Effects of blood-feeding on olfactory sensitivity of the malaria mosquito Anopheles gambiae: Application of mixed linear models to account for repeated measurements

Olfaction plays an important role in the host-seeking behavior of the malaria mosquito Anopheles gambiae. After a complete blood meal, female mosquitoes will not engage in host-seeking behavior until oviposition has occurred. We investigated if peripheral olfactory sensitivity changed after a blood meal by recording electroantennograms (EAGs) of female mosquitoes at three time points (2 h, 48 h and 72 h) to 15 volatile kairomones of either human origin or documented to emanate from oviposition sites. The EAG-sensitivity was compared with that of females of similar age post eclosion. As is common practice in electrophysiological studies, the EAG recordings were obtained by repeated stimulation of the same antennal preparations. We introduce mixed linear modeling as an improved statistical analysis for electrophysiological data. Two hours after blood ingestion, olfactory sensitivity as quantified through EAG-recording increased significantly and selectively, i.e. for seven compounds, compared to unfed females of the same age. Such short-term electrophysiological sensitization in the olfactory system as a result of feeding has not been documented before for insects. Sensitization to six compounds persisted until 48 h or 72 h post-blood meal at one or more concentrations. Desensitization was observed at 48 and 72 h pbm in response to two and three kairomones, respectively. For several compounds, sensitization at the EAG-level corresponded with sensitization found previously in single sensillum studies on olfactory neurons in antennal sensilla trichodea of An. gambiae females. These effects are likely to reflect sensitization to oviposition cues, as eggs have matured 48–72 h pbm. Knowledge of changes in olfactory sensitivity to kairomones can be applied to increase trap catches of malaria mosquitoes that have taken a blood meal and need to locate oviposition sites.     

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.     

Refugia and habitat partitioning among midges (Diptera: Chironomidae) in rain-pools

ABSTRACT.

• 1 Rain-pools on granite exposures are common in tropical Africa and they are inhabited by the larvae of two midge species, Chironomus imicola Kieffer and Chironomus pulcher Wiedemann.
• 2 These pools differ from other ephemeral island habitats such as carrion, fallen fruit, sap flows, mushrooms and dung in their high degree of spatial predictability. Most pools are predictably inhabited by either C.imicola or C.pulcher. This finding is considered in the light of contemporary competition theory.
• 3 An arrangement whereby the same species repeatedly invades the same temporary habitat indicates that there is something about the habitat itself causing the discrimination. I suggest that degree of exposure to sunlight may provide a cue allowing females to discriminate between pools as oviposition sites.
• 4 Both species survive the long dry season by retreating to pools of river water left by receding rivers. Dispersal to rain-pools by emerging adults occurs during the following season with imicola occupying rain-pools remote from shaded rivers. Appropriately, measured against pulcher, the biology of imicola is that of a colonizing species.
• 5 The possibility is discussed that the chironomids in rock-pools do not survive to breed. Instead, these populations may be maintained externally, for example by invasions from breeding populations in the Great Lakes of Africa, where larval densities may be too low to be detected by sampling. The evolutionary implications of such a situation are considered.

     

Use of climatic data and satellite imagery to model the abundance of Culicoides imicola, the vector of African horse sickness virus, in Morocco

African horse sickness (AHS) is a vector-borne, infectious disease of equids caused by African horse sickness virus (AHSV) . The only proven field vector of the virus is the biting midge Culicoides imicola. Following a recent epizootic (1989–91) of AHS in Morocco, light traps and automatic weather stations were operated for 2 years at twenty-two sites distributed over much of the country. The annually-averaged mean daily trap catch of C. imicola at these sites was negatively correlated with wind speed, and positively correlated with the average and mean annual minimum NDVI (Normalized Difference Vegetation Index, a remotely sensed measure of vegetation activity). There were no significant correlations between the mean daily trap catch and air temperature, soil temperature, relative humidity, saturation deficit, rainfall, altitude or the mean annual maximum or range of NDVI. The best two-variable model, which combined Windspeed_MnAvMn (the average daily minimum wind speed of the least windy month) and NDVI_min (the average annual minimum NDVI) as predictors, explained over 50% of the variance in the annually-averaged mean daily trap catch of C. imicola. There was a significant, positive correlation between minimum wind speed at night and the daily mortality rate of adult female C. imicola and it is suggested that the relationship between wind speed and the abundance of C. imicola arises from effects on adult mortality or dispersal. Considering several climatic variables, in North Africa NDVI_min was most significantly correlated with total annual rainfall. It is suggested that the relationship between NDVI_min and the abundance of C. imicola arises from the impact of soil moisture on both. It is proposed that areas of Morocco with higher levels of soil moisture in late summer or autumn provide more, larger and/or more enduring breeding sites for C. imicola, as well as supporting more photosynthetically active vegetation and hence having higher NDVI.     

Oviposition behaviour in Lygus rugulipennis: a morpho-functional study

Before oviposition, a Lygus rugulipennis (Poppius) (Heteroptera: Miridae) female explores the substrate with her mouthparts, performing ‘probing’ behaviour which eventually leads to a choice of oviposition site. In this study, morphological and behavioural observations were carried out in order to define the mechanisms through which female bugs recognise suitable oviposition sites. The morphological study was conducted using electron‐microscopy techniques, while the behavioural experiments aimed at the temporary (using ZnSO₄) or permanent (i.e., cauterization of the rostrum and ablation of both stylets and labium) inactivation of sensory structures located in the labium tip and in the stylets, which are probably involved in oviposition site selection. The tip of the labium of L. rugulipennis females is characterised by the presence of 11–12 uniporous gustatory sensilla which are innervated by 3–6 sensory neurones. One aporous mechanoreceptor is located ventrally on both areas of the labium tip. Other aporous mechanosensilla are found more proximally on the labium. ZnSO₄ labium tip treatment did not affect oviposition site selection, while stylet amputations, as well as rostrum cauterisations, resulted in almost complete oviposition failure. Labium tip‐ablated females oviposited similarly to control females. These results suggest that the sensory neurones associated with the stylets are involved in the location of the oviposition site. Uniporous gustatory sensilla on the labium tip are not involved in the final steps of the oviposition behaviour. However, they seem to play a role in assessing the suitability of the substrate as a food source.     

Behavioural and chemosensory responses of the turnip root fly (Delia floralis) to glucosinolates

This study investigates the behavioural and neural mechanisms involved in the oviposition behaviour of the turnip root fly,Delia floralis (Fallen). Behavioural studies showed that glucosinolates modulated the oviposition behaviour of the flies on artificial leaves as well as the number of eggs laid in the soil at the base of these leaves. Electrophysiological responses to glucosinolates were obtained from type A and type D sensilla on the prothoracic and mesothoracic tarsi, as well as from the long contact sensilla on the labellum. The neural responses from these sensilla were positively correlated with the oviposition behaviour of the flies and with the number of eggs laid. Of the eleven glucosinolates tested in the behavioural and electrophysiological tests, the flies were most responsive to glucobrassicanapin, gluconapin and glucobrassicin. The type D tarsal sensilla were more responsive to the glucosinolates than either the type A tarsal sensilla or the labellar sensilla. The structure-activity investigations showed that slight modifications to the chemical composition of the glucosinolates resulted in changes in neural activity.     

Protection of livestock against bluetongue virus vector Culicoides imicola using insecticide‐treated netting in open areas

The protection of livestock against Culicoides species (Diptera: Ceratopogonidae) using physical barriers or chemically treated barriers is difficult owing to the small size of these biting midges and animal welfare concerns associated with the reduction of air flow. Culicoides imicola Kieffer is the main bluetongue virus vector in the Mediterranean basin, including the southern Iberian peninsula, where livestock is mainly housed in open pens or sheds which offer no physical protection against C. imicola. In this study we assessed the efficacy of surrounding yearling ewe pens with a canvas barrier or a cypermethrin‐treated canvas barrier in reducing the entry of Culicoides spp. and C. imicola. Analyses were based on comparisons of Culicoides catches in traps in pens with and without barriers, and in traps located outside pens. Although there was no clear reduction in the abundance of Culicoides other than C. imicola in pens with either barrier, the C. imicola presence was markedly reduced by the insecticide‐treated barrier compared with the untreated barrier; the latter did not reduce the abundance of this species in pens. Estimates of the protection conferred against C. imicola by the treated barrier differed depending on whether catch comparisons were based on outside traps or on traps located inside no‐barrier pens. The results suggest that the use of insecticide‐treated barriers may reduce contact between livestock and C. imicola in open areas or sheds. More research is necessary to assess the degree of protection as a function of barrier height, C. imicola abundance, and the size of the area to be protected.     

Contact chemoreception related to host selection and oviposition behaviour in the monarch butterfly, Danaus plexippus

Abstract. Behavioural events during host selection by ovipositing monarch butterflies (Danaus plexippus (L.), Danainae, Nymphalidae) include tapping the leaf surface with fore-tarsi and touching this surface with mid-tarsi (‘drumming’) and antennae. Flavonoids identified from host plant extracts are known to stimulate oviposition. Scanning electron microscopy revealed the presence of contact-chemoreceptor sensilla on all appendages that contact the leaf surface. This electrophysiological study was conducted to identify the contact chemoreceptors that are sensitive to the known oviposition stimuli and are therefore probably involved in host recognition. Receptor cells of conspicuous sensilla grouped in clusters on fore-tarsi of females were sensitive to the behaviourally active butanol fraction of host plant (Asclepias curassavica) extract. However, these receptors generally had low sensitivity to three oviposition-stimulating flavonoids identified from this fraction, but they were also sensitive to the butanol fraction of a non-host (Brassica oleracea). Chemoreceptors in sensilla of the tarsomers 2–4 of the mid-legs also responded to the behaviourally active fraction of host plant extract and showed some sensitivity to two of the flavonoids that stimulate oviposition. Similar results were obtained from receptor cells in sensilla on the tip of the antennae. Most of these sensilla had cells responding to the butanol fraction of A. curassavica extract but only 25% of them were also sensitive to one of the behaviourally active flavonoids. These electrophysiological results, in combination with behavioural observations, suggest that host selection in monarch butterflies relies on a complex pattern of peripheral sensory information from several types of tarsal and antennal contact chemoreceptors.     

Sensilla and Cuticular Appendages on the Female Abdomen of Lasioptera rubi (Schrank) (Diptera,Cecidomyiidae)

Studies by SEM and TEM revealed 6 types of integumental appendages on female uromeres VIII-X in Lasioptera rubi: microtrichia, not innervated; spines, probably without sensory function; nonporous sensory hairs, each containing one dendrite ending with a tubular body indicating a tactile function; uniporous sensory hairs, each innervated partly by 3 dendrites indicating a chemosensory function, partly by an additional dendrite with a tubular body indicating a tactile function; scoop-like sensilla, each containing partly a branched structure of dendrites in the distal half of the sensillum indicating an olfactory function, partly an unbranched dendrite ending at a pore near the base of the sensillum, most probably registrating chemical stimuli by contact or gustation; finally, nonporous bristles, all or some of them innervated, in a manner indicating a tactile function. In addition, two scolopophorous proprioceptors were found inside uromere X. The nonporous sensory hairs, the uniporous sensory hairs and the scolopophores may be used by the midge to determine the mechanical and chemical properties of potential oviposition sites. The spines and nonporous bristles may function as conidia carriers.     

Functional bases of host-acceptance behaviour in the aphid parasitoid Aphidius ervi

Abstract.  The host acceptance behaviour in Aphidius ervi is investigated, assessing the role of both external and internal host-associated cues, offered to the experimental parasitoids with parafilm-made aphid dummies. The reaction to internal cues present in the host haemolymph is clearly evident, and its intensity is enhanced by external cues. Parasitoid females lay few eggs in aphid dummies filled with host haemolymph. A significant increase in the number of both oviposition reactions (host stinging) and egg laying is observed only when these dummies are coated with cornicle secretion. However, this enhancement is not observed when the aphid dummies contain distilled water. Thus, the host acceptance behaviour of A. ervi females appears to be controlled by the integration of both external and internal chemical cues. The physiological basis of this behavioural response is investigated with a detailed study on the anatomy and ultrastructure of A. ervi ovipositor. The detection of chemical cues present in the host haemolymph that act as kairomones is made possible by the presence of gustatory sensilla on the tip of the ovipositor. These sensilla consist of porous areas, reached by unbranched dendrites running inside both the lower valves (i.e. first valvulae) and the unpaired upper valve (i.e. second valvulae). The mechanosensory function during oviposition appears to be provided mostly by the basiconic sensilla found on the tip of external valves (i.e. third valvulae). A tentative functional model accounting for the observed oviposition behaviour of A. ervi is proposed.     

Influence of light and kairomone baiting systems on trap collections of biting midges in southern Sweden

Effective surveillance is essential for protecting livestock from Culicoides biting midges and the viruses they transmit. The objective of this study was to determine how the baiting system used in traps (UV, incandescent light, incandescent light with CO₂, and incandescent light with CO₂ and 1‐octen‐3‐ol) influences estimates of midge population abundance, parity, and diel activity. This was achieved through a standardized trapping protocol conducted in three habitats in Sweden. UV light traps caught the most Culicoides species and more C. obsoletus complex females than incandescent light traps. Traps baited with CO₂ plus 1‐octen‐3‐ol caught more female C. impunctatus than incandescent light traps. No consistent effect of bait type was found on C. obsoletus parity rate, as estimated from the proportion of midges with presence or absence of pigmentation. Midge activity, as reflected by trap catches, peaked between ‐3 h and +3 h relative to sunset, with UV traps catching significantly more female C. obsoletus complex and C. impunctatus at and after sunset than before sunset. We conclude that baiting system can influence biting midge collections, even using identical traps. Effective surveillance may require more than one bait type and kairomones to attract species that do not feed exclusively on cattle.     

Oviposition preferences in Grapholita molesta: the relative importance of visual and olfactory cues

The oriental fruit moth, Grapholita molesta (Busck) (Lepidoptera: Tortricidae), is a major pest of fruit trees worldwide. Females oviposit on the young leaves of peach [Prunus persica (L.) Batsch (Rosaceae)] shoots at twilight, when light intensity changes markedly. Previous researches have shown that the oviposition response of this moth is guided particularly by visual and olfactory cues from host plants. However, the relative importance of visual and olfactory cues in their oviposition preference is largely unknown. As a crepuscular moth, how do choices change as light intensity drops rapidly from day to night? In the present study, through two‐choice behavioural experiments, the oviposition responses of G. molesta to visual and olfactory cues (alone or in combination) from peach shoots were tested at four light intensities ranging from daylight to starlight. Grapholita molesta showed similar oviposition responses to plant cues under 1 000, 100, 1 and 0.01 mW m^?2 illumination. Olfactory cues from peach shoots attracted significantly more oviposition than the negative control, whereas visual cues alone did not. Furthermore, olfactory cues were more attractive than visual cues, and no significant interaction was observed between the responses to the two cues. Our findings indicate that G. molesta females mainly rely on olfactory cues to recognize oviposition sites, regardless of differences in light intensity. These results do not provide evidence for attraction to visual cues, but the possibility that the brightness of leaves might be used to guide oviposition is discussed.     

Morphological and functional heterogeneity in olfactory perception between antennae and maxillary palps in the pumpkin fruit fly,Bactrocera depressa

The morphology and ultrastructure of the olfactory sensilla on the antennae and maxillary palps were investigated through scanning electron microscopy (SEM) and transmission electron microscopy (TEM), and their responses to five volatile compounds were measured using electroantenogram (EAG) and electropalpogram (EPG) techniques in the pumpkin fruit fly, Bactrocera depressa (Shiraki; Diptera: Tephritidae). Male and female B. depressa displayed distinct morphological types of olfactory sensilla in the antennae and maxillary palps, with predominant populations of trichoid, basiconic, and coeloconic sensilla. Basiconic sensilla, the most abundant type of olfactory sensilla in the antennae, could be further classified into two different types. In contrast, the maxillary palps exhibited predominant populations of a single type of curved basiconic sensilla. High‐resolution SEM observation revealed the presence of multiple nanoscale wall‐pores on the cuticular surface of trichoid and basiconic sensilla, indicating that their primary function is olfactory. In contrast, coeloconic sensilla displayed several longitudinal grooves around the sensillum peg. The TEM observation of individual antennal olfactory sensilla indicates that the basiconic sensilla are thin‐walled, while the trichoid sensilla are thick‐walled. The profile of EAG responses of male B. depressa was different from their EPG response profile, indicating that the olfactory function of maxillary palps is different from that of antennae in this species. The structural and functional variation in the olfactory sensilla between antennae and maxillary palps suggests that each plays an independent role in the perception of olfactory signals in B. depressa.     

Characterisation of the arrestment responses of Trichogramma evanescens

Summary Contact kairomones and oviposition in a host egg stimulated arrestment behaviour in Trichogramma evanescens, characterised by a reduction in walking speed and increased turning. Previous oviposition experience did not influence a parasitoid's response to contact kairomones, but successive encounters with kairomone patches resulted in parasitoids habituating to the contact chemical. Oviposition on a kairomone patch did not reverse this habituation effect. It was concluded that contact kairomones and host eggs will both contribute independently to the duration of a patch visit. The selection of patches by T. evanescens will depend on its response to kairomones. Results from this study indicate that the application of contact kairomones to field crops will not necessarily increase the probability of parasitoids finding hosts.     

Volatile cues can drive the oviposition behavior in Odonata

Selection for the oviposition site represents the criterion for the behavioral process of habitat selection for the next generation. It is well known that in Odonata the most general cues are detected visually, but laboratory investigations on the coenagrionid Ischnura elegans showed through behavioral and electrophysiological assays that adults were attracted by olfactory cues emitted by prey and that males of the same species are attracted by female odor.The results of the present behavioral and electrophysiological investigations on I. elegans suggest the involvement of antennal olfactory sensilla in oviposition behavior. In particular, I. elegans females laid in the laboratory significantly more eggs in water from larval rearing aquaria than in distilled or tap water. Moreover, the lack of preference between rearing water and tap water with plankton suggests a role of volatiles related to conspecific and plankton presence in the oviposition site choice. I. elegans may rely on food odor for oviposition site selection, thus supporting the predictions of the “mother knows best” theory. These behavioral data are partially supported by electroantennographic responses. These findings confirm a possible role of olfaction in crucial aspects of Odonata biology.     

The influence of potential stressors on oviposition site selection and subsequent growth,survival and emergence of the non‐biting midge (Chironomus tepperi)

Theory predicts that animals should prefer habitats where their fitness is maximized but some mistakenly select habitats where their fitness is compromised, that is, ecological traps. Understanding why this happens requires knowledge of the habitat selection cues animals use, the habitats they prefer and why, and the fitness costs of habitat selection decisions. We conducted experiments with a freshwater insect, the non‐biting midge Chironomus tepperi to ask: (a) whether females respond to potential oviposition cues, (b) to explore whether oviposition is adaptive in relation to metal pollution and conductivity, and (c) whether individuals raised in poor quality sites are more likely to breed in similarly poor locations. We found the following: (a) females responded to some cues, especially conductivity and conspecifics, (b) females preferred sites with higher concentrations of bioavailable metals but suffered no consequences to egg/larval survival, (c) females showed some avoidance of high conductivities, but they still laid eggs resulting in reduced egg hatching, larval survival, and adult emergence, and (d) preferences were independent of natal environment. Our results show that C. tepperi is susceptible to ecological traps, depending on life stage and the relative differences in conductivities among potential oviposition sites. Our results highlight that (a) the fitness outcomes of habitat selection need to be assessed across the life cycle and (b) the relative differences in preference/suitability of habitats need to be considered in ecological trap research. This information can help determine why habitat preferences and their fitness consequences differ among species, which is critical for determining which species are susceptible to ecological traps.     

Can two species of midge coexist in a single puddle of rain-water?

Larvae of two midge species, Chironomus pulcher and Chironomus imicola normally occur in different puddles of rain-water; C. pulcher in shaded and C. imicola in sunny ones. When the sun/shade signal is unclear, both species can coexist for most of each rainy season. Yet, in the laboratory, C. pulcher shows total competitive exclusion of C. imicola. The puzzle is resolved by experiments and observations which suggest that some naturally occurring puddles possess characteristics not present in laboratory pools. These include a spatial refuge (deep water) and separate shaded and sunny parts. Both variables are normally found only in larger puddles, so coexistence is a function of patch size. On its own, each variable is a necessary but not a sufficient condition for coexistence.     

Imperfect oviposition decisions by the pitcher plant mosquito (Wyeomyia smithii)

Summary Wyeomyia smithii mosquitoes distribute their eggs across available oviposition sites (water-holding pitcher plant leaves) of varying quality. I experimentally examined responses to three components of site quality: conspecific larval density, larval density of the pitcher plant midge,Metriocnemus knabi, and pitcher size. Responses to larval treatments were complex and apparently suboptimal. Although mosquito larval performance is better in leaves with fewer conspecific and more midge larvae, females did not lay more eggs in such pitchers. Instead, more eggs were laid in experimental pitchers containing either midge or mosquito larvae, but fewer eggs in pitchers with neither or both. More eggs were laid in larger pitchers, which tend to accumulate more resources and dry out less often. Therefore, although the oviposition decisions made were suboptimal, they were better than random.     

Influence of carbon dioxide on numbers of Culicoides midges collected with suction light traps in South Africa

To implement risk management against diseases transmitted by species of Culicoides Latreille, 1809 (Diptera: Ceratopogonidae), it is essential to identify all potential vectors. Light traps are the most commonly used tool for the collection of Culicoides midges. Given the indiscriminate artificial attraction of light, traps will collect all night‐flying insects rather than only livestock‐associated Culicoides midges. Factors that may increase the efficacy of traps, especially for livestock‐associated Culicoides midges, require investigation. In the present study, results obtained with Centers for Disease Control (CDC) and Onderstepoort light traps baited with carbon dioxide (CO₂) were compared with those of unbaited controls. Comparisons were made using two replicates of a 4 × 4 randomized Latin square design. With both trap types, the mean numbers of Culicoides midges collected in 16 baited traps were higher than those caught in 16 unbaited traps. Although exceptionally low numbers were collected with the CDC traps, the increases in the numbers and frequency of collection of Culicoides imicola Kieffer, 1913 were more pronounced in the CDC traps compared with the Onderstepoort traps. These results indicate that the addition of CO₂ may increase the efficiency of these traps for the collection of C. imicola and other livestock‐associated Culicoides species.