Diversity of the Enzymatic Activity in the Lipoxygenase Gene Family of <Emphasis Type="Italic">Arabidopsis thaliana</Emphasis>

Lipoxygenases (LOX) catalyze the oxygenation of polyunsaturated fatty acids, the first step in the biosynthesis of a large group of biologically active fatty acid metabolites collectively named oxylipins. In the present study we report the characterization of the enzymatic activity of the six lipoxygenases found in the genome of the model plant Arabidopsis thaliana. Recombinant expressed AtLOX-1 and AtLOX-5 had comparable oxygenase activity with either linoleic acid or linolenic acid. AtLOX-2, AtLOX-3, AtLOX-4 and AtLOX-6 displayed a selective oxygenation of linolenic acid. Analyses by high-performance liquid chromatography and gas chromatography-mass spectrometry demonstrated that AtLOX-1 and AtLOX-5 are 9S-lipoxygenases, and AtLOX-2, AtLOX-3, AtLOX-4 and AtLOX-6 are 13S-lipoxygenases. None of the enzymes had dual positional specificity. The determined activities correlated with that predicted by their phylogenetic relationship to other biochemically-characterized plant lipoxygenases. Gerard Bannenberg, Marta Martínez shared first authors.     

Geographic Variation in Alkaloid Production in <Emphasis Type="Italic">Conium maculatum</Emphasis> Populations Experiencing Differential Herbivory by <Emphasis Type="Italic">Agonopterix alstroemeriana</Emphasis>

Conium maculatum, a Eurasian weed naturalized in North America, contains high concentrations of piperidine alkaloids that act as chemical defenses against herbivores. C. maculatum was largely free from herbivory in the United States, until approximately 30 yr ago, when it was reassociated via accidental introduction with a monophagous European herbivore, the oecophorid caterpillar Agonopterix alstroemeriana. At present, A. alstroemeriana is found in a continuum of reassociation time and intensities with C. maculatum across the continent; in the Pacific Northwest, A. alstroemeriana can cause severe damage, resulting in some cases in complete defoliation. Studies in biological control and invasion biology have yet to determine whether plants reassociated with a significant herbivore from the area of indigeneity increase their chemical defense investment in areas of introduction. In this study, we compared three locations in the United States (New York, Washington, and Illinois) where C. maculatum experiences different levels of herbivory by A. alstroemeriana to determine the association between the intensity of the interaction, as measured by damage, and chemical defense production. Total alkaloid production in C. maculatum was positively correlated with A. alstroemeriana herbivory levels: plants from New York and Washington, with higher herbivory levels, invested two and four times more N to alkaloid synthesis than did plants from Illinois. Individual plants with lower concentrations of alkaloids from a single location in Illinois experienced more damage by A. alstroemeriana, indicative of a preference on the part of the insect for plants with less chemical defense. These results suggest that A. alstroemeriana may act either as a selective agent or inducing agent for C. maculatum and increase its toxicity in its introduced range.     

Within-Plant Variation In Glucosinolate Concentrations of <Emphasis Type="Italic">Raphanus sativus</Emphasis> Across Multiple Scales

Variation in chemical defenses remains underexplored. In particular, little is known about patterns of variation at small scales within leaves and spatial variation of induction. I examined variation in glucosinolate concentrations in the leaves of Raphanus sativus at several different spatial scales in two related experiments. I used samples equivalent in area to the amount an intermediate-sized caterpillar might eat in 1 d, a smaller scale than used in most previous studies. I examined variation due to induction and leaf age and small-scale spatial variation within leaves. The mean and variance of glucosinolate concentrations were higher in induced plants, young leaves, and the proximal half of leaves. Higher glucosinolate concentrations in the proximal half of leaves are previously unreported. Small-scale variation was extreme, accounting for 57% of the total random variation, and spatially random. There was no spatial autocorrelation found at scales as small as 1–2 cm. The high degree of small-scale, spatially random variation in glucosinolate concentrations in leaves is previously unreported. This small-scale variation and the variation caused by induction may have significant effects on herbivores and could be an important component of plant defense.     

<Emphasis Type="Italic">Arabidopsis thaliana:</Emphasis> A New Test Species for Phytotoxic Bioassays

Lettuce seeds (Lactuca sativa L.) and other crop species are often used in phytotoxic bioassays that are designed to detect allelochemicals. The seeds of these species are considered ideal because they are readily available, germinate rapidly and uniformly, and are routinely used in laboratories around the world. Despite their common use, however, the seeds of these species are often not as sensitive or responsive to some phytotoxic chemicals as are the seeds of other species. While searching for a more sensitive test species for phytotoxic bioassays, the Columbia ecotype of Arabidopsis thaliana exhibited greater sensitivity to seven potent allelochemicals than did lettuce seeds, which, in some cases, did not respond at all to those substances. Sensitivity satisfies one of the criteria for selecting a test species for bioassays. We now report on the results of our study and offer additional reasons for using A. thaliana seeds.     

The Desert Locust, <Emphasis Type="Italic">Schistocerca gregaria</Emphasis>, Detoxifies the Glucosinolates of <Emphasis Type="Italic">Schouwia purpurea</Emphasis> by Desulfation

Desert locusts (Schistocerca gregaria) occasionally feed on Schouwia purpurea, a plant that contains tenfold higher levels of glucosinolates than most other Brassicaceae. Whereas this unusually high level of glucosinolates is expected to be toxic and/or deterrent to most insects, locusts feed on the plant with no apparent ill effects. In this paper, we demonstrate that the desert locust, like larvae of the diamondback moth (Plutella xylostella), possesses a glucosinolate sulfatase in the gut that hydrolyzes glucosinolates to their corresponding desulfonated forms. These are no longer susceptible to cleavage by myrosinase, thus eliminating the formation of toxic glucosinolate hydrolysis products. Sulfatase is found throughout the desert locust gut and can catalyze the hydrolysis of all of the glucosinolates present in S. purpurea. The enzyme was detected in all larval stages of locusts as well as in both male and female adults feeding on this plant species. Glucosinolate sulfatase activity is induced tenfold when locusts are fed S. purpurea after being maintained on a glucosinolate-free diet, and activity declines when glucosinolates are removed from the locust diet. A detoxification system that is sensitive to the dietary levels of a plant toxin may minimize the physiological costs of toxin processing, especially for a generalist insect herbivore that encounters large variations in plant defense metabolites while feeding on different species.     

Phytotoxic Effects of 21 Plant Secondary Metabolites on <Emphasis Type="Italic">Arabidopsis thaliana</Emphasis> Germination and Root Growth

This study investigated potential phytotoxic effects on germination and root growth of 21 plant secondary metabolites (sinapinic, syringic, vanillic, ferulic, p-coumaric, chlorogenic, gallic, gentisic, protocatechuic, p-hydroxybenzoic, and trans-cinnamic acids, and eucalyptol, quercetin, vanillin, syringaldehyde, rutin, 2-benzoxazolinone, protocatechualdehyde, tyrosol, juglone, and l-mimosine) in the plant model Arabidopsis thaliana. Eleven of the 21 molecules showed significant inhibitory effects on germination, and 17 inhibited root growth. Inhibitory effects on root growth were more evident when nutrients were not added. We present dose–response curves for germination effects and IC₅₀ values for each compound, along with possible explanations of the observed inhibitory actions in terms of molecular structure.     

Interspecific Variation Within the Genus <Emphasis Type="Italic">Asclepias</Emphasis> in Response to Herbivory by a Phloem-feeding Insect Herbivore

Induced plant responses to leaf-chewing insects have been well studied, but considerably less is known about the effects of phloem-feedings insects on induction. In a set of laboratory experiments, we examined density-dependent induction by the milkweed-oleander aphid, Aphis nerii, of putative defenses in four milkweed species (Asclepias incarnata, Asclepias syriaca, Asclepias tuberosa, and Asclepias viridis). We hypothesized that high aphid density would lead to increased cardenolide expression in species with low constitutive levels of cardenolides (e.g., A. tuberosa), but that there would be no induction in high constitutive cardenolide species (e.g., A. viridis). Based on previous studies, we did not expect cardenolide induction in A. incarnata. Contrary to our predictions, we observed feeding-induced declines of cardenolide concentrations in A. viridis. Cardenolide concentrations did not respond to aphid feeding in the other three milkweed species. Aphids also caused reductions in biomass accumulation by two of four Asclepias species, A. viridis and A. incarnata. High aphid density led to a decrease in A. viridis foliar nitrogen concentration. However, aphids had no effect on the defensive chemistry, growth, or nutritional quality of either A. syriaca or A. tuberosa. Our results highlight that congeneric plant species may respond differently to the same levels of herbivore damage.     

Effect of Volatile Constituents from <Emphasis Type="Italic">Securidaca</Emphasis><Emphasis Type="Italic">Longepedunculata</Emphasis> on Insect pests Of Stored Grain

Securidaca longepedunculata Fers (Polygalaceae) is commonly used as a traditional medicine in many parts of Africa as well as against a number of invertebrate pests, including insects infesting stored grain. The present study showed that S. longepedunculata root powder, its methanol extract, and the main volatile component, methyl salicylate, exhibit repellent and toxic properties to Sitophilus zeamais adults. Adult S. zeamais that were given a choice between untreated maize and maize treated with root powder, extract, or synthetic methyl salicylate in a four-way choice olfactometer significantly preferred the control maize. Methyl salicylate vapor also had a dose-dependant fumigant effect against S. zeamais, Rhyzopertha dominica, and Prostephanus truncates, with a LD₁₀₀ achieved with a 60 l dose in a 1-l container against all three insect species after 24 hr of exposure. Probit analyses estimated LD₅₀ values between 34 and 36 l (95% CI) for all insect species. Furthermore, prolonged exposure for 6 days showed that lower amounts (30 l) of methyl salicylate vapor were able to induce 100% adult mortality of the three insect species. The implications are discussed in the context of improving stored product pest control by small-scale subsistence farmers in Africa.     

Relationship Between the Endophyte <Emphasis Type="Italic">Embellisia</Emphasis> spp. and the Toxic Alkaloid Swainsonine in Major Locoweed Species (<Emphasis Type="Italic">Astragalus</Emphasis> and <Emphasis Type="Italic">Oxytropis)</Emphasis>

Locoweeds (Astragalus and Oxytropis spp. that contain the toxic alkaloid swainsonine) cause widespread poisoning of livestock on western rangelands. There are 354 species of Astragalus and 22 species of Oxytropis in the US and Canada. Recently, a fungal endophyte, Embellisia spp., was isolated from Astragalus and Oxytropis spp. and shown to produce swainsonine. We conducted a survey of the major locoweeds from areas where locoweed poisoning has occurred to verify the presence of the endophyte and to relate endophyte infection with swainsonine concentrations. Species found to contain the fungal endophyte and produce substantial amounts of swainsonine were A. wootoni, A. pubentissimus, A. mollissimus, A. lentiginosus, and O. sericea. Astragalus species generally had higher concentrations of swainsonine than Oxytropis. Swainsonine was not detected in A. alpinus, A. cibarius, A. coltonii, A. filipes, or O. campestris. The endophyte could not be cultured from A. mollissimus var. thompsonii or A. amphioxys, but was detected by polymerase chain reaction, and only 30% of these samples contained trace levels of swainsonine. Further research is necessary to determine if the endophyte is able to colonize these and other species of Astragalus and Oxytropis and determine environmental influences on its growth and synthesis of swainsonine.     

<Emphasis Type="Italic">Santalum </Emphasis><Emphasis Type="Italic">insulare</Emphasis> Acetylenic Fatty Acid Seed Oils: Comparison within the <Emphasis Type="Italic">Santalum</Emphasis> Genus

The sandalwood kernels of Santalum insulare (Santalaceae) collected in French Polynesia give seed oils containing significant amounts of ximenynic acid, E-11-octadecen-9-oic acid (64–86%). Fatty acid (FA) identifications were performed by gas chromatography/mass spectrometry (GC/MS) of FA methyl esters. Among the other main eight identified fatty acids, oleic acid was found at a 7–28% level. The content in stearolic acid, octadec-9-ynoic acid, was low (0.7–3.0%). An inverse relationship was demonstrated between ximenynic acid and oleic acid using 20 seed oils. Results obtained have been compared to other previously published data on species belonging to the Santalum genus, using multivariate statistical analysis. The relative FA S. insulare composition, rich in ximenynic acid is in the same order of those given for S. album or S. obtusifolium. The other compared species (S. acuminatum, S. lanceolatum, S. spicatum and S. murrayanum) are richer in oleic acid (40–59%) with some little differences in linolenic content.     

Laboratory Evaluation of <Emphasis Type="Italic">Artemisia annua</Emphasis> L. Extract and Artemisinin Activity against <Emphasis Type="Italic">Epilachna paenulata</Emphasis> and <Emphasis Type="Italic">Spodoptera eridania</Emphasis>

Ethanolic extract of aerial parts of Artemisia annua L. and artemisinin were evaluated as anti-insect products. In a feeding deterrence assay on Epilachna paenulata Germ (Coleoptera: Coccinellidae) larvae, complete feeding rejection was observed at an extract concentration of 1.5 mg/cm² on pumpkin leaf tissue. The same concentration produced a feeding inhibition of 87% in Spodoptera eridania (Cramer) (Lepidoptera: Noctuidae). In a no-choice assay, both species ate less and gained less weight when fed on leaves treated with the extract. Complete mortality in E. paenulata and 50% mortality in S. eridania were observed with extract at 1.5 mg/cm². Artemisinin exhibited a moderate antifeedant effect on E. paenulata and S. eridania at 0.03–0.375 mg/cm². However, a strong effect on survival and body weight was observed when E. paenulata larvae were forced to feed on leaves treated at 0.03 and 0.075 mg/cm². Artemisia annua ethanolic extract of aerial parts at 1.5 mg/cm² showed no phytotoxic effect on pumpkin seedlings.     

Chemotaxis of the Pinewood Nematode, <Emphasis Type="Italic">Bursaphelenchus xylophilus</Emphasis>, to Volatiles Associated with Host Pine, <Emphasis Type="Italic">Pinus massoniana</Emphasis>, and its Vector <Emphasis Type="Italic">Monochamus alternatus</Emphasis>

The pinewood nematode (PWN), Bursaphelenchus xylophilus, the most important invasive species in pine forests of Asia, is transported to new pine hosts by beetles of the genus Monochamus. Third-stage dispersal juveniles (J_III) aggregate in pupal chambers around the vector as it matures. We demonstrated that the ratio of three terpenes (α-pinene, β-pinene, and longifolene at 1:2.7:1.1) released by larval Monochamus alternatus strongly attract J_III, whereas the different ratio (1:0.1:0.01) of these three terpenes found in healthy xylem of Pinus massoniana attracts only the propagative stage (J_n) of the nematode. We suggest that the volatiles produced by the host plants could be the basis of a chemoecological relationship between plant parasitic nematodes and their vector insects. Capture of J_III with terpene-baited trap tubes deployed for 2 hr in the field was demonstrated. This technique may lead to the development of rapid sampling methodologies for use at either ports-of-entry or in the field.     

Plant Growth Inhibition By <Emphasis Type="Italic">Cis</Emphasis>-Cinnamoyl Glucosides and <Emphasis Type="Italic">Cis</Emphasis>-Cinnamic Acid

Spiraea thunbergii Sieb. contains 1-O-cis-cinnamoyl--_d-glucopyranose (CG) and 6-O-(4-hydroxy-2-methylene-butyroyl)-1-O-cis-cinnamoyl--_d-glucopyranose (BCG) as major plant growth inhibiting constituents. In the present study, we determined the inhibitory activity of CG and BCG on root elongation of germinated seedlings of lettuce (Lactuca sativa), pigweed (Amaranthus retroflexus), red clover (Trifolium pratense), timothy (Phleum pratense), and bok choy (Brassica rapa var chinensis) in comparison with that of two well-known growth inhibitors, 2,4-dichlorophenoxyacetic acid (2,4-D) and (+)-2-cis-4-trans-abscisic acid (cis-ABA), as well as two related chemicals of CG and BCG, cis-cinnamic acid (cis-CA) and trans-cinnamic acid (trans-CA). The EC₅₀ values for CG and BCG on lettuce were roughly one-half to one-quarter of the value for cis-ABA. cis-Cinnamic acid, which is a component of CG and BCG, possessed almost the same inhibitory activity of CG and BCG, suggesting that the essential chemical structure responsible for the inhibitory activity of CG and BCG is cis-CA. The cis-stereochemistry of the methylene moiety is apparently needed for high inhibitory activity, as trans-CA had an EC₅₀ value roughly 100 times that of CG, BCG, and cis-CA. Growth inhibition by CG, BCG, and cis-CA was influenced by the nature of the soil in the growing medium: alluvial soil preserved the bioactivity, whereas volcanic ash and calcareous soils inhibited bioactivity. These findings indicate a potential role of cis-CA and its glucosides as allelochemicals for use as plant growth regulators in agricultural fields.     

EFFECT OF PBAN ON PHEROMONE PRODUCTION BY MATED <Emphasis Type="Italic">Heliothis virescens</Emphasis> AND <Emphasis Type="Italic">Heliothis subflexa</Emphasis> FEMALES

Mated female Heliothis virescens and H. subflexa were induced to produce sex pheromone during the photophase by injection of pheromone biosynthesis activating neuropeptide (PBAN). When injected with 1 pmol Hez-PBAN, the total amount of pheromone that could be extracted from glands of mated females during the photophase was similar to that extracted from virgin females in the scotophase. The PBAN-induced profile of pheromone components was compared between mated, PBAN-injected females and virgin females during spring and fall. Virgin females exhibited some differences in the relative composition of the pheromone blend between spring and fall, but no such temporal differences were detected in PBAN-injected, mated females. Because the temporal variation in pheromone blend composition was greater for virgin females than for PBAN-injected females, PBAN can be used to determine a females native pheromone phenotype. This procedure has the advantages that pheromone glands can be extracted during the photophase, from mated females that have already oviposited.     

Effects of Root Isoquinoline Alkaloids from <Emphasis Type="Italic">Hydrastis canadensis</Emphasis> on <Emphasis Type="Italic">Fusarium oxysporum</Emphasis> Isolated from <Emphasis Type="Italic">Hydrastis</Emphasis> Root Tissue

Goldenseal (Hydrastis canadensis L.) is a popular medicinal plant distributed widely in North America. The rhizome, rootlets, and root hairs produce medicinally active alkaloids. Berberine, one of the Hydrastis alkaloids, has shown antifungal activity. The influence of a combination of the major Hydrastis alkaloids on the plant rhizosphere fungal ecology has not been investigated. A bioassay was developed to study the effect of goldenseal isoquinoline alkaloids on three Fusarium isolates, including the two species isolated from Hydrastis rhizosphere. The findings suggest that the Hydrastis root extract influences macroconidia germination, but that only the combined alkaloids—berberine, canadine, and hydrastine—appear to synergistically stimulate production of the mycotoxin zearalenone in the Fusarium oxysporum isolate. The Hydrastis root rhizosphere effect provided a selective advantage to the Fusarium isolates closely associated with the root tissue in comparison with the Fusarium isolate that had never been exposed to Hydrastis.     

(7<Emphasis Type="Italic">Z</Emphasis>,9<Emphasis Type="Italic">E</Emphasis>)-2-Methyl-7,9-Octadecadiene: A Sex Pheromone Component of <Emphasis Type="Italic">Lymantria Bantaizana</Emphasis>

Our objective was to identify the sex pheromone of Lymantria bantaizana (Lepidoptera: Lymantriidae) whose larvae feed exclusively on walnut, Juglans spp., in China, and Japan. Coupled gas chromatographic–electroantennographic detection (GC-EAD) analyses of pheromone gland extracts revealed a single EAD-active component. Retention index calculations of this compound on four GC columns suggested that it was a methyl-branched octadecadiene with conjugated double bonds. In GC-EAD analyses of 2-methyloctadecenes, (Z)-2-methyl-7-octadecene and (E)-2-methyl-7-octadecene elicited the strongest antennal responses, suggesting that the double bond positions were at C7 and C9. In comparative GC-EAD analyses of pheromone gland extract and stereoselectively synthesized isomers (E,E; E,Z; Z,E; Z,Z) of 2-methyl-7,9-octadecadiene, the (E,Z)- and (Z,E)-isomer had retention times identical to that of the candidate pheromone, but only the latter isomer elicited strong EAD activity. Results of field experiments in Japan substantiated that (7Z,9E)-2-methyl-7,9-octadecadiene is the L. bantaizana sex pheromone, a compound previously unknown in the Lepidoptera. Detection surveys in North America for exotic Eurasian forest defoliators could include traps baited with the L. bantaizana pheromone.     

Phytotoxic and Antifungal Compounds from Two <Emphasis Type="Italic">Apiaceae</Emphasis> Species, <Emphasis Type="Italic">Lomatium californicum</Emphasis> and <Emphasis Type="Italic">Ligusticum hultenii</Emphasis>, Rich Sources of Z-ligustilide and Apiol,Respectively

The seeds of two Apiaceae species, Ligusticum hultenii and Lomatium californicum, were investigated. Preliminary bioassays indicated that methylene chloride extracts of seeds of both species contained selective phytotoxic activity against monocots and antifungal activity against Colletotrichum fragariae. Active constituents were isolated by bioassay-guided fractionation, and the structures were elucidated by NMR and GC-MS as apiol and Z-ligustilide, isolated from L. hultenii and L. californicum, respectively. Apiol and Z-ligustilide had I₅₀ values of about 80 and 600 μM, respectively, for inhibition of the growth of Lemna paucicostata. The methylene chloride (CH₂Cl₂) extracts of the seeds and the isolated and purified compounds were tested against the 2-methylisoborneol-producing cyanobacterium (blue-green alga) Oscillatoria perornata, and the green alga Selenastrum capricornutum. The CH₂Cl₂ extracts of both Apiaceae species and apiol were weakly toxic to both species of phytoplankton, while Z-ligustilide was toxic to both with a lowest complete inhibitory concentration (LCIC) of 53 μM. Seeds of L. californicum and L. hultenii were found to be rich sources of Z-ligustilide (97 mg/g of dry seed) and apiol (40 mg/g of dry seed), respectively.     

Attraction of the Parasitoid <Emphasis Type="Italic">Anagrus nilaparvatae</Emphasis> to Rice Volatiles Induced by the Rice Brown Planthopper <Emphasis Type="Italic">Nilaparvata lugens</Emphasis>

Anagrus nilaparvatae, an egg parasitoid of the rice brown planthopper Nilaparvata lugens, was attracted to volatiles released from N. lugens-infested plants, whereas there was no attraction to volatiles from undamaged plants, artificially damaged plants, or volatiles from N. lugens nymphs, female adults, eggs, honeydew, and exuvia. There was no difference in attractiveness between plants infested by N. lugens nymphs or those infested by gravid females. Attraction was correlated with time after infestation and host density; attraction was only evident between 6 and 24 hr after infestation by 10 adult females per plant, but not before or after. Similarly, after 24 hr of infestation, wasps were attracted to plants with 10 to 20 female planthoppers, but not to plants with lower or higher numbers of female planthoppers. The attractive time periods and densities may be correlated with the survival chances of the wasps' offspring, which do not survive if the plants die before the wasps emerge. Wasps were also attracted to undamaged mature leaves of a rice plant when one of the other mature leaves had been infested by 10 N. lugens for 1 d, implying that the volatile cues involved in host location by the parasitoid are systemically released. Collection and analyses of volatiles revealed that 1 d of N. lugens infestation did not result in the emission of new compounds or an increase in the total amount of volatiles, but rather the proportions among the compounds in the blend were altered. The total amounts and proportions of the chemicals were also affected by infestation duration. These changes in volatile profiles might provide the wasps with specific information on host habitat quality and thus could explain the observed behavioral responses of the parasitoid.     

Role of (3<Emphasis Type="Italic">Z</Emphasis>,6<Emphasis Type="Italic">Z</Emphasis>,8<Emphasis Type="Italic">E</Emphasis>)-Dodecatrien-1-ol in Trail Following,Feeding, and Mating Behavior of <Emphasis Type="Italic">Reticulitermes hesperus</Emphasis>

Trail pheromones mediate communication among western subterranean termites, Reticulitermes hesperus Banks. Repetitive passages of ≥28 termites were required to establish a pheromone trail and trails needed to be reinforced because they lasted <48 hr. The minimal threshold concentration for inducing responses from termite workers and secondary reproductives was between 0.01 and 0.1 fg/cm of (3Z,6Z,8E)-dodecatrien-1-ol (henceforth, dodecatrienol). Workers showed optimal trail-following behavior to dodecatrienol at a concentration of 10 fg/cm. Trails with concentrations >10 pg/cm were repellent to workers. Workers did not detect pheromone gradients, responding equally to increasing or decreasing gradients of dodecatrienol, and termite workers were not able to differentiate between different concentrations of dodecatrienol. Termites preferred dodecatrienol trails to 2-phenoxyethanol trails. Antennae played a key role in trail pheromone perception. Dodecatrienol acted as an arrestant for worker termites (10 fg/cm²) and male alates (5 ng/cm²), whereas sternal gland extracts from females attracted male alates. Workers and alates, upon contact with filter paper disks treated with higher doses (10 fg/cm² and 5 ng/cm², respectively) of dodecatrienol, were highly excited (increased antennation and palpation) and repeatedly returned to the treated disks. Dodecatrienol did not act as a phagostimulant when offered on a paper towel disk. Reticulitermes hesperus is highly responsive to dodecatrienol, and it may play an important role in orientation of workers and alates.     

Roquefortine/Oxaline Biosynthesis Pathway Metabolites in <Emphasis Type="Italic">Penicillium</Emphasis> ser. <Emphasis Type="Italic">Corymbifera</Emphasis>: <Emphasis Type="Italic">In Planta</Emphasis> Production and Implications for Competitive Fitness

Three strains of each of the seven taxa comprising the Penicillium series Corymbifera were surveyed by direct injection mass spectrometry (MS) and liquid chromatography–MS for the production of terrestric acid and roquefortine/oxaline biosynthesis pathway metabolites when cultured upon macerated tissue agars prepared from Allium cepa, Zingiber officinale, and Tulipa gesneriana, and on the defined medium Czapek yeast autolysate agar (CYA). A novel solid-phase extraction methodology was applied for the rapid purification of roquefortine metabolites from a complex matrix. Penicillium hordei and P. venetum produced roquefortine D and C, whereas P. hirsutum produced roquefortine D and C and glandicolines A and B. P. albocoremium, P. allii, and P. radicicola carried the pathway through to meleagrin, producing roquefortine D and C, glandicolines A and B, and meleagrin. P. tulipae produced all previously mentioned metabolites yet carried the pathway through to an end product recognized as epi-neoxaline, prompting the proposal of a roquefortine/epi-neoxaline biogenesis pathway. Terrestric acid production was stimulated by all Corymbifera strains on plant-derived media compared to CYA controls. In planta, production of terrestric acid, roquefortine C, glandicolines A and B, meleagrin, epi-neoxaline, and several other species-related secondary metabolites were confirmed from A. cepa bulbs infected with Corymbifera strains. The deposition of roquefortine/oxaline pathway metabolites as an extracellular nitrogen reserve for uptake and metabolism into growing mycelia and the synergistic role of terrestric acid and other Corymbifera secondary metabolites in enhancing the competitive fitness of Corymbifera species in planta are proposed.