Responses of Flea Beetle <Emphasis Type="Italic">Phyllotreta cruciferae</Emphasis> to Synthetic Aggregation Pheromone Components and Host Plant Volatiles in Field Trials

Male-specific compounds, previously identified from Phyllotreta cruciferae and synthesized or isolated from natural sources, attracted both sexes of the beetle in field trials and therefore function as components of a male-produced aggregation pheromone. Six field experiments of 7 to 10 d duration each were conducted over 2 yr using modified boll weevil traps and two doses of pheromone. Treatments containing two doses of allyl isothiocyanate (AITC), a breakdown product of glucosinolates in Brassica napus L., a host plant of the beetles, were included in the study. A dose response was observed for both the pheromone components and AITC, and combinations of the pheromone and AITC generally attracted greater numbers of flea beetles than did either component itself. This increased attraction to a combination of beetle-produced compounds and host odors has not been previously demonstrated in halticine beetles and could help explain patterns of movement by P. cruciferae into field crops.     

Effect of allyl isothiocyanate on field behavior of crucifer-feeding flea beetles (Coleoptera: Chrysomelidae)

When water traps baited with allyl isothiocyanate (AIC)diffusing through polyvinyl chloride (PVC) and rubber membranes were used to monitor four species of crucifer-feeding flea beetle adults in a rutabaga field at L'Assomption, Que. in 1980–1981, differential responses to AIC were observed.Phyllotreta cruciferae was more attracted to AIC thanP. striolata, whereas the behavior ofPsylliodes punctulata was not affected by the presence of AIC. The traps with the PVC membrane caught significantly more flea beetles than the traps with the rubber membrane in 1980, but caught a similar number in 1981. Sticky traps covered with AIC mixed with Tangletrap® caught significantly more flea beetles than control sticky traps.     

Response of flea beetles,Phyllotreta spp., to mustard oils and nitriles in field trapping experiments

Trapping experiments were carried out near Winnipeg, Canada, in the spring of 1987 and 1988 to test attraction of crucifer-feeding flea beetles to volatile glucosinolate (GS) hydrolysis products released from glass vials. Nine isothiocyanates (IC) or mustard oils and three nitriles (CN) were tested. The pattern of attraction was the same for both flea beetle species,Phyllotreta cruciferae, andP. striolata. Captures in traps baited with allyl IC increased as release rates increased from 0.04 mg/day to 40 mg/day. The lowest rate that attracted large numbers of beetles was 4 mg/day; therefore this rate was used for further experiments. More beetles were captured in traps baited with allyl IC than with any other compound. In 1988 only, four IC in addition to allyl IC were attractive to both species; namely, benzyl IC, ethyl IC, and a mixture of ethyl and methyl 4-isothiocyanatobutyrate (ICB). When captures of the two species were pooled, 3-methylthiopropyl IC, methyl ICB, andn-butyl IC were also found to be significantly attractive. Nitriles were the least attractive compounds. The high release rates of mustard oils required to attract flea beetles indicate that fields ofBrassica crops would release sufficient quantities of IC to attract flea beetles from a distance but individual or small groups of plants normally would not. It is concluded thatBrassica varietal resistance to flea beetles is unlikely to be affected by manipulating IC release.     

Developmental profile of sinalbin (p-hydroxybenzyl glucosinolate) in mustard seedlings,Sinapis alba L., and its relationship to insect resistance

Sinalbin was identified as a chemical component of insect anti-xenosis and antibiosis resistance mechanisms in seedlings ofSinapis alba by DEAE-Sephadex chromatography, HPLC, treatment with sulfatase and myrosinase, various feeding tests using artificial and natural substrates, and by measuring sinalbin concentrations in cotyledons and leaves during seedling development. The effects of sinaibin on feeding were dependent upon the insect species and upon the rapidly changing profile of sinaibin concentrations in the developing seedling. The high concentrations of sinalbin found in young cotyledons (up to 20 mM) and leaves (up to 10 mM) deterred the feeding of the flea beetle,Phyllotreta cruciferae Goeze and larvae of the bertha armyworm,Mamestra configurata Walker. The protection that sinalbin confers upon the vulnerable, newly emerged seedling (and upon tiny, young leaves) appears critical for the first few days of survival ofS. alba under feeding pressure from flea beetles in the field. The lower concentrations of sinaibin found in older cotyledons and leaves (2–3 mM) offer little or no protection againstP. cruciferae and may actually stimulate the feeding of this crucifer specialist. These concentrations of sinaibin, however, are still effective in reducing the level of feeding by larvae of the more generalist feederM. configurata.     

Pheromonal Activity of Compounds Identified from Male <Emphasis Type="Italic">Phyllotreta cruciferae</Emphasis>: Field Tests of Racemic Mixtures,Pure Enantiomers,and Combinations with Allyl Isothiocyanate

Four himachalene sesquiterpenes and (+)-γ-cadinene, previously identified as possible pheromone components from males of a North American population of Phyllotreta cruciferae Goeze (Coleoptera, Chrysomelidae), were tested for attractiveness in field trapping experiments in Hungary. A mixture of the four synthetic racemic himachalene derivatives and (+)-γ-cadinene from a botanical source was slightly attractive to beetles, but much more attractive when blended with the known host-plant-derived attractant allyl isothiocyanate. This result was consistent with a previous study in North America. In tests with optically pure synthetic compounds, a blend of the same himachalene enantiomers found from male beetles was equivalent to the corresponding blend of racemic compounds, whereas a blend of the opposite enantiomers was not active. Through subtraction tests, it was found that the single compound, (6R,7S)-2,2,6,10-tetramethylbicyclo[5.4.0.]undeca-9,11-diene [compound (+)-A in this study], was as active as the whole mixture, suggesting that this compound is the key pheromone component of the European population of P. cruciferae. During field trials, several congeneric species, including P. vittula, P. nemorum, P. nodicornis, and P. ochripes, also were caught, suggesting that the same compound(s) may be relatively widespread as pheromone components in this genus.     

Aggregation pheromone ofCarpophilus antiquus (Coleoptera: Nitidulidae) and kairomonal use ofC. lugubris pheromone byC. antiquus

Males ofCarpophilus antiquus Melsheimer (Coleoptera: Nitidulidae) emit an aggregation pheromone that was found to be a novel hydrocarbon, (3E,5E,7E,9E)-6,8-diethyl-4-methyl-3,5,7,9-dodecatetraene. A synthetic scheme and spectra (mass and proton NMR) are given for the compound. Beetles produced the pheromone when feeding on a variety of media, including the brewer's yeast-based artificial diet, fermenting whole-wheat bread dough, corn, and prunes; live baker's yeast was generally added to the food media. Males held individually produced, on average, 25 × more pheromone per beetle than males held in groups of 10 or more. Pheromone was not produced until males were at least 5 days old but was still detected from the oldest beetles tested (47 days). In field tests, the pheromone was attractive to both sexes ofC. antiquus, and it was synergized by food volatiles: A combination of pheromone and fermenting whole wheat dough attracted 2.5× more beetles than pheromone alone, but dough by itself was not significantly more attractive than the control. Semiochemical interactions were studied amongC. antiquus and two other sympatric species for which pheromones are known,C. lugubris Murray andC. freemani Dobson.C. antiquus responded readily to the pheromone ofC. lugubris, but all other interspecific responses to the pheromones were weak. In a sample of naturally infested corn ears, the presence ofC. antiquus was strongly associated with the presence ofC. lugubris, as would be expected if the pheromone ofC. lugubris serves as a kairomone forC. antiquus.     

Attraction of pinyon pine bark beetle,Ips hoppingi,to conspecific andI. confusus pheromones (Coleoptera: Scolytidae)

Females of a pinyon pine bark beetle,Ips hoppingi Lanier, were less attracted by the aggregation pheromone produced by conspecific males than by the pheromone produced by the neighboring sibling species,I. confusus (LeConte). Cross-attraction was elicited by males infesting the regional pinyon pine hosts (P. discolor andP. edulis) of eitherIps species in south-eastern Arizona. Pheromonal specificity has not accompanied speciation in this species pair.     

Male-produced aggregation pheromone ofCarpophilus obsoletus (Coleoptera: Nitidulidae)

Males ofCarpophilus obsoletus Erichson produce an aggregation pheromone to which both sexes respond. The pheromone was identified by GC-MS as (2E,4E,6E,8E)-3,5,7-trimethyl-2,4,6,8-undecatetraene (1), which is also a minor constituent of the pheromone blends ofC. hemipterus (L.),C. freemani Dobson, andC. lugubris Murray. The pheromone was synergized in wind-tunnel bioassays by propyl acetate, a host-type coattractant. In a dose-response study, 50 pg of1, plus propyl acetate, was significantly more attractive than just propyl acetate. Pheromone emission from groups of 65 males, feeding on artificial diet, averaged 2.2 ng/male/day. Emissions from individual males were larger, averaging 72 ng/day and ranging as high as 388 ng/day. Synthetic1 was tested in a date garden in southern California (500 µg/rubber septum), using fermenting whole-wheat bread dough as the coattractant. The pheromone plus dough attracted significantly more beetles than dough alone (means were 4.2 and 0.0 beetles per week per trap). Captured beetles were 54% females. Field trap catches were highest during the months of July and August.All programs and services of the U.S. Department of Agriculture are offered on a nondiscriminatory basis without regard to race, color, national origin, religion, sex, age, marital status, or handicap. Mention of firm names or trade products does not imply endorsement by the U.S. Department of Agriculture over similar firms or products not mentioned.     

Field attraction ofBiprorulus bibax Breddin (Hemiptera: Pentatomidae) to synthetic aggregation pheromone and (E)-2-hexenal,a pentatomid defense chemical

A synthetic blend of the aggregation pheromone [(3R,4S,1E)-3,4-bis(1-butenyl)tetrahydro-2-furanol, linalool, farnesol, and nerolidol] of the spined citrus bug,Biprorulus bibax, and the pentatomid defense chemical, (E)-2-hexenal, both attracted adultB. bibax to individual trees in citrus orchards. Lemon trees containing single glass vials with aggregation pheromone or (E)-2-hexenal were colonized by significantly greater numbers of reproductiveB. bibax than unbaited trees. There was no significant difference between the treatments and bug recruitment was not improved by using both treatments.B. bibax did not enter cylinder/funnel traps baited with aggregation pheromone but colonized trees containing the traps. Orange or lemon trees containing aggregation pheromone on orchard perimeters recruited significantly larger populations of emigrating, nonreproductiveB. bibax during fall than untreated trees. Nonreproductive bugs were not attracted to trees containing (E)-2-hexenal. The potential for using these semiochemicals as management tools forB. bibax is discussed.     

Ancestral semiochemical attraction persists for adjoining populations of siblingIps bark beetles (Coleoptera: Scolytidae)

Three species ofIps pine bark beetles in Hopping's group IX (S.L. Wood'sgrandicollis group),Ips confusus, I. lecontei, andI. paraconfusus, are parapatrically distributed in the American Southwest. They share post-Pleistocene altitudinal ecotones with their host pines. Adjacent to these areas of host overlap, we tested the hypothesis that aggregation pheromones produced by male beetles and/or host volatiles are sufficient to elicit the speciesspecific colonization behaviors typical of these threeIps species in nature. A more distantly related species,I. pini (Hopping's group IV, S.L. Wood'spini group) was used for outgroup comparison. Under the influence of pheromone, males ofI. confusus andI. paraconfusus do not discern among unin-fested log bolts of host and nonhost pine prior to bark contact. Males responding to pheromones emanating from infested bolts are similarly undiscriminating. Females ofI. confusus andI. lecontei olfactorily discern the combination of conspecific males in host pine from other possible beetlepine combinations; females ofI. paraconfusus do not. FemaleI. pini discerned conspecific pheromone from that ofI. lecontei. The bark beetle predator,Enoclerus lecontei, is attracted by, but does not discriminate among, the male-produced volatiles of these Group IXIps species. These results support a hypothesis that divergence in pheromonal responses by these group IXIps species has evolved following their speciation, having been manifested first in the female sex. Evolutionarily, the derived pheromonal messages have preceded their behavioral discrimination by these beetles. Additional speciesspecific cues may operate between the sexes in the field that may preclude heterospecific pairings.     

Aggregation pheromone of driedfruit beetle,Carpophilus hemipterus Wind-tunnel bioassay and identification of two novel tetraene hydrocarbons

A male-produced aggregation pheromone was demonstrated inCarpophilus hemipterus (L.) (Coleoptera: Nitidulidae) using a wind-tunnel bioassay. Both sexes responded to the pheromone, but the beetles flew in the wind tunnel only after they had been starved for at least several hours. The attractiveness of the pheromone was greatly enhanced by volatiles from a food source, and combinations of pheromone and food volatiles typically attracted 3–10 times more beetles than either source by itself. A variety of food-related sources of volatiles were effective. These included apple juice; a mixture of baker's yeast plus banana; the pinto bean diet used for rearing this beetle; the chemicals propyl acetate, ethanol; and a mixture of acetaldehyde, ethyl acetate, and ethanol. The pheromonal activity resided with a series of 10 male-specific, unsaturated hydrocarbons of 13, 14, and 15 carbon atoms. These were partially separated by HPLC. No single compound was absolutely required for pheromonal activity to be observed, and various subsets of these compounds were active. The most abundant component was (2E,4E,6E,8E)-3,5,7-trimethyl-2,4,6,8-decatetraene. One minor component was (2E,4E,6E,8E)-3,5,7-trimethyl-2,4,6,8-undecatetraene. These structures were proven by synthesis. Together, the synthetic compounds were as active in the wind tunnel as the beetle-derived pheromone.     

Effects of Enantiomeric Blend of Verbenone on Response ofIps paraconfusus to Naturally Produced Aggregation Pheromone in the Laboratory

The aggregation pheromone produced by maleIps paraconfusus (Coleoptera: Scolytidae) tunneling in Monterey pine,Pinus radiata, logs was trapped on Porapak Q. A concentration of an extract of trapped volatiles that attracted beetles was determined in the laboratory through a concentration–response analysis of walking behavior of males and females. The interruptant effects of four concentrations of verbenone on response to a constant concentration of this naturally produced aggregation pheromone were tested with males and females. Independent of its enantiomeric composition [99.5% (S)-(–), 93.1 % (S)-(–), and 98.3% (R)-(+)], verbenone significantly reduced the percentage of females (but not males) reaching the attractant source. However, when the time required for beetles of both sexes taken together to reach the attractant source was considered, verbenone of higher enantiomeric purity had a greater effect on beetle behavior. Solutions of 99.5% (S)-(–)- and 98.3% (R)-(+)-verbenone increased the time required for beetles to reach the attractive source when compared to 93.1% (S)-(–)-verbenone. When pooled across enantiomeric blends, increasing concentrations of verbenone resulted in slower responses in beetles that reached the attractant source within 2 min. Males and females did not respond to verbenone alone.     

(S)-2-pentadecyl acetate and 2-pentadecanone Components of aggregation pheromone ofDrosophila busckii

(S)-2-Pentadecyl acetate and 2-pentadecanone were identified as the major aggregation pheromone components, inDrosophila busckii. Both sexes of flies were attracted equally in a wind-tunnel olfactometer. The flies also responded to racemic 2-pentadecyl acetate but not to the pureR enantiomer. In bioassay, (S)-2-pentadecyl acetate and 2-pentadecanone were each active alone, and a mixture of both increased the number of flies responding ca. twofold. The aggregation pheromone components are found in the ejaculatory bulb of sexually mature males and are transferred primarily to the female cuticle during mating. One third of the pheromone transferred is released by the female to the surrounding environment in a few hours after mating. None of the aggregation pheromone components remained on the mated female's cuticle, leaving two thirds unaccounted for. The same results were obtained when racemic 2-pentadecyl acetate was topically applied to immature and mature virgin males and females. BothD. mulleri andD. busckii were attracted to (S)-2-acetates of 13, 14 and 15 carbons, butD. mulleri preferred (S)-2-tridecyl acetate andD. busckii preferred (S)-2-pentadecyl acetate.     

Male-Specific Sesquiterpenes from Phyllotreta and Aphthona Flea Beetles

It was previously reported that males of the crucifer flea beetle, Phyllotreta cruciferae, feeding on host foliage are attractive to both males and females in the field. Based on this evidence for an aggregation pheromone, volatiles were collected from male and female P. cruciferae feeding on cabbage (Brassica oleracea) and analyzed. For comparison, volatiles were also collected from males and females of three other flea beetle species, Aphthona flava, A. czwalinae, and A. cyparissiae, all feeding on their host, leafy spurge foliage (Euphorbia esula). Six male-specific compounds were isolated from P. cruciferae, and the same compounds plus two additional ones were isolated from males of Aphthona flava, A. czwalinae, and A. cyparissiae. The blends of compounds were relatively consistent within species, but there were characteristic differences between species. Compound structures were studied by mass spectrometry, NMR spectroscopy, UV spectroscopy, polarimetry, chiral and achiral gas chromatography, molecular modeling, and microchemical tests. Three of the compounds were identified as (+)-ar-himachalene; (+)-trans--himachalene; (+)--cadinene. Two others were new enantiomers of himachalene hydrocarbons that were previously identified from the fir trees, Abies alba and Abies nordmanniana. Finally, there were two himachalene alcohols and one norsesquiterpene ketone that is a himachalene analog. Only (+)-ar-himachalene and (+)--cadinene are previously known natural products. Electrophysiological activity was demonstrated for five of the compounds. The chemical and electrophysiological patterns are consistent with, but do not prove, a pheromonal function.     

Identification of major component of male-produced aggregation pheromone of larger grain borer,Prostephanus truncatus (Horn) (Coleoptera: Bostrichidae)

Male beetles of the larger grain borer,Prostephanus truncatus Horn (Coleoptera: Bostrichidae) were shown to produce material causing an electroantennographic (EAG) response from both male and female beetles. Volatiles from mixed sex cultures were collected on Porapak Q and fractionated by liquid chromatography (LC) with gradient elution. The total volatiles and the fractions were analyzed by gas chromatography (GC) linked to EAG, and two compounds eluting in the same LC fraction elicited EAG responses from male and female beetles. These were assumed to be components of the male-produced aggregation pheromone. Amounts of these compounds obtained were very low and less than 10^–3 times the amounts of the aggregation pheromone components produced by the related species,Rhyzopertha dominica, under similar conditions. The most abundant of these EAG-active compounds was identified as 1-methyiethyl (2E)-2-methyl-2-pentenoate by comparison of its GC retention times and mass spectrum with those of synthetic analogs. The synthetic compound elicited electrophysiological and behavioral responses in both male and female beetles significantly greater than those to structural analogs. This compound is attractive toP. truncatus beetles in the field, and has been given the trivial name of trunc-call 1.     

Glucosinolate levels in cotyledons of mustard,Brassica juncea L. and rape,B. napus L. do not determine feeding rates of flea beetle,Phyllotreta cruciferae (Goeze)

Sinigrin (allyl glucosinolate), the major glucosinolate in the cotyledons ofBrassica juncea cv. Cutlass, occurred in the highest concentration and amount at seedling emergence and declined during growth. Glucobrassicin (3-indolylmethyl glucosinolate), the major glucosinolate in the cotyledons ofB. napus cv. Westar, occurred in the lowest concentration and amount at seedling emergence. The amount of glucobrassicin per cotyledon pair increased about fourfold during 14 days of growth, but its concentration remained relatively unchanged because of dilution by increasing cotyledon biomass. These different glucosinolate profiles indicate a different metabolic control and different biological function for sinigrin and glucobrassicin. The flea beetle,Phyllotreta cruciferae Goeze, does not discriminate between cotyledons having sinigrin or glucobrassicin since the two crucifers were fed upon equally in choice tests. Restricting the concentration of sulfur in the nutrient medium accelerated the decline of sinigrin inB. juncea cv. Cutlass but did not alter the feeding rate ofP. cruciferae compared to controls. Sulfur restriction reduced glucobrassicin inB. napus cv. Westar to undetectable levels and somewhat reduced the feeding rate of P.Cruciferae. Nevertheless,P. cruciferae still fed actively on cotyledons ofB. napus cv. Westar depleted of glucosinolates and severely damaged many of them. Since glucosinolate type and concentration had little effect on feeding response, reduction or elimination of foliar glucosinolates alone would not seem a useful strategy for protecting seedlings of these two crucifers from flea beetle damage.     

Male-Produced Aggregation Pheromone Compounds from the Eggplant Flea Beetle (Epitrix fuscula): Identification, Synthesis, and Field Biossays

Volatiles from the eggplant flea beetle, Epitrix fuscula Crotch (Coleoptera: Chrysomelidae), feeding on host foliage, were investigated. Six male-specific compounds were detected and were identified through the use of mass spectrometry, nuclear magnetic resonance (NMR) spectrometry, chiral and achiral gas chromatography, high-performance liquid chromatography, electrophysiology (gas chromatography-electroantennography, GC–EAD), and microchemical tests. The two most abundant of the six compounds were (2E,4E,6Z)-2,4,6-nonatrienal (1) and (2E,4E,6E)-2,4,6-nonatrienal (2). The other four compounds, present in minor amounts, were identified as himachalene sesquiterpenes; two of these, 3 and 4, were hydrocarbons and two, 5 and 6, were alcohols. All four sesquiterpenes were previously encountered from male flea beetles of Aphthona spp. and Phyllotreta cruciferae. Synthetic 1 and 2 matched the natural products by GC retention times, mass spectra, and NMR spectra. Sesquiterpenes 3–6 similarly matched synthetic standards and natural samples from the previously studied species in all ways, including chirality. Both natural and synthetic 1 and 2 gave positive GC–EAD responses, as did sesquiterpenes 3, 5, and 6. Field trials were conducted with a mixture of 1 and 2, and the baited traps were significantly more attractive than control traps to both male and female E. fuscula. The E. fuscula pheromone has potential for monitoring or controlling these pests in eggplants.Mention of trade names or commercial products in this article is solely for the purpose of providing specific information and does not imply recommendation or endorsement by the Department of Agriculture.     

Mechanism of aggregation behavior inMaladera matrida Argaman (Coleoptera: Scarabaeidae)

AdultMaladera matrida Argaman (Coleoptera: Scarabaeidae: Melolonthinae) males emerge from soil for an active period at dusk, a few minutes before the females. Adults are found during most of the active hours on the foliage in aggregations composed of an equal sex ratio. The mechanism of aggregation behavior ofM. matrida beetles was studied in a Y-shaped olfactometer. No evidence was found for the existence of an aggregation pheromone released either by males or by females, but behavior tests indicate that adultM. matrida beetles, males as well as females, are attracted to volatiles of an injured host plant. The following scenario is suggested: Males emerge daily from soil at dusk, a few minutes before the females, and immediately start feeding. Additional males are attracted to the injured host's volatiles and form aggregations. When females emerge from soil, the attractant volatiles are concentrated in spots, and the females join the aggregations, forming an equal sex ratio.     

Ipsenol: an aggregation pheromone forIps latidens (Leconte) (Coleoptera: Scolytidae)

Ipsenol was identified from the frass of male, but not female,Ips latidens from British Columbia, feeding in phloem tissue of lodgepole pine,Pinus contorta var.latifolia. The responses ofJ. latidens to sources of ips-enol andcis-verbenol were determined with multiple-funnel traps in stands of lodgepole pine in British Columbia. Ipsenol attracted both male and femaleI. latidens, verifying that it is a pheromone for this species. MaleI. latidens showed a slight preference for (S)-(–)-ipsenol.cis-Verbenol was not produced by beetles of either sex and, in contrast to an earlier report, both enantiomers inhibited attraction to ipsenol-baited traps. The predators,Enoclerus sphegeus andThanasimus undatulus (Cleridae), were attracted to traps baited withcis-verbenol and ipsenol.     

Identification of the Sex Pheromone of Holotrichia reynaudi

The male attractant pheromone of the scarab beetle Holotrichia reynaudi, an agricultural pest native to southern India, was extracted from abdominal glands of females with hexane and analyzed by gas chromatography–mass spectrometry. Field testing of the candidate chemicals, indole, phenol, and anisole, both alone and as binary mixtures, led us to conclude that anisole was the major component of the sex pheromone. Neither male nor female beetles were attracted to indole or phenol on their own. Similarly, when indole and anisole were combined, the attractiveness of the solution did not increase over that obtained with anisole alone. However, combination of phenol and anisole did alter the attractiveness of anisole, with fewer male beetles attracted to the binary mixture than to anisole on its own. The behavior of female beetles was not altered by any of the chemicals tested. Anisole is also the sex pheromone of H. consanguinea, making this the first known example of two melolonthine scarabs sharing the same pheromone.