Mango headspace volatiles trigger differential responses of the mango fruit fly Ceratitis cosyra and its parasitoids.

Before the introduction of Bactrocera dorsalis (Hendel) to sub-Saharan Africa, Ceratitis cosyra (Walker) was economically the most important pest in mango farming. Its native natural enemy, the solitary parasitoid Psyttalia cosyrae (Wilkinson), played a crucial role in C. cosyra bio-control, later complemented by the exotic parasitoids Diachasmimorpha longicaudata (Ashmead) and Fopius arisanus (Sonan) among Integrated Pest Management (IPM) systems. To understand the in situ mango-C. cosyra-parasitoid tritrophic interaction, we assessed the responses of the fruit fly and the three parasitoids to headspace volatiles from various mango conditions. These conditions included non-infested mature unripe mangoes, C. cosyra-infested mangoes, 7th- and 9th-day post-infestation mangoes, non-infested ripe mangoes of three varieties (Kent, Apple, and Haden), and clean air (blank). We also compared the fruit fly's performance in the mango varieties and identified the chemical profiles of mango headspace volatiles. Ceratitis cosyra was attracted to both infested and non-infested mangoes (66-84 % of responsive C. cosyra) and showed superior performance in Kent mango (72.1 % of the 287 puparia recovered) compared to Apple and Haden varieties. Fopius arisanus displayed a stronger attraction to the volatiles of C. cosyra-infested mangoes (68-70 %), while P. cosyrae and D. longicaudata were significantly attracted to the 9th-day post-infestation mangoes (68-78 %) compared to non-infested mango volatiles. Gas chromatography-mass spectroscopy showed substantial quantitative and qualitative differences in volatile profiles among mango treatments. Esters predominated in non-infested ripe, 7th- and 9th-day post-infestation mangoes, while monoterpenes and sesquiterpenes were most dominant in the other treatments. The in situ experiments underscored varying preferences of the species for mango headspace volatiles and their subsequent treatments. These results provide valuable insights for further exploration, specifically in identifying the key volatiles responsible for species responses, to facilitate the development of applicable selective semiochemicals for managing species of African fruit fly.

The maxillary palps of Tephritidae are selectively tuned to food volatiles and diverge with ecology.

The maxillary palp is an auxiliary olfactory organ in insects, which, different from the antennae, is equipped with only a few olfactory sensory neuron (OSN) types. We postulated that these derived mouthpart structures, positioned at the base of the proboscis, may be particularly important in mediating feeding behaviors. As feeding is spatio-temporally segregated from oviposition in most Tephritidae, this taxonomic group appears quite suitable to parse out sensory breadth and potential functional divergence of palps and antennae. Scanning electron microscopy and anterograde staining underlined the limited palpal olfactory circuit in Tephritidae: only three morphological subtypes of basiconic sensilla were found, each with two neurons, and project to a total of six antennal lobe glomeruli in Bactrocera dorsalis. Accordingly, the palps detected only few volatiles from the headspace of food (fermentation and protein lures) and fruit (guava and mango) compared to the antennae (17 over 77, using gas-chromatography coupled electrophysiology). Interestingly, functionally the antennae were more tuned to fruit volatiles, detecting eight times more fruit than food volatiles (63 over 8), whereas the number of fruit and food volatile detection was more comparable in the palps (14 over 8). As tephritids diverge in oviposition preferences, but converge on food substrates, we postulated that the receptive ranges of palpal circuits would be more conserved compared to the antennae. However, palpal responses of three tephritid species that differed in phylogenetic relatedness and ecologically niche, diverged across ecological rather than phylogenetic rifts. Two species with strongly overlapping ecology, B. dorsalis and Ceratitis capitata, showed inseparable response profiles, whereas the cucurbit specialist Zeugodacus cucurbitae strongly diverged. As Z. cucurbitae is phylogenetically placed between B. dorsalis and C. capitata, the results indicate that ecology overrides phylogeny in the evolution of palpal tuning, in spite of being predisposed to detecting food volatiles.

The push-pull intercrop Desmodium does not repel, but intercepts and kills pests.

Over two decades ago, an intercropping strategy was developed that received critical acclaim for synergizing food security with ecosystem resilience in smallholder farming. The push-pull strategy reportedly suppresses lepidopteran pests in maize through a combination of a repellent intercrop (push), commonly Desmodium spp., and an attractive, border crop (pull). Key in the system is the intercrop's constitutive release of volatile terpenoids that repel herbivores. However, the earlier described volatile terpenoids were not detectable in the headspace of Desmodium, and only minimally upon herbivory. This was independent of soil type, microbiome composition, and whether collections were made in the laboratory or in the field. Furthermore, in oviposition choice tests in a wind tunnel, maize with or without an odor background of Desmodium was equally attractive for the invasive pest Spodoptera frugiperda. In search of an alternative mechanism, we found that neonate larvae strongly preferred Desmodium over maize. However, their development stagnated and no larva survived. In addition, older larvae were frequently seen impaled and immobilized by the dense network of silica-fortified, non-glandular trichomes. Thus, our data suggest that Desmodium may act through intercepting and decimating dispersing larval offspring rather than adult deterrence. As a hallmark of sustainable pest control, maize-Desmodium push-pull intercropping has inspired countless efforts to emulate stimulo-deterrent diversion in other cropping systems. However, detailed knowledge of the actual mechanisms is required to rationally improve the strategy, and translate the concept to other cropping systems.

Species-Specific Induction of Plant Volatiles by Two Aphid Species in Apple: Real Time Measurement of Plant Emission and Attraction of Lacewings in the Wind Tunnel.

Upon damage by herbivores, plants release herbivory-induced plant volatiles (HIPVs). To find their prey, the pest's natural enemies need to be fine-tuned to the composition of these volatiles. Whereas standard methods can be used in the identification and quantitation of HIPVs, more recently introduced techniques such as PTR-ToF-MS provide temporal patterns of the volatile release and detect additional compounds. In this study, we compared the volatile profile of apple trees infested with two aphid species, the green apple aphid Aphis pomi, and the rosy apple aphid Dysaphis plantaginea, by CLSA-GC-MS complemented by PTR-ToF-MS. Compounds commonly released in conjunction with both species include nonanal, decanal, methyl salicylate, geranyl acetone, (Z)-3-hexenyl acetate, (Z)-3-hexenyl butanoate, (Z)-3-hexenyl 2-methyl-butanoate, (E)-ß-caryophyllene, ß-bourbonene and (Z)-3-hexenyl benzoate. In addition, benzaldehyde and (E)-ß-farnesene were exclusively associated with A. pomi, whereas linalool, (E)-4,8-dimethyl-1,3,7-nonatriene were exclusively associated with D. plantaginea. PTR-ToF-MS additionally detected acetic acid (AA) and 2-phenylethanol (PET) in the blends of both trees attacked by aphid species. In the wind tunnel, the aphid predator, Chrysoperla carnea (Stephens), responded strongly to a blend of AA and PET, much stronger than to AA or PET alone. The addition of common and species-specific HIPVs did not increase the response to the binary blend of AA and PET. In our setup, two host-associated volatiles AA + PET appeared sufficient in the attraction of C. carnea. Our results also show the importance of combining complementary methods to decipher the odor profile associated with plants under pest attack and identify behaviourally active components for predators.

bric à brac controls sex pheromone choice by male European corn borer moths.

The sex pheromone system of ~160,000 moth species acts as a powerful form of assortative mating whereby females attract conspecific males with a species-specific blend of volatile compounds. Understanding how female pheromone production and male preference coevolve to produce this diversity requires knowledge of the genes underlying change in both traits. In the European corn borer moth, pheromone blend variation is controlled by two alleles of an autosomal fatty-acyl reductase gene expressed in the female pheromone gland (pgFAR). Here we show that asymmetric male preference is controlled by cis-acting variation in a sex-linked transcription factor expressed in the developing male antenna, bric à brac (bab). A genome-wide association study of preference using pheromone-trapped males implicates variation in the 293 kb bab intron 1, rather than the coding sequence. Linkage disequilibrium between bab intron 1 and pgFAR further validates bab as the preference locus, and demonstrates that the two genes interact to contribute to assortative mating. Thus, lack of physical linkage is not a constraint for coevolutionary divergence of female pheromone production and male behavioral response genes, in contrast to what is often predicted by evolutionary theory.

Dispersal and competitive release affect the management of native and invasive tephritid fruit flies in large and smallholder farms in Ethiopia.

African horticulture is seriously affected by fruit flies, both native and invasive. Novel sustainable control methods need testing against the backdrop of smallholder-dominated farming of Africa. We evaluated the potential of male-specific attractants (parapheromones) laced with insecticide to suppress the alien invasive Bactrocera dorsalis and native Ceratitis capitata. In large-scale guava, methyl-eugenol (ME)-bait stations combined with toxic protein baits suppressed B. dorsalis within 8 months but resulted in a resurgence of the displaced Ceratitis capitata. In smallholder farms, intervention using SPLAT-ME laced with spinosad was surprisingly unsuccessful. Subsequent mark-release-recapture experiments showed high dispersal rates of flies, covering many times a typical farm size, leading to a continuous influx of flies from surrounding areas. Several other factors important for intervention were evaluated. SPLAT-MAT-ME dollops remained attractive for over two weeks, although gradually becoming less attractive than fresh baits. Further, competitive displacement was observed: C. capitata selectively emerged from fruits in which B. dorsalis infestation was low. Finally, we evaluated whether ME could be combined with C. capitata male attractants [trimedlure (TML) and terpinyl acetate (TA)] without affecting attraction. Combining male lures did not affect catches directly, although at very high populations of B. dorsalis attracted to ME interfered with C. capitata trap entry. Although ME-based methods can effectively suppress B. dorsalis, they were not effective at single smallholder scale due to the high dispersive propensity of tephritids. Further, competitive release implies the need for a combination of lures and methods. These observations are important for developing control schemes tailored for African smallholder settings.

Is Anopheles gambiae attraction to floral and human skin-based odours and their combination modulated by previous blood meal experience?

BACKGROUND: Mosquitoes use odours to find energy resources, blood hosts and oviposition sites. While these odour sources are normally spatio-temporally segregated in a mosquito's life history, here this study explored to what extent a combination of flower- and human-mimicking synthetic volatiles would attract the malaria vector Anopheles gambiae sensu stricto (s.s.) METHODS: In the laboratory and in large (80 m2) outdoor cages in Tanzania, nulliparous and parous A. gambiae s.s. were offered choices between a blend of human skin volatiles (Skin Lure), a blend of floral volatiles (Vectrax), or a combination thereof. The blends consisted of odours that induce distinct, non-overlapping activation patterns in the olfactory circuitry, in sensory neurons expressing olfactory receptors (ORs) and ionotropic receptors (IRs), respectively. Catches were compared between treatments. RESULTS: In the laboratory nulliparous and parous mosquitoes preferred skin odours and combinations thereof over floral odours. However, in semi-field settings nulliparous were significantly more caught with floral odours, whereas no differences were observed for parous females. Combining floral and human volatiles did not augment attractiveness. CONCLUSIONS: Nulliparous and parous A. gambiae s.s. are attracted to combinations of odours derived from spatio-temporally segregated resources in mosquito life-history (floral and human volatiles). This is favourable as mosquito populations are comprised of individuals whose nutritional and developmental state steer them to diverging odours sources, baits that attract irrespective of mosquito status could enhance overall effectiveness and use in monitoring and control. However, combinations of floral and skin odours did not augment attraction in semi-field settings, in spite of the fact that these blends activate distinct sets of sensory neurons. Instead, mosquito preference appeared to be modulated by blood meal experience from floral to a more generic attraction to odour blends. Results are discussed both from an odour coding, as well as from an application perspective.

Designing a species-selective lure based on microbial volatiles to target Lobesia botrana.

Sustainable, low impact control methods, including mating disruption and microbial insecticides against L. botrana have been available for decades. Yet, successful implementation has been restricted to only a few grapevine districts in the world. A limiting factor is the lack of a female attractant to either monitor or control the damaging sex. Volatile attractants for both female and male insects can be used to assess when L. botrana populations exceed economic thresholds, and to decrease the use of synthetic pesticides within both conventional and pheromone programs. Rather than using host-plant volatiles, which are readily masked by background volatiles released by the main crop, we tested the attractiveness of volatiles that signify microbial breakdown and more likely stand out against the background odour. A two-component blend of 2-phenylethanol (2-PET) and acetic acid (AA) caught significant numbers of both sexes. Catches increased with AA and, to a minimal extent, 2-PET loads. However, a higher load of 2-PET also increased bycatches, especially of Lepidoptera and Neuroptera. Major (ethanol, ethyl acetate, 3-methyl-1-butanol) or minor (esters, aldehydes, alcohols and a ketone) fermentation volatiles, did surprisingly not improve the attraction of L. botrana compared to the binary blend of 2-PET and AA alone, but strongly increased bycatches. The most attractive lure may thus not be the best choice in terms of specificity. We suggest that future research papers always disclose all bycatches to permit evaluation of lures in terms of sustainability.

A zooprophylaxis strategy using L-lactic acid (Abate) to divert host-seeking malaria vectors from human host to treated non-host animals.

BACKGROUND: Zooprophylaxis is a technique in which blood-seeking vectors are diverted to non-host animals in order to lower blood-feeding rates on human hosts. The success of this technique depends on the host preference of the vector being targeted. The objective of this study was to evaluate the effect of L-lactic acid (Abate) to divert malaria mosquito, Anopheles gambiae from feeding on human host. METHODS: A 14-month-old female goat was treated with Abate, a formulation incorporating L-lactic acid into a slow-release matrix. This formulation was applied on the fur of the goat's back and neck. The treated animal was then presented to Anopheles gambiae sensu stricto (s.s.) as a prospective host in a semi-field environment ('mosquito sphere') together with either an untreated animal or a human. The number of mosquitoes caught to each host choice offered were compared. RESULTS: Goat treated with the L-lactic acid formulation successfully attracted An. gambiae at higher rates (70.2%) than the untreated ones (29.8%). Furthermore, An. gambiae s.s. were attracted to a treated goat at an equivalent degree (47.3%) as to their preferred human host (52.7%), even when the preferred host was present in the same environment. CONCLUSIONS: The findings indicate that human host-seeking mosquitoes can be diverted into feeding on non-preferred hosts despite the close proximity of their favoured host, hence reducing chances for the transmission of blood-borne parasites.

Hold your breath - Differential behavioral and sensory acuity of mosquitoes to acetone and carbon dioxide.

Host seeking in the yellow fever mosquito, Aedes aegypti, and the African malaria mosquito, Anopheles coluzzii, relies on specific and generic host-derived odorants. Previous analyses indicate that the behavioral response of these species depends differentially on the presence of carbon dioxide (CO2) and other constituents in human breath for activation and attraction. In this study, we use a flight tube assay and electrophysiological analysis to assess the role of acetone, a major component of exhaled human breath, in modulating the behavioral and sensory neuronal response of these mosquito species, in the presence and absence of CO2. When presented alone at ecologically relevant concentrations, acetone increases attraction in Ae. aegypti, but not in An. coluzzii. Moreover, in combination with CO2, human breath-equivalents of acetone ranging between 0.1 and 10 ppm reproduces a behavioral response similar to that observed to human breath in host-seeking Ae. aegypti, but not in An. coluzzii. Acetone does, however, reduce attraction to CO2 in An. coluzzii, when presented at a higher concentration of 10 ppm. We identify the capitate peg A neuron of the maxillary palp of both species as a dual detector of CO2 and acetone. The sensory response to acetone, or binary blends of acetone and CO2, reflects the observed behavioral output in both Ae. aegypti and An. coluzzii. We conclude that host recognition is contextual and dependent on a combination of ecologically relevant odorants at naturally occurring concentrations that are encoded, in this case, by differences in the temporal structure of the neuronal response. This information should be considered when designing synthetic blends for that optimally attract mosquitoes for monitoring and control.

False positives from impurities result in incorrect functional characterization of receptors in chemosensory studies.

The discovery of chemoreceptors and technological advances have greatly increased our understanding of chemosensory mechanisms. However, some of this rapid progress may have been severely compromised by insufficient attention given to the possible effects of impurities in the chemical standards used in identifying ligands for target receptors. Here, we show that even trace amounts of impurities in test stimuli can completely obscure true ligand-receptor relationships. Responses to impurities may go unrecognized because of two main factors. First, the sensitivity of receptors to ligands may be greater than that of the instruments used to check sample purity. Second, the concentrations of impurities actually reaching the chemoreceptor during experiments may be orders of magnitude higher than that of the putative stimulus, due to large differences in vapour pressure between the impurities and the putative stimulus. Errors caused by impurities are not limited to receptor-ligand studies, but can also affect related areas of chemosensory research, such as neural processing, downstream behaviours, and "in-silico" bioinformatics predictions of response profiles. The purity of standards is always implied but must be checked rigorously to prevent skewed or invalid results or conclusions, such as we exemplify here for Drosophila melanogaster and its olfactory receptor DmOr7a.

Novel odor-based strategies for integrated management of vectors of disease.

The proven ability of vector mosquitoes to adapt to various strategies developed to control them has enabled mosquito-borne diseases such as malaria, dengue, and lymphatic filariasis to remain entrenched as public health threats all over the world. Rather than continuing to seek a miracle cure for all mosquito vector problems among the ranks of single mode-of-action chemical pesticides, today's developers of vector control strategies are increasingly turning to more integrated, varied techniques, relying on pheromones and other semiochemicals to effect vector control through behavioral manipulation of the vector. Examples of this focus include attract-and-kill technologies utilizing floral odors and vertebrate host-associated scent cues to achieve control of adult mosquitoes, and selective oviposition attractants and larval phagostimulants to improve the efficacy of bacterial larvicides.

Recruiting on the Spot: A Biodegradable Formulation for Lacewings to Trigger Biological Control of Aphids.

Upon herbivory, plants release herbivore-induced plant volatiles (HIPVs), which induce chemical defenses in the plant as well as recruit natural enemies. However, whether synthetic HIPVs can be employed to enhance biological control in a cultivated crop in the field is yet to be explored. Here we show that a biodegradable formulation loaded with induced and food-signaling volatiles can selectively recruit the common green lacewing, Chrysoperla carnea, and reduce pest population under field conditions. In apple orchards, the new formulation attracted lacewing adults over a 4-week period, which correlated well with independent assessments of the longevity of the slow-release matrix measured through chemical analyses. In barley, lacewing eggs and larvae were significantly more abundant in treated plots, whereas a significant reduction of two aphid species was measured (98.9% and 93.6% of population reduction, for Sitobion avenae and Rhopalosiphum padi, respectively). Results show the potential for semiochemical-based targeted recruitment of lacewings to enhance biological control of aphids in a field setting. Further research should enhance selective recruitment by rewarding attracted natural enemies and by optimizing the application technique.

Translating olfactomes into attractants: shared volatiles provide attractive bridges for polyphagy in fruit flies.

Tephritid flies are serious fruit pests. Despite clear niche differences, many species show considerable overlap in fruit preferences, of which we here analysed the olfactory correlate. Using the volatiles of four unrelated fruit species, antennal responses were quantified to construct a fruit-odour response database for four tephritid species. Although responses were distinct with a significant niche-correlated bias, the analyses show that the probability of detection of a volatile strongly increased with its sharedness across fruits. This also held for the unrelated fruit fly Drosophila melanogaster (DoOR repository-based analyses). We conjectured that shared volatiles signify 'host' to the fly 'nose' and induce attraction. Indeed, blends of volatiles shared by fruit and detected by all four species were very attractive for tephritid species, more than fruits. Quantitative whole antennal recordings en lieu of, or complementing bottom-up molecular neurogenetic approaches, enables comparative olfactomics in non-model species, and facilitate interpretation of olfaction in evolutionary, ecological, and applied contexts.

Potential of locally sustainable food baits and traps against the Mediterranean fruit fly Ceratitis capitata in Bolivia.

BACKGROUND: Small-scale farmers often take a double hit from pests such as tephritid fruit-flies. The high price of products against fruit-flies, together with the higher risk of reinfestation from neighboring orchards, limits options for control. Therefore, management requires low-cost local products and concerted action. Peach production in central Bolivia is increasingly affected by invasive Ceratitis capitata. To provide locally sustainable techniques that could incentivize area-wide cooperation of growers, we tested efficiency and specificity of low-cost lures and traps compared with commercial lures and traps (Tephritrap). RESULTS: In the laboratory, the local fermented beverage 'chicha' and baker's yeast were equally or more attractive than commercial lures. Both chicha and baker's yeast trapped more flies in field (average FTD 10.31 and 9.49), whereas commercially hydrolyzed protein lure (4.71) or Torula yeast (6.82). However, many non-target species were caught (57.3% and 53.4%). Of the six PET bottles-based traps used, the T-trap caught a similar number of flies (average FTD 5.55), but fewer beneficial insects (0.16) compared to the Tephritrap (0.92). CONCLUSIONS: This study provides fruit growers with an economical and effective method to capture large numbers of C. capitata, suitable to be part of integrated pest management programs for fruit fly control. © 2018 Society of Chemical Industry.

Detection of Volatile Constituents from Food Lures by Tephritid Fruit Flies.

Tephritid fruit flies require protein for sexual and gonotrophic development. Food-based lures are therefore widely used in strategies to detect and control fruit flies in the Tephritidae family. However, these baits are attractive to a broad range of insect species. We therefore sought to identify volatiles detected by the fly antennae, with the goal to compose lures that more specifically target tephritids. Using gas chromatography-coupled electroantennographic detection (GC-EAD) we screened for antennal responses of four important tephritid species to volatile compounds from five commercially available protein-based baits. Antennal active compounds were reconstituted in synthetic blends for each species and used in behavioral assays. These species-based blends were attractive in olfactometer experiments, as was a blend composed of all antennally active compounds from all the four species we observed (tested only in Bactrocera dorsalis, Hendel). Pilot field tests indicate that the blends need to be further evaluated and optimized under field conditions.

Detection and perception of generic host volatiles by mosquitoes: responses to CO2 constrains host-seeking behaviour.

Natural selection has favoured specialization in anthropophilic mosquito host choice, yet in the absence of human hosts, females feed on a selected range of vertebrates. For host recognition, we hypothesize that mosquitoes primarily rely on generic host volatiles. Detection and perception of such compounds would provide the mosquito with a flexible, yet constrained, odour coding system that could delineate host preference. In this study, we show that the quintessential generic volatile for host-seeking, carbon dioxide, activates and attracts the malaria mosquito, Anopheles coluzzii, and the arbovirus vectors, Aedes aegypti and Culex quinquefasciatus, within boundaries set by the dynamic range and coding capacity of the CO2-sensitive olfactory receptor neurons. These boundaries are sufficiently broad to elicit behavioural responses to various hosts within their preferred host range. This study highlights the significance of the sensitivity of the carbon dioxide detection system and its regulation of host seeking and recognition.

Genetic mapping of male pheromone response in the European corn borer identifies candidate genes regulating neurogenesis.

The sexual pheromone communication system of moths is a model system for studies of the evolution of reproductive isolation. Females emit a blend of volatile components that males detect at a distance. Species differences in female pheromone composition and male response directly reinforce reproductive isolation in nature, because even slight variations in the species-specific pheromone blend are usually rejected by the male. The mechanisms by which a new pheromone signal-response system could evolve are enigmatic, because any deviation from the optimally attractive blend should be selected against. Here we investigate the genetic mechanisms enabling a switch in male response. We used a quantitative trait locus-mapping approach to identify the genetic basis of male response in the two pheromone races of the European corn borer, Ostrinia nubilalis Male response to a 99:1 vs. a 3:97 ratio of the E and Z isomers of the female pheromone is governed by a single, sex-linked locus. We found that the chromosomal region most tightly linked to this locus contains genes involved in neurogenesis but, in accordance with an earlier study, does not contain the odorant receptors expressed in the male antenna that detect the pheromone. This finding implies that differences in the development of neuronal pathways conveying information from the antenna, not differences in pheromone detection by the odorant receptors, are primarily responsible for the behavioral response differences among the males in this system. Comparison with other moth species reveals a previously unexplored mechanism by which male pheromone response can change in evolution.

Combining Attractants and Larvicides in Biodegradable Matrices for Sustainable Mosquito Vector Control.

BACKGROUND: There is a global need for cost-effective and environmentally friendly tools for control of mosquitoes and mosquito-borne diseases. One potential way to achieve this is to combine already available tools to gain synergistic effects to reduce vector mosquito populations. Another possible way to improve mosquito control is to extend the active period of a given control agent, enabling less frequent applications and consequently, more efficient and longer lasting vector population suppression. METHODOLOGY/PRINCIPAL FINDINGS: We investigated the potential of biodegradable wax emulsions to improve the performance of semiochemical attractants for gravid female culicine vectors of disease, as well as to achieve more effective control of their aquatic larval offspring. As an attractant for gravid females, we selected acetoxy hexadecanolide (AHD), the Culex oviposition pheromone. As toxicant for mosquito larvae, we chose the biological larvicides Bacillus thuringiensis israelensis (Bti) and Bacillus sphaericus (Bs). These attractant and larvicidal agents were incorporated, separately and in combination, into a biodegradable wax emulsion, a commercially available product called SPLAT (Specialized Pheromone & Lure Application Technology) and SPLATbac, which contains 8.33% Bti and 8.33% Bs. Wax emulsions were applied to water surfaces as buoyant pellets of 20 mg each. Dose-mortality analyses of Culex quinquefasciatus Say larvae demonstrated that a single 20 mg pellet of a 10-1 dilution of SPLATbac in a larval tray containing 1 L of water caused 100% mortality of neonate (1st instar) larvae for at least five weeks after application. Mortality of 3rd instar larvae remained equally high with SPLATbac dilutions down to 10-2 for over two weeks post application. Subsequently, AHD was added to SPLAT (emulsion only, without Bs or Bti) to attract gravid females (SPLATahd), or together with biological larvicides to attract ovipositing females and kill emerging larvae (SPLATbacAHD, 10-1 dilution) in both laboratory and semi-field settings. The formulations containing AHD, irrespective of presence of larvicides, were strongly preferred as an oviposition substrate by gravid female mosquitoes over controls for more than two weeks post application. Experiments conducted under semi-field settings (large screened greenhouse, emulating field conditions) confirmed the results obtained in the laboratory. The combination of attractant and larvicidal agents in a single formulation resulted in a substantial increase in larval mosquito mortality when compared to formulations containing the larvicide agents alone. CONCLUSIONS/SIGNIFICANCE: Collectively, our data demonstrate the potential for the effective use of wax emulsions as slow release matrices for mosquito attractants and control agents. The results indicate that the combination of an oviposition attractant with larvicides could synergize the control of mosquito disease vectors, specifically Cx. quinquefasciatus, a nuisance pest and circumtropical vector of lymphatic filariasis and encephalitis.

Do Fruit Ripening Volatiles Enable Resource Specialism in Polyphagous Fruit Flies?

Frugivorous tephritid fruit flies have lineages with high levels of host generalism. These insects use olfaction to locate fruits, but how they are able to recognize the odors of so many different host species is poorly understood. We used a series of behavioral experiments to investigate the role of fruit ripening volatiles as host cues in the Queensland fruit fly, Bactrocera tryoni (Froggatt), a polyphagous pest in Australia. Odors of mature guava (Psidium guajava) attracted female and male flies more strongly than three other ripening stages and guava pulp. We analyzed volatiles from guava odor and selected eleven compounds, all of which elicited an electrophysiological response in the antenna of female flies. Three of these, ethyl acetate, ethyl butyrate, and ethyl propionate, were released at the highest rates from the most attractive ripening stage. In behavioral trials, these three esters were not attractive individually, whereas a combination was necessary and sufficient in attracting female flies. The three-component blend was as attractive as the entire 11-component blend, which without these key volatiles was not attractive. Moreover, injecting low ranking hosts (squash and cucumber) with the three volatiles increased attraction in ovipositing female flies. These fruit flies are classed as generalists, but like many polyphagous insects they could be regarded as resource specialists, preferring specific plant reproductive stages with predictable odor cues. Exploring olfaction from this perspective could improve our understanding of host choice in polyphagous insects, and the selection of volatiles to be used as attractants in insect pest management.