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The sex pheromone of the azalea mealybug, Crisicoccus azaleae (Tinsley, 1898) (Hemiptera: Pseudococcidae), includes esters of a methyl-branched medium-chain fatty acid, ethyl and isopropyl (E)-7-methyl-4-nonenoate. These compounds are exceptional among mealybug pheromones, which are commonly monoterpenes. Determination of the absolute configuration is challenging, because both chromatographic and spectrometric separations of stereoisomers of fatty acids with a methyl group distant from the carboxyl group are difficult. To solve this problem, we synthesized the enantiomers via the Johnson-Claisen rearrangement to build (E)-4-alkenoic acid by using (R)- and (S)-3-methylpentanal as chiral blocks, which were readily available from the amino acids L-(+)-alloisoleucine and L-(+)-isoleucine, respectively. Each pure enantiomer, as well as the natural pheromone, was subsequently derivatized with a highly potent chiral labeling reagent used in the Ohrui-Akasaka method. Through NMR spectral comparisons of these derivatives, the absolute configuration of the natural pheromone was determined to be S. Field-trap bioassays showed that male mealybugs were attracted more to (S)-enantiomers and preferred the natural stereochemistry. Moreover, the synthetic pheromones attracted Anagyrus wasps, indicating that the azalea mealybug pheromone has kairomonal activity., (© 2024. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

Mealybug females release sex pheromones to attract conspecific males for mating. It is critical for mealybug males, which are fragile and short-lived, to respond to the pheromone of their species without time- and energy-consuming cross-attractions to other species. Thus, mealybug pheromone systems are considered to have evolved to be species-specific with unique structures in each species and offer an opportunity to study the diversity of pheromone chemistry that mediates intersexual courtship signals. More than 20 mealybug pheromones are reported to be monoterpenes in general, with only one exception, a hemiterpene alcohol esterified with a medium-chain fatty acid (MCFA), found in the Matsumoto mealybug, Crisicoccus matsumotoi. However, it is unknown whether this is truly exceptional, or if similar compounds are used in other related mealybugs. In this study, we isolated and characterized the pheromone of an allied species, the azalea mealybug C. azaleae. Using gas chromatography-mass spectrometry, nuclear magnetic resonance spectroscopy, and bioassays with synthetics, the pheromone was shown to be composed of isopropyl (E)-7-methyl-4-nonenoate, isopropyl (E)-7-methyl-4-octenoate, and ethyl (E)-7-methyl-4-nonenoate. Surprisingly, the structures of these compounds do not include hemiterpene nor monoterpene motifs but have methyl-branched MCFA parts that are similar to an acid moiety of the C. matsumotoi pheromone. This study implies irregular events for the divergence of pheromone structures in ancestors of the genus Crisicoccus and other mealybugs., (© 2022. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

Insect sex pheromones are examples of semiochemicals that trigger the most conspicuous biological activities, and they have attracted the interest of chemical ecologists since the dawn of this multidisciplinary field. For a deeper understanding of the ecological and evolutionary scenario of pheromones, as well as other targets of chemical ecology, it is essential to analyze the chemicals produced by individual organisms along with sound chemical identifications using reference compounds. Prof. Kenji Mori and his colleagues have developed various synthetic routes and have provided their products as authentic standards to many researchers. Using such a legacy, the tiny amounts of pheromones emitted by individual mealybug females were successfully analyzed and quantified by selected-ion-monitoring mode of gas chromatography-mass spectrometry. The results of the analyses of the monoterpene pheromones from Planococcus citri, P. minor, and their hybrids suggested that shift of the cyclobutane structure in P. citri and its acyclic form in P. minor is largely attributable to a single genetic locus., (© 2022. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

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 cotton mealybug, Phenacoccus solenopsis, the distribution of which was formerly limited to Nearctic and Neotropical regions, recently invaded many countries in various regions including Asia, Africa, and the Pacific. More recently, P. solenopsis was newly recorded in Japan and is currently an emerging pest of agricultural crops. In this study, we determined the structure of a sex pheromone of P. solenopsis in order to develop an effective lure for monitoring this pest. From volatiles emitted by virgin adult females, we isolated a compound attractive to males. By means of coupled gas chromatography-mass spectrometry and nuclear magnetic resonance spectroscopy, we identified this as (2,2-dimethyl-3-isopropylidenecyclobutyl)methyl 3-methylbut-2-enoate. This compound was synthesized and shown to be attractive to male P. solenopsis. Analysis by gas chromatography using an enantioselective stationary phase and polarimetry analyses of the natural pheromone and synthetic enantiomers showed the natural compound to be the (R)-(-)-enantiomer. This compound is an ester of maconelliol, which has an unusual cyclobutane structure found in sex pheromones of other mealybug species, and senecioic acid, also found in the pheromones of other mealybug species. However, this is the first example of the ester of maconelliol and senecioic acid as a natural product.

The grey pineapple mealybug, Dysmicoccus neobrevipes, is a serious pest that attacks a variety of crops in tropical regions. Recently, it was recorded on an island in southwestern Japan, suggesting that its distribution is expanding. As a measure against this expansion, a monitoring tool is urgently needed. In this study we determined the structure of the sex pheromone of D. neobrevipes in order to develop an efficient lure for monitoring traps. Volatiles collected from virgin adult females were fractionated by high performance liquid chromatography (HPLC) and gas chromatography, and fractions were tested for attractiveness to males in a laboratory bioassay. A single compound was isolated which was as attractive to males as the crude collections, and this was proposed to be the main, if not the only, component of the female-produced sex pheromone. The structure of this was determined to be (E)-2-isopropyl-5-methylhexa-3,5-dienyl acetate by gas chromatography/mass spectrometry and nuclear magnetic resonance analyses. This compound was synthesized through four steps, and the synthetic chemical was as attractive as the natural product in a greenhouse bioassay. The enantiomers of the synthetic acetate were obtained by enantioselective HPLC fractionation of the corresponding alcohols, and the natural pheromone was shown to be the (+)-isomer. The carbon skeleton of this novel compound is related to lavandulol, a monoterpene with an unusual non-head-to-tail connection of isoprene units that is often found in mealybug pheromones.

Females of the solitary endoparasitoid Cotesia vestalis respond to a blend of volatile organic compounds (VOCs) released from plants infested with larvae of their host, the diamondback moth (Plutella xylostella), which is an important pest insect of cruciferous plants. We investigated the flight response of female parasitoids to the cruciferous plant Brassica rapa, using two-choice tests under laboratory conditions. The parasitoids were more attracted to plants that had been infested for at least 6 hr by the host larvae compared to intact plants, but they did not distinguish between plants infested for only 3 hr and intact plants. Although parasitoids preferred plants 1 and 2 days after herbivory (formerly infested plants) over intact plants they also preferred plants that had been infested for 24 hr over formerly infested plants. This suggests that parasitoids can distinguish between the VOC profiles of currently and formerly infested plants. We screened for differences in VOC emissions among the treatments and found that levels of benzyl cyanide and dimethyl trisulfide significantly decreased after removal of the host larvae, whereas terpenoids and their related compounds continued to be released at high levels. Benzyl cyanide and dimethyl trisulfide attracted parasitoids in a dose-dependent manner, whereas the other compounds were not attractive. These results suggest that nitrile and sulfide compounds temporarily released from plants under attack by host larvae are potentially more effective attractants for this parasitoid than other VOCs that are continuously released by host-damaged plants.

We analyzed the sex pheromone of the pear fruit moth, Acrobasis pyrivorella, by means of gas chromatography-electroantennographic detection (GC-EAD) and GC-mass spectrometry. Two EAD-active compounds were detected in the pheromone gland extract of females. They were identified as (Z)-9-pentadecenyl acetate (Z9-15:OAc) and pentadecyl acetate (15:OAc). The amounts per female gland (mean +/- standard error) of these compounds were 12.9 +/- 2.8 and 0.8 +/- 0.1 ng, respectively. Synthetic Z9-15:OAc (300 microg) attracted conspecific males in field trapping experiments. When 15:OAc (21 microg; 7% of Z9-15:OAc quantity) was added, the number of males trapped increased significantly. Catch in traps baited with the mixture of these compounds was greater than that in traps baited with 1-3-day-old virgin females. We, therefore, conclude that Z9-15:OAc and 15:OAc are sex pheromone components of this species.

In addition to 2,13- and 3,13-octadecadien-1-ols and their acetates, aldehyde analogs have been identified from lepidopteran species in the family Sesiidae. To establish a reliable analytical method for determining the positions and configurations of the two double bonds in natural pheromone components, all geometric isomers of the 2,13- and 3,13-octadecadienals were synthesized by Dess-Martin oxidation of the corresponding alcohols with limited isomerization of the double bond at the 2- or 3-position. GC-MS analysis of these aldehydes showed isomerization of (Z)-2-, (Z)-3-, and (E)-3-double bonds to an (E)-2-double bond, even with a cool on-column injection. In contrast, HPLC analysis with an ODS column was accomplished without isomerization. The geometric isomers of each dienal eluted in the order ZZ --> EZ --> ZE --> EE. The conjugated 2,13-dienals were detectable in nanogram amounts with a UV detector at 235 nm. Whereas the detection of 3,13-dienals was difficult because of the lack of a chromophore, a highly sensitive analysis was achieved after derivatization with 2,4-dinitrophenylhydrazine. LC-MS with atmospheric pressure chemical ionization showed a strong [M-1](-) at m/z 443 for the derivatives. Based on these analytical data, a pheromone extract of a sesiid moth, Macroscelesia japona, was examined by HPLC and LC-MS, and it was confirmed that the octadecadienal tentatively identified by a previous GC-MS analysis did indeed have the 2E,13Z configuration. Furthermore, field evaluation of four synthetic geometric isomers of the 2,13-dienal revealed specific attraction to a lure with the (2E,13Z)-isomer as a main component.

Crossing experiments between two closely related moths, Ostrinia scapulalis and O. zealis, were conducted to gain insight into the genetic basis of the divergence of female sex pheromones. The sex pheromone of O. scapulalis comprises (E)-11- and (Z)-11-tetradecenyl acetates (E11 and Z11), and distinct genetic variation is found in the blend of components. This variation is largely controlled by a single autosomal locus with two alleles, AE(sca) and AZ(sca). E-type (AE(sca)AE(sca)) females produce a pheromone with a mean E11:Z11 ratio of 99:1, whereas Z-type (AZ(sca)AZ(sca)) and 1-type (AE(sca)AZ(sca)) females produce a pheromone with a mean of 3:97 and 64:36, respectively. O. zealis is distinctive in that it has a third pheromone component, (Z)-9-tetradecenyl acetate (Z9), in addition to E11 and Z11, and the typical blend ratio is 60:35:5 (Z9:E11:Z11). Our study revealed that Z9 production in O. zealis is mainly regulated by an autosomal recessive gene phr(zea), which is suggested to be involved in the chain-shortening of a pheromone precursor fatty acid, and linked to AE(zea), a gene corresponding to AE(sca) in O. scapulalis. A few mutations in a gene involved in pheromone production could explain the dramatic shift between a two-component pheromone communication system in O. scapulalis and a three-component system in O. zealis.

The adzuki bean borer, Ostrinia scapulalis, has distinct genetic variation in the blend of two sex pheromone components, (E)- and (Z)-11-tetradecenyl acetates. This variation is largely controlled by a single autosomal locus with two alleles, E and Z. E-type (EE) females produce a pheromone with a mean E:Z ratio at 99:1 whereas Z-type (ZZ) and I-type (ZE) produce pheromones with mean of 3:97 and 64:36, respectively. Interestingly, in many natural populations of O. scapulalis in Japan, this pheromone polymorphism appears to be stably maintained. We tried to predict the changes in relative abundance of each pheromone type by estimating the pheromone production genotype of wild females and their male mates. The pheromone titer in the wild, mated females was increased without changing the blend ratio by an injection of a peptide with pheromone biosynthesis activating activity (TKYFSPRL-NH2). The frequencies of E-, I-, and Z-types at Matsudo were 15, 52, and 33%, respectively, and did not deviate from the Hardy-Weinberg expectations. The estimated mating patterns were concordant with the assumption that no assortative mating was occurring in this population, and this is suggested as a cause of sustained polymorphism at Matsudo.