Pathogen vectors at the wildlife-livestock interface: molecular approaches to elucidating Culicoides (Diptera: Ceratopogonidae) biology

Abstract

Emerging infectious diseases (EIDs) continue to threaten human and animal welfare worldwide. Vector-borne pathogens are particularly concerning because of their ubiquity, pathogenicity, and lack of predictably due to insufficient ecological and evolutionary knowledge regarding vector species. Biting midges in the genus Culicoides (Diptera; Ceratopogonidae) have been implicated in the transmission of highly pathogenic viruses and parasites. In North America, the complete transmission cycles of many of these pathogens need further elucidation as outbreaks occur in the absence of known vector species. Further, our knowledge about the evolution and ecology of most Culicoides species is limited at best. The focus of research has been morphological identification and distribution of some North American Culicoides species, particularly those that are known pathogen vectors, but little is known about the phylogeny, ecology, behavior, and vector competence of many other species, some of which may prove to be important vectors. Ultimately, risk assessments and management of pathogens transmitted by Culicoides in North America are limited by this lack of knowledge and these limitations prove to be disastrous. Large and damaging outbreaks of Culicoides-transmitted viruses around the globe in the last two decades have raised concerns that North American livestock and wildlife are at risk. These threats, in conjunction with the lack of knowledge about vector species, led to the current research so that we can more insight into Culicoides biology and be proactive in preparing for future outbreaks. In chapter 1, I argue for more research into the biology of Culicoides in North America. In particular, I highlight our lack of information about all potential vector species, draw attention to the issues with the current taxonomic designations, discuss the main pathogens of concern transmitted by Culicoides, and then explore outbreaks of bluetongue virus in Europe as a case study illustrating how insufficient knowledge can lead to significant impacts on agriculture and the economy. I also summarize all known and potential vector species in North America to establish recognition of the tenuous nature of data used to incriminate or exclude some Culicoides species as vectors. I end with a discussion of the importance of systematics to research and management of vector-borne diseases and make a call for more Culicoides-based studies to minimize the biological, ecological, and economic damages caused by the introduction of exotic pathogens. In Chapter 2, I investigated the phylogentic relationships of the three known and described species in the C. variipennis species complex. In North America, two currently recognized vector species, C. sonorensis and C. variipennis, are thought to drive the transmission of bluetongue virus (BTV) and epizootic hemorrhagic disease virus (EHDV), respectively. Both species, along with a non-vector species C. occidentalis, belong to a morphologically similar species complex, the C. variipennis complex. The taxonomic history of this complex is convoluted due to the limited number of morphological characters that are species-diagnostic. My objective was to evaluate the current taxonomy of the C. variipennis species complex using multi-locus molecular phylogenetics and population genetic approaches in order to understand evolutionary relationships and historical biogeography within the complex. I determined that molecular data do not support the current taxonomy of the three species in the C. variipennis complex. In fact, phylogenetic approaches demonstrated very little resolution at the species level within the C. variipennis complex, except for a single clade that represented C. occidentalis. I detected weak population genetic structure using two nuclear loci, but this weak structure did not correlate with morphological species identification. I suggest a taxonomic reevaluation of this species complex is sorely needed so that more accurate species identification and distributions can be obtained. Improved species designations lead to more precise inferences about ecology and evolution of vector species. Through this taxonomic demystification, we can better grasp sylvatic transmission cycles and prepare for outbreaks of both domestic and foreign pathogenic viruses. In Chapter 3, I investigated host choice of Culicoides species collected in North America using DNA-based bloodmeal analyses. In North America, we currently lack the ability to understand transmission networks and assess outbreak risk for Culicoides-borne pathogens as our knowledge base of trophic ecology is meager at best. Knowing the diversity of vertebrate hosts for Culicoides can help identify susceptible species and pathogen reservoirs, and can lead to the detection of new vector species. The objective of this study was to identify the diversity of Culicoides hosts in North America. I sequenced two vertebrate mitochondrial genes (cytochrome oxidase I and cytochrome B) from blood-engorged Culicoides. I detected the mitochondrial DNA of 12 host species from five different Culicoides species and the C. variipenis species complex. The hosts included both mammals and birds. I documented new host records for some of the Culicoides species collected. The majority of the mammalian hosts were large ungulate species but I also detected a lagomorph and a carnivore. The bird species that were detected included House Finch and Emu, the latter is evidence that the species in the C. variipennis species complex are not strictly mammalophilic. These results demonstrate that Culicoides species will feed on multiple classes of vertebrates and may be more opportunistic in regards to host choice than previously thought. Throughout the dissertation, I have drawn attention to the areas for which we lack the most knowledge regarding North American Culicoides, illuminated the evolutionary relationships and taxonomic discrepancies in the C. variipennis complex, and improved our understand of Culicoides host choice. These are the first steps towards developing an improved understanding of Culicoides biology, in general, and the sylvatic cycles of Culicoides-borne pathogens, in particular. My research has also raised other important questions that deserve more attention so that we can better grasp the impacts that anthropogenic change has on Culicoides ecology, evolution, and pathogen transmission. As we increase our knowledge about this important group of insects, the ability to predict, prepare, and mitigate the economic and ecological damage from disease outbreaks will be greatly improved.