Insect Sex Pheromones Research Articles

Expression patterns and colocalization of two sensory neurone membrane proteins in Ectropis obliqua Prout, a geometrid moth pest that uses Type-II sex pheromones.

Sensory neurone membrane proteins (SNMPs) function as essential cofactors for insect sex pheromone detection. In this study, we report two SNMPs in Ectropis obliqua Prout, a serious geometrid pest that produces typical Type-II sex pheromones. Sequence alignments and phylogenetic analyses showed that EoblSNMP1 and EoblSNMP2 belong to two distinct SNMP subfamilies. Quantitative real-time PCR suggested that EoblSNMP1 was male antennae-biased, whereas EoblSNMP2 was highly expressed on male antennae but was also expressed on female antennae and other chemosensory tissues. Additionally, EoblSNMP1 and EoblSNMP2 differed in their developmental expression profiles. In situ hybridization revealed that EoblSNMP1 was sensilla trichodea I specific, whereas EoblSNMP2 was expressed in sensilla trichodea I and the sensilla basiconica; furthermore, EoblSNMP1 and EoblSNMP2 were co-expressed in sensilla trichodea I but in different cells. This study suggests that EoblSNMP1 is functionally distinct from EoblSNMP2 in E.obliqua; EoblSNMP1 may specifically contribute to the recognition of sex pheromones, whereas EoblSNMP2 exhibits multiple olfactory roles. Our findings comprehensively reveal the expression patterns of SNMPs in a lepidopteran species that uses Type-II sex pheromones, providing new insights into the functional evolution of SNMPs from lepidopteran moths with Type-I sex pheromones to those with Type-II sex pheromones.

Ring‐Closing Strategy Utilizing Nitrile α‐Anions: Chiral Synthesis of (+)‐Norchrysanthemic Acid and Expeditious Asymmetric Total Synthesis of (+)‐Grandisol

Chiral syntheses of two distinct small cycloalkanes, (1R,3R)‐(1Z)‐norchrysanthemic acid and (+)‐grandisol, were performed by characteristic ring‐closing methodologies using carbanions at the α‐position of nitriles (nitrile α‐anions). (i) (1R,3R)‐(1Z)‐Norchrysanthemic acid, a highly potent ingredient of synthetic pyrethroid containing a cyclopropane structure, was synthesized from readily available (S)‐epoxide derived from 3‐methyl‐but‐2‐en‐1‐ol in 7 steps in 23 % overall yield and with > 98 % ee. This sequence involves a trans‐selective cyclopropane formation using the nitrile α‐anion of (S)‐3‐mesyloxynitrile as the key step. The present chiral synthesis was performed with effective stereocontrol of both the chirality in the 1,3‐positions on the cyclopropane and the Z‐geometry of the propenyl group. (ii) (+)‐Grandisol, an insect sex pheromone possessing a characteristic cyclobutane structure, was synthesized from commercially available cyclopropyl methyl ketone (route A) or from commercially available 3‐cyanopropylzinc bromide and 1‐bromo‐1‐methylpropene (route B) in 10 or 8 steps in 6 % or 8 % overall yield and with 80 % ee. This sequence involves a Shi asymmetric epoxidation of a trisubstituted olefin and a straightforward Stork‐type asymmetric cyclobutane formation with clean SN2 stereoinversion using the nitrile α‐anion of the chiral epoxynitrile. The present expedient method is the second asymmetric total synthesis starting from achiral compounds.

Research Progress and Application Prospect of Insect Sex Pheromone Mating Disruption

Research Progress and Application Prospect of Insect Sex Pheromone Mating Disruption

The Use of Mercury Compounds in the Synthesis of Some Lepidoptera Insect Sex Pheromones with Monoenic Structure

New and practical synthesis of (E)-11-tetradecen-1-yl acetate and (Z)-11-hexadecen-1-yl acetate were developed. The synthesis were based on a C12+C2=C14 and C12+C4=C16 coupling scheme. The routes involve, as the key step, the use of the same mercury derivative of the terminal-alkyne w-functionalised as intermediate.The first coupling reaction was effected by adding 1-tert-butoxy-10-bromodecane to monosodate acetylene, obtained in situ from DMSO and sodium hydride. It was prepared 1-tert-butoxy-dodec-11-yne, which is transformed in di[tert-butoxy-dodec-11-yne]mercury, the common intermediate in the synthesis of the two pheromones. In order to obtain (E)-11-tetradecen-1-yl acetate, the mercury derivative was directly lithiated and then alkylated with 1-bromoethane obtaining 1-tert-butoxy-tetradec-11-yne. After reduction with lithium aluminium hydride of 1-tert-butoxy-tetradec-11-yne and acetylation gave (E)-11-tetradecen-1-yl acetate with 99 % isomeric purity. In order to obtain (Z)-11-hexadecen-1-yl acetate, the mercury derivative was directly lithiated and then alkylated with 1-bromobutane obtaining 1-tert-butoxy-hexadec-11-yne. After acetylation of 1-tert-butoxy-hexadec-11-yne and stereoselective reduction in the presence of NiP-2 catalyst gave (Z)-11-hexadecen-1-yl acetate with 90 % isomeric purity.

Structure-Activity Studies of Semiochemicals from the Spider Orchid Caladenia plicata for Sexual Deception.

Sexually deceptive orchids attract specific pollinators by mimicking insect sex pheromones. Normally this mimicry is very specific and identical compounds have been identified from orchids and matching females of the pollinators. In this study, we conduct a detailed structure-activity investigation on isomers of the semiochemicals involved in the sexual attraction of the male pollinator of the spider orchid Caladenia plicata. This orchid employs an unusual blend of two biosynthetically unrelated compounds, (S)-β-citronellol and 2-hydroxy-6-methylacetophenone, to lure its Zeleboria sp. thynnine wasp pollinator. We show that the blend is barely attractive when (S)-β-citronellol is substituted with its enantiomer, (R)-β-citronellol. Furthermore, none of the nine-possible alternative hydroxy-methylacetophenone regioisomers of the natural semiochemical are active when substituted for the natural 2-hydroxy-6-methylacetophenone. Our results were surprising given the structural similarity between the active compound and some of the analogues tested, and results from previous studies in other sexually deceptive orchid/wasp systems where substitution with analogues was possible. Interestingly, high-level ab initio and density functional theory calculations of the hydroxy-methylacetophenones revealed that the active natural isomer, 2-hydroxy-6-methylacetophenone, has the strongest intramolecular hydrogen bond of all regioisomers, which at least in part may explain the specific activity.

KlebsiellaBacteria Isolated from the Genital Chamber ofPhyllophaga obsoleta

Females of Phyllophaga obsoleta Blanchard (Coleoptera: Melolonthidae) display a typical “calling” behavior by protruding the abdominal tip and exposing their genital chamber. This is the moment when the sex pheromone is released into the environment. The genital chamber and accessory glands have a specialized epithelia involved in production of the chemicals. In other species of the group (e.g., Costelytra zealandica White), bacteria have been confirmed to be involved. We identified the microorganisms in the genital chamber of females of P. obsoleta. Two types of bacterial colonies were isolated from the inside of the genital chamber and identified using the 16S rRNA gene sequence. The sequence of one of the bacterial colonies was a 97% match with the Klebsiella oxytoca strain HP1 sequence, while the sequence of the other bacterial colony was a 98% match with the Klebsiella michiganensis strain VITSW4 sequence. Microbiological and biochemical tests were used for taxonomic identification. Both bacteria produce indole and acetoin, chemicals that have been reported as attractants and sex pheromones in other insects, showing they can potentially participate in production of sex pheromone of P. obsoleta.

Effect of temperature on the sex pheromone communication of Chilo suppressalis Walker (Lepidoptera: Pyralidae)

Environmental factors affect the insect sex pheromone communication, and subsequently the attractiveness of sex pheromone lures used in pest control. In order to provide the guidance for the sex pheromone use in control of Chilo suppressalis, effects of temperature on the sex pheromone production in female moths and electro-physiological response of male moths to sex pheromones, were explored in our present study. The insects were treated with different temperatures (15, 20, 25, 30 and 35 ℃) at pupa and adult stages, then the contents and relative proportion of each of three pheromone components were determined by a gas chromatography (GC), and the electro-physiological response to sex pheromones were detected by an electroantennograph (EAG). Results showed that the contents of all three sex pheromone components (Z9-16:Ald, Z11-16:Ald and Z13-18:Ald) were highest in 25 ℃ treated females, significantly higher than those in other temperature treated females, and the relative proportion of Z13-18:Ald was significantly reduced in 25 ℃ treated females. On the male part, EAG responses to each of the three sex pheromone components and the tertiary blend showed no significant difference at temperatures between 15-25 ℃, but EAG va-lues decreased significantly as the temperature increased to 30 ℃ and further to 35 ℃ for component Z9-16:Ald, Z11-16:Ald and the blend, and to 35 ℃ for Z13-18:Ald. Taken together, the optimum temperature for the sex pheromone communication was 20-25 ℃ in C. suppressalis, and too higher or lower temperature will affect the normal communication. The study provided an important reference for the use of sex pheromone in pest control, and for prediction of the development of pest population at extremely low or high temperature.

Production, identification, and field evaluation of sex pheromone from calling females in Diaphania angustalis (Lepidoptera: Crambidae).

Insect sex pheromones play a crucial role in the mate finding and calling behavior of Lepidoptera pests. Currently, little is known about the chemical ecology of Diaphania angustalis Snellen (Lepidoptera: Crambidae), a severe and important defoliator attacking the medicinal plant, Alstonia scholaris. In the present study, the pheromone components of D. angustalis females were investigated using electrophysiological and behavioral methods. Distilled hexane extracts of female pheromone glands were analyzed through electroantennogram (EAG) and gas chromatography-electroantennogram detector (GC-EAD), and the active compounds were identified through gas chromatography-mass spectrometry (GC-MS). Production peak of female sex pheromone occurred on the third day of age at 5h into the scotophase with the EAG test, and the hexane extracts were attractive to males in the wind tunnel test. GC-EAD analysis of virgin males to gland extracts that were subsequently evaluated showed two active compounds, (E,E)-10,12-hexadecadienal (E10E12-16:Ald) and (E,E)-10,12-hexadecadien-1-ol (E10E12-16:OH), based on comparison of retention time and mass spectrum, with suitable synthetic compounds. Under laboratory conditions, the blend of E10E12-16:Ald and E10E12-16:OH in a ratio of 9:1 elicited a stronger EAG response than other treatments or a single component. In the field, more male moths were captured by traps baited with the mixture of E10E2-16:Ald and E10E2-16:OH in a ratio of 9:1, whereas a mixture of 8:1 and 10:1 also caught males. Accordingly, E10E2-16:Ald and E10E2-16:OH were regarded as the major sex pheromone components in D. angustalis females.

(1S,3R)-cis-Chrysanthemyl Tiglate: Sex Pheromone of the Striped Mealybug, Ferrisia virgata.

Derivatives of 2,2-dimethyl-3-(2-methylprop-1-enyl)cyclopropanecarboxylic acid (chrysanthemic acid) are classic natural pyrethroids discovered in pyrethrum plants and show insecticidal activity. Chrysanthemic acid, with two asymmetric carbons, has four possible stereoisomers, and most natural pyrethroids have the (1R,3R)-trans configuration. Interestingly, chrysanthemic acid-related structures are also found in insect sex pheromones; carboxylic esters of (1R,3R)-trans-(2,2-dimethyl-3-(2-methylprop-1-enyl)cyclopropyl)methanol (chrysanthemyl alcohol) have been reported from two mealybug species. In the present study, another ester of chrysanthemyl alcohol was discovered from the striped mealybug, Ferrisia virgata (Cockerell), as its pheromone. By means of gas chromatography-mass spectrometry, nuclear magnetic resonance spectrometry, and high-performance liquid chromatography analyses using a chiral stationary phase column and authentic standards, the pheromone was identified as (1S,3R)-(-)-cis-chrysanthemyl tiglate. The (1S,3R)-enantiomer strongly attracted adult males in a greenhouse trapping bioassay, whereas the other enantiomers showed only weak activity. The cis configuration of the chrysanthemic acid-related structure appears to be relatively scarce in nature, and this is the first example reported from arthropods.

The developmental transcriptome of the bamboo snout beetle Cyrtotrachelus buqueti and insights into candidate pheromone-binding proteins.

Cyrtotrachelus buqueti is an extremely harmful bamboo borer, and the larvae of this pest attack clumping bamboo shoots. Pheromone-binding proteins (PBPs) play an important role in identifying insect sex pheromones, but the C. buqueti genome is not readily available for PBP analysis. Developmental transcriptomes of eggs, larvae from the first instar to the prepupal stage, pupae, and adults (females and males) from emergence to mating were built by RNA sequencing (RNA-Seq) in the present study to establish a sequence background of C. buqueti to help understand PBPs. Approximately 164.8 million clean reads were obtained and annotated into 108,854 transcripts. These were assembled into 24,338, 21,597, 24,798, 21,886, 24,642, and 83,115 unigenes for eggs, larvae, pupae, females, males, and the combined datasets, respectively. Unigenes were annotated against NCBI non-redundant protein sequences, NCBI non-redundant nucleotide sequences, Gene Ontology (GO), Protein family, Clusters of Orthologous Groups of Proteins/ Clusters of Eukaryotic Orthologous Groups (KOG), Swiss-Prot, and KEGG Orthology databases. A total of 17,213 unigenes were annotated into 55 sub-categories belonging to three main GO categories; 10,672 unigenes were classified into 26 functional categories by KOG classification, and 8,063 unigenes were classified into five functional KEGG categories. RSEM software for RNA sequencing showed that 4,816, 3,176, 3,661, 2,898, 4,316, 8,019, 7,273, 5,922, 5,844, and 4,570 genes were differentially expressed between larvae and males, larvae and eggs, larvae and pupae, larvae and females, males and females, males and eggs, males and pupae, females and eggs, females and pupae, and eggs and pupae, respectively. Of these, three were confirmed to be significantly differentially expressed between larvae, females, and males. Furthermore, PBP Cbuq7577_g1 was highly expressed in the antenna of males. A comprehensive sequence resource of a desirable quality was constructed from developmental transcriptomes of C. buqueti eggs, larvae, pupae, and adults. This work enriches the genomic data of C. buqueti, and facilitates our understanding of its metamorphosis, development, and response to environmental change. The identified candidate PBP Cbuq7577_g1 might play a crucial role in identifying sex pheromones, and could be used as a targeted gene to control C. buqueti numbers by disrupting sex pheromone communication.

Knockdown of a metathoracic scent gland desaturase enhances the production of (E)-4-oxo-2-hexenal and suppresses female sexual attractiveness in the plant bug Adelphocoris suturalis.

Insect sex pheromones (SPs) are central to mate-finding behaviour, and play an essential role in the survival and reproduction of organisms. Understanding the roles, biosynthetic pathways and evolution of insect chemical communication systems has been an exciting challenge for biologists. Compared with Lepidoptera, little is known about the mechanisms underlying pheromone biosynthesis in Hemiptera. Inthis study, we isolated and characterized two newdesaturase-like genes, termed Asutdes1 and Asutdes2, from Adelphocoris suturalis, an important agricultural pest in China. Although the two genes encode an identical protein, Southern blot analysis revealed that they are duplicated genes. The Asutdes2 transcript is more abundant than Asutdes1 in the tissues tested, in particular the metathoracic scent gland and fat body. Silencing Asutdes expression in females by injecting double-stranded RNA (dsAsutdes) against a portion of the coding sequence shared by the two genes enhanced the production of (E)-4-oxo-2-hexenal, a component of the A.suturalis SP blend, and dramatically suppressed the sexual attractiveness of A.suturalis females. We conclude that dsAsutdes is associated with the SP biosynthetic pathway in A.suturalis.

Synthesis of the enantiomers of (3Z,9Z)-cis-6,7-epoxy-3,9-octadecadiene, one of the major components of the sex pheromone of Ectropis oblique Prout

Both enantiomers of (3Z,9Z)-cis-6,7-epoxy-3,9-octadecadiene, one of which is the major component of the sex pheromone of Ectropis oblique Prout, were synthesized in 23% overall yield for the (−)-(6S,7R)-enantiomer and 18% yield for the (+)-(6R,7S)-isomer. This protocol uses a sequential regioselective ring-opening strategy and provides a convenient and reliable access to other structurally related insect sex pheromones. Preliminary biological studies revealed that (−)-(6S,7R)-2a was roughly as active as the natural pheromone, while racemic (±)-2 was less bioactive and (+)-2b was much less bioactive.

A cytochrome P450 gene plays a role in the recognition of sex pheromones in the tobacco cutworm, Spodoptera litura.

Cytochrome P450 (P450 or CYP) genes are involved in fundamental physiological functions, and might be also associated with the olfactory recognition of sex pheromones in beetles and moths. A P450 gene, Spodoptera litura CYP4L4 (SlituCYP4L4), was cloned for the first time from the antennae of S.litura. SlituCYP4L4 was almost exclusively expressed in the adult stage and predominantly expressed in the adult antennae. In situ hybridization showed that SlituCYP4L4 localized mainly at the base of the long sensilla trichoidea, which responds to sex pheromone components. Pretreatment with an S.litura sex pheromone significantly reduced the expression levels of SlituCYP4L4, consistent with other genes involved in sex pheromone recognition. The expression level of SlituCYP4L4 was different in moths collected with different ratios of sex pheromone lures and collected in different geographical locations. After gene knockdown of SlituCYP4L4 in the antennae, the electroantennogram (EAG) responses of male and female moths to (9Z,11E)-tetradecadienyl acetate or (9Z,12E)-tetradecadienyl acetate were significantly decreased. In contrast, EAG responses to plant volatiles and sex pheromones of other moth species were not significantly influenced in these moths. SlituCYP4L4 was also expressed in the gustatory tissues and sensilla, which suggests that SlituCYP4L4 may have other functions in the chemosensory system. Our results have shown for the first time the function of a CYP gene with appendage-specific expression in insect sex pheromone recognition, especially in adult moths.

Management of Rice Yellow Stem Borer Scirpophaga insertulas (Walker) using Different Formulations of Insect Sex Pheromone in West Bengal

Management of Rice Yellow Stem Borer Scirpophaga insertulas (Walker) using Different Formulations of Insect Sex Pheromone in West Bengal

Plant Volatiles Increase Sex Pheromone Attraction of Holotrichia parallela (Coleoptera: Scarabaeoidea).

Holotrichia parallela (Coleoptera: Scarabaeoidea) is a notorious pest of many crops. To improve the effectiveness of its female-produced sex pheromone (L-isoleucine methyl ester:(R)-(-)-linalool=6:1), 14 plant volatiles, including dodecanoic acid, dodecanal, farnesol, α-farnesene, (Z)-3-hexen-1-ol, (E)-2-hexen-1-ol, (Z)-3-hexenyl acetate, (E)-2-hexenyl acetate, (R)-(+)-limonene, α-phellandrene, α-pinene, ocimene, methyl benzoate, and benzaldehyde, were individually evaluated using electroantennography and olfactometer assays. (E)-2-Hexenyl acetate and (Z)-3-hexenyl acetate were found to elicit the strongest responses in both males and females. Further testing of these two compounds in mixtures with the sex pheromone indicated that (E)-2-hexenyl acetate had a stronger synergistic effect than (Z)-3-hexenyl acetate. Field evaluations showed that mixtures of (E)-2-hexenyl acetate and the sex pheromone resulted in significantly higher catches than the sex pheromone alone. Using a 5:1 mixture of the sex pheromone and (E)-2-hexenyl acetate, the maximum number of females per trap per day was 14, showing a synergistic effect of a factor of four. For males, a 3:1 mixture of the sex pheromone and (E)-2-hexenyl acetate yielded a maximum number of 310 individuals per trap per day, equivalent to a synergistic effect of 175%. These results may provide the basis for the development of efficient pest management systems against H. parallela using plant volatiles and insect sex pheromones.

A Practical Synthesis of (Z)- and (E)-8-Dodecene-1-yl Acetate, Components of Lepidoptera Insect Sex Pheromones

New and practical synthesis of (Z)-8-dodecene-1-yl acetate and (E)-8-dodecene-1-yl acetate were developed. The synthesis were based on a C3+C6=C9 and C9+C3=C12 coupling scheme, the starting material being 2-propyn-1-ol and 1,6-hexandiol. The routes involve, as the key step, the use of the same mercury derivative of the terminal-alkyne w-functionalised as intermediate. The first coupling reaction took place between methoxyallene and Grignard reagent of 1-tert-butoxy-6-bromo-hexan obtaining 1-tert-butoxy-non-8-yne, which is transformed in di[1-tert-butoxy-non-8-yne]mercury, the common intermediate in the synthesis of the two pheromones. In order to obtain (Z)-8-dodecene-1-yl acetate and (E)-8-dodecene-1-yl acetate, the mercury derivative was directly lithiated and then alkylated with 1-bromo-propan obtaining 1-tert-butoxy-dodec-8-yne. After acetylation of 1-tert-butoxy-dodec-8-yne and stereoselective reduction in the presence of NiP-2 catalyst gave (Z)-8-dodecene-1-yl acetate with 85 % isomeric purity. After reduction with lithium aluminium hydride of 1-tert-butoxy-dodec-8-yne and acetylation was obtained (E)-8-dodecene-1-yl acetate with 90% isomeric purity.

Convenient Synthesis of (Z)-7- and (E)-9-dodecene-1-yl Acetate, Components of Some Lepidoptera Insect Sex Pheromone

Were developed new and practical synthesis of (Z)-7-dodecene-1-yl acetate and (E)-9-dodecene-1-yl acetate. The routes involve, as the key step, the use of the mercury derivative of the terminal-alkyne w-functionalised as intermediate. The synthesis of (Z)-7-dodecene-1-yl acetate was based on a C6+C2=C8 and C8+C4=C12 coupling scheme, starting from 1,6-hexane-diol. The first coupling reaction took place between 1-tert-butoxy-6-bromo-hexane and lithium acetylide-ethylendiamine complex obtaining 1-tert-butoxy-oct-7-yne, which is transformed in di[tert-butoxy-oct-7-yne]mercury. The mercury derivative was directly lithiated and then alkylated with 1-bromobutane obtaining 1-tert-butoxy-dodec-7-yne. After acetylation and reduction with lithium aluminium hydride of 7-dodecyne-1-yl acetate gave (Z)-7-dodecene-1-yl acetate with 96 % purity. The synthesis of (E)-9-dodecene-1-yl acetate was based on a C8+C2=C10 and C10+C2=C12 coupling scheme, starting from 1,8-octane-diol. The first coupling reaction took place between 1-tert-butoxy-8-bromo-octane and lithium acetylide-ethylendiamine complex obtaining 1-tert-butoxy-dec-9-yne, which is transformed in di[tert-butoxy-dec-9-yne]mercury. The mercury derivative was directly lithiated and then alkylated with 1-bromoethane obtaining 1-tert-butoxy-dodec-9-yne. After reduction with lithium aluminium hydride of 1-tert-butoxy-(E)-9-dodecene and acetylation was obtained (E)-9-dodecene-1-yl acetate with 97 % purity.

RNAi-Induced Electrophysiological and Behavioral Changes Reveal two Pheromone Binding Proteins of Helicoverpa armigera Involved in the Perception of the Main Sex Pheromone Component Z11-16:Ald.

Pheromone binding proteins (PBPs) are thought to play key roles in insect sex pheromone recognition; however, there is little in vivo evidence to support this viewpoint in comparison to abundant biochemical data in vitro. In the present study, two noctuid PBP genes HarmPBP1 and HarmPBP2 of the serious agricultural pest, Helicoverpa armigera were selected to be knocked down by RNA interference, and then the changes in electrophysiological and behavioral responses of male mutants to their major sex pheromone component (Z)-11-hexadecenal (Z11-16:Ald) were recorded. There were no significant electrophysiological or behavioral changes of tested male moths in response to Z11-16:Ald when either single PBP gene was knocked down. However, decreased sensitivity of male moths in response to Z11-16:Ald was observed when both HarmPBP1 and HarmPBP2 genes were silenced. These results reveal that both HarmPBP1 and HarmPBP2 are required for the recognition of the main sex pheromone component Z11-16:Ald in H. armigera. Furthermore, these findings may help clarify physiological roles of moth PBPs in the sex pheromone recognition pathway, which in turn could facilitate pest control by exploring sex pheromone blocking agents.

Convergent evolution of sexual deception via chromatic and achromatic contrast rather than colour mimicry

The Orchidaceae is characterised by the repeated evolution of sexual deception, one of the most specialised pollination strategies. In orchids, sexual deception involves long-range pollinator attraction via mimicry of female insect sex pheromones. At close range, visual signals involving colour mimicry, contrast to the background, and exploitation of pollinator sensory biases could attract pollinators, but remain largely untested. Here we focus on a remarkable system in which species from two only distantly related sexually deceptive orchid genera with strikingly different flowers (Drakaea livida and three species of Caladenia) share the same pollinator, males of the thynnine wasp Zaspilothynnus nigripes. We used spectral reflectance measurements and modelling to investigate pollinator perception of colour, including the first examination of overall colour patterns in flowers via colour pattern geometry analyses. Rather than closely matching the colours of female Z. nigripes, these orchids had strong chromatic and achromatic contrast against their backgrounds. For Caladenia, the sepals and petals show high contrast, while in D. livida, which has diminutive petals and sepals, it is the labellum that contrasts strongly against the background. Despite varying in colour, the Caladenia species all had strong within-flower contrast between a UV-bright central target (column and labellum) and a corolla of radiating stripes (petals and sepals). The colour pattern geometry analyses also indicated that the orchids’ overall colour patterns are highly conspicuous against their backgrounds. Contrast, UV, and target patterns could all enhance detection, and exploit pollinators’ innate preferences. Since colour contrast may function with a range of colours and floral forms, attracting pollinators via contrast rather than visual mimicry may be a critical but previously overlooked process facilitating the evolution of sexual deception.

Morganella morganii bacteria produces phenol as the sex pheromone of the New Zealand grass grub from tyrosine in the colleterial gland.

Costelytra zealandica (Coleoptera: Scarabeidae) is a univoltine endemic species that has colonised and become a major pest of introduced clover and ryegrass pastures that form about half of the land area of New Zealand. Female beetles were previously shown to use phenol as their sex pheromone produced by symbiotic bacteria in the accessory or colleterial gland. In this study, production of phenol was confirmed from the female beetles, while bacteria were isolated from the gland and tested for attractiveness towards grass grub males in traps in the field. The phenol-producing bacterial taxon was identified by partial sequencing of the 16SrRNA gene, as Morganella morganii. We then tested the hypothesis that the phenol sex pheromone is biosynthesized from the amino acid tyrosine by the bacteria. This was shown to be correct, by addition of isotopically labelled tyrosine ((13)C) to the bacterial broth, followed by detection of the labelled phenol by SPME-GCMS. Elucidation of this pathway provides specific evidence how the phenol is produced as an insect sex pheromone by a mutualistic bacteria.