Ana Mutis

Volatiles Released From Vaccinium corymbosum Were Attractive to Aegorhinus superciliosus (Coleoptera: Curculionidae) in an Olfactometric Bioassay

ABSTRACT The objective of this study was to evaluate the role of host volatiles in the relationship between a blueberry plant Vaccinium corymbosum L. and the raspberry weevil Aegorhinus superciliosus (Guérin) (Coleoptera: Curculionidae), the principal pest of blueberry in the south of Chile. Volatiles from the aerial part of different phenological stages of the host were collected on Porapak Q and analyzed by coupled gas chromatography-mass spectrometry (GC-MS). Several chemical groups were identified including green leaf volatiles, aromatic compounds, and terpenes. The olfactometric responses of A. superciliosus toward different odor sources were studied in a four-arm olfactometer. Blueberry shoots at the phenological stages of fruit set, and blue-pink fruit color elicited the greatest behavioral responses from weevils. Five compounds (2-nonanone, eucalyptol, R- and S-limonene, and 4-ethyl benzaldehyde) elicited an attractant behavioral response from A. superciliosus. The results suggest the host location behavior of A. superciliosus could be mediated by volatiles derived from V. corymbosum. This work has identified a number of compounds with which it is possible to develop a lure for the principal pest of blueberry in southern Chile.

Evidence of Contact Pheromone Use in Mating Behavior of the Raspberry Weevil (Coleoptera: Curculionidae)

ABSTRACT Numerous studies of insect species have shown that a subset of female cuticular hydrocarbons is used as short-range or contact pheromones. Here, we studied the possible use of contact pheromones in the mating behavior of the weevil Aegorhinus superciliosus, a native species of Chile. Males mounted females only after antennal contact with the females cuticle, and only 33% of the males attempted to mate with dead females washed with solvent. When a glass rod (dummy) was coated with female cuticular extracts, males exhibited behaviors similar to those observed with females. A preliminary gas chromatography-mass spectrometry (GC-MS) analysis of cuticular extracts indicated that males and females share a series of aliphatic hydrocarbons but that the relative abundance of some of these compounds differ between the sexes. These results suggest that cuticular lipids mediate mating behavior of the raspberry weevil and provide the first evidence of contact pheromones in curculionids.

Ligand binding and homology modelling of insect odorant‐binding proteins

This review describes the main characteristics of odorant‐binding proteins (OBPs) for homology modelling and presents a summary of structure prediction studies on insect OBPs, along with the steps involved and some limitations and improvements. The technique involves a computing approach to model protein structures and is based on a comparison between a target (unknown structure) and one or more templates (experimentally determined structures). As targets for structure prediction, OBPs are considered to play a functional role for recognition, desorption, scavenging, protection and transportation of hydrophobic molecules (odourants) across an aqueous environment (lymph) to olfactory receptor neurones (ORNs) located in sensilla, the main olfactory units of insect antennae. Lepidopteran pheromone‐binding proteins, a subgroup of OBPs, are characterized by remarkable structural features, in which high sequence identities (approximately 30%) among these OBPs and a large number of available templates can facilitate the prediction of precise homology models. Approximately 30 studies have been performed on insect OBPs using homology modelling as a tool to predict their structures. Although some of the studies have assessed ligand‐binding affinity using structural information and biochemical measurements, few have performed docking and molecular dynamic (MD) simulations as a virtual method to predict best ligands. Docking and MD simulations are discussed in the context of discovery of novel semiochemicals (super‐ligands) using homology modelling to conceive further strategies in insect management.

Influence of genotype and harvest year on polyphenol content and antioxidant activity in murtilla (Ugni molinae Turcz) fruit

Polyphenol content and antioxidant activity in murtilla (Ugni molinae Turcz) fruit from three genotypes (the 14-4 genotype and the Red Pearl-INIA and South Pearl-INIA varieties) were studied over five growing seasons. Our results showed significant differences in total polyphenol content among yearly harvests. The lowest value (283 ± 72 mg GAE/100 g dw) was obtained for the 14-4 genotype in the 2008 harvest, and the highest value (2,152 ± 290 mg GAE/100 g dw) was observed for the variety South Pearl-INIA in 2007. The lowest value for antioxidant activity (2,234 ± 337 μmol TE/100 g dw) was obtained for the Red Pearl-INIA variety in 2008, and the highest value (4,073 ± 76 μmol TE/100 g dw) was observed for the 14-4 genotype in 2007. There was a significant effect of genotype and growing season on polyphenol content, antioxidant activity and dry matter content for the murtilla fruits evaluated in this study, but additional studies examining other abiotic and biotic factors are required to fully explain causality.

Behavioral and electrophysiological responses of Hylastinus obscurus to volatiles released from the roots of Trifolium pratense L.

Root volatiles from field-collected red clover plants, Trifolium pratense L., of five different ages were trapped by solid-phase microextraction (SPME) and analyzed by gas chromatography-mass spectrometry (GC-MS). Ethanol, E-2-hexenal, hexanal, 3-octanone, limonene and α-pinene were identified. The electroantennographic (EAG) and olfactometric responses of clover root borer, Hylastinus obscurus, to the identified compounds were studied. The GC-MS with SPME as a solvent-free col- lection method identified ethanol and hexanal compounds that have not previously been reported in T. pratense. The EAG experiments showed that all of the tested compounds were perceived by H. obscurus, but mixed responses were found in be- havioral assays. For females, ethanol and E-2-hexenal were attractive at one or more of the tested doses, while hexanal, 3-octanone, R-limonene and S-limonene were repellent at one or more of the tested doses. Females responded to a larger number of tested compounds and a wider range doses compared to males. This behavior could reflect a more active role of females in host finding and colonization.

Behavioral Responses of Clover Root Borer to Long-Chain Fatty Acids from Young Red Clover (Trifolium pratense) Roots

ABSTRACT The olfactory and contact behavioral responses of clover root borer, Hylastinus obscurus (Marsham) (Coleoptera: Curculionidae), against fatty acid components present in 9-mo-old red clover, Trifolium pratense L., roots were investigated by using an automated behavioral observation system. From GC-MS analysis of dichloromethane extracts of T. pratense roots, of 15 compounds identified in total, four long-chain free fatty acids lauric, palmitic, oleic, and stearic acids were found to be main components in the extracts. In a four-arm olfactometer the clover root extract significantly attracted both male and female H. obscurus. When solutions of the four individual synthetic fatty acids and their blend at the ratio found in the root extract (10 µg/ml) were assayed with H. obscurus, lauric, palmitic, and oleic acid elicited an attractant behavioral response from females, whereas all substances tested did not elicit a response from males. In contact bioassays, wood dummies coated with root extract, the four fatty acids, or their respective blend of the same composition found in the root extract at 100 µg/ ml, were significantly preferred by female H. obscurus. In contrast, males showed preference only for dummies treated with root extract, or palmitic or oleic acid. The behavioral evidences suggest that long-chain free fatty acids present in 9-mo-old red clover roots may play a role as close-range olfactory, tactile cues, or both in the host-finding process of H. obscurus.

Growth promotion of Lactuca sativa in response to volatile organic compounds emitted from diverse bacterial species

Agrochemicals are currently used in horticulture to increase crop production. Nevertheless, their indiscriminate use is a relevant issue for environmental and legal aspects. Alternative tools for reducing fertilizers and synthetic phytohormones are being investigated, such as the use of volatile organic compounds (VOCs) as growth inducers. Some soil bacteria, such as Pseudomonas and Bacillus, stimulate Arabidopsis and tobacco growth by releasing VOCs, but their effects on vegetables have not been investigated. Lactuca sativa was used as model vegetable to investigate bacterial VOCs as growth inducers. We selected 10 bacteria strains, belonging to Bacillus, Staphylococcus and Serratia genera that are able to produce 3-hydroxy-2-butanone (acetoin), a compound with proven growth promoting activity. Two-day old-seedlings of L. sativa were exposed to VOCs emitted by the selected bacteria grown in different media cultures for 7 days. The results showed that the VOCs released from the bacteria elicited an increase in the number of lateral roots, dry weight, root growth and shoot length, depending on the media used. Three Bacillus strains, BCT53, BCT9 and BCT4, were selected according to its their growth inducing capacity. The BCT9 strain elicited the greatest increases in dry weight and primary root length when L. sativa seedlings were subjected to a 10-day experiment. Finally, because acetoin only stimulated root growth, we suggest that other volatiles could be responsible for the growth promotion of L. sativa. In conclusion, our results strongly suggest that bacteria volatiles can be used as growth-inducers as alternative or complementary strategies for application in horticulture species.

Type and Distribution of Sensilla in the Antennae of the Red Clover Root Borer, Hylastinus obscurus

Abstract In order to determine the type, distribution, and structures of sensilla, the antennae of the red clover root borer, Hylastinus obscurus Marsham (Coleoptera: Curculionidae: Scolytinae), were examined by light and electron microscopy (both scanning and transmission). Four different types of sensilla were identified in the club, and one type of chaetica was found in the scape and funicle of both male and female individuals. Chaetica and basiconica were the most abundant sensilla types in the club. They were present in the three sensory bands described, totaling approximately 80% of sensilla in the antennal club of H. obscurus. Chaetica were predominantly mechanoreceptors, although gustatory function could not be excluded. Basiconica forms showed characteristics typical of olfactory sensilla. Trichoidea were not found in the proximal sensory band, and they exhibited abundant pores, suggesting olfactory function. Styloconica were the least abundant sensillum type, and their shape was similar to that reported as having hygro- and thermoreceptor functions. There was no difference in the relative abundance of antennal sensilla between males and females. Finally, the sensillar configuration and abundance of receptors in the H. obscurus antennae suggest that these sensilla have chemoreceptive and other functions.

Electroantennographic and Behavioral Responses of Adults of Raspberry Weevil Aegorhinus superciliosus (Coleoptera: Curculionidae) to Odors Released From Conspecific Females

ABSTRACT The raspberry weevil, Aegorhinus superciliosus (Guérin) (Coleoptera: Curculionidae), is the most important pest in blueberry and raspberry fields in the south of Chile. In this study, we investigated the electroantennographic and behavioral responses of A. superciliosus to semiochemicals released from conspecific individual adults, with particular attention to male attraction to females. Odors released from females significantly attracted males in a Y-tube olfactometer. Gas chromatographic and mass spectral analysis of female volatile extracts revealed the presence of limonene and &agr;-pinene. Electroantennogram recordings from both sexes indicated that males of A. superciliosus possess olfactory sensitivity for the R isomer of limonene and &agr;-pinene, whereas females only perceived R-limonene. Behavioral assays using synthetic compounds showed that only R-limonene elicited an attraction response from male weevils. Field experiments confirmed the laboratory results, showing that R-limonene was attractive to weevils. This is the first report of intraspecific chemical communication in this weevil. We discuss the origin of these compounds, their possible role in the sexual behavior of this species, and their potential use in a pest control strategy.

β-Ionone as putative semiochemical suggested by ligand binding on an odorant-binding protein of Hylamorpha elegans and electroantennographic recordings

Currently, odorant‐binding proteins (OBPs) are considered the first filter for olfactory information for insects and constitute an interesting target for pest control. Thus, an OBP (HeleOBP) from the scarab beetle Hylamorpha elegans (Burmeister) was identified, and ligand‐binding assays based on fluorescence and in silico approaches were performed, followed by a simulated binding assay. Fluorescence binding assays showed slight binding for most of the ligands tested, including host‐plant volatiles. A high binding affinity was obtained for β‐ionone, a scarab beetle‐related compound. However, the binding of its analogue α‐ionone was weaker, although it is still considered good. On the other hand, through a three‐dimensional model of HeleOBP constructed by homology, molecular docking was carried out with 29 related ligands to the beetle. Results expressed as free binding energy and fit quality (FQ) indicated strong interactions of sesquiterpenes and terpenoids (α‐ and β‐ionone) with HeleOBP as well as some aromatic compounds. Residues such as His102, Tyr105 and Tyr113 seemed to participate in the interactions previously mentioned. Both in silico scores supported the experimental affinity for the strongest ligands. Therefore, the activity of α‐ionone, β‐ionone and 2‐phenyl acetaldehyde at antennal level was studied using electroantenography (EAG). Results showed that the three ligands are electrophysiologically active. However, an aliquot of β‐ionone (represented by 3.0 ng) elicited stronger EAG responses in antennae of males than of females. Finally, the role of these ligands as potential semiochemicals for H. elegans is discussed.