Location: Arthropod-borne Animal Diseases Research
2017 Annual Report
Objectives
Objective 1: Perform risk assessment of bacterial pathogen transmission by house flies.
Sub-objective 1.A: Develop more effective larval control techniques by understanding the role of microbes in larval development and fitness.
Sub-objective 1.B: Evaluate the role of fly-bacteria and bacteria-bacteria interactions in house fly pathogen transmission.
Objective 2: Determine biological characteristics of mosquito vectors influencing animal health in a changing climate.
Sub-objective 2.A: Model mosquito ecological niches and impact of climate change.
Sub-objective 2.B: Characterize the biology of discrete mosquito populations.
Objective 3: Develop methods to reduce biting midge transmission of animal pathogens.
Sub-objective 3.A: Identify and characterize the salivary protein components of Culicoides sonorensis.
Sub-objective 3.B: Identify potential Culicoides vectors of epizootic hemorrhagic disease and bluetongue.
Sub-objective 3.C: Determine breeding site characteristics of Culicoides spp..
Sub-objective 3.D: Evaluate efficacy of candidate pesticides against C. sonorensis.
Approach
An extremely small percentage of insect species transmit disease-causing pathogens to animals and humans. Specific biological and behavioral characteristics allow these vector insect species to be efficient means of pathogen propagation and transmission; however these same characteristics may be targeted by control measures to limit pathogen spread or disease vector abundance. The common purpose of these projects is to understand key components of the host-pathogen-vector cycle to reduce or prevent pathogen transmission by the most common disease vectors: house flies, mosquitoes, and biting midges (Fig. 1). House flies associate with bacteria-rich environments due to the nutritional requirements of their larvae. This research defines the role of bacteria in fly development, bacterial persistence during microbe and insect interactions, and pathogen dissemination. Natural selection for increased Culex tarsalis mosquito fitness for various habitats and animal hosts has left genetic markers (single nucleotide polymorphisms) throughout the genome. These markers can be associated with traits and used to predict regional entomological risk in a changing climate throughout the mosquito’s large geographic range. The identification of biting midges or Culicoides saliva components that facilitate pathogen transmission will lead to improved transmission and pathogenesis models. This information will enhance development of vaccines and other countermeasures to reduce disease transmission. Lastly, not all Culicoides are competent vectors and this study will determine vector species and their habitats to help estimate risk in specific geographic regions. This plan aims to limit pathogen transmission by targeting the connections between hosts, vectors, and their environments via the insects’ unique characteristics using novel disease control methods.
Progress Report
Related to Objective 1. A new method for comparing gene expression across fly species was developed by ARS and university partners and is being used to identify genes common or unique to larvae of veterinary importance (house, stable, horn and face flies.) This analysis was paired with “snapshots” of the microbiome profiles in the manure over time. These analyses will reveal the way these fly species differentially utilize manure during their development.
House flies have a remarkable expansion in genes involved in the innate immune response, especially antimicrobial peptides like defensins. ARS scientists determined that copies of defensins are used differently by house fly life stages (larvae, pupae, adults) and some are used in a tissue-specific manner (gut but not body). These studies give insight into the remarkable way that flies have adapted to live and thrive in septic environments like manure. These effector molecules could serve as possible alternatives to traditional antimicrobials.
Related to Objective 2. The Invasive Mosquito Project (IMP), a national mosquito monitoring program conducted by citizen scientists collected 500 mosquito samples from across the U.S. The submissions indicate the IMP has successfully been integrated into educational curricula around the country, which will enable niche mosquito sampling from any location within the continental U.S. The submitted mosquito samples have been used to identify the movement of mosquitoes between areas such as California, the Rocky Mountains, and the Great Plains. Precipitation tends to be higher in the West and decrease towards the East until reaching the Mississippi river, whereas temperature has a clear North-South trend with the northern areas being cooler than the southern. Based on collections, mosquito populations are divided into seven East-West clusters and none are North-South splits, suggesting adaptation to dryer environments has permitted the spread of Culex tarsalis and Aedes vexans disease vector mosquitoes to new geographic ranges and not adaptation to temperature. This will allow for predictions of which mosquito species will adapt to a changing climate and human disturbance on the landscape.
Related to Objective 3. Culicoides populations and their associations with specific breeding habitats were studied at multiple sites that differ in their animal use patterns (beef cattle, dairy units, farmed deer, and bison and cattle grazed prairie). The distribution of known and putative disease vectors in several landscapes of Eastern Kansas has been characterized and data is being analyzed to elucidate relationships with habitat type, soil characteristics, animal use patterns, microbial populations, and variation in seasonal rainfall. All studies on habitat use were completed in 2017. Site-specific soil chemistry and associated microbiomes are currently being analyzed and will be correlated to midge collections from those sites in the last several years.
The innate mammalian immune response to Culicoides midge feeding and saliva has been characterized in a mouse model and related to bite transmission of orbiviruses.
Accomplishments
1. Female house flies acquire and accumulate more bacteria from manure than male house flies. House flies develop within and fed upon livestock manure, which often contains bacteria that are pathogenic to humans. ARS scientists in Manhattan, Kansas and Kansas State University collaborated to demonstrate that male and female house flies differentially acquire and harbor bacteria from cattle manure. Bacteria that were tested included both non-pathogenic E. coli and the pathogen Salmonella Typhimurium. Female flies obtained and accumulated more bacteria then male flies, both internally in their digestive tract and on their surface. These sex-based differences may be related to females spending more time associated with manure due to nutritional needs and egg-laying interest. This finding may help in predicting and determining risk of pathogen dissemination by house flies.
2. Dose-dependent fate of the pathogenic bacterium Salmonella typhimurium in house flies. Salmonella typhimurium is a pathogenic bacterium often associated with food borne illness, and previously found in wild-caught house flies. Adult house flies acquire Salmonella bacteria when they feed on contaminated substrates such as animal dung and garbage. The survival and persistence of bacteria within the fly gut directly impacts transmission potential. ARS scientists in Manhattan, Kansas described for the first time a dose-dependent effect on the survival of the Salmonella Typhimurium in house flies, where, surprisingly, lower doses of bacteria proliferated to a greater extent within the fly than higher doses. Additionally, irrespective of dose, S. typhimurium both persisted and proliferated in house flies, making them important reservoirs and potential vectors of this pathogen. This finding can help better understand the risk flies pose in harboring and transmitting bacterial pathogens such as Salmonella to livestock and humans.
3. Bacterial dose and fly sex both affect pathogen excretion from house flies. House flies ingest, harbor and excrete pathogenic bacteria, transmitting (or vectoring) them in the process. How the amount, or “dose”, of bacteria a fly ingests as well as the sex of the fly impacts bacterial excretion has not been studied. ARS scientists in Manhattan, Kansas demonstrated for the first time that both fly sex and bacteria dose impact excretion of the pathogen Salmonella typhimurium from house flies. Male house flies fed a high dose of bacteria not only shed the highest proportion of infected droplets, but also shed the most bacterial cells per droplet. Interestingly, both male and female house flies fed the dose of bacteria ten times lower shed a higher percentage of the amount that had been ingested, compared to the high dose treatment. Collectively these results show that male flies who ingest large doses of bacteria may be more significant in transmitting pathogens in their excreta. Further, both male and female flies that ingest low doses of bacteria could serve as more substantial long-term disseminators of bacteria in the environment, thereby causing a greater risk to animal and human health.
4. Double knock-down: Using RNA-interference (RNAi) to suppress then restore gene function in the biting midge. Culicoides biting midges transmit pathogenic viruses to livestock and deer. Understanding the interactions between these insect vectors and the viruses they transmit is the first step in developing methods for blocking virus transmission. Because very few molecular tools for studying midges exist, understanding virus-vector interactions can be difficult. Scientists at ARS in Manhattan, Kansas and Kansas State University showed for the first time that the molecular tool RNA-interference (RNAi) can be used both to knock down, and subsequently to restore, the activation of genes in midges. This type of technology will allow scientists to experimentally manipulate midges which can help in understanding the genetic components that underlie the midge’s vector ability to transmit viruses.
Review Publications
Thomson, J., Yeater, K.M., Zurek, L., Nayduch, D. 2016. Abundance and accumulation of Escherichia coli and Salmonella Typhimurium procured by male and female house flies (Diptera: Muscidae) exposed to cattle manure. Annals of the Entomological Society of America. 110:37-44.
Mills, M.K., Nayduch, D., McVey, D.S., Michel, K. 2017. Functional validation of Apoptosis genes IAP1 and DRONC in midgut tissue of the biting midge Culicoides sonorensis (Diptera: Ceratopogonidae) by RNAi. Journal of Medical Entomology. 54:559-557.
Mills, M., Michel, K., Ruder, M., Pfannenstiel, R., Veronesi, E., Nayduch, D. 2017. Culicoides-virus interactions: infection barriers and possible factors underlying vector competence. Current Opinion in Insect Science. 22:7-15.
Nayduch, D., Burrus, R. 2017. Flourishing in filth: house fly-microbe interactions across life history. Annals of the Entomological Society of America. 110:6-18.
