Monoterpenes and epicuticular waxes help female Autumn Gum Moth differentiate between waxy and glossy Eucalyptus and leaves of different ages

The autumn gum moth, Mnesampela privata, is a native Australian species whose preferred host, Eucalyptus globulus (Myrtaceae), is an aromatic evergreen tree that has long-lived waxy leaves during the juvenile phase of growth. We compared the behavioral and antennal responses of female moths to whole leaves (new and old) and samples of foliar chemicals (from new and old leaves) from a typical E. globulus subsp. pseudoglobulus with responses to a glossy, half-sibling E. g. subsp. pseudoglobulus putative hybrid (the result of natural cross-pollination). We also studied larval survival and development on leaves from the same trees. In laboratory binary-choice assays, female M. privata laid more eggs on waxy leaves than on glossy leaves thereby confirming the nonpreference for the glossy tree that was observed in the field. Analyses of the monoterpenes and waxes of both trees revealed that they had comparable suites of monoterpenes and total oil contents but different suites of epicuticular waxes. Headspace extracts differed in the intensity of component monoterpenes. Gas chromatographic analyses with electroantennographic detection showed different patterns of monoterpene detection. Leaves of the glossy tree had a less diverse array of epicuticular waxes than those of the waxy tree. Electroantennographic screening of responses to wax extracts from leaves (new and old) from either tree revealed positive dose-dependent responses of female antennae to waxes from new leaves only. Binary-choice assays also revealed a strong preference by ovipositing females for new, compared to old, leaves whether they were from the waxy or the glossy tree. However, new leaves from either tree could be manipulated (by physical abrasion of epicuticular waxes) so that females would lay almost no eggs on them. Larval survival did not differ between groups reared on leaves from both trees (new and old). Over 70% of all larvae survived to pupation. However, larvae reared on leaves from the glossy tree took longer to pupate than those reared on leaves from the waxy tree. Also, larvae reared on new leaves from either tree did not perform as well as those reared on old leaves. Monoterpene and wax cues are suggested as helping female M. privata locate preferred hosts in native forests.