Tolerance to fermentation products in sugar reception: gustatory adaptation of adult butterfly proboscis for feeding on rotting foods

Adult Vanessa indica and Argyreus hyperbius frequently forage on flower nectar, but the former also utilizes tree sap and rotting fruits. Compared to flower nectar, these rotting foods are characterized by low sugar concentrations and the presence of fermentation products (ethanol and acetic acid). We suspected that gustatory responses by the receptors on the proboscis might differ in these species. Among the three sugars tested, sucrose elicited the greatest probing (behavioral) responses and was followed by fructose and glucose. A. hyperbius showed higher sugar sensitivity than V. indica in probing responsiveness. In electrophysiological responses of the proboscis sensilla, V. indica was slightly more sensitive than A. hyperbius to glucose and lower concentrations of the other sugars. The sugar reception in A. hyperbius was strongly inhibited by fermentation products, particularly acetic acid at natural concentrations. In contrast, V. indica was noticeably less susceptible to them than A. hyperbius, and its behavioral and sensory responses to sucrose were enhanced by 5-20% (w/v) ethanol. Thus, V. indica not only possesses tolerance to fermentation products but may perceive them as synergists for sugar reception. To utilize rotting foods, such tolerance might be more necessary than high sugar sensitivity.     

Identification of a gustatory receptor tuned to sinigrin in the cabbage butterfly Pieris rapae

Glucosinolates are token stimuli in host selection of many crucifer specialist insects, but the underlying molecular basis for host selection in these insects remains enigmatic. Using a combination of behavioral, electrophysiological, and molecular methods, we investigate glucosinolate receptors in the cabbage butterfly Pieris rapae. Sinigrin, as a potent feeding stimulant, elicited activity in larval maxillary lateral sensilla styloconica, as well as in adult medial tarsal sensilla. Two P. rapae gustatory receptor genes PrapGr28 and PrapGr15 were identified with high expression in female tarsi, and the subsequent functional analyses showed that Xenopus oocytes only expressing PrapGr28 had specific responses to sinigrin; when ectopically expressed in Drosophila sugar sensing neurons, PrapGr28 conferred sinigrin sensitivity to these neurons. RNA interference experiments further showed that knockdown of PrapGr28 reduced the sensitivity of adult medial tarsal sensilla to sinigrin. Taken together, we conclude that PrapGr28 is a gustatory receptor tuned to sinigrin in P. rapae, which paves the way for revealing the molecular basis of the relationships between crucifer plants and their specialist insects.     

Effect of age on the responsiveness of peripheral chemosensory sensilla of the turnip root fly (Delia floralis)

The effect of insect age on the neural responsiveness of gustatory sensilla was investigated. Electrophysiological recordings were obtained from type A and type D sensilla on the pro- and meso-thoracic tarsi, and from sensilla on the labellum of the turnip root fly,Delia floralis (Fallen) in response to potassium chloride, sucrose and sinigrin. The age of the fly did have an effect on the numbers of sensilla responding to the test stimuli and on the magnitude of the response, but there was no consistent pattern in these effects among sensilla. The labellar sensilla were more responsive to sucrose than the tarsal sensilla and the proportion of flies whose labellar sensilla responded to sucrose was initially low, but increased after day 2 of adult life. In contrast, the number of flies whose tarsal sensilla responded to stimulation with sucrose was initially high and decreased as the flies aged. There was a similar decline in the proportion of tarsal sensilla responding to potassium chloride. Neither the proportion of flies whose tarsal sensilla responded to sinigrin nor the magnitude of the response was influenced by the age of the fly. These finding are discussed in relation to the feeding and host selection behaviour of the fly.     

Morphological and electrophysiological differences in tarsal chemosensilla between the wild silkmoth Bombyx mandarina and the domesticated species Bombyx mori

Gustatory and olfactory senses of phytophagous insects play important roles in the recognition of host plants. In the domestic silkmoth Bombyx mori and its wild species Bombyx mandarina, the morphologies and responses of adult olfactory organs (antennae) have been intensely investigated. However, little is known about these features of adult gustatory organs and the influence of domestication on the gustatory sense. Here we revealed that both species have two types of sensilla (thick [T] and slim [S] types) on the fifth tarsomeres of the adult legs. In both species, females have 3.6–6.9 times more T-sensilla than males. Therefore, T-sensilla seem to play more important roles in females than in males. Moreover, gustatory cells of T-sensilla of B. mandarina females responded intensely to mulberry leaf extract in electrophysiological experiments, while T-sensilla of B. mori females (N4 strain) hardly responded to mulberry leaf extract. These results suggest that T-sensilla of B. mandarina females are involved in the recognition of oviposition sites. We also observed that, in three B. mori strains (N4, p50T, and Kinshu × Showa), the densities of sensilla on the fifth tarsomeres were much lower than in B. mandarina. These results indicate that domestication has influenced the tarsal gustatory system of B. mori.     

Antennal sugar sensitivity in the click beetle Agriotes obscurus

The occurrence of salt‐, sugar‐sensitive neurones and a mechanoreceptor neurone in the antennal hair‐like gustatory sensilla of the click beetle Agriotes obscurus L. (Coleoptera, Elateridae) is demonstrated using the electrophysiological sensillum tip‐recording technique. The stimulating effect of 13 water soluble sugars at 100 mm is tested on the neurones of these sensilla. Sucrose and fructose are the two most stimulating sugars for the sugar‐sensitive neurone, evoking almost 30 spikes s^?1 at 100 mm . The stimulating effect of arabinose, glucose, mannose, maltose and raffinose is three‐ to five‐fold lower, in the range 5.9–9.6 spikes s^?1. The remaining six sugars, xylose, galactose, rhamnose, cellobiose, trehalose and lactose, have very low (<1 spikes s^?1) or no ability to stimulate the sugar‐sensitive neurone. Concentration/response curves of the sugar‐sensitive neurone to sucrose, fructose and glucose at 0.01–100 mm overlap to a large extent in hibernating, cold reactivated and reproductively‐active beetles. A remarkable 9–50% decrease in the number of spikes evoked by 100 mm fructose and 10–100 mm sucrose occurs, however, in reproductively‐active beetles in June compared with beetles at the beginning of hibernation in October. These findings show that A. obscurus is capable of sensing a wide range sugars via their antennal gustatory sensilla.     

Sex-specific non-pheromonal taste receptors in Drosophila

Taste receptorshave recently been reported in Drosophila [1] and [2], but little is known of the relation between receptor and response. Morphological studies of the distribution of chemosensory sensilla indicate that the fruit fly has two major sites of gustation: the proboscis and the legs [3]. The taste sensilla on both these sites are similar in structure and each sensillum generally houses four gustatory neurons [4]. Early anatomical observations have demonstrated a sexual dimorphism in the number of tarsal sensilla [5] and in their central projections [6]. We measured the electrophysiological responses of the prothoracic taste sensilla to non-pheromonal substances—salts, sugars and water—and found a clear sexual dimorphism. From the response profile of individual sensilla, we were able to distinguish three types of tarsal sensilla in females as against only two types in males. The female-specific type, which responded specifically to sugar, was absent in males except when male gustatory neurons were genetically feminised. The fact that tarsal gustatory hairs exhibit a sexual dimorphism that affects the perception of non-pheromonal compounds suggests that sexual identity is more complex than has previously been thought [7] and [8].     

Alterations of ultrastructure and physiology of chemoreceptor dendrites in blowfly taste hairs treated with vinblastine and colchicine

Vinblastine (10^?3 M) and colchicine (10^?2 M) dissolved in fly Ringer solution were applied to chemoreceptor dendrites in individual tarsal taste hairs of blowflies (Phaenicia serricata Meigen), and the resultant changes in the morphology and physiological responsiveness of the receptors were examined. While Ringer solution alone did not have any morphological or physiological effects, treatment with drugs dissolved in Ringer solution increased the latency of responses to sucrose and NaCl and decreased spike amplitude and frequency. The drugs also disrupted microtubules and caused the destruction of the distal end of the dendrites. The observations provide evidence that the microtubules have a role in maintaining the structural integrity of the dendrites. Anodal current applied along with a chemical stimulus (NaCl or sucrose) increased the action potential frequency of treated and untreated receptors, and the electrical current elicited spikes only from those receptors which were activated by the chemical stimulus alone. When drug treatment resulted in a complete cessation of response to chemical stimuli, the receptors were also unresponsive to electrical stimulation. The results of this study are discussed in relation to hypotheses of transduction in chemoreception.     

Tarsal chemoreception in the polyphagous grasshopper Schistocerca americana: behavioural assays, sensilla distributions and electrophysiology

ABSTRACT Behavioural and electrophysiological responses of Schistocerca americana (Drury) (Orthoptera: Acrididae) to chemical stimulation of the tarsi were investigated. Using restrained insects, differences in leg-waving behaviour were observed following stimulation by sucrose and nicotine hydrogen tartrate (NHT), compared to control stimulations by water. Furthermore, free-walking insects were able to detect NHT on leaf surfaces, resulting in leg-raising to avoid tarsal contact.
SEM studies showed the presence of numerous peg chemoreceptor sensilla on the ventral surface of the tarsus. Tip recordings from such pegs showed activity from up to three chemosensitive neurones, plus a mechanoreceptor neurone. Stimulation by NaCl and KC1 elicited similar responses from two or three neurones in all sensilla tested, with increased firing rates at higher concentrations. Sucrose caused an increase in firing rate in few sensilla. In such cases several neurones were stimulated, and there was no evidence of a specific neurone sensitive to sucrose. In contrast, NHT elicited rapid firing in a single neurone, which was not sensitive to NaCl. Stimulation by NHT also inhibited the activity of the NaCl-sensitive neurones.
Possible mechanisms for chemical discrimination in S. americana tarsi are compared with those previously proposed for grasshopper mouthpart sensilla, and the significance of a NHT-sensitive neurone in tarsal sensilla is discussed.     

Ultrastructure of chemoreceptive tarsal sensilla in an armored harvestman and evidence of olfaction across Laniatores (Arachnida,Opiliones)

Harvestmen (Arachnida, Opiliones) are especially dependent on chemical cues and are often regarded as animals that rely mainly on contact chemoreception. Information on harvestman sensilla is scarce when compared to other arachnid orders, especially concerning internal morphology. Using scanning (SEM) and transmission (TEM) electron microscopy, we investigated tarsal sensilla on the distal tarsomeres (DT) of all leg pairs in Heteromitobates discolor (Laniatores, Gonyleptidae). Furthermore, we explored the typological diversity of sensilla present on the DT I and II in members of the suborder Laniatores, which include two thirds of the formally described opilionid fauna, using species from 17 families representing all main laniatorian lineages. Our data revealed that DT I and II of H. discolor are equipped with wall-pored falciform hairs (two types), wall-pored sensilla chaetica (two types) and tip-pored sensilla chaetica, while DT III and IV are mainly covered with trichomes (non-sensory) and tip-pored sensilla chaetica. The ultrastructural characteristics support an olfactory function for all wall-pored sensilla and a dual gustatory/mechanoreceptive function for tip-pored sensilla chaetica. Based on our comparative SEM survey, we show that wall-pored sensilla occur in all investigated Laniatores, demonstrating their widespread occurrence in the suborder and highlighting the importance of both legs I and II as the sensory appendages of laniatorean harvestmen. Our results provide the first morphological evidence for olfactory receptors in Laniatores and suggest that olfaction is more important for harvestmen than previously thought.     

Restricted diet breadth of the larvae of Antheraea assamensis and the role of the labrum‐epipharynx and galeal sensilla

The silkworm, Antheraea assamensis Helfer (Lepidoptera: Saturniidae), grows primarily on Persea bombycina and Litsea polyantha. To understand if the restricted diet breadth is due to the specific role of gustatory sensilla of the larvae of A. assamensis, the same fifth instar larvae retaining only labrum‐epipharynx or galeal sensilla were subjected to food choice tests. The foods used were leaves of two host‐plant and two non‐host‐plant species. Mean per cent consumption and per cent of choosing larvae were used as parameters for drawing conclusions. The finding indicated involvement of the labrum‐epipharynx for acceptance and galeal sensilla for rejection of a non‐host‐plant species. Scanning electron microscope studies revealed the presence of two sensilla on the galea, one lateral and one medial sensilla styloconicum and two gustatory sensilla in the epipharynx of A. assamensis. The study revealed the key role of galeal sensilla in the restrictive diet‐breadth of A. assamensis.     

Morphology and function of ovipositorial and tarsal sensilla of female Asian citrus psyllid

The Asian citrus psyllid (ACP), Diaphorina citri Kuwayama (Hemiptera:Psyllidae), is one of the most important pests of citrus plants worldwide, due to be the dominant vector of the devastating citrus disease Huanglongbing (HLB). Selection of suitable oviposition site is a crucial event of phytophagous insect life. Usually, the chemical and physical characteristics of a host plant were recognized by sensilla on the ovipositor and tarsi of a phytophagous insect after settling on a plant surface. In this study, the morphology, number, and distribution pattern of the ovipositorial and tarsal sensilla of adult female psyllids, were observed by scanning electron microscopy (SEM) techniques. The effect of these sensilla on oviposition was investigated by sensilla deactivation using hydrochloric acid (HCl). The results showed that sensilla trichoid (ST), sensilla chaetica (ChS) and sensilla basiconica (SB) were distributed on ovipositor, while sensilla trichoid (ST) were distributed on the external genitalia. Sensilla chaetica (ChS) and basiconica (SB) were mainly distributed on the end of the ovipositor back plate, and on the base of the sternum. On tarsus, sensilla trichoid (ST), sensilla chaetica (ChS), sensilla triangular (TeS) and sensilla ear ball (SE) were observed. However, whole tarsal segments were covered with Böhm's mane, which was arranged in a circular line. Oviposition assay revealed that a significantly lower number of eggs were laid on the Murraya paniculata seedlings confined by the psyllids with deactivated ovipositorial sensilla followed by protarsal sensilla, mesotarsal sensilla and hindtarsal sensilla. These results suggested that the ovipositorial and tarsal sensilla of citrus psyllid have an important role in the oviposition.     

Fine Structure of Antennal Sensilla of Paysandisia archon and Electrophysiological Responses to Volatile Compounds Associated with Host Palms

Paysandisia archon (Lepidoptera: Castniidae) is a serious pest of palm trees. A comprehensive knowledge of the insect olfactory system is essential for the development of efficient semiochemical-based control methods. The olfactory sensilla are located particularly on the antennae, and these can detect plant volatiles that provide important cues for the insects in the search for their host plants. To date, the fine structure of P. archon antennal sensilla studies and their role in host-plant perception have not been investigated in great detail. Using light microscopy and scanning and transmission electron microscopy, the antennae of both sexes of P. archon are described here in detail, according to the different types, quantities and distributions of the sensilla. Six types of sensilla were identified. The most widespread are sensilla trichoidea, sensilla basiconica and sensilla auricilica, which are associated with olfactory function. These have cuticular shafts characterised by numerous pores, and they are innervated by two or three sensory neurons. Sensilla coeloconica, sensilla chaetica and sensilla ampullacea are associated with olfactory or olfactory-thermoreception, mechano-gustatory, and thermo-hygroreception functions, respectively. Moreover, the role of P. archon antennae in locating of the host palms was evaluated using electroantennograms, to monitor responses to ester and terpene compounds previously identified as volatiles of damaged/fermenting palm tissues. P. archon showed responses to all of the synthetic chemicals tested, with greater responses in the females, providing a significant sex*dose effect. Among the compounds tested, ethyl isobutyrate elicited the strongest antenna responses. The fine structure of the cuticular and cellular components of the P. archon antenna sensory equipment is described for the first time. The results of this study form an important starting point and complement physiological and behavioural studies, to provide valuable information of practical importance for the development of efficient semiochemical-based control methods.     

Electrophysiological studies of gustation in lepidopterous larvae

Summary The electrophysiological responses of the maxillary gustatory sensilla of ten species of lepidopterous larvae to sugars, polyhydric alcohols, amino acids, organic acids, glycosides, and sterols were tested. Of the caterpillars examined three were monophagous (Calpodes ethlius, Ceratomia catalpae, Danaus plexippus), three were oligophagous (Pieris rapae, Papilio polyxenes, Malacosoma americana), and four were polyphagous (Helioihis zea, Estigmene acrea, Isia isabella, Porthetria dispar). The sensilla of all species are structurally similar to the extent of having four chemosensory bipolar neurons in each peg.It is not possible to account for the wide range of responsiveness to diverse compounds in terms of narrowly specific receptors characterized as water, sugar, salt, inositol, or amino acid receptors. Bach receptive cell was found to have only a limited degree of specificity. Not all amino acids stimulated the same cell. Some amino acids stimulated only one; some stimulated two; some inhibited all cells; some inhibited one while stimulating another. Differential sensitivity to amino acids varied from species to species and even between the two sensilla in an individual.The diversity of relationships found with amino acids existed also with respect to acids and glycosides. Ascorbic and hydrochloric acids at equivalent pH values each stimulated a different cell. Glycosides stimulated one or two cells depending on the species and sensillum.Synergism occurred nearly as often as inhibition. Among the instances were synergistic reactions between sucrose and inositol, proline and inositol, sucrose and sinigrin, glycine and sucrose. The occurrence of the phenomenon was specific to certain species and sensilla.Results point to the conclusion that the basis of difference in sensitivity to different compounds from species to species and from receptor to receptor reflects variations in the molecular structure of the receptor substrate. Whatever the mechanisms involved it is clear that caterpillars are able to detect not only compounds that may be of nutritive value but also non-nutritive compounds some of which are secondary plant substances. When the total taste sensitivities of the various species of caterpillars are compared, there is no obvious difference between polyphagous, oligophagous, and monophagous species.This work was supported by National Science Foundation Grant GB 1472.     

The interaction of external and internal receptors on the feeding behaviour of the blowfly, Phormia regina

The interaction of sensory activity from internal gut stretch receptors and from external labellar chemosensory hairs has been studied both behaviourally and electrophysiologically in the control of proboscis extension of the blowfly, Phormia regina. Labellar thresholds for proboscis extension, tested behaviourally, do not change significantly up to an hour after feeding in contrast to tarsal thresholds which rise quickly after feeding. Motor activity of the extensor muscle of the haustellum was recorded simultaneously with sensory activity from labellar sensilla. The mean number of muscle spikes per response and the sensory input necessary to trigger a response do not vary with starvation, feeding, or sectioning of the recurrent nerve. Activity of internal stretch receptors seem to interact with tarsal sensory input but apparently do not modulate motor responses triggered by labellar sensory input.     

Deterrence of feeding and oviposition responses of adult Heliothis virescens by some compounds bitter-tasting to humans

Some compounds that are bitter-tasting to humans, both alkaloidal (quinine, quinidine, atropine, caffeine) and non-alkaloidal (denatonium benzoate, sucrose octaacetate, naringin), deterred feeding and oviposition by Heliothis virescens (F.) in laboratory and field cage experiments. Preliminary electrophysiological studies of gustatory sensilla on the ovipositor of H. virescens provided evidence of 3 neurons, one of which is responsive to sucrose. Preliminary indications are that responses of this neuron may be inhibited by quinine and denatonium benzoate.     

Antennal gustatory perception and behavioural responses in Trissolcus brochymenae females

Trissolcus brochymenae (Hymenoptera: Platygastridae) is an egg parasitoid that could be used to control stink bugs like Murgantia histrionica (Heteroptera: Pentatomidae), a pest of brassicaceous crops. Before laying their eggs, parasitoid females spend considerable time examining the substrate with their antennae, which are also used during feeding and mating behaviours. This suggests that contact chemoreception plays a prominent role in many aspects of parasitoid ecology. Therefore, we examined the sensitivity of antennal taste sensilla chaetica to several classical tastants including those that are appetitive or aversive. In addition we studied the taste preferences of wasps when presented with these chemicals alone or mixed. The sensilla chaetica of T. brochymenae responded to serial concentrations of sucrose, salts, and quinine, but no concentration-dependent effect was observed when testing sinigrin, a secondary metabolite found in many brassicaceae. However, both sinigrin and quinine inhibited responses to 0.1 M sucrose when mixed with this sugar. Behavioural taste preference assays confirmed that wasps showed a dose dependent preference for sucrose over agarose. In addition, a behavioural avoidance of sucrose solutions containing quinine was observed. This effect was not observed when sinigrin was used as a feeding deterrent. In the two-choice tests the wasp did not discriminate between sucrose solutions mixed with salts and sucrose alone. Further no preference for salts or sinigrin compared to agarose alone was observed. This work represents the first step towards the identification of gustatory receptor neurons implicated in the detection of different types of chemical cues in egg parasitoids.     

The role of contact chemoreception in the host location process of an egg parasitoid

Taste allows insects to detect palatable or toxic foods, identify a mate, and select appropriate oviposition sites. The gustatory system strongly contributes to the survival and reproductive success of many species, yet it is rarely studied in insect parasitoids. In order to locate and assess a host in which they will lay their eggs, female wasps actively search for chemical cues using their sensory organs present mainly on the antennae. In this paper, we studied the role of antennal taste sensilla chaetica in the perception of contact semiochemicals in Trissolcus brochymenae (Hymenoptera: Platygastridae), an egg parasitoid of the brassicaceae pest Murgantia histrionica (Heteroptera: Pentatomidae). Methanolic extracts obtained from male and female hosts elicited action potentials in taste neurons housed in antennal sensilla chaetica, indicating that these sensilla are involved in the perception of non volatile host kairomones. In behavioural assays, wasp females displayed an intense searching behaviour in open arenas treated with host extracts, thus confirming that these kairomones are soluble in polar solvents. We further investigated the extracts by Gas Chromatography-Mass Spectrometry (GC-MS) and found that they contain several compounds which are good candidates for these contact kairomones. This study contributes to better understanding contact chemoreception in egg parasitoids and identifying gustatory receptor neurons involved in the host location process.     

Sense organs on palps and fore tarsi of gamasid mites (Parasitiformes, Rhinonyssidae), parasites of the nasal cavity of the Great Tit, the Rock Dove, and the Eurasian Coot

Palpal and tarsal sensilla of mites parasitizing in the nasal cavity of the Great Tit Parus major (Ptilonyssus sairae, Ptilonyssus pari), the Rock Dove Columba livia (Mesonyssus melloi), and the Eurasian Coot Fulica atra (Rallinyssus caudistigmus), were examined under a scanning electron microscope. Differences in the topography of the tarsal organ reflect phylogenetic relations of species and genera, whereas differences in the structure of palpal receptors reflect the ecological peculiarities of parasitism.     

Morphological and electrophysiological characteristics of the epipharyngeal sensilla of the silkworm, Bombyx mori

One pair of gustatory sensilla was found on the epipharynx ofBombyx mori larvae, and some morphological and electrophysiological characteristics of the epipharyngeal sensilla were investigated. They are sensilla coeloconica composed of a small papilla with a pore at the tip and a swelling of cuticle encircling the papilla. Three bipolar neurons innervate each sensillum. One neuron is an inositol receptor which responds to inositol only. Another cell responds with action potentials of relatively large amplitude to some feeding deterrent substances, such as strychnine nitrate. The thresholds of these cells for inositol and strychnine nitrate are approximately 10⁻⁴ M and 10⁻⁷ M, respectively. At least two kinds of spikes can be observed when these sensilla are stimulated with some salts and acids. Dose-response relationships and time courses of responses to inositol and strychnine nitrate were also examined in this study.     

Morphology and distribution of antennal sensilla in the pine wood nematode insect vectors Monochamus alternatus Hope and Monochamus saltuarius Gebler (Coleoptera: Cerambycidae)

As insect vectors of pine wood nematodes, Monochamus alternatus Hope and Monochamus saltuarius Gebler have different levels of attractiveness to semiochemicals. Although active use of chemical communication in both species is well known, there was no report on the differences in morphology and distribution of antennal sensilla between the two Monochamus species. The present study was carried out to explore the morphology, distribution, and ultrastructure of antennal sensilla of the two species using scanning and transmission electron microscopy. The overall shapes of the antennae were not different between M. alternatus and M. saltuarius. However, both M. alternatus and M. saltuarius exhibited sexual dimorphism overall. The antennae of each species were composed of the scape, pedicel, and nine flagellomeres, with four morphological types of sensilla: trichodea, basiconica, chaetica, and coeloconica. The four types occurred in varying numbers and distributions. The sensilla chaetica type was the most abundant in both species. Three types of sensilla (trichodea, basiconica, and chaetica) could be subdivided according to the two Monochamus species; sensilla trichodea could be divided into two subtypes, and sensilla basiconica and sensilla chaetica into three. All sensilla basiconica subtypes were multiporous, indicating that their primary function was olfactory. Sensilla trichodea subtype 2 was uniporous, indicating that the function of these sensilla might be gustatory. The morphological information obtained in our study provides a basis for future investigation of the sensory physiological function of each type of sensilla in the two species.