A novel terpene synthase produces an anti-aphrodisiac pheromone in the butterfly Heliconius melpomene

Terpenes, a group of structurally diverse compounds, are the biggest class of secondary metabolites. While the biosynthesis of terpenes by enzymes known as terpene synthases (TPSs) has been described in plants and microorganisms, few TPSs have been identified in insects, despite the presence of terpenes in multiple insect species. Indeed, in many insect species, it remains unclear whether terpenes are sequestered from plants or biosynthesised de novo. No homologs of plant TPSs have been found in insect genomes, though insect TPSs with an independent evolutionary origin have been found in Hemiptera and Coleoptera. In the butterfly Heliconius melpomene, the monoterpene (E)-β-ocimene acts as an anti-aphrodisiac pheromone, where it is transferred during mating from males to females to avoid re-mating by deterring males. To date only one insect monoterpene synthase has been described, in Ips pini (Coleoptera), and is a multifunctional TPS and isoprenyl diphosphate synthase (IDS). Here, we combine linkage mapping and expression studies to identify candidate genes involved in the biosynthesis of (E)-β-ocimene. We confirm that H. melpomene has two enzymes that exhibit TPS activity, and one of these, HMEL037106g1 is able to synthesise (E)-β-ocimene in vitro. Unlike the enzyme in Ips pini, these enzymes only exhibit residual IDS activity, suggesting they are more specialised TPSs, akin to those found in plants. Phylogenetic analysis shows that these enzymes are unrelated to previously described plant and insect TPSs. The distinct evolutionary origin of TPSs in Lepidoptera suggests that they have evolved multiple times in insects.Significance statementTerpenes are a diverse class of natural compounds, used by both plants and animals for a variety of functions, including chemical communication. In insects it is often unclear whether they are synthesised de novo or sequestered from plants. Some plants and insects have converged to use the same compounds. For instance, (E)-β-ocimene is a common component of floral scent and is also used by the butterfly Heliconius melpomene as an anti-aphrodisiac pheromone. We describe two novel terpene synthases, one of which synthesises (E)-β-ocimene in H. melpomene, unrelated not only to plant enzymes but also other recently identified insect terpene synthases. This provides the first evidence that the ability to synthesise terpenes has arisen multiple times independently within the insects.

1-Octen-3-ol is repellent to Ips pini (Coleoptera: Curculionidae: Scolytinae) in the midwestern United States

In field experiments at three sites in Michigan and Ohio we tested the activity of 1-octen-3-ol in combination with ipsdienol, the aggregation pheromone of the pine engraver, Ips pini (Say). When 1-octen-3-ol was added to funnel traps baited with ipsdienol, significantly fewer beetles of either sex were captured than in traps baited with ipsdienol alone. This result suggests that the compound is potentially repellent and interrupts the response of beetles to their aggregation pheromone, and is consistent with previous reports of its inhibition of aggregation behaviour in other bark beetles.

Assemblage of Hymenoptera arriving at logs colonized by Ips pini (Coleoptera: Curculionidae: Scolytinae) and its microbial symbionts in western Montana

Colonization of a tree by bark beetles and their symbionts creates a new habitat for a diverse assemblage of arthropods, including competing herbivores, xylophages, fungivores, saprophages, predators, and parasitoids. Understanding these assemblages is important for evaluating nontarget effects of various management tactics and for subsequently evaluating how changes in climate, the presence of invasive species, and altered forestry practices and land-use tenure may affect biodiversity. We characterized the assemblage of hymenopterans attracted to logs of ponderosa pine (Pinus ponderosa C. Lawson (Pinaceae)) colonized by the bark beetle Ips pini (Say) and its microbial symbionts. In one experiment, the composition and relative abundances of species arriving at hosts colonized by I. pini, and possible sources of attraction, were determined. Treatments consisted of a log containing I. pini with its natural complement of microorganisms, a log alone, and a blank control. A second experiment was carried out to determine whether or not Hymenoptera were attracted to microbial symbionts of I. pini. Treatments consisted of a blank control, a log alone, a log containing I. pini with its natural complement of microorganisms, either Ophiostoma ips, Burkholderia sp., or Pichia scolyti, and a log inoculated with a combination of these three microorganisms. Over 2 years, 5163 Hymenoptera were captured, of which over 98% were parasitoids. Braconidae, Platygastridae, Encyrtidae, Pteromalidae, and Ichneumonidae were the most abundant. Seven known species of bark beetle parasitoids (all Pteromalidae) were captured. However, parasitoids of Diptera, Lepidoptera, Hymenoptera, and non-wood-boring Coleoptera were also common. Nineteen species showed preferential attraction to host plants infested with I. pini and its complement of microorganisms, host plants inoculated with I. pini microbial symbionts, or host plants alone. Interestingly, many of these species were parasitoids of phytophagous, fungivorous, and saprophytic insects rather than of bark beetles themselves. These results suggest that a diverse assemblage of natural enemies that attack various feeding guilds within a common habitat exploit common olfactory cues.