Behavioral Phylogeny of Corbiculate Apidae (Hymenoptera; Apinae), with Special Reference to Social Behavior

Home , Apidae, Apinae, Bombini, Euglossa cordata, Euglossini, Mass provisioning

Cladistics 18, 137–153 (2002) doi:10.1006/clad.2001.0191, available online at http://www.idealibrary.com on

Fernando B. Noll1 Museum of Biological Diversity, Department of Entomology, The Ohio State University, 1315 Kinnear Road, Columbus, Ohio 43212

Accepted September 25, 2000

The phylogenetic relationships among the four tribes of Euglossini (orchid bees, about 175 species), Bombini corbiculate bees (Euglossini, Bombini, Meliponini, and (bumblebees, about 250 species), Meliponini (stingless Apini) are controversial. There is substantial incongru- bees, several hundred species), and Apini (honey bees, ence between morphological and molecular data, and the about 11 species) (Michener, 2000), that belong to the single origin of eusociality is questionable. The use of Apinae, a subfamily of long-tongued bees. They all behavioral characters by previous workers has been possess apical combs (the corbiculae) on the females’ restricted to some typological definitions, such as soli- tibiae as well as several other morphological synapo- tary and eusocial. Here, I expand the term “social” to 42 morphies (with the exception of the parasitic forms, characters and present a tree based only on behavioral and the queens of highly eusocial species). In addition, characters. The reconstructed relationships were similar with the exception of the Euglossini and the social to those observed in morphological and “total evidence” parasite Psithyrus in Bombini, all corbiculate bees are ؉ ؉ ؉ analyses, i.e., Euglossini (Bombini (Meliponini social. Apini)), all of which support a single origin of euso- Despite many studies that have focused on the classi- ciality. ᭧ 2002 The Willi Hennig Society ﬁcation and phylogeny of bees, many authors do not agree on one classiﬁcation and one phylogeny. Ac- cording to Michener (1990) few morphological characters exist for resolving tribal relationships. In fact, four different phylogenetic hypotheses were proposed INTRODUCTION based on morphological characters: (1) (Euglossini (Bombini (Meliponini ϩ Apini) (Michener, 1944; Pren- Corbiculate Apidae (Roig-Alsina and Michener, tice, 1991; Roig-Alsina and Michener, 1993). (2) (Euglos- 1993; Michener, 2000) comprises four tribes (Fig. 1), sini ϩ Bombini) (Meliponini ϩ Apini) (Michener, 1990). (3) (Meliponini (Apini (Bombini ϩ Euglossini) (Win- 1Present address: Departamento de Biologia, FFCLR-USP, Ave. ston and Michener, 1977; Kimsey, 1984). (4) (Euglossini Bandeirantes 3900, 14040-901 Riberaˇo, Pretio-SP, Brazil. (Meliponini (Bombini ϩ Apini) (Plant and Paulus,

FIG. 1 Nest examples from the four tribes of corbiculate Apidae: Eulaema nigrita, Euglossini (A); Bombus atratus, Bombini (B); Melipona ininterrupta grandis, Meliponini (C); and Apis mellifera, Apini (D). A, B, C, modiﬁed from Michener (1974); D, modiﬁed from Wilson (1971).

1987). Based on molecular data, two different phyloge- Michener (1969) divided the social bees into two nies were determined: Euglossiniϩ Apini as a sister main groups: the primitively social bees, which include group of Meliponini ϩ Bombini (Cameron, 1991) and the Bombini (and more distantly related bees not con- Apini as the sister group of the other corbiculate bees sidered here, such as Allodapini and Halictinae), and with Meliponini and Bombini as sister groups (Cam- the highly social bees, which comprise the Apini and eron, 1993; Cameron and Mardulyn, 2001; Kouli- the Meliponini. By definition, the castes of primitively anos et al., 1999; Mardulyn and Cameron, 1999; Shep- social bees are externally indistinguishable except for pard and McPheron, 1991). In both molecular data sets, allometric macrocephaly in some of the queens. Larval strong support was found for the branch Bombini ϩ food is stored in brood cells, and colonies in the major- Meliponini but not for this clade and the other two ity of the cases are small (no more than a few hundred tribes (Cameron, 1993; Koulianos et al., 1999; Sheppard adults), although in Bombus there is caste differentia- and McPheron, 1991). On the other hand, a “total evi- tion according to size and food is stored outside of the dence” analysis using molecular and morphological brood cells. The highly social bees show well-defined data (Chavarrıá and Carpenter, 1994) resulted in the behavioral, physiological, and anatomical caste differ- same structure as found by Roig-Alsina and Michener entiation. For example, the queens “lack structures for (1993) in which Apini and Meliponini are sister groups, collecting and manipulating pollen” (Michener, 1969). and recent analyses of one molecular data set (opsin) Queens are unable to survive away from the colony, have suggested that the gene may be providing weak and new colonies are exclusively established via support at the tribal level (Ascher et al., 2001). swarming. Colonies are perennial, persisting for years,

᭧ 2002 by The Willi Hennig Society All rights reserved. Behavioral Phylogeny of Corbiculate Bees 139 and food is stored in the nest. The evolution of ad- The usefulness of ecological characters in phyloge- vanced eusociality in bees was considered as a unique netic analysis is twofold: they may be used in phylog- event (Michener, 1944). However, because all molecu- eny reconstruction or evaluated based upon existing lar and some morphological analyses (see above) were cladograms. Explicit statements regarding definition unable to support Meliponini and Apini as a single of alternative character states and direction of state clade, multiple origins for advanced eusociality in the change can trace evolution through hypothetical ances- corbiculate Apidae have been suggested (Winston and tors, establishing the most parsimonious explanation Michener, 1977; Cameron, 1993). for observed data. Applications of these methods sug- The use of behavioral classifications to determine the gest that, when properly used and defined, behavioral social status in insects remains controversial. Typologi- and ecological traits may accurately reflect phylogeny cal characterization can lead to wrong generalizations (reviewed in Wenzel, 1992; Miller and Wenzel, 1995). In fact, constructing phylogenies based only on behav- or the lack of useful information (Wcislo, 1997). For ioral characters, and comparing the resulting tree(s) to example, at least in some sweat bees, such classifica- phylogenies based on other data, seems to be best way tions seem to be appropriate only at the colony level, to test the robustness of behavioral data (courtship but not at the species levels. Even though such defini- behavior in salamanders of the genus Triturus (Arntzen tions are not so easily applicable to the social apine and Sparreboon, 1989), reproductive behavior in gas- tribes because of the complex social organization of teroid fishes (McLennan, 1993), social life traits in pri- their societies (see above), it is important to keep in mates (DiFiore and Rendall, 1994), social behavior and mind that standardized classifications are most stable life history in albatrosses, petrels, and penguins (Pater- if they make reference to historical patterns (e.g., phy- son et al., 1995), social behavior in pelicaniforms (Ken- logenies) (Wcislo, 1997). nedy et al., 1996), cocoon spinning behavior in black The units of behavior that together compose social flies (Stuart and Hunter, 1998), grooming behaviors behavior in corbiculate bees present many informative in Hymenoptera (Basibuyuk and Quicke, 1999), and characters (Michener, 1974, discussed in Wcislo, 1997), courtship displays in storks (Slikas, 1998)). Only in this but they have been rarely considered in phylogenetic last case were behavioral data considered less reliable analyses. In fact, when these characters are used, they than other data. Behavioral data were used to recon- are in a typological form, like “Sociality: none (0); prim- struct the phylogeny of orb-weaving spiders (Orbicu- itive (1); highly eusocial (2)” (reviewed in Chavarrıá lariae) (Coddington, 1990). Qualitatively different and Carpenter, 1994). steps of frame, radius, hub, temporary spiral, and It has been suggested that the acquisition of social sticky spiral construction (Coddington, 1990) have not behavior has greatly increased the rate of evolution only supported the monophyly of Orbiculariae, but and, in its turn, provided a large amount of variation also determined the homology in some derived groups in which web architecture was no longer recognized among eusocial bees (Michener, 1974). In fact, it is as an orb (Griswold et al., 1998). widely known, for example, that animal architecture It would seem that with all the interest in the deriva- evolves as an extension of builders’ phenotype (Han- tion of social habit among corbiculate apines, it would sell, 1984). Just as morphological adaptations may ac- be profitable to study the behaviors that constitute quire new functions through time, so can architectural sociality. As stated above, most analyses to date col- traits. The architecture of many social insects includes lapse all behavioral variation into a single character structures serving functions other than retaining the with perhaps three states. Unfortunately, to find single brood (Fig. 1). These functions may allow the builder to or multiple origins in such a study provides little infor- exploit new opportunities and, when evolving rapidly, mation and certainly no meaningful statement about may obscure the purpose of the original structure. Re- the stepwise modification of behavior that leads to the cently an analysis of nest architecture in social wasps charismatic taxa we see today. Accordingly, I replaced has shown that such information is highly valuable the single character “sociality” with 42 characters in and is in agreement with morphological data (Wenzel, 65 derived, states, many of these homologized for these 1993; Wenzel and Carpenter, 1994). taxa for the first time. Using characters based on nest

᭧ 2002 by The Willi Hennig Society All rights reserved. 140 Fernando B. Noll construction, food storage, worker reproduction, fe- (Felsenstein, 1985) and Jackknife (Farris et al., 1996) male–female interactions, and food provisioning, a cla- in Winclada and Bremer support (Bremer, 1988, 1994) distic analysis of corbiculate Apidae (Euglossini, Meli- analyses in NONA. ponini, Apini, Bombini) is provided. Ten of 19 multistate characters were treated as additive either because there seems to be a logical intermediate condition between two end-points or because MATERIALS AND METHODS tradition has interpreted the characters as progressing through the transformations shown here. The rationale for coding is explained on a character-by-character ba- The 42 characters chosen for the analysis (Table 1; sis in the Appendix. In three cases (characters 8, 24, and Appendix) were based on the literature (Michener, 25) a form of polymorphism (where bees will perform 1974) and original observations. Outgroup taxa, Epi- either of two behaviors from time to time) is coded as charis zonata and Centris pallida, were chosen from the a step intermediate between the alternatives. Although Centridini, the nearest tribal relative of corbiculate Api- the coding shown in this study includes additive dae within Apinae (Roig-Alsina and Michener, 1993; multistate characters, the resulting cladogram is identi- Michener, 2000). Cladistic analyses were performed us- cal when all characters are treated as nonadditive. Po- ing programs NONA version 1.8 (Goloboff, 1998) and larity of all characters is determined by outgroup com- Winclada version 0.9 (Nixon, 1999) using procedures parison. “mult* 50” and “max*” for tree searches and the The 23 taxa studied here were chosen to represent branch-swapping option tree bisection–reconnection. several terminals in each tribe. Some species were Branch support was estimated using Bootstrap coded identically, but they were included to illustrate

TABLE 1 Data matrix of character states in Appendix

1234 123456789012345678901234567890123456789012 Source

Epicharis zonata 0000000000000000±0±0000000000±0000000±0±00 Michener (2000) Centris pallida 0000000000000000±0±0000000000±0000000±0±00 Michener (2000) Eulaema nigrita 1111000111020000±0000000010100001100000000 Zucchi et al. (1969); Pereira-Martins and Kerr (1991); Santos and Garo´falo (1994) Eufrisea auriceps 1100000110000000±000000001000000??000±0±00 Zucchi et al. (1969); Garo´falo (pers. comm.) Euglossa cordata 1111000121020000±1010000010100001100000000 Garo´falo (1985, 1992) Bombus morio 0111011231010110±0211111111211100010010010 Zucchi (1973) Bombus atratus 0111011231010110±1111111111211100010010010 Zucchi (1973); Bourke (1994) Bombus nevadensis 0111011231010010±??11111111211100010010010 Hobbs (1965); Plowright (1977) Bombus terrestris 0111011231010110±1111101111211100010010010 Michener (1974); Honk et al. (1980); Fisher and Pomeroy (1990); Bourke (1994) Apis mellifera 011202033212100112222102212322210001120120 Michener (1974) Apis dorsata 011002033212100112222102212322210001120120 Michener (1974) Scaptotrigona postica 011212123201100112320002111322201111111121 Sakagami and Zucchi (1963) Plebeia minima 011112023101100112320002111322201111111121 Zucchi et al. (1999) Melipona quadrifasciata 011212123201100102320002111322201111111121 Sakagami et al. (1965) Frieseomellita varia 011212023101100112220002111322201111110121 Zucchi et al. (1999) Nannotrigona testaceicornis 011112123201100112320002111322201111111121 Sakagami et al. (1993) Meliponula bocandei 011212123201100112320002111322201111111121 Sakagami et al. (1977) Leurotrigona mulleri 011112023101100112220002111322201111110121 Sakagami (1982) Trigonisca duckei 011112023101100112320002111322201111111121 Sakagami (1982) Geotrigona mombuca 011212123201100112320002111322201111111121 Lacerda et al. (1998) Oxytrigona tataira 011212123201100112320002111322201111111121 Silva-Matos et al. (1997) Paratrigona lineata 011212123201100112320002111322201111111121 Zucchi et al. (1997) Trigona spinipes 011212123201100112320002111322201111111121 Zucchi and Sakagami (1972; pers. obs.)

᭧ 2002 by The Willi Hennig Society All rights reserved. Behavioral Phylogeny of Corbiculate Bees 141 the limits of this analysis with respect to separating (Bombini ϩ (Meliponini ϩ Apini)). Behavioral charac- the most apical taxa. These taxa represent all those for ters supported the monophyly of all tribes, except for which information extends beyond anecdotal reports. the Euglossini. Homoplasies were found in the follow- ing characters: collection and storage of aromatic chemicals by the males (character 1, Eufriesea auriceps and ϩ RESULTS Eulaema nigrita Euglossa cordata), nest coating (character 4, Apis mellifera, Frieseomelitta varia, and Nannotri- gona testaceicornesϩ Trigona spinipes), wax envelope The analysis found a single most parsimonious cla- covering the brood area (character 7, Bombini and N dogram, with a length of 81, a consistency index (CI) testaceicornesϩ T. spinipes), brood cell architecture (char- of 81, and a retention index of 94 (Fig. 2). The resulting acter 10, Apini and N. testaceicornesϩ T. spinipes), cell cladogram (Fig. 2) agrees with a previous analysis us- construction and oviposition (character 18, Euglossa ing morphological characters (Roig-Alsina and Miche- cordata and Bombus atratusϩ Bombus terrestris), feeding ner, 1993) and combined morphological and molecular pockets (character 23, reversion in B. terrestris), B. ter- data (Chavarrı´a and Carpenter, 1994): Euglossini ϩ restris and Apiniϩ Meliponini), cell closure and cell

FIG. 2 Single most parsimonious cladogram from Apinae bees based on behavioral characters. Length ϭ 81, consistency index ϭ 81, retention index ϭ 94. The data matrix upon which this cladogram is based is presented in Table 1; character descriptions are given in the Appendix. Values in the boxes represent bootstrap, jackknife, and Bremer support, respectively. Black circles represent uncontroverted synapomorphies and white circles represent homoplasies based on a delayed optimization.

᭧ 2002 by The Willi Hennig Society All rights reserved. 142 Fernando B. Noll collar (characters 32 and 33, Eul. nigritaϩ Eug. cordata Euglossini (Fig. 2; Appendix) because most data relate and Meliponini) (Fig. 2; Appendix). to social interactions and this poorly known tribe con- High support for the corbiculate Apidae and also for sists of solitary, communal, and parasitic species (Kim- the Bombini, Apini, and Meliponini was found using sey, 1984). In the most recent study of Euglossini based bootstrap, jackknife, and Bremer branch support analy- on morphology (Engel, 1999), parasitic forms appeared ses (Fig. 2). twice and the solitary Eufriesea is not basal. Thus, it seems plausible that the communal behavior observed in Eulaema and Euglossa could be basal for euglossines DISCUSSION and the origin of social behavior could be basal for all corbiculate bees. In this way, the “solitary” behavior of Eufriesea would be secondarily derived. Some species Dissecting sociality into the 42 behavioral compo- have both communal and solitary colonies (Garo´falo nents (Appendix) shows that these characters evolve et al., 1998) and, in fact, several females were observed neatly with little homoplasy. A CI of 0.81 is high for a sharing the same nest site in Euf. auriceps (Kimsey, study of 23 taxa (Sanderson and Donoghue, 1989). This 1982), suggesting some kind of communal behavior. result is consistent with the general principle that care- However, because most colonies studied behaviorally fully defined behavioral characters perform as well come from trap nests (Garo´falo et al., 1998), it is possible as (de Queiroz and Winberger, 1993) or better than that this collection method may affect their behaviors. (Coddington, 1990; Wenzel, 1993) morphological char- Also, it is plausible that social behavior in euglossines acters. Furthermore, direct study of these characters varies intraspecifically. marks a decided improvement over plotting on a tree The clade Bombini ϩ Meliponini ϩ Apini is sup- based on molecular data (e.g., Cameron, 1993; Cam- ported by 12 synapomorphies (Fig. 2; Appendix). eron and Mardulin, 2001; Lockhart and Cameron, Some social-behavior characters can show how the 2001), and claims that such molecular studies provide basic pattern of social organization has possibly a better “test” are mistaken (Wenzel, 1997) because evolved. Bombines, meliponines, and apines have col- they are based on other characters that may represent ony hygienic habits; they build single-layered cells other selective forces and do not constitute a examina- that are made of wax and resin, and food (pollen and tion of the characters of interest (Wenzel, 1997). nectar) is stored before being used (although pollen Even though similar trees, at the tribal level, are itself can be used in bombines). Most Bombini show presented here (behavioral data) and in Chavarrıá and both solitary and social behavior. In the solitary phase, Carpenter (1994) (total evidence analysis), it is im- many plesiomorphic character states were found, es- portant to point out that only three behavioral charac- pecially those for nest construction. Also, several char- ters were used in the Chavarrıá and Carpenter (1994) acter states in Bombus are plesiomorphic in relation to matrix, and only one character is shared between both Apini ϩ Meliponini but apomorphic as compared with matrices (character 2—nest site; see Appendix). Also, a Euglossini. Although Bombus is considered primitively morpho-behavioral connection between both matrices eusocial, it shares many behavioral apomorphies with can be made only on the corbicula (Chavarrıá and Apini and Meliponini (Table 1; Appendix). The inter- Carpenter, 1994) and pollen collection behavior (char- mediate nature of Bombini that makes it the sister acter 26—pollen transportation; see Appendix). group of Apini and Meliponini is usually overlooked The monophyly of the corbiculate Apidae is sup- because several autapomorphies (especially those re- ported by four synapomorphies mainly based on nest lated to the nest ontogeny) make comparisons with and cell construction (Fig. 2; Appendix). Unlike other other social bees difficult. Bombus species present ex- bees, apines take advantage of preexisting cavities in- pandable cells and unusual larval feeding behavior if stead of excavating their own nests. Also, they prepare compared to the other Apines. However, at least in the cavity before cell construction and they use vegeta- some species such as Bombus nevadensis (Hobbs, 1965), ble material in their construction, a state not exten- the first brood cells are built simultaneously and ex- sively observed in other bees. panded after the queen lays one egg in each cell. This Behavioral data did not support monophyly of the is the same as observed in Apini ϩ Meliponini. In

᭧ 2002 by The Willi Hennig Society All rights reserved. Behavioral Phylogeny of Corbiculate Bees 143 addition, in the same first brood cells, the queen primes This problem is no longer the case for corbiculate Api- the cell with a pollen lump. In this case, it is possible dae. The present results show that behavioral data are that Apini and Bombini have acquired progressive very useful for phylogenetic reconstruction in corbicu- provisioning independently. late bees and they should be used (wherever possible) In both highly eusocial groups, Apini and Melipon- in future analyses. ini, tasks performed by queens and workers are very specific and castes are dependent upon one another. For some traits, interpretation of homology is difficult APPENDIX: CHARACTER CODING because of the specialized habits in honey bees (i.e., aerial nests, swarms occurring with the presence of the mother queen, absence of real storage pots). However, 1. Collection and storage of aromatic chemicals by attention to these traits should not obscure other possi- the males. [0] absent; [1] present. ble homologies, like hygienic habits, building behavior Euglossines are markedly different from other bees of the brood cells, food storage behavior including the because the males not only have structures involved patterns in pollen storage behavior and honey produc- with collection and storage of aromatic chemicals tion, division of labor, food exchange, presence of royal (Kimsey, 1980, 1984) but they also perform the same cells, morphological caste differences (not only size– movements as females in the acquisition of such sub- based differences), obligatorily queenright colonies, stances (Kimsey, 1984; Roubik, 1989). and no task overlap between queen and workers. 2. Nest site and general structure. [0] cells excavated The relationships shown in Fig. 2 support not only in burrows; [1] free-standing constructions of im- ported material. the single origin of eusociality in Bombini, Apini, and Zucchi et al. (1969) and Michener (1974) emphasized Meliponini but also the single origin of the highly euso- the importance of this character. Most solitary bees dig cial habits in Apini and Meliponini. This is consistent burrows for nests. They usually excavate their own with the hypothesis that eusociality originated from a cells connected to the burrow or a system of branching primitive stage, probably similar to that observed in burrows and more than one cell is built at the same some Euglossini (i.e., matrifilial associations). The site. As many of these characters have not been defined foundress female nests in a preexisting cavity and pre- in a cladistic sense, a rationale for statements of homol- pares the space with resin and other materials. Later, ogy and identity is offered here. Apines always con- she stores building material. After the emergence of struct one or more cells usually in preformed cavities the first daughters, the dominant, the mother or the in wood, pithy stems, human constructions, or vertical oldest daughter, stays in the nest and she is usually banks of various sizes and shapes. aggressive toward subordinates. Dominance is based In Euglossini, Euglossa (except the parasite species on the oophagy of the subordinates’ eggs. However, and the subgenus Glossura (Kimsey, 1987), Eufriesea, it is not clear whether the ritualized behavior and the and Eulaema follow the pattern mentioned (Fig. 1A). origin of trophic eggs as observed in stingless bees are Even Eufriesea takes advantage also of preexisting cavi- homologous with the Euglossini or whether they have ties when their nests are partly exposed (Kimsey, 1987). secondarily evolved because in Apini and Bombini, Bombini usually nest in a variety of sites—above chemical control of the workers’ ovarian development ground and underground (Fig. 1B)—and they can be by queens seems plausible (Ro¨seler et al., 1981). found in bird boxes and tree hollows, but, as stated by It is important to note how congruent the characters Sakagami (1976), they never excavate their own nests. of Appendix 1 are when they are interpreted not only Apini build both aerial and cavity nests. According as simple definitions but also as a source of characters to Alexander (1991), nesting in cavities is the plesio- different from those used before. For example, the mo- morphic state (Fig. 1D). Engel and Schultz (1997) sug- nophyly of the highly eusocial Apini and Meliponini gest that cavity-nesting behavior in honey bees is sec- is supported by 14 apomorphies (Fig. 2; Appendix 1). ondarily derived, but the plesiomorphic state of the Proctor (1996) found that most systematists exclude whole tribe is also nesting in cavities. behavioral characters because they are not available. Meliponini nest preferentially in tree trunks (Fig. 1C),

᭧ 2002 by The Willi Hennig Society All rights reserved. 144 Fernando B. Noll but some species are found in termite, ant, and wasp found exclusively in stingless bees (Wille, 1983). Con- nests, and even human constructions (Sakagami, 1982). nectives are horizontal and pillars are vertical. They Aerial nests are observed in some Trigona such as T. are responsible for supporting all other structures spinipes, Trigona hyalinata, and Trigona fuscipennis. within the lining batumen (Fig. 1C). The cells of the 3. Storing of nest construction material. [0] absent; combs are in contact with one another or connected [1] present. by a small pillar or connective of soft cerumen. Pillars Materials employed in nest construction are conser- connect the combs with one another and with sur- vative in the taxa examined here. They are composed rounding structures. Pillar-like structures can be found mostly of vegetable resin that can be used in various in some Apis, but homology is questionable and will ways such as mixed with other vegetable substances not be proposed here. or with endogenous secretions. In the Euglossini mate- 6. Hygienic habits. [0] waste materials are kept in rials brought from the field are deposited in a common the nest; [1] waste materials are kept in the nest but place and later are used in tasks like lining the cavity are clearly removed from the brood area to the periph- and cell construction. In Bombini resin collection is ery, [2] waste materials are carried away from the nest, practically unknown. Based on direct observations in [3] waste materials are removed and deposited in the B. atratus, no external material, except nectar and pol- lowermost involucral sheet. Nonadditive. len, is collected by the foragers. This suggests that if Because nests in solitary bees are not reused, waste something is mixed with wax, it is probably pollen materials like feces, corpses, and pieces of cocoon are (Michener, 1974; Sakagami, 1976). In the Apini large kept in the nest. The same pattern, i.e., waste materials amounts of propolis are collected and used mainly in kept in the nest, is observed in euglossines. In bom- nest coating. As in Apini, Meliponini also collect some bines, although waste material is kept in the nest, it is propolis, but a variety of different materials are used removed from the brood area to the periphery, clearly in various ways (reviewed in Roubik, 1989). demonstrating hygienic behavior. In meliponines and 4. Nest coating. [0] absent; [1] lining of all or part apines, waste material is carried away from the nest: of the cavity with resin; [2] thick layer of propolis the workers usually fly away with the material in their covering the nest cavity floor, walls, and ceiling. mandibles. However, in some meliponines like T. spin- Solitary bees usually line their cells with glandular ipes and Cephalotrigona capitata, waste materials are also substances, but because they basically excavate their removed but, instead of being carried away, they are cells only, there are no other structures of spaces in their deposited in the lowermost involucral sheet. Such ma- nests. In the in-group taxa analyzed, lining cavities is terials will form a structure called a scutellum that, widely distributed. In euglossines (except Eufriesea) according to Zucchi and Sakagami (1972), could help and bombines all or part of the cavity is lined with in nest thermoregulation. resin while in apines and meliponines a thick layer of 7. Wax envelope covering the brood area. [0] absent; propolis covers the nest cavity floor, walls, and ceiling. [1] present. Lining the cell cavity seems to be an important step An envelope is absent in Euglossini (envelope-build- in the evolution of corbiculate apids, because the addi- ing Euglossa do not use a preexisting cavity and homol- tion of resin in the cavity would seem to be an im- ogy of this envelope with others is in doubt), in Apini portant trait in the maintenance of a barrier and hy- (even in those that nest in cavities), and in some sting- gienic conditions in the nest. A phylogenetic tree based less bees. It is present in Bombini and most of the on morphological data (Camargo and Pedro, 1992) Meliponini. In meliponines the envelope usually de- shows that stingless bees have secondarily lost this limits the brood area (Fig. 1C) only, while in bumble- trait. This seems to be a reversal especially because bees both combs and storage pots are covered with the such species nest in very narrow places, involving sev- envelope. It was observed in Bombus mendax (Haas, eral modifications like the loss of comb structure. 1976) that the envelope covers only the brood area and 5. Vertical pillars and connectives extensively built that the storage pots are kept outside. In B. atratus the in the nest used for stabilizing brood cells. [0] absent; envelope appears only in periods with low tempera- [1] present, ture, suggesting its thermoregulatory function (Zucchi, Among corbiculate bees, connectives and pillars are 1973). An envelope is absent in some stingless bees but

᭧ 2002 by The Willi Hennig Society All rights reserved. Behavioral Phylogeny of Corbiculate Bees 145 it probably is a reversal (see character 4), because most observed have cell walls with only one layer, not in- basal species possess an envelope. In addition, it is not cluding a special lining made of a self-produced sub- clear whether the envelope originated once (in this stance. In Euglossa, cells are usually composed of resin case it has appeared in the branch Bombini ϩ Apini ϩ while in the other apines they are composed mostly Meliponini and secondarily lost in Apini) or twice, of wax and some other secreted materials. independently in Bombini and Meliponini. 10. Brood cell architecture. [0] independent cells; [1] 8. Materials employed in cell construction. [0] non- cells constructed independently but clustered together; vegetable material from the substrate; [1] materials [2] cells serially constructed as a comb so that the cells other than substrate, composed of vegetable material have at least one wall in common. brought from outside the nest such as resin and a small Most solitary bee cells are made by excavation while quantity of secreted material; [2] secreted material in corbiculate bees cells are made by building. Cells mixed with resin or pollen; [3] secreted material. Non- are radially symmetrical and oval, made more or less additive. in contact. Such traits are clearly observed in euglos- Solitary bees usually excavate their nests. Besides sines. In addition, species of Euglossa and Eulaema build secretions employed in cell lining, usually soil or other cells clustered together (Fig. 1A). Bombus nevadensis nonvegetable material from the substrate is employed. (which differs from other Bombus) makes one cell for Apparently, the most striking feature in these taxa is each egg as in euglossines. Most Bombus lay more than the change to an extensive use of vegetable material one egg per cell (Fig. 1B), at least for the second brood, but that habit is unique to them and unlike other bees. brought from outside the nest, like resin to make brood (See character 14). In meliponines and apines, cells are cells, and the mixture of this with secretions—namely, serially constructed as a comb so that the cells have at wax. Apparently the proportions were gradually least one wall in common (Figs. 1C and 1D). In the changed—initially much more resin and little wax, and former, cells are still radially symmetric and oval while later much more wax than resin. In fact, in euglossines in the latter cells are hexagonal. Meliponines, such as wax glands are present but much less developed than some species of Plebeia, and Frieseomellita have the in the other cobirculate bees (Cruz-Landim, 1963). In comb structure disintegrated although their cells usu- euglossines, cells of Euglossa are basically made of resin ally share walls (see character 4). (Garo´falo, 1985, 1992). Some variations are observed 11. Cell shape. [0] elliptical; [1] hexagonal. in Eufrisea and Eulaema—the former uses pieces of bark Unlike other bees, corbiculate apids nest in places extensively and the latter collects large quantities of much larger than their own cells (see character 1). In mud and feces. In bumblebees and stingless bees cells this way they have a speciﬁc shape difference from are composed mostly of secreted material mixed with those of solitary bees that mold their cells according to pollen and resin, respectively, while in Apini they are their excavated burrows. Euglossines, bombines, and composed mostly of secreted material even though meliponines have cells that are ovoid, elliptical in cross materials like pollen, pieces of cocoon, and propolis section (Figs. 1A–1C). In Apini, cells are hexagonal have been extensively found in their nests (Miche- (Fig. 1D), but as stated by Seeley (1985) such conforma- ner, 1974). tion has evolved from a nonhexagonal shape favoring 9. Cell walls. [0] cells are excavated and are lined the economy of wax. with endogenous substances; [1] cell walls differenti- 12. Reuse of cells or space where larvae are reared. ated into outer and inner layers with different texture, [0] absent; [1] material reused but cells reconstructed; without special lining; [2] cell walls have only one layer [2] cells or spaces always reused. Nonadditive. made of resin; [3] cell walls have only one layer made Another important characteristic found in this group of secreted material. Nonadditive. is nest reuse by various generations. It is also found in Because cells are excavated in solitary bees, these other bees with some degree of sociality. In corbiculate spaces have a special lining made by endogenous secre- bees, brood cells can be reused as observed in some tions. In Eulaema and Eufrisea, however, cells are lined euglossines (Eul. nigrita Santos and Garo´falo, 1994, and with resin and not with secretions. All the other apines Eug. cordata Garo´falo, 1992) and apines (the classical

᭧ 2002 by The Willi Hennig Society All rights reserved. 146 Fernando B. Noll honey bee habit) or only the material used in the previ- cell construction is also seen when some workers de- ous cells is reused but new cells are reconstructed as velop ovaries. In apines and meliponines more than is widely observed in bombines and meliponines. one cell can be built before oviposition. Homology between cell reuse in Euglossini and 17. Royal cells. [0] absent; [1] present. Apini is unclear. Perhaps material reuse has evolved This character is problematic. In bombines cell size independently in meliponines and bombines. is not fixed (see character 14) and cells are expanded 13. Brood cell construction. [0] each cell construction constantly according to the development of the larvae. is performed from start to finish by the continuous The two remaining groups, apines and meliponines, work of a single female; [1] various individuals are have royal cells but built in two different ways. In engaged in the construction of one cell at the same time. meliponines, except Melipona, royal cells are larger than In solitary bees cell construction is performed from worker and male cells (Sakagami, 1982). In honey bees, royal cells differ strikingly from regular cells not only start to finish by the continuous work of a single fe- in size but also in shape (Fig. 1D). Gynes in all species male. Such behavior is widely observed in euglossines. are reared in much larger, irregularly shaped cells, Even in bombines, queens initiate cells and complete never forming any sort of comb, and typically hanging the first stage of construction in mature nests (although from the edge of the comb or from the upper margin a cell may be expanded by workers much later when of any holes in the comb (Michener, 1974). In addition, the growing larva requires more space). Thus, this as- drones are produced in larger cells than workers. Thus, pect of progressive provisioning (which contrasts with the homology of the character states remains specula- mass provisioning of solitary bees) begins in a compa- tive. rable way. In apines and meliponines various individu- 18. Cell construction and oviposition. [0] always by als are always engaged in the construction of a single the same female; [1] by the same female but she can cell and under normal conditions these individuals open a cell of other females, eat the egg, and lay her will not lay eggs in that cell. own egg; [2] always in cells built by other females. Non- 14. Number of eggs per cell. [0] one; [1] more than additive. one. The same single female in solitary bees always per- Several eggs are laid in each cell in Bombini (Fig. forms cell construction and oviposition. This is also 1B). This is a synapomorphy for Bombini. Although observed in euglossines and bombines in the solitary some Allodapini build communal chambers (Miche- phase but, when more than one female is present one ner, 1974), all other bees lay one egg per cell as is also female can eventually open a cell from another female, observed in B. nevadensis (Hobbs, 1965). eat the inside egg, and lay her own egg. In apines and 15. Cell walls. [0] fixed; [1] expansible. meliponines the queen always lays in cells made by In most bees, cells are constructed with a predeter- other females. 19. Oviposition in the presence of more than one mined size so that the brood will fill the whole inner female. [0] more than one female can lay eggs at the space of the cell. In bombines, cells are distended ac- same time; [1] only the queen lays eggs initially, but cording to the growth of the brood (reviewed in Saka- workers start building their own cells and ovipositing gami, 1976). in the end of the cycle (usually when the queen is 16. Brood cell construction. [0] each cell is followed rearing males); [2] only the queen lays eggs and worker by oviposition; [1] more than one cell can be built reproduction is rare; [3] queens and workers lay eggs before oviposition. but in the latter the eggs are mostly nonviable. Nonad- In most solitary bees oviposition follows cell con- ditive. struction immediately. Such a trait is observed also in Because the outgroups are usually solitary, outgroup bombines and euglossines in the solitary phase. How- comparisons are not possible for this character. How- ever, when several individuals are living together, it is ever, communal behavior observed in other groups possible to see more than one cell being constructed and even euglossines suggests that the plesiomorphic in euglossines and in bombines (Garo´falo, 1992, and state is more than one female being able to lay eggs at Sakagami, 1976, respectively). In bombines, multiple the same time. In Eug. cordata many individuals lay

᭧ 2002 by The Willi Hennig Society All rights reserved. Behavioral Phylogeny of Corbiculate Bees 147 eggs but the dominant female opens cells and eats progressively to developing larvae are observed in the subordinates’ eggs (Garo´falo, 1992). In bumblebees apines and bombines. only the queen lays eggs initially, but workers start 23. Feeding pockets. [0] absent; [1] present. building their own cells and ovipositing toward the In some bumblebees pollen is fed to larvae through end of the cycle (usually when the queen is rearing the pocket made at the side of the larval cell. In non- males). In apines only the queen lays eggs and worker pocket-makers, pollen is given directly by perforating reproduction is rare. In meliponines queens and work- the cell. As stated by Williams (1994), the use of such ers lay eggs but in the latter the eggs are mostly sterile. a character in phylogenetic analysis is controversial 20. Cell provisioning. [0] always by the same female especially because some species like B. nevadensis can that oviposited in the cell; [1] by the same female that behave as pocket-makers and non-pocket makers at oviposited in the cell in the solitary phase only; [2] successive stages of colony development (Hobbs, 1965; always by other females than that which oviposited in Plowright, 1977). However, as stated by Sakagami the cell. (1976), the explanation can be derived from the habits Euglossines and bombines in the solitary phase pro- for provisioning food to the first cell. Usually, the larvae vision and oviposit in the cell alone, of course, as ob- receive additional pollen through gaps in the canopies served in solitary bees. In bombines in the social phase, of both ends of the cells in all subgenera so far ob- apines, and meliponines, workers always perform pro- served. This resembles the feeding by means of visioning. pockets. 21. First brood cell provisioning. [0] provisions pro- 24. Brood food origin. [0] comes straight from the vided in toto prior to oviposition and cell closure; [1] field; [1] comes from both field and storing places; [2] is always from storing places. provision of the egg chamber of first brood is done In euglossines (and in solitary bees), brood food before the eggs are laid but more food is added before comes straight from the field (plesiomorphic state). An the larvae mature; [2] provisions composed entirely or intermediate step is expressed in Bombus: brood food almost entirely of secreted material fed to developing can be brought by the females from the field and di- larva in the open cell. rectly offered to the larvae or it is stored in pots and Provisioning brood is divided in two types, mass later offered to the larvae. In a third step, observed in provisioning and progressive provisioning. According apines and meliponines, brood food is always depos- to Michener (1974) and Sakagami (1976), founder ited first and later used to feed the larvae. queens in Bombini show high variation. In the plesio- 25. Brood food composition. [0] pollen and nectar; morphic state, provisions are provided in toto prior to [1] pollen, honey, and glandular secretions; [2] glandu- oviposition and cell closure. Such mass provisioning lar origin. observed in solitary bees is also observed in euglos- In solitary bees and euglossines food is mostly pollen sines and meliponines. In some bumblebees, provi- and nectar. In bombines, apines, and meliponines, be- sions for the egg chamber of the first brood are supplied sides pollen and nectar, glandular secretions are an before the eggs are laid, but more food is added before important part of larval diet, while in apines it seems the larvae mature. In apines, the provisions are com- to be mostly glandular (Webster and Peng, 1988). In posed entirely or almost entirely of secretions that are fact, according to Cruz-Landim and Costa (1998), hypo- offered to the developing larvae in open cells. pharingeal glands in Euglossini are similar to those 22. Feeding the second brood. [0] provisions pro- of other solitary bees (plesiomorphic state). Bombini, vided in toto prior to oviposition and cell closure; [1] Meliponini, and Apini present similar patterns, but provisions provided more or less progressively to de- Apini and Meliponini possess glands in a different veloping larva. apomorphic condition. In many bombines, cells of the second and subse- 26. Pollen transportation. [0] dried; [1] in a firm mass quent brood do not receive food deposits before ovipo- together with a little nectar. sition. In this case, provisions provided in toto after Pollen as food for the individual bee is found in oviposition and cell closure as observed in euglossines the crop, but pollen for use in the nest is ordinarily and meliponines. Provisions provided more or less transported on the pollen-carrying hairs or scopa of

᭧ 2002 by The Willi Hennig Society All rights reserved. 148 Fernando B. Noll females. It is carried in dry, somewhat powdery form This is also observed in Eul. nigrita (Zucchi et al., 1969), in solitary bees or in a firm mass stuck together with but Santos and Garo´falo (1992) and Pereira-Martins a little nectar as observed in Apinae (Michener, 1974). and Kerr (1991) have not confirmed such observations. 27. Food storage. [0] absent, food is deposited di- In apines and meliponines, young workers start to rectly in brood cells; [1] honey and pollen are separately work with wax and cerumen away from the brood deposited in storage pots; [2] honey and pollen are area of the nest, and then they shift toward building separately deposited in combs. and provisioning brood cells. From brood care they As previously observed, in solitary bees and euglos- go to foraging: first they receive incoming nectar and sines food storage is absent and its deposition is direct dehydrate it before becoming actual foragers. This in brood cells. Food storage is largely observed in well-known form of temporal polyethism has evolved apines, bombines, and meliponines. In bombines, and independently in ants, bees, and wasps (Wilson, 1971). meliponines, honey and pollen are separately depos- 30. Food exchange among adults. [0] absent; [1] via ited in storage pots (Figs. 1B and 1C), which are special- brood cells or brood food only; [2] direct via oral troph- ized cells to store food. In some Bombus, cocoons can allaxis. also be used as a food-storing place (Michener, 1974; The amount of contact among adults in a colony Sakagami, 1976). In honey bees, honey and pollen are varies greatly in the different taxa. Contact is com- separately deposited in combs. This group has lost pletely absent in euglossines. In Bombus, foragers re- the specialized structures observed in Bombini and turning to the nest put the food in special pots or Meliponini. Probably the comb has evolved to be a pockets, instead of placing it in brood cells. In such multiuse structure to accommodate both brood and places other adults can feed themselves. In highly euso- food separately. cial bees, nectar is not ordinarily put into storage places 28. Foraging behavior. [0] all individuals forage; [1] or into brood cells or fed to larvae by the returning all individuals forage but dominants stay in the nest foragers. Instead, the foragers transfer the nectar to much more than forage; [2] queen forages only until other individuals; most of them specialized in domestic the first brood emerges; [3] queens never forage. Non- tasks (Michener, 1974, Webster and Peng, 1988). additive. 31. Nectar exchange among the individuals before In solitary bees and Eufrisea all individuals forage. being stored. [0] nectar is deposited directly in the In Eulaema and Euglossa, all individuals forage, but brood cell; [1] nectar is directly deposited in the pots; dominant individuals stay in the nest more than forage. [2] nectar is exchanged among workers and is dehy- In bombines, the founding queen forages only until drated before being stored. the first brood emerges. After that, she never forages again. Experimental removal of all workers in B. atratus In euglossines nectar is deposited directly in the showed that the queen never starts foraging behavior brood cell. In bombines, a returning forager with nectar again (Zucchi, personal communication). In apines and deposits it directly in the honey pots. In eusocial bees, meliponines, queens never forage. foragers transfer nectar to receivers that are workers 29. Division of nonreproductive labor. [0] absent; [1] that have not yet begun foraging activities. Receiving larger individuals perform activities like foraging and bees offer nectar to other bees. At the same time, the smaller individuals perform nest activities; [2] young nectar is being converted to honey by treatment with individuals start to work with wax and cerumen, away invertase from the hypopharyngeal glands of the bees from the brood nest, and then some shift toward build- (Michener, 1974; Webster and Peng, 1988). ing and provisioning brood cells (i.e., age polyethism). 32. Time of cell closure. [0] immediately after an egg From brood care they go to foraging: first they receive is laid; [1] upon pupation. incoming nectar and dehydrate it before becoming ac- In most bees cells are closed immediately after an tual foragers. egg is laid. The only exception is observed in Apis.In In solitary bees and euglossines nonreproductive di- this group, cells are capped with wax only 2 or 3 days vision of labor is absent. In bombines, larger workers after pupation (Michener, 1974). perform activities like foraging and smaller workers 33. Cell closure. [0] with mandibles; [1] with abdomi- do nest activities (Zucchi, personal communication). nal insertation and rotating behavior.

Bees that fashion complete cells from substrate mate- conditions before selecting the final nest site, which is rials (Colletidae, Andrenidae, Melittidae, Halictidae, sought after temporary settlement of the mass in an most Anthophorini) usually construct their cell caps intermediate bivouac. Association with the old nest is in the form of flat spirals or as inverted conical plugs. terminated by departure of the mass, resembling more The cap is constructed with the mandibles by placing the swarm-founding polistine wasps (Jeanne, 1991) small strips of material on the upper rim of the open than stingless bees. In stingless bees, swarming is char- cell (Stephen et al., 1969; Rozen, 1984). Apines form acterized by the construction of the new nest over a dome-shaped cell caps from masticated material (wax period of days or weeks by workers who transport or resin). In euglossines and meliponines, capping cells resources (wax, nectar, pollen) to the chosen site repeat- can be divided in two phases: (1) rotation phase: in- edly from the mother colony. Eventually, when the nest serting the metasoma into the cell and rotating the is ready the daughter queen arrives at the nest site body around the cell, the female closes the orifice by unaccompanied by a large mass of excited workers. bending the collar inward with mandible and forelegs; The connection with the old nest continues for a consid- (2) side work: the metasoma is withdrawn and the erable time. In both cases, queens are incapable of orifice is closed by pressing the collar with mandibles. founding a new nest, being completely dependent on 34. Cell collar. [0] absent; [1] present. worker care. Considering swarming in both groups as A preoviposition behavior, similarly observed in eu- homologs, it is possible to assume that the plesiomor- glossines and meliponines, begins with the improve- phic state could be observed in meliponines. Observa- ment of the cell collar. At this phase, the collar is thick tions of some swarming processes in stingless bees and short, and it is gradually lengthened through fur- demonstrate that workers prepare all nest structures ther manipulation (Zucchi et al., 1969). before initiating cells. Lining cavities and building the 35. Removal of cell walls after cocoon spinning. [0] nest entrance and storage pots are the first tasks of the absent; [1] present. workers. In some way, this resembles euglossines that Materials employed in cell construction are rarely build the nest entrance and line cavities before starting reused in all bees as a regular trait. However, workers cell construction (Garo´falo, 1992; Santos and Garo´- in meliponines and bombines remove the wax from falo, 1994). cell walls after cocoon spinning. This material is reused 37. Queen (or lone ␣) foraging. [0] Pollen-collecting in the construction of new cells or in other structures and carrying apparatus as in ␣; [1] with such structures in the nests (Sakagami, 1982; Michener, 1974). Ac- absent, must start colony with swarm of workers. cording to Webster and Peng (1988), cell destruction See character 36. is due to the amount of feces remaining at the bottom 38. Social regulation. [0] dominance behaviorally of the cells. Because the brood food in Apini is mostly mediated; [1] dominance pheromonally–behaviorally glandular (see character 26), it was suggested that the mediated; [2] dominance pheromonally mediated. amount of feces is less than in other groups. This sug- Nonadditive. gested that hygienic habits based on cell destruction Female–female interactions usually display domi- are not as important as in other tribes (Webster and nance–subordination behavior in various groups of Peng, 1988). Also, not removing cell walls after cocoon social insects, and this apparently is the first step in spinning can be secondarily derived. the evolution of social behavior. In euglossines, all indi- 36. Colony foundation. [0] solitary; [1] swarm. viduals are inseminated and have approximately the Solitary foundation is the basic mode of starting a same type of ovarian development, but in groups in new colony in most social insects. Swarming is rela- which females live together (Eul. nigrita, Zucchi et al., tively rare and found only in army ants, apines, meli- 1969; Eug. cordata, Garo´falo, 1992), interactions among ponines, and some social vespid wasps. As extensively females are practically absent; however, interactions discussed by various authors, many features differ be- are aggressive when one female comes close to a cell tween swarms of apines and meliponines. In honey constructed by another female or if she tries to “help.” bees, swarms can be characterized as a rather abrupt In Eug. cordata, one female exerts her dominance and quick departure. A large mass of workers and against other females behaviorally through a repertoire the mother queen leave the nest under highly excited of different behaviors (Garo´falo, 1992). In bumblebees,

᭧ 2002 by The Willi Hennig Society All rights reserved. 150 Fernando B. Noll dominance is maintained by means of pheromones size and discrete morphological differences (queens (Honk et al., 1980; Pomeroy and Plowright, 1982; Fisher lack apparatus for collection pollen, producing wax). and Pomeroy, 1990): the young queen, using phero- Nonadditive. mones and some aggressiveness, acts as the sole domi- Castes in bees can be physiologically, behaviorally, nant individual in the nest. In fact, overt aggressiveness and sometimes morphologically differentiated. Castes toward queens is rare in normal colonies. However, are absent in euglossines, but dominance behavior is when the queen becomes older, she probably loses the found in Eug. cordata. In bumblebees, castes differ strik- ability to control ovary development in workers chemi- ingly in mean size and sometimes also in coloration cally, and they start rearing their own brood. During (Michener, 1974). However, there are records of mated this phase, violent aggressive interactions are ob- workers and of colonies dominated by workers (Pla- served, and in some cases, workers kill the old queen. teaux-Queńu, 1961; Silva-Matos and Garo´falo, 1995). In Apis, pheromones secreted by the queen inhibit ovar- The differences between castes in highly eusocial bees ian development of workers (more details see Miche- are quite evident. The queen has no pollen-colleting ner, 1974; Zucchi et al., 1999). Queens have full ovarian apparatus or apparatus for producing wax or handling development, and workers’ ovaries are inhibited pher- construction materials. She can feed on stored honey omonally until the colony is queenless (some workers but during times of laying activity is dependent on develop ovaries). Chemical inhibition in stingless bees secretions produced by the workers. She also is not seems to be present also. Queens have full ovarian aggressive with natural enemies. On the other hand, development and workers have minor ovarian devel- workers have specialized pollen-collecting apparatus, opment. However, instead of having their ovaries in- wax-producing glands, and apparatus for handling hibited, their eggs are, in various cases, morphologi- construction materials. They also exhibit behaviors re- cally or behaviorally trophic. In queen-right colonies, lated to collecting food and colony defense. Workers, workers eggs are always laid away from the brood however, have lost the behavior patterns and anatomy food and, in some cases, they are laid outside the cells related to mating. (Zucchi, 1993; Zucchi et al., 1999). Before a queen ovi- 42. Provision and oviposition process. [0] absent; posits, various behavioral interactions occur between [1] present. queen and workers. Such behaviors can be ritualized One of the most interesting oviposition behaviors (Zucchi, 1993). among social insects is found in stingless bees. It in- 39. Morphologically trophic eggs. [0] absent; [1] volves highly complex, stereotyped, species-typical present. queen–worker interactions and oophagy. Called “pro- See character 38. vision and oviposition process” (Sakagami, 1982), it 40. Dominant female or queen disappearance. [0] has four main steps: (1) attraction of the workers and subordinates or workers behave as dominant [1] a sub- queen to growing and completed cells; (2) discharge ordinate or worker(s) never behaves as dominant. of larval food by workers and inspection of the cell Dominance is not documented in euglossines such by queens and workers; (3) oviposition by a worker, as Eul. nigrita (Zucchi et al., 1969; Santos and Garo´falo, oophagy by the queen, and queen oviposition; (4) cell 1994) or Euf. surinamensis (Zucchi et al., 1969). In Eug. closure. Although such a complex behavioral pattern cordata, if a dominant female disappears, a subordinate seems unique, its precursors may be found in Eug. can assume dominant status. Such succession also oc- cordata according to Zucchi et al. (1999). In this species, curs in Bombus (Bourke, 1994). In B. atratus, some work- the dominant individual guards and oviposits in the ers can be inseminated and carry on the colony until cells constructed and provisioned by the subordinates. the gyne production phase (Silva-Matos and Garo´falo, Before her oviposition, the dominant eats the subordi- 1995). In stingless and honey bees, other than a few nate’s egg (Garo´falo, 1985). workers involved in male production in orphan colonies (Michener, 1974), workers never act as dominant ACKNOWLEDGMENTS females or replace queens. 41. Morphological differences between castes. [0] no The author acknowledges financial support by FAPESP (Fundaçaõ castes; [1] castes differ strikingly in mean size; [2] mean de Amparo a` Pesquisa do Estado de Saõ Paulo PROC.01/02491-4)

᭧ 2002 by The Willi Hennig Society All rights reserved. Behavioral Phylogeny of Corbiculate Bees 151 and The Ohio State University PostDoctoral Fellowship. Thanks are Di Fiore, A., and Rendall, D. (1994). Evolution of social organization: extended to Ronaldo Zucchi and Carlos A. Garo´falo (Sa˜o Paulo A reappraisal for primates by using phylogenetic methods. Proc. University) for discussions and important information, John W. Wen- Natl. Acad. Sci. USA 91, 9941±9945. zel and Kurt M. Pickett (The Ohio State University), and three anony- Engel, M. S. (1999). The ®rst fossil Euglossa and phylogeny of the mous reviewers for reading the manuscript and for important sug- orchid bees (Hymenoptera: Apidae; Euglossini). Am. Mus. Nov. gestions. 3272, 1±14. Engel, M. S., and Schultz, T. R. (1997). Phylogeny and behavior in honey bees (Hymenoptera: Apidae). Ann. Entomol. Soc. Am. REFERENCES 90, 43±51. Farris, J. S., Albert, V. A., Kallersjo, M., Lipscomb, D., and Kluge, A. (1996). Parsimony jackkni®ng outperforms neighbor-joining. Alexander, B. (1991). A cladistic analysis of the genus Apis. In ªDiver- Cladistics 12, 99±124. sity in the Genus Apisº (D. R. Smith, Ed.), pp.1±28. Westview Press, Felsenstein, J. (1985). Con®dence limits on phylogenies: An approach Boulder, CO. using the bootstrap. Evolution 39, 783±791. Arntzen, J. W., and Sparreboom, M. (1989). A phylogeny for the old Fisher, R. M., and Pomeroy, N. (1990). Sex discrimination and infanti- world newts, genus Triturus: Biochemical and behavioral data. J. cide by queens of the bumble bee Bombus terrestris (Hymenoptera: Zool. 219, 645±664. Apidae). Anim. Behav. 39, 801±802. Ascher, J. S., Danforth, B. N., and Ji, S. (2001). Phylogenetic utility GaroÂfalo, C. A. (1985). Social-structure of Euglossa cordata nests of the major opsin in bees (Hymenoptera: Apoidea): A reassessment. (Hymenoptera, Apidae, Euglossini). Entomol. Gen. 11, 77±83. Mol. Phylogenet. Evol. 19, 76±93. GaroÂfalo, C. A. (1992). Nesting behaviour and nest structure of Basibuyuk, H. H., and Quicke, D. L. (1999). Grooming behaviours in the Euglossine bee Euglossa cordata. (Hymenoptera: Apidae: the Hymenoptera (Insecta): Potential phylogenetic signi®cance. Euglossini). Rev. Bras. Biol. 52, 187±198. Zool. J. Linnean Soc. 125, 349±382. GaroÂfalo, C. A., Camillo, E., Augusto, S.C., Jesus, B. M. V., and Bourke, A. F. G. (1994). Worker matricide in social bee and wasps. Serrano, J. C. (1998). Nest structure and communal nesting in J. Theor. Biol. 167, 283±292. Euglossa (Glossura) annectans Dressler (Hymenoptera, Apidae, Bremer, K. (1988). The limits of amino acid sequence data in angio- Euglossini). Rev. Bras. Zool. 15, 589±596. sperm phylogenetic reconstruction. Evolution 42, 795±803. Goloboff, P. A. (1998). NONA: A tree searching program. Program Bremer, K. (1994). Branch support and tree stability. Cladistics 10, and documentation. Version 1.8. [Available at http://www.cladistics. 295±304. com] Camargo, J. M. F., and Pedro, S. R. M. (1992). Systematics, phylogeny Griswold, C. E., Coddington, J. A., Hormiga, G.; and Scharff, N. (1998). and biogeography of the Meliponinae (Hymenoptera, Apidae): A Phylogeny of the orb-web building spiders (Araneae, Orbiculariae: mini-review. Apidologie 23, 509±522. Deinopoidea, Araneoidea). Zool. J. Linn. Soc. 123, 1±99. Cameron, S. A. (1991). A new tribal phylogeny of the Apidae inferred from mitochondrial DNA sequences. In ªDiversity in the Genus Haas, A. (1976). Paarungsverhalten und Nestbau der alpinen Hummel- Apisº (D. R. Smith, Ed.), pp. 71±87. Westview Press, Boulder, CO. art Bombus mendax (Hymenoptera: Apidae). Entomol. Germanica 3, 248±259. Cameron, S. A. (1993). Multiple origins of advanced eusociality in bees inferred from mitochondrial DNA sequences. Proc. Natl. Acad. Hansell, M. H. (1984). ªAnimal Architecture and Building Behaviour.º Sci. USA 90, 8687±8691. Longman, London/New York. Cameron, S. A., and Mardulyn, P. (2001). Multiple data sets suggest Hobbs, G. A. (1965). Ecology of species of Bombus Latr. in southern independent origins of highly eusocial behavior in bees (Hymenop- Alberta. II. Subgenus Bombias. Can. Entomol. 97, 120±129. tera: Apinae). Syst. Biol. 50, 194±214. Honk, C. G. J., Velthuis, H. H. W., Roseler, P-F., and Malotaux, M. ChavarrõÂa, G., and Carpenter, J. M. (1994). ªTotal evidenceº and the E. (1980). The mandibular glands of Bombus terrestris queens as a evolution of highly social bees. Cladistics 10, 229±258. source of queen pheromones. Entomol. Exp. Appl. 28, 191±198. Coddington, J. A. (1990). Cladistics and spider classi®cation: Araneo- Jeanne, R. L. (1991). The swarm-founding Polistinae. In ªThe Social morph phylogeny and the monophyly of orb-weavers (Araneae: Biology of Wasps.º (K. G. Ross and R. W. Matthews, Eds.), pp. Araneomorphae; Orbiculariae). Acta Zool. Fenn. 190, 75±88. 191±231. Cornell Univ. Press, Ithaca, NY. Cruz-Landim, C. (1963). Evolution of the wax and scent glands of the Kennedy, M., Spencer, H. G, and Gray, R. D. (1996). Hop, step and Apinae. J. N. Y. Entomol. Soc. 71, 2±13. gape: Do the social displays of the Pelecaniformes re¯ect phylogeny? Cruz-Landim, C., and Costa, R. A. C. (1998). Structure and function Anim. Behav. 51, 273±291. of the hypopharyngeal glands of Hymenoptera: A comparative ap- Kimsey, L. S. (1980). The behavior of male orchid bees (Insecta; proach. J. Comp. Biol. 3, 151±163. Hymenoptera: Apidae) and the question of leks. Anim. Behav. 28, de Queiroz, A., and Wimberger, P. H. (1993). The usefulness of behav- 996±1004. ior for phylogeny estimation: Levels of homoplasy in behavioral Kimsey, L. S. (1982). Systematics of bees of the genus Eufriesea. and morphological characters. Evolution 47, 46±60. Univ. Cal. Pub. Entomol. 95, 1±125.

Kimsey, L. S. (1984). A reevaluation of the phylogenetic relationships Pomeroy, N., and Plowright, R.C. (1982). The relation between worker in the Apidae Hymenoptera. Syst. Entomol. 9, 435±442. numbers and the production of males and queens in the bumble bee Kimsey, L. S. (1987). Generic relationships within the Euglossini Hy- Bombus perplexus. Can. J. Zool. 60, 954±957. menoptera Apidae. Syst. Entomol. 12, 63±72. Prentice, M. (1991). Morphological analysis of the tribes of Apidae. Koulianos, S., Schmid-Hempel, R., Roubik, D. W., and Schmid-Hem- In ªDiversity in the Genus Apisº (D. R. Smith, Ed.), pp. 51±69. pel, P. (1999). Phylogenetic relationships within the corbiculate Westview Press, Boulder, CO. Apinae (Hymenoptera) and the evolution of eusociality. J. Evol. Proctor, H. C. (1996). Behavioral characters and homoplasy: Perception Biol. 12, 380±384. versus practice. In ªHomoplasy: The Recurrence of Similarity in Evolutionº (M. J. Sanderson and L. Hufford, Eds.), pp. 131±148. Lacerda, L. M., Zucchi, R., and Sakagami, S. F. (1998). Oviposition Academic Press, San Diego, CA. behavior of the stingless bees. XXV. Ethological relationships of Geotrigona mombuca to other stingless bees taxa (Insecta: Hyme- Roig-Alsina, A., and Michener, C. D. (1993). Studies of the phylogeny noptera; Apidae, Meliponinae). Nat. Hist. Bull. Ibaraky Univ. 2, and classi®cation of long-tongued bees (Hymenoptera: Apoidea). 263±276. Univ. Kansas Sci. Bull. 55, 124±162. Lockhart, P. J., and Cameron, S. A. (2001). Trees for bees. TREE RoÈseler, P-F., RoÈseler, I., and van Honk, C.G. (1981). Evidence for 16, 84±88. inhibition of corpora allata activity in workers of Bombus terrestris by a pheromone from the queen's mandibular glands. Experientia Mardulyn, P., and Cameron, S. A. (1999). The major opsin in bees 37, 348±351. (Insecta: Hymenoptera): A promising nuclear gene for higher level Roubik, D. W. (1989). ªEcology and Natural History of Tropical Bees.º phylogenetics. Mol. Phylogenet. Evol. 12, 168±176. Cambridge Univ. Press., Cambridge, MA. McLennan, D. A. (1993). Phylogenetic relationships in the Gasterostei- Rozen, J. G. (1984). Nesting biology of diphaglossine bees (Hymenop- dae: An updated tree based on behavioral characters with a discussion tera: Colletidae). Am. Mus. Nov. 2786, 1±33. of homoplasy. Copeia 2, 318±326. Sakagami, S. F. (1976). Speci®c differences in the bionomics characters Michener, C. D. (1944). Comparative external morphology, phylogeny, of bumblebees. A comparative review. J. Fac. Sci. Hokkaido Univ. and a classi®cation of the bees. Bull. Am. Mus. Nat. Hist. 82, Ser. VII 20, 390±447. 151±326. Sakagami, S. F. (1982). Stingless bees. In ªSocial Insects IIIº (R. H. Michener, C. D. (1969). Comparative social behavior of bees. Annu. Hermann, Ed.), pp. 361±423. Academic Press, New York. Rev. Entomol. 14, 299±342. Sakagami, S. F., and Zucchi, R. (1963). Oviposition process in a Michener, C. D. (1974). ªThe Social Behavior of Bees: A Comparative stingless bee, Trigona (Scaptotrigona) postica Latreille (Hymenop- study.º Harvard Univ. Press, Cambridge, MA. tera). Studia Entomol. 6, 497±510. Michener, C. D. (1990). Classi®cation of the Apidae (Hymenoptera). Sakagami, S. F., Montenegro, M. J., and Kerr, W. E. (1965). Behavior Univ. Kansas Sci. Bull. 54, 75±164. studies of the stingless bee, with special reference to the oviposition Michener, C. D. (2000). ªThe Bees of the World.º Johns Hopkins Univ. process. V. Melipona quadrifasciata anthidioides Lepeletier. J. Fac. Press, Baltimore, MD. Sci. Hokkaido Univ. Ser. VI Zool. 15, 578±607. Miller, J. S., and Wenzel, J. W. (1995). Ecological characters and Sakagami, S. F., Zucchi, R. and Portugal-ArauÂjo, V.(1977). Oviposition phylogeny. Annu. Rev. Entomol. 40, 389±415. behaviour of an aberrant African stingless bee Meliponula bocandei, with notes on the mechanism and evolution of oviposition behavior in Nixon, K. C. (1999). Winclada (beta) version 0.9. Published by the stingless bees. J. Fac. Sci. Hokkaido Univ. Ser. VI Zool. 20, 647±690. author. Ithaca, NY. [Available at http://www.cladistics.com] Sakagami, S. F., Zucchi, R., Bego, L. R., De Sanctis, M., and Mateus, S. Paterson, A., Wallis, G. P., and Gray, R. D. (1995). Penguins, petrels, (1993). Oviposition behavior of the stingless bees. XV. Nannotrigona and parsimony: Does cladistic analysis of behavior re¯ect seabird testaceicornis and its strictly integrated oviposition process (Apidae, phylogeny? Evolution 49, 974±989. Meliponinae). Jpn. J. Ent. 61, 323±340. Pereira-Martins, S. R., and Kerr, W. E. (1991). Biologia de Eulaema Sanderson, M. J., and Donoghue, M. J. (1989). Patterns of variation nigrita. 1. ConstrucËaÄodeceÂlulas, oviposicËaÄo e desenvolvimento. in levels of homoplasy. Evolution 43, 1781±1795. Pap. Avuls. Zool. 37, 227±235. Santos, M. L., and GaroÂfalo, C. A. (1994). Nesting biology and nest Plant, J. D., and Paulus, H. F. (1987). Comparative morphology of the reuse of Eulaema nigrita (Hymenoptera, Apidae, Euglossini). Insect postmentum of bees (Hymenoptera: Apoidea) with special remarks Soc. 41, 99±110. on the evolution of the lorum. Z. Zool. Syst. Evol. Forsch. 25, Seeley, T. D. (1985). ªHoney Bee Ecology: A Study in Adaptation in 81±103. Social Life.º Princeton Press, Princeton, NJ. Plateaux-QueÂnu, C. (1961). Les sexues de remplacement ches les in- Sheppard, W. S., and McPheron, B. A. (1991). Ribosomal DNA diver- sects sociaux. Ann. Biol. 37, 177±216. sity in the Apidae. In ªDiversity in the Genus Apisº (D. R. Smith, Plowright, R. C. (1977). Nest architecture and the biosystematics of Ed.), pp. 89±102. Westview Press, Boulder, CO. bumble bees. In ªProceedings of the 8th International Congress and Silva-Matos, E. V., and GaroÂfalo, C. A. (1995). Observations on the International Union for the Study of Social Insects, Wageningen,º development of queenless colonies of Bombus atratus (Hymenop- pp. 183±185. tera, Apidae). J. Apicult. Res. 34, 177±185.

Silva-Matos, E. V., Zucchi, R., and Yamane, SoÃ. (1997). Oviposition Wille, A. (1983). Biology of the stingless bees. Annu. Rev. Entomol. behavior of the stingless bees, XXII. Oxytrigona tataira and its 28, 41±64. successive oviposition process in the presence of generalized agita- Williams, P. H. (1994). Phylogenetic relationships among bumble bees tion (Insecta: Hymenoptera: Apidae). Nat. Hist. Bull. Ibaraky Univ. (Bombus Latr.): A reappraisal of morphological evidence. Syst. Ento- 1, 121±134. mol. 19, 327±344. Slikas, B. (1998). Recognizing and testing homology of courtship Wilson, E. O. (1971). ªThe Insect Societies.º Belknap Press and Har- displays in storks (Aves: Ciconiiformes: Ciconiidae). Evolution vard Univ. Press, Cambridge, MA. 52, 884±893. Winston, M. L., and Michener, C. D. (1977). Dual origin of highly social Stephen, W. P., Bohart, G. E., and Torchio, P. F. (1969). ªThe Biology behavior among bees. Proc. Natl. Acad. Sci. USA 74, 1135±1137. and External Morphology of Bees.º Agricultural Experiment Station, Zucchi, R. (1973). ªAspectos bionoÃmicos de Exomalopsis aureopilosa Oregon State University, Corvallis. e Bombus atratus inlcuindo consideracËoÄes sobre a evolucËaÄo do com- portamento social (Hymenoptera, Apoidea).º Doctoral thesis. Stuart, A. E., and Hunter, F. F. (1998). End-products of behaviour FFCLRP±USP, SP, Brazil. versus behavioural characters: A phylogenetic investigation of pupal Zucchi, R. (1993). Ritualized dominance, evolution of queen±worker cocoon construction and form in some North American black ¯ies interactions and related aspects in stingless bees (Hymenoptera: (Diptera: Simuliidae). Syst. Ent. 23, 387±398. Apidae). In ªEvolution of Insect Societies.º (T. Inoue and S. Yamane, Wcislo, W. T. (1997). Are behavioral classi®cations blinders to studying Eds.), pp. 207±249. Hakuhin-sha, Tokyo. natural variation? In ªThe Evolution of Social Behavior in Insects Zucchi, R., and Sakagami, S. F. (1972). Capacidade termo-reguladora and Arachnidsº (J. C. Choe and B. Crespi, Eds.), pp. 8±13. em Trigona spinipes e em algumas outras espeÂcies de abelhas sem Cambridge Univ. Press, New York. ferraÄo. ªHomenagem a Warwick E. Kerr,º pp. 301±309. Rio Webster, T. C., and Peng., Y.S. (1988). The evolution of food-producing Claro, Brazil. glands in eusocial bees (Apoidea, Hymenoptera). J. Evol. Biol. Zucchi, R., Sakagami, S. F., and Camargo, J. M. F. (1969). Biological 2, 165±76. observations on a neotropical parasocial bee, Eulaema nigrita, with Wenzel, J. W. (1992). Behavioral homology and phylogeny. Annu. J. a review of the biology of Euglossinae. A comparative study. J. Rev. Ecol. Syst. 23, 361±381. Fac. Sci. Hokkaido Univ. VI Zool. 17, 271±380. Zucchi, R., Silva-Matos, E. V., Yamane, SoÃ., and Mateus, S. (1997). Wenzel, J. W. (1993). Application of the biogenetic law to behavioral Oviposition behavior of the stingless bees. XXI. Paratrigona lineata ontogenyÐA test using nest architecture in paper wasps. J. Evol. and P. subnuda with reference to their highly integrated oviposition Biol. 6, 229±247. processes (Apidae, Meliponinae). Nat. Hist. Bull. Ibaraki Univ. 1, Wenzel, J. W. (1997). When is a phylogenetic test good enough? Mem. 107±120. Mus. Natl. Hist. Nat. 173, 31±45. Zucchi, R, Silva-Matos, E. V., Nogueira-Ferreira, F. H., and Azevedo, Wenzel, J. W., and Carpenter, J. M. (1994). Comparing methods: Adap- G. G. (1999). On the cell provisioning and oviposition process (POP) tive traits and tests of adaptation. In ªPhylogenetics and Ecology.º of the stingless bees, nomenclature reappraisal and evolutionary (P. Eggleton and R. Vane-Wright, Eds.), pp. 79±101. Academic considerations (Hymenoptera, Apidae, Meliponinae). Sociobiology Press, London. 34, 65±86.