Patrick M. Guerin

The Anopheles gambiae Odorant Binding Protein 1 (AgamOBP1) Mediates Indole Recognition in the Antennae of Female Mosquitoes

Haematophagous insects are frequently carriers of parasitic diseases, including malaria. The mosquito Anopheles gambiae is the major vector of malaria in sub-Saharan Africa and is thus responsible for thousands of deaths daily. Although the role of olfaction in A. gambiae host detection has been demonstrated, little is known about the combinations of ligands and odorant binding proteins (OBPs) that can produce specific odor-related responses in vivo. We identified a ligand, indole, for an A. gambiae odorant binding protein, AgamOBP1, modeled the interaction in silico and confirmed the interaction using biochemical assays. RNAi-mediated gene silencing coupled with electrophysiological analyses confirmed that AgamOBP1 binds indole in A. gambiae and that the antennal receptor cells do not respond to indole in the absence of AgamOBP1. This case represents the first documented instance of a specific A. gambiae OBP–ligand pairing combination, demonstrates the significance of OBPs in odor recognition, and can be expanded to the identification of other ligands for OBPs of Anopheles and other medically important insects.

Identification of host plant attractants for the carrot fly, Psila rosae

Cold-trapped carrot leaf volatiles were analyzed by gas chro-matography with an outlet splitter to a flame ionization detector and to a carrot fly antennogram preparation as the second detector (GC-EAD). Strongest EAD responses were elicited by products whose elution temperatures corresponded to the propenylbenzenes,trans-methylisoeugenol (3,4-dimethoxy-1-propenylbenzene) andtrans-asarone (2,4,5-trimethoxy-1-propenylbenzene) and, to a lesser extent, by-products matching the elution temperatures of the leaf aldehydes hexanal, (E)-2-hexenal, and heptanal, and of the terpenes linalool and caryophyllene. The identity of the propenylbenzenes was confirmed by gas chromatography-mass spectrom-etry. GC-EAD permitted accurate estimation of the olfactory thresholds; it was lowest fortrans-asarone at 500 attogram (5 × 10−16g)/ml of air passing over the antenna. Both the leaf aldehydes and propenylbenzenes were attractive when tested individually in the field with yellow sticky traps; fly captures were linearly related to the quantity of propenylbenzenes applied per trap. A combination oftrans-asarone and hexanal was more attractive than either compound singly, suggesting that the fly is adaptively equipped to respond to a mixture of compounds emanating from carrot foliage. In laboratory choice tests, flies were more attracted by vapors from intact carrot foliage than by that from a nonhost; leaf odor alone also mediated oviposition. We conclude that through the selectivity and sensitivity of its response to foliar volatiles, the carrot fly may achieve host-plant orientation and also at close range, in union with its response to less volatile leaf surface components, selection of an oviposition site.

The composition and timing of flower odour emission by wild Petunia axillaris coincide with the antennal perception and nocturnal activity of the pollinator Manduca sexta

In the genus Petunia, distinct pollination syndromes may have evolved in association with bee-visitation (P. integrifolia spp.) or hawk moth-visitation (P. axillaris spp). We investigated the extent of congruence between floral fragrance and olfactory perception of the hawk moth Manduca sexta. Hawk moth pollinated P. axillaris releases high levels of several compounds compared to the bee-pollinated P. integrifolia that releases benzaldehyde almost exclusively. The three dominating compounds in P. axillaris were benzaldehyde, benzyl alcohol and methyl benzoate. In P. axillaris, benzenoids showed a circadian rhythm with an emission peak at night, which was absent from P. integrifolia. These characters were highly conserved among different P. axillaris subspecies and P. axillaris accessions, with some differences in fragrance composition. Electroantennogram (EAG) recordings using flower-blends of different wild Petunia species on female M. sexta antennae showed that P. axillaris odours elicited stronger responses than P. integrifolia odours. EAG responses were highest to the three dominating compounds in the P. axillaris flower odours. Further, EAG responses to odour-samples collected from P. axillaris flowers confirmed that odours collected at night evoked stronger responses from M. sexta than odours collected during the day. These results show that timing of odour emissions by P. axillaris is in tune with nocturnal hawk moth activity and that flower-volatile composition is adapted to the antennal perception of these pollinators.

Behavioral attributes and parental care of Varroa mites parasitizing honeybee brood

Varroa jacobsoni, an ectoparasite of the Asian honeybee Apis cerana, has been introduced world-wide, and is currently decimating colonies of the European honeybee Apis mellifera. Varroas reproductive cycle is tuned to that of drone cells, those mainly parasitized in the original host. We describe here how a single fertilized female, infesting a brood cell, can produce two to four adult fertilized females within the limited time span of bee development (270 h in worker and 320 h in drone cells), despite the disturbance caused by cocoon spinning and subsequent morphological changes of the bee. From observations on transparent artificial cells we were able to show how the mite combats these problems with specialized behaviors that avoid destruction by the developing bee, prepares a feeding site for the nymphs on the bee pupa, and constructs a fecal accumulation on the cell wall which serves as a rendezvous site for matings between its offspring. The proximity of the fecal accumulation to the feeding site facilitates feeding by the maturing progeny. However, communal use of the feeding site leads to competition between individuals, and protonymphs are most disadvantaged. This competition is somewhat compensated by the timing of oviposition by the mites. Use of a common rendezvous and feeding site by two or more Varroa mothers in multiinfested cells may have developed from the parental care afforded to them as nymphs.

Darkness induces mobility, and saturation deficit limits questing duration, in the tick Ixodes ricinus.

SUMMARY The behaviour of Ixodes ricinus nymphs was recorded in 10-day experiments using computer-assisted video-tracking, in the absence of any host stimuli. These ticks switch spontaneously from questing in a desiccating atmosphere to quiescence in a water-saturated atmosphere after dark. Quantification of both questing and quiescence duration demonstrates that questing duration is inversely related to saturation deficit whereas quiescence duration is not. Distance walked after quiescence increased with desiccating conditions, while the distance walked after questing remained unchanged. Almost all locomotor activities of I. ricinus occurred during darkness under either a 14 h:10 h L:D or a 8 h:4 h L:D cycle. We established that all life stages of I. ricinus are equipped to sense shifts in light intensity with bilaterally placed strings of photoreceptors. This permits I. ricinus to use onset of darkness to trigger mobility when desiccation risk is reduced in nature.

Ammonia attracts the haematophagous bug Triatoma infestans : behavioural and neurophysiological data on nymphs

Abstract 1) Nymphs of the haematophagous bug Triatoma infestans (Heteroptera: Reduviidae) are attracted to volatiles from their own faeces on a servosphere. 2) Biological substrates attractive to triatomines release NH3: wetted triatomine faecal papers release NH3 at 256 ppb NH3 from a 60-g source and stale rabbit urine at 394 ppb from 200 ml. Ammonia released from aqueous NH3 also attracts bugs at doses of 3 ppb and 17 ppb on the servosphere. 3) Bugs typically show negative anemotaxis in a stimulus-free air-stream on the servosphere. At onset of stimulation with ammonia from either biological substrates or aqueous NH3 the bugs stop, move their antennae, turn and walk upwind, i.e. odour-mediated anemotaxis. 4) At lower NH3 doses a latency in attraction is recorded, but this latency disappears when the relative humidity of the stimulus delivery air-stream is dropped from 90 to 35%. 5) Electrophysiological recordings from single olfactory sensilla on antennae of Triatoma nymphs reveal two different types of NH3-excited receptors, both within grooved-peg sensilla. The responses of one of these receptor cells to NH3 has been studied in detail and shows that the action potential discharge rate is dose-dependent over the range 2–200 ppb. 6) The amplitudes of electroantennograms recorded from Triatoma nymphs to NH3 are dose dependent over the range 5–550 ppb.

Neurophysiological and behavioural evidence for an olfactory function for the dorsal organ and a gustatory one for the terminal organ in Drosophila melanogaster larvae.

Multicellular electrophysiological responses from the dorsal organ on the cephalic lobes of third instar Drosophila melanogaster larvae (wild-type Canton S) stimulated with a cold-trapped banana volatile extract showed that this structure has an olfactory function in the fruit fly. Responses of the dorsal organ were also recorded to constituents of the banana volatile extract as they eluted from a gas chromatographic column (GC-coupled dorsal organ electrophysiology). The active chemostimulants were identified as 2-heptanone, isoamyl alcohol, hexyl acetate, hexanol and hexyl butyrate by gas chromatography-coupled mass spectrometry. Applying the same recording system to the terminal organ sensilla, no responses were obtained to either the banana volatile bouquet or its constituents. By contrast, high frequency multicellular responses were recorded in response to touching the terminal organ with the gustatory stimuli KCl and grapefruit juice; responses were absent on similar stimulation of the dorsal organ with either NaCl or KCl. This suggests a role for olfaction by the dorsal organ and for gustation by the terminal organ in Drosophila larvae.In a 7-mm high wind tunnel with a thin 1.2% agar floor, the Drosophila larvae showed odour-conditioned upwind responses in an air stream of 0.1 m/s bearing banana volatiles. Drosophila larvae responded best to the odour of cut bananas. A 1:1 mixture of the banana odour constituents 2-heptanone and hexanol (at either 50 or 100 &mgr;g source dose each) proved as attractive as the known larval attractants propionic acid and isoamyl acetate on their own at 100 &mgr;g, whereas hexanol and 2-heptanone on their own at a 100 &mgr;g source dose were less attractive. The stronger behavioural response to the banana volatile bouquet and to the binary mixture serves to underline the multireceptor nature of the dorsal organ response to food odour in Drosophila.

Effect of mating frequency and brood cell infestation rate on the reproductive success of the honeybee parasite Varroa jacobsoni

Abstract. 1 The reproductve biology of Varroa jacobsoni, whose females infest honeybee brood, was studied in natural and transparent artificial brood cells. These investigations were made under the headings of maturation behaviour and fertilization, and the influence of infestation rate of brood cells on the number of mated females produced per infesting Varroa. 2 Mating of Varroa daughters, observed in the transparent brood cells with time‐lapse video, occurs just after ecdysis and as soon as they arrive on the faecal accumulation prepared by the mother. Such females are remated for as long as no other freshly moulted daughter arrives on the faecal accumulation. 3 The number of spermatozoa stocked in the spermatheca increases with remating, a strong indication for sperm mixing in this species when brood cells contain more than one Varroa foundress. 4 The number of daughters per infesting mother decreases at higher rates of infestation per cell, but the proportion of such daughters with a mate rises sharply due to the higher probability of finding a male within multi‐infested cells. The number of mated daughters per mother is maximal in cells with two foundress Varroa females. 5 The frequency distributions of infesting mites in drone cells are aggregated, and approximate to negative binomial distributions. 6 We postulate from the above that the observed non‐random infestation by Varroa in drone brood augments the mites mean reproductive success through the production of a higher number of mated daughters than the corresponding Poisson distributions would.

Oriented responses of the triatomine bugs Rhodnius prolixus and Triatoma infestans to vertebrate odours on a servosphere

Oriented responses of both R. prolixus and T. infestans adults were recorded on a servosphere to mouse-odour, one of its components (CO2), and to rabbit urine-odour. The volatiles were delivered in an air-stream under controlled conditions which excluded other sensory modalities. In stimulus-free air the triatomines walked preferentially downwind in straight bouts interrupted by stops or periods at relatively low speeds, all of variable duration. In odour-laden air, bugs maintained their typical walking habit but switched from negative to positive anemotaxis. The characteristic response to odour onset was to stop, sample the air with the antennae, turn upwind in situ, and then walk off in the direction of the source for at least a few seconds, i.e., odour mediated anemotaxis. Mouse-odour caused T. infestans to increase its speed to 5.3 cms-1. Both species continued with the upwind response for some time after odour delivery ceased, but the crosswind component of the tracks was more prominent during this period — an effort, we presume, by the bugs to re-contact an odour plume. This investigation provides unequivocal evidence for host finding in triatomines by olfactory cues alone.

Field attraction ofAgrotis segetum males in four european countries to mixtures containing three homologous acetates

Abstract(Z)-5-Decenyl, (Z)-7-dodecenyl, and (Z)-9-tetradecenyl acetate were tested as single compounds and blends for field attraction to the turnip moth in Denmark, France, Hungary, and Switzerland. Best attraction was obtained when all three components were present. Attractants for other noctuids are described, e.g.,Meristis trigrammica, Hoplodrina alsines, Erastria trabealis, Euxoa nigricans, Axylia putris, andAutographa gamma, which were caught with some of the components and their blends.