Live calcium imaging of Aedes aegypti neuronal tissues reveals differential importance of chemosensory systems for life-history-specific foraging strategies.

BackgroundThe mosquito Aedes aegypti has a wide variety of sensory pathways that have supported its success as a species as well as a highly competent vector of numerous debilitating infectious pathogens. Investigations into mosquito sensory systems and their effects on behavior are valuable resources for the advancement of mosquito control strategies. Numerous studies have elucidated key aspects of mosquito sensory systems, however there remains critical gaps within the field. In particular, compared to that of the adult form, there has been a lack of studies directed towards the immature life stages. Additionally, although numerous studies have pinpointed specific sensory receptors as well as responding motor outputs, there has been a lack of studies able to monitor both concurrently.ResultsTo begin filling aforementioned gaps, here we engineered Ae. aegypti to ubiquitously express a genetically encoded calcium indicator, GCaMP6s. Using this strain, combined with advanced microscopy, we simultaneously measured live stimulus-evoked calcium responses in both neuronal and muscle cells with a wide spatial range and resolution.ConclusionsBy coupling in vivo live calcium imaging with behavioral assays we were able to gain functional insights into how stimulus-evoked neural and muscle activities are represented, modulated, and transformed in mosquito larvae enabling us to elucidate mosquito sensorimotor properties important for life-history-specific foraging strategies

Classical Engineering Systems Provide Behavioral Analog for Ephemeral Insect and Plant Biomechanics

In this dissertation we consider ephemeral behaviors of two small-scale living systems, mosquitoes and citrus fruit reservoirs. While these two systems share few obvious commonalities, they both express life events that are complex and conclude within approximately 50 milliseconds. We utilize high-speed videography, between 1,000-16,000 fps, to detail how complex behavior can be modeled as classical engineering systems. Beginning with the larger organism we assessed the landing and takeoff behavior of Aedes aegypti mosquitoes to ascertain the secrets of their covert interaction with humans. At takeoff, mosquitoes decrease pushing contact time with substrates of low friction through a modified takeoff behavior of striking the substrate with a hind-leg prior to a classic push phase. We propose a 2D analog where the striking leg acts as a rotating cantilever about a fixed end that generates upward momentum with a small penalty in body rotation. Landing mosquitoes are filmed in 2D and modeled as a mass-spring-damper system whose natural frequency, damping coefficient, ratio, and spring constant are determined experimentally and validated through a nonlinear least square solver fitting of the free vibration ODE\u27s general solution. Results indicate mosquitoes behave as an underdamped system to scrub their incoming momentum through extending impact duration, effectively reducing temporal impact force. Shrinking in scale we proceed to characterize citrus reservoir rupture as a passive system capable of microjetting oil through expanding orifices at accelerations greater than 5000 gravities. Citrus reservoirs are modeled as ellipsoidal pressure vessels capped by a thin membrane of contrasting stiffness to the surrounding ductile compressible albedo

Summer Research Fellowship Project Descriptions 2019

A summary of research done by Smith College’s 2019 Summer Research Fellowship (SURF) Program participants. Ever since its 1967 start, SURF has been a cornerstone of Smith’s science education. Supervised by faculty mentor-advisors drawn from the Clark Science Center and connected to its eighteen science, mathematics, and engineering departments and programs and associated centers and units. At summer’s end, SURF participants were asked to summarize their research experiences for this publication.https://scholarworks.smith.edu/clark_womeninscience/1008/thumbnail.jp

Multi-Modal Effects of the Repellent Deet Across Protostomia

DEET (N, N-diethyl-m-toluamide) is the most broadly effective and widely used personal repellent available, yet we do not understand what makes it so effective. Even in well-studied species like Drosophila melanogaster flies and Aedes aegypti mosquitoes, many mysteries remain as to how DEET can affect behavior in these species. For example, Ae. aegypti mosquitoes are attracted to human arms. When an arm is covered in DEET, wild-type mosquitoes are not attracted to the arm, while mutant mosquitoes that lack the odorant receptor co-receptor (orco), approach the arm, but rarely bite. We investigated this orco-independent DEET repellency in Ae. aegypti and found that these mosquitoes can sense DEET with their tarsi as well as their proboscis. The tarsi are required for mosquitoes to be repelled after contact with a DEET-treated arm. The proboscis is required for the rejection of DEET-laced liquid food. These results suggest that DEET acts on multiple sensory modalities to repel insects. Both this work and most prior literature has focused on studying how DEET affects Arthropods, yet one of the major open questions in the field is how DEET can be effective across so many different species. To identify genes and neurons required for DEET-sensitivity outside of Arthropoda, we turned to the nematode C. elegans. Here, we demonstrated that DEET affects chemotaxis to some odors but not others. We used this behavior as the basis for a forward genetic screen, and identified two genes as candidates required for complete DEET-sensitivity. We identified a natural isolate of C. elegans that was also resistant to DEET, and found that it contains a large deletion in one of the hits from our screen, the G protein-coupled receptor str-217. This gene is required for DEET-sensitivity in both wild-type and wild isolate strains. str-217 is expressed in a single pair of chemosensory neurons called ADL, which are required for complete DEET sensitivity, and respond to DEET as assayed by calcium imaging. Although we identified additional sensory neurons that respond to DEET, their behavioral contributions are unknown. Both ADL and str-217 are required for a specific, DEET-induced behavior during chemotaxis and exploration: an increase in average pause duration. Through optogenetic stimulation of ADL, we demonstrated that ADL activity alone is sufficient to increase average pause duration. Taken together, these experiments provide insights into the genetic and neural mechanisms underlying DEET-sensitivity in C. elegans, and allow for comparisons across Protostomes. We also establish C. elegans as a model non-Arthropod species for further investigation into the effects of DEET

The Neuropeptide Regulation of Host-Seeking Behavior in Aedes Aegypti Mosquitoes

Aedes aegypti mosquitoes are the principal vectors for several human diseases including Dengue Fever, which causes ~400 million cases and ~24,000 deaths per year (Bhatt et al., 2013; WHO, 2002). Novel strategies to combat mosquito-borne diseases are needed for A. aegypti and other mosquitoes such as the malaria vector Anopheles gambiae. Our goal was to discover new ways to interfere with the ability of a mosquito to locate a human host for a blood meal. Currently, the mechanistic basis of host-seeking and its regulation remain incompletely understood. Although it is known that mosquitoes require human odor cues to locate a human host, the critical odor components and associated olfactory receptors have not been identified (Klowden, 1995; Takken and Knols, 1999). Previous work showed that mosquito host-seeking behavior is inhibited by a hemolymph-borne humoral factor for three days following a blood meal. Subsequent studies identified Head Peptide-I as a candidate neuropeptide modulating this suppression in host-seeking behavior. This conclusion was strengthened by the observation that Head Peptide-I injection into non-blood-fed females triggered the inhibition of host-seeking. The mechanism by which this important peptide alters mosquito behavior and the receptor through which it signals are unknown (Brown et al., 1994). We used a cell-based calcium-imaging screen to identify the G-protein coupled receptor NPY-Like Receptor-1 (NPYLR1) as a candidate Head Peptide-I receptor. We found that multiple NPYLR1 agonists, including the feeding-related Short-Neuropeptide-3 (sNPF3), are capable of inhibiting host-seeking behavior when injected into non-blood-fed females. To investigate whether NPYLR1 is required for Head Peptide-I inhibition, we pioneered targeted mutagenesis with zinc-finger nucleases to create multiple NPYLR1 null-mutant mosquito lines. We predicted that these mutants would no longer show inhibition of host-seeking behavior after a blood meal. While we can say with certainty that NPYLR1 is a receptor for Head Peptide-I, we found no behavioral effects for NPYLR1 mutants in locomotion, egg-laying, sugar feeding, blood feeding, or host-seeking behavior. Our results suggest that NPYLR1 is not required in vivo for Head Peptide-I action and that a redundant signaling mechanism for behavioral inhibition exists. Future work will determine the necessity of Head Peptide-I during host-seeking inhibition and attempt to identify additional Head Peptide-I and sNPF receptors. This research will clarify the mechanism of Head Peptide-I inhibition and could form the basis for novel strategies to control mosquito host-seeking behavior

Distinct Olfactory Signaling Mechanisms in the Malaria Vector Mosquito Anopheles gambiae

A combination of gene silencing and behavioral studies in the malaria vector mosquito Anopheles gambiae sheds light on the olfactory basis of DEET repulsion as well as reveals the role of another family of chemosensory receptors that facilitate olfaction in An. gambiae

Lincoln University Cooperative Extension and Research Annual Report 2010

This is LUCER-MC Report #03-11 Published by Lincoln University Cooperative Extension and Research (LUCER) Media Center; 900 Chestnut Street, 301 Allen Hall; Jefferson City, MO 65101.https://bluetigercommons.lincolnu.edu/lucer_reports/1000/thumbnail.jp

Abstracts of Papers, 89th Annual Meeting of the Virginia Academy of Science, May 25-27, 2011, University of Richmond, Richmond VA

Full abstracts of the 89th Annual Meeting of the Virginia Academy of Science, May 25-27, 2011, University of Richmond, Richmond V

ZOOS AS EXPERIMENT ENVIRONMENTS: BIOLOGY OF LARVAL AND ADULT MOSQUITOES (DIPTERA: CULICIDAE)

Zoos are a unique environment where humans and animals are in close daily contact, potential mosquito habitats exist, exotic plants and animals are introduced regularly, and wild animals roam. Studies of mosquito behaviors in zoos will lead to a better understanding, both within and outside zoos, of disease transmission routes and mosquito biology. To investigate whether the unique assemblage of habitats in zoos affects mosquito behavior, I sampled larvae and adults in the Greenville Zoo and the Riverbanks Zoo, South Carolina, USA, from March 2008 to January 2011. The objectives of my study were to investigate mosquito oviposition behavior, blood-host usage, and transmission of the causative agent of dog heartworm (Dirofilaria immitis); document the structure of the mosquito pyloric armature; and provide zoos with suggestions for mosquito control. My results underscore the medical and veterinary importance of studying mosquito blood feeding ecology in zoos, and the experimental utility of zoos for studying mosquito behavior. A total of 1,630 larvae and 4,349 adults representing 16 species was collected and identified. The most common species were Aedes albopictus, Ae. triseriatus, Culex erraticus, Cx. restuans, and Cx. pipiens complex. Principal components and multiple logistic regression analyses showed that across both zoos the overall larval mosquito presence (regardless of species) was predicted by ambient and site temperature, precipitation, dissolved oxygen, presence of natural habitats, and absence of aquatic vegetation. Pairwise species associations indicated significant habitat-based relationships between larvae of Ae. albopictus and Ae. triseriatus, and Cx. pipiens complex and Cx. restuans. Recommendations to zoo personnel, regarding larval mosquito habitat management, were to reduce or eliminate artificial containers and shade sources greater than or equal to 2 m above standing water, use mosquito larvicides when source reduction is not possible, and receive training in recognizing and mitigating larval mosquito habitats. Mosquitoes fed on captive animals, humans, and wild animals, and took mixed bloodmeals. Blood hosts included 1 amphibian species, 16 bird species, 10 mammal species (including humans), and 2 reptile species. Minimum flight distances (dispersal) from host locations ranged from 15.5 m to 327.0 m, with a mean of 94.1 m ± 13.4 m. No mosquitoes tested (n = 45) were positive for D. immitis. The pyloric spines of Ae. albopictus, Ae. j. japonicus, Ae. triseriatus, An. punctipennis, Cx. pipiens complex, Cx. restuans, Or. signifera, and Tx. rutilus were photographed and measured. Differences exist in qualitative and quantitative spine structure, with Aedes spp. forming one general group, Culex spp. another, and An. punctipennis and Or. signifera a third. The one specimen of Toxorhynchites rutilus examined was most like Culex spp. mosquitoes. Larval mosquito-habitat, adult mosquito-host associations, and pyloric armature and spine structures generally conformed to previously published accounts, indicating that mosquito biology inside zoos represents mosquito biology outside zoos. Therefore, zoos can be used for experiments not feasible in the field. However, novel variation (e.g., new, exotic host records) recorded in mosquito species warrants further investigation in zoos. My study demonstrates that zoos can be used as experiment environments to study mosquito behaviors (e.g., oviposition cues, innate versus learned host preferences, mosquito dispersal, and home range memory), and that findings can be extrapolated to non-zoo areas, while also providing medical and veterinary benefits to zoo animals, visitors, and the public