1 (owlet ) 1 (hook tip moths) (butter ies) 2 Calliduloidea 2 Cimelioidea (gold moths) ( moths, moths) 3 (lappet moths) Mimallonoidea (sack bearer moths) 3 Thyridoidea (picture-winged leaf moths) 4 Hyblaeoidea (teak moths) Copromorphoidea (fruitworm moths) Immoidea 4 Whalleyanoidea Pterophoroidea (plume moths)

Alucitoidea (many-plumed moths) Epermenioidea 5 Schreckensteinioidea (false burnet moths) Tortricoidea (leafroller moths) 5 Choreutoidea

Sesioidea (clearwing moths) 6 (carpenter moths) (burnet moths, forester moths) 6 Galacticoidea Symaethistoidea (case-bearers, twirler moths) 7 (ermine moths) 7 (leaf miners) (cloth moths) 8

Tischerioidea 8 Palaephatoidea 9 Andesianoidea (ghost moths, swift moths) 10 9 Mnesarchaeoidea Neopseustoidea Lophocoronoidea Acanthopteroctetoidea 10

MYA Eriocranioidea Heterobathmioidea HETEROBATHMIINA c. 160 Agathiphagoidea AGLOSSATA 29 Micropterigoidea ZEUGLOPTERA

Tree of Lepidopterans showing the phylogenetic relationships among the most significant groups (45 taxonomic superfamilies). Colored boxes indicate high taxonomic ranks. Branches with thick lines indicate robust clades, and branches with thin lines indicate less-supported clades. The number in the green circle indicates the chapter in which lepidopterans are also included. The orange circle marks the most internal node and its age. Photographs illustrate principal clades; boxed numbers associate photographs with clades. © 2014 Sinauer Associates, Inc. This material cannot be copied, reproduced, manufactured or disseminated in any form without express written permission from the publisher.

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summary Both professional researchers and aficionados, motivated by the appeal and popularity of (butterflies and moths), have compiled a huge amount of knowl- edge on their biology, , and distribution over the centuries, comparable only to the amount of information we have on vertebrates. However, it is only recently that a clear picture of the branch of the tree of life belonging to these has begun to emerge. The huge diversity of lepidopterans poses considerable difficulties for studying their evolu- tion: with 170,000 described, Lepidoptera represents the largest lineage of herbiv- orous organisms and one of the four most diverse orders on the planet. The limited fossil record has been another obstacle, both for understanding the evolution of morphological characters and for calibrating molecular clocks. The results of phylogenetic studies based on nucleotide sequences generally support hypotheses based on morphological charac- ters. Indeed, both methods are advancing in parallel, and as more characters are added, the results are generally congruent. The taxonomy is gradually being adapted to phyloge- netic results as they are consolidated and confirmed in numerous independent studies. To- day, the most relationships are quite well resolved, as is the general structure of the tree. Along with fossil information, it tells the story of lepidopterans: their emergence in the or Early from a common ancestor resembling the (Trichoptera), the evolution from mandibulates to with a proboscis, and the progressive acquisition of other specialized characters throughout one of the lineages, leading to the huge radiations of Ditrysia, which include the large moths and butterflies. Most of the rela- tionships within the Ditrysia have yet to be satisfactorily resolved, due largely to this group’s extremely rapid diversification. Projects under way include research on the phylogeny of the order based on morphological data and a large number of nuclear genes, the use of phylogenomics to resolve the most difficult cases, and investigation of multiple phylogenies of superfamilies and lower taxa. These promise exciting results that will clear up many ob- scure points in lepidopteran phylogeny and evolution.

With some 170,000 species described, lepidopterans, the most extensive for any group of such great biodi- along with coleopterans, hymenopterans, and dipter- versity. Their evolutionary success is based at least in ans, constitute one of the most diverse orders on the part on their holometabolism, which allows for a high planet. Their beauty has captured the interest of scien- degree of specialization in their different life stages tists and aficionados alike, and the information com- (see Figure 36.1). Another key factor is their special- piled on their biology, taxonomy, and distribution is ized mouthpart (proboscis or haustellum), which al-

What is a lepidopteran? A lepidopteran (from the Greek lepis [scale] and contain pigments and microstructures with optical pteron [wing]) is a holometabolous (undergoing properties, giving their color patterns enormous complete ) whose larvae, plasticity. The order Lepidoptera, one of the most called , are herbivorous, with rare diverse orders on the planet, is the sister clade exceptions. It has six true legs and multiple prolegs of the trichopterans or (Trichoptera), with diminutive hooks on them. The adults develop and together these clades form the superorder two pairs of wings densely covered in scales that Amphiesmenoptera.

© 2014 Sinauer Associates, Inc. This material cannot be copied, reproduced, manufactured or disseminated in any form without express written permission from the publisher.

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(A) (B) insects. Although most species have this chromosome number, there are notable exceptions. For example, the family has a modal number of 23–24. In some clades, the variability in chromosome (C) number is huge. For example, the subgenus Agrodiae- tus (Papilionoidea, Lycaenidae) contains species with chromosome numbers ranging from 10 to 134. Poly- ommatus atlanticus (Figure 36.2), also a lycaenid, is the metazoan with the highest chromosome number: its haploid number is around 223. Leptidea sinapis (Papil- ionoidea, ) boasts another record: it exhibits a cline (continuous gradient) in number of chromosomes from n = 53 on the Iberian Peninsula to n = 28 in Central Figure 36.1 Postembryonic stages of a holometabolous insect, the lepidopteran Melitaea didyma (Papilionoidea: Asia. Surprisingly, the extremes have been observed to ). (A) , (B) , and (C) adult. maintain a notable level of fertility when crossed. An- other curiosity of lepidopteran cytogenetics is that they Figure 36.1 have achiasmatic oogenesis; in other words, there is lows them to sip from flowering plants (angio- no meiotic recombination of maternal genetic mate- sperms). Lepidopterans are the largest lineage of phy- rial. Knowledge of this group’s chromosomal evolu- tophagous organisms in existence, as their larvae feed tion, the type of rearrangements, and the reasons for mostly on plants, especially angiosperms. These two their occurrence is practically nonexistent. It seems that groups have therefore evolved in tandem, and it is not karyotype changes occur as a result of fusion and frag- surprising that the explosive radiation of angiosperms mentation, given that the total amount of genetic mate- (see Chapter 12, Angiosperms) coincides with that of rial apparently remains constant (see also Chapter 12, butterflies and moths, as well as that of other pollinat- Angiosperms). The DNA content (C-value) has been ing insects like (see Chapter 33, Hymenopterans). determined for 60 species of lepidopterans, with an av- Another very important characteristic in the basic anat- erage value of 0.66 ± 0.04 picogram (pg), ranging from omy of lepidopterans is the dense covering of scale on a minimum of 0.29 to a maximum of 1.9 pg. their wings. This serves a function in both flight and The most commonly used markers in phylogenet- thermoregulation, and also gives their color patterns ics include mitochondrial genes (cox1, cox2, nad1 and enormous plasticity, a trait with profound implications nad5) and single-copy nuclear genes (CAD, EF-1a, for intra- and inter-specific relationships. H3, wg, etc.), as well as ribosomal regions (mainly the Knowledge of the lepidopteran evolutionary tree subunit 28S), and noncoding regions such as spacer is still only partial (see Tree of Lepidopterans), and un- regions (ITS-2) of the nuclear genome. Mitochondrial til recently was based only on morphological charac- markers have a greater substitution rate than nuclear ters. Studies using modern techniques of phylogenetic markers, and are therefore very helpful for studying reconstruction have appeared more recently, as have the relationship between only slightly divergent taxa, studies based on DNA sequences, or on a combination of morphological and molecular data. This chapter (A) (B) describes the phylogeny and evolution of lepidopter- ans, with special reference to results of the most recent studies.

Characteristics of Lepidopteran Genomes Our knowledge of lepidopteran cytogenetics, as well as (C) that of trichopterans with whom they share the char- acteristics described below, is limited. Their sex-de- termination system is ZW (heterogametic females). Their chromosomes are holocentric, appearing almost 10 μm circular during metaphase, and generally small and Figure 36.2 Polyommatus atlanticus, the species with the numerous. Lepidopterans have the widest range of highest chromosome number of all metazoans studied. chromosome numbers of all , ranging from 7 (A) Adult male. (B) Adult female. (C) Microscopic image of to approximately 223. The haploid modal number for the nucleus of a male gamete in formation (metaphase 1) lepidopterans is 30–31, which is unusually high for (n = approximately 233). © 2014 Sinauer Associates, Inc. This material cannot be copied, reproduced, manufactured or Figure 36.2 disseminated in any form without express written permission from the publisher.

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Basic terms Achiasmatic: Not forming a chiasma. The chiasma is Metapopulation: A group of unstable local the region where the filaments of non-sister chro- populations interconnected by migratory processes. matids cross during genetic recombination. This concept is of great significance for the study of Apodeme: An infolding of the integument (exoskel- habitat fragmentation and conservation. eton) serving as an insertion point for musculature. Microlepidopteran: Belonging to one of the 36 non- Aposematic coloration: Contrasted coloration, typi- superfamilies. A nonmonophyletic cally yellow and black, or red and black, that alerts group made up mostly of small-sized species (see potential predators and serves as an indication of Figure 36.4). toxicity or danger. Myrmecophily: The ability to live in association C-Value: The amount of DNA contained in a haploid with . In lepidopterans, myrmecophily is very nucleus of a eukaryotic organism. common in the larvae and pupae of the families Cerci (sing., cercus): A pair of appendages on the ter- Lycaenidae and . minal segment of many ’ abdomens. Ovipore: The female ovipositor opening. Epiphysis: Each of the ends of a long bone or joint. Polyphenism: The expression of different phenotypes Frenulum: A bristle located on the base of the hind by a single genotype, depending on environmental wing, inserted into a hook or fold (retinaculum) at conditions. the base of the forewing. Forms part of the system Preimaginal stages: Insect stages prior to the adult joining the forewings and hind wings of most Het- () stage: egg, larva, and pupa (some of these eroneura lepidopterans. are shown in Figure 36.1). Galeae (sing., galea): External lobes on the maxilla Proboscis: An elongated appendage on the head of an which, in the case of lepidopterans, has been modi- . fied to form the haustellum. Retinaculum: A hook or fold on the base of the Gonopore: The genital copulatory opening. forewing into which a bristle (frenulum) extending Haustellum: Mouthparts of the vast majority of lepi- from the base of the hindwing is inserted. It forms dopterans, consisting of maxillae modified into a part of the system joining the forewings and hind long tube that is kept rolled up in a spiral when not wings of most lepidopterans. in use. Rs veins: Veins in the wings of insects running from Holometabolous: Undergoing complete metamor- the cell down to the apex or apical area. phosis, through the postembryonic phases of larva, Setae: Bristles on any part of an organism. pupa, and adult (see Figure 36.1). Sternite: Each of the ventral plates that form part of M veins: Central veins in the wings of insects run- the cuticular lining of the segments in arthropods. ning from the cell down to the termen, or external Tergite: Each of the dorsal plates forming part of the margin. cuticular lining of the segments on arthropods. Macrolepidopteran: Belonging to one of the nine ZW sex determination system: A system whereby the superfamilies that form the derived group Macro- sex of offspring is determined by the ovum, not by lepidoptera. Most are medium-sized to large species the spermatozoid. The males are the homogametic that have been recovered as a monophyletic clade in sex (ZZ) and the females are the heterogametic sex some studies, but not in others. (ZW). This system is also found in birds and certain actinopterygian fishes.

but saturation makes them much less useful for clari- evolutionary relationships. Information from the fossil fying deeper relationships. The substitution rate in the record, valuable both for understanding the evolution nuclear markers is quite variable, and some also com- of morphological characters and for calibrating mo- bine more or less conserved regions. Noncoding mark- lecular clocks, is very limited in the case of lepidop- ers have insertions and deletions that make it difficult terans. The relative scarcity of lepidopteran fossils is to align sequences between taxa at the level of tribe or due to the soft tissue of their larvae and the capacity of higher for ITS-2 and at the level of family for 28S. dead adult bodies to remain floating on water surfaces for long periods rather than sedimenting quickly. The Phylogenetic Results Contrasted oldest known lepidopteran fossil, Archaeolepis mane (190 million years old), belongs to the ancestral branch with Previous Classifications common to all extant lepidopterans. Based on this fos- Molecular phylogenies are contributing a huge amount sil, it would be reasonable to place the origin of the of new data on the origins of lepidopterans and their order in the Early Jurassic, some 200 million years ago. © 2014 Sinauer Associates, Inc. This material cannot be copied, reproduced, manufactured or disseminated in any form without express written permission from the publisher.

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• larvae with modified salivary glands that produce Box 36.1 Unique morphological silk. characters of the Lepidoptera • The most striking ecological specialization of the Adult lepidopteran with some of the autapomorphies trichopterans is that their larvae construct highly of the order indicated using the numbering from the elaborate casings or shelters, for which they list below. The characters listed are among the most use silk and material from their environment. evident of the more than 20 that have been described: Lepidopterans also have these glands, and some groups use them to build shelters for their larvae 1. Wings with a dense scale covering on veins and membrane or, more commonly, for their pupae (see Tree of Lepidopterans). 2. M Veins of wings with three branches, instead of four or more (except in the only two known species The basal structure of the Tree of Lepidopterans is of Agathiphagoidea) now quite well defined, and the results of phylogenies based on molecular data generally concur with the 3. Tibia of the forelegs with only one or no apical spine results based on morphological characters. The most widely accepted classification today divides the order 4. Loss of central ocellus (simple eye) into four suborders (Zeugloptera, Aglossata, Hetero- 5. First tergite very slightly sclerotized bathmiina, and Glossata), although there are still some 6. Loss of cerci on the abdomen questions left unanswered, such as the position of Ag- 7. Tibia of front legs with epiphysis articulated on athiphagoidea (Aglosata). Other divisions have been inner surface. posited based on morphological characteristics (Het- erocera and Rhopalocera; Microlepidoptera and Mac- rolepidoptera; and Ditrysia); all of these are still used at times, although more detailed mor- 1 phological studies and results based on molecular data have shown that they contain paraphyletic groups. There are currently 45 accepted superfamilies and around 124 accepted families, whose monophyly has been proven, in many cases, by molecular studies. In 2 4 addition, in the last few years, robust molecular phy- logenies and revisions of the systematics of most ma- 3, 7 5 6 jor superfamilies have been published. However, the deep relationships between the lepidopteran Ditrysia, a monophyletic group representing 98% of the order’s species, have proven hard to resolve, largely due to the fact that its prodigious diversity contains numerous adaptive radiations that occurred almost simultane-

However,Figure the onlyBox 36.1 molecular-clock dating performed for the order so far places its origin in the Triassic, (A) (C) some 240 million years ago. The results of all the phylogenetic studies based on DNA sequences published to date agree with pre- vious classifications that placed Trichoptera as the sis- ter order to Lepidoptera (Figure 36.3); together, they form the superorder called Amphiesmenoptera. This is one of the very few relationships between orders of (B) (D) holometabolous insects on which there is unanimous agreement. The monophyly of the Amphiesmenop- tera is supported by more than 15 morphological and vital synapomorphies, notable among which are the following: • ZW sex-determination system (i.e., the females are the heterogametic sex); • presence of setae, more or less modified into Figure 36.3 Trichoptera is the sister order of Lepidoptera. scales, on the wing surface; and (A) Adult trichopteran and (B,C) Trichopteran larvae in their shelters. (D) Psychidae (Lepidoptera) larva in its shelter. © 2014 Sinauer Associates, Inc. This material cannot be copied,Figure reproduced, 36.3 manufactured or disseminated in any form without express written permission from the publisher.

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the proboscis. They have also improved their flight Box 36.2 Lepidopterans by the by modifying the venation, design, and attachment numbers of their forewings and hind wings. The females have • Number of species described: approximately developed independent reproductive and excretory 170,000 systems, and the copulatory opening has also been separated from the ovipositor opening for greater ef- • Estimated number of species: 280,000–500,000 ficiency and control. It is also worth noting that on at • Number of families: approximately 124 least eight independent occasions, Ditrysia lepidop- • Family with the largest number of species: terans have developed tympanal organs that enable (more than 35,000 species described), although the them to hear the echolocation signals of bats in order exact definition of this family is still being debated to avoid them. • Origin: Triassic or Early Jurassic, 240–200 million Listed below are the most significant apomorphic years ago characters, from the oldest apparently monophyletic • Oldest fossil: 190 million years old, Archaeolepis groups to the most subordinate: mane (Early Jurassic) • Glossata: Vestigial, nonfunctional mandibles; • Largest species: Wingspan of 280 mm in the adult elongated galeae forming a proboscis (also called agrippina (Noctuidae) (Central and South haustellum); larva with a spinneret, an organ America). The largest wing surface, more than 400 used to create silk thread, located on the top of cm2, is that of () (Southeast the prelabial–hypopharyngeal lobe Asia). • : Hollow scales on the wing surface • Smallest species: Wingspan of <2 mm in adults of several species of the family • Heteroneura: Female genitalia with the gonopore • Approximate number of species described and separate from the ovipore, and no connection estimated for the main groups: between the two; reduction in the number of branches of Rs veins in the hind wings; fore- Species Species wings and hind wings coupled by the frenulum Species estimated estimated and retinaculum system; loss of the first abdomi- Taxon described (minimum) (maximum) nal sternite Basal groups 10,000 15,000 20,000 • Ditrysia: Internal connection between gonopore Gelechioidea 18,000 60,000 100,000 and ovipore; second abdominal sternite with Tortricoidea 10,000 15,000 20,000 large apodemes; muscles in the proboscis in short Pyraloidea 16,000 30,000 30,000 bands instead of long fibers Papilionoidea 18,000 18,000 20,000 • Apoditrysia: Second abdominal sternite with spe- Geometroidea 21,000 30,000 45,000 cialized apodemes, with large, short bases Noctuoidea 70,000 100,000 150,000 • : Pupa whose first to sixth abdomi- Other 10,000 15,000 20,000 nal segments are immobile; loss of dorsal rows of Total 173,000 283,000 405,000 spines on pupal tergites • Macrolepidoptera: Complete loss of the vein that ends at the anal angle (CuP vein); larval prolegs ously. Although several relationships among Ditrysia with diminutive hooks forming a crescent-shaped superfamilies have been resolved in specific studies, series on the inner part each new study identifies contradictory relationships, making it almost impossible to reach any conclusion. Most of the discrepancies between traditional classifi- Evolutionary Tendencies cations and the new molecular phylogenies are found A comparison of the Lepidoptera with other orders, precisely in these highly diverse groups. especially their sister order, makes it possible to de- fine evolutionary tendencies specific to butterflies and moths. One of the most obvious is the fact that lepi- Evolution of Characters dopterans have specialized in terrestrial larval devel- Over the course of their evolution, lepidopterans opment, while caddisflies have specialized their larval (except for a few lineages whose habits and forms development in aquatic environments. Only a few lepi- seem to have changed very little) have adapted their dopterans have aquatic larvae, and only a few trichop- larvae and pupae to terrestrial environments. The terans have terrestrial larvae (although they are always adult forms have specialized in obtaining nectar and associated with damp leaf litter). Another evolution- other fluids from , with the development of ary feature of butterflies and moths is their extremely © 2014 Sinauer Associates, Inc. This material cannot be copied, reproduced, manufactured or disseminated in any form without express written permission from the publisher.

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Butterflies: In search of a robust phylogeny In spite of their peculiarities, both in appearance and in sister group to the rest (Papilionoidea). The five families habits, butterflies form a clade with relatively little di- within Papilionoidea were recovered as monophyletic, versity within the immense variety of lepidopterans. The and the relationships between them appeared to be al- number of butterfly species (18,000) is approximately most completely established. 10% of the total, although it is the most extensively However, the most recent phylogenies, which in- studied group. Its position in the phylogenetic tree indi- clude a greater number of nuclear markers and revised cates that it is a group of macrolepidopterans special- morphological characters, have brought a new surprise: izing in diurnal activity that probably first appeared in the the family Papilionidae is the sister group to the rest; early , some 110 million years ago. and Hedylidae, in spite of its -like appearance (see The beauty of butterflies has made them an object Box Macrosoma: The missing link?), is a sister group of study for nature lovers since ancient times. For this of Hesperiidae. The new relationships between families reason, data on their taxonomy, biology, and distribu- have resulted in the unification of all butterflies into a tion is the most complete of all invertebrates. In spite of single superfamily: Papilionoidea (see Figure). this wealth of information, the phylogenetic relationships In the same way, by joining efforts and combining between the main groups have been a source of consid- data, researchers are also trying to improve our under- erable debate. In studies based mainly on morphological standing of the tree of all lepidopterans. In addition to data on adult insects, various hypotheses have been the numerous groups studying the evolutionary relation- posited with little empirical support. The first studies ships between subfamilies or lower groups, there are to include molecular data to address the problem were two especially noteworthy international collaboration based on very short nucleotide fragments, and the con- projects. LepTree, a project funded by the US National clusions were far from clear. Science Foundation’s Assembling the Tree of Life In light of this situation, an exemplary international Program (http://www.phylo.org/atol/projects), aims to collaboration project was established that brought to- resolve the phylogenetic tree by sequencing 24 genes gether all the main molecular laboratories (approximately 18 kb), from representatives of all of the specializing in Rhopalocera, with one objective: to work families; and the European project LepSys (www.Lep- together on the creation of a solid phylogenetic hy- Sys.eu) seeks to facilitate the integration of traditional pothesis based on a combination of morphological and morphological methods with the new molecular meth- molecular data. The resulting phylogeny identified Hes- ods, as well as creating create a catalogue and a clas- perioidea (made up of a single family, Hesperiidae) as a sification of all lepidopterans.

(a) (b) (c) (d) (e) (f)

Papilionidae Hedylidae Hesperiidae Pieridae Papilionoidea Nymphalidae Lycaenidae Riodinidae

Relationships among Papilionoidea families based on the tree. (a) Papilionidae, (b) Hesperiidae, (c) Pieridae, (d) Nym- combination of morphological and molecular data. Photo- phalidae, (e) Lycaenidae, and (f) Riodinidae. graphs of adult representatives of some of the families on the

UNBox 36.1 high rate of diversification. In spite of being a relatively tant species. Three of the four suborders (Zeugloptera, young order, lepidopterans have diversified consider- Aglossata, and Heterobathmiina) contain only around ably since their origins, especially in comparison with 250 known species. By placing the fossils and morpho- trichopterans. There are close to 170,000 known lepi- logical characters on the evolutionary tree, it can be de- dopteran species, compared with 11,500 described spe- duced that these lineages retain many ancestral charac- cies of trichopterans. This difference of more than one ters and have remained almost unchanged for millions order of magnitude might be associated with the adap- of years (evolutionary stasis). These species are super- tation of butterflies and moths to diverse environments, ficially similar to the smallest trichopterans and prob- to the high level of specialization of many species, and ably resemble the common ancestor of all lepidopter- to the importance of sexual selection in this order. ans. The adults still have mandibles (functional in some However, the Tree of Lepidopterans is extremely asym- groups, atrophied in others), an ancestral character that metrical, and several basal lineages have very few ex- has been replaced by the proboscis in all other lepidop- © 2014 Sinauer Associates, Inc. This material cannot be copied, reproduced, manufactured or disseminated in any form without express written permission from the publisher.

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(A) (B) • The greatest difficulties in phylogenetic studies, and most of the discrepancies between the results of different methods, are found in the most di- verse groups, which have undergone significant adaptive radiations.

Biogeography and Figure 36.4 Microlepidopterans. (A) Adult micropterigid, of a and species not yet described, in northern Lepidoptera is the fourth most diverse order on the Madagascar. (B) Mastic (Pistacia lentiscus) leaf mined by a planet. Most of its biodiversity is concentrated in the larva of the species Acalyptris minimella (Nepticuloidea) in the tropics, no doubt because of the good conditions for Sierra de Grazalema, Spain. butterflies and moths found in these regions. The Figure 36.4 biogeographical origin of the group is unknown. The oldest known fossil, which appears to be a common terans. (Figure 36.4A) are the only ex- ancestor of all extant lepidopterans, was found in tant representatives of the suborder Zeugloptera, and Great Britain. The two main lineages into which lepi- seem to be the sister lineage to all other lepidopterans. dopterans are divided (micropterigids and the rest Their larvae are ; they feed on plant remains of the lepidopterans) are distributed across all conti- and fungi in damp soil. The only two known species nents. It is important to remember that lepidopterans of the suborder Aglossata, whose phylogenetic posi- probably appeared at the time that the supercontinent tion is not yet resolved, have leaf-miner larvae that feed Pangea was breaking apart. It is therefore not surpris- on seeds of relict conifers (Araucariaceae) in Oceania, ing that they were able to spread very quickly over and which have probably maintained characters from the planet. Although no biogeographical study based an evolutionary stage prior to the angiosperm radia- on the whole lepidopteran tree has been conducted, tion. On the other hand, the Heterobathmiina special- there are numerous studies of families and lower lev- ize in angiosperms, and their known species, number- els. These studies have made it possible to determine ing around 10, have leaf-mining larvae that feed on the the origin, for example, of Parnassiinae (Papilionoidea: austral relict genus Nothofagus. In short, lepidopterans Papilionidae), in Central Asia, and of Pseudopontiinae seem to have followed the evolution of the plants, and + Dismorphiinae (Papilionoidea: Pieridae) in western for much of their evolutionary history they have been Gondwana. However, in some cases the interpretation small in size (microlepidopterans) and generally with of the results continues to be debated due to the dis- leaf-miner larvae (Figure 36.4B). persal capacity of butterflies and moths. Indeed, recent One lineage in particular has acquired a large molecular studies with groups have confirmed the number of apomorphies, key innovations that led dispersal of butterflies and moths between continents, to its ecological success: the Ditrysia lepidopterans, as well as their colonization of remote islands. This which probably appeared in the mid-Cretaceous at the capacity for dispersal needs to be taken into account beginning of the angiosperm radiation. The Ditrysia when calibrating the molecular clock based on vicari- represent the vast majority of species of the order and paleogeographic events. include all of the major radiations, such as that of the Noctuoidae (some 70,000 known species) (Box 36.2). Following is a summary of other evolutionary ten- Differentiation and Speciation dencies specific to lepidopterans: The study of speciation processes in lepidopterans has • There is a very clear tendency toward an increase focused almost exclusively on butterflies. Sexual se- in average size over the course of their evolution. lection associated with the pronounced sexual dimor- phism of wing coloration in butterflies (see for example • The chromosome number is relatively stable, Figure 36.2) provided Darwin with a key example for although it is highly variable in specific genera. his theory. Although much less studied, there is also • The cases of chromosome number variability a chemical component to sexual selection and specia- appear to be due to fusion or fragmentation. tion in lepidopterans. Butterflies and moths produce • Speciation is nearly always allopatric or para- characteristic of each species that enable patric. the sexes of the same species to find and recognize one • Obligate myrmecophily and ant parasitism en- another. However, hybridization sometimes occurs, tail a high degree of specialization that poses a and a large number of natural hybrids can be found greater risk of extinction, while also appearing to in collections. Several of the few proven cases of ho- increase the rate of speciation. moploid speciation resulting from the hybridization of two parental species in animals are butterflies (for © 2014 Sinauer Associates, Inc. This material cannot be copied, reproduced, manufactured or disseminated in any form without express written permission from the publisher.

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Lepidopterans as model organisms Silkworm () New Zealand. It is also one of the few butterfly species This moth from the superfamily Bombycoidea has whose genome has been sequenced. been grown in China for silk for at least 5000 years. It Squinting Bush Brown (Bicyclus anynana) is the only insect that has been totally domesticated This African woodland butterfly (Papilionoidea: Nym- to the point that it is incapable of living in the wild. Be- phalidae) exhibits a marked seasonal polyphenism (phe- cause it is easy to raise, and because of its economic notypic plasticity) in its wing color pattern. It is used as importance, the silkworm has come to be used as a a model for studying the development and genetics of . This has led to some major discover- wing patterns. The sequencing of its genome is currently ies about pheromones, , and the structure of underway. the nervous system. Its , called bombykol, was the first to be characterized. For this study, Nobel Melitaeini Prize winner Adolph Butenandt had to collect extracts Several species of this group (Papilionoidea: Nymphali- from 500,000 individuals. Since then, hundreds of lines dae), especially Euphydryas editha in the United States and mutations have been described, and Bombyx mori and Melitaea cinxia in Europe, have been used for years was the first lepidopteran to have its nuclear genome in ecological and population dynamics research. Their sequenced. Several genetic engineering projects have usefulness for study lies mainly in their structure in enabled silkworms to produce proteins for medical or metapopulations, along with the fact that they are eas- industrial uses, instead of for silk. ily marked and monitored in the field. Heliconians Large blues (genus Maculinea) Most heliconians (Papilionoidea: Nymphalidae) are toxic The larvae of the families Lycaenidae and Riodinidae or foul tasting. This characteristic has favored the de- (Papilionoidea), like many hemipterans, often sustain velopment of a conspicuous aposematic coloration mutualistic relationships with ants (myrmecophily) (see and widespread among heliconian species. The Chapter 32, Hemipterans). However, the relationship genetics and genomics of speciation and wing patterns sustained with ants by the genus Maculinea is parasitic. in this group are being studied in depth. In several European countries, large blues are either Monarch butterfly (Danaus plexippus) extinct or are in extreme danger of extinction. They are used as a model organism for studying parasitism and Monarch butterflies (Papilionoidea: Nymphalidae) are a featured organism in invertebrate conservation ef- make spectacular migrations between Mexico, where forts. The number of studies conducted on large blues, they gather in a few select forests to spend the winter, as well as the number of initiatives to monitor, protect, and the northern regions of North America. Both their and even reintroduce these butterflies in Europe, is com- extraordinary journey of thousands of kilometers and parable only with the campaigns carried out for certain their beauty have made them one of the world’s most endangered vertebrates. popular butterflies. The monarch is a model for research on migration in invertebrates; marking and recapture Tobacco hornworm ( sexta) campaigns and sightings of specimens in migration are This moth (Bombycoidea: ) is used as a conducted with the help of both amateurs and scholars. model species in studies of invertebrate neurobiology. It is a lesser-known fact that this species is widely dis- Its large size facilitates the study and manipulation of its tributed over the planet; it includes colonies on the nervous and hormonal systems. Atlantic and Pacific Islands, as well as in Australia and

(a) (b) (c) (d)

Lepidopterans used as model organisms. (a) Monarch butter- (c) heliconians (Actinote sp.), and (d) large blue (Maculinea (Danaus plexippus), (b) silkworm (Bombyx mori), alcon).

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Macrosoma: The missing link? The genus Macrosoma Hübner, with close to 40 spe- cies described (all restricted to Neotropical forests), is (a) the only known representative of the family Hedylidae. Its external appearance (pictured) is typical of moths of the geometrid group (Geometroidea: Geometridae). It is therefore no surprise that the group was described in 1857 as belonging to Geometridae, a taxonomic position they occupied until 1986, when Scoble published his (b) revision of the group. Scoble’s conclusion was astonish- ing: in spite of their appearance and their mainly noctur- nal habits, the Hedylidae are not moths, but butterflies. This extraordinary hypothesis identified Macrosoma as the only extant genus of a superfamily of butterflies, and a sister group to the rest. A detailed analysis reveals a large number of possibly synapomorphic characters for The Hedylidae (genus Macrosoma) are butterflies (Papilionoi- butterflies including Macrosoma. Indeed, their appear- dea) in spite of their moth-like appearance. Shown are Macro- ance in immature stages is surprisingly similar to that soma tipulata (a) adult and (b) larva. of several groups of butterflies. The first researcher to UNBox 36.3 study the development of Macrosoma wrote, “I thought that the larvae might belong to a species of nymphalid data have corroborated that the Hedylidae are butterflies [Papilionoidea: Nymphalidae], but when the first larva and identify them as a sister group to Hesperiidae. The hatched I was sure that it was a pierid [Papilionoidea: last revision of the group posits a single superfamily that Pieridae]. The first adult to emerge was a complete sur- encompasses all butterflies (Papilionoidea), and thus prise.” Recent studies based on molecular data and on Macrosoma has ceased to be viewed as a missing link a combination of molecular and morphological and is now considered a true butterfly.

example, the heliconian Heliconius heurippa, the swal- sify by specializing on ants or plants, but the converse lowtail Papilio appalachiensis, and lycaenids of the ge- relationship rarely occurs. The famous case of the co- nus Plebejus). The heliconians, a group that serves as a evolution of yucca moths (Adeloidea: ) model for research into speciation, have also provided with their host plant is a notable exception. evidence of the possibility of a species dividing into two, due to specialization in mimicking two different Principal Questions Remaining toxic models. A recent study with lycaenids of the sub- • Will it be possible to resolve the topology of genus Agrodiaetus used their phylogeny to demonstrate the Tree of Lepidopterans and establish solid the reinforcement of prezygotic barriers. Specifically, it relationships among the 45 superfamilies? demonstrated that natural selection favors differentia- • Given their rapid diversification, relationships tion in colors of males between nascent species that are within the Ditrysia clade seem practically in secondary geographic contact, and their selection by impossible to resolve with current methods. females of the same species, so as to prevent creating in- Will phylogenomics provide new data on this fertile hybrids. Chromosome fragmentation and fusion group? are common in this subgenus’s process of speciation. With around 120 species produced over some three • Is it possible to create a complete matrix of million years, it has one of the highest known rates of morphological characters that would encompass speciation, probably due to its great chromosomal in- the whole order and combine it with molecular stability. Studies conducted to date suggest that allo- data? patric speciation is the most common type of speciation • Can the origin of lepidopterans be dated more among lepidopterans, although parapatric speciation precisely by calibrating the molecular clock from has also been documented in a few cases. the fossil record? In spite of the close relationship between lepidop- • What is the biogeographical origin of the order? terans and food plants, and between ants and some • Can the taxonomy continue to be updated in butterfly larvae, hardly any cases of coevolution have response to new discoveries about the group’s been documented. In other words, the usual evolution- evolution? ary pattern involves lepidopterans being able to diver- © 2014 Sinauer Associates, Inc. This material cannot be copied, reproduced, manufactured or disseminated in any form without express written permission from the publisher.

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• Will the phylogenetic tree of the order help to and biogeography. Berlin, New York: Walter de explain the origin and evolution of ecological Gruyter. specializations such as changes in diet, Kristensen, N. P. (ed.) 2003. Handbook of Zoology IV. myrmecophily, diurnality, the aquatic life of Arthropoda: Insecta, Part 36: Lepidoptera, moths preimaginal stages, and so on? Will it also help and butterflies. Volume 2: Morphology, physiology, to explain their effects on generating biodiversity and development. Berlin, New York: Walter de and its correlation with morphological Gruyter. adaptations? Mutanen, M., Wahlberg, N., and Kaila, L. 2010. Comprehensive gene and taxon coverage Basic Bibliography elucidates radiation patterns in moths and Heikkilä, M., Kaila, L., Mutanen, M., Peña, C., and butterflies. Proceedings of the Royal Society B: Wahlberg, N. 2011. Cretaceous origin and repeated Biological Sciences 277: 2839–2848. tertiary diversification of the redefined butterflies. Regier, J. C., Mitter, C., Zwick, A., Bazinet, A. L., Proceedings of the Royal Society B: Biological Sciences Cummings, M. P., et al. 2013. A Large-Scale, 279: 1093–1099. Higher-Level, Molecular Phylogenetic Study of the Kristensen, N. P. (ed.) 1999. Handbook of Zoology IV. Insect Order Lepidoptera (Moths and Butterflies). Arthropoda: Insecta, Part 35: Lepidoptera, moths PLoS ONE 8: e58568. doi:10.1371/journal. and butterflies. Volume 1: Evolution, systematics, pone.0058568.

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36_TOL1e_36.indd 438 5/8/14 11:13 AM © 2014 Sinauer Associates, Inc. This material cannot be copied, reproduced, manufactured or disseminated in any form without express written permission from the publisher.

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