Changes in the Chemical Profile of Cephalic Salivary Glands of Scaptotrigona Postica (Hymenoptera, Meliponini) Workers Are Phase Related Silvana B
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© 2015. Published by The Company of Biologists Ltd | The Journal of Experimental Biology (2015) 218, 2738-2744 doi:10.1242/jeb.124180 RESEARCH ARTICLE Changes in the chemical profile of cephalic salivary glands of Scaptotrigona postica (Hymenoptera, Meliponini) workers are phase related Silvana B. Poiani1,*, E. David Morgan2, Falko P. Drijfhout2 and Carminda da Cruz-Landim1 ABSTRACT adult bees, salivary glands are paired structures located in the Most advanced eusocial bees recruit their nest mates to food thorax, but some species from the Apinae subfamily have developed resources. Recent studies in Meliponini species have revealed that a branch of salivary glands in the head called cephalic salivary the cephalic salivary (labial) glands (CSGs) are responsible for the glands (CSGs) (Cruz-Landim, 1967). production of scent trail pheromones. Studies on CSGs have shown All salivary glands, including the CSGs, have several functions in that changes occur in worker glandular cell morphology from eusocial bees that go beyond food ingestion (Poiani and da Cruz- emergence from brood combs until forager phase, which may be Landim, 2010a). Heselhaus (1922) was one of the first researchers correlated to changes in the composition of the CSG secretion. to suggest the function of the CSG secretion of Apis mellifera; However, the composition of the CSG secretion and the chemical specifically, softening wax during nest building. Later, Simpson changes that occur in it according to the worker’s life phase or tasks (1960) described lubrication of the mouthparts as the function of the performed are unknown for many species, including Scaptotrigona CSGs. In stingless bees, Santos et al. (2009) suggested that the CSG postica. In this study, the chemical profile of CSG secretion in S. secretions of Plebeia emerina nurses aid in the handling in the postica workers was studied. Glands were taken from specimens that colony of resins collected in the field, and other recent studies have were newly emerged (NE), working in the brood comb area (CA) and shown that Trigona recursa (Jarau et al., 2004, 2006), Trigona foraging (FO), and were analyzed by gas chromatography–mass spinipes (Schorkopf et al., 2007), Geotrigona mombuca (Stangler spectrometry. The results showed that the glandular secretion et al., 2009) and Trigona corvine (Jarau et al., 2010a) use the consists of oxygenated compounds of middle volatility (acids, secretion of the CSGs as a scent trail to indicate a food source to nest alcohols, aldehydes, ketones, esters and ether), and their quantity mates, by producing ester compounds. In another stingless bee varies among the different life phases, increasing as the individual species, Melipona beecheii, the CSGs produce geraniol, which has moves from intra- to extra-colonial activities. The NE phase contained been found to affect caste determination by significantly increasing the smallest variety and quantity of compounds. Because of the the number of larvae that develop into queens (Jarau et al., 2010b). variability of compounds, the CA workers were separated into three The functions of the CSGs are known to vary among species, sex, subgroups according to the chemical constitution of their secretion. caste, physiological state and roles of individuals in the colony. In Forager workers showed the largest quantity and variety of chemical many Bombus species, variation between sexes and castes occurs compounds. The major compounds in forager gland secretion were 7- not only in the development of the CSGs but also in the identity and hexadecen-1-yl acetate and 5-tetradecen-1-yl acetate. Statistical function of the secreted compounds. In general, the CSGs of sterile analysis indicates that the chemical composition of glandular females of Bombus terrestris produce larger amounts of wax esters secretion is phase related. (mainly dodecyl esters) than the CSGs of fertile females of this species (Amsalem et al., 2014). Males of Bombus species use KEY WORDS: GC-MS, Ester, Labial glands, Life phases, Multivariate secretions of the CSGs for territorial marking and female attraction statistics, Stingless bee (Amsalem et al., 2014; Bergman and Bergström, 1997; Bertsch et al., 2005; Kullenberg et al., 1973; Terzo et al., 2007). In INTRODUCTION A. mellifera, the hydrocarbons in the glandular secretions are also Stingless bees are a large group of eusocial bees with approximately present in the cuticle of forager workers (Arnold and Delage- 500 species, comprising the tribe Meliponini (Michener, 2007). As Darchen, 1978; Arnold et al., 1996; Poiani, 2012), suggesting that for all eusocial bees, members of a colony are divided into castes. the CSGs might be the source of cuticular hydrocarbons in this The caste of workers is responsible for brood care, building, species. Histochemical and ultrastructural studies of CSGs indicate attending the queen, guarding the entrance and activities outside the that a lipid-like secretion is produced in stingless bees (Poiani and nest. Cruz-Landim, 2009, 2010c), and Elias-Santos et al. (2013) in a The salivary system of bees comprises the salivary (also known as proteome analysis of CSG of Melipona quadrifasciata anthidioides the labial), mandibular and hypopharingeal glands, named after the found enzymes relevant to lipid metabolism. mouth appendices to which they are connected. In all species of In Scaptotrigona postica and A. mellifera workers, the CSGs were shown to begin to produce secretion soon after worker 1Departamento de Biologia, Universidade Estadual Paulista – UNESP, Instituto de emergence and accumulate the secretion continually in the alveolar Biociências, Rio Claro, SP 13506-900, Brazil. 2Chemical Ecology Group, Lennard- lumen throughout their life, and thus subsequent changes in both the Jones Laboratories, Keele University, Staffordshire ST5 5BG, UK. intensity of secretory activity and the chemical composition of the *Author for correspondence ([email protected]) secretion would be expected (Poiani and Cruz-Landim, 2009, 2010a,b,c). Francke et al. (1983) studied the CSGs of S. postica Received 23 April 2015; Accepted 23 June 2015 workers, but they used whole-head extracts, including other glands The Journal of Experimental Biology 2738 RESEARCH ARTICLE The Journal of Experimental Biology (2015) 218, 2738-2744 doi:10.1242/jeb.124180 and cuticle compounds, so the identified compounds are not passes through a phase in which it remains at the nest entrance exclusive to the CSGs. Therefore, little is known about the chemical functioning as guard. The labour division is plastic. The age at properties of the CSGs of S. postica. Addressing this gap in which the duties are performed and time expended in each task vary evidence might allow understanding of the dynamics of secretion with the species and colony state. synthesis of these glands and how it may affect social behaviour as In view of the above, the present study aimed to identify and the tasks performed are related to the bees’ physiological state. quantify the volatile compounds that are present in the CSG secretions The tasks of workers in eusocial bee colonies are performed of S. postica during different life phases using gas chromatography– according to a division of labour based on age polyethism. The mass spectrometry (GC-MS), and subsequently identify possible ability to perform a given task is achieved through physiological correlations between the changes in secretion composition and the maturity, acquired in steps with the advancement of age (Ribbands, status of the worker in the colony by statistical analysis. 1953; Oster and Wilson, 1978). The pertinent literature reporting several aspects of labour division in S. postica (Akahira et al., 1970; RESULTS Dias and Simões, 1972; Bego, 1982) shows that the sequence of The CSG secretion of S. postica workers varied qualitatively and worker activities is similar to that of other eusocial bees: newly quantitatively not only across the analysed phases (NE, CA and FO) emerged (NE) workers clean themselves of brood debris and spend but also within them. In general, the identified compounds included: much of the time inactive or grooming. In the subsequent age step, acids, alcohols, aldehydes, ketones, esters and ether (Table 1). in which they work in the nest brood area (CA, generally named the The CSG of the NE phase produce little secretion. Among the nurse phase), they display a variety of actions directed mainly, but nine identified compounds in this life phase, 2-tridecanone, ethyl not solely, at aiding the queen in laying her eggs and brood care. The oleate and nonanal were the most abundant. phase of work inside the nest ends when workers became foragers The CA phase showed large qualitative and quantitative (FO), at which point they go out to obtain resources for the colony. variations in glandular secretion composition. As a result, the Between the duties inside the nest and those outside, the worker phase was separated into three groups, CA1, CA2 and CA3, Table 1. Quantities of chemical compounds found in the cephalic salivary gland (CSG) secretions of Scaptotrigona postica workers ID KI Compound NE (N=8) CA1 (N=3) CA2 (N=5) CA3 (N=14) FO (N=8) Acid A1 894 Nonanoic acid ––0.70±0.95 – 0.20±0.53 A2 1069 Decanoic acid ––5.30±3.58 – 1.00±2.82 A3 1383 Dodecanoic acid ––0.46±1.01 –– A4 1686 Tetradecanoic acid ––8.42±5.22 – 0.32±0.85 A5 1820 Pentadecanoic acid –––1.80±1.76 0.15±0.41 A6 1970 Hexadecanoic acid 5.79±1.36 – 30.88±12.09 – 0.48±1.34 Alcohol Al7 1307 2-Tridecanol – 7.96±4.35 0.45±0.98