Gianfranco Anfora

Putative Chemosensory Receptors of the Codling Moth, Cydia pomonella, Identified by Antennal Transcriptome Analysis

The codling moth, Cydia pomonella, is an important fruit pest worldwide. As nocturnal animals, adults depend to a large extent on olfactory cues for detection of food and mates, and, for females, oviposition sites. In insects, odor detection is mediated by odorant receptors (ORs) and ionotropic receptors (IRs), which ensure the specificity of the olfactory sensory neuron responses. In this study, our aim was to identify chemosensory receptors in the codling moth as a means to uncover new targets for behavioral interference. Using next-generation sequencing techniques, we identified a total of 43 candidate ORs, one gustatory receptor and 15 IRs in the antennal transcriptome. Through Blast and sequence similarity analyses we annotated the insect obligatory co-receptor ORco, five genes clustering in a conserved clade containing sex pheromone receptors, one homolog of the Bombyx mori female-enriched receptor BmorOR30 (but no homologs of the other B. mori female-enriched receptors) and one gene clustering in the sugar receptor family. Among the candidate IRs, we identified homologs of the two highly conserved co-receptors IR8a and IR25a, and one homolog of an IR involved in phenylethyl amine detection in Drosophila. Our results open for functional characterization of the chemosensory receptors of C. pomonella, with potential for new or refined applications of semiochemicals for control of this pest insect.

ANTENNAL AND BEHAVIORAL RESPONSES OF GRAPEVINE MOTH Lobesia botrana FEMALES TO VOLATILES FROM GRAPEVINE

Grapevine moth Lobesia botrana is the economically most important insect of grapevine Vitis vinifera in Europe. Flower buds, flowers, and green berries of Chardonnay grapevine are known to attract L. botrana for oviposition. The volatile compounds collected from these phenological stages were studied by gas chromatography—mass spectrometry, and the antennal response of L. botrana females to these headspace collections was recorded by gas chromatography—electroantennography. The compounds found in all phenological stages, which consistently elicited a strong antennal response, were pentadecane, nonanal, and α -farnesene. In a wind tunnel, gravid L. botrana females flew upwind to green grapes, as well as to headspace collections from these berries released by a piezoelectric sprayer release device. However, no females landed at the source of headspace volatiles, possibly due to inappropriate concentrations or biased ratios of compounds in the headspace extracts.

A predicted sex pheromone receptor of codling moth Cydia pomonella detects the plant volatile pear ester

Plant volatiles mediate host discrimination and host finding in phytophagous insects. Understanding how insects recognize these signals is a current challenge in chemical ecology research. Pear ester, ethyl (E,Z)-2,4-decadienoate, is a powerful, bisexual attractant of codling moth Cydia pomonella (Lepidoptera, Tortricidae) and strongly synergizes the male response to female-produced sex pheromone. We show here that the codling moth odorant receptor (OR) CpomOR3 is dedicated to detecting this plant volatile. Heterologous expression of CpomOR3 in Drosophila T1 trichoid and ab3A basiconic sensilla, followed by a screening with codling moth pheromone compounds and known plant volatile attractants, confirms that CpomOR3 binds to pear ester. Although CpomOR3 does not respond to any of the pheromone components tested, a phylogenetic analysis of lepidopteran chemosensory receptor genes reveals a close relationship of CpomOR3 with pheromone receptors (PRs) in moths. This corroborates the interaction of ecological and social chemosensory cues during premating communication. The finding that a plant volatile compound, pear ester, is a specific ligand for a PR-like lepidopteran receptor adds to our understanding of insect-plant interactions and emphasizes the interaction of natural and sexual selection during the phylogenetic divergence of insect herbivores.

Chemical Ecology and Management of Lobesia botrana (Lepidoptera: Tortricidae)

ABSTRACT The moth Lobesia botrana (Denis & Schiffermüller) (Lepidoptera: Tortricidae) feeds on grapes (Vitis vinifera L.), reducing yield and increasing susceptibility to fungal infections. L. botrana is among the most economically important insects in Europe and has recently been found in vineyards in Chile, Argentina, and California. Here, we review L. botrana biology and behavior in relation to its larval host (the grapevine) and its natural enemies. We also discuss current and future control strategies in light of our knowledge of chemical ecology, with an emphasis on the use of the sex pheromone-based strategies as an environmentally safe management approach. Pheromone-mediated mating disruption is the most promising technique available on grapes and is currently implemented on ≈140,000 ha in Europe. Experience from several growing areas confirms the importance of collaboration between research, extension, growers, and pheromone-supply companies for the successful implementation of the mating disruption technique. In the vineyards where mating disruption has been successfully applied as an areawide strategy, the reduction in insecticide use has improved the quality of life for growers, consumers, as well as the public living near wine-growing areas and has thereby reduced the conflict between agricultural and urban communities.

Behavioural and electrophysiological lateralization in a social (Apis mellifera) but not in a non-social (Osmia cornuta) species of bee.

Recent evidence suggests that asymmetry between the left and right sides of the brain is not limited to vertebrates but extends to invertebrates as well. We compared olfactory lateralization in two species of Hymenoptera Apoidea, the honeybee (Apis mellifera), a social species, and the mason bee (Osmia cornuta), a solitary species. Recall of the olfactory memory 1 h after training to associate an odour with a sugar reward, as revealed by the bee extending its proboscis when presented with the trained odour, was better in honeybees trained with their right than with their left antenna. No such asymmetry was observed in mason bees. Similarly, electroantennographic responses to a floral volatile compound and to an alarm pheromone component were higher in the right than in the left antenna in honeybees but not in mason bees. These findings seem to support recent game-theoretical models suggesting that population-level lateralization is more likely to have evolved in social than in non-social species.

Coding and interaction of sex pheromone and plant volatile signals in the antennal lobe of the codling moth Cydia pomonella.

SUMMARY In the codling moth Cydia pomonella (Lepidoptera: Tortricidae) plant volatiles attract males and females by upwind flight and synergise the male response to the female-produced sex pheromone, indicating a close relationship between the perception of social and environmental olfactory signals. We have studied the anatomical and functional organisation of the antennal lobe (AL), the primary olfactory centre, of C. pomonella with respect to the integration of sex pheromone and host-plant volatile information. A three-dimensional reconstruction of the glomerular structure of the AL revealed 50±2 and 49±2 glomeruli in males and females, respectively. These glomeruli are functional units involved in the coding of odour quality. The glomerular map of the AL was then integrated with electrophysiological recordings of the response of individual neurons in the AL of males and females to sex pheromone components and behaviourally active plant volatiles. By means of intracellular recordings and stainings, we physiologically characterised ca. 50 neurons in each sex, revealing complex patterns of activation and a wide variation in response dynamics to these test compounds. Stimulation with single chemicals and their two-component blends produced both synergistic and inhibitory interactions in projection neurons innervating ordinary glomeruli and the macroglomerular complex. Our results show that the sex pheromone and plant odours are processed in an across-fibre coding pattern. The lack of a clear segregation between the pheromone and general odour subsystems in the AL of the codling moth suggests a level of interaction that has not been reported from other insects.

Integrating temperature-dependent life table data into a matrix projection model for Drosophila suzukii population estimation

Temperature-dependent fecundity and survival data was integrated into a matrix population model to describe relative Drosophila suzukii Matsumura (Diptera: Drosophilidae) population increase and age structure based on environmental conditions. This novel modification of the classic Leslie matrix population model is presented as a way to examine how insect populations interact with the environment, and has application as a predictor of population density. For D. suzukii, we examined model implications for pest pressure on crops. As case studies, we examined model predictions in three small fruit production regions in the United States (US) and one in Italy. These production regions have distinctly different climates. In general, patterns of adult D. suzukii trap activity broadly mimicked seasonal population levels predicted by the model using only temperature data. Age structure of estimated populations suggest that trap and fruit infestation data are of limited value and are insufficient for model validation. Thus, we suggest alternative experiments for validation. The model is advantageous in that it provides stage-specific population estimation, which can potentially guide management strategies and provide unique opportunities to simulate stage-specific management effects such as insecticide applications or the effect of biological control on a specific life-stage. The two factors that drive initiation of the model are suitable temperatures (biofix) and availability of a suitable host medium (fruit). Although there are many factors affecting population dynamics of D. suzukii in the field, temperature-dependent survival and reproduction are believed to be the main drivers for D. suzukii populations.

Linking Genomics and Ecology to Investigate the Complex Evolution of an Invasive Drosophila Pest

Drosophilid fruit flies have provided science with striking cases of behavioral adaptation and genetic innovation. A recent example is the invasive pest Drosophila suzukii, which, unlike most other Drosophila, lays eggs and feeds on undamaged, ripening fruits. This not only poses a serious threat for fruit cultivation but also offers an interesting model to study evolution of behavioral innovation. We developed genome and transcriptome resources for D. suzukii. Coupling analyses of these data with field observations, we propose a hypothesis of the origin of its peculiar ecology. Using nuclear and mitochondrial phylogenetic analyses, we confirm its Asian origin and reveal a surprising sister relationship between the eugracilis and the melanogaster subgroups. Although the D. suzukii genome is comparable in size and repeat content to other Drosophila species, it has the lowest nucleotide substitution rate among the species analyzed in this study. This finding is compatible with the overwintering diapause of D. suzukii, which results in a reduced number of generations per year compared with its sister species. Genome-scale relaxed clock analyses support a late Miocene origin of D. suzukii, concomitant with paleogeological and climatic conditions that suggest an adaptation to temperate montane forests, a hypothesis confirmed by field trapping. We propose a causal link between the ecological adaptations of D. suzukii in its native habitat and its invasive success in Europe and North America.

Synthetic Grape Volatiles Attract Mated Lobesia botrana Females in Laboratory and Field Bioassays

In laboratory experiments, we identified and quantified volatiles emitted by inflorescences and berries of two grape varieties (Trebbiano and Sangiovese) and examined the effects of the volatiles on oviposition by the grapevine moth Lobesia botrana. Compared to Trebbiano, Sangiovese is relatively more susceptible to L. botrana infestations under natural conditions. Chemical and electrophysiological analysis indicated only quantitative differences between the volatiles released by the two varieties. In a dual-choice oviposition bioassay based only on volatile cues, females did not show any preference between the two varieties. The six major components of the odor profiles that were GC-EAD-active to female antennae included: limonene, 4,8-dimethyl-1,(E)-3,7-nonatriene, (±)-linalool, (E)-caryophyllene, (E,E)-α-farnesene, and methyl salicylate. At the beginning of the berry touch phenological stage, their proportions were about 10:0.6:0.4:0.5:0.9:0.6 in Trebbiano and 10:1:0.4:1.5:0.4:0.3 in Sangiovese. A six-component synthetic lure (with the proportion 10:1:1:1:1:1, which approximated the ratio of components released by both varieties) was used in further laboratory oviposition bioassays. Depending on its dosage, the synthetic lure either attracted or repelled oviposition. L. botrana females laid significantly more eggs in the presence of either the grape bunches or the synthetic lure at the attractive dosage. In a release-capture experiment conducted in a field cage that covered two grapevine rows, the synthetic lure was more attractive than a grape cluster or a blank control, and it stimulated oviposition on the vegetation near the lure. The results indicate that L. botrana uses olfactory cues to select oviposition sites and that an artificial lure, containing the major volatiles released by two grape varieties, may be useful in monitoring female activity in the field.

Exploitation of insect vibrational signals reveals a new method of pest management.

Food production is considered to be the main source of human impact on the environment and the concerns about detrimental effects of pesticides on biodiversity and human health are likely to lead to an increasingly restricted use of chemicals in agriculture. Since the first successful field trial, pheromone based mating disruption enabled sustainable insect control, which resulted in reduced levels of pesticide use. Organic farming is one of the fastest growing segments of agriculture and with the continuously growing public concern about use of pesticides, the main remaining challenge in increasing the safety of the global food production is to identify appropriate alternative mating disruption approaches for the numerous insect pests that do not rely on chemical communication. In the present study, we show for the first time that effective mating disruption based on substrate-borne vibrational signals can be achieved in the field. When disruptive vibrational signals were applied to grapevine plants through a supporting wire, mating frequency of the leafhopper pest Scaphoideus titanus dropped to 9 % in semi-field conditions and to 4 % in a mature vineyard. The underlying mechanism of this environmentally friendly pest-control tactic is a masking of the vibrational signals used in mate recognition and location. Because vibrational communication is widespread in insects, mating disruption using substrate vibrations can transform many open field and greenhouse based farming systems.