Koji Noge

Parasitic wasp responses to symbiont-based defense in aphids

BackgroundRecent findings indicate that several insect lineages receive protection against particular natural enemies through infection with heritable symbionts, but little is yet known about whether enemies are able to discriminate and respond to symbiont-based defense. The pea aphid, Acyrthosiphon pisum, receives protection against the parasitic wasp, Aphidius ervi, when infected with the bacterial symbiont Hamiltonella defensa and its associated bacteriophage APSE (Acyrthosiphon pisumsecondary endosymbiont). Internally developing parasitoid wasps, such as A. ervi, use maternal and embryonic factors to create an environment suitable for developing wasps. If more than one parasitoid egg is deposited into a single aphid host (superparasitism), then additional complements of these factors may contribute to the successful development of the single parasitoid that emerges.ResultsWe performed experiments to determine if superparasitism is a tactic allowing wasps to overcome symbiont-mediated defense. We found that the deposition of two eggs into symbiont-protected aphids significantly increased rates of successful parasitism relative to singly parasitized aphids. We then conducted behavioral assays to determine whether A. ervi selectively superparasitizes H. defensa-infected aphids. In choice tests, we found that A. ervi tends to deposit a single egg in uninfected aphids, but two or more eggs in H. defensa-infected aphids, indicating that oviposition choices may be largely determined by infection status. Finally, we identified differences in the quantity of the trans-β-farnesene, the major component of aphid alarm pheromone, between H. defensa-infected and uninfected aphids, which may form the basis for discrimination.ConclusionsHere we show that the parasitic wasp A. ervi discriminates among symbiont-infected and uninfected aphids, and changes its oviposition behavior in a way that increases the likelihood of overcoming symbiont-based defense. More generally, our results indicate that natural enemies are not passive victims of defensive symbionts, and that an evolutionary arms race between A. pisum and the parasitoid A. ervi may be mediated by a bacterial symbiosis.

Adults and nymphs do not smell the same: the different defensive compounds of the giant mesquite bug (Thasus neocalifornicus: Coreidae).

Heteropteran insects often protect themselves from predators with noxious or toxic compounds, especially when these insects occur in aggregations. The predators of heteropteran insects change from small insect predators to large avian predators over time. Thus, a chemical that is deterrent to one type of predator at one point in time may not be deterrent to another type of predator at another point in time. Additionally, these predator deterrent compounds may be used for other functions such as alarm signaling to other conspecifics. Defensive secretion compounds from the adult and the nymph giant mesquite bug (Thasus neocalifornicus: Coreidae) were isolated and identified by gas chromatography–mass spectrometry and NMR. The predominant compounds isolated from the nymph mesquite bugs during a simulated predator encounter were (E)-2-hexenal and 4-oxo-(E)-2-hexenal. In adults, the major compounds released during a simulated predator encounter were hexyl acetate, hexanal, and hexanol. Results from predator bioassays suggest the nymph compounds are more effective at deterring an insect predator than the adult compounds. By using behavioral bioassays, we determined the role of each individual compound in signaling to other mesquite bugs. The presence of the nymph secretion near a usually compact nymph aggregation caused nymph mesquite bugs to disperse but did not affect adults. Conversely, the presence of the adult secretion caused the usually loose adult aggregation to disperse, but it did not affect nymph aggregation. The compounds that elicited nymph behavioral responses were (E)-2-hexenal and 4-oxo-(E)-2-hexenal, while those that elicited adult behavioral responses were hexyl acetate and hexanal. The differences between the chemical composition of nymph and adult defensive secretions and alarm behavior are possibly due to differences in predator guilds.

Germacrene D, a common sesquiterpene in the genus Bursera (Burseraceae).

The volatile components of the leaves of five Bursera species, B. copallifera, B. exselsa, B. mirandae, B. ruticola and B. fagaroides var. purpusii were determined by gas chromatography–mass spectrometry (GC–MS). Germacrene D was one of the predominant components (15.1–56.2%) of all of these species. Germacrene D has also been found in other Bursera species and some species of Commiphora, the sister group of Bursera, suggesting that the production of germacrene D might be an ancient trait in the genus Bursera.

Defensive roles of (E)-2-alkenals and related compounds in heteroptera.

We examined whether shared volatiles found in various heteropteran species and developmental stages function to repel predators. The nymphal dorsal abdominal gland secretions of Riptortus pedestris (Heteroptera: Alydidae) and Thasus acutangulus (Heteroptera: Coreidae), and the metathoracic scent gland secretion of Euschistus biformis (Heteroptera: Pentatomidae) adults were identified by gas chromatography/mass spectrometry (GC/MS). (E)-2-Hexenal, 4-oxo-(E)-2-hexenal (4-OHE), and (E)-2-octenal were found in all three species and deemed likely candidates for repelling predators. In addition to (E)-2-alkenals, the adult E. biformis secreted (E)-2-hexenyl acetate, (E)-2-octenyl acetate, and four hydrocarbons. We evaluated the potential predator repellent properties of these compounds and compound blends against a generalist, cosmopolitan insect predator, the Chinese praying mantid (Mantodea: Mantidae: Tenodera aridifoliasinensis). Mantids that experienced (E)-2-hexenal, (E)-2-octenal, and (E)-2-octenyl acetate moved away from the site of interaction, while 4-OHE and (E)-2-hexenyl acetate did not affect mantid behavior. The compound blends did not have additive or synergistic repellency effects on predator behavior. Compound repellency was not related to compound volatility. Instead, the repellent effect is likely related to predator olfaction, and the affinity of each compound to receptors on the antennae. Our results also suggest the repellents might intensify the visual defensive signals of aposematism (T. acutangulus nymphs) and mimicry (R. pedestris nymphs) in heteropteran bugs.

Gut Microbiota in Nymph and Adults of the Giant Mesquite Bug (Thasus neocalifornicus) (Heteroptera: Coreidae) Is Dominated by Burkholderia Acquired De Novo Every Generation

ABSTRACT The coreid bug Thasus neocalifornicus Brailovsky and Barrera, commonly known as the giant mesquite bug, is a ubiquitous insect of the southwestern United States. Both nymphs and adults are often found aggregated on mesquite trees (Prosopis spp.: Fabaceae) feeding on seedpods and plant sap. We characterized the indigenous bacterial populations of nymphs and adults of this species by using molecular and phylogenetic techniques and culturing methods. Results show that this insects bacterial gut community has a limited diversity dominated by Burkholderia associates. Phylogenetic analysis by using 16s rRNA sequences suggests that these &bgr;-Proteobacteria are closely related to those symbionts obtained from other heteropteran midgut microbial communities but not to Burkholderia symbionts associated with other insect orders. These bacteria were absent from the eggs and were not found in all younger nymphs, suggesting that they are acquired after the insects have hatched. Rearing experiments of nymphs with potentially Burkholderia contaminated soil suggested that if this symbiont is not acquired, giant mesquite bugs experience higher mortality. Egg, whole-body DNA extractions of younger nymphs, and midgut DNA extractions of fifth-instar nymphs and adults also revealed the presence of &agr;-Proteobacteria from the Wolbachia genus. However, this bacterium was also present in reproductive organs of adults, indicating that this symbiont is not specific to the gut.

Herbivore-induced phenylacetonitrile is biosynthesized from de novo-synthesized l-phenylalanine in the giant knotweed, Fallopia sachalinensis

Plants emit a series of characteristic volatile blends when damaged by insect feeding. Phenylacetonitrile is one of the volatiles from the leaves of the giant knotweed, Fallopia sachalinensis, infested by the Japanese beetle, Popillia japonica, or treated with exogenous airborne methyl jasmonate (MeJA). We examined the precursor of the nitrile and its origin in this system. l‐Phenylalanine was determined to be a precursor of the nitrile in F. sachalinensis leaves, and the phenylalanine was also induced by beetle feeding and MeJA treatment. We also found that exogenous MeJA enhanced the biosynthesis of several amino acids in F. sachalinensis leaves.

Methyl jasmonate is transported to distal leaves via vascular process metabolizing itself into JA-Ile and triggering VOCs emission as defensive metabolites.

Plants have developed multifaceted defensive systems against adverse environmental factors. One such recognized system is the production of metabolites in plants. Jasmonic acid (JA) and its metabolite methyl jasmonate (MeJA) are known to play key roles in metabolites production. The role of MeJA as a mobile signal has been established in Arabidopsis and Solanaceae plants. However, it remains largely unclear how MeJA-based signaling is organized via its elicited metabolites. Here, we investigated the signaling ability of MeJA by means of vascular transport using Achyranthes bidentata as an experimental plant. Results showed that MeJA was transported and essentially metabolized into its active form JA-Ile in the distal undamaged leaves accompanied by emission of volatile organic compounds. Results presented and discussed therein provide convincing evidence that MeJA acts as a transportable inter-cellular mobile compound in plants self-defense scheme.

Cytochrome P450 CYP71AT96 catalyses the final step of herbivore-induced phenylacetonitrile biosynthesis in the giant knotweed, Fallopia sachalinensis

The giant knotweed Fallopia sachalinensis (Polygonaceae) synthesizes phenylacetonitrile (PAN) from l-phenylalanine when infested by the Japanese beetle Popillia japonica or treated with methyl jasmonate (MeJA). Here we identified (E/Z)-phenylacetaldoxime (PAOx) as the biosynthetic precursor of PAN and identified a cytochrome P450 that catalysed the conversion of (E/Z)-PAOx to PAN. Incorporation of deuterium-labelled (E/Z)-PAOx into PAN emitted from the leaves of F. sachalinensis was detected using gas chromatography–mass spectrometry. Further, using liquid chromatography–tandem mass spectrometry, we detected the accumulation of (E/Z)-PAOx in MeJA-treated leaves. These results showed that (E/Z)-PAOx is the biosynthetic precursor of PAN. MeJA-induced mRNAs were analysed by differential expression analysis using a next-generation sequencer. Of the 74,329 contigs obtained from RNA-seq and de novo assembly, 252 contigs were induced by MeJA treatment. Full-length cDNAs encoding MeJA-induced cytochrome P450s CYP71AT96, CYP82AN1, CYP82D125 and CYP715A35 were cloned using 5′- and 3′-RACE and were expressed using a baculovirus expression system. Among these cytochrome P450s, CYP71AT96 catalysed the conversion of (E/Z)-PAOx to PAN in the presence of NADPH and a cytochrome P450 reductase. It also acted on (E/Z)-4-hydroxyphenylacetaldoxime and (E/Z)-indole-3-acetaldoxime. The broad substrate specificity of CYP71AT96 was similar to that of aldoxime metabolizing cytochrome P450s. Quantitative RT-PCR analysis showed that CYP71AT96 expression was highly induced because of treatment with MeJA as well as feeding by the Japanese beetle. These results indicate that CYP71AT96 likely contributes the herbivore-induced PAN biosynthesis in F. sachalinensis.

Phenylacetonitrile from the Giant Knotweed, Fallopia sachalinensis, Infested by the Japanese Beetle, Popillia japonica, Is Induced by Exogenous Methyl Jasmonate

Phenylacetonitrile, (E)-β-ocimene, linalool, (E)-4,8-dimethyl-1,3,7-nonatriene and (E,E)-α-farnesene were identified as Japanese beetle, Popillia japonica, feeding-induced volatiles from the leaves of the giant knotweed, Fallopia sachalinensis, but not by mechanical damage. Volatile emission was also induced by treatment with a cellular signaling molecule, methyl jasmonate. These results suggest that volatiles will be synthesized de novo by a biotic elicitor from P. japonica oral secretion.

Deuterium labeling for investigating de novo synthesis of terpene volatiles in Achyranthes bidentata.

Purpose of workPlants synthesize and accumulate secondary metabolites as defensive volatiles against diverse stresses. We aim to unravel the jasmonate-inducible volatile de novo synthetic metabolites in plants using a deuterium-labeling technique. Jasmonic acid and its methyl ester (MeJA) are well-documented for inducing defensive volatiles. Here, we have developed an efficient deuterium oxide (D2O)-based labeling approach to determine the extent of de novo synthetic metabolites in a model plant A. bidentata bidentata. The labeling approach was demonstrated on quantitative profiling of terpene volatile organic compounds (VOCs) elicited by airborne MeJA in Achyranthes plants. We show, for the first time that airborne MeJA-elicited terpene VOCs are predominantly and differentially de novo synthesized except for a homoterpene, (3E)-4,8-dimethyl-1,3,7-nonatriene, which is weakly and least labelled with deuterium. D2O is therefore an efficient labeling source for investigating de novo synthetic metabolites of terpene VOCs in planta.