RESUMOXVIII Simpósio EXPANDIDO de Mirmecologia 031 379A

FEEDING BEHAVIOR OF SEXDENS (: FORMICIDAE) QUEENS DURING ESTABLISHMENT OF NEW COLONIES IN LABORATORY

Comportamento alimentar de rainhas de (Hymenoptera: Formicidae) durante o estabelecimento de novas colônias em condições de laboratório

J.O. Augustin* & J.F.S. Lopes

Universidade Federal de Juiz de Fora, Instituto de Ciências Biológicas, Campus Universitário Martelos, CEP 36036-330, Juiz de Fora, MG, . E-mail: [email protected]

In , colony founding is the most vulnerable the in the mother colony, but also the stage in their life cycle (Hymenoptera, Formicidae) establishment, growth and care of the initial garden (HÖLLDOBLER & WILSON, 1990). In response, a number (HÖLLDOBLER & WILSON, 1990; FERNÁNDEZ-MARÍN et al., of strategies that allowed colonial success were 2003, 2004). selected (PEETERS & ITO, 2001; BROWN & BONHOEFFER, Considering that the establishment of a new colony 2003). Ants were favored with two main reproductive is a key feature to understand the evolutionary strategies. The dependent strategy, in which a adaptations by which many social went founding queen depends on their nestmates to start a through (HÖLLDOBLER & WILSON, 1990; PEETERS & ITO, new colony, and the independent strategy, in which 2001; BROWN & BONHOEFFER, 2003; FERNÁNDEZ-MARÍN et the founding queen does not require the help of al., 2003, 2004; JOHNSON, 2006), this study focuses on nestmates (HÖLLDOBLER & WILSON, 1990). Independent the feeding behavior of Atta sexdens queens during founding strategies can be of two types. First, in the both colony founding and early ergonomic stages in semiclaustral founding, queens leave the nest to forage laboratory. for food for themselves and their brood. Second, in the Just after their in early november, 211 fully claustral founding, queens seal themselves in reproductive of A. sexdens were collected in the nest after mating and never leave the nest to forage. pasture fields within the City of Juiz de Fora (21°46‘S– It is generally assumed that fully claustral founding 43°21‘W, 678m average altitude), southeastern Brazil. queens rely solely on their body reserves to rear the Queens were collected with the aid of tweezers and first generation of workers (KELLER & PASSERA, 1989; individually placed in transparent plastic containers WHEELER, 1994; LIU et al., 2001; HAHN et al., 2004). The (12 cm of diameter and 10,5 cm of height) with a 1 cm definition of claustral colony founding, however, is layer of dampened plaster at their base to keep humidity more precise than that. It has been proposed by inside the chambers. With the aid of needle tip, small HÖLLDOBLER & WILSON (1990) and is given as “the holes on the chamber covers were made in order to procedure during which queens (or royal pairs in the allow the entrance of oxygen in the colonies. Labelled case of termites) seal themselves off in cells and rear containers permitted the identification of the colonies the first generations of workers on nutrients obtained to be observed every week. Laboratory conditions mostly or entirely from their own storage tissues, were kept in 25 ± 5° C, 70 to 80% of relative humidity including fat bodies and histolysed wing muscles”. and photoperiod of 12 hours. The claustral founding in ants is performed by the The Focal Sampling method (ALTMANN, sub-families Dolichoderinae, Formicinae and 1974) was conducted in 30 colonies, starting from the (HÖLLDOBLER & WILSON, 1990; PEETERS & following day since the collection of the queens. It took ITO, 2001; BROWN & BONHOEFFER, 2003), which are the colonies 11 weeks before the first workers emerged, considered morphologically more complex ants. so that observations were extended for more 11 weeks Particularly more elaborated is the founding behavior after workers emerged in each colony. Six colonies of the fungus-growing ants (Formicidae: Attini) were observed per day, during 30 uninterrupted (HÖLLDOBLER & W ILSON, 1990; WIRTH et al., 2003). Great minutes each. After 22 weeks, colonies had been part of this merit is due to the fact that Attini founding observed for 11 hours each, during both the founding queens must alone accomplish tasks such as both the and the ergonomic stages. care of the offspring and the fungus garden. The later Queens that eventually perished were replaced by includes not only the collection of an initial portion of other ones from the same sample.

*Programa de Pós-Graduação em Comportamento e Biologia Animal.

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Behaviors were recorded as single occurrences using unidirectional movements of the glossa (from regardless of their duration. Data were found to be not within to the outside of the mouth) they ingested the normally distributed according to the Kolmogorov- corporal fluids that come out of the body of the . Smirnov modified by Lilliefors test. In order to evaluate In the ergonomic stage, the queens were observed to differences in the absolute frequencies of the receive from a worker’s jaws a portion of the body of behavioural acts both in different phases within the a larva, that they ingested by movements of both the founding stage as well as in between the founding glossa (from within to the outside of the mouth) and and ergonomic stages, all data were analyzed by jaws (in open-and-close movements). means of c2 test. Only once were the queens observed to perform Queens were found to perform five behavioral acts pupophagy (Table 1). Similarly to larvophagy, queen related to the feeding behavior: were observed to receive from a worker’s jaws a portion 1. Ingesting trophic of the pupal abdomen, which the queens ingested by 2. Ingesting fungi staphylae movements of both the glossa (from within to the 3. Ingesting reproductive egg outside of the mouth) and jaws (in open-and-close 4. Larvophagy movements). 5. Pupophagy Although it is generally believed that fully The ingestion of trophic was the most claustral queens rear their first clutch of workers frequently recorded behavior (Table 1). Queens were solely on internal reserves (KELLER & PASSERA, 1989; observed to bend their bodies in a dorso-ventral WHEELER, 1994; LIU et al., 2001; H AHN et al., 2004), our direction before a newly- was laid. While work shows that this may not always be the case, the egg was still on the anal pore, the queens ingested given the fact that A. sexdens queens were found to it with unidirectional movements of the glossa. Queens ingest both trophic and reproductive eggs as well as also ingested trophic eggs that happened to be on the fungi staphylae. Here we show that the main food mouth of the larvae. sources for the queens were both the trophic eggs and The second most frequently recorded behavior the fungi staphylae. Ingesting extra nutritious was ingesting staphylae (Table 1). The queens were sources would strongly mean that A. sexdens queens observed to grab a portion of the simbiont fungus required more energy for founding new colonies in using their jaws and with unidirectional movements laboratory than their internal reserves could offer. of the glossa (from within to the outside of the mouth) This could largely be because founding attine queens they ingested the fungus. Despite being the most face a challenge that other fully claustral queens do frequently recorded behaviors, no significant not: they must accomplish fungiculture, otherwise differences were found between the ingesting of their colonies will not last for long. Indeed, because trophic eggs and ingesting staphylae in any of the of the obligatory -fungus mutualism (CURRIE, 2001), colonial development stages (c2 = 3,42; p = 0,0644). Atta colonies have been found not to last for more Comparing both colonial development stages, than 78 days without the fungus garden (AUTUORI, however, ingesting fungi staphylae (c2 = 60,552; p < 1941, WEBER, 1972; FERNÁNDEZ-MARÍN et al., 2005, 0,001) and ingesting trophic egg (c2 = 54,256; p < AUGUSTIN, 2007). Although claustral queens that 0,001) were more frequent in the founding stage than exhibit the independent mode of colony founding in the ergonomic stage. are capable of accumulating energy reserves to Only twice were the queens observed to ingest support mating, dispersal, and either digging nests reproductive eggs (Table 1). Such behavior was only (KELLER & PASSERA, 1989; JOHNSON, 1996; HAHN et al., observed in the founding stage. Queens were observed 2004) the oophagy behavior is not uncommon among to grab a reproductive egg between her jaws, licking Atta queens (AUTUORI, 1940, 1942; HÖLLDOBLER & it repeated times until it was ingested, by means of WILSON, 1990; AUGUSTIN, 2007). In fact, our study unidirectional movements of the glossa from within shows the ingesting of trophic eggs as the most to the outside of the mouth. The frequency of ingesting frequently recorded behavior, which did not trophic eggs was significantly higher than the significantly differ from the ingesting of staphylae. ingesting of reproductive eggs both in the founding This could mean that both behaviors not only share stage (c2 = 118,127; p <0,001) and the ergonomic stage equivalent costs but also equally benefits for colony (c2 = 32; p <0,001). maintenance. Considering that the claustral Larvophagy was once observed in the founding founding is energetically demanding for newly mated stage and once observed in the ergonomic stage (Table queens that found new colonies independently, this 1). This behavior was observed in two different colonies result strongly indicates that A. sexdens queens do and was performed in two different ways. In the not rely solely on nutrients obtained entirely from founding stage, the queens were observed to perforate their own storage tissues when founding new with their jaws the cuticle of the body a larva and, colonies.

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Table 1 - Absolute frequency of each behavioral category registered for Atta sexdens queens during both the founding and ergonomic stages, with the result of the c 2 test. Value in parentheses refers to the proportion of the behavioral acts in each stage. Behavioral act Colonial development stages Total c2 p Founding Ergonomic Ingesting trophic egg 124 32 156 54.256 <0.001 (79.50%) (20.50%) (54.55%) Ingesting fungi staphylae 106 19 125 60.552 <0.001 (84.80%) (15.20%) (43.71%) Ingesting reproductive egg 2 0 2 2.00 0.1573 (100%) (0%) (0.70%) Larvophagy 1 1 2 - - (50%) (50%) (0.70%) Pupophagy 0 1 1 1.00 0.3173 (0%) (100%) (0.35%) Total 234 52 286 115.818 <0.001 (81.82%) (18.18%) (100%)

Ingesting comparatively more trophic eggs than point of view of the queen, there is obviously a high reproductive eggs was already expected for this cost in performing such behavior. Nevertheless, if because trophic egg-laying in eusocial insects has a larvophagy is performed, it is probably because the unique purpose: to serve as alternative food source for benefit with its expression overcomes the costs of mothers and offspring (PERRY & ROITBERG, 2006). performing it (ALCOCK , 2 001). This hypothesis predicts Trophic eggs have been formally defined by CRESPI that, when facing insufficient stored body reserves, (1992) as non-developing eggs or egg-like structures queens would use larvae as alternative food source. produced for offspring consumption. According to One way to test such hypothesis would be to compare this definition, trophic eggs are not simply unavoidable the behavior of newly-mated A. sexdens queens that non-developing eggs that happen to be eaten by eventually weighted less than the average 0,7g offspring but rather an evolved maternal phenotype, previously recorded by AUGUSTIN (2007) for this species, which, in the case of Atta queens, is clearly an with the behavior of A. sexdens queens that eventually adaptation (considering Alcock’s (2001) definition of weighed more than 0,7g. In this study, the occurrence adaptation) because (1) trophic eggs in this are of larvophagy coincidentally with the colony founding morphologically distinct from reproductive eggs stage may be a supporting evidence to this hypothesis. (AUTUORI, 1940; B AZIRE-BENAZET, 1957; AUGUSTIN, 2007); Secondly, larvophagy could also be related to the (2) Atta queens’ behavior appears to be fine-tuned for possible contamination of the colonies by parasitoid facilitating egg-consumption by offspring (AUTUORI, Neodohrniphora flies (Diptera: ), although no 1942; MARICONI, 1970; AUGUSTIN, 2007) and (3) as this flies were observed in the colonies. Such phorid flies study shows, trophic eggs are, together with the fungi commonly parasitize colonies of Atta sp., as it has staphylae, the most important energy source for newly- previously been verified by other authors (WEBER, mated A. sexdens queens during colony establishment. 1972; HÖLLDOBLER & WILSON, 1990; BRAGANÇA et al., Further work is needed to elucidate the dynamics of 2002). The adult fly lays its eggs inside of the body of trophic egg-laying and consumption by A. sexdens the worker ant usually through the introduction of its queens and their offspring during colony ovipositor either in the posterior part of the host’s establishment. Such work is in progress (AUGUSTIN head or in the final portion of the ant’s gaster, according and SANTOS, in prep.) to observations of MARICONI (1970), WEBER (1972) AND Larvophagy, on the other hand, is not common BRAGANÇA et al. (2002). Recent studies reveal, however, under normal situations and the literature does not that incipient A. sexdens colonies raised in laboratory register its occurrence among the Attini. Two can also be parasitized by Phoridae flies in the hypotheses could be formulated to explain the founding stage (AUGUSTIN, 2007). Ingesting a occurrence of such behavior. Firstly, the queens could parasitized larva would likely mean an urgent attempt be facing low levels of energy reserves and, in an of the founding queen to “make the best of a bad urgent attempt of recomposing their nutritional needs, situation” (KREBS & DAVIS, 1996; ALCOCK, 2001) by they would sacrifice their own offspring. From the adopting the behavior with a lower reproductive

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payoff. A founding queen would not normally feed on Arquivos do Instituto Biológico, São Paulo, v.12, p.197- her own offspring, but when stimulated by any 228, 1941. environmental modification (, for example) AUTUORI, M Contribuição para o conhecimento da saúva she may exhibit larvophagy as an alternative strategy. (Atta spp. - Hymenoptera: Formicidae). II. O sauveiro This hypothesis predicts that A. sexdens queens that inicial (Atta sexdens rubropilosa Forel, 1908). Arqui- consume parasitized larvae (1) would not only ingest vos do Instituto Biológico, São Paulo, v.13, p.67-86, an extra source of nutrients but also contribute to the 1942. protection of the colony, once the eggs of the parasite BAZIRE-BENAZET, M. Sur la formation de l’oeuf alimentaire would not evolve, and (2) are capable of discriminating chez Atta sexdens rubropilosa Forel, 1908 (Hymenoptera: between parasitized and non-parasitized larvae. Such Formicidae). Comptes Rendus de l´Académie des Science, hypothesis remains to be tested. v.244, p.1277-1280, 1957. BRAGANÇA, M.A.L.; TONHASCA, J.R.A.; MOREIRA, D.O.Parasitism Literature shows no reports on pupophagy among caracteristics of two phorid fly species in relation to the Attini. This behavioral act has been observed in their host, the -cutting ant (Smith) natural colonies of Polistes simillimus Zikán, 1951 (Hymenoptera: Formicidae). Neotropical Entomology, (Hymenoptera: Vespidae) for a single time in 37 hours v.31, n.2, p.305-311, 2002. of observation (GRAZINOLI, 2006), similarly to the arti- BROWN, M.J.F. & BONHOEFFER, S. On the origin of claustral ficial colonies of A. sexdens in this study. Pupophagy colony founding in ants. Evolutionary Ecology Research, seems to be an odd behavior that probably happens v.5, p.305-313, 2003. under very particular circumstances, to mention, low CRESPI, B.J. Cannibalism and trophic eggs in subsocial and levels of energy reserves. 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