Shield Defense of a Larval Tortoise Beetle

Larvae of the folivorous tortoise beetle, Plagiometriona clavata, carry shields formed from feces and exuviae above their bodies. We used an ecologically relevant predatory ant, Formica subsericea, in a bioassay to determine if shields functioned as simple barriers, as previous studies indicated, or whether they were chemical defenses. Shields were necessary for larval survival; shield removal rendered larvae vulnerable. Shields produced by larvae reared on a substitute diet failed to provide protection. Solvent-leached shields also failed to deter ants, indicating the shield had a host-derived chemical component likely located in the feces, not in the exuviae. Solanum dulcamara, the larval host plant, contained free phytol, steroidal glycoalkaloids, and saponins. Shields contained partially deglycosylated metabolites of host steroidal glycoalkaloids and saponins, a suite of fatty acids, and derivatives of phytol, which together formed a deterrent barrier against ant attack. We compared the mobile shield of P. clavata to the stationary shield of another S. dulcamara-feeding leaf beetle, Lema trilinea. Both larval shield defenses were formed from a very similar array of host-derived compounds with deterrent properties. We concluded that convergent patterns of limited chemical transformation and selective incorporation of particular deterrent metabolites in shield defenses of two unrelated taxa represented responses to selection from invertebrate predators.     

Sequestration of Furostanol Saponins by <Emphasis Type="Italic">Monophadnus</Emphasis> Sawfly Larvae

Sawfly larvae of the tribe Phymatocerini (Hymenoptera: Tenthredinidae), which are specialized on toxic plants in the orders Liliales and Ranunculales, exude a droplet of deterrent hemolymph upon attack by a predator. We investigated whether secondary plant metabolites from Ranunculaceae leaves are sequestered by phymatocerine Monophadnus species, i.e., Monophadnus alpicola feeding upon Pulsatilla alpina and Monophadnus monticola feeding upon Ranunculus lanuginosus. Moreover, two undescribed Monophadnus species were studied: species A collected from Helleborus foetidus and species B collected from Helleborus viridis. Comparative high-performance liquid chromatographic–photodiode array detection–electrospray ionization–mass spectrometric analyses of plant leaf and insect hemolymph extracts revealed the presence of furostanol saponins in all samples. Larvae of species A and B actively sequestered (25R)-26-[(α-l-rhamnopyranosyl)oxy]-22α-methoxyfurost-5-en-3β-yl O-β-d-glucopyranosyl-(1→3)-O-[6-acetyl-β-d-glucopyranosyl-(1→3)]-O-β-d-glucopyranoside (compound 1). This compound occurred at a 65- to 200-fold higher concentration in the hemolymph of the two species (1.6 and 17.5 μmol/g FW, respectively) than in their host plant (0.008 and 0.268 μmol/g FW, respectively). In M. monticola, compound 1 was found at a concentration (1.2 μmol/g FW) similar to that in the host plant (1.36 μmol/g FW). The compound could not be detected consistently in M. alpicola larvae where, however, a related saponin may be present. Additional furostanol saponins were found in H. foetidus and H. viridis, but not in the two Monophadnus species feeding on them, indicating that sequestration of compound 1 is a highly specific process. In laboratory bioassays, crude hemolymph of three Monophadnus species showed a significant feeding deterrent activity against a potential predator, Myrmica rubra ant workers. Isolated furostanol saponins were also active against the ants, at a concentration range similar to that found in the hemolymph. Thus, these compounds seem to play a major role for chemical defense of Monophadnus larvae, although other plant secondary metabolites (glycosylated ecdysteroids) were also detected in their hemolymph. Physiological and ecological implications of the sequestered furostanol saponins are discussed. Dedicated to the memory of Professor Ivano Morelli (1940–2005)     

Chemical Defense in Larval Tortoise Beetles: Essential Oil Composition of Fecal Shields of Eurypedus nigrosignata and Foliage of its Host Plant, Cordia curassavica

Larvae of the tortoise beetle Eurypedus nigrosignata construct fecal shields using cast skins and fecal strands. Survival of larvae with intact shields was higher in the field than for larvae with shields removed. In the laboratory, E. nigrosignata feculae had a deterrent effect on feeding in the ant Myrmica rubra as did an extract of the host plant, Cordia curassavica. Three chemical types were identified in the host-plant foliage and were named -terpinene, -pinene, and sabinene, depending on their mono- and sesquiterpene composition. This is the first report of lower terpenes (essential oils) in foliage of Cordia. Fecal shields of E. nigrosignata displayed the same terpene pattern as larval host-plant leaves. The absolute concentration of mono- and sesquiterpenes in the dorsal fecal shield depended on the plant chemical type and tended to decrease with larval age. No oxidation or detoxification products of ingested terpenes were detected in the larval fecula, indicating that the chemical composition of the larval fecal shield is influenced primarily by the host-plant secondary chemistry.     

Fumigant and Contact Toxicities of Monoterpenes to <Emphasis Type="Italic">Sitophilus oryzae</Emphasis> (L.) and <Emphasis Type="Italic">Tribolium castaneum</Emphasis> (Herbst) and their Inhibitory Effects on Acetylcholinesterase Activity

A comparative study was conducted to assess the contact and fumigant toxicities of eleven monoterpenes on two important stored products insects—, Sitophilus oryzae, the rice weevil, and Tribolium castaneum, the rust red flour beetle. The monoterpenes included: camphene, (+)-camphor, (−)-carvone, 1-8-cineole, cuminaldehyde, (l)-fenchone, geraniol, (−)-limonene, (−)-linalool, (−)-menthol, and myrcene. The inhibitory effect of these compounds on acetylcholinesterase (AChE) activity also was examined to explore their possible mode(s) of toxic action. Although most of the compounds were toxic to S. oryzae and T. castaneum, their toxicity varied with insect species and with the bioassay test. In contact toxicity assays, (−)-carvone, geraniol, and cuminaldehyde showed the highest toxicity against S. oryzae with LC₅₀ values of 28.17, 28.76, and 42.08 μg/cm², respectively. (−)-Carvone (LC₅₀ = 19.80 μg/cm²) was the most effective compound against T. castaneum, followed by cuminaldehyde (LC₅₀ = 32.59 μg/cm²). In contrast, camphene, (+)-camphor, 1-8-cineole, and myrcene had weak activity against both insects (i.e., LC₅₀ values above 500 μg/cm²). In fumigant toxicity assays, 1-8-cineole was the most effective against S. oryzae and T. castaneum (LC₅₀ = 14.19 and 17.16 mg/l, respectively). Structure-toxicity investigations revealed that (−)-carvone—, a ketone—, had the highest contact toxicity against the both insects. 1-8-Cineole—, an ether—, was the most potent fumigant against both insects. In vitro inhibition studies of AChE from adults of S. oryzae showed that cuminaldehyde most effectively inhibited enzyme activity at the two tested concentrations (0.01 and 0.05 M) followed by 1-8-cineole, (−)-limonene, and (l)-fenchone. 1-8-Cineole was the most potent inhibitor of AChE activity from T. castaneum larvae followed by (−)-carvone and (−)-limonene. The results of the present study indicate that (−)-carvone, 1,8-cineole, cuminaldehyde, (l)-fenchone, and (−)-limonene could be effective biocontrol agents against S. oryzae and T. castaneum.     

Identification of Host Attractants for the Ethiopian Fruit Fly, <Emphasis Type="Italic">Dacus ciliatus</Emphasis> Loew

The Ethiopian fruit fly, Dacus ciliatus, is an oligophagous pest of cucurbit crops, particularly melons, cucumbers, and marrows (summer squash). The present study aimed to identify host attractants for D. ciliatus and was guided by a behavioral bioassay and an electrophysiological assay. We tested volatile compounds from the fruits of a host plant, ripe and unripe Galia melon, Cucumis melo var. reticulates. Both sexes were attracted to melon volatiles. Those of ripe melon were preferred. Gas chromatography-electroantennographic detection analysis of the behaviorally active ripe melon volatiles consistently showed that 14 compounds elicited similar antennal responses from both sexes. Twelve compounds were identified by gas chromatography-mass spectrometry (GC-MS) using GC-MS libraries, retention indices (RI), and authentic standards. The electrophysiological activities of the compounds that were present at sufficient levels for identification, benzyl acetate, hexanyl acetate, (Z)-3-hexenyl acetate, (Z)-3-octenyl acetate, octanyl acetate, (Z)-3-decenyl acetate, and (E)-β-farnesene, were evaluated at six different dosage levels by using electroantennography (EAG). Benzyl and hexanyl acetates elicited dose responses only in males, while other tested compounds elicited dose responses in both sexes. The strongest responses were observed for doses between 100 ng and 10 μg. The dose response, in terms of attractiveness to synthetic compounds within the active range (as determined by EAG), also was evaluated in the behavioral bioassay. Synthetic acetates were attractive to both sexes when tested individually. Significant attraction was observed when individual compounds were applied in the bioassay arena at doses of 0.5–1 μg/dispenser. Blends of compounds in equal proportions also were attractive to the insects. The most attractive blend was a mixture of four or five identified acetates. The addition of an equal proportion of (E)-β-farnesene to this mixture had a deterrent effect.     

Sex Pheromone Components of the Pear Fruit Moth, <Emphasis Type="Italic">Acrobasis pyrivorella</Emphasis> (Matsumura)

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

Larval Beetles Form a Defense from Recycled Host-Plant Chemicals Discharged as Fecal Wastes

Larvae of the leaf-feeding beetles Neolema sexpunctata and Lema trilinea carry feces on their backs that form shields. We used the generalist predatory ant, Formica subsericea, in a bioassay to determine whether shields were a physical barrier or functioned as a chemical defense. Fecal shields protected both species against ant attack. Larvae of both species reared on lettuce produced fecal shields that failed to deter ants. Commelina communis, N. sexpunctata's host, lacks noxious secondary compounds but is rich in phytol and fatty acids, metabolites of which become incorporated into the fecal defense. In contrast, the host plant of L. trilinea, Solanum dulcamara, contains steroidal glycoalkaloids and saponins, whose partially deglycosylated metabolites, together with fatty acids, appear in Lema feces. Both beetle species make modifications to host-derived precursors before incorporating the metabolites into shields. Synthetic chemicals identified as shield metabolites were deterrent when applied to baits. This study provides experimental evidence that herbivorous beetles form a chemical defense by the elimination of both primary and secondary host-derived compounds. The use of host-derived compounds in waste-based defenses may be a more widely employed strategy than was hitherto recognized, especially in instances where host plants lack elaborate secondary compounds.     

Differential Attractiveness of Potato Tuber Volatiles to <Emphasis Type="Italic">Phthorimaea operculella</Emphasis> (Gelechiidae) and the Predator <Emphasis Type="Italic">Orius insidiosus</Emphasis> (Anthocoridae)

The behavioral responses of the potato tuberworm moth Phthorimaea operculella and the polyphagous predator Orius insidiosus to volatiles emanating from exposed tubers were studied by four-arm olfactometer bioassays. Mated females of P. operculella distinguished volatiles released by intact potato tubers from volatiles damaged mechanically or by conspecific larvae. Volatiles from intact potato tubers were attractive to them. On the other hand, unmated females of P. operculella did not respond to tuber volatiles. Adults of O. insidiosus were attracted to volatiles from tubers damaged by P. operculella larvae, but did not respond to intact or mechanically damaged tubers. Methyl jasmonate (MeJA) was the only compound identified from the headspace of potato tubers (GC-MS of direct headspace sampling). The amount varied with the type of induction, being 0.001 ± 0.0003 ng g⁻¹ in tissues of intact fresh tubers, 0.002 ± 0.0007 ng g⁻¹ in mechanically damaged tubers, and showing a six- to tenfold increase in P. operculella damaged tubers (0.090 ± 0.006 ng g⁻¹). Behavioral bioassays with synthetic MeJA confirmed that the response of the insects is dependent on MeJA concentration. Mated females of P. operculella showed the highest response at 0.001 ng g⁻¹ (concentration released by intact tubers), whereas O. insidiosus showed the highest response, between 0.01 and 0.05 ng g⁻¹, which is close to the concentration released by P. operculella damaged tubers. Based on these results, we postulate that P. operculella and O. insidiosus have adapted their responses to plant volatiles differently, enabling them to locate suitable hosts or prey.     

Flavonoid glycosides and naphthodianthrones in the sawfly Tenthredo zonula and its host-plants, Hypericum perforatum and H. hirsutum

Larvae of the sawfly Tenthredo zonula are specialized on Hypericum. Whether the sawfly is able to sequester plant metabolites was unknown. Aerial materials of Hypericum perforatum and H. hirsutum, as well as dissected larvae and prepupae of T. zonula, were analyzed by HPLC to determine the presence and content of flavonoid glycosides (rutin, hyperoside, isoquercitrin, and quercitrin) and naphthodianthrones (pseudohypericin and hypericin). All flavonoid glycosides were detected in both Hypericum species, with hyperoside as major compound in H. perforatum (ca. 1.7 μmol/g fresh weight, FW) and isoquercitrin in H. hirsutum (0.7 μmol/g FW). Naphthodianthrones were present at low concentrations (0.02 μmol/g FW) in the former, and almost undetected in the latter species. In the body parts (i.e., hemolymph, digestive tract, salivary glands, or miscellaneous organs) of T. zonula, the surveyed compounds were detected more frequently in prepupae than in larvae. The compounds were not present in every sample, and flavonoid glycosides especially occurred in highly variable amounts, with maximal concentrations of 41 μg rutin/prepupa in salivary glands, 8 μg hyperoside/prepupa in hemolymph (= 0.36 μmol/g FW), 32 μg isoquercitrin/prepupa in salivary glands, and 63 μg quercitrin/larva in miscellaneous organs (mainly composed of the integument). We conclude that flavonoid glycosides are sequestered since they were detected in organs other than the digestive tract of larvae, and because prepupae are a non-feeding stage. The naphthodianthrone pseudohypericin, but not hypericin, occurred generally in the digestive tract (up to 0.25 μg/larva). Both naphthodianthrones and related unidentified compounds, but not flavonoid glycosides, were found in the larval excrement. The highly variable distributions of flavonoid glycosides and naphthodianthrones in T. zonula larvae and prepupae make it difficult to determine the ecological significance of these metabolites.     

Sequestration ofVeratrum alkaloids by specialistRhadinoceraea nodicornis konow (Hymenoptera,Tenthredinidae) and its ecoethological implications

The larvae of the specialist sawflyRhadinoceraea nodicornis Konow (Hymenoptera, Tenthredinidae) store in their hemolymph ceveratrum alkaloids originating from the host plantVeratrum album L. (Liliales, Melanthiaceae). The major alkaloid found in the hemolymph is 3-acetyl-zygadenine. Qualitative and quantitative data showed that the plant alkaloid 3-angeloylzygadenine is most probably metabolized in the larval gut to zygadenine and then acetylated. A still unidentified alkaloid with a molecular weight of 591 Da was detected in plant leaves as well as in the gut, hemolymph, and excrement of larvae. Protoveratrine A and B, on the other hand, seem to be degraded by the larvae. These findings indicate that the pathway of ceveratrum alkaloids inR. nodicornis larvae is fourfold: direct sequestration, metabolism followed by sequestration, excretion of intact alkaloids, and degradation. In contrast, no ceveratrum alkaloids were detected in the hemolymph and excrement of larvae of the generalist sawflyAglaostigma sp. fed withV. album leaves. Bioassays with the antMyrmica rubra L. proved that the hemolymph ofR. nodicornis larvae is highly deterrent and toxic. In bioassays evaluating defensive efficiency against predators (ants, spiders, and bushcrickets), no larvae were eaten. Ceveratrum alkaloids were also detected in the hibernating prepupae ofR. nodicornis. In feeding bioassays, the shrewCrocidura russula Hermann rarely fed upon prepupae, suggesting that this stage is also protected from predation to some degree. In field surveys, the only parasitoids recorded were two ichneumonid species that are believed to be specialized onR. nodicornis. Bioassays and field observations enable us to suppose thatR. nodicornis and its enemies produce a food web of ion connectance.     

Vestitol: A phytoalexin with insect feeding-deterrent activity

A major feeding deterrent forCostelytra zealandica larvae was isolated from the root of the resistant pasture legumeLotus pedunculatus and was identified as 3R-(—)-vestitol. This compound was also identified in feeding deterrent-activeL. pedunculatus leaf extracts. (—)-Vestitol and a secondLotus isoflavan, sativan, have been reported to have phytoalexin activity, and the implications of this for the study and understanding of insect resistance are briefly discussed.     

Identification of Sex Pheromone Components of the Hessian Fly, <Emphasis Type="Italic">Mayetiola destructor</Emphasis>

Coupled gas chromatographic (GC)–electroantennographic detection (EAD) analyses of ovipositor extract of calling Hessian fly, Mayetiola destructor, females revealed that seven compounds elicited responses from male antennae. Four of the compounds—(2S)-tridec-2-yl acetate, (2S,10Z)-10-tridecen-2-yl acetate, (2S,10E)-10-tridecen-2-yl acetate, and (2S,10E)-10-tridecen-2-ol—were identified previously in female extracts. Two new EAD-active compounds, (2S,8Z,10E)-8,10-tridecadien-2-yl acetate and (2S,8E,10E)-8,10-tridecadien-2-yl acetate, were identified by GC–mass spectroscopy (MS) and the use of synthetic reference samples. In a Y-tube bioassay, a five-component blend (1 ng (2S)-tridec-2-yl acetate, 10 ng (2S,10E)-10-tridecen-2-yl acetate, 1 ng (2S,10E)-10-tridecen-2-ol, 1 ng (2S,8Z,10E)-8,10-tridecadien-2-yl acetate, and 1 ng (2S,8E,10E)-8,10-tridecadien-2-yl acetate) was as attractive to male Hessian flies as a similar amount of female extract (with respect to the main compound, (2S,10E)-10-tridecen-2-yl acetate). The five-component blend was more attractive to male flies than a three-component blend lacking the two dienes. Furthermore, the five-component blend was more attractive than a blend with the same compounds but that contained one tenth the concentration of (2S,8E,10E)-8,10-tridecadien-2-yl acetate (more accurately mimicking the ratios found in female extract). This suggests that the ratios emitted by females might deviate from those in gland extracts. In a field-trapping experiment, the five-component blend applied to polyethylene cap dispensers in a 100:10 μg ratio between the main component and each of the other blend components attracted a significant number of male Hessian flies. Also, a small-plot field test demonstrated the attractiveness of the five-component blend to male Hessian flies and suggests that this pheromone blend may be useful for monitoring and predicting Hessian fly outbreaks in agricultural systems.     

Isolation of Three Diterpenoid Acids from Sunflowers,as Oviposition Stimulants for the Banded Sunflower Moth, <Emphasis Type="Italic">Cochylis hospes</Emphasis>

The banded sunflower moth (BSFM), Cochylis hospes Walshingham (Lepidoptera: Cochylidae) is a specialist insect, the larvae of which feed on sunflowers, Helianthus spp., and a few other species of Compositae. It is one of the most important pests of sunflower in the USA. Previous work on H. annuus, the cultivated sunflower, revealed two diterpenoids that function as oviposition stimulants for female BSFM, and that other, more polar compounds also stimulated oviposition. Using a bioassay-guided approach, we isolated three additional diterpenoids, grandifloric acid (1), 15β-hydroxy-ent-trachyloban-19-oic acid (2), and 17-hydroxy-16α-ent-kauran-19-oic acid (3), from polar fractions of pre-bloom sunflower head extracts. In laboratory bioassays, purified natural samples of each of these compounds stimulated oviposition by female BSFM. Structure–activity relationships of the five diterpenoids known to stimulate oviposition by female BSFM are discussed.     

Behavioral and Physiological Effects of Neem (Azadirachta indica) Seed Kernel Extracts on Larval Parasitoid, Bracon hebetor

An aqueous suspension and an ethanolic extract of neem seed kernel (NSK) at 0.3, 0.6, 1.2, 2.5, and 5.0% concentrations were tested for ovipositional deterrency, feeding deterrency, toxicity, sterility, and insect growth regulatory effects on a larval parasitoid, Bracon hebetor. Neither NSK extracts delivered in the food or by contact influenced the B. hebetor oviposition (parasitization). They also did not cause parasitoid sterility through feeding, but they showed feeding deterrent effects for a limited period. Parasitoid eggs and pupae also were unaffected by the extracts tested. The parasitoid larvae, however, were killed by feeding contaminated host larvae and also through contact with neem extracts. Thus, a minimum safety period is suggested for inundative releases of B. hebetor in integrated pest management.     

Antimicrobial Activity of Essential Oils Isolated from Phlomis crinita Cav. ssp. mauritanica Munby

The essential oil extracted from the leaves and flowers of Phlomis crinita Cav. ssp. mauritanica Munby were obtained by steam distillation and analyzed by gas chromatography coupled with mass spectrometry. The major constituents of flower oil were β-caryophyllene (58.1%) and germacrene D (35.1%). This oil inhibited the growth of Staphylococcus aureus, Enterococcus faecalis, and Salmonella typhimurium with minimal inhibition concentrations (MIC) varying between 39 and 625 μg/ml. The essential oil obtained from the leaves was mainly composed of trans-caryophyllene (40.8%) and germacrene D (39.1%) and exhibited an antimicrobial profile against the same strains mentioned above with MIC between 156 μg/ml and 2.5 mg/ml.     

Fate of quinolizidine alkaloids through three trophic levels:Laburnum anagyroides (Leguminosae) and associated organisms

The quinolizidine alkaloids (QA) of golden rain,Laburnum anagyroides, and those of phytophagous insects associated with the plant, as well as of parasitoids of the latter, were analyzed by capillary GLC and GLC-MS. The alkaloid content in samples of vegetative plant parts was high at the beginning of the season, then decreased, while that of reproductive organs was high throughout flowering, pod formation, and maturation. The analyses showed that the QA of the plant passed through two higher trophic levels (herbivorous insects and their parasitoids) and that the alkaloid pattern changed little during the passage. The alkaloids were present in two phytophagous insect species associated with golden rain: the predispersal seed predator,Bruchidius villosus [5–13g/g fresh weight (fw)], andAphis cytisorum (182–1012g/g fw), an aphid that feeds on shoots, leaves, and inflorescences. Braconid and chalcidoid parasitoids emerging from the bruchid host also contained alkaloids (1.3–3g/g fw), as did three foraging ant species,Lasius niger, Formica rufibarbis, andF. cunicularia (45g/g fw), that visited the aphid colonies or honeydew-covered leaves of aphid-infested plants. The hypothesis that developing bruchid larvae and/or the plant manipulate QA supply to infested seeds was not supported, because QA content of leftover endosperm in seeds after bruchid development was similar to that of uninfested seeds. The frass of developing bruchid larvae was rich in QA (31g/ g dry weight). While aphids sequestered, the bruchid larvae took up and eliminated QA with the frass without chemical transformation.     

Amphibian Chemical Defense: Antifungal Metabolites of the Microsymbiont <Emphasis Type="Italic">Janthinobacterium lividum</Emphasis> on the Salamander <Emphasis Type="Italic">Plethodon cinereus</Emphasis>

Disease has spurred declines in global amphibian populations. In particular, the fungal pathogen Batrachochytrium dendrobatidis has decimated amphibian diversity in some areas unaffected by habitat loss. However, there is little evidence to explain how some amphibian species persist despite infection or even clear the pathogen beyond detection. One hypothesis is that certain bacterial symbionts on the skin of amphibians inhibit the growth of the pathogen. An antifungal strain of Janthinobacterium lividum, isolated from the skin of the red-backed salamander Plethodon cinereus, produces antifungal metabolites at concentrations lethal to B. dendrobatidis. Antifungal metabolites were identified by using reversed phase high performance liquid chromatography, high resolution mass spectrometry, nuclear magnetic resonance, and UV-Vis spectroscopy and tested for efficacy of inhibiting the pathogen. Two metabolites, indole-3-carboxaldehyde and violacein, inhibited the pathogen’s growth at relatively low concentrations (68.9 and 1.82 μM, respectively). Analysis of fresh salamander skin confirmed the presence of J. lividum and its metabolites on the skin of host salamanders in concentrations high enough to hinder or kill the pathogen (51 and 207 μM, respectively). These results support the hypothesis that cutaneous, mutualistic bacteria play a role in amphibian resistance to fungal disease. Exploitation of this biological process may provide long-term resistance to B. dendrobatidis for vulnerable amphibians and serve as a model for managing future emerging diseases in wildlife populations. Electronic supplementary material  The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

Temperature and Spatiotemporal Variability of Salicylihalamide A in the Sponge <Emphasis Type="Italic">Haliclona</Emphasis> sp.

The production of salicylihalamide A by the marine sponge Haliclona sp. was investigated. Samples of the two morphologies (green and brown) were collected from four locations covering approximately 1,200 km of coastline. Temporal variation between winter and summer was also examined at Bremer Bay. Chemical profiling by using liquid chromatography coupled with ultra violet detection and mass spectrometry showed that salicylihalamide A was produced only by the green morphology. Salicylihalamide A concentration was significantly correlated to water temperature but not to the size or depth of the sponge. Salicylihalamide A concentration was found to differ significantly among locations (Bremer Bay 13.5 μg g⁻¹, Hamelin Bay 11 μg g⁻¹, Rottnest Island 9.9 μg g⁻¹, and Jurien Bay 8.5 μg g⁻¹) partially accounted for by the influence of water temperature. A difference between seasons was also observed in Bremer Bay (summer concentration of 13.5 μg g⁻¹ vs. winter concentration of 8.2 μg g⁻¹). Environmental and physiological factors appear to be important in the production of salicylihalamide A by the green morphology. Additionally, the brown morphology does not produce salicylihalamide A, thus adding to the evidence that this morphology may be a different species.     

Tolerance of <Emphasis Type="Italic">Drosophila</Emphasis> Flies to Ibotenic Acid Poisons in Mushrooms

The mushroom genus Amanita has a spectrum of chemical compounds affecting survival and performance of animals. Ibotenic acid is one of such compounds found in some Amanita mushrooms. We studied the effects of ibotenic acid and its derivative, muscimol, on egg-to-pupa survival, pupation time, and pupal size in five Drosophila species (Diptera: Drosophilidae), Drosophila bizonata, Drosophila angularis, Drosophila brachynephros, Drosophila immigrans, and Drosophila melanogaster. The first three species are mycophagous and use a wide range of mushrooms for breeding, whereas D. immigrans and D. melanogaster are frugivorous. We reared fly larvae on artificial medium with 500, 250, 125, and 62.5 μg/ml of ibotenic acid and/or musimol. The three mycophagous species were not susceptible to ibotenic acid, whereas the two frugivorous species were affected. In experiments with D. melanogaster, muscimol was less toxic than ibotenic acid.     

Responses of maxillary sensilla styloconica inBombyx mori to glucosides fromOsmunda japonica,a nonhost plant

We isolated glucosides from the royal fern,Osmunda japonica, which elicit a deterrent response in larvae ofBombyx mori. These compounds were absent in taro (Colocasia antiquorum) and castor-oil plant (Ricinus communis) leaves and did not evoke responses of sensory cells in the lateral and medial sensilla styloconica ofSpodoptera litura. This glucoside extract of the royal fern leaves stimulates receptors generally associated with deterrent. It is also possible that this compound may act as a behavioral deterrent, and from actual feeding tests, it is suggested that this compound may prevent feeding in some monophagous insects, such asBombyx mori. The deterrent glucoside possesses a noncyclic aglycon.