Cuticular hydrocarbons (HCs) play important roles in insect communication but few studies clearly demonstrate the direct link between HCs and nestmate recognition. Therefore, cuticular lipids were extracted from ants, their HC and non-HC fractions as well as the three principal classes of HCs (n-alkanes, branched alkanes and alkenes) were purified and tested using an immobilizing "joust" device which allowed quantification of early pairwise behavioural responses, mandibular opening and antennal retraction, without occurrence of subsequent damages as in classic dyadic encounters. Chemical recognition of ants was studied at three levels of interactions (intra-colonial, intra-specific and inter-specific). Three closely related species already chemically characterized were used: Pachycondyla villosa (Pv), P. inversa (Pi) and P. subversa (Ps). Each species had its own behavioural responses. Moreover, responses of Pi and Ps towards Pv were significantly longer, than they were between themselves whereas Pv ants were equally aggressive towards Pi and Ps. These differences are in agreement with the results of the cluster analysis of the cuticular HCs profiles that place Pi closer to Ps. In support of the idea that components of cuticular lipids profiles are important for recognition, we found that only the HC fraction and its branched subfraction elicited a behavioural response of Ps workers. It is suggested that internally branched methyl- and dimethylalkanes are involved in recognition behaviour.

Nestmate recognition in ants is based on perceived differences in a multi-component blend of hydrocarbons that are present on the insect cuticle. Although supplementation experiments have shown that some classes of hydrocarbons, such as methyl branched alkanes and alkenes, have a salient role in nestmate recognition, there was basically no information available on how ants detect and perceive these molecules. We used a new conditioning procedure to investigate whether individual carpenter ants could associate a given hydrocarbon (linear or methyl-branched alkane) to sugar reward. We then studied perceptual similarity between a hydrocarbon previously associated with sugar and a novel hydrocarbon. Ants learnt all hydrocarbon-reward associations rapidly and with the same efficiency, regardless of the structure of the molecules. Ants could discriminate among a large number of pairs of hydrocarbons, but also generalised. Generalisation depended both on the structure of the molecule and the animal's experience. For linear alkanes, generalisation was observed when the novel molecule was smaller than the conditioned one. Generalisation between pairs of methyl-alkanes was high, while generalisation between hydrocarbons that differed in the presence or absence of a methyl group was low, suggesting that chain length and functional group might be coded independently by the ant olfactory system. Understanding variations in perception of recognition cues in ants is necessary for the general understanding of the mechanisms involved in social recognition processes based on chemical cues.

Two sibling species, Solenopsis richteri and S. invicta, were both introduced into the southern USA from South America in the early 20th century. Today, S. richteri occupies higher latitudes and colder areas, while S. invicta occupies lower latitudes. Between the distributions of the two species, there is a large area of viable hybrid (S. richteri × S. invicta) populations. This study aimed to characterize the forces driving this distribution pattern and the underlying mechanisms. Cuticular hydrocarbons (CHCs) of freshly killed workers of S. invicta, hybrids, and S. richteri were removed using hexane. Both intact and CHCs-extracted workers were subjected to a constant rate of increasing temperature from 10 to 60 °C to obtain relative water loss and the water loss transition temperature (Tc-ant). Mass loss and Tc-ant were both significantly increased with CHCs removal. We then examined the CHC composition of three species. CHC profiles of S. richteri are characterized by significant amounts of short-chain (C23-C27) saturated and unsaturated hydrocarbons. In contrast, profiles of S. invicta consist primarily of long-chain (C27-C29) saturated hydrocarbons; unsaturated alkenes are completely lacking. Hybrid fire ants show intermediate profiles of the two parent species. We measured the melting point (Tm) and water-loss transition temperature of CHC blends (Tc-CHC) of different ant species colonies using differential scanning calorimetry (DSC) and an artificial membrane system, respectively. There were 3-5 Tms of each CHCs sample of different ant colonies due to their complex chemistry. The highest Tms (Tm-maxs) of CHCs samples from S. invicta and the hybrid were significantly higher than that from S. richteri. The correlation between Tc-CHC and Tm-max obtained from the same CHCs sample was highly significant. These results reveal that species having higher Tc and Tm-max retain more water under relatively higher temperature, and consequently are able to occupy warmer environments. We conclude that CHC chemistry plays a role in shaping current distribution patterns of S. richteri, S. invicta and their hybrid in the United States.