Manipulating two olfactory cues causes a biological control beetle to shift to non‐target plant species

1. Olfactory cues can determine the host preferences of herbivorous insects, but their role in host shifting is unclear. Host specificity and the potential for host shifts are important criteria for screening and post-release evaluation of biological control agents for invasive plants. However, the role of olfactory cues in mediating host shifts in biological control agents is not well understood. 2. To investigate the role of olfactory cues in host selection of a reportedly monophagous flea beetle (Agasicles hygrophila), an important biocontrol agent for invasive alligator weed (Alternanthera philoxeroides), we extracted and analysed the volatiles produced by the host-plant A. philoxeroides and the non-host plants Alternanthera sessilis, Beta vulgaris and Amaranthus mangostanus. Moreover, we used electrophysiological techniques, behavioural bioassays and field trials to test the antennal responses and behavioural preferences of A. hygrophila to combinations of different plant volatiles and treatments, and pure compounds in different dosages and combinations. 3. We show that A. hygrophila female beetles indeed use olfactory cues to select plants for feeding and oviposition and that the survivorship of larvae on the second preferred non-host plant A. sessilis, a close relative of the first preferred host plant A. philoxeroides, was over 75% in a field trail. Although female beetles responded to many volatile compounds from host and non-host plants, (E)-4,8-dimethyl-1,3,7-nonatriene (DMNT) positively encouraged the beetle's feeding and oviposition preferences, whereas (Z)-3-hexenol displayed repellent effect. Remarkably, complementation assays with (Z)-3-hexenol on host plant or DMNT on non-host plants significantly shifted A. hygrophila host preferences to non-host plants and resulted in oviposition and egg hatching on the non-host plant A. sessilis in field trials. 4. Synthesis. We demonstrate an olfactory mechanism by which a specialized herbivorous beetle uses the ratio of two common plant volatiles, (E)-4,8-dimethyl-1,3,7-nonatriene and (Z)-3-hexenol, to discriminate between its host and non-host plants in nature. This study highlights an important mechanism by which olfactory cues could lead to undesired host range expansion in biocontrol agent, thus representing an important warning of the potential for a host shift and development of invasiveness in a common biocontrol agent, the flea beetle.