Junwei Zhu

Leaf Volatiles from Nonhost Deciduous Trees: Variation by Tree Species, Season and Temperature, and Electrophysiological Activity in Ips typographus

The leaf volatiles emitted from four nonhost tree species of Ips typographus, i.e. Betula pendula, B. pubescens, Populus tremula, and Sambucus nigra, were collected outdoors by headspace sampling in situ and analyzed by GC-MS. Three major classes of compounds, aliphatics [mainly green-leaf volatiles (GLVs)], monoterpenes, and sesquiterpenes, existed in all the deciduous tree species investigated. In June, when the bark beetles are searching in flight for host trees, GLVs mainly consisting of (Z)-3-hexenyl acetate and (Z)-3-hexen-1-ol were the dominant constituents in B. pendula and S. nigra. In B. pubescens and P. tremula, sesquiterpenes (and their derivatives) and monoterpenes made up the major part of whole volatile blends, respectively. Surprisingly, sesquiterpene alcohols and other oxides released from B. pubescens in considerable amounts were not found in the closely related species, B. pendula. By August, both the total volatiles and individual compounds significantly decreased, mainly due to the maturation of leaves, since the light intensity and temperatures during sampling were the same as in June. There were almost no volatiles detected from P. tremula and S. nigra leaves in August. The total emissions from these deciduous species were significantly different among the species, with B. pubescens releasing 5–10 times more than other species. Under the conditions of constant light intensity and humidity, emissions of both total volatiles and most individual components of severed B. pendula and S. nigra branches (with fresh leaves) increased according to a saturation curve from 16°C to 40°C. Ips typographus antennae responded strongly to green leaf alcohols: (Z)-3-hexen-1-ol, 1-hexanol, and (E)-2-hexen-1-ol, but not to aldehydes or acetates in GC-EAD analyses of B. pendula and B. pubescens leaf volatiles. No antennal responses to monoterpenes, sesquiterpenes, or sesquiterpene oxides were found. These three antennally active GLVs emitted from nonhost tree leaves might be indicators of a wrong habitat in the host selection of conifer bark beetles.

Reductase specificity and the ratio regulation of E/Z isomers in pheromone biosynthesis of the European corn borer, Ostrinia nubilalis (Lepidoptera: Pyralidae)

Species specificity of moth sex pheromones is in many cases achieved by means of specific blends rather than by specific components. Two pheromone strains of the European corn borer, Ostrinia nubilalis, use (E)- and (Z)-ll-tetradecenyl acetate in different ratios as their pheromone, but show the same ratio of the pheromone precursors (70:30 EIZ-11-tetradecenoic acid). The hypothesis that the ratio of the pheromone components in the two strains and their hybrids is controlled by the specificity of the reductase system, responsible for conversion of acid to the corresponding alcohol precursors, was tested. Deuterium-labeled alcohols, aldehydes and fatty acids corresponding to the two pheromone components were topically applied to the pheromone glands in different ratios and their selective incorporation into pheromone components was determined by gas chromatography with mass selective detection. Acetylation of the (E)- and (Z)-ll-tetradecenols was unselective, whereas the corresponding aldehydes and acids were selectively incorporated into the pheromone components. Z strain females selectively metabolized the Z-isomers whereas E strain females converted the E-isomers. The E strain being the most selective of the two strains. Hybrids converted both geometric isomers. The relative conversion rate of both E- and Z-isomers of all tetradecenoic acids with the double bond in positions from 7-12, was also determined. In addition to the All-isomers, the E strain females converted (E)-8-tetradecenoic acid into acetate and the Z strain females converted (E)-12-tetradecenoic acid. None of these substrates occur naturally in the pheromone gland, but (E)-12-tetradecenyl acetate is a pheromone component of the Asian corn borer O. furnacalis. Thus the possibility for conversion of (E)-12-tetradecenoic acid to acetate in the Z strain, as well as the earlier reported conversion of (Z)-ll-tetradecenoic acid to acetate in O. furnacalis, suggests that O. furnacalis is closest related to the Z strain of O. nubilalis. Pheromone biosynthesis Ratio regulation Reductase specificity Deuterium-labeled precursors Ostrinia nubilalis

Pheromone specificity in Eriocrania semipurpurella (Stephens) and E. sangii (Wood) (Lepidoptera: Eriocraniidae) based on chirality of semiochemicals

The fifth abdominal segment of femaleEriocrania semipurpurella (Stephens) andE. sangii (Wood) contains a pair of exocrine glands. Hexane extracts of this segment were prepared from both species and analyzed by gas chromatography with simultaneous flame ionization and electroantennographic detection (EAD). For both species, the EAD active peaks were identified as nonan-2-one, (Z)-6-nonen-2-one, and (Z)-6-nonen-2-ol by means of mass spectrometry and comparison of retention indices with those of synthetic standards. Enantiomeric separation of chiral alcohols from the female extracts was achieved by gas chromatographic analysis on a cyclodextrin column. InE. semipurpurella, a mixture of (2S,6Z)-nonen-2-ol and (2R,6Z)-nonen-2-ol (2: I) was found, whereas inE. sangii (2S,6Z)-nonen-2-ol was the predominant enantiomer and only traces of theR enantiomer were indicated by the antennal response. In field tests, a blend of the three compounds was not attractive to conspecific males. A subtractive assay showed that the alcohol in various enantiomeric mixtures was the only attractive compound, whereas addition of (Z)-6-nonen-2-one to the alcohol completely inhibited the attraction of both species. A trapping experiment including a wide range of ratios between theR andS enantiomers showed that baits containing 95–100% of theS enantiomer were attractive to maleE. sangii, whereas males ofE. semipurpurella were attracted to all tested ratios of the enantiomers. However, the response profiles of maleE. semipurpurella differed between populations from southern Sweden, south Finland, and the Kola Peninsula in Russia. In south Sweden males were maximally attracted to a racemic mixture of the alcohols. At the Kola PeninsulaE. semipurpurella was attracted to baits containing 95–100% of theR enantiomer. In south Finland all tested ratios between 0 and 100%R enantiomer trappedE. semipurpurella, but the trap catches appeared to be bimodally distributed with peaks around 15 and 70%R enantiomer. The trapping results suggest the existence of pheromone races or sibling species among the specimens identified asE. semipurpurella.

Identification of a novel moth sex pheromone inEriocrania cicatricella (Zett.) (Lepidoptera: Eriocraniidae) and its phylogenetic implications

Extracts from different body parts of adult femaleEriocrania cicatricella (Zett.) were tested for electrophysiological activity on conspecific male antennae. Extracts from the Vth abdominal segment, containing a pair of exocrine glands, elicited the largest electroantennographic response when compared to extracts of other body parts. Female extracts were analyzed by gas chromatography with simultaneous flame ionization and electroantennographic detection (EAD). The EAD active peaks were identified as (Z)-4-hepten-2-one, (2R)-heptane-2-ol, and (2R)-(Z)-4-hepten-2-ol by coinjection on a gas chromatography and by comparison of mass spectra with those of synthetic standards. In field tests, a blend of these three pheromone components was highly attractive to conspecific males, and a subtractive assay confirmed that the unsaturated alcohol is the major pheromone component, whereas no definite behavioral activity could be assigned to the ketone or the saturated alcohol. A bait containing the two alcohols withS-configuration was attractive to maleE. sparrmannella (Bosc), whereas no males ofE. cicatricella were found in these traps. The sex pheromone compounds inE. cicatricella are chemically similar to pheromones reported in Trichoptera and they are produced in homologous glands.

Identification of the Sex Pheromone of the Currant Shoot Borer Lampronia capitella

Under an artificial light:dark cycle, females of Lampronia capitella were observed calling, with extended terminal abdominal segments, during the first 2 hr of the photoperiod. Extracts of terminal abdominal segments from females elicited large electroantennographic responses from male antennae. Gas chromatography with electroantennographicDetection revealed three active peaks. Based on comparison of retention times and mass spectra of synthetic standards, these compounds were identified as (Z,Z)-9,11-tetradecadienol and the corresponding acetate and aldehyde. The electroantennographic activity of the four geometric isomers of all three compounds was investigated, and the respective (Z,Z)-isomer was found to be the most active in all cases. Aldehydes generally elicited larger antennal responses than alcohols, whereas acetates were the least active compounds. A subtractive trapping assay in the field, based on a 13:26:100 μg mixture of (Z,Z)-9,11-tetradecadienal, (Z,Z)-9,11-tetradecadienyl acetate, and (Z,Z)-9,11-tetradecadienol confirmed that all three compounds are pheromone components. Subtraction of (Z,Z)-9,11-tetradec- adienol from the blend completely eliminated its attractiveness, whereas the other two-component blends showed reduced activity. This is the first pheromone identification from the monotrysian superfamily Incurvarioidea, confirming that the common pheromones among ditrysian moths (long-chain fatty acidDerivatives comprising alcohols, acetates, and aldehydes with one or more double bonds) is not an autapomorphy of Ditrysia, but a synapomorphy of the more advanced heteroneuran lineages.

SPERMATOPHORE SIZE AND MULTIPLE MATING: EFFECTS ON REPRODUCTIVE SUCCESS AND POST-MATING BEHAVIOUR IN THE INDIAN MEAL MOTH

Ensuring fatherhood is an important issue in polygamous species. In many lepidopteran species, the male inserts a large package (spermatophore) consisting of sperm, accessory gland fluids and nutrients into the females bursa copulatrix during copulation. In most species it has been shown that a large male donation causes females either to stop calling, or to prolong the time until re-mating, hereby ensuring fatherhood. We investigated the changes in size of the donation in multiple mating and how the size affected the female post mating behaviour in the Indian meal moth, Plodia interpunctella. In concordance with other previous studies, we found that a male inserted a significantly larger donation during the first mating compared to following mating occasions. The larger donation was not coupled with larval output since it did not decline during mating occasions. Some males were able to mate at least eight times and male fitness, counted as larval output, showed no association with sequence of mating number. Even though females received larger donations during the first mating, there was no effect on female post mating behaviour in terms of pheromone production and calling behaviour. The weight of the mated females bursa copulatrix decreased linearly over the first eight days after mating, suggesting that females absorb material other than sperm. We discuss the female advantage of receiving a large donation and why males invest more energy into the first mating while female post-mating behaviour is not affected.

A comparative study of sex pheromone biosynthesis in two strains of the turnip moth, Agrotis segetum, producing different ratios of sex pheromone components

Abstract Among the sex pheromone components of the turnip moth, Agrotis segetum, (Z)-5-decenyl acetate, (Z)-7-dodecenyl acetate and (Z)-9-tetradecenyl acetate are biosynthetically derived from palmitic acid by Δ11-desaturation, chain-shortening, reduction and acetylation. Females of a Zimbabwean population produce the three components in a 78:20:2 ratio, while Swedish females produce the three components in a 12:59:29 ratio. We found that the titers of primary pheromone precursors, such as 16:Acyl and Z11-16:Acyl, did not differ significantly between the two populations. However, the amounts of intermediate precursors Z5-10:Acyl, Z7-12:Acyl and Z9-14:Acyl were significantly higher in the Swedish female extracts. There was no obvious correlation between the ratios of pheromone precursors and the ratios of pheromone components. By application of D3-16:COOH, D9-Z11-16:COOH, D9-Z9-14:COOH and D9-Z7-12:COOH to the female pheromone glands, we found that Zimbabwean females produced more labelled D9-Z5-10:OAc than Swedish females. In contrast, in Swedish females, the labelled precursors were mainly converted to D9-Z9-14:OAc and D9-Z7-12:OAc, rather than to D9-Z5-10:OAc. Moreover, the conversion rate of D9-Z5-10:COOH to D9-Z5-10:OAc, was significantly higher in Zimbabwean females than in Swedish females. Our results indicate that differences in chain shortening and/or the preferential reduction of acids with different chain lengths may lead to the production of different ratios. Brain-SOG homogenates from the two populations increased the pheromone production of decapitated females of both populations, but did not change the pheromone ratios of the two populations.

A sex pheromone component novel to the Geometridae identified from Epirrita autumnata.

(Z,Z,Z)‐1, 3, 6, 9‐Heneicosatetraene was identified as a sex pheromone component of Epirrita autumnata (Borkhausen), (Lepidoptera: Geometridae). Gas chromatography coupled with electroantennographic detection revealed one active peak‐from female pheromone gland extracts. The chemical identification of the compound was based on gas chromatography‐mass spectrometry, chemical micro‐reactions and unambiguous synthesis. In a field test, a rubber septum loaded with 3000 μg of this compound showed highest attraction of male E. autumnata moths compared to lower doses and the control. The attraction of males to a 300 μg bait was equivalent to that obtained using live virgin females.

Identification of polyenic hydrocarbons from the northern winter moth, Operophtera fagata , and development of a species specific lure for pheromone traps

Summary.In order to elucidate the composition of the female sex pheromone of the northern (beech) winter moth, Operophtera fagata Scharf. (Lepidoptera: Geometridae), ovipositor extracts of unmated, calling females were analysed by gas chromatography with simultaneous electroantennographic and flame ionization detection (GC-EAD/FID). Male antennal responses indicated three active components, two of which had distinct matching peaks in the FID trace. Using coupled gas chromatography- mass spectrometry (GC-MS), these two compounds were identified as (9Z)-nonadecene (9Z-19:Hy), and (6Z,9Z)-nonadecadiene (6Z9Z-19:Hy), respectively. The third component, present in very small amounts only, was identified as (1,3Z,6Z,9Z)-nonadecatetraene (1,3Z6Z9Z-19:Hy), known as the sex pheromone of the common winter moth, O. brumata. Field tests revealed that traps baited with 6Z9Z-19:Hy and 1,3Z6Z9Z-19:Hy caugth large numbers of male O. fagata. Both compounds were found to be essential for attraction of O. fagata. In addition, the diene prevented captures of co-occurring O. brumata. In contrast, 9Z-19:Hy neither influenced the attractiveness of the two-component mixture towards O. fagata nor contributed to bait specificity. A binary mixture of 6Z9Z-19:Hy and 1,3Z6Z9Z-19:Hy in a ratio of 10:1, applied to pieces of rubber tubing, constituted a highly attractive and species-specific bait for O. fagata, which can be used for monitoring of the flight of this defoliator pest of deciduous forests.

Olefinic acetates, Delta-9, 11-14:OAc and Delta-7,9-12:OAc used as sex pheromone components in three geometrid moths, Idaea aversata, I-straminata, and I-biselata (Geometridae, Lepidoptera)

Pheromone compounds so far identified from most geometrid moths consist of all-Z diene, triene, or tetraene hydrocarbons with chain lengths of C17 to C21, and their monoepoxide derivatives biosynthesized from linoleic and linolenic acids. The present study reports the occurrence of olefinic acetates as sex pheromones in three species of Geometridae. (Z,Z)-9,11-tetradecadienyl acetate and (Z,Z)-7,9-dodecadienyl acetate found in female gland extracts ofIdaea aversata elicited significant responses from conspecific male antennae in gas chromatography with electroantennographic detection (GCEAD). In extracts ofI. straminata, (Z,Z)-7,9-dodecadienyl acetate, (E,Z)-7,9-dodecadienyl acetate, and (Z,Z)-7,9-dodecadienyl acetate were found, and the synthetic compounds elicited strong responses from conspecific male antennae. In the third species,I. biselata, only (Z,Z)-7,9-dodecadienyl acetate was found in the female extracts, and this compound elicited a strong EAD response from the conspecific male antenna. The identities of the pheromone components inI. aversata andI. straminata were further confirmed according to their characteristic ions after GC-MS analyses. Single sensillum recordings fromI. aversata showed two types of pheromone-detecting sensilla present on the male antenna. One type contained two receptor neurons, one of which was specifically tuned to (Z,Z)-9,11-tetradecadienyl acetate, the other to (Z,E)-9,11-tetradecadienyl acetate. A second type contained one neuron responding to (Z,Z)-7,9-dodecadienyl acetate. The two types were clearly different also with respect to external morphology, the former being considerably longer and having a larger base diameter. Also inI. straminata two physiological types of sensilla could be distinguished. One type contained two neurons, one of which responded to (Z,Z)-7,9-dodecadienyl acetate, the other to (Z,E)-9,11-tetradecadienyl acetate. The second type contained one neuron, responding to (Z,Z)-7,9-dodecadienyl acetate. No correlation between external morphology and physiological response of the investigated sensilla was observed inI. straminata. In field tests, a two-component blend containing (Z,Z)-9,11-tetradecadienyl acetate and (Z,Z)-7,9-dodecadienyl acetate in a ratio of 10:1 was attractive to males ofI. aversata. This two-component blend was also attractive to males ofI. straminata, but in a ratio of 1:1. High numbers of maleI. biselata were caught in traps baited with (Z,Z)-7,9-dodecadienyl acetate alone. The incorporation of deuterium labels into pheromone components after topical application of deuterium-labeled palmitic acid confirmed that the pheromone components ofI. aversata could be synthesized from this precursor, as has been previously observed for acetate pheromone components of many other moth species. Our results suggest that an evolutionary reversal back to the production of palmitic acid-derived pheromone components has occurred within the geometrid subfamily Sterrhinae.