Andrew J. Myrick

Interspecific Pheromone Plume Interference Among Sympatric Heliothine Moths: A Wind Tunnel Test Using Live, Calling Females

Three species of North American heliothine moths were used to determine the level at which interspecific female interference of male attraction to conspecific females occurs. We used live calling females of Heliothis virescens, H. subflexa, and Helicoverpa zea, as lures for conspecific males in a wind tunnel, and then placed heterospecific females on either side of the original species such that the plumes of the three females overlapped downwind. In nearly all combinations, in the presence of heterospecific females, fewer males flew upwind and contacted or courted the source than when only conspecific females were used in the same spatial arrangement. Males did not initiate upwind flight to solely heterospecific female arrangements. Our results show that the naturally emitted pheromone plumes from heterospecific females of these three species can interfere with the ability of females to attract conspecific males when multiple females are in close proximity. However, the fact that some males still located their calling, conspecific females attests to the ability of these male moths to discriminate point source odors by processing the conflicting information from interleaved strands of attractive and antagonistic odor filaments on a split-second basis.

Chopper-stabilized gas chromatography-electroantennography: Part I. Background, signal processing and example

A new method that can improve gas-chromatography-electroantennographic detection (GC-EAD) by orders of magnitude through a technique known as chopper stabilization combined with matched filtering in colored noise is presented. The EAD is a physiological recording from the antenna of an insect which can be used to find compounds in the GC effluent that the antenna is able to detect, having important applications for pest control and understanding of chemical communication in nature. The new method is demonstrated with whole-animal male Helicoverpa zea antennal preparations for detection of major pheromone component (cis-11-hexadecenal) and compared to results obtained using traditional EAD recording techniques. Results indicate that chopper stabilization under these circumstances can increase odorant detection performance by a factor of approximately 10(4) over traditional methods. The time course of the response of the antenna is also better resolved under chopped conditions. Although the degree of improvement is expected to vary with insect species, odor, and insect preparation, under most circumstances a more sensitive and robust GC-EAD instrument will result from the application of this technique.

Locating a compact odor source using a four-channel insect electroantennogram sensor.

Here we demonstrate the feasibility of using an array of live insects to detect concentrated packets of odor and infer the location of an odor source (∼15 m away) using a backward Lagrangian dispersion model based on the Langevin equation. Bayesian inference allows uncertainty to be quantified, which is useful for robotic planning. The electroantennogram (EAG) is the biopotential developed between the tissue at the tip of an insect antenna and its base, which is due to the massed response of the olfactory receptor neurons to an odor stimulus. The EAG signal can carry tens of bits per second of information with a rise time as short as 12 ms (K A Justice 2005 J. Neurophiol. 93 2233-9). Here, instrumentation including a GPS with a digital compass and an ultrasonic 2D anemometer has been integrated with an EAG odor detection scheme, allowing the location of an odor source to be estimated by collecting data at several downwind locations. Bayesian inference in conjunction with a Lagrangian dispersion model, taking into account detection errors, has been implemented resulting in an estimate of the odor source location within 0.2 m of the actual location.

Chopper-Modulated Locked in Amplified Gas Chromatography—Electroantennography Part II: Signal Processing and Performance Comparisons

A new method that improves gas-chromatography-electroantennographic detection through lock-in amplification is demonstrated. Here, measurements of antennal responses to major pheromone component,

Isolation of a Female-Emitted Sex Pheromone Component of the Fungus Gnat, Lycoriella ingenua, Attractive to Males

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Olfactory Sensory Neurons of the Asian Longhorned Beetle, Anoplophora glabripennis, Specifically Responsive to its two Aggregation-Sex Pheromone Components

-16:Ald, are performed under more optimal conditions, using saline electrical connections to excise antennae from male H. subflexa. Matched filtering in colored noise is applied to traditional enhanced graphics adapter recordings, allowing the signal to noise ratio of to be increased by about 6.1 dB (uncertainty is dependent on dosage). A dose-response model, including model parameter uncertainties is then used to estimate and compare performance in terms of naïve error rates involved in the detection of insect responses to GC peaks. Results indicate that relative performance is dosage dependent. Without a visible flame ionization detector (FID) reference peak for determining elution time, the model predicts the detection limit (placed at 5.0% expected naïve error rate) to be approximately 12 times lower using chopper modulation than when using traditional methods. At the highest traditional dosage tested, 10 pg, the equivalent chopper modulated dosage is estimated to be about 92 times lower. When a reference FID peak is clearly visible, the predicted detection limit (at 5.0% expected naïve error rate) is expected to be approximately 7 times lower using chopper modulation than when using traditional methods. At the traditional dosage of 10 pg, the predicted equivalent chopped dosage is estimated to be about 66 times lower.

Chopper-modulated gas chromatography electroantennography enabled using high-temperature MEMS flow control device

Lycoriella ingenua Dufour (Diptera: Sciaridae) is acknowledged as the major pest species of the white button mushroom, Agaricus bisporus, throughout the world. Components of the female-produced sex pheromone of this species were identified previously as C15-C18n-alkanes, with the major component n-heptadecane, and shown to be attractive to L. mali. However, a subsequent report could not repeat this work. We reinvestigated the sex pheromone of this species by confirming that virgin females were attractive to males in a Y-tube bioassay and by collection of extracts from virgin females. Extracts were analyzed by gas chromatography coupled to electroantennographic detection, and by the less widely-used technique of gas chromatography coupled to a behavioral bioassay to detect compounds causing wing-fanning and copulatory abdomen curling in males. A single, behaviorally-active pheromone component was isolated and characterized by gas chromatography coupled to mass spectrometry. This component was definitively not n-heptadecane or any of the other C15-C19n-alkanes reported previously, but is proposed to be a sesquiterpene alcohol having analytical characteristics that closely matched those of reference germacradienols.

Differences in spectral selectivity between stages of visually guided mating approaches in a buprestid beetle

We performed single-sensillum recordings from male and female antennae of the Asian longhorned beetle, Anoplophora glabripennis, that included as stimuli the two components of this species’ aggregation-sex pheromone in addition to various general odorants. We compared the aggregation-sex-pheromone-component responses of olfactory sensory neurons (OSNs) to those of OSNs that responded to a variety of plant-related odorants. In the smooth-tipped, tapered, trichoid sensilla on the most distal antennal flagellomeres nos. 10 or 11 of both males and females, we found OSNs with high-amplitude action potentials that were tuned to the aldehyde and alcohol pheromone components and that did not respond to various plant-related volatiles. Because this OSN type responded to both the alcohol and aldehyde components it cannot be considered to be specifically tuned to either component. These large-spiking OSNs were co-compartmentalized in these sensilla with a second, smaller-spiking OSN responding to plant-related volatiles such as geraniol, citronellal, limonene, 1-octanol, nerol and citral. The large-spiking OSNs thus appear to be a type that will be involved in aggregation-sex pheromone pathways targeting a specific glomerulus in the antennal lobe and in generating pheromone-related behavioral responses in A. glabripennis. In other sensilla located in these distal antennal flagellomeres as well as those located more proximally, i.e., mid-length along the antenna on flagellomere nos. 4–7, we found OSNs in blunt-tipped basiconic sensilla that were responsive to other plant-related volatiles, especially the terpenoids, (E,E)-alpha farnesene, (E)-β-farnesene, β-caryophyllene, and eugenol. Some of these terpenoids have been implicated in improving attraction to pheromone-baited traps. Some of these same OSNs responded additionally to either of the two sex pheromone components, but because these OSNs also responded to some of the above plant volatiles as shown by cross-adaptation experiments, these OSNs will not be the types that convey sex-pheromone-specific information to the antennal lobe.

Optimizing the Point-Source Emission Rates and Geometries of Pheromone Mating Disruption Mega-Dispensers

We report the design, fabrication and characterization of a microelectromechanical systems (MEMS) flow control device for gas chromatography (GC) with the capability of sustaining high-temperature environments. We further demonstrate the use of this new device in a novel MEMS chopper-modulated gas chromatography-electroantennography (MEMS-GC-EAG) system to identify specific volatile organic compounds (VOCs) at extremely low concentrations. The device integrates four pneumatically actuated microvalves constructed via thermocompression bonding of the polyimide membrane between two glass substrates with microstructures. The overall size of the device is 32 mm×32 mm, and it is packaged in a 50 mm×50 mm aluminum housing that provides access to the fluidic connections and allows thermal control. The characterization reveals that each microvalve in the flow control chip provides an ON to OFF ratio as high as 1000:1. The device can operate reliably for more than 1 million switching cycles at a working temperature of 300 °C. Using the MEMS-GC-EAG system, we demonstrate the successful detection of cis-11-hexadecenal with a concentration as low as 1 pg at a demodulation frequency of 2 Hz by using an antenna harvested from the male Helicoverpa Virescens moth. In addition, 1 μg of a green leafy volatile (GLV) is barely detected using the conventional GC-EAG, while MEMS-GC-EAG can readily detect the same amount of GLV, with an improvement in the signal-to-noise ratio (SNR) of ~22 times. We expect that the flow control device presented in this report will allow researchers to explore new applications and make new discoveries in entomology and other fields that require high-temperature flow control at the microscale.

Working Range of Stimulus Flux Transduction Determines Dendrite Size and Relative Number of Pheromone Component Receptor Neurons in Moths

ABSTRACT Spectral mating preferences were examined in male Agrilus angustulus (Buprestidae: Coleoptera), a member of a taxon known for its high species diversity and striking metallic coloration. The spectral emission profile of a typical A. angustulus female displays low chroma, broadly overlapping that of the green oak leaves they feed and rest upon, while also including longer wavelengths. To pinpoint behaviorally significant spectral regions for A. angustulus males during mate selection, we observed their field approaches to females of five Agrilus planipennis color morphs that have greater chroma than the normal conspecific female targets. Agrilus angustulus males would initially fly equally frequently toward any of the three longest wavelength morphs (green, copper and red) whose spectral emission profiles all overlap that of typical A. angustulus females. However, they usually only completed approaches toward the two longest wavelength morphs, but not the green morphs. Thus, spectral preference influenced mate selection by A. angustulus males, and their discrimination of suitable targets became greater as these targets were approached. This increasing spectral discrimination when approaching targets may have evolved to allow female emissions to remain somewhat cryptic, while also being visible to conspecifics as distinct from the background vegetation and heterospecific competitors. Summary: The stereotypical visually guided mating approaches of male buprestid beetles include two stages that use different spectral cues.