Summary Sexually deceptive orchids are predicted to represent a special case of plant speciation where strong reproductive isolation may be achieved by differences in floral scent. In this study of Australian sexually deceptive Chiloglottis orchids, we performed choice experiments to test for wasp pollinator specificity in the field; identified the compounds involved in pollinator attraction by gas chromatography with electroantennographic detection (GC‐EAD), gas chromatography with mass selective detection (GC‐MS), chemical synthesis and behavioural bioassays; and mapped our chemical findings on to a phylogeny of the orchids. Field experiments confirmed pollination is a highly specific interaction, but also revealed a pool of nonpollinating ‘minor responder’ wasps. Six novel compounds, all 2,5‐dialkylcyclohexan‐1,3‐diones, called ‘chiloglottones’, were discovered to be involved in pollinator attraction. Bioassays confirmed that pollinator specificity has a strong chemical basis, with specificity among sympatric orchids maintained by either different single compounds or a variation in a blend of two compounds. The phylogenetic overlay confirmed that speciation is always associated with pollinator switching and usually underpinned by chemical change. If the chemical differences that control reproductive isolation in Chiloglottis have a strong genetic basis, and given the confirmed pool of potential pollinators, we conclude that pollinator‐driven speciation appears highly plausible in this system.

Orchids employing sexual deceit attract males of their pollinator species through specific volatile signals that mimic female-released sex pheromones. One of these signals proved to be 2-ethyl-5-propylcyclohexan-1,3-dione (chiloglottone1), a new natural product that was shown to be most important in the relations between orchids of the genus Chiloglottis, native to Australia, and corresponding pollinator species. Systematic investigations on the mass spectrometric fragmentation pattern of 2,5-dialkylcyclohexan-1,3-diones identified key ions providing information about the structures of the substituents at positions 2 and 5. Results enabled us to identify 2-ethyl-5-pentylcyclohexan-1,3-dione (chiloglottone2) and 2-butyl-5-methylcyclohexan-1,3-dione (chiloglottone3) as new natural products that play a decisive role in the pollination syndrome of some Chiloglottis species. During field bioassays, pure synthetic samples of chiloglottone1-3 or mixtures thereof proved to be attractive to the corresponding orchid pollinators. Because of their likely biogenesis from ubiquitous fatty acid precursors, 2,5-dialkylcyclohexan-1,3-diones may represent a hitherto overlooked, widespread class of natural products.2-butyl-5-methylcyclohexan-1,3-dione ͉ 2-ethyl-5-pentylcyclohexan-1,3-dione ͉ Chiloglottis ͉ semiochemical ͉ mass spectrometry

UV-B light is required for the synthesis of chiloglottones - the semiochemicals used by Chiloglottis orchids to sexually lure their male pollinators. This discovery appears to be the first case to our knowledge where plant floral odour production depends on UV-B radiation at normal levels of sunlight. In the future, identification of the genes and enzymes involved, will allow us to understand better the role of UV-B light in the biosynthesis of chiloglottones.

The sepals and petals of Chiloglottis orchids strongly block UV-B wavelengths of light, preventing chiloglottone production inside the bud. While initiation of chiloglottone biosynthesis requires only UV-B light, sustained chiloglottone biosynthesis requires both UV-B and de novo protein biosynthesis. The internal amounts of chiloglottone in a flower reflect the interplay between developmental stage, duration and intensity of UV-B exposure, de novo protein synthesis, and feedback loops linked to the starting amount of chiloglottone. It is concluded that UV-B light contributes directly to chiloglottone biosynthesis. These findings suggest an entirely new and unexpected biochemical reaction that might also occur in taxa other than these orchids.

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A five-step synthesis of monoalkyl- and 2,5-dialkyl-1,3-cyclohexanediones (1) is described via a sequence involving sequential Birch reductions and alkylations from the readily accessible and inexpensive starting material, 3,5-dimethoxybenzoic acid. Two approaches were considered in which alkylation at C-2 occurs either prior or subsequent to the proposed reduction. The successful route, in which Birch reduction of a 3-alkyl resorcinol derivative (3) precedes alkylation was applied in the synthesis of chiloglottone 1 (1dc), in 58% overall yield. Chiloglottone 1 is a member of a new class of natural products, representing a known sex pheromone of the thynnine wasp Neozeleboria cryptoides and pollinator attractant in the Australian sexually deceptive orchid genus Chiloglottis. The synthetic homologues were assessed for their biological activity via electroantennographic detection.

Orchids of the genus Chiloglottis rely on sexual deception to attract a male thynnine wasp, whereupon a courtship routine results in the deposition and transfer of pollinia and subsequent pollination of the orchid. The chemical cues responsible for this behavior were recognized as a new class of natural products and efforts to synthetically prepare them, to assist in structural identification and to aid in ecological studies, have been ongoing. Most recently the first alkenylated member of this class was proposed based on GC–MS analysis of a physiologically active compound present in an orchid extract. The current work reveals synthetic efforts that provide access to diastereomerically pure unsaturated chiloglottone analogues and in combination with laboratory‐based electrophysiological assays and field‐based studies have unambiguously allowed identification of 5‐allyl‐2‐ethyl‐1,3‐cyclohexanedione (1d, chiloglottone 4) as the first alkenyl congener in the chiloglottone family. Convergent synthetic approaches, involving copper(I)‐mediated conjugate addition to a glutaconic ester converging on cadmium‐mediated desymmetrization of cyclic anhydrides or a Johnson–Claisen rearrangement of a mixed ketene acetal provided the appropriately substituted δ‐keto acids suitable for carbocyclization to furnish the required chiloglottone analogues.

Many arthropod pests of humans and other animals select their preferred hosts by recognising volatile odour compounds contained in the hosts’ ‘volatilome’. Although there is prolific literature on chemical emissions from humans, published data on volatiles and vector attraction in other species are more sporadic. Despite several decades since the identification of a small number of critical volatiles underpinning specific host–vector relationships, synthetic chemicals or mixtures still largely fail to reproduce the attractiveness of natural hosts to their disease vectors. This review documents allelochemicals from non-human terrestrial animals and considers where challenges in collection and analysis have left shortfalls in animal volatilome research. A total of 1287 volatile organic compounds were identified from 141 species. Despite comparable diversity of entities in each compound class, no specific chemical is ubiquitous in all species reviewed, and over half are reported as unique to a single species. This review provides a rationale for future enquiries by highlighting research gaps, such as disregard for the contribution of breath volatiles to the whole animal volatilome and evaluating the role of allomones as vector deterrents. New opportunities to improve vector surveillance and disrupt disease transmission may be unveiled by understanding the host-associated stimuli that drive vector-host interactions.