Michael J. Domingue

Reduction in host-finding behaviour in fungus-infected mosquitoes is correlated with reduction in olfactory receptor neuron responsiveness.

BackgroundChemical insecticides against mosquitoes are a major component of malaria control worldwide. Fungal entomopathogens formulated as biopesticides and applied as insecticide residual sprays could augment current control strategies and mitigate the evolution of resistance to chemical-based insecticides.MethodsAnopheles stephensi mosquitoes were exposed to Beauveria bassiana or Metarhizium acridum fungal spores and sub-lethal effects of exposure to fungal infection were studied, especially the potential for reductions in feeding and host location behaviours related to olfaction. Electrophysiological techniques, such as electroantennogram, electropalpogram and single sensillum recording techniques were then employed to investigate how fungal exposure affected the olfactory responses in mosquitoes.ResultsExposure to B. bassiana caused significant mortality and reduced the propensity of mosquitoes to respond and fly to a feeding stimulus. Exposure to M. acridum spores induced a similar decline in feeding propensity, albeit more slowly than B. bassiana exposure. Reduced host-seeking responses following fungal exposure corresponded to reduced olfactory neuron responsiveness in both antennal electroantennogram and maxillary palp electropalpogram recordings. Single cell recordings from neurons on the palps confirmed that fungal-exposed behavioural non-responders exhibited significantly impaired responsiveness of neurons tuned specifically to 1-octen-3-ol and to a lesser degree, to CO2.ConclusionsFungal infection reduces the responsiveness of mosquitoes to host odour cues, both behaviourally and neuronally. These pre-lethal effects are likely to synergize with fungal-induced mortality to further reduce the capacity of mosquito populations exposed to fungal biopesticides to transmit malaria.

Field observations of visual attraction of three European oak buprestid beetles toward conspecific and heterospecific models

Agrilus biguttatus Fabricius, Agrilus sulcicollis Lacordaire, and Agrilus angustulus Illiger (Coleoptera: Buprestidae) are three beetle species associated with oak trees [Quercus spp. (Fagaceae)] in Europe. In Hungary, all three species were observed in the foliage near freshly cut oak log piles. Agrilus biguttatus was active later in the afternoon, whereas the other species were observed earlier in the day. Dead female models of these three native Agrilus species, as well as the native species Agrilus cyanescens Ratzeburg and the non‐native Agrilus planipennis Fairmaire, were pinned onto adjacent leaves in direct sunlight to observe the visual mating approaches of the local male populations. Agrilus biguttatus and A. sulcicollis males flew toward and landed directly on the models from a distance of 1 m. Agrilus angustulus flew toward the models from a similar distance, but rather than landing directly on a model would alight on the leaf, 1–2 cm away, before walking closer to the model while antennating it. For all three species, there was substantial cross‐attraction to models of other species. Both A. biguttatus and A. sulcicollis males chose A. angustulus models less often than their respective conspecific models. Likewise, A. angustulus males approached A. sulcicollis models less often than their normal conspecific models. Agrilus biguttatus males attempted to copulate with both A. biguttatus and A. planipennis models, afterward remaining with them for several minutes. Agrilus biguttatus males spent more time on A. planipennis models than on conspecific models. Thus, there is substantial cross‐species attraction in visually mediated mating approaches and copulation behavior. These findings suggest a common behavioral template for visual mate‐finding among buprestids and a large degree of close‐range mating compatibility between A. biguttatus and A. planipennis.

Genetic Control of the Enantiomeric Composition of Ipsdienol in the Pine Engraver, Ips pini

The genetic nature of pheromone variation within species has rarely been studied, and never for male-produced long-range pheromones. Males from western North American populations of Ips pini produce predominantly (R)-(−)-ipsdienol, whereas those from eastern North American populations produce higher proportions of (S)-(+)-ipsdienol. From a population in the hybrid zone, we divergently selected lines for the opposing pheromonal types and then created F1, F2, and backcross lines. We formed additional F1, F2, and backcross lines, first by using populations with low (+)-ipsdienol enantiomeric ratios near to and distant from the hybrid zone, and then by using populations with high (+)-ipsdienol enantiomeric ratios near to and distant from the hybrid zone. Three types of analysis were employed: (1) line means analysis; (2) Mendelian analysis of assigned high and low (+)-ipsdienol enantiomeric ratio phenotypes when applicable; and (3) Castle–Wright estimation of the number of effective factors. Dominance at one autosomal locus explains much of the variation in ipsdienol blend between the divergently selected lines. Thus, as in all previously studied female long-range pheromone systems, a major genetic element is implicated. The populations with low (+)-ipsdienol enantiomeric ratios near and distant to the hybrid zone differ negligibly for this trait. We confirmed previous studies showing slightly higher ratios of (+)-ipsdienol at the hybrid zone than in a distant eastern population and reveal a genetic basis for this difference.

Visual and chemical cues affecting the detection rate of the emerald ash borer in sticky traps

Using sticky traps, we compared the efficacy of chemical and visual lures, both alone and in combination, for improving the detection of populations of the emerald ash borer (EAB), Agrilus planipennis. Ash leaflets to which EAB visual decoys were pinned and coated with sticky material were able to trap EAB with as high a rate of detection as large sticky visually unbaited ‘prism traps’ currently used in wide‐scale EAB surveillance programs in North America, in a high‐density area. Both the sticky leaf traps and prism traps captured more EAB when a point source of plant odours, either manuka or phoebe oil, was deployed with the trap. For the sticky leaf traps, the shape of the EAB visual decoy lure was found to be important in optimizing the detection rate. Either an entire dead beetle or else two elytra placed side by side to mimic a resting beetle resulted in optimal trap performance. When two elytra were placed end to end or else other body parts were deployed, the traps lost their efficacy. Small green plastic surfaces to which EAB visual decoys were pinned were found to be fairly good substitutes for live ash leaflets, but the rate of beetle detection was reduced significantly from that of the ash leaflet plus EAB decoy. Throughout all experiments, a clear male bias occurred in sticky leaf traps when EAB visual decoys were placed on the traps. The implications of these findings for developing new trapping designs for EAB and other forest buprestids are discussed.

Detecting emerald ash borers (Agrilus planipennis) using branch traps baited with 3D-printed beetle decoys

Small visual-decoy-baited traps for the emerald ash borer (EAB), Agrilus planipennis Fairmaire (Coleoptera: Buprestidae), have been researched as an alternative to current technologies, but have relied on real beetles rather than synthetic materials. We hypothesized that visual decoys created by three-dimensional (3D) printing can provide such a substitute. Branch traps displaying decoys consisting of real EAB females or 3D-printed decoys were compared to controls without decoys. Traps of the three varieties were placed on neighboring branches along with one (Z)-3-hexen-1-ol lure per tree and checked daily. Both real and 3D-printed decoys similarly increased EAB trap captures compared to controls. The numbers of both sexes were higher on the decoy-baited traps, but the increase in male captures was more pronounced. Males were also ensnared closer to the decoys than females. Daily trap–capture patterns showed sparse activity of EAB adults before June 18, 2013 followed by a peak in captures of both males and females until June 28, 2013. Beginning at approximately July 1, 2013, there was a second peak of EAB captures, which consisted almost entirely of males caught on the decoy-baited traps. The native ash borer Agrilus subcinctus was found earlier in the season and was also significantly attracted to both the real EABs and the 3D-printed decoys compared with control traps. Four purple prism traps were also deployed concurrently and captures tallied on three different days within the season. The results demonstrate efficacy of a small, inexpensive, and fully synthetic decoy-based branch trap system for EAB.

Trapping of European buprestid beetles in oak forests using visual and olfactory cues

Trapping approaches developed for the emerald ash borer (EAB), Agrilus planipennis Fairmaire (Coleoptera: Buprestidae), were adapted for trapping several European oak buprestid species. These approaches included the use of natural leaf surfaces as well as green and purple plastic in sticky trap designs. Plastic surfaces were incorporated into novel ‘branch‐trap’ designs that each presented two 5 × 9‐cm2 rectangular surfaces on a cardboard structure wrapped around the leaves of a branch. We used visual adult Agrilus decoys in an attempt to evoke male mating approaches toward the traps. Our first experiment compared the attractiveness of visual characteristics of the surfaces of branch‐traps. The second looked at the effect on trap captures of adding semiochemical lures, including manuka oil, (Z)‐3‐hexen‐1‐ol, and (Z)‐9‐tricosene. In total, 1 962 buprestid specimens including 14 species from the genus Agrilus were caught on 178 traps in a 22‐day time‐span. Overall, the green plastic‐covered branch‐traps significantly out‐performed the other trap designs. We further found that the presence of an EAB visual decoy placed on the trap surface often increased captures on these green traps, but this effect was stronger for certain Agrilus species than for others. The visual decoy was particularly important for the most serious pest detected, Agrilus biguttatus Fabricius, which was captured 13 times on traps with decoys, but only once without a decoy. There were some small but significant effects of odor treatment on the capture of buprestids of two common species, Agrilus angustulus Illiger and Agrilus sulcicollis Lacordaire. There were also 141 Elateridae specimens on these traps, which were not influenced by trap type or decoys. The results suggest that small branch‐traps of this nature can provide a useful new tool for monitoring of buprestids, which have the potential to be further optimized with respect to visual and olfactory cues.

Olfactory neuron responsiveness and pheromone blend preference in hybrids between Ostrinia furnacalis and Ostrinia nubilalis (Lepidoptera: Crambidae)

The olfactory receptor neuron (ORN) and behavioral responses of hybrids between the Asian corn borer (ACB), Ostrinia furnacalis, and the E-strain European corn borer (ECB(E)), Ostrinia nubilalis were examined and compared to the parental populations. In hybrids and both parents, the large-spike-size ORN was capable of responding to all four pheromone components of ACB and ECB, despite differences in which compounds elicited the greatest spike frequency in each population. There was a small-spiking ORN more narrowly tuned to the minor pheromone components in both ACB and ECB(E). In hybrids the homologous small-spiking ORN was tuned primarily to the ECB(E) minor pheromone component, with some responsiveness to the ACB minor component. Both species and all the hybrids had an intermediate spike-size ORN tuned primarily to their common behavioral antagonist. Dominance of responsiveness to the ECB(E) versus the ACB minor pheromone component on the small-spiking ORN may explain the greater tendency of hybrids to fly upwind to the ECB(E) pheromone blend than the ACB blend. This finding points toward a distinct evolutionary role for this ORN in allowing a pheromone shift.

Support for (Z)-11-Hexadecanal as a Pheromone Antagonist in Ostrinia nubilalis: Flight Tunnel and Single Sensillum Studies with a New York Population

The flight-tunnel response of male Z-strain European corn borer moths (ECB), Ostrinia nubilalis, from a population in New York State (USA), was significantly antagonized by addition of 1% (Z)-11-hexadecanal (Z11-16:Ald) to their sex pheromone (a 97:3 mix of (Z)- and (E)-11-tetradecenyl acetate [Z/E11-14:OAc]). The level of antagonism was equivalent to that observed for the previously identified ECB antagonist, (Z)-9-tetradecenyl acetate (Z9-14:OAc), and supports a recent report showing that Z11-16:Ald, a minor pheromone component of the Noctuid moth, Sesamia nonagrioides, caused antagonism of ECB pheromone communication in sympatric populations in the Iberian Peninsula. Single-sensillum recordings from ECB antennae, which included cross-adaptation experiments, showed that the same olfactory receptor neuron processing Z9-14:OAc inputs was responsible for detecting Z11-16:Ald, and that this neuron was not responsive to two other aldehydes, (Z)-9-tetradecanal (Z9-14:Ald) and (Z)-9-hexadecanal (Z9-16:Ald), found in other moth sex pheromones. Our results show that the antagonism is not confined to one geographic region, is specific for Z11-16:Ald, and that antagonist pathways might have the potential for processing a number of structurally similar compounds.

Bioreplicated visual features of nanofabricated buprestid beetle decoys evoke stereotypical male mating flights

Significance Advances in material processes for bioreplication have led to the use of bioinspired designs in a wide variety of practical applications, often at a scale involving nanofabrication. Such techniques also provide the opportunity to examine the functional significance of nanostructured organismal properties within biological systems. This paper describes the replication of fine-scale elements of the exoskeleton of buprestid beetles that produce a visually interpreted mating signal. A nanofabricated replica of the beetle was exploited to cause wild male beetles to land on synthetic decoy beetles. The development of such bioreplicated decoys opens new avenues for the study of the nature of insect visual responses, as well as applications for detection technologies that target pest organisms. Recent advances in nanoscale bioreplication processes present the potential for novel basic and applied research into organismal behavioral processes. Insect behavior potentially could be affected by physical features existing at the nanoscale level. We used nano-bioreplicated visual decoys of female emerald ash borer beetles (Agrilus planipennis) to evoke stereotypical mate-finding behavior, whereby males fly to and alight on the decoys as they would on real females. Using an industrially scalable nanomolding process, we replicated and evaluated the importance of two features of the outer cuticular surface of the beetle’s wings: structural interference coloration of the elytra by multilayering of the epicuticle and fine-scale surface features consisting of spicules and spines that scatter light into intense strands. Two types of decoys that lacked one or both of these elements were fabricated, one type nano-bioreplicated and the other 3D-printed with no bioreplicated surface nanostructural elements. Both types were colored with green paint. The light-scattering properties of the nano-bioreplicated surfaces were verified by shining a white laser on the decoys in a dark room and projecting the scattering pattern onto a white surface. Regardless of the coloration mechanism, the nano-bioreplicated decoys evoked the complete attraction and landing sequence of Agrilus males. In contrast, males made brief flying approaches toward the decoys without nanostructured features, but diverted away before alighting on them. The nano-bioreplicated decoys were also electroconductive, a feature used on traps such that beetles alighting onto them were stunned, killed, and collected.

The genetic architecture of pheromone production between populations distant from the hybrid zone of the pine engraver, Ips pini

Summary.Across North America, populations of the pine engraver, Ips pini, differ in their expressed ratios of the two enantiomers of ipsdienol, the main component of its aggregation pheromone. We confirm previous studies, showing that the percentage of (+)-ipsdienol ranged from approximately 40 to 70% for New York (NY) males and less then 5% for California (CA) males. We performed line crosses including the F1, F2, and backcross generations between these populations. These line crosses showed that most F1 hybrids produced an intermediate enantiomeric blend of ipsdienol that was closer to the CA blend, with a frequency distribution peak near 15% (+)-ipsdienol. There was also strong segregation to either parental type in the F2 and backcross generations, but not in a pattern that could be clearly described by a single autosomal locus. There was also an X-linked effect that caused some individuals in the F1 to have phenotypes more characteristic of NY populations. Generation means analysis confirms this X-linked effect, and also suggests a complicated autosomal dominance by dominance epistatic interaction. Despite the appearance of segregation, these results suggest a more complicated system of pheromone blend control than the single major gene previously found between divergent lines for high and low (+)-ipsdienol blends from the hybrid zone in British Columbia.