Deorphanizing an odorant receptor tuned to palm tree volatile esters in the Asian palm weevil sheds light on the mechanisms of palm tree selection.

The Asian palm weevil, Rhynchophorus ferrugineus, is a tremendously important agricultural pest primarily adapted to palm trees and causes severe destruction, threatening sustainable palm cultivation worldwide. The host plant selection of this weevil is mainly attributed to the functional specialization of odorant receptors (ORs) that detect palm-derived volatiles. Yet, ligands are known for only two ORs of R. ferrugineus, and we still lack information on the mechanisms of palm tree detection. This study identified a highly expressed antennal R. ferrugineus OR, RferOR2, thanks to newly generated transcriptomic data. The phylogenetic analysis revealed that RferOR2 belongs to the major coleopteran OR group 2A and is closely related to a sister clade containing an R. ferrugineus OR (RferOR41) tuned to the non-host plant volatile and antagonist, α-pinene. Functional characterization of RferOR2 via heterologous expression in Drosophila olfactory neurons revealed that this receptor is tuned to several ecologically relevant palm-emitted odors, most notably ethyl and methyl ester compounds, but not to any of the pheromone compounds tested, including the R. ferrugineus aggregation pheromone. We did not evidence any differential expression of RferOR2 in the antennae of both sexes, suggesting males and females detect these compounds equally. Next, we used the newly identified RferOR2 ligands to demonstrate that including synthetic palm ester volatiles as single compounds and in combinations in pheromone-based mass trapping has a synergistic attractiveness effect to R. ferrugineus aggregation pheromone, resulting in significantly increased weevil catches. Our study identified a key OR from a palm weevil species tuned to several ecologically relevant palm volatiles and represents a significant step forward in understanding the chemosensory mechanisms of host detection in palm weevils. Our study also defines RferOR2 as an essential model for exploring the molecular basis of host detection in other palm weevil species. Finally, our work showed that insect OR deorphanization could aid in identifying novel behaviorally active volatiles that can interfere with weevil host-searching behavior in sustainable pest management applications.

Pheromone receptor of the globally invasive quarantine pest of the palm tree, the red palm weevil (Rhynchophorus ferrugineus).

Palm trees are of immense economic, sociocultural, touristic, and patrimonial significance all over the world, and date palm-related knowledge, traditions, and practices are now included in UNESCOs list of the Intangible Cultural Heritage of Humanity. Of all the pests that infest these trees, the red palm weevil (RPW), Rhynchophorus ferrugineus (Olivier), is its primary enemy. The RPW is a category-1 quarantine insect pest that causes enormous economic losses in palm tree cultivation worldwide. The RPW synchronizes mass gathering on the palm tree for feeding and mating, regulated by a male-produced pheromone composed of two methyl-branched compounds, (4RS, 5RS)-4-methylnonan-5-ol (ferrugineol) and 4(RS)-methylnonan-5-one (ferrugineone). Despite the importance of odorant detection in long-range orientation towards palm trees, palm colonization, and mating, the pheromone receptor has not been identified in this species. In this study, we report the identification and characterization of the first RPW pheromone receptor, RferOR1. Using gene silencing and functional expression in Drosophila olfactory receptor neurons, we demonstrate that RferOR1 is tuned to ferrugineol and ferrugineone and binds five other structurally related molecules. We reveal the lifetime expression of RferOR1, which correlates with adult mating success irrespective of age, a factor that could explain the wide distribution and spread of this pest. As palm weevils are challenging to control based on conventional methods, elucidation of the mechanisms of pheromone detection opens new routes for mating disruption and the early detection of this pest via the development of pheromone receptor-based biosensors.

Gravimetric biosensors.

Gravimetric transducers produce a signal based on a change in mass. These transducers can be used to construct gas sensors or biosensors using odorant binding proteins (OBPs) as recognition elements for small volatile organic compounds. The methods described in this chapter are based on the immobilization of the OBPs onto functionalized (activated) self-assembled monolayer (SAMs) on gold and on nanocrystalline diamond surfaces. Depending on the surface immobilization methods used to fabricate the biosensor, recombinant proteins can be engineered to express six histidine tags either on the N-terminal or C-terminal of the proteins and these can also be used to facilitate protein immobilization. These methods are used to produce functional sensors based on quartz crystal microbalances or surface acoustic wave devices and are also applicable to other types of gravimetric transducers.

Modification of an Anopheles gambiae odorant binding protein to create an array of chemical sensors for detection of drugs.

The binding pockets of odorant binding proteins from Anopheles gambiae (OBP1 and OBP47) were analysed using in silico modelling. The feasibility of creating mutant proteins to achieve a protein array capable of detecting drugs of abuse in solution or in vapour phase was investigated. OBP1 was found to be easily adapted and several mutant proteins were expressed and characterised. AgamOBP1_S82P was found to have high affinities to cannabinol, 3,4-methylenedioxy methamphetamine (MDMA/Ecstasy) and cocaine hydrochloride. When these proteins were immobilised on a quartz crystal microbalance, saturated cocaine hydrochloride vapour could be detected. The sensors were stable over a period of at least 10 months in air. The approach taken allows flexible design of new biosensors based on inherently stable protein scaffolds taking advantage of the tertiary structure of odorant binding proteins.