John G. Hildebrand - US grants

Olfaction is decisively important for the control of many insect behaviors. Orientation toward hosts, feeding, egg-laying, aggregation, and courtship and mating in many species of insects -- including vectors of parasitic and infectious diseases and agricultural pests -- are strongly influenced or controlled by these antennal senses. Very little is known, however, about the processes in the insect central nervous system (CNS) through which olfactory information influences behavior and which might be exploitable for improved strategies of insect management. This research has as an ultimate goal the understanding and exploitation of central processes in olfaction and other antennal senses, first in the experimentally favorable "model" insect Manduca sexta and eventually in medically and economically important (but experimentally less favorable) species, which are key determinants of harmful or otherwise important insect behaviors. Present knowledge of the structure and physiology of sensilla and their receptor cells in the antennae and other sensory organs of insects provides a basis for a detailed neurophysiological and structural study of neural pathways subserving olfaction and other antennal senses in the insect CNS. In this project, we plan to continue our established research effort aimed at revealing neural mechanisms of information processing in the insect brain, through which biologically meaningful odors as well as mechanosensory, humidity, and temperature stimuli detected by the antennae generate "higher-order" neural signals that ultimately trigger or sustain significant behaviors. Our studies aim to trace the projections and connections of specific types of central neurons in the antennal-sensory pathway; to characterize by means of intracellular recording the electrophysiological responses of neurons in the antennal lobes of the brain to defined and biologically meaningful antennal stimuli; to analyze the processes of central integration of sensory information and the underlying synaptic organization and mechanisms in the antennal lobes; to probe for neuromodulatory mechanisms in the central olfactory pathway; to explore further the usefulness of 2-deoxyglucose radiochemical activity-labeling to map patterns of neural activity in the antennal sensory pathways in the CNS; and to pursue further our collaborative efforts to characterize the sex-pheromone system of Manduca, to unravel ultrastructurally the details of synpatic "wiring" in the olfactory glomeruli of the antennal lobes, and to characterize fully the responsiveness of individual receptor cells in antennal olfactory sensilla.

To understand how olfactory information controls certain behaviors, it is necessary identify neural elements in the central nervous system that process and relay olfactory information and to determine the mechanisms by which such elements exert their influence over particular motor acts. In favorable invertebrate preparations, the sensory afferent pathways and the neural elements that govern stereotyped behaviors are accessible to cellular study. These systems therefore serve as useful models for studies of neural mechanisms leading from sensory inputs to motor outputs. As has been demonstrated in numerous important cases, insights gained from such model systems can provide a basis for, and guide research on, less accessible vertebrate systems. One such useful model is the moth Manduca sexta, in which detection of certain chemical cues by antennal olfactory receptors stimulates stereotyped behaviors. For example, the sex pheromones released by a receptive female trigger a characteristic flight response in a male, by which he orients and moves toward, and eventually finds the signalling female for mating. Olfactory information about sex pheromones is processed in a specialized part of the brain that is found only in the antennal lobes of normal males and gynandromorphic females (with antennal lobes innervated by sensory axons from male antennae). Olfactory information integrated in these and other, "higher" brain centers ultimately descends to thoracic motor centers and to control flight. By means of intracellular recording and staining, this research will explore the physiology and structure of nerve cells in the pathway from olfactory centers in the brain to the flight-motor outputs in the thorax. The proposed studies will focus on: (1) the control of flight motor activity by odors (particularly sex pheromones) delivered to the antennae; (2) descending projection neurons that carry the integrated olfactory information from the brain to motor centers in the thoracic ganglia of males and gynandromorphic females; and (3) the roles of identifiable neurons in the thoracic ganglia as targets of the descending projection neurons and as possible modulators of flight motor activity. This research promises to expand our understanding of neural mechanisms of olfaction as well as of motor control. Learning about the mechanisms by which olfactory information exerts control over important behaviors in such a model system will help to fill a major gap in our understanding of the chemical senses and their roles in normal and abnormal neural and behavioral functions.

This proposal requests funds for the purchase of a computer- aided system for the reconstruction, measurement and statistical analysis of neuronal processes from sectioned or whole-mounted nervous tissue. The central goal of all the projects described is to understand how the detailed structure of neurons and their anatomical relationship?s with one another translate into the precise functioning of the nervous system. Five of the major users have projects that utilize various aspects of the insect nervous system as test subjects, a sixth investigates neural development in the amphibian nervous system.

This award provides funds to a group of neuroscientists at the University of Arizona for the purchase of a computer, networking hardware, and associated software. This equipment will be used to analyze experimental data resulting from comparative studies of olfaction and spatial orientation in lower and higher animals. Though collaborations with mathematicians, these investigators plan to generate models for nerve cell interactions that appear fundamental to these processes. Models will be based on anatomical and electrophysiological properties of the nervous system. Other models will have an explicitly behavioral basis. The models will developed using a neural network simulation program called GENESIS and the computer. Interspecific comparisons have traditionally provided a useful tool in understanding the underlying mechanisms of biological processes. Increasingly, the use of experimental data and theoretical schemes for the synthesis of models that make detailed predictions has played an equally important role in modern biology. Olfaction and spatial orientation are both problems of nervous integration that have interested neurobiologists for some time. The use of computational models, in particular neural network models, to get at the underlying integrative mechanisms is a promising approach to these classical problems.

The three participating research groups share a common interest in understanding the postembryonic development of neurons, the neural systems in which they function, and the muscles they control. Growing collaboration among these groups over the past three years and increasing mutual focus on the mechanisms that control the developmental plasticity of nerve and muscle cells have led to the emergence of this new Program Project. Among the research areas represented by these laboratories are: the roles and mechanisms of short-distance and contact-mediated intercellular influences in neural development; hormonally regulated, postembryonic differentiation and respecification of sensory and motor neurons and interneurons; ultrastructural and physiological aspects of synapse formation in developing neural systems; correlation and coordination of cellular, physiological, and neurochemical events in neurogenesis; the role of glial cells in the development of organized neuropil in the central nervous system; and developmental, physiological, anatomical, and molecular studies of olfactory, visual, mechanosensory, and motor systems. The participants propose to mount multidisciplinary, highly collaborative investigations of mechanisms underlying hormonal and trans- cellular regulation of the survival and development of nerve cells and muscles during postembryonic life. The five component projects in this program project probe for cellular and physiological mechanisms of: (1) influences of primary-sensory neurons, glial cells, and development- regulating hormones and neurotransmitters ont he development of arborizations and voltage- and ligand-gated membrane ion channels in olfactory interneurons in primary cell culture; (2) steroid-hormonal regulation of development of identified motor neurons in primary cell culture; (3) influences of steroid hormones on cellular, neurochemical, and systemic characteristics of the first-order olfactory center in the brain. All of the proposed studies are based on the use of non-vertebrate model systems that are economical, readily available, and experimentally tractable. Because they have been extensively studied by many investigators, including the participants in this program project, the insect preparations to be used in the proposed studies are especially well understood and favorable for these project, the insect preparations to be used in the proposed studies are especially well understood and favorable for these experiments. Taking advantage of this background, the component projects focus on neurons and glial cells that are identifiable as individuals or as specific functional types from animal. The basic mechanisms of development have been conserved during evolution. We expect, therefore, that information obtained in these projects about neural and muscular development in insects will illuminate related phenomena in other animals including human beings.

DESCRIPTION (adapted from applicant's abstract): The primary olfactory centers in the brains of diverse animals, including humans, are characterized by an array of synaptic modules called glomeruli. These centers are thought to be organized chemotopically, such that odor information is represented spatially among glomeruli, but this important hypothesis has not been tested comprehensively in a species offering the advantages of anatomical simplicity, identifiable glomeruli, accessible receptor cells and central neurons, and chemically identified, behaviorally relevant odors. Moreover, despite dramatic advances in chemosensory research, we still do not understand how complex odor stimuli are encoded in neural activity, within and among glomeruli, that ultimately leads to appropriate behavioral responses. This project builds on a firm foundation of technical experience and knowledge about an experimentally favorable model, the olfactory system of Manduca sexta, which is comparable to its vertebrate counterpart in organization and function and permits the hypothesis of glomerular chemotopy to be tested with greater precision than has been possible in other species. This model system also offers an exceptional opportunity to unravel the synaptic neural circuitry within and between identified glomeruli in order to reveal how specific odor information is processed at its first way-station in the brain. By means of intracellular recording and staining, extracellular multichannel recording, laser-scanning confocal and transmission electron microscopy, and computer-assisted mathematical modeling, we will focus on identified glomeruli in recognizable clusters to: (1) test the hypothesis that glomeruli are organized chemotopically by determining what chemical, temporal, and intensity properties of behaviorally significant odor stimuli are analyzed and encoded by individual neurons innervating particular glomeruli; (2) characterize synaptic circuits within and between glomeruli to learn how the various types of neurons associated with glomeruli interact to shape the signas conveyed by output neurons projecting to higher centers in the brain; and (3) develop mathematical models of characterized neurons to generate testable hypotheses about their physiology and synaptic interactions. This research will promote understanding of basic olfactory mechanisms in all animals, including mankind, and promises to contribute toward explanation of sensory disorders such as parosmia, hyposmia, and anosmia.

An important challenge in developmental neurobiology is to unravel mechanisms responsible for the influences exerted by afferent innervation on the development of its targets. Good examples of such phenomena are found in the olfactory system. In the brains of all species studied to date, development of the primary olfactory center with its characteristic array of synaptic glomeruli depends dramatically on innervation by primary- afferent axons of olfactory receptor cells. How sensory axons control the formation of glomeruli, and how odotopy -- the orderly spatial representation of attributes of odor molecules -- arises in the glomerular array, are pressing problems. This project will focus on the development of individual glomeruli and their uniglomerular projection neurons, endowed with characteristic "molecular receptive range" properties and will take advantage of a remarkable model system, the sexually dimorphic olfactors lobe in the brain of Manduca sexta. In particular, we will investigate the postembryonic development of the prominent macroglomerular complex in the male's olfactory lobes. This unique complex receives primary-afferent inputs solely from male-specific olfactory receptor cells, which induce the formation of the macroglomerular complex and are specialized to detect individual components of the female's sex-pheromone blend. The complex comprises two glomerular substructures, each of which receives and processes primary-afferent input about a different one of the two key components of the blend. These identified, odotopically defined glomeruli also contain neurites of male-specific central neurons, most notably uniglomerular projection neurons, which participate in specialized synaptic circuitry for processing sensory input about pheromone. As specific aims, the proposed studies ask: (1) how the macroglomerular complex develops, (2) what role(s) glial cells play in its development, (30 whether there are quantitative and temporal requirements for male-afferent control of its development, and (4) whether male-specific sensory axons have molecular specializations that might be responsible for induction of the macroglomerular complex. The ultimate goals of the line of research represented by this proposal are; (a) to ascertain the cellular and molecular mechanisms underlying the decisive role of sensory axons in the development of glomeruli, (b) to discover how primary-afferent and central elements ultimately destined to have similar, distinctive molecular receptive ranges come to associate with each other, and (c) to determine whether glial cells participate in the development of all of the glomeruli in a particular species. This is the first effort, in any species, to ascertain developmental processes and mechanisms underlying morphogenesis of anatomically identified olfactory glomeruli with known functional specificity. This research promises to add significantly to understanding of cell-cell interactions in the development of functionally specialized, modular neuropil in the central nervous system and thus to fuel progress toward improved understanding and treatment of developmental disorders of sensory systems.

4NSF IBN-9983302 8 February 2000


Lay Abstract

This research focuses on determining how female moths use their sense of smell to find appropriate plants on which to lay their eggs. We study an experimentally favorable and extensively investigated model insect, the tobacco hornworm moth Manduca sexta. Because little is known about hostplant finding by female moths, and in particular which compounds released by plants facilitate this interaction between moths and their specific hostplants, this research comprises two main tasks: (a) chemical separation of volatile compounds ("plant volatiles") collected from the air surrounding living plants, and (b) behavioral testing of the responses of female moths to these hostplant volatiles under controlled laboratory conditions. In the next year of this research, we will focus on utilizing a chemical separation technique called preparative gas-chromatography (GC) to fractionate headspace-volatile mixtures obtained from living plants and on testing the behavioral responses of adult female M. sexta moths to the isolated fractions. Currently we are constructing a special fraction collector that will allow us to trap volatile compounds separated by GC as they emerge from the GC column. As a fraction emerging from the GC column pass into a cooled collection tube in the fraction collector, the volatile compounds in the fraction will be trapped. The isolated fraction then can be tested for its effects on the flight behavior of female moths, in the laboratory, and if the fraction elicits appropriate behaviors, compounds making up that fraction will be identified by means of GC coupled with mass spectroscopy (GC-MS). We will focus on using one hostplant species, named devil's claw (Proboscidea louisianica var. fragrans), and one non-hostplant species, foliage of potato plants (Solanum tuberosum). Headspace-volatiles from potato foliage trigger strong behavioral responses in the moths both in the flight tunnel and in a bench-top bioassay that tests abdomen-curling responses (a simple behavior that immediately precedes egg-laying). Potato plants are closely related to hostplants of M. sexta and are attractive to female moths, but these plants are not good hostplants themselves because they are rejected for oviposition after a female moth has contacted the leaves of the plant. From our previous work in this project, we know that the behavioral responses of female moths to the volatiles released by greenhouse-grown potato plants are stronger than behavioral responses to several documented hostplants. Devil's claw plants are not related to the previously known hostplants of M. sexta, but we recently found that this plant is a surprisingly good hostplant nevertheless. We expect that both of these plant species, whose volatiles elicit strong behavioral responses from the female moths, will emit the same or very similar behaviorally active volatile compounds. By studying and comparing their volatiles and the behavioral effects those compounds elicit, we furthermore expect to be able to discover what volatile compounds are responsible for the attractiveness of hostplants. Because our research plan is directed toward obtaining publishable results within one year, our goal will be to focus on volatiles that elicit one of the two key behavioral responses that are elicited by hostplants and volatiles collected from them -- either odor-modulated upwind flight or the abdomen- curling behavior that precedes egg-laying.

The carbon dioxide (CO2) produced by the respiration of most organisms is a fundamental constituent of their odor. Many insects can sense the concentration of CO2 in the air around them, and they are thought to use that information in vital tasks such as locating food sources (e.g., appropriate plants) and possibly identifying desirable sites for oviposition (egg laying). Whereas considerable research has focused on how the sensory information about CO2 is acquired, little is known about how that information is processed in the central nervous system (CNS). In order to address this issue, this project will take advantage of an experimentally favorable and extensively studied model plant-eating insect, the moth Manduca sexta, using methods already developed and in practice in this laboratory. Pilot studies that led to this project suggested that a particular group of sensory receptor cells, situated in a sensory organ (the labial-palp pit organ, LPO) recessed in a deep invagination on each labial palp (a mouthpart) of the adult moth, are specialized to detect and quantitatively assess CO2 in the air around that organ. Anatomical evidence shows that although the LPO receptor cells are located in that mouthpart, they send their axons to the antennal lobe (AL), the primary-olfactory center resembling the vertebrate olfactory bulb, in the insect's brain. Furthermore, our pilot work clearly shows that some of the central neurons in the AL receive and process synaptic inputs, and thus information about CO2, from the LPO receptor cells. Thus, the project centers on recording of the electrical signals (coded sensory information about CO2) generated by LPO receptor cells and the responding neurons in the AL that receive inputs from the LPO sensory cells. In addition, the morphology of those AL neurons is studied by means of intracellular staining in order to reveal the types and patterns of branching of neurons contributing to processing of CO2 information. This research is the first study of central processing of environmental-CO2 information, and it promises to add significantly to our understanding of CNS olfactory mechanisms in insects. CO2, temperature, and humidity are environmental variables of importance to insects, and the sensory systems for those three kinds of stimuli share certain physiological properties. Therefore it may be that the processing of CO2 information has aspects in common with the processing of information about temperature and/or humidity, such that general principles of information processing in the brain could emerge from the proposed project. These studies are expected to lead to understanding of how information about environmental CO2 is first processed in the AL of the insect brain. Manduca sexta is an excellent experimental animal for this research because it: (a) possesses a highly developed CO2-detecting organ; (b) is large and hence ideal for neurophysiological studies, easily reared in the laboratory, and favorable for experimentation at the molecular, physiological, and organismal levels; (c) has yielded a wealth of information about the neurobiology of insect olfaction and the neuroethology of odor-guided behavior, much of it through previous research done in this laboratory; and (d) is an agricultural pest which, although not very important economically, exhibits behavior and sensory mechanisms that are similar to those of important, herbivorous pest insects. Thus, the findings from this project, complementing other research under way in this laboratory, will benefit ongoing basic research aimed at understanding how nervous systems analyze, recognize, and respond to odors, and should also yield insights that will be useful in the agricultural arena for designing new strategies for protection of crop plants from insect predation. Finally, in addition to its scientific impact and potential benefit to agriculture, this project will contribute to the research training of a postdoctoral associate (the key investigator) and also involve one or more undergraduate students in aspects of the studies.

COLLABORATIVE RESEARCH: DETECTION, PERCEPTION AND USE OF FLORAL CO2 IN NECTAR FEEDING BY MANDUCA SEXTA
John G. Hildebrand and Robert A. Raguso
University of Arizona and University of South Carolina

Many insects are sensitive to environmental carbon dioxide (CO2), and it is known to be important for attraction of blood-feeding species to their hosts, but the behavioral significance of CO2 for insects that feed on floral nectar is largely unclear. Preliminary studies yielded findings suggesting that the hawkmoth Manduca sexta (Lepidoptera: Sphingidae) may use elevated CO2, emitted by newly opened flowers, to help it find unexploited nectar sources during foraging. The principal goal of the proposed research is to confirm and establish the roles of floral CO2 in moths' foraging behavior, and hence pollination of flowers, by investigating ecological, behavioral, and neurophysiological aspects of the sensory detection of CO2 in Manduca. In particular, the project aims to discover: (1) the relationship between a flower's nectar content and CO2 concentration in front of the flower, both before and after a moth's visit; (2) whether and how Manduca uses floral CO2 during foraging; and (3) how sensory information about CO2 is processed and integrated with information about floral scent in the moth's brain. The investigation of foraging behavior in aim (2) addresses, in separate experiments, whether moths evaluate profitable individual flowers or plant patches, and whether foraging moths use sensory information about floral CO2 to increase their nectar intake. We also will test whether moths use floral CO2 alone or in conjunction with floral odor.
To achieve these aims, we will use a combination of experimental approaches. We will measure behavioral responses of individual moths, both in the laboratory (for experiments on flying moths in a tunnel and in a climate-controlled flight arena) and in the field (in outdoor flight enclosures), to artificial and natural flowers that vary in the amounts of nectar, CO2, and floral scent that they emit. We will use the techniques of sensory neurophysiology to record responses of nerve cells in the olfactory center of the moth's brain, when the moths are stimulated with CO2 and floral-scent compounds, both separately and together. This approach should yield insights about the adaptive responses of foraging moths to floral stimuli and their underlying neurobiological mechanisms.
Among its broader impacts, this research is expected to be useful to the agricultural community, as many moths and other plant-associated insects are beneficial pollinators and/or economically important pests. Manipulation of insect performance via CO2 cues is already used to control blood-feeding insects such as mosquitoes and, on the basis of this research, may be developed similarly to help control moths in an agricultural context. This work may also help to assess the consequences for ecological interactions (food webs, mutualisms) if ambient levels of CO2 continue to increase, as is predicted for global climate change. Importantly, moreover, participants in the project will profit from its multidisciplinary approach and gain experience with a range of methods and technical challenges. Involvement of two undergraduate students is planned. Both collaborating laboratories have records of supporting and training minority students and will continue to do so. The undergraduate student in the South Carolina laboratory will be chosen from the state-wide SC Alliance for Minority Participation in Research (SCAMP) and will visit Tucson each year to meet with project participants and participate in outreach activities. Research findings will be disseminated in written and oral form at scientific conferences and through institutions targeting the public, e.g. the Arizona-Sonora Desert Museum.

DESCRIPTION (provided by applicant): Triatomine bugs, commonly known as kissing bugs, vinchuca, chipo, barbeiro, are blood-sucking insects, vectors of the protozoan parasite Trypanosoma cruzi, the causative agent of Chagas Disease. This Disease is endemic in Mexico, Central and South America and affects about 11-13 million people. Control of Chagas Disease depends mainly on the elimination of vectors through use of residual insecticides and screening of blood banks for infection with T. cruzi. Interruption of vectorial transmission of Chagas Disease requires continuous entomological surveillance even in countries where the domestic vectors are in the process of being eliminated or have been eliminated because many wild animals act as reservoir hosts and many species of triatominae that could become domiciliated transmit the parasite. Surveillance could be effectively achieved by methods that actively detect the presence of insects, e.g. using odor-baited traps. Such methods would allow efficient monitoring of houses for the presence of the insects prior to application of intervention measures. In addition, the use of traps could contribute to control efforts under certain circumstances (e.g., under conditions of low insect population density), thus reducing the use of undesirable insecticides. Triatomine insects rely mainly on olfactory cues to find their hosts. Studies of the physiology and role of the olfactory system of these insects will provide knowledge that could be used to develop odor-baited traps. An initial step to develop these surveillance and control tools is to identify the chemical constituents of the attractive animal odors. We will use gas chromatography (GC) coupled to electrophysiological recording from olfactory receptor cells (ORCs) to characterize the chemical constituents of natural host odors that are detected by the olfactory system of Rhodnius prolixus, one of the main vectors of Chagas Disease. Moreover, we will couple this technique to multi-channel electrophysiological recordings from neurons in the antennal lobe (AL;the insect's primary olfactory center). This state-of-the-art technique (which we have been using successfully in our laboratory) will allow us to probe how odor information is represented in the AL. Moreover, because neural responses to odors at this central level of olfactory processing are highly sensitive owing to the high degree of convergence of ORCs into AL neurons, AL recordings will allow us to detect active constituents of natural odors efficiently and with high sensitivity. We will identify the active compounds using GC coupled to mass spectrometry (GC-MS), a technique also established in our laboratory. After identifying bioactive odors, we will develop attractive blends of synthetic odorants. To accomplish this, the efficiency of different blends will be evaluated by means of electrophysiological recordings of AL neurons and behavioral assays using dual-choice olfactometers. The use of recordings from brain neurons coupled to analytical chemical techniques to identify efficient attractant blends is novel and is being developed in our laboratory. Ultimately, efficient chemical attractants could serve as lures in traps for sensitive detection and trapping of triatomines in or around houses. PUBLIC HEALTH RELEVANCE: Triatomine bugs are blood-sucking vectors of Chagas Disease, a parasitic infection that affects more than 11 million people in the Americas. Complete and continuous interruption of disease transmission by these insects requires improvement of entomological surveillance, which could be effectively achieved by methods (e.g. odor-baited traps) that use natural attractants (e.g. host-odors) actively to detect the presence of the insects. We propose to use neurophysiological, analytical chemical, and behavioral methods to identify odor attractants used by Rhodnius prolixus, one of the main vectors of the disease, to find its hosts and that can be used as trap lures.

This project creates resources and a conceptual schema for developing informed policies that support ethical and responsible research and data practices at academic institutions working with Indigenous Peoples. Although public investment in science has led to numerous societal benefits, research has not always been conducted in ethical and responsible ways, especially in studies involving Indigenous populations. Moreover, efforts to address harmful research practices have tended to be reactive and superficial rather than systematic and collaborative. These issues are addressed by using an innovative Indigenous data governance framework to review institutional norms and practices that promote or inhibit ethical design, outcomes, and approaches across the STEM research landscape and address barriers to developing institutional policies and practices that are responsive to the ethical concerns and priorities of Indigenous Peoples. In so doing it provides research leadership opportunities for early career, women, and Indigenous scholars, and improves institutional guidelines and practices by providing tools to address historical barriers to inclusive and ethical research infrastructures.

This project develops an Indigenous data governance framework by drawing on international norms (i.e., the United Nations Declaration on the Rights of Indigenous Peoples) and relevant data governance standards (i.e., the CARE Principles for Indigenous Data Governance). These sources are used to form a normative matrix that guides project activities. Major goals include: (1) analyzing scholarly literature and institutional policies to develop indicators for assessing ethical research and data practices when working with Indigenous communities; (2) conducting surveys and interviews to identify strategies adopted in similar colonial contexts (e.g., New Zealand, Australia, Canada); and (3) identifying factors that support or inhibit ethical research and data practices with respect to Indigenous communities. A central feature of the project is bi-directional engagement of relevant constituents, rights-holders, and stakeholders in developing indicators, factors, and resources. Indigenous and mainstream experts and Indigenous leaders and community members will help inform the project?s early stages and participate in co-developing guidelines. These activities will advance knowledge of ethical research practices with respect to marginalized communities, outline methods for increasing ethical and responsible research more generally, and promote ethical and responsible research by engaging diverse knowledge systems and communities to increase research data relevance, quality, and reproducibility.

This project was co-funded by the BIO and CISE directorates.

This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.