Olfactory mechanisms of host selection in phytophagous insects

The most challenging tasks for phytophagous insects are the location and selection of mates, food sources, and oviposition sites, all crucial for survival and reproduction. To perform these tasks insects rely largely on their sense of smell (olfaction). I address how the insect olfactory system discriminates between components of complex odor mixtures, modulating behavior and fitness. I have studied modulation of attraction in the moth Spodoptera littoralis and the bark beetle Ips typographus by separation of pheromone (Ph) and anti-attractants, and of Ph components alone. An antagonist reduced male moth attraction towards the female sex Ph, and a blend of non-host volatiles (NHV) reduced attraction of both sexes of I. typographus towards their Ph, insect catches decreased with decreasing odor-source distance. Conversely, increasing distance between Ph components decreased attraction in both insect species. However, moths were more sensitive to small-scale spacing. Reproductive behaviors as well as fecundity and longevity of S. littoralis moths were negatively affected in the presence of volatiles from leaves of non-host plants, Picea abies or Adhatoda vasica. The presence of non-host plants strongly modulated male moths’ behavior, reducing their attraction towards the Ph source in flight assays. Gas chromatography-electroantenno-graphic detection (GC-EAD) by female S. littoralis antennae with headspace volatile collections from P. abies and A. vasica revealed eight active compounds, with seven new actives. Single sensillum recordings (SSR) created a functional-morphological map of 49 olfactory sensory neuron (OSN) functional types in six morphological sensillum types in female S. littoralis. Proximally located OSNs showed a higher sensitivity, shorter latency, and displayed more phasic responses than distally located OSNs of the same class. GC-SSRs with volatiles from a larval host, cotton plants, and the adult nectar source, lilac flowers, revealed 38 active compounds for female OSNs, including 12 new actives. The odor response specificities of four olfactory receptor (OR) genes of S. littoralis were deorphanized by expression in the Empty Neuron System (ENS) of Drosophila melanogaster using SSR and GC-SSR (GC-SSR-ENS). Two of the ORs responded specifically to single odorants, while the other two responded similarly to the same 9 compounds, but dose-response experiments with new compounds, identified by GC-SSR, revealed specific odor-response profiles

Plume interaction and odour source spacing of pheromone and non-host volatiles

Insects have a remarkable ability to sense whether odours are released from one point source or from two closely separated ones. Therefore it is of interest to study the interactions of pheromone component odour plumes to affect the insect behaviour when the release points of different components are physically separated. It helps us to understand the mechanism that insects use to find their host or any attractive source. We studied the behavioural response of Spodoptera littoralis (Lepidoptera, Noctuidae) males, in a walking bioassay, to two components of the female sex pheromone blend, (9Z,11E-tetradecadienyl acetate) and(9Z,12E-tetradecadienyl acetate), released from two separate dispensers at various distances (0-5 cm) from each other. The main pheromone component, 9Z-11E-14:OAc was tested at 1 and 10 ng in combination with the minor component 9Z-12E-14:OAc, at 1 % of the main component. The low dose always showed a lower behavioural response than the high dose at the same separation distances (0-5 cm). When low doses were used, the response was higher at 0 and 3 cm separation than other separations whereas when high doses were used there was not much difference in response at 0, 2, and 3 cm separations. A Photo Ionization Detector(PID) was used to investigate plume width and overlap. The PID showed that 5 cm separation of odour sources was enough to totally separate the odour plumes from each other, whereas at 3 cm separation, plumes partially overlapped. By comparing the behavioural results with PID data it is concluded that insects are much more sensitive than the PID, since they responded at distances where the PID showed zero ppb. Male insects may respond well to doses that are 20 times lower than one female equivalent. A field experiment for S. litura was done in Pakistan on cotton crop by spacing sex pheromone components. Pheromone components were spaced horizontally at 0, 3, 5 and 15 cm on plastic moth pheromone traps. The high dose attracted more males than the medium and low dose. Males were highly attracted at 0 cm separation but spacing decreased the catch and at 15 cm spacing there was no catch. The behavioural effect of separating ph. components was further studied on the spruce bark beetle, Ips typographus L., in the field. The attraction of I. typographus to traps baited with the two aggregation pheromone components (cis-verbenol and 2-metyl-3-buten-2-ol) separated, was investigated in a Norway spruce clear-cut. The pheromone components were separated both vertically (0-112 cm) on extended Lindgren (19 funnel) traps and horizontally (0-80 cm) on modified windvane traps. I. typographus was strongly attracted when the two components were released from the same point but spacing between components decreased the trap catch. However, at 16 cm distance, in both the vertical and horizontal test, the trap catch was not much different from the positive control (0 cm separation). The effect of odour source spacing in the field on I. typographus was also studied with regards to repellent non-host volatiles (NHV). In this test, the aggregation pheromone were separated from a blend of repellents (trans-conopthorin, 1,8-cineol, 3-octanol, 1-octen-3-ol, 1-hexanol and verbenone) using the same experimental design. NHV showed strong inhibitory effect up to 48 cm spacing but there was not much effect at 80 and 112 cm separations

A novel lineage of candidate pheromone receptors for sex communication in moths

Sex pheromone receptors (PRs) are key players in chemical communication between mating partners in insects. In the highly diversified insect order Lepidoptera, male PRs tuned to female-emitted type I pheromones (which make up the vast majority of pheromones identified) form a dedicated subfamily of odorant receptors (ORs). Here, using a combination of heterologous expression and in vivo genome editing methods, we bring functional evidence that at least one moth PR does not belong to this subfamily but to a distantly related OR lineage. This PR, identified in the cotton leafworm Spodoptera littoralis, is highly expressed in male antennae and is specifically tuned to the major sex pheromone component emitted by females. Together with a comprehensive phylogenetic analysis of moth ORs, our functional data suggest two independent apparitions of PRs tuned to type I pheromones in Lepidoptera, opening up a new path for studying the evolution of moth pheromone communication

Effect of Prepartum Dietary Energy Level on Production and Reproduction in Nili Ravi Buffaloes

The objective of this study was to evaluate the effect of prepartum dietary energy level on postpartum production and reproduction in Nili Ravi buffaloes (n = 21). The buffaloes were offered low energy (LE: 1.31 Mcal/kg DM NEL (net energy for lactation)), medium energy (ME: 1.42 Mcal/kg DM NEL) or high energy (HE: 1.54 Mcal/kg DM NEL) diet for 63 days prepartum, and received the same lactation diet (LD: 1.22 Mcal/kg DM NEL) during 14 weeks postpartum. The effects of dietary energy level and week were analyzed with Proc GLIMMIX model. Dry matter intake (DMI) was lower in buffaloes fed the LE diet compared with buffaloes fed the ME or HE diet. Calf birth weight (CBW) was higher in buffaloes fed the HE diet compared with buffaloes fed the ME or LE diet. Milk production was similar in buffaloes fed the HE, ME or LE diet within 14 weeks postpartum and throughout the lactation. Milk fat was higher in buffaloes fed the LE diet compared with buffaloes fed the ME or HE diet. Milk protein and lactose yields was high in buffaloes fed the HE diet compared with buffaloes fed the ME or LE diet. Body condition score was high in HE and was affected by diet × week interactions during pre and postpartum period. The concentrations of β-hydroxybutyrate (BHBA) and triglycerides in serum was lowest in buffaloes fed the HE diet compared with the buffaloes fed the ME or LE diet. The buffaloes fed the HE diet had early uterine involution (UI), first estrus, short dry days, and calving interval (CI) compared with buffaloes fed the ME or LE diet. None of buffaloes fed the LE diet exhibited estrus during the first 14 weeks postpartum compared with buffaloes fed the ME or HE diet. In conclusion, prepartum feeding of high energy diet can be helpful in improving the postpartum productive and reproductive performance in Nili Ravi buffaloes

Attraction modulated by spacing of pheromone components and anti-attractants in a bark beetle and a moth

Orientation for insects in olfactory landscapes with high semiochemical diversity may be a challenging task. The partitioning of odor plumes into filaments that are interspersed with pockets of ‘clean air’ may help filament discrimination and upwind flight to attractive sources in the face of inhibitory signals. We studied the effect of distance between odor sources on trap catches of the beetle, Ips typographus, and the moth, Spodoptera littoralis. Insects were tested both to spatially separated pheromone components [cis-verbenol and 2-methyl-3-buten-2-ol for Ips; (Z,E)- 9,11-tetradecadienyl acetate and (Z,E)-9,12-tetradecadienyl acetate for Spodoptera], and to separated pheromone and anti-attractant sources [non-host volatile (NHV) blend for Ips; (Z)-9-tetradecenyl acetate for Spodoptera]. Trap catch data were complemented with simulations of plume structure and plume overlap from two separated sources using a photo ionization detector and soap bubble generators. Trap catches of the beetle and the moth were both affected when odor sources in the respective traps were increasingly separated. However, this effect on trap catch occurred at smaller (roughly by an order of magnitude) odor source separation distances for the moth than for the beetle. This may reflect differences between the respective olfactory systems and central processing. For both species, the changes in trap catches in response to separation of pheromone components occurred at similar spacing distances as for separation of pheromone and anti-attractant sources. Overlap between two simulated plumes depended on distance between the two sources. In addition, the number of detected filaments and their concentration decreased with downwind distance. This implies that the response to separated odor sources in the two species might take place under different olfactory conditions. Deploying multiple sources of anti-attractant around a pheromone trap indicated long-distance (meter scale) effects of NHVon the beetle and a potential use for NHV in forest protection

Chemical composition, larvicidal and repellent activities of wild plant essential oils against Aedes aegypti

Bio-degradable and eco-friendly essential oils (EOs) extracted from Mentha longifolia, Salsola imbricata, Erigeron bonariensis, E. canadensis, Ailanthus altissima, and Zanthoxylum armatum were investigated for their repellent and larvicidal potential against Aedes aegypti mosquitoes. The EOs of M. longifolia, S. imbricata, E. bonariensis, E. canadensis, A. altissima, and Z. armatum exhibited 99.0%, 96.8%, 40.2%, 41.7%, 29.1%, and 13.2% repellency against mosquitoes at a tested dose of 33.3 μg/cm2, respectively. In time span bioassays, the EOs of M. longifolia, S. imbricata, E. bonariensis, and E. canadensis showed more than 40% repellency for 60 min at a tested dose of 330 μg/cm2. Larvicidal bioassays revealed that larvae of Ae. aegypti were the most susceptible to M. longifolia (LC50, 39.3 mg/L), E. bonariensis (LC50, 26.0 mg/L), E. canadensis (LC50, 35.7 mg/L), and Z. armatum (LC50, 35.9 mg/L) EOs upon 48 h exposure. The most abundant constituents in the EOs of M. longifolia, S. imbricata, E. bonariensis, E. canadensis and A. altissima were piperitone oxide (45.5%), carvone (39.9%), matricaria ester (43.1%), (31.7%) and eugenol (24.4%), respectively. Our study demonstrates that EOs of M. longifolia, S. imbricata, E. bonariensis, and E. canadensis might be used to control Ae. aegypti mosquitoes without harming humans or the environment

Styrene, (+)-trans-(1R,4S,5S)-4-Thujanol and Oxygenated Monoterpenes Related to Host Stress Elicit Strong Electrophysiological Responses in the Bark Beetle Ips typographus

Bark beetles kill apparently vigorous conifers during epidemics by means of pheromone-mediated aggregation. During non-endemic conditions the beetles are limited to use trees with poor defense, like wind-thrown. To find olfactory cues that help beetles to distinguish between trees with strong or weak defense, we collected volatiles from the bark surface of healthy felled or standing Picea abies trees. Furthermore, living trees were treated with methyl jasmonate in order to induce defense responses. Volatiles were analyzed by combined gas chromatography and electroantennographic detection (GC-EAD) on Ips typographus antennae. Compounds eliciting antennal responses were characterized by single sensillum recording for identification of specific olfactory sensory neurons (OSN). Release of monoterpene hydrocarbons decreased, while oxygenated compounds increased, from spring to early summer in felled trees. In both beetle sexes particular strong EAD activity was elicited by trace amounts of terpene alcohols and ketones. 4-Thujanol gave a very strong response and the absolute configuration of the tested natural product was assigned to be (+)-trans-(1R,4S,5S)-thujanol by stereoselective synthesis and enantioselective gas chromatography. One type of OSN responded to all ketones and five other OSN were characterized by the type of compounds that elicited responses. Three new OSN classes were found. Of the eight EAD-active compounds found in methyl jasmonate-treated bark, the known anti-attractant 1,8-cineole was the one most strongly induced. Our data support the hypothesis that highly active oxygenated host volatiles could serve as positive or negative cues for host selection in I. typographus and in other bark beetles

Prolonged Repellent Activity of Plant Essential Oils against Dengue Vector, Aedes aegypti

Repellents are effective personal protective means against outdoor biting mosquitoes. Repellent formulations composed of EOs are finding increased popularity among consumers. In this study, after an initial screening of 11 essential oils (EOs) at the concentration of 33 μg/cm2, five of the most repellent EOs, Perovskia atriplicifolia, Citrus reticulata (fruit peels), C. reticulata (leaves), Mentha longifolia, and Dysphania ambrosioides were further investigated for repellent activity against Aedes aegypti mosquitoes in time span bioassays. When tested at the concentrations of 33 μg/cm2, 165 μg/cm2 and 330 μg/cm2, the EO of P. atriplicifolia showed the longest repellent effect up to 75, 90 and 135 min, respectively, which was followed by C. reticulata (peels) for 60, 90 and 120 min, M. longifolia for 45, 60 and 90 min, and C. reticulata (leaves) for 30, 45 and 75 min. Notably, the EO of P. atriplicifolia tested at the dose of 330 μg/cm2 showed complete protection for 60 min which was similar to the commercial mosquito repellent DEET. Gas chromatographic-mass spectrometric analyses of the EOs revealed camphor (19.7%), limonene (92.7%), sabinene (24.9%), carvone (82.6%), and trans-ascaridole (38.8%) as the major constituents of P. atriplicifolia, C. reticulata (peels), C. reticulata (leaves), M. longifolia, and D. ambrosioides, respectively. The results of the present study could help develop plant-based commercial repellents to protect humans from dengue mosquitoes