Capture of female Hylotrupes bajulus as influenced by trap type and pheromone blend.

Three different types of traps were evaluated for a semiochemical-based trapping method for Hylotrupes bajulus (L.) (Cerambycidae). One, designated the ground trap, was the most efficient under both laboratory and natural conditions and had an active space of about 3.5 m. Significantly more beetles were captured in traps baited with a mixture of synthetic pheromones, (3R)-3-hydroxy-2-hexanone [(3R)-ketol] + 1-butanol, than in those with either single compound or with racemic mixtures. Furthermore, the synthetic lure captured more females than two virgin males in a laboratory bioassay. In addition, doubling the concentration of the synthetic pheromone significantly increased trap catches. The present findings have potential value of the management of this pest.

Biodegradation and transfer of ingested 2,4-D herbicide by a polyphagous saturniid caterpillar.

A herbicide containing 2,4-dichlorophenoxyacetic acid (2,4-D) and related chemicals was fed to caterpillars of Eupackardia calleta, and the fate of the substances in the larvae and during further ontogenesis was followed by combined gas chromatography/mass spectrometry. The compounds were found in differing amounts in larval midgut, faeces, fat body/haemolymph, and even in an exocrine secretion produced by integumental glands. Furthermore, they were detected in samples from the resulting adult moths, indicating an intraindividual transfer. Since the individual development of E. calleta was distinctly accelerated by 2,4-D, possible impacts of the herbicide on the life history of the animals in the field are discussed. Based on the chemical data, hypothetical metabolic pathways for 2,4-D in E calleta larvae are proposed.

Iridoid biosynthesis in staphylinid rove beetles (Coleoptera: Staphylinidae, Philonthinae).

The biosynthesis of chrysomelidial and plagiodial was studied in the rove beetle subtribe Philonthina (Staphylinidae). Glandular homogenates were found to convert synthetic (2E,6E)-[trideuteromethyl-5,5-(2)H(5)]octa-2,6-diene-1,8-diol (10) into nor-chrysomelidial (14) and nor-plagiodial (13). The overall transformation requires; i) oxidation of the substrate at C(1) and C(8), ii) cyclization of the resulting dialdehyde to nor-plagiodial followed by iii) isomerization to give nor-chrysomelidial. The oxidase requires molecular oxygen as a cofactor and operates with removal of the pro-R hydrogen from C(1) and C(8) of synthetic (1R,8R,2E,6E)-[1,8-(2)H(2)]-2,6-dimethyl-octa-2,6-diene-1,8-diol (15), producing a dialdehyde along with H(2)O(2). Unlike enzymes from iridoid-producing leaf beetle larvae, the Philonthus enzyme is able to oxidize saturated substrates such as citronellol. Crude protein extracts prepared from Philonthus glands by ammonium sulfate precipitation, were found to produce hydrogen peroxide at a rate of 0.085+/-0.003 ng H(2)O(2) (ng protein)(-1) hr(-1) with nerol as an oxidase substrate. The cyclase operates with opposite stereochemistry to the enzyme(s) from Phaedon cochleariae and other herbivorous leaf beetles, specifically removing the C(5)-H(R) hydrogen atom from (4R,5S,2E,6E)-[4,5-(2)H(2)]-2-methyl-octa-2,6-diene-1,8-diol (17). These findings have enabled us to construct a detailed account of iridoid biosynthesis in rove beetles, which resembles the biosynthetic route in leaf beetle larvae, but exhibits distinct stereochemical differences.

Comparative morphology and evolution of the funnel warts of larval Lymantriidae (Lepidoptera).

The larval funnel warts of 14 species of Lymantriidae (tussock moths) were investigated by means of scanning electron microscopy. The smaller, paired osmeteria on other abdominal segments were also inspected in four of the species. It seems that the bilaterally symmetrical funnel warts have developed by fusion of two smaller, single osmeteria. Based on the morphological findings from surface views and inside views of the structures, we made an attempt to reconstruct the pathway of evolution of the funnel warts. The high degree of specific modifications of the funnel warts appears valuable for analyses of the phylogenetic relationship of the investigated species.

Gut bacteria may be involved in interactions between plants, herbivores and their predators: microbial biosynthesis of N-acylglutamine surfactants as elicitors of plant volatiles.

N-Acylamino acids are dominant and widespread constituents of insect oral secretions (regurgitants), serving the insect as biosurfactants in the digestive process. During feeding the conjugates may be introduced into damaged leaves and contribute there to the elicitation of plant defenses such as the induction of volatile biosynthesis. From gut segments of Spodoptera exigua, Mamestra brassicae and Agrotis segetum 23 bacterial strains were isolated, ten of which were able to synthesise typical lepidopteran N-acylamino acids from externally added precursors. Four strains, Providencia rettgeri, Ochrobactrum spec., Myroides odoratus and Acinetobacter sp. genospecies 11 were identified on the basis of their 16 S rDNA. The organisms displayed a very broad substrate tolerance, since fatty acids of different chain length and different degree of saturation were converted into N-acylamino acids. Moreover, most of the proteinogenic amino acids, but not glutamic and aspartic acid, were used as substrates. The dominant occurrence of fatty acids conjugated with glutamine may result from a preferred transport of glutamine from the hemolymph into the gut of the insects. The involvement of bacteria in the biosynthesis of compounds which play a pivotal role in the interaction of plants, herbivores and their predators adds a new trophic level to this complex network of interactions. Due to their short generation cycle and the ease of adaptation endosymbiontic bacteria may have an outstanding importance for the coevolution of plant-insect interactions.

Polyunsaturated monoglycerides and a pregnadiene in defensive glands of the water beetle Agabus affinis.

In addition to the C21 steroid 15alpha-hydroxypregna-4,6-dien-3,20-dione, four 1- or 2-acylated polyunsaturated monoglycerides, 1- or 2-(cis-5,8,11,14-eicosatetraenoyl)-glycerol and 1- or 2-(cis-5,8,11,14,17-eicosapentaenoyl)glycerol were identified as constituents of the prothoracic defensive gland secretion of the dytiscid beetle Agabus affinis by gas chromatography-mass spectrometry of trimethylsilylated gland extracts. In a feeding assay with minnows, synthetic samples of the two 2-acylated monoglycerides showed only a weak activity as a feeding deterrent. For that reason, other possible functions of the monoglycerides are discussed, such as roles as emulsifiers of cannabimimetics.

The secretion of the ventral glands in Cladius, Priophorus and Trichiocampus sawfly larvae.

The volatile secretion from ventral glands of the larvae of Cladius pectinicornis, Priophorus morio, P. pallipes and Trichiocampus grandis was found to be principally composed of long-chain acetogenins, in majority of the esters and hydrocarbons, with more than 15 carbon atoms. The scarcity of more volatile compounds may be considered as plesiomorphic for the tribe Cladiini to which the four species belong. Further chemotaxonomic significance and chemical ecological implications of the glandular secretions are discussed. Moreover, the function of the well-developed pubescence covering the body of Cladiini larvae is discussed as a part of their defensive mechanism.

Transformation of steroids by Bacillus strains isolated from the foregut of water beetles (Coleoptera: Dytiscidae): II. Metabolism of 3 beta-hydroxypregn-5-en-20-one (pregnenolone).

The in vitro metabolism of pregnenolone by two Bacillus strains (HA-V6-3 and HA-V6-11) isolated from the foregut of the water beetle Agabus affinis (Payk.) was examined in the course of our studies about a possible participation of gut micro-organisms in the biosynthesis of prothoracic defensive steroids of dytiscids. The transformation products were identified by EI GC--MS of culture extracts after derivatization. The dominating reactions were hydroxylations, with 7 alpha-hydroxypregnenolone as the major product. With considerably lower yields, 7 beta- and 15xi-hydroxypregenolone were formed by both strains, while 11, 17 and 16 alpha-hydroxypregnenolone were produced only by HA-V6-3. The occurrence of 7, 11 alpha- and 7 beta, 11 alpha-dihydroxypregnenolone as well as several minor products containing a 17 alpha-OH group proved the capability of HA-V6-11 to hydroxylate pregenenolone at C(11) and C(17) as well. The monohydroxylated 7-OH-pregnenolones were partly oxidized to 7-oxopregnenolone by both strains. In trace amounts, HA-V6-3 performed 3 beta-acetylation of pregnenolone.

Transformation of steroids by Bacillus strains isolated from the foregut of water beetles (Coleoptera:Dytiscidae): I. Metabolism of androst-4-en-3,17-dione (AD).

Two Bacillus strains were isolated from the foregut of the water beetle Agabus affinis (Payk.) and tested for their steroid transforming ability. After incubation with androst-4-en-3,17-dione (AD), 13 different transformation products were detected. AD was hydroxylated at C6, C7, C11 and C14, resulting in formation of 6beta-, 7alpha-, 11alpha- and 14alpha-hydroxy-AD. One strain also produced small amounts of 6beta,14alpha-dihydroxy-AD. Partly, the 6beta-hydroxy group was further oxidized to the corresponding 6-oxo steroids. In addition, a specific reduction of the delta4-double bond was observed, leading to the formation of 5alpha-androstane derivatives. In minor yields the carbonyl functions at C3 and C17 were reduced leading to the formation of 3zeta-OH or 17beta-OH steroids. EI mass spectra of the trimethylsilyl and O-methyloxime trimethylsilyl ether derivatives of some transformation products are presented for the first time.

Role of cuticular hydrocarbons of aphid parasitoids in their relationship to aphid-attending ants.

Lysiphlebus cardui, the dominant aphidiid parasitoid of the black bean aphid,Aphis fabae cirsiiacanthoidis (Afc), on creeping thistle, is able to forage in ant-attended aphid colonies without being attacked by ants. Several behavioral observations and experimental studies led to the hypothesis thatL. cardui mimics the cuticular hydrocarbon profile of its host aphid. Chemical analysis of the cuticular extracts revealed that bothL. cardui and Afc exclusively possess saturated hydrocarbons:n-alkanes, monomethyl (MMA), dimethyl (DMA), and trimethyl alkanes (TMA). Comparison of the hydrocarbon profiles of parasitoid and aphid showed great qualitative resemblance between parasitoid and host:L. cardui possesses almost all host-specific compounds in addition to species-specific hydrocarbons of mainly higher molecular weight (>C30). However, there is a lesser quantitative correspondence between parasitoid and host aphid. Furthermore, we analyzed the cuticular hydrocarbon profile of another parasitoid of Afc,Trioxys angelicae. This aphidiid species is vigorously attacked and finally killed by honeydewcollecting ants when encountered in aphid colonies. Its cuticular hydrocarbon profile is characterized by the presence of large amounts of (Z)-11-alkenes of chain lenghts C27, C29, C31, and C33, in addition to alkanes and presumably trienes. The role of the unsaturated hydrocarbons onT. angelicae as recognition cues for aphid-attending ants is discussed.

Chemical defense of giant springtailTetrodontophora bielanensis (Waga) (Insecta: Collembola).

The giant springtail,Tetrodontophora bielanensis (Waga), is characterized by integumental openings (pseudocells) from which small droplets of a sticky defensive fluid are secreted after molestation. The secretion originates initially from secretory cells below the pseudocellae; subsequent irritations result in release of hemolymph, which was identified by both chemical and microscopical methods as well as by scanning electron microscopy. Bioassays with topically treated ground beetlesNebria brevicollis showed that the pseudocellular fluid evokes a total disorientation and cleansing behavior of the beetle. The main constituents were identified as the following pyridopyrazines: 2,3-dimethoxpyrido[2,3-b]pyrazine (1), 3-isopropyl-2-methoxypyrido[2,3-b]pyrazine (2), and 2-methoxy-4H-pyrido[2,3-b]pyrazine-3-one (3). These alkaloids are mainly present in the pseudocellar fluids of female and male springtails but are absent in their food or feces. Minor amounts are found in the hemolymph of adults, while larvae contain traces of2 only. All compounds were synthesized and tested for activity. In natural concentrations, the synthetic alkaloids elicited the same effects from the ground beetles as the pseudocellar fluid.

Allocation of pederin during lifetime ofPaederus rove beetles (Coleoptera: Staphylinidae): Evidence for polymorphism of hemolymph toxin.

Pederin, a hemolymph toxin found in the rove beetle genusPaederus, is quantified in individual specimens ofP. fuscipes andP. riparius. Whereas males always contain only small quantities of the toxin (0.1-1.5 µg), females are not so restricted (0.2-20.5 µg) and in most cases possess roughly 10-fold that in males. There are, however, females containing as little pederin as the males, and so two clearly separate groups of females can be discerned. These two groups hint at two types of females defined by the eggs they lay. About 85% of the females, i.e., those containing much of the substance, transfer pederin into every egg and thus are denoted (+)-females. The remaining 15%, the (-)-females, contain at best small amounts of the toxin, which is transferred in minute quantities into the first eggs while the subsequent ones lack pederin. With respect to the pederin content of their eggs, there is no overlap between these two types of females. If fed with pederin, (-)-females can transfer it into the eggs like (+)-females. After hatching the larvae store pederin when present. Larvae are not able to biosynthesize the toxin on their own, but storage of that received in the egg is very efficient, and the difference between larvae with and without pederin is preserved until imaginal eclosion. In (+)-females, pederin increases, probably reflecting a biosynthetic capacity, but in males and presumptive (-)-females the amount of pederin never exceeds the quantity transferred by parental (+)-females. Consequently, males and (-)-females probably are unable to biosynthesize pederin. This polymorphism of females may have a genetic basis. Individuals of all stages nevertheless sequester pederin if it is supplied with the diet.

Interactions betweenAlloxysta brevis (Hymenoptera, Cynipoidea, Alloxystidae) and honeydew-collecting ants: How an aphid hyperparasitoid overcomes ant aggression by chemical defense.

Foraging females of the aphid hyperparasitoidAlloxysta brevis were attacked by honeydew-collecting workers of the antLasius niger at the first encounter. However, ants abandoned their attacks quickly, and foragingA. brevis remained unmolested for a subsequent time interval of approximately 5 min, which is long enough for the hyperparasitoid to oviposit successfully. Furthermore, freshly killed intactA. brevis were disregarded by ants, while decapitated specimens were readily removed. We present evidence thatA. brevis females release a mandibular gland secretion, which contains 6-methyl-5-hepten-2-one, actinidin, and unidentified iridoids, in response to an ant attack. This secretion functions both as a measure of self-defense if the female is seized by an ant worker and as a repellent, which prevents ant attacks during subsequent encounters. This is the first evidence for chemical defense in a hymenopterous parasitoid. It enablesA. brevis females to hyperparasitize ant-attended aphids that constitute a major proportion of their hosts and significantly reduces mortality by ectohyperparasitoids.

Quantification of cantharidin in canthariphilous ceratopogonidae (Diptera), anthomyiidae (Diptera) and cantharidin-producing oedemeridae (Coleoptera).

Cantharidin contents were determined in several canthariphilous insects by means of quantitative gas chromatography. Usually the ceratopogonidsAtrichopogon oedemerarum andA. trifasciatus caught in the field contained low concentrations of cantharidin, with concentrations in males, in most cases, being lower than in females. When fed in the laboratory with synthetic cantharidin, these species concentrated cantharidin by as much as 100-fold (males) and 40-fold (females). Accumulation in the different body tagmata (head, thorax, abdomen) of these species is similar. Maximal concentrations of cantharidin in tissues ofAtrichopogon are comparable to those known from oedemerid and meloid beetles. InA. trifasciatus about 90% of total cantharidin content is bound in tissues. Investigations using the canthariphilous anthomyiid flyAnthomyia pluvialis and three cantharidin-producing oedemerid species revealed the same pattern of distribution in different body tagmata as inAtrichopogon.

Attacus atlas caterpillars (Lep., Saturniidae) spray an irritant secretion from defensive glands.

The ability ofAttacus atlas caterpillars to spray a defensive secretion seems to be due to the fine structure of the integumental glands that produce it. The giant gland cells are fixed to stable cuticular rings surrounding the gland openings and tightly closed by cuticular lids. Probably by increasing hemolymph pressure, the lids are blasted off and the secretion spouts out. The fluid contains several aromatics, biogenic amines (e.g., acetylcholine, histamine), glycerol, and trehalose and exhibits tyrosinase activity. Deterrent effects of caterpillar secretion and hemolymph on predatory ants could be shown. Presumably the spraying process serves to apply the secretion to sensitive sites of vertebrate target organisms.

Recognition of aphid parasitoids by honeydew-collecting ants: The role of cuticular lipids in a chemical mimicry system.

The aphidiid waspLysiphlebus cardui parasitizes in ant-attendedAphis fabae cirsiiacanthoidis colonies without causing aggressive behavior in the antLasius niger. By contrast,Trioxys angelicae, another aphidiid parasitoid of aphids, is rapidly recognized and vigorously attacked by the ants.L. niger workers also responded differently to dead individuals ofL. cardui andT. angelicae. DeadL. cardui parasitoids were often ignored when encountered byL. niger, whereas deadT. angelicae individuals were immediately grasped by ants that discovered them. However, hexane-washed parasitoids caused a similar reaction pattern in the ants, in that both aphidiid species were tolerated in the aphid colony. Lure experiments demonstrated that chemical stimuli on the cuticle are major cues for the ants to distinguish between the parasitoids. The hexane extract ofL. cardui transferred to washed individuals ofT. angelicae resulted in ant responses characteristic towardsL. cardui, andL. niger workers displayed the typical removal pattern they normally showed towardsT. angelicae whenT. angelicae extract was applied toL. cardui individuals. Both parasitoid species treated with the hexane extract ofA. fabae cirsiiacanthoidis were similarily treated by the ants as were aphid control individuals. The suggestion that the aphidiid waspL. cardui uses chemical mimicry is discussed.

Dabbing and shooting of benzo- and naphthoquinone secretions: Defensive strategies of bark-inhabiting aleocharine (col.: Staphylinidae) and tenebrionid (col.: Tenebrionidae) beetle larvae.

The abdominal glands of three bark-inhabiting larvae of generaBolitochara, Leptusa (Staphylinidae), andHypophloeus (Tenebrionidae) were studied chemically and morphologically. Behavior of the larvae indicated that secretion is emitted only after severe disturbance of the larvae. These mechanical contacts may also occur incidentally with coinhabiting nonpredatory arthropods when the beetle larvae move within small interstices under bark. Depending on the species, the secretions contained 1,4-benzoquinone derivatives and three 6-alkyl-naphthoquinones dissolved within various alkanes, alkenes, ethyl-, isopropyl-, and isoamylesters. More erratically distributed gland constituents also detected were acetophenone, benzyl propionate, and methyl hydroxybenzoate. In the laboratory, synthetic quinone-containing solutions simulating those found inLeptusa andBolitochara larvae acted as strong topical irritants and caused further damage to last-stageCalliphora vomitoria larvae if hydrocarbons or esters were used as solvents. The natural secretions ofHypophloeus versipellis elicited considerable mortality in two subcortical sciarid larvae cooccurring with tenebrionid larvae. Bioassay and secretion chemistry of the Staphylinidae/Tenebrionidae larval secretions indicated that they are typical defensive secretions that act topically. Morphological data characterized Bolitocharini larvae as possessing protuberant abdominal tergites supplied with an interiorly situated gland reservoir. After mechanical contact, the defensive secretion is topically applied to other arthropods by dabbing this tergal protuberance on targets. The defensive gland ofHypophloeus versipellis is unusual in possessing a movable reservoir opening situated at the anterior border of tergite IX. By this peculiar gland morphologyHypophloeus larvae are capable of shooting secretion droplets frontally from their slightly depressed dorsal abdominal surface without bending their abdominal tips dorsally. This seems an adaptation to the interstitial habitat of the larvae. The types of defensive glands and their phylogenetic value in Aleocharinae/Tenebrionidae larvae are discussed.

Defense chemicals from abdominal glands of 13 rove beetle species of subtribe staphylinina (Coleoptera: Staphylinidae, Staphylininae).

Forty-one volatile constituents have been characterized from the abdominal defensive glands of 13 Staphylinina rove beetle species (Staphylinidae). The major secretion component, a rapidly polymerizing iridoid dialdehyde (in most cases iridodial), is mainly accompanied by variable amounts of some iridoid lactones, traces of actinidine, some possible monoterpene precursors, short-chain ketones, cyclic compounds, and spiroacetals. The secretion mixtures even of single Staphylinina genera are very heterogenously composed of a variety of active substances with different structures. This situation may be a typical feature of iridoid defense systems based upon a primary fixative major compound and clearly differs from other chemical defense systems (for example the quinone system) which have a toxic main component.

Sticky secretion from two pairs of defensive glands of rove beetleDeleaster dichrous (Grav.) (Coleoptera: Staphylinidae) : Gland morphology, chemical constituents, defensive functions, and chemotaxonomy.

The phylogenetically primitive rove beetleDeleaster dichrous (Grav.) (Oxytelinae) has been shown for the first time to possess two pairs of neighboring abdominal glands which are depleted simultaneously on molestation. The morphology of these glands is described. The defensive constituents of theDeleaster glands were elucidated directly from the mixtures by gas chromatographic-mass spectroscopic methods and microchemical reactions. The paired whitish glands secrete iridodial, which polymerizes on exposure to air to form an adhesive that probably deters small predatory arthropods. The red gland system ofD. dichrous contains the toxicp-toluquinone and a variety of isopropyl andsec-butyl esters. Artificial quinoid ester mixtures simulating the secretion ofD. dichrous showed only weak effects on mortality ofLucilia larvae in comparison with more effective secretions of phyiogenetically derived Oxytelinae. The secretion of the primitive genusDeleaster is characterized chemotaxonomically by ß, γ-unsaturated C12 acids and esters, which are postulated as precursors for the characteristic defensive compounds of the derived species, thus indicating a clear evolutionary trend at the micromolecular level.

Pygidial defensive titer and population structure ofAgabus bipustulatus L. andAgabus paludosus F. (Coleoptera, Dytiscidae).

Aromatic pygidial gland constituents ofAgabus bipustulatus L. andA. paludosus F. were quantitatively determined. Concentration fluctuations were found to be dependent on age and season. Both in quantity and quality of the secretion, young beetles differ from mature beetles by storing only small amounts of gland material with different portions of constituents. Seasonal variations are mainly due to the changing population structure of the species.