Biosystematics and Biodiversity of Pathogens and Parasites

APPROACH: Phylogenetic analyses of comparative morphological and molecular databases will be applied to new classifications, diagnostic probes, and recognition of unique morphological markers, and as the basis for a predictive frame work to understand patterns of parasite distribution and the potential for emergence and disease. Focus of research: a. medium and large stomach worms, nomenclature, diagnostic, cryptic species, and ecosystem interfaces. b. definition of helminth faunas of bovids and cervids in the Holarctic. c. phylogeny and nomenclature of Strongyloidea and Cyathostominae in horses. Research will be cased on materials held in the US National Parasite Collection (USNPC) and those derived from collaborations and through field studies. In the USNPC activities will focus on: a. Acquisition/curation of specimens-based collections; b. development of primary information systems, databases and historical baselines providing a predictive and preventive framework for documenting and understanding geographic distribution and host-association for parasites within the context of global change.

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PROGRESS: 2001/11 TO 2006/10<BR>
Progress Report 1. What major problem or issue is being resolved and how are you resolving it (summarize project aims and objectives)? How serious is the problem? Why does it matter? Helminth parasites are economically significant pathogens in ruminants, other food animals and equines, contributing to over $2 billion in losses annually in the United States. Efforts to understand epidemiology, distribution and disease and the most efficacious basis for control have often been hampered by difficulties in identification of pathogens and parasites. A paucity of knowledge about the species and populations involved and often limited information on geographic and host- distribution hinders our ability to predict the outcome of host-parasite interactions. Long-term challenges are evident that require resolution even as we develop more refined morphological and molecular capabilities for parasite identification. Identity and phylogenetic relationship are baseline information for defining host and geographic distribution and patterns of parasite diversity. Recognition of cryptic, emergent, exotic and invasive pathogens, determination of the importance of reservoir hosts such as wildlife, and elucidation of the influence of interfaces between agricultural and wild ecosystems on the structure of helminth faunas follow from biodiversity research. Additionally, among ruminants, the mosaic nature of the helminth fauna, particularly hidden species diversity, impacts our abilities to recognize emerging pathogens and invasive species or those increasingly influenced by anthropogenic and climatological global change. Consequently, continued strategic biodiversity survey and inventory in conjunction with phylogenetic systematic studies of parasitic groups are necessary to provide for accurate identification, diagnostics and definition of the parasitic fauna. Ecological perturbation linked to global change and its influence on the distribution of pathogens and emergence of disease can only be understood with a context provided by systematics. These issues form the framework of the current program for a series of integrated studies in systematics and parasite biodiversity among the crown clades of the strongylate nematodes primarily in ruminants and equines. Concurrently we will expand, and curate the specimens-collections of the US National Parasite Collection (USNPC). An infrastructure for Biodiversity Bioinformatics will link interactive information systems and diagnostic identification keys, with phylogenetic and epidemiological information for dissemination on the internet. We also provide a resource for reference specimens and information to support parasitology and animal health nationwide and globally. Specific goals and objectives include: Objective 1- Characterize parasite biodiversity in ruminants, and equines by integration of comparative morphology and molecular systematics. Detect, identify, and classify parasites, define geographic distribution, and host associations to recognize, and reduce the impact of, cryptic, emergent, exotic and invasive pathogens. Objective 1a- Trichostrongyloidea- Haemonchinae & Ostertagiinae- Describe and redescribe structural attributes for haemonchines and ostertagiines including the synlophe, genital cone, spicules and other characters; (2.) Examine generic level relationships, and hypotheses for host-parasite coevolution and distribution; (3) Examine relationships for species of Haemonchus, Ashworthius, and Mecistocirrus; construct keys for genera and species. (4.) Construct keys for Ostertagiinae at generic level; (5) Conduct morphological and/or molecular evaluations among putative cryptic species of ostertagiines. (6.) Stabilize nomenclature for polymorphic ostertagiines. Objective 1b. Metastrongyloidea, Protostrongylidae- Determine the identity and distribution for Elaphostrongylinae and Protostrongylinae in northern ruminants. (1.) Utilize molecular approaches, sequencing and comparative phylogenetics of ITS and mtDNA loci, to examine putative geographic partitioning in populations of parasites and their caprine hosts. Molecular and morphological criteria for identification of larval and adult parasites contribute substantially to resolving host and geographic distribution and the exchange of parasites between domestic and wild hosts. Objective 1c. Parasite Fauna, Holarctic Bovidae and Cervidae- Survey and inventory for parasites in wild and domestic ruminants leads to definition of the history and structure of the North American/Holarctic fauna: (1.) Develop comprehensive species lists for parasites in domestic and wild bovids and cervids. (2.) Summarize data for geographic range and host associations and synthesize information for each nematode species (literature and collections based information system), linked to specimen-based data and bioinformatics in the US National Parasite Collection. Objective 1d. Strongyloidea, Phylogeny and Nomenclature- Among major pathogens in equines:(1.) Improve identification and classification and complete phylogenetic studies of small strongyle (Cyathostominae) nematodes in horses. (2.) Improve terminology, nomenclature and diagnostics. Objective 2- U.S. National Parasite Collection- Expand, and curate the specimens-collections of the U.S. National Parasite Collection (USNPC). Develop interactive information systems linking diagnostic identification keys, with phylogenetic and epidemiological information for dissemination on the internet; build an infrastructure for Biodiversity Bioinformatics Relevance to ARS National Program Action Plan- Research is conducted under National Program 103, Animal Health (100%), involving 3 program components defined in the National Program Action Plan: 3.2- Protection of Animal and Human Health; 3.2.1- Animal Disease Diagnosis; and 3.2.2.1- Detection and Monitoring for Parasites. Components of this program contribute to: 1) Preventing the introduction of Exotic or Foreign Animal Diseases through the movement of domesticated animals and wildlife; 2) Identifying and predicting the impact of Emerging and Invasive Animal Pathogens and Parasites, and 3) Identifying hosts and distribution for Zoonotic Parasites and Diseases. Of special significance, as reflected in current Action Plans, is a focus invasive species at ecotones on the interface of agricultural/managed ecosystems and natural systems and the interaction of parasite faunas in domesticated food animals and wildlife. Parasitic worms and associated diseases continue to cause major economic impacts, and ultimately control is linked to a better understanding of the identity, along with the host and geographic distribution of various species. Improving methods for diagnosing and reducing the economic impact of parasitic worms depend on the accurate identification of the species involved. Definitive identifications are generally based only on detailed microscopic examination of adult parasites collected after the death of the host. Such has been further complicated recently by the recognition of cryptic species that are often morphologically indistinguishable from known pathogens. Thus, the development of diagnostic tools useful for eggs and immature stages in the environment or in fecal samples from the host, in addition to adult parasites, are needed to target appropriate methods for intervention. Accurate determination of the species causing economic impacts and articulation of phylogenies to predict new, emerging or invasive pathogens depends on a comparative approach integrating morphological and molecular data. Species-identity is linked to recognition of diagnostic criteria within a phylogenetic context. Long-term, basic level research on related groups of parasitic nematodes is the primary foundation for such synoptic studies. Identity and phylogenetic relationship are baseline information applied to understand patterns of parasite biodiversity and distribution requisite for documenting invasive and emergent pathogens. Following a century of often intensive investigations on the North American fauna, it is apparent that major lacunae continue to exist with respect to our knowledge of species diversity and host and geographic distributions. Also continually emphasized is the pervasive influence of translocation, introduction, and management practices on the distribution of parasites. Emergence of disease is linked to a breakdown in ecological isolating mechanisms, and primary controls are often anthropogenic in nature. This defines the need to completely document the North American fauna- to simultaneously understand what is endemic and to be able to recognize what is and can be introduced within a global context. Benefits/ Impact- There is nothing more fundamental than a comprehensive understanding of parasite biodiversity, including accurate taxonomy and species identity, evolutionary relationships, geographic distribution and host associations. Systematics research is the foundation. Accurate morphological characterization of parasites is critical for the reliability of any approaches to molecular diagnostics, comparative genomics and population genetics. Direct benefits to U.S. agriculture from the proposed research program: (1.) Refined knowledge of the North American fauna, and the interface of managed and natural ecosystems; (2.) Accurate definition of species, development of diagnostic criteria; (3.) Predictive databases and development of biodiversity-baselines to recognize the introduction and to limit the impact of exotic parasite species; (4.) Standardization of nomenclature for major taxa and species of economically important nematodes; (5.) Acquisition, maintenance and curation of the U.S. National Parasite Collection and databases. With continuing expansion of sustainable agriculture and organic approaches to animal production, and with the advent of resistance to anthelmintics by nematodes in cattle, it is increasingly critical to understand species diversity for pathogens and parasites that can emerge in the absence of established procedures for control. At the most basic level, improved diagnostics, anthelmintics, vaccines and other measures of control safeguard animal health and ultimately result in more efficient production of animals and animals products- all are linked to effective systematics. 2. List by year the currently approved milestones (indicators of research progress) Milestones and Expected Outcomes (1 October 2001- 30 September 2006/ FY 02- FY06) Milestone Time Lines. Objective 1a/ Trichostrongyloidea- Haemonchinae & Ostertagiinae Milestone 1 (12 months) - (a.) Comparative morphology of Haemonchus spp., Ashworthius spp., Spiculopteragia spp, Longistrongylus spp.; (b.) Sequencing for Ostertagia, Marshallagia and Teladorsagia. Milestone 2 (24 months) - (a.) Phylogeny of Haemonchinae and Haemonchus spp.; (b.) Morphology of Camelostrongylus, Marshallagia. (c.) Generic phylogeny Ostertagiinae. Milestone 3 (36 months) - (a.) Complete nomenclature Ostertagiinae; (b.) Key for Haemonchus spp.; (c.) Key for genera Ostertagiinae. Milestone 4 (48 months) - (a.) Completion molecular comparisons of ostertagiines. (b.) Assessment Mecistocirrus spp. Milestone 5 (60 months) - (a.) Completion illustrated keys, linkage to USNPC. Objective 1b/ Metastrongyloidea- Protostrongylidae Milestone 1 (12 months) - (a.) Confirm identity Parelaphostrongylys & Protostrongylus; field collections for adult and larval specimens, initiate sequencing. Milestone 2 (24-36 months)- (a.) Complete Sequence comparisons for protostrongylids. Milestone 3 (48 months)- (a.) Complete phylogeographic analyses for hosts and parasites. Objective 1c/ Holarctic Bovidae & Cervidae Milestone 1 (12-60 months) - (a.) Survey and inventory of nematode faunas, field collections supporting Objectives 1a and 1b. Milestone 2 (24-36 months) - (a.) Development host-parasite checklists. Milestone 3 (60 months) - (a.) Integration illustrated keys, phylogenetic, host & geographic data linkage to USNPC. Objective 1d/ Strongyloidea, Cyathostominae Milestone 1 (8 months) - Preliminary phylogeny of the Cyathostominea. Milestone 2 (24 months) - Comparative morphology of ovejectors of Cyathostominea and related genera. Milestone 3 (36 months) - Morphological phylogeny of the Cyathostominea. Milestone 4 (48 months) - Key to species of the Cyathostominea. Objective 2, US National Parasite Collection Milestone 1 (12-60 months). (a.) Continue acquisition and accessioning of specimens, and development of collections database. Maintain collections records on line. Milestone 2 (48 months). (a.) Refine geographic information in USNPC, develop GIS linkages. Milestone 3 (60 months). (a.) Linkage of specimens data in bioinformatics infrastructure. 4a List the single most significant research accomplishment during FY 2006. A. Single most significant accomplishment for FY 2006. Relevance to ARS National Program Action Plan- NP 103 Animal Health- This program is addressed in the Animal Health Action Plan through Objective 3.2: Develop and Deliver Science-Based Information and Technologies to Reduce the Number and Severity of Agricultural Pest, Insect, Weed, and Disease Outbreaks. Specifically this research supports Performance Measure 3.2.1: Provide scientific information to protect animals from pests, infectious diseases, and other disease-causing entities that affect animal and human health. Evolution of Trichinella nematodes defined: Completed first comprehensive phylogenetic and historical biogeographic evaluation of pathogenic Trichinella nematodes, agents of trichinellosis in humans, swine and wild mammals, birds and reptiles. Trichinella nematodes, are agents of parasitic disease in humans and food animals and an understanding of their evolution contributes to measures for management and control. Collaborations (APDL and the Trichinella Reference Center, Rome Italy) focused on molecular phylogenetic analyses of all extant species, and revealed origins for this group in central Eurasia and a complex history for geographic colonization and host switching among omnivorous and carnivorous mammalian hosts.With respect to infections in humans, the ancestral trait for all Trichinella is one of broad infectivity for mammals. The geographic distributions for Trichinella has been strongly influenced by anthropogenic drivers such as transport of infected swine and global introduction of parasites. Significantly this study is the foundation for detailed research in population genetics which for the first time allows molecular epidemiological approaches to be applied to these parasites and to understand patterns and distribution of human infection. 5. Describe the major accomplishments to date and their predicted or actual impact. Relevance to ARS National Program Action Plan- NP 103 Animal Health- This research is addressed in the Animal Health Action Plan through Objective 3.2: Develop and Deliver Science-Based Information and Technologies to Reduce the Number and Severity of Agricultural Pest, Insect, Weed, and Disease Outbreaks. Specifically this research supports Performance Measure 3.2.1: Provide scientific information to protect animals from pests, infectious diseases, and other disease-causing entities that affect animal and human health. (1) Globally species of Haemonchus, large stomach worms, are the most economically important gastrointestinal parasites of ruminants; veterinarians and scientists require a synoptic understanding of host and geographic associations and diagnostic criteria. Scientists of the US National Parasite Collection, and the Royal Veterinary College, London completed phylogenetic studies of all species of Haemonchus based on original descriptions and redescriptions along with the first coevolutionary and historical biogeographic analyses for this group. Specific Accomplishment: The phylogeny for Haemonchus is the foundation for a comprehensive reference system for diagnostic criteria and development of interactive identification keys; historically, coevolutionary and biogeographic analyses demonstrate the pervasive role of host translocation, and parasite introduction/ emergence in the global distributions of H. contortus, H. placei and H. similis the dominant pathogenic species in domestic stock. Impact: Phylogenetic resolution for Haemonchus is the foundation for development of accurate and reliable keys for identification, and a refined understanding of the geographic distribution and host association, all information required globally to control these parasites. (2) Large stomach worms include the most significant pathogens in domestic ruminants in North America and globally. Collaborations between scientists of the U.S. National Parasite Collection and the Royal Veterinary College, London, resulted in descriptions of new species,and redescriptions of genera and species of large stomach worms (Haemonchus, Ashworthius). Comparative morphological information will be utilized as diagnostic characters in the construction of illustrated keys for identification of large stomach worms. Impact: New information provided will be critical to prevent importation or to help manage the distribution of large stomach worms in ruminant hosts. (3) The comparative anatomy of ovejectors will provide key information required to determine evolutionary relationships among strongylid nematodes, the most important nematode pathogens of farm animals. A light microscope study of representatives of 22 subfamilies of strongylid nematodes was conducted at the U.S. National Parasite Collection in collaboration with Ian Beveridge, University of Melbourne; Lynda Gibbons, University of London; and Hideo Hasegawa, Oita Medical University, Hasama, Japan. The collaborators discovered significant new information on the recognition of homologous parts of ovejectors, and organized an International Workshop on Comparative Structure of Strongylid Nematode Ovejectors in Vancouver, August 5, 2002 to recommend a uniform terminology for the structures. Impact: The new information contributes to analyses of evolutionary relationships among strongylid nematodes providing a predictive tool for scientists developing diagnostic and control methods for these important pathogens of farm animals and man. (4) Nematodirinae are trichostrongyloid nematodes that globally include serious pathogens of ruminants, particularly caprines. Scientists of the U.S. National Parasite Collection completed the first phylogenetic analyses to reveal relationships among the 5 genera of nematodirines. Impact: Information from these studies contributes to species-level analyses of Nematodirus and Nematodirella, including species known as serious pathogens in domestic sheep and cattle, and to improved diagnostics for these Holarctic nematodes. (5) Biodiversity bioinformatics, represents an effective and synergistic foundation to develop, utilize and present diverse knowledge about the distribution, host associations, and history for parasites represented by specimens-based collections such as the U.S. National Parasite Collection. The Curator of the US National Parasite Collection in collaboration with colleagues at the Southwestern Museum of Vertebrate Biology, University of New Mexico, the Western College of Veterinary Medicine, University of Saskatchewan, Department of Ecology and Evolutionary Biology, Cornell University, and the Vantaa Forest Research Centre, Rovaniemi, Finland developed protocols for comparative biodiversity studies focusing on parasite-host assemblages in high latitude ecosystems of the Holarctic. Studies serve as models for expansion of specimens-based collections as information systems integrating strategic field collections and databasing for development of comparative baselines for biodiversity in a contemporary and historical context. Impact: Specimens collections become portals for access to critical information linking interactive keys, phylogenetic, biological and epidemiological data through species homepages for helminth parasites in Holarctic mammals, including those that circulate among domestic and wild ruminants. (6) U.S. National Parasite Collection, Curatorial Activities (FY 03, 04, 05, 06)- Supervision of national and international loans- approximately 1, 000 lots of specimens annually. New accessions and databasing for 1500- 2000 lots of specimens annually. Repair and maintenance of slides representing 1500-2000 specimens on slides annually. Transfer of significant collections to the USNPC (2003)- R.L. and V.R. Rausch Collection, University of Washington estimated 30,000 lots. Bench support for national and international visitors who examined in excess of 3,000 specimens. (7) Phylogenetic studies by a collaborative team including scientists at the University of Colorado and The Natural History Museum, London indicate that the occurrence of Taenia tapeworms in humans pre-dates the development of agriculture, animal husbandry and the domestication of cattle or swine. Taeniid tapeworms in African carnivores twice independently colonized hominids and the genus Homo, prior to the origin of modern humans coincidental with dietary and behavioral shifts from herbivory to scavenging and carnivory. Parasitological data provide a unique means of elucidating historical ecology, foraging behavior, and food habits of hominids during their evolution. Impact: The typical parasite fauna in humans is far older than previously considered and indicates a very prolonged association between humans and their characteristic food-borne pathogens; such insights are important in understanding epidemiology and distribution of Taenia in humans and food animals. (8) Discovered that the large stomach worm genus Haemonchus includes at least 2 additional species than the 10 previously recognized. One of the newly distinguished species was previously included within Haemonchus contortus, the cosmopolitan parasite of sheep and other domestic ruminants. Improved knowledge of the species composition of this economically important genus provides information critical to understanding patterns of host association and geographic distribution and contributes to the development of methods and agents for controlling these nematodes. (9) The taxonomy and systematics of medium stomach worms remains confused due to our yet incomplete understanding of polymorphism among males in a diversity of genera. In collaboration with scientists at the Universidad Complutense, Madrid molecular sequence analyses of Spiculopteragia asymmetrica and S. quadrispiculata demonstrated that these nominal taxa represent 2 males within a single polymorphic species. Demonstration of polymorphism in the genus Spiculopteragia had not been previously based on molecular criteria. Observations from this study serve to further validate the hypothesis for polymorphism among certain genera of the medium stomach worms, and provides a sound basis for identification and diagnostics of these nematodes in cervids introduced to North America. (10) The history for the Nematodirinae across the Holarctic, and definition of the coevolutionary and biogeographic associations for Nematodirella and Nematodirus in ruminant hosts was resolved. Based on the phylogeny for this group of parasites, historical analyses were completed that indicate a complex history for nematodirines in the Eurasia and North America. The fauna is composed of old or endemic species, versus those that are young and have been introduced following European contact. This study contributed to further definition of the mosaic nature of the nematode fauna of bovids and cervids in North America. It was further determined that introductions of parasite groups that originated in Africa (e.g., Haemonchinae, and Cooperiinae) form the core of a temperate fauna in domestic cattle and sheep. Responses to habtitat perturbation and global change by old or endemic assemblages (typically Boreal to Arctic) versus young and introduced parasites (typically Temperate to SubTropical) are predicted to differ. Significantly those parasites of the Temperate fauna may undergo range expansion to the north coincidental with climate change. Implications are apparent for understanding the potential responses and impacts of these parasites and the interactions of domestic and wild hosts at dynamic and changing ecotones under a regime of climate change. (11) Definitive redescriptions and keys for 65 species of equine strongyles were articulated. Redescriptions including synoptic line drawings and photomicrography of diagnostic structural characters for small and large strongyles of horses serve as the foundation for a synthetic comprehensive key for identification and diagnostics of adult parasites. Preparation of the key as hard copy and on line will make critical information available to the veterinary community. A draft of the keys for cyathostomins is available on the internet at: www.nas.gov. ua/iz/strong/. (12) New methods were developed and validated for geographically extensive and site intensive survey and inventory of parasites in ungulate hosts based on application of molecular sequence data. Protostrongyle nematodes include pathogenic parasites that reside in the pulmonary system, skeletal musculature, or the central nervous system of their ruminant hosts. Identification based on either adults in tissue and tissue spaces, or larval parasites in feces has remained problematic, and has hampered a detailed understanding of host distribution and geographic range. Such information is critical in defining the potential for disease, and the degree to which parasites may be shared among a number of different ungulates. We applied a combination of comparative morphology and molecular analyses to define the host and geographic range for Parelaphostrongylus odocoilei in North America. Molecular identification of dorsal-spined larvae (DSL) indicates that the protostrongylid parasite Parelaphostrongylus odocoilei occupies a broader geographic range in western North America than previously reported; covering 30 degrees of latitude. We analyzed 2,124 fecal samples at 29 locations from thinhorn sheep (Ovis dalli dalli and O. d. stonei), bighorn sheep (Ovis canadensis canadensis and O. c. californiana), mountain goats (Oreamnos americanus), woodland caribou (Rangifer tarandus caribou), mule deer (Odocoileus hemionus hemionus), and black-tailed deer (O. h. columbianus). DSL were recovered from populations of thinhorn sheep south, but not north, of the Arctic Circle, and were not recovered from any population of bighorn sheep examined. DSL were identified as P. odocoilei by comparing sequences of the second internal transcribed spacer (ITS2) region of ribosomal RNA among 9 protostrongylid species validated by adult comparative morphology. ITS2 sequences were markedly different among Parelaphostrongylus and other protostrongylid genera. IMPACT: The concepts demonstrated in our study are significant in the realm of molecular epidemiology and in circumstances where ruminant hosts cannot be collected for detailed necropsy. Our research represents the first study to combine extensive fecal surveys, comparative morphology, and molecular diagnostic techniques to comprehensively describe the host associations and geographic distribution of a parasitic helminth. The development of such epidemiological probes will have significant applications in veterinary and conservation medicine. (13) Developed first empirical model for responses of complex host- parasite system to globlal climate change and directional warming. Global warming, one of the most important drivers of emergence of disease in people and domestic livestock, is anticipated to have significant impacts on the occurrence of parasitic disease in arctic and subarctic wildlife. Many parasites of wildlife have life stages that develop in the environment or require invertebrate vectors for transmission but at northern latitudes these parasites are typically constrained by the long cold winters and short, cool summers. Climate warming is expected to relax some of these constraints and alter the epidemiology of the parasites. A simple predictive model for parasite development rates in the Arctic was developed from laboratory data and validated by field experiments on Umingmakstrongylus pallikuukensis, a protostrongylid nematode lungworm of muskoxen. This model indicates that during summers with normal temperatures larval development to the infective stage (L3) in the gastropod intermediate host requires two years, whereas if mean summer temperatures increase by as little as 1C development to L3 will occur in a single summer. Survival to L3 is significantly higher if larvae develop within a single season as opposed to having to over-winter, thus single season development may result in increased infection pressure for muskoxen. A retrospective analysis of temperature data from 1978-2003 shows development to L3 within a single year has become more common over time. Applying this same model to climate warming scenarios reveals that with increasing temperatures L3 will be available earlier and the transmission period will be longer. Current research on Parelaphostrongylus odocoilei, a related muscle-dwelling protostrongylid in Dalls sheep, suggests that this model also accurately predicts development rates for this parasite in a sub-arctic mountain environment. Caribou, a keystone species in the Canadian Arctic, are also infected by several protostrongylid nematode species that may be similarly affected by climate warming. Possible impacts of climate warming on host-parasite dynamics in the north include increased infection levels, emergence of sub-clinical and clinical disease, and expansion of the geographic distribution and host ranges of parasites. Such changes may threaten the stability and sustainability of wildlife populations. Additionally, many wildlife diseases are transmissible to people and changes in the dynamics of these infections may have public health implications. We continue to explore possible effects of climate change on northern host-parasite relationships and new initiatives to identify and address potential vulnerabilities in the health of caribou and wild sheep related to climate change and infectious disease. (14) Demonstrated structural polymorphism in first stage larvae of a Protostrongylid nematode. Nematodes of the family Protostrongylidae have significant economic impact among domestic and wild ungulates throughout the world. In North America various species are documented as serious pathogens that occur in the pulmonary system, skeletal musculature or central nervous system of domestic and wild sheep, and cervid hosts. Identification of larval parasites has remained problematic and this has limited our ability to carryout extensive geographic surveys or to fully understand epidemiology and the dynamics of transmission among intermediate hosts (gastropods) and final hosts that harbor adult worms. Recently molecular-based methods have been developed for definitive diagnosis and have been applied to studies of Parelpahostrongylus odocoilei. We have applied these techniques, based on comparisons of sequences from the mitochondrial and nuclear genomes to resolve the identification of certain larval parasites. We demonstrated unequivocally the occurrence of polymorphism in the structure of the tail among first-stage larvae representing a single species, Parelaphostrongylus odocoilei, among the Protostrongylidae. Two distinct larvae, with a characteristic dorsal spine, include: (1.) a morphotype with a conical tail marked by 3 distinct transverse folds or joints and a symmetrical terminal tail-spike; and (2.) a morphotype with a digitate terminal region lacking folds or joints and with an asymmetrical, subterminal tail-spike. These divergent larval forms had been postulated as representing distinct species of elaphostrongyline nematodes. IMPACT: Application of a multi-locus approach using ITS-2 sequences from the nuclear genome and COX-II sequences from the mitochondrial genome served to confirm the identity of these dorsal-spined larvae as P. odocoilei. Ours is the first demonstration of structural polymorphism among larvae of a single species in the Metastrongyloidea and Strongylida. The basis for this polymorphism remains undetermined, but such phenomena if discovered to be more widespread may contribute to continued confusion in discriminating among first-stage larvae for species, genera and subfamilies within the economically important Protostrongylidae. (15) Defined the linkage of historical and ecological processes in dispersal and establishment of parasites and complex host-parasite assemblages through natural and anthropogenic events. As knowledge has expanded about the structure, history and distribution of a global biota we have come to understand that myriad mechanisms and circumstances have served as determinants of organismal diversity. Among complex host- parasite systems we have shifted from a mechanistically simplistic view where diversification has been strongly linked to processes of maximum cospeciation and maximum vicariance, to one where empirical evidence describes a complex mosaic in which host switching and geographic colonization have often prevailed. Concepts are examined and framed by equating colonization with a breakdown in mechanisms for ecological isolation such as those driven by periodic global extinction, or episodic and cyclical climate fluctuation and ecological perturbation that characterized marine and terrestrial systems. Multiple events of biotic expansion in earth history serve as powerful model systems that are applicable in evolutionary and ecological time. Significantly, processes of invasion, establishment and emergence of pathogens and parasites are the same in both historical and contemporary systems. Highlighted is a new dimension essential for understanding patterns of introduction, dissemination, and emergence of pathogens and parasites in the current regime of directional change in global climate with attendant disruption of ecological continuity. 6. What science and/or technologies have been transferred and to whom? When is the science and/or technology likely to become available to the end- user (industry, farmer, other scientists)? What are the constraints, if known, to the adoption and durability of the technology products? (1) U.S. National Parasite Collection- Databases for specimen records including host and geographic distribution are on line (www.anri.barc. usda/bnpcu/index.html). These support the global parasitological community. (2) Identification keys to strongylid nematodes of horses released on the internet at: www.nas.gov.ua/iz/strong/ (3) DNA sequences from protostrongylid nematodes deposited in GenBank. 7. List your most important publications in the popular press and presentations to organizations and articles written about your work. (NOTE: List your peer reviewed publications below). Articles/ Television Interviews About Our Research- Washington Post [Kids Post]. F. Shen Dont open this can of worms- Leeches and Tapeworms and Lice- Oh Gross! October 26, 2005. Lectures (invited)- Hoberg E.P. Trichinella- archaic pathogens in space and time: Evolutionary interfaces for parasites, hosts and biogeography. Department of Biology, University of New Mexico. November 2005.