Cuticular hydrocarbons of Drosophila montana: Geographic variation, sexual dimorphism and potential roles as pheromones Jackson H. Jennings a,d,⇑ , William J. Etges a , Thomas Schmitt b,c , Anneli Hoikkala d a Program in Ecology and Evolutionary Biology, Department of Biological Sciences, University of Arkansas, AR 72701, USA b Department of Animal Ecology and Tropical Biology, Faculty of Biology, University of Würzburg, Würzburg 97074, Germany c Ecological Networks, Faculty of Biology, Technical University of Darmstadt, Germany d Department of Biological and Environmental Science, University of Jyväskylä, Jyväskylä 40014, Finland article info Article history: Received 8 July 2013 Received in revised form 21 November 2013 Accepted 3 December 2013 Available online 25 December 2013 Keywords: CHCs Pheromones Sexual selection Mate choice Speciation abstract Sexual selection within populations can play an important role in speciation when divergence in mating signals and their corresponding preferences occur along different coevolutionary trajectories in different populations. In insects, one potential target of sexual selection is the blend of cuticular hydrocarbons (CHCs), which often show intra- and interspecific variation, sexual dimorphism and may act as phero- mones. In Drosophila montana, a cold-adapted, circumboreal member of the Drosophila virilis species group, flies from different populations have been found to show significant premating isolation as well as variation in male mating signal (song) and female preference. While the role of male courtship song in mate choice has been studied extensively, CHCs in this species have received little attention. In this study, we identified most of the CHCs found on the cuticle of D. montana and characterized population divergence and sexual dimorphism of CHC profiles among flies established from three natural popula- tions – one European and two North American. We also studied their potential role as pheromones by analyzing CHCs of flies used in female-choice mating experiments. We report significant population  sex effects on CHC profiles, as well as significant relationships between some CHC principal components and particular mating behaviours, such as female attractiveness and male mating success, providing evidence that CHCs may play a role in mate choice in this species. The study also provides evidence for variation in the degree to which CHCs play a role in chemical communication among these populations, which may have an influence on the speciation process itself, and could be due to variation in interactions with other closely-related species that occur sympatrically with D. montana in some, but not other, parts of its distribution. Ó 2013 Elsevier Ltd. All rights reserved. 1. Introduction Sexual selection is based on genetic variation in traits that are important in mate choice, giving some phenotypes an advantage over others when one sex exercises choice among possible mates (Lande, 1981; Andersson, 1994). In Drosophila, mating success may be determined by phenotypes like body size and courtship vigour, and/or by male mating signals such as courtship song or sex pheromones, which coevolve with the corresponding female preferences for these traits (Ewing and Bennet-Clark, 1968; Par- tridge et al., 1987a,b; Markow and O’Grady, 2005; Ferveur, 2005). If these signals and preferences diverge along different evolution- ary trajectories in different conspecific populations, e.g. during extended periods of allopatry, they may contribute to premating isolation upon secondary contact, providing the initial grounds for speciation (Gavrilets, 2000; Kirkpatrick and Ravigné, 2002; Coy- ne and Orr, 2004; van Doorn et al., 2009; Kraaijeveld et al., 2011). Insects have evolved complex cuticular chemistry that allows them to cope with life in terrestrial habitats. The chemicals found on their body surface are generally hydrophobic hydrocarbons of varying chain lengths (cuticular hydrocarbons, or CHCs) and may play important roles in waterproofing, desiccation or disease resis- tance, and/or mate choice (Edney, 1977; Jallon, 1984; Gibbs, 1998; Wagner et al., 2001; Howard and Blomquist, 2005; Ferveur, 2005; Ferveur and Cobb, 2010). CHCs may also be under sexual selection if particular components confer a mating advantage or increase the fitness of resulting offspring. The evolution of CHC variation and chemistry may also be constrained due to phylogenetic affinity (Oliveira et al., 2011). The evolution of olfactory communication in generating reproductive isolation among populations or species, however, is still not fully understood (Smadja and Butlin, 2009). 0022-1910/$ - see front matter Ó 2013 Elsevier Ltd. All rights reserved. http://dx.doi.org/10.1016/j.jinsphys.2013.12.004 ⇑ Corresponding author at: Department of Biological Sciences, SCEN 601, 1 University of Arkansas, Fayetteville, AR 72701, USA. Tel.: +1 479 531 5781; fax: +1 479 575 4010. E-mail address: jjenning@uark.edu (J.H. Jennings). Journal of Insect Physiology 61 (2014) 16–24 Contents lists available at ScienceDirect Journal of Insect Physiology journal homepage: www.elsevier.com/locate/jinsphys