rhagoletis cornivora was basal in the complex, distinguished by fixed substitutions at all loci. Gene trees did not provide reciprocally monophyletic relationships among US populations of R. pomonella, R. mendax, R. zephyria and the undescribed flowering dogwood fly. However, private alleles were found for these taxa for certain loci. We discuss the implications of the results with respect to identifiable genetic signposts (stages) of speciation, the mosaic nature of genomic differentiation distinguishing formative species and a concept of speciation mode plurality involving a biogeographic contribution to sympatric speciation in the R. pomonella complex.     

Rapid and repeatable shifts in life‐history timing of Rhagoletis pomonella (Diptera: Tephritidae) following colonization of novel host plants in the Pacific Northwestern United States

Host shifts of phytophagous insect specialists to novel plants can result in divergent ecological adaptation, generating reproductive isolation and potentially new species. Rhagoletis pomonella fruit flies in eastern North America underwent a host shift ~160 ya from native downy hawthorn (Crataegus mollis) to introduced, domesticated apple (Malus domestica). Divergent selection on diapause phenology related to the earlier fruiting time of apples versus downy hawthorns resulted in partial allochronic reproductive isolation between the fly races. Here, we test for how rapid and repeatable shifts in life‐history timing are driving ecological divergence of R. pomonella in the Pacific Northwestern USA. The fly was introduced into the region via larval‐infested apples 40–65 ya and now attacks native black hawthorn (Crataegus douglasii) and introduced ornamental hawthorn (Crataegus monogyna), in addition to early‐ and late‐maturing apple varieties in the region. To investigate the life‐history timing hypothesis, we used a field‐based experiment to characterize the host‐associated eclosion and flight activity patterns of adults, and the feeding times of larvae at a field site in Vancouver, Washington. We also assessed the degree to which differences in host‐fruiting time generate allochronic isolation among apple‐, black hawthorn‐, and ornamental hawthorn‐associated fly populations. We conclude that host‐associated fly populations are temporally offset 24.4% to 92.6% in their seasonal distributions. Our results imply that R. pomonella possesses the capacity for rapid and repeatable shifts in diapause life history to match host‐fruiting phenology, which can generate ecologically based reproductive isolation, and potentially biodiversity in the process.     

Intra- and interspecific competition and host race formation in the apple maggot fly,Rhagoletis pomonella (Diptera: Tephritidae)

Intra- and interspecific resource competition are potentially important factors affecting host plant use by phytophagous insects. In particular, escape from competitors could mediate a successful host shift by compensating for decreased feeding performance on a new plant. Here, we examine the question of host plant-dependent competition for apple (Malus pumila)- and hawthorn (Crataegus mollis)-infesting larvae of the apple maggot fly, Rhagoletis pomonella (Diptera: Tephritidae) at a field site near Grant, Michigan, USA. Interspecific competition from tortricid (Cydia pomonella, Grapholita prunivora, and Grapholita packardi) and agonoxenid (subfamily Blastodacninae) caterpillars and a curculionid weevil (Conotrachelus crataegi) was much stronger for R. pomonella larvae infesting the ancestral host hawthorn than the derived host apple. Egg to pupal survivorship was estimated as 52.8% for fly larvae infesting hawthorn fruit without caterpillars and weevils compared to only 27.3% for larvae in harthorns with interspecific insects. Survivorship was essentially the same between fly larvae infesting apples in the presence (44.8%) or absence (42.6%) of interspecific insects. Intraspecific competition among maggots was also stronger in hawthorns than apples. The order or time that a larva exited a hawthorn fruit was a significant determinant of its pupal mass, with earlier emerging larvae being heavier than later emerging larvae. This was not the case for larvae in apples, as the order or time that a larva exited an apple fruit had relatively little influence on its pupal mass. Our findings suggest that decreased performance related to host plant chemistry/nutrition may restrict host range expansion and race formation in R. pomonella to those plants where biotic/ecological factors (i.e. escape from competitors and parasitoids) adequately balance the survivorship equation. This balance permits stable fly populations to persist on novel plants, setting the stage for the evolution of host specialization under certain mitigating conditions (e.g. when mating is host specific and host-associated fitness trade-offs exist).     

The population genetics of the apple maggot fly,Rhagoletis pomonella and the snowberry maggot,R. zephyria: implications for models of sympatric speciation

True fruit flies belonging to the Rhagoletis pomonella (Diptera:Tephritidae) sibling species complex possess several attributes consistent with a history of sympatric divergence via host plant shifts. Here, we investigate whether hybridization and genetic introgression is occurring between two members of the group, Rhagoletis pomonella (Walsh), whose primary hosts are domestic apples (Malus pumila) and hawthorns (Crataegus spp., and R. zephyria (Snow) whose host is snowberries (Symphoricarpos spp.). These flies are important because they appear to be at a transition stage between taxa reproductively isolated solely on the basis of host plant-related adaptations and those that have evolved additional non-host dependent sterility and inviability. Observing extensive hybridization and introgression between R. pomonella and R. zephyria would have major repercussions for current models of sympatric speciation.In a survey of allozyme and mitochondrial DNA (mtDNA) variation for 1105 flies collected throughout the northern United States we found two results suggesting that low level hybridization occurs between R. pomonella and R. zephyria. (1) Two flies reared from snowberries and one fly reared from hawthoms had genotypes indicative of them being R. pomonella and R. zephyria, respectively. Rhagoletis pomonella and R. zephyria adults may therefore occasionally frequent each others host plant, providing the opportunity for hybridization. (2) Four flies collected from hawthorns and one from snowberries had genotypes that made them likely to be F1 hybrids.Likelihood analysis revealed the data were also consistent with an hypothesis of shared ancestral alleles (i.e., the pattern of genetic variation could also be explained by R. pomonella and R. zephyria sharing alleles/haplotypes whose origins date to a common ancestor). We estimated that, in the absence of interspecific mating, random assortment of genes within R. pomonella and R. zephyria populations would produce an average of 5.4 flies with genotypes suggesting they were F1 hybrids – a number equivalent to the 5 putative F1 hybrids observed in the study. Our results therefore underscore the difficulty in distinguishing between hypotheses of low level introgression and shared ancestral polymorphism. But even if hybridization is occurring, the data suggest that it is happening at a very low and probably evolutionarily insignificant level (perhaps 0.09% per generation), consistent with sympatric speciation theory. Future tests are discussed that could help resolve the hybridization issue for R. pomonella and R. zephyria.     

Recursive adaptation in action: allochronic isolation and divergence of host-associated populations of the apple maggot fly,Rhagoletis pomonella,following its recent introduction to the western USA

An outstanding issue in the study of insect host races concerns the idea of ‘recursive adaptive divergence’, whereby adaptation can occur repeatedly across space and/or time, and the most recent adaptive episode is defined by one or more previously similar cases. The host plant shift of the apple maggot fly, Rhagoletis pomonella (Walsh) (Diptera: Tephritidae, Carpomyini), from ancestral downy hawthorn [Crataegus mollis (Torr. & A. Gray) Scheele] to introduced, domesticated apple (Malus domestica Borkh.) in the eastern USA has long served as a model system for investigating ecologically driven host race formation in phytophagous insect specialists. Here, we report results from an annual geography survey of eclosion time demonstrating a similar ecological pattern among nascent host-associated populations of the fly recently introduced ca. 40 years ago from its native range in the east into the Pacific Northwest (PNW) region of the USA. Specifically, using data collected from 25 locations across 5 years, we show that apple-infesting fly populations in the PNW have rapidly and repeatedly shifted (and maintained differences in) their adult eclosion life-history timing to infest two novel hawthorn hosts with different fruiting phenologies – a native species (Crataegus douglasii Lindl.) and an introduced species (Crataegus monogyna Jacq.) – generating partial allochronic reproductive isolation in the process. The shifts in the PNW parallel the classic case of host race formation in the eastern USA, but have occurred bi-directionally to two hawthorn species with phenologies slightly earlier (black hawthorn) and significantly later (ornamental hawthorn) than apple. Our results imply that R. pomonella can both possess and retain extensive-standing variation (i.e., ‘adaptive memory’) in diapause traits, even following introductions, to rapidly and temporally track novel phenological host opportunities when they arise. Thus, ‘specialized’ host races may not constitute evolutionary dead ends. Rather, adaptive phenotypic and genetic memory may carry over from one host shift to the next, recursively facilitating host race formation in phytophagous insects.