Sex Pheromone of the Saturniid Moth Hemileuca nevadensis from Southern California.

The major and possibly only component of the sex attractant pheromone of the moth Hemileuca nevadensis (Lepidoptera: Saturniidae) from southern California was determined to be (E10,Z12)-hexadecadienal (E10,Z12-16:Ald). Detectable quantities of the analogs (E10,Z12)-hexadecadien-1-yl acetate (E10,Z12-16:Ac) and (E10,Z12)-hexadecadien-1-ol (E10,Z12-16:OH) were also present in solvent extracts of sex pheromone glands, and stimulated male antennae in coupled gas chromatography-electroantennogram detector (GC-EAD) assays. GC-EAD traces from solid phase microextraction (SPME) wipe samples of sex pheromone glands of calling females confirmed the presence of E10,Z12-16:Ald and traces of E10,Z12-16:OH on the gland surface, but E10,Z12-16:Ac was not detected. Despite evidence for the presence of all three compounds in extracts, behavioral responses to synthetic compounds in the field suggested that only E10,Z12-16:Ald is required for optimal attraction.

Phylogeography of an endemic California silkmoth genus suggests the importance of an unheralded central California province in generating regional endemic biodiversity.

The California Floristic province is a biodiversity hotspot. Understanding the phylogeographic patterns that exist in this diverse region is essential to understanding its evolution and for guiding conservation efforts. Calosaturnia is a charismatic silkmoth genus endemic to large portions of the region with three described species, C. mendocino, C. walterorum, and C. albofasciata. We sampled all three species from across their ranges, sequenced 1463 bp of mitochondrial COI and 1941 bp of nuclear DNA from three genes, and reconstructed phylogenetic relationships and estimated divergence times within the lineages. All three species show pronounced evidence of isolation and, in two cases, secondary reconnection. An unexpected monophyletic mtDNA lineage was found in the Central Coast region, in a region thought to represent an intergrade between C. mendocino and C. walterorum. Our genetic data also significantly revise previous hypotheses as to which species occur in which regions, suggesting that historical ecological changes around four Ma ago isolated some lineages, and a secondary isolation event two Ma ago led to isolation of populations both in the Central Coast region and the southern Sierra Nevada. Our results add to a currently under-appreciated pattern suggesting that coastal Central California is not a transition zone between Northern and Southern California Floristic Province faunas but rather its own unique, periodically isolated, biogeographic region. They also suggest cryptic diversity may be present in many other currently unrecognized groups. Additional research should focus on this central California region because many species may be highly restricted in range and in need of conservation attention.

Deceptive signals and behaviors of a cleptoparasitic beetle show local adaptation to different host bee species.

Chemosensory signals play a key role in species recognition and mate location in both invertebrate and vertebrate species. Closely related species often produce similar but distinct signals by varying the ratios or components in pheromone blends to avoid interference in their communication channels and minimize cross-attraction among congeners. However, exploitation of reproductive signals by predators and parasites also may provide strong selective pressure on signal phenotypes. For example, bolas spiders mimic the pheromones of several moth species to attract their prey, and parasitic blister beetle larvae, known as triungulins, cooperatively produce an olfactory signal that mimics the sex pheromone of their female host bees to attract male bees, as the first step in being transported by their hosts to their nests. In both cases, there is strong selection pressure on the host to discriminate real mates from aggressive mimics and, conversely, on the predator, parasite, or parasitoid to track and locally adapt to the evolving signals of its hosts. Here we show local adaptation of a beetle, Meloe franciscanus (Coleoptera: Meloidae), to the pheromone chemistry and mate location behavior of its hosts, two species of solitary bees in the genus Habropoda We report that Mfranciscanus' deceptive signal is locally host-adapted in its chemical composition and ratio of components, with host bees from each allopatric population preferring the deceptive signals of their sympatric parasite population. Furthermore, in different locales, the triungulin aggregations have adapted their perching height to the height at which local male bees typically patrol for females.

Identification of Aggregation-Sex Pheromone Components for a "Living Fossil", the False Click Beetle, Palaeoxenus dohrni Horn (Coleoptera: Eucnemidae).

Insect pheromones have rarely been exploited in surveys or studies of rare and endangered species, despite their potential as effective and highly selective attractants for target species. Here, we report the identification, synthesis, and field bioassays of a male-produced aggregation-sex pheromone blend of a rare false click beetle species endemic to southern California, Dohrn's elegant eucnemid beetle, Palaeoxenus dohrni Horn (Coleoptera: Eucnemidae). This species is the only extant species in its genus and subfamily. Analyses of extracts of headspace volatiles collected from adult beetles revealed several male-specific compounds. Two of these compounds, identified as (E)-2-nonen-4-one and (R)-2-nonanol, elicited electroantennographic responses from antennae of beetles of both sexes. In field bioassays, a blend of the two compounds attracted both sexes, whereas the individual compounds were not attractive. The identification of an attractant pheromone should provide a useful tool for bioconservation and ecological studies of this iconic species.

Effects of Pheromone Dose and Conspecific Density on the Use of Aggregation-Sex Pheromones by the Longhorn Beetle Phymatodes grandis and Sympatric Species (Coleoptera: Cerambycidae).

Many species of longhorn beetles (Coleoptera: Cerambycidae) utilize male-produced aggregation-sex pheromones that attract both sexes. However, the reasons why and the details of how this type of pheromone is used by cerambycids and other coleopteran species that utilize analogous male-produced pheromones remain unclear. Thus, our goals were to test the hypotheses that 1) cerambycids respond to pheromones in a dose-dependent (= release rate-dependent) manner and 2) pheromone emission is density-dependent. If true, these characteristics of pheromone use could suggest that cerambycids utilize an optimal density strategy to limit competition for scarce and ephemeral hosts, i.e., the stressed or dying trees that typically constitute their larval hosts. Attraction of beetles to a range of release rates of two common pheromone components - 2-methylbutanol and 3-hydroxyhexan-2-one - was tested in field trials. Responses, as measured by the number of beetles caught in pheromone-baited traps, increased with release rates for five endemic species, even at the highest rates tested (~1450 µg/h for 2-methylbutanol and ~720 µg/h for 3-hydroxyhexan-2-one). The effect of density of conspecific males on per capita pheromone production was tested by collecting the volatiles produced by individuals, pairs, or groups of three or four male Phymatodes grandis Casey. Frequency of pheromone production was significantly different among the treatment densities, and emission rates of the pheromone (R)-2-methylbutanol decreased with increasing density. These results are discussed in the context of a possible optimal density strategy used by cerambycids, and more broadly, in relation to the use of male-produced aggregation-sex pheromones by other coleopterans. In addition, we report the identification of the pheromones of four of our five test species, specifically, Phymatodes obliquus Casey, Brothylus conspersus LeConte, Brothylus gemmulatus LeConte, and Xylotrechus albonotatus Casey.

An Unstable Monoterpene Alcohol as a Pheromone Component of the Longhorned Beetle Paranoplium gracile (Coleoptera: Cerambycidae).

We report the identification of p-mentha-1,3-dien-8-ol, an unstable monoterpene alcohol, as a male-produced aggregation-sex pheromone component of the cerambycid beetle Paranoplium gracile (Leconte) (subfamily Cerambycinae, tribe Oemini), a species endemic to California. Headspace volatiles from live males contained a blend of nine cyclic terpenoids that were not detected in analogous samples from females. Volatiles produced by male Eudistenia costipennis Fall, also in the tribe Oemini, contained the same suite of nine compounds. Four compounds, dehydro-p-cymene, p-mentha-1,3-dien-8-ol, p,α,α-trimethylbenzyl alcohol, and an unidentified compound were found to elicit responses from antennae of P. gracile females in coupled gas chromatography-electroantennogram detection (GC-EAD) assays, whereas only p-mentha-1,3-dien-8-ol elicited responses from antennae of males. In field assays, p-mentha-1,3-dien-8-ol stabilized with the antioxidant butylated hydroxytoluene (BHT) attracted P. gracile of both sexes, indicating it functions as an aggregation-sex pheromone, as with other pheromones identified from its subfamily, the Cerambycinae. Adding four of the other compounds found in headspace samples to the dienol lure had no effect on attraction. Because of the instability of p-mentha-1,3-dien-8-ol, it seems likely that at least some of the compounds seen in the extracts of volatiles from both species are artefacts, rather than being components of the pheromone.

Sex Attractant Pheromones of Virgin Queens of Sympatric Slave-Making Ant Species in the Genus Polyergus, and their Possible Roles in Reproductive Isolation.

Species of the ant genus Polyergus are social parasites that steal brood from colonies of their hosts in the closely related genus Formica. Upon emergence as adults in a mixed population, host Formica workers carry out all the normal worker functions within the Polyergus colony, including foraging, feeding, grooming, and rearing brood of the parasitic Polyergus ants. Some unmated Polyergus gynes (queens) run in the raiding columns of their colonies and attract males by releasing a pheromone from their mandibular glands. There are two Polyergus species groups in North America: an eastern P. lucidus group and a western P. breviceps group. One species of each of these groups, P. lucidus Mayr and P. mexicanus Emery, are sympatric in Missouri. In this study, we characterized the sex pheromones of virgin queens of two species of the P. lucidus group (P. lucidus sensu stricto and P. sanwaldi) and one species of the P. breviceps group (P. mexicanus), and compared these with the previously identified sex pheromone of P. topoffi of the P. breviceps group. We then used sex pheromone blends reconstructed from synthesized components of the two groups to test their efficacy at reproductively isolating these species. We found that methyl 6-methylsalicylate is conserved as the major component of the pheromone blends for both Polyergus species groups; however, methyl (R)-3-ethyl-4-methylpentanoate is the species-specific minor component produced by P. lucidus group queens, and (R)-3-ethyl-4-methylpentan-1-ol is the crucial minor component for P. breviceps group queens. The optimal ratio of the major and minor components for P. lucidus group queens was about 100:1 salicylate to ester. In concurrent field trials in Missouri, males of P. lucidus sensu stricto and P. mexicanus (a member of the P. breviceps group) were attracted almost exclusively to their particular blends of sex pheromone components. To our knowledge, this is the first example of a possible sex-pheromone-based reproductive isolating mechanism in ants.

Isolation and determination of absolute configurations of insect-produced methyl-branched hydrocarbons.

Although the effects of stereochemistry have been studied extensively for volatile insect pheromones, little is known about the effects of chirality in the nonvolatile methyl-branched hydrocarbons (MBCHs) used by many insects as contact pheromones. MBCHs generally contain one or more chiral centers and so two or more stereoisomeric forms are possible for each structure. However, it is not known whether insects biosynthesize these molecules in high stereoisomeric purity, nor is it known whether insects can distinguish the different stereoisomeric forms of MBCHs. This knowledge gap is due in part to the lack of methods for isolating individual MBCHs from the complex cuticular hydrocarbon (CHC) blends of insects, as well as the difficulty in determining the absolute configurations of the isolated MBCHs. To address these deficiencies, we report a straightforward method for the isolation of individual cuticular hydrocarbons from the complex CHC blend. The method was used to isolate 36 pure MBCHs from 20 species in nine insect orders. The absolute stereochemistries of the purified MBCHs then were determined by digital polarimetry. The absolute configurations of all of the isolated MBCHs were determined to be (R) by comparison with a library of synthesized, enantiomerically pure standards, suggesting that the biosynthetic pathways used to construct MBCHs are highly conserved within the Insecta. The development of a straightforward method for isolation of specific CHCs will enable determination of their functional roles by providing pure compounds for bioassays.

The Influence of Host Plant Volatiles on the Attraction of Longhorn Beetles to Pheromones.

Host plant volatiles have been shown to strongly synergize the attraction of some longhorn beetle species (Coleoptera: Cerambycidae) to their pheromones. This synergism is well documented among species that infest conifers, but less so for angiosperm-infesting species. To explore the extent of this phenomenon in the Cerambycidae, we first tested the responses of a cerambycid community to a generic pheromone blend in the presence or absence of chipped material from host plants as a source of host volatiles. In the second phase, blends of oak and conifer volatiles were reconstructed, and tested at low, medium, and high release rates with the pheromone blend. For conifer-infesting species in the subfamilies Spondylidinae and Lamiinae, conifer volatiles released at the high rate synergized attraction of some species to the pheromone blend. When comparing high-release rate conifer blend with high-release rate α-pinene as a single component, species responses varied, with Asemum nitidum LeConte being most attracted to pheromones plus α-pinene, whereas Neospondylis upiformis (Mannerheim) were most attracted to pheromones plus conifer blend and ethanol. For oak-infesting species in the subfamily Cerambycinae, with the exception of Phymatodes grandis Casey, which were most attracted to pheromones plus ethanol, neither synthetic oak blend nor ethanol increased attraction to pheromones. The results indicate that the responses to combinations of pheromones with host plant volatiles varied from synergistic to antagonistic, depending on beetle species. Release rates of host plant volatiles also were important, with some high release rates being antagonistic for oak-infesting species, but acting synergistically for conifer-infesting species.

Sex Attractant Pheromone of the Luna Moth, Actias luna (Linnaeus).

Giant silk moths (Lepidoptera: Saturniidae) typically are not well represented as larvae or adults in community level inventories of Lepidoptera, and as a result, little is known about their population dynamics. Furthermore, in recent years, many species of silk moths appear to have experienced population declines. Volatile sex pheromones are powerful sampling tools that can be used in operational conservation and monitoring programs for insects. Here, we describe the identification of the sex attractant pheromone of a giant silk moth, the luna moth Actias luna. Coupled gas chromatography-electroantennographic detection and gas chromatography-mass spectrometric analyses of extracts from pheromone glands of female luna moths supported the identification of (6E,11Z)-6,11-octadecadienal (E6,Z11-18:Ald), (6E)-6-octadecenal (E6-18:Ald), and (11Z)-11-octadecenal (Z11-18:Ald) as the compounds in extracts that elicited responses from antennae of male moths. These identifications were confirmed by synthesis, followed by testing of blends of the synthetic compounds in field trials in Ontario, Canada, and Kentucky, USA. Male moths were attracted to synthetic E6,Z11-18:Ald as a single component. Attraction appeared to be enhanced by addition of E6-18:Ald but not Z11-18:Ald, suggesting that the luna moth pheromone consists of a blend of E6,Z11-18:Ald and E6-18:Ald.

North American Species of Cerambycid Beetles in the Genus Neoclytus Share a Common Hydroxyhexanone-Hexanediol Pheromone Structural Motif.

Many species of cerambycid beetles in the subfamily Cerambycinae are known to use male-produced pheromones composed of one or a few components such as 3-hydroxyalkan-2-ones and the related 2,3-alkanediols. Here, we show that this pheromone structure is characteristic of the cerambycine genus Neoclytus Thomson, based on laboratory and field studies of 10 species and subspecies. Males of seven taxa produced pheromones composed of (R)-3-hydroxyhexan-2-one as a single component, and the synthetic pheromone attracted adults of both sexes in field bioassays, including the eastern North American taxa Neoclytus caprea (Say), Neoclytus mucronatus mucronatus (F.), and Neoclytus scutellaris (Olivier), and the western taxa Neoclytus conjunctus (LeConte), Neoclytus irroratus (LeConte), and Neoclytus modestus modestus Fall. Males of the eastern Neoclytus acuminatus acuminatus (F.) and the western Neoclytus tenuiscriptus Fall produced (2S,3S)-2,3-hexanediol as their dominant or sole pheromone component. Preliminary data also revealed that males of the western Neoclytus balteatus LeConte produced a blend of (R)-3-hydroxyhexan-2-one and (2S,3S)-2,3-hexanediol but also (2S,3S)-2,3-octanediol as a minor component. The fact that the hydroxyketone-hexanediol structural motif is consistent among these North American species provides further evidence of the high degree of conservation of pheromone structures among species in the subfamily Cerambycinae.

A tetraene aldehyde as the major sex pheromone component of the promethea moth (Callosamia promethea (Drury)).

The promethea moth Callosamia promethea is one of three species of silkmoths from the genus Callosamia that occur in North America. Cross attraction of males to heterospecific calling females has been observed in the field, and hybrid progeny have been produced by pairing heterospecifics in captivity. These observations suggest that all three species share or have considerable overlap in the sex attractant pheromones produced by females, so that other prezygotic isolating mechanisms, such as diel differences in reproductive activity, limit hybridization in the field. Coupled gas chromatography-electroantennogram detection and gas chromatography- mass-spectrometry analyses of extracts of volatiles collected from female promethea moths supported the identification of (4E,6E,11Z,13Z)-hexadeca-4,6,11,13-tetraenal [(4E,6E,11Z,13Z)-16:Ald] as the compound in extracts that elicited the largest responses from antennae of males. The identification was confirmed by non-selective synthesis of several isomers as analytical standards, and stereoselective synthesis of (4E,6E,11Z,13Z)-16:Ald for testing in field trials. Male moths were strongly attracted to synthetic (4E,6E,11Z,13Z)-16:Ald, suggesting that this compound is the major and possibly the only component of the sex pheromone of these large saturniid moths. Based on the cross-attraction of heterospecifics, it is likely that this is also a major pheromone component of the other two North American Callosamia species as well.

(R)-desmolactone, a female-produced sex pheromone component of the cerambycid beetle Desmocerus californicus californicus (subfamily Lepturinae).

We report the identification, synthesis, and field bioassays of a female-produced sex attractant pheromone for the cerambycid beetle Desmocerus californicus californicus Horn. Headspace volatiles from females contained a sex-specific compound, (R)-desmolactone [(4R,9Z)-hexadec-9-en-4-olide], which elicited strong responses from the antennae of adult males in coupled gas chromatography-electroantennogram analyses. Short syntheses of both enantiomers were developed from commercial chiral synthons. In field bioassays, significant numbers of males were collected in traps baited with (R)-desmolactone, whereas the (S)-enantiomer attracted no males. The racemate was less attractive than the pure (R)-enantiomer, indicating some degree of antagonism by the unnatural enantiomer. This compound is the first example of a new structural class of cerambycid pheromones, and is the second pheromone identified for a species in the subfamily Lepturinae.

2,3-Hexanediols as sex attractants and a female-produced sex pheromone for cerambycid beetles in the prionine genus Tragosoma.

Recent work suggests that closely related cerambycid species often share pheromone components, or even produce pheromone blends of identical composition. However, little is known of the pheromones of species in the subfamily Prioninae. During field bioassays in California, males of three species in the prionine genus Tragosoma were attracted to 2,3-hexanediols, common components of male-produced aggregation pheromones of beetles in the subfamily Cerambycinae. We report here that the female-produced sex pheromone of Tragosoma depsarium "sp. nov. Laplante" is (2R,3R)-2,3-hexanediol, and provide evidence from field bioassays and electroantennography that the female-produced pheromone of both Tragosoma pilosicorne Casey and T. depsarium "harrisi" LeConte may be (2S,3R)-2,3-hexanediol. Sexual dimorphism in the sculpting of the prothorax suggests that the pheromone glands are located in the prothorax of females. This is the second sex attractant pheromone structure identified from the subfamily Prioninae, and our results provide further evidence of pheromonal parsimony within the Cerambycidae, in this case extending across both subfamily and gender lines.

Identification of the sex pheromone of the invasive scale Acutaspis albopicta (Hemiptera: Diaspididae), arriving in California on shipments of avocados from Mexico.

As a result of relaxation of importation restrictions ordered by the Animal and Plant Health Inspection Service of the U.S. Department of Agriculture, shipments of fresh avocados from Mexico began entering California year-round in 2007, despite the fact that these shipments were heavily infested with a number of exotic and potentially invasive armored scale species that are not thought to be present in California. Here, we report the identification of the sex pheromone of one of these species, Acutaspis albopicta (Cockerell), from a quarantine colony of these insects initiated from specimens collected from commercial shipments of Mexican avocados. The compound was identified as [(1S,3S)-2,2-dimethyl-3-(prop-1-en-2-yl)cyclobutyl)]methyl (R)-2-methylbutanoate, and was similar in structure to the pheromones of several other scale and mealybug species. In laboratory bioassays, the pheromone was highly attractive to male scales in microgram doses. The pheromone will provide a very sensitive and selective tool for detection of the scale to try and prevent its permanent establishment in California.

cis-Vaccenyl acetate, a female-produced sex pheromone component of Ortholeptura valida, a longhorned beetle in the subfamily Lepturinae.

We report the identification, synthesis, and field bioassays of a female-produced sex attractant pheromone component of the cerambycid beetle Ortholeptura valida (LeConte). Headspace volatiles from females contained a female-specific compound, (Z)-11-octadecen-1-yl acetate, which elicited a strong response from antennae of adult males in coupled gas chromatography-electroantennogram analyses. In field bioassays, significant numbers of males were collected by traps baited with this compound. The pheromone represents a new structural class of cerambycid pheromones, and is the first pheromone identified for a cerambycid species in the subfamily Lepturinae.

Determination of the relative and absolute configurations of the female-produced sex pheromone of the cerambycid beetle Prionus californicus.

We previously identified the basic structure of the female-produced sex attractant pheromone of the cerambycid beetle, Prionus californicus Motschulsky (Cerambycidae: Prioninae), as 3,5-dimethyldodecanoic acid. A synthesized mixture of the four stereoisomers of 3,5-dimethyldodecanoic acid was highly attractive to male beetles. Here, we describe stereoselective syntheses of three of the four possible stereoisomers, and the results of laboratory and field bioassays showing that male beetles are attracted specifically to (3R,5S)-3,5-dimethyldodecanoic acid, but not to its enantiomer, (3S,5R)-3,5-dimethyldodecanoic acid, indicating that the (3R,5S)-enantiomer is the active pheromone component. The diastereomeric (3R,5R)- and (3S,5S)-enantiomers were excluded from consideration because their gas chromatographic retention times were different from that of the insect-produced compound. The mixture of the four stereoisomers of 3,5-dimethyldodecanoic acid was as attractive to male P. californicus as the (3R,5S)-enantiomer, indicating that none of the other three stereoisomers inhibited responses to the active enantiomer. Beetles responded to as little as 10 ng and 10 µg of synthetic 3,5-dimethyldodecanoic acid in laboratory and field studies, respectively. Field studies indicated that capture rate did not increase with dosages of 3,5-dimethyldodecanoic acid greater than 100 µg. In field bioassays, males of a congeneric species, P. lecontei Lameere, were captured in southern California but not in Idaho.

Identification of the sex pheromone of a protected species, the Spanish moon moth Graellsia isabellae.

Sex attractant pheromones are highly sensitive and selective tools for detecting and monitoring populations of insects, yet there has been only one reported case of pheromones being used to monitor protected species. Here, we report the identification and synthesis of the sex pheromone of a protected European moth species, Graellsia isabellae (Lepidoptera: Saturniidae), as the single component, (4E,6E,11Z)-hexadecatrienal. In preliminary field trials, lures loaded with this compound attracted male moths from populations of this species at a number of widely separated field sites in France, Switzerland, and Spain, clearly demonstrating the utility of pheromones in sampling potentially endangered insect species.

Field trials of aggregation pheromones for the stink bugs Chlorochroa uhleri and Chlorochroa sayi (Hemiptera: Pentatomidae).

In field trials, adult Chlorochroa uhleri (Stål) (Heteroptera: Pentatomidae) of both sexes were caught in significant numbers in cylindrical screen traps baited with gray rubber septum lures loaded with the main component of the male-produced pheromone, methyl (E)-6-2,3-dihydrofarnesoate. Addition of the two possible minor components of the pheromone, methyl (E)-5-2,6,10-trimethyl-5,9-undecadienoate and methyl (2E,6E)-farnesoate, did not affect attraction. Combining the pheromone with different concentrations of volatiles mimicking the odors of a known host plant, alfalfa (Medicago sativa L.), had no significant effect on attraction of adult bugs, whereas combining the pheromone with the pheromones of two sympatric stink bug species, Chlorochroa sayi (Stål) and Euschistus conspersus Uhler, decreased trap captures, suggesting interference between the pheromones. Small numbers of Chlorochroa ligata (Say) adults also were attracted, but numbers caught were too low to allow statistical comparisons between lure blends. In field trials with C. sayi, all three of the male-specific pheromone compounds [methyl geranate, methyl citronellate, and methyl (E) -6-2,3-dihydrofarnesoate] were required for optimal attraction. As with C. uhleri, adults of both sexes were attracted to pheromone lures in approximately equal numbers. Because of the decreased volatility (=release rate) of methyl (E)-6-2,3-dihydrofarnesoate in comparison with the other two, lower molecular weight pheromone components, lures needed to be loaded with a disproportionately high amount of methyl (E)-6-2,3-dihydrofarnesoate to obtain the best trap catch. There was no indication that the pheromone components of C. uhleri or E. conspersus interfered with the attractiveness of the C. sayi pheromone in lures containing a blend of all three pheromones.

Identification of critical secondary components of the sex pheromone of the navel orangeworm (Lepidoptera: Pyralidae).

We identified a four-component sex pheromone blend that is as attractive or more attractive to male navel orangeworm moths, Amyelois transitella (Walker) (Lepidoptera: Pyralidae), than either unmated females or hexane extracts of pheromone glands of females. The blend consisted of (11Z,13Z)-hexadecadienal; (11Z,13Z)-hexadecadien-1-ol; (11Z,13E)-hexadecadien-1-ol; and a hydrocarbon, (3Z,6Z,9Z,12Z,15Z)-tricosapentaene (C23 pentaene), in ratios of approximately 100:100:5:5. Other minor components of pheromone gland extracts included (11Z,13E)-hexadecadienal; (11E,13Z)-hexadecadienal; (11Z,13Z)-hexadecadien-1-yl acetate; (Z)-11-hexadecenal; hexadecanal; hexadecan-1-ol; and a second pentaene, (3Z,6Z,9Z,12Z,15Z)-pentacosapentaene (C25 pentaene). These minor components did not increase attraction of male navel orangeworm to the basic four-component blend. The use of another, cross-attracted pyralid moth, Pyralisfarinalis L., as a model species was crucial in implicating the C23 pentaene as an important component of the navel orangeworm pheromone blend. The four-component navel orangeworm pheromone blend was optimized using a combination of wind tunnel and field bioassays. Attractiveness of field deployed synthetic pheromone lures decreased rapidly despite incorporation of stabilizers and use of different release devices, suggesting that degradation products antagonize male navel orangeworm responses. Overall, the combination of type I lepidopteran pheromone components consisting of C16 aldehydes and alcohols with type II components consisting of long-chain polyunsaturated hydrocarbons has now been documented in several lepidopteran species and may indicate a paradigm shift in the range of compounds that constitute sex pheromone blends for individual lepidopteran species. This suggests that careful reexamination of pheromone gland contents for both type I and type II compounds may prove fruitful in species that have been studied but for which full attractant blends have eluded identification.