Progress 12/15/18 to 12/14/24
Outputs
Target Audience:The target audience of these studies include policy makers (national and regional), veterinary diagnosticians, livestock producers and veterinarians, and industry stakeholders (dairy, beef cattle, sheep, deer, bison and wildlife). We are engaging with leaders of Future Farmers of America (FFA) programs in our local school district in order to develop interactive field exercises that incorporate aspects of vector-borne disease and ruminant host species. The goal of this exercise is to integrate problem solving for understanding and preventing vector-borne diseases that affect the livestock industry. We have identified and assigned an individual to maintain our laboratory website so that the project and project team can publicize research findings and distribute educational materials. Team members will continue to be involved in developing and leading symposia and workshop based on advancing next generation sequencing techniques. Some of these techniques were used for this project. These symposia and workshops are aimed at fostering the adoption of genomics approaches by diagnosticians and wildlife researchers and includes participation from many members of the team including the PI's (Mark Stenglein and Christie Mayo). Important aspects of reaching our target audience through our broader impact activities include reinforcing the study of a globally emerging arbovirus infections of livestock and wildlife, including the potential impact of climate change thereon. Changes/Problems:There are no changes to report for the final report and all pertinent changes have been reported in previous annual progress reports. What opportunities for training and professional development has the project provided?The project has served as a venue for the involvement and training/education of undergraduate, professional (DVM), producers, extension agents, graduate academic students, and postdoctoral scholars. The breadth of opportunities for these individuals included field, laboratory, and modeling studies. One of our recent graduates will continue collaborative Culicoides work as a faculty member at the Universidad Ana G. Mendez in Puerto Rico. Another graduate student will be defending in May and moving on to a postdoctoral research position that will involve mathematical and statistical modeling of SARS-CoV2 infection dynamics in ruminants in the Western United States. This will continue her work in this host community system and was made possible through skills she did not previously have but acquired through her involvement in this grant. Many of the postdoctoral scholars that worked on the modeling aspects of the grant allowed for them to have the opportunity to collaborate with such a large multidisciplinary team. Additionally, our work has allowed us to develop international collaborations with the Pirbright institute in the UK and National University of San Marcos in Peru. How have the results been disseminated to communities of interest?We have continued to grow as a team of individuals dedicated to accomplishing our project goals in a collaborative space. We have also participated in several opportunities that support the broader impacts of this grant. We have also participated in two 4-H events and participated in some K-12 programs with Dr. Wade Ingle showcasing this work in his outreach program. One of our graduate students has participated in the letters to a pre-scientist program sharing about our laboratorie's research. Results of the laboratory studies have been published in peer reviewed journals and presented to relevant professional and commodity groups including regional (County-based) livestock producer groups, the American Association of Veterinary Laboratory Diagnosticians, the Conference for Research Workers in Animal Disease (CRWAD), 4-H groups, genomics workshops, Society for Vector Ecology (SOVE), and European Congress of Entomology (ECE). We have been able to present in person and shift to the virtual platform for most of these opportunities and have remained actively engaged in discussions to promote website development and additional outreach. PI's and post docs have been interviewed for relevant articles in order to explain the effects of climate change on vector-borne disease and graduate students have been engaged in STEM, presenting our materials to elementary school children in Loveland. Dr. Mayo was interviewed for an article published in Science (2024) entitled: "Livestock virus hits Europe with a vengeance" What do you plan to do during the next reporting period to accomplish the goals?
Nothing Reported
Impacts
What was accomplished under these goals?
Objective 1: The primary goal of this objective was to quantify gene reassortment frequency in Culicoides sonorensis midges under various conditions. Our initial study examined the effect of temperature (20, 25, 30C) on bluetongue virus (BTV) coinfection in midges. The key findings included higher temperatures led to faster BTV replication but shorter midge survival and limited reassortment observed, with most sequences aligning to BTV-17. This work was published in Molecular Sciences (2024). Our second study investigated reassortment with different BTV-10 and BTV-17 coinfection rations. Key findings allowed us to understand most plaques aligned with the parental strain and higher titer. There was a significant amount of reassortment observed in midges at a 75:25 ratio and all of this work was published in Viruses (2024). Our final study explored coinfection at the cellular level using in situ hybridization. Our key findings allowed us to understand detection patterns varied, with some midges showing both serotypes. There was limited overlap of BTV-10 and BTV-17 at the cellular level and all of this work was published in Pathogens (2023). Overall, these studies enhance our understanding of BTV reassortment and its implications for viral evolution in the context of climate change. Objective 2: The initial plans for modeling the reassortment of BTV and EHDV under Objective 2 were based on observing a common reassortment pattern under Objective 1, which did not occur. Instead, it was found that dominant strains limit reassortment opportunities. This led to the development of a novel model of BTV infection and reassortment dynamics with a Culicoides midge, published in Viruses Evolution. The model, based on established biological knowledge and empirical studies, suggests reassortment is most likely when two strains are inoculated simultaneously or close together and have similar production rates. If production rates differ significantly, reassortment is less likely unless more productive strain is inoculated second and there is a long eclipse phase. The role of temperature in these dynamics remains unclear, but the study provided new modeling tools and recommendations for future research. Objective 3: Field data on interactions between Culicoides sonorensis midges and ruminant mammals were collected to assess how mammal densities impact life traits and the potential for density-dependent transmission of BTV and EHDV. The findings, synthesized into a manuscript under review by co-authors, indicate that as host abundance increases, so does the number of vectors per host. This could lead to a higher transmission potential. Collectively, this supports the assumption of density-dependent transmission. Additionally, midges from the 2021 field season were tested for orbivirus infection, revealing differences in host communities, infections prevalence, and vector species composition between sites. Statistical analyses are ongoing to quantify the direct impacts of host diversity. We analyzed results from a final field survey of potential larval habitats across different environments, to see where and how oftenCulicoidescan develop and emerge. We found thatCulicoidesin general were found in natural areas as much as or more than on livestock operations, thoughCulicoidessonorensiswere more associated with livestock habitats. The manuscript detailing this work has been submitted to the Journal of Vector Ecology and is currently under revision. Objective 4: To quantify the potential for different vertebrate host species to transmit BTV and EHDV, researchers evaluated the immunological and pathogenic responses to BTV infection in sheep and Reeves' muntjac deer using two serotypes (BTV-10 and BTV-17). They found that sheep showed robust infections with both serotypes, while muntjac had a more transient BTV-10 infection but a similarly robust BTV-17 infection. Midges feeding on infected sheep had higher BTV uptake compared to those feeding on muntjac. The study also explored the role of the immune response, noting differences in serological and cellular responses between species and serotypes. A mathematical model of BTV infection dynamics in ruminants was developed, which accurately recreated the temporal dynamics of infection and immune response, highlighting the significant role of lymphocytes in controlling infection. These findings suggest differences in transmission efficacy between livestock and wildlife ruminants and provide insights into host-vector interaction and viral pathogenesis. Objective 5: To determine whether reassortment of BTV and EHDV is more likely in midges or ruminants, we extended the within-midge infection model to account for temperature dependence, noting that warmer temperatures increase blood-feeding rates and mortality. This suggests quicker co-infection but shorter midge lifespan. They also extended the within-ruminant model to allow for co-infection and reassortment, finding a short window for successive infections to result in reassortment. Using an SEIR-SEI model, we're able to quantify co-infections during an epidemic, concluding that midges have a greater potential to produce reassortant viruses due to their longer-lasting infections. Objective 6: Our team developed a temperature-driven model of midge population dynamics and BTV transmission to understand how mean temperature and seasonal temperature amplitude affect outbreak sizes. We've identified that outbreak sizes are larger when mean temperature is close to the optimal for BTV transmission and seasonal amplitude is small, and larger when mean temperature is farther for optimal and seasonal amplitude is high. In Northern Colorado, where mean temperature is relatively cool, outbreak sizes are sensitive to seasonal amplitude. Additionally, an agent-based model tailored to Larimer and Weld Counties suggests that deer may significantly contribute to BTV transmission, though other factors like midge distribution among farms cannot be ruled out. Both studies under data analysis for manuscript preparation and publication.
Publications
|
Progress 12/15/23 to 12/14/24
Outputs
Target Audience:The target audience of these studies include policy makers (national and regional), veterinary diagnosticians, livestock producers and veterinarians, and industry stakeholders (dairy, beef cattle, sheep, deer, bison and wildlife). We are engaging with leaders of Future Farmers of America (FFA) programs in our local school district in order to develop interactive field exercises that incorporate aspects of vector-borne disease and ruminant host species. The goal of this exercise is to integrate problem solving for understanding and preventing vector-borne diseases that affect the livestock industry. We have identified and assigned an individual to maintain our laboratory website so that the project and project team can publicize research findings and distribute educational materials. Team members will continue to be involved in developing and leading symposia and workshops based on advancing next generation sequencing techniques thatwere optimized and used for this project.These symposia and workshops areaimed at fostering the adoption of genomics approaches by diagnosticians and wildlife researchers and includes participation from many members of the team including the PI's (Mark Stenglein and Christie Mayo). Important aspects of reaching our target audience through our broader impact activities include reinforcing the study of a globally emerging arbovirus infections of livestock and wildlife, including the potential impact of climate change thereon. Changes/Problems:
Nothing Reported
What opportunities for training and professional development has the project provided?The project has served as a venue for the involvement and training/education of undergraduate, professional (DVM), producers, extension agents, graduate academic students, and postdoctoral scholars. The breadth of opportunities for these individuals included field, laboratory, and modeling studies. One of our recent graduates will continue collaborative Culicoides work as a faculty member at the Universidad Ana G. Mendez in Puerto Rico.Another graduate student will be defending in May and moving on to a postdoctoral research position that will involve mathematical and statistical modeling of SARS-CoV-2 infection dynamics in ruminants in the Western United States. This will continue her work in this host community system and was made possible through skills she did not previously have but acquired through her involvement in this grant. Many of the postdoctoral scholars that worked on the modeling aspects of the grant allowed for them to have the opportunity to collaborate with such a large multidisciplinary team. Additionally, our work has allowed us to develop international collaborations with the Pirbright institute in the UK and National University of San Marcos in Peru. How have the results been disseminated to communities of interest?We have continued to grow as a team of individuals dedicated to accomplishing our project goals in a collaborative space. Wehave participated inseveral opportunities that support the broader impacts of this grant including4-H events and some K-12 programs with Dr. Wade Ingle showcasing this work in his outreach program. One of our graduate students has participated in the letters to a pre-scientist program sharing about our laboratorie's research.Results of the laboratory studies have been published in peer reviewed journals andpresented to relevant professional and commodity groups including regional (County-based) livestock producer groups, the American Association of Veterinary Laboratory Diagnosticians, the Conference for Research Workers in Animal Disease (CRWAD), 4-H groups, genomics workshops, Society for Vector Ecology (SOVE), and European Congress of Entomology (ECE). We have been able to present in person and shift to the virtual platform for most of these opportunities and have remained actively engaged in discussions to promote website development and additional outreach. PI's and post docs have been interviewed for relevant articlesin order to explain the effects of climate change on vector-borne disease and graduate students have been engaged in STEM, presenting our materials to elementary school children in Loveland. Dr. Mayo was interviewed for an article published in Science (2024)entitled: "Livestock virus hits Europe with a vengeance" What do you plan to do during the next reporting period to accomplish the goals?This is the final year of this project so there is nothing to report for the next reporting period.
Impacts
What was accomplished under these goals?
Objective 1: The 3 studies performed for objective 1 sought to characterize reassortment in colony raised Culicoides sonorensismidgesthat were coinfected with 2 different endemic strains of bluetongue virus (BTV-10 and BTV-17). The remaining work is now completed and published in International Journal of Molecular Sciences and Viruses in 2024. Objective 2: Modeling efforts for this objective were adjusted based on findings from Objective 1. Here, we elected to explore how often hypothetical pairs of strains with differing relative growth rates and other properties would be expected to undergo reassortment. To address this question, we developed a novel model of BTV infection and reassortment dynamics within an individual midge (published in Virus Evolution). Mathematical models of within-host infection dynamics are relatively well established for human infections, but there are only a very small number of published models of infection dynamics within an insect vector. Those few models are limited to mosquito-borne pathogens of humans, and none of them have explored reassortment. Based on a combination of established biological knowledge of BTV infection dynamics within midges and published empirical studies that observed more frequent reassortment than was observed under objective 1, we developed a mathematical model of two co-infection strains of BTV in a Culicoides midge. Overall, our modeling results suggest that reassortment is most likely to occur when inoculations of the two strains occur simultaneously or close to one another and when the production rates of the viruses are similar. Objective 3: We collected field data about interactions between C. sonorensis midges and ruminant mammal species to assess how ruminant mammal densities impact C. sonorensis life traits with the potential to cause density-dependent transmission of BTV andEHDV. Previous work on host density effects in C. sonorensis has been analyzed, and the findings have been synthesized into a manuscript currently under coauthor review and being finalized for submission. Our results find that as host abundance increases, the number of vectors per host animal also increases, leading to an increase in transmission potential with an increase in hosts. Overall, work carried out under this objective supports an assumption of density-dependent transmission in this system. In addition,midges from 2021 field season were sorted byspeciesand testedfor orbivirus infection via qRT-PCR. Statistical analyses are ongoing to quantify direct impacts of host diversity, but preliminaryfindings show differences in host communities, differences in infection prevalence, and differences in vector species composition between sites. Objective 4: To address this objective, we evaluated the immunological and pathogenic response to BTV infection in sheep and Reeves' muntjac deer using two serotypes endemic to the US. We found that across both viral serotypes and animal host species, all positively inoculated animals showed detectable circulating virus during a longitudinal time course (30 days). Differences were seenin the rate of initial detection and duration of infection between both species and serotypes investigated. This work has been published in Viruses. Additionally, we developed a mathematical model of BTV infection dynamics within a ruminant host. Similar to the within-host model of BTV infection dynamics developed under Objective 2, this model begins at the time of virus inoculation of a ruminant animal and tracks the concentration of free virus and the infection status of cells over the course of the infection. Importantly, the model for ruminant hosts also integrates aspects of the host immune response. We found that this model was able to simultaneously recreate the temporal dynamics of all of these empirically measured variables, suggesting that it provides a biologically realistic portrayal of the dynamics of BTV infection and immune response in ruminants. Overall, these studies address the objective by evaluating the host response, vector uptake, and viral pathogenesis across multiple host species and two BTV serotypes. Objective 5: We approached Objective 5 by focusing on the question of whether reassortment of BTV and EHDV is more likely to occur in midges or ruminants. First, we extended the within-midge infection model developed under Objective 2 to account for temperature dependence. We considered the empirical observation that blood-feeding rates and mortality rates are temperature-dependent, with blood-feeding occurring more quickly and mortality elevated under warmer temperatures. This suggests that two strains co-infecting a midge can happen in quicker succession under warmer temperatures (which increases reassortant production) but that midges will not live as long (which decreases reassortant production). Second, we extended the within-ruminant model developed under Objective 4 to allow for co-infection and reassortment. Using this model, we found that there is a particularly short window of time in which successive infections must occur for appreciable reassortment to happen. Third, to put these individual-level tendencies into a population-level context, we used an SEIR-SEI model for ruminant and midge BTV infection dynamics to provide guidance on the expected number of co-infections that could occurin a limited window, a part in midges and ruminants during a realistic epizootic of two strains of BTV. Combining that expected number of co-infections with the expected production of reassortants arising from a co-infected individual allowed us to quantify the total numbers of reassortant infections arising from midges and ruminants, respectively. In general, we found that midges have greater potential to produce reassortant viruses due to their potential for longer lasting infections. Objective 6: A temperature-driven model of midge population dynamics and BTV transmission dynamics was first developed to understand how two aspects of temperature seasonality--mean temperature and seasonal temperature amplitude--affect outbreak sizes. To do this, we developed an ordinary differential equations model of coupled midge population dynamics and BTV transmission dynamics, with midge and BTV traits driven by temperature according to published laboratory experiments. We found a strong interaction between mean temperature and seasonal temperature amplitude such that when mean temperature is close to the optimal temperature for BTV transmission, outbreak sizes are larger when seasonal temperature amplitude is small. When mean temperature is farther from the optimal temperature, outbreak sizes are larger when seasonal temperature amplitude is high. In Northern Colorado, mean temperature is relatively cool, suggesting that BTV outbreak sizes are likely sensitive to seasonal temperature amplitude. This work is currently being processed for publication. Additionally, we developed an agent-based model of BTV transmission in the NetLogo programming language that was tailored to Larimer and Weld Counties in Northern Colorado. Using a highly realistic and detailed model, we explored what the contribution of each wildlife species is to overall transmission by performing simulation experiments in which we removed a single wildlife species in each of several treatments. These simulations suggest that deer may play a significant role in facilitating BTV transmission across this landscape, although there are factors not considered by the model that cannot be ruled out as alternatives, such as spread among farms by midges. This work is currently being processed for publication.
Publications
- Type:
Conference Papers and Presentations
Status:
Accepted
Year Published:
2024
Citation:
Westrich, J.A, McNulty, E.E., Stoltz, M., Sherman, T.J., Carpenter, M., Burton, M., Nalls, A., Rubio, H.S., Sandoval, A., Mayo, C. and Mathiason, C.K. Determining longitudinal viral progression and immunological responses to Bluetongue Virus in experimentally infected ruminants; Oral presentation at CSU MIP Celebration; 2024 June; Fort Collins, CO.
- Type:
Conference Papers and Presentations
Status:
Accepted
Year Published:
2024
Citation:
Westrich, J.A., Warren, C.J., McNulty, E.E., Nalls, A.V., Mayo, C., Mathiason, C.K.. RNA flow cytometry as a reliable compliment to antibody based immune cell detection in nontraditional model systems; Poster presentation at CSU MIP Celebration; 2024 June; Fort Collins, CO.
- Type:
Conference Papers and Presentations
Status:
Accepted
Year Published:
2024
Citation:
Westrich, J.A., McNulty, E.E., Stoltz, M., Sherman, T.J., Carpenter, M., Burton, M., Nalls, A., Rubio, H.S., Sandoval, A., Mayo, C. and Mathiason, C.K. Serotype specific differences in experimental Bluetongue virus infection of two disparate host species; Oral presentation at 24th annual meeting of Rocky Mountain Virology Conference; 2024 September; Bellvue, CO.
- Type:
Conference Papers and Presentations
Status:
Accepted
Year Published:
2024
Citation:
Perkins TA, Cavany S, Majumder A, Westrich J, Sherman T, Carpenter M, Stenglein M, Matthiason C, Mayo C. Viral reassortment: An explanation of its drivers with models at different scales; Oral presentation at Society for Mathematical Biology Annual Meeting; 2024 July; Seoul, South Korea.
- Type:
Peer Reviewed Journal Articles
Status:
Published
Year Published:
2025
Citation:
Navarro-Mamani D, Jurado J, Vargas-Calla A, Ponce K, Sherman T, Zarate Y, Murga-Moreno C, Perez I, Villacaqui R, Ara M, Ortiz P, Rivera H, Mayo C, 2025. National Seroprevalence and Risk Factors of Bluetongue Virus in Domestic Ruminants of Peru. Transboundary and Emerging Diseases. 10 January 2025 https://doi.org/10.1155/tbed/2690231.
- Type:
Peer Reviewed Journal Articles
Status:
Published
Year Published:
2024
Citation:
Westrich, J.A., McNulty, E.E., Stoltz, M., Sherman, T.J., Carpenter, M., Burton, M., Nalls, A., Rubio, H.S., Sandoval, A., Mayo, C. and Mathiason, C.K., 2024. Immunological and Pathogenic Differences of Two Experimental Bluetongue Virus Serotype Infections Evaluated in Two Disparate Host Species. Viruses, 16(10), p.1593. DOI: 10.3390/v16101593; PMID: 39459926; PMCID: PMC11512378
- Type:
Conference Papers and Presentations
Status:
Accepted
Year Published:
2024
Citation:
Carpenter M, Benavides Obon A, Kopanke J, Lee J, Reed K, Sherman T, Rodgers C, Stenglein M, Mayo C. In situ hybridization (RNAscope) detection of bluetongue virus serotypes 10 and 17 in experimentally co-infected Culicoides sonorensis; Poster presentation at American Association of Veterinary Laboratory Diagnosticians Meeting; 2024 October; Nashville, TN.
- Type:
Peer Reviewed Journal Articles
Status:
Published
Year Published:
2024
Citation:
Carpenter M, Kopanke J, Lee J, Rodgers C, Reed K, Sherman TJ, Graham B, Cohnstaedt LW, Wilson WC, Stenglein M, Mayo C. Evaluating Temperature Effects on Bluetongue Virus Serotype 10 and 17 Coinfection in Culicoides sonorensis. Int J Mol Sci., 2024. Mar 6;25(5):3063. doi: 10.3390/ijms25053063. PMID: 38474308; PMCID: PMC10932384.
- Type:
Peer Reviewed Journal Articles
Status:
Published
Year Published:
2024
Citation:
Carpenter M, Kopanke J, Lee J, Rodgers C, Reed K, Sherman TJ, Graham B, Stenglein M, Mayo C., 2024. Assessing Reassortment between Bluetongue Virus Serotypes 10 and 17 at Different Coinfection Ratios in Culicoides sonorenesis. Viruses. 2024 Feb 2;16(2):240. doi: 10.3390/v16020240. PMID: 38400016; PMCID: PMC10893243
- Type:
Conference Papers and Presentations
Status:
Accepted
Year Published:
2024
Citation:
Carpenter M, Benavides Obon A, Kopanke J, Lee J, Reed K, Sherman T, Rodgers C, Stenglein M, Mayo C. In situ hybridization (RNAscope) detection of bluetongue virus serotypes 10 and 17 in experimentally co-infected Culicoides sonorensis; Poster presentation at CSU MIP Celebration; 2024 June; Fort Collins, CO.
- Type:
Conference Papers and Presentations
Status:
Accepted
Year Published:
2024
Citation:
Carpenter MJ, Kopanke JH, Rodgers C, Lee JS, Graham B, Stenglein M, Mayo, CE. Evaluating temperature effects on bluetongue virus serotype 10 and 17 coinfection in Culicoides sonorensis; Oral presentation at CRWAD; 2024 January; Chicago, IL.
- Type:
Conference Papers and Presentations
Status:
Accepted
Year Published:
2024
Citation:
Burton M, Carpenter M, Rodgers C, Torchetti M, Fox K, Sherman T, Mayo C. Initial detection and spread of bluetongue virus serotype 6 (BTV-6) in wild and domestic ruminants in Colorado. Oral presentation at the CRWAD; 2024 January; Chicago, IL.
- Type:
Conference Papers and Presentations
Status:
Accepted
Year Published:
2024
Citation:
Barbera C, Mayo C, Perkins TA, Rohr JR. A Field Survey of Larval Development Habitats of Culicoides Midges in Colorado Oral presentation at Society for Vector Ecology (SOVE) annual meeting, Fort Collins, CO September 2024
- Type:
Conference Papers and Presentations
Status:
Accepted
Year Published:
2024
Citation:
Barbera C, Mayo C, Rohr JR, Perkins TA. Modeling the Impact of Seasonal Temperature Variation on Culicoides Population Dynamics and BTV Transmission Poster presentation at Ecology and Evolution of Infectious Disease (EEID) conference, Palo Alto, CA June 2024.
- Type:
Conference Papers and Presentations
Status:
Accepted
Year Published:
2024
Citation:
Barbera C, Mayo C, Rohr JR, Perkins TA. Assessing the impact of Host Diversity on Transmission of Bluetongue Virus Oral Presentation at Biofrass student seminar, University of Notre Dame Department of Biological Sciences April 2024.
|
Progress 12/15/22 to 12/14/23
Outputs
Target Audience:The target audience of these studies include policy makers (national and regional), veterinary diagnosticians, livestock producers and veterinarians, and industry stakeholders (dairy, beef cattle, sheep, deer, and wildlife). We are engaging with leaders of Future Farmers of America (FFA) programs in our local school district in order to develop interactive field exercises that incorporate aspects of vector-borne disease and ruminant host species. The goal of this exercise is to integrate problem solving for understanding and preventing vector-borne diseases that affect the livestock industry. We developed and offered a course-based undergraduate research experience (CURE) at CSU in the Spring of 2022. The primary focus of the course content was reinforcing the collaboration and depth of projects (i.e. sequencing, fieldwork, modeling, in vivo work) we are currently conducting. In 2023, we were able to publish a manuscript based on this course. We have identified and assigned an individual to initiate a laboratory website so that the project and project team can publicize research findings and distribute educational materials. Team members will continue to be involved in developing and leading an annual "Genomics of Disease in Wildlife" workshop that was offered for the first time in June 2017 (Co-PI Stenglein is co-developer and co-lead). This workshop, which is aimed at fostering the adoption of genomics approaches by wildlife researchers, leverages existing NSF EEID funding (to PI Sue VandeWoude) and includes participation from many members of the team including the PI (Christie Mayo). Important aspects of reaching our target audience through our broader impact activities include reinforcing the study of a globally emerging arbovirus infections of livestock and wildlife, including the potential impact of climate change thereon. Changes/Problems:There are no significant changes or problems that have been encountered for the project. Due to the SARS-CoV-2 pandemic, research activities at Colorado State University were suspended in March and April of 2020. As principle investigators, we were advised to apply for a return to research review in May. Research activities returned to a level of 50%; however, the pandemic continued to have an impact on our overall performance during that time. In the interest of making the most efficient progress, we were able to identify the critical needs and expertise in order to continue the progress. We are remaining flexible based on the uncertainty of the pandemic and potential subsequent re-closures or reduction in research effort at the department or university level. Due to some restrictions for our in vivo work (objective 4), we have expanded our field surveillance to incorporate deer and cattle so that we can obtain the parameter estimates needed for modeling efforts. This would represent a substantial change in our in vivo work but expands our field surveillance (objective 6) and the data we gain from this opportunity allows us to understand infection dynamics in natural settings. What opportunities for training and professional development has the project provided?The project has served as a venue for the involvement and training/education of undergraduate, professional (DVM), producers, extension agents, graduate academic students, and postdoctoral scholars. The breadth of opportunities for these individuals included field, laboratory, and modeling studies. The work has continued to expand from temperature dependent reassortment studies in Culicoides sonorensis to in vivo and field studies this past summer. We have been working with livestock producers and wildlife agencies to expand and include free-ranging species field sites in addition to our domestic beef cattle and sheep sites. Laboratory studies have included virologic investigations, laboratory evaluation of vector midges, and immunologic investigation of data obtained in our in vivo studies. The professional development in our students has been rewarding as they learn from each other and their mentors in this multi-disciplinary program. The combination of skillsets and expertise have allowed us to develop and offer a Culicoides midge focused course. Specifically, we offerred a course-based undergraduate research experience (CURE) at CSU in the Spring of 2022. Since this time, we have published works focused on pedagogy and the impacts of this course on science based outcomes (Journal of Microbiology & Biology Education, 2023). The primary focus of the course content will be reinforcing the collaboration and depth of projects we are currently conducting in this project. Overall, we have also incorporated aspects of diversity, equity, and inclusion as we continue to foster these collaborations and promote members of our team. Additionally, our work has allowed us to develop international collaborations with the Pirbright institute. Most recently, we hosted a visiting student and scholar (Rhiannon Moody) and further developed her sequencing analysis. Our fieldwork has opened the doors to collaboration with Peru where we're actively in the process of writing manuscripts and grants for future funding. We have initiated conversations with Sam Wisely (University of Florida) as well to submit for a USDA Sustainable Agriculture Systems grant. How have the results been disseminated to communities of interest?We have continued to grow as a team of individuals dedicated to accomplishing our project goals in a collaborative space. We have also participated in several opportunities that support the broader impacts of this grant. An individual made me a customized Culicoides midge costume that I have worn proudly at the Open House on the Oval. The president of our university (Rick Miranda) appreciated the novelty of a midge giving out blue suckers to emphasize the impact of bluetongue transmission. I have also participated in two 4-H events and participated in some K-12 programs with Dr. Wade Ingle showcasing this work in his outreach program. Results of the laboratory studies presented in Objectives 1 and 2 have been published in peer reviewed journals. Results from objectives 1-4 have been presented to relevant professional and commodity groups including regional (County-based) livestock producer groups, the American Association of Veterinary Laboratory Diagnosticians, the Conference for Research Workers in Animal Disease (CRWAD), 4-H groups, genomics workshops, Society for Vector Ecology (SOVE), and European Congress of Entomology (ECE). We have been able to present in person and shift to the virtual platform for most of these opportunities and have remained actively engaged in discussions to promote website development and additional outreach. PI's and post docs have been interviewed for relevant podcasts (American Sheep Industry Association) in order to explain the effects of climate change on vector-borne disease and graduate students have been engaged in STEM, presenting our materials to elementary school children in Loveland. Graduate students have also expressed their creativity in creating and presenting a music video entitled: "Ram of Constant Sorrow" and "Culicoides by lady Baba." This platform was well-received by our department and succinctly explained studies outlined in objective 1 in a creative way. What do you plan to do during the next reporting period to accomplish the goals?Administrative: We will continue to build and refine our infrastructure for the primary teams focused on reassortment studies, fieldwork, in vivo studies, and modeling efforts. We have been successful in enhancing our communication by forming a monthly meeting and yearly retreats for all members of the team. This allows for us to build strong collaboration, share updates and data among groups. One of our Co-PI's from Notre Dame visited the Colorado State University campus for a sabbatical this year and this was an amazing opportunity to exchange data and enhance our modeling approaches. This has allowed several other Co-PI's to recognize the success of sabbaticals and we are encouraging this activity to focus on future grant writing. Scientific Metrics: We have successfully finalized a few objectives with published outcomes. Data from these objectives have been used to inform models for estimating the relationship between temperature, vertebrate/invertebrate interactions and the probability of transmission of reassortant viruses (objectives 5-6). We will continue to encourage all team members to present their work at national and international meetings. The results from the fieldwork in 2021-2023 will help to broaden our knowledge of risk factors of vector-borne diseases and is informing the planning for fieldwork in 2024. As outlined in our no-cost extension paperwork, we will conduct a targeted, final field collection this upcoming season that would satisfy the data collection needs described in our original proposal. Notably, we recently adopted a new sequencing workflow (SISPA/Oxford Nanopore application) that will streamline diagnostics and reduce costs (5 hours less run time and $66.07 cheaper per test) for livestock producers, thereby fostering greater incentive for testing and applied research. Greater testing incentive will also enhance surveillance efforts and contribute to the pool of available whole genome sequences which can be used for further evolutionary analysis. We are also using this time during the NCE to write and submit manuscripts. Broader Impacts: We will continue to update our laboratory website that allows the project team to publicize research findings and to distribute educational materials. Team members will continue developing and participating in the Genomics of Disease in Wildlife workshop that's held annually at CSU. We continue to grow as a team of individuals dedicated to accomplishing our project goals in the collaborative spaces of the VDL and Center for Vector-Borne Diseases (CVID). The resulting research supports the mission of our program by protecting the health of livestock and building strong relationships with ranchers and clients. An example of the level of investment our community has in our work; a colleague custom-made us a Culicoides midge (the invertebrate vector of BTV) costume that I proudly wear at different community events. This year, she made our group an additional sheep costume and we use these costumes to connect with students and the greater community. Simple things like costumes and blue suckers (for the blue tongue disease caused by BTV) bring enthusiasm to my trainees and team and creates a safe space where we can connect over humor and use fun to motivate learning. Further, our laboratory partners with Dr. Wade Ingle to share STEM with 4-H clubs and K-12 programs showcasing our broader impacts and outreach programs.
Impacts
What was accomplished under these goals?
Objective 1:The studies of objective 1 sought to characterize reassortment in colony raisedCulicoides sonorensis(also known as midges) that were coinfected with 2 different endemic strains of bluetongue virus (BTV-10 and BTV-17). It was found that while BTV replicated faster in midges at higher temperatures midge survival times were shorter. Results from representative plaques have revealed limited reassortment as most reads align to BTV-17. These findings were recently published in theInternational Journal of Molecular Sciences (2024)and suggest that parental strain fitness of the virus may play a role in reassortment outcomes. To further interrogate determinants of BTV reassortment in the midge, our next study investigated if different coinfection ratios between BTV-10 and BTV-17. Overall, reassortment events were limited and suggest that reassortant prevalence may be maximized upon the occurrence of reassortant genotypes that can outcompete the parental genotypes (publishedViruses, 2024). The limited reassortant events in the aforementioned studies, prompted the exploration of coinfection at the cellular level using in situ hybridization techniques. An RNAscope chromogenic platform was used to detect and distinguish segment 2 of BTV-10 and BTV-17 in histologic sections of coinfectedC. sonorensis. Observations of the mosaic patterns between BTV-10 and BTV-17, in which serotypes did not often overlap, suggest that coinfection at the cellular level may not be abundant with these two serotypes inC. sonorensis(publishedPathogens, 2023).Overall, BTV reassortment patterns and their biological consequences will add an important dimension to the modeling of viral expansion and evolution in the context of climate change.Objective 2:We developed two within-host models of bluetongue virus infection: one for midge vectors and one for mammalian hosts. The models shared some features, such as the depletion of target cells, an eclipse phase that permitted coinfection by two distinct strains, an infectious phase, and free virus for each infecting strain and all reassortant strains combined. The models also had some differences, including in their detail about tissue types and parameterization. The models were fitted to all pertinent data sources that could be identified. Net infectiousness was a focal outcome for both models. In both models, net infectiousness of reassortants was sensitive to the relative production of reassortants during cellular co-infection. An important difference between the models was that the timing of inoculations with distinct strains could be much closer in vertebrate hosts, resulting in a higher likelihood of reassortants arising. At the same time, vector infections are not self-limiting, providing more time for reassortants to grow to appreciable levels in longer-lived vectors. To place these findings in the context of epizootic transmission, we simulated an epizootic with an SEIR-SEI model, which quantified time-varying force of infection for both midges and vertebrate hosts. Force of infection of midges is relevant to reassortment, because it determines the frequency with which successive bloodmeals contain viruses of different strains that can result in reassortment. Similarly, the force of infection of vertebrates determines the rate at which successive infections occur within a short enough timeframe for reassortment to occur. In general, we found that more intense transmission increased the likelihood of reassortment, and that reassortants were generated more commonly in midges under higher transmission scenarios. Objective 3:We made progress on the analysis of data collected during summer 2020 to provide insight about the effect of cattle density on midge population density. This provides a basis for inferring whether transmission in this system is frequency- or density-dependent. Our progress on the analysis was to develop a hierarchical Bayesian model of these data, with the goal of accounting for interactions among variables and nested structure within the data set. We implemented this model in the rstan software package to enable good convergence of our estimates. This analysis pipeline is now complete, and we have run the analysis on a number of alternative models to assess robustness of our conclusions to alternative assumptions and model structures. In general, this analysis appears to provide support for the hypothesis that transmission is density-dependent in this system. That implies that larger farms are expected to support disproportionately larger midge populations and, consequently, be subject to disproportionately more intense midge-borne virus transmission.Objective 4:In this last year, we have concluded severalin vivostudies and successfully experimentally infected two disparate ruminant species using two distinct BTV serotypes. We were able to monitor these animals for signs of infection, and through intermittent blood draws, monitor the host response of these animals using a suite of virological and immunological techniques (published Virus Research, 2023). The remaining study cohorts will be published in the next quarter. Briefly taking our findings together, we have found that the CD8 T cell response, potentially driven proinflammatory cytokines (CXCL10 and IFN-gamma), is critical to control the infection after the previously described pan-lymphopenia. Interestingly, we have also identified that the species that was used to model cervid infections (Reeves' muntjac) had a much more subdued response to the infections, with one exception of an animal that acquired a secondary bacterial lung infection and had to be humanely euthanized. Secondary infections have been speculated (and published) as contributing factors to death within BTV- infected free-ranging populations.Objective 5:We derived a stage-structured simulation model with temperature-dependent life-history traits of the vector, based on laboratory estimates of stage-specific thermal performance, to simulate seasonalCulicoidesabundances. We then used an SEI-SEIR model, incorporating viral thermal responses, to simulate the impact of inter-annual seasonal variation on relevant epidemiological outcomes. Specifically, we simulated across a range of annual mean temperatures, seasonal variation, and temperature at the time of pathogen introduction. We found that transmission depends jointly on annual mean temperatures and seasonal temperature variation, with the largest outbreaks caused by high mean temperatures when seasonal variation is low, or by intermediate mean temperatures when variation is high. Additionally, we found that higher temperature at time of pathogen introduction led to more rapid epidemics. As bluetongue range has been infringing on previously unaffected areas, our results could be used in future work to identify areas with seasonal temperature profiles that might result in elevated risk for bluetongue outbreaks under present-day and future climate-change scenarios. In particular, cooler areas may be more at risk for outbreaks as seasonal temperature variation increases and transmission seasons shift to earlier in the year.Objective 6:The 2023 vertebrate surveillance work was completed and consisted of both longitudinal and cross-sectional sampling strategies. The longitudinal sampling consisted of four cattle sites and four sheep sites were sampled monthly from July through December. Overall prevalence across all sites and species for BTV via serology was 46% and RNA prevalence was 5%. This was a decrease compared to what we had observed previously in 2022 which was 57% and 23% for seroprevalence and RNA prevalence, respectively. Cross-sectional sampling took place across 29 different sheep and cattle sites and analysis is still in progress for those samples. Model development and data analysis are ongoing for all field seasons (2021-2023).
Publications
- Type:
Conference Papers and Presentations
Status:
Accepted
Year Published:
2023
Citation:
Barbera C, Bill H, Perkins, A, Modeling the Impact of Seasonal Temperature Variation on Culicoides Population Dynamics and Bluetongue Virus Transmission, Epidemics - 9th International conference on infectious disease dynamics, Bologna, Italy, November, 2023, Poster presentation.
- Type:
Conference Papers and Presentations
Status:
Accepted
Year Published:
2023
Citation:
Yu G, Burton M, Barbera C, Mayo C, Perkins A, A Spatial, Agent-based Model to Explore Mechanisms of Bluetongue Virus Persistence at the Interface of Domestic and Wildlife Animal Populations, Epidemics - 9th International conference on infectious disease dynamics, Bologna, Italy, November, 2023, Poster presentation.
- Type:
Conference Papers and Presentations
Status:
Accepted
Year Published:
2023
Citation:
Westrich, J., McNulty E, Carpenter M, Burton M, Reed K, Nalls A, Sandoval A, Mayo C, Mathiason C, Longitudinal viral progression and immunological responses to Bluetongue virus in experimentally infected ruminants. Conference of Reseach Workers in Animal Diseases (CRWAD), Chicago, IL, January 2023. Oral Presentation
- Type:
Conference Papers and Presentations
Status:
Accepted
Year Published:
2023
Citation:
Westrich, J., McNulty E, Carpenter M, Burton M, Reed K, Nalls A, Sandoval A, Mayo C, Mathiason C, Longitudinal viral progression and immunological responses to Bluetongue virus in experimentally infected ruminants. TAMASAG, Laporte, Colorado, April, 2023, Oral Presentation
- Type:
Other
Status:
Published
Year Published:
2023
Citation:
Podcast: Sheep Stuff Ewe Should Know by Ryan Mahoney/Dan Macon/Dr. Roselle Busch. August 25, 2023 episode #137 Bluetongue Research Update (US) Interview with Dr. Christie Mayo and Dr. Molly Carpenter.
- Type:
Journal Articles
Status:
Published
Year Published:
2023
Citation:
Carpenter MJ, Rodgers CR, Torchetti MK, Fox KA, Burton M, Sherman TJ, Mayo CE. Recovery of multireassortant bluetongue virus serotype 6 sequences from a mule deer (Odocoileus hemionus) and Dorset sheep (Ovis aries) in Colorado. Vet Microbiol. 2024 Feb;289:109944. doi: 10.1016/j.vetmic.2023.109944. Epub 2023 Dec 12. PMID: 38141398.
- Type:
Journal Articles
Status:
Published
Year Published:
2023
Citation:
Sparrer MN, Hodges NF, Ragan I, Yamashita T, Reed KJ, Sherman TJ, Mayer T, Maichak C, Adney DR, Carpenter M, Webb TL, Mayo C. SARS-CoV-2 surveillance in a veterinary health system provides insight into transmission risks. J Am Vet Med Assoc. 2023 Oct 13;262(1):93-99. doi: 10.2460/javma.23.05.0229. PMID: 38103381.
- Type:
Journal Articles
Status:
Published
Year Published:
2023
Citation:
Carpenter M, Benavides Obon A, Kopanke J, Lee J, Reed K, Sherman T, Rodgers C, Stenglein M, McDermott E, Mayo C. In Situ Hybridization (RNAscope) Detection of Bluetongue Virus Serotypes 10 and 17 in Experimentally Co-Infected Culicoides sonorensis. Pathogens. 2023 Sep 30;12(10):1207. doi: 10.3390/pathogens12101207. PMID: 37887723; PMCID: PMC10609982.
- Type:
Journal Articles
Status:
Published
Year Published:
2023
Citation:
Sparrer MN, Hodges NF, Sherman T, VandeWoude S, Bosco-Lauth AM, Mayo CE. Role of Spillover and Spillback in SARS-CoV-2 Transmission and the Importance of One Health in Understanding the Dynamics of the COVID-19 Pandemic. J Clin Microbiol. 2023 Jul 20;61(7):e0161022. doi: 10.1128/jcm.01610-22. Epub 2023 Apr 26. PMID: 37098970; PMCID: PMC10358177.
- Type:
Journal Articles
Status:
Published
Year Published:
2023
Citation:
Simpson KM, Depenbrock SM, Oman RE, Mayo CE. When dermatologic diseases are devastating: differentiating common endemic conditions in the United States from sheep and goat pox. J Am Vet Med Assoc. 2023 Mar 25;261(S1):S95-S102. doi: 10.2460/javma.22.12.0553. PMID: 36944220.
- Type:
Journal Articles
Status:
Published
Year Published:
2023
Citation:
Bertram MR, Rodgers C, Reed K, Velazquez-Salinas L, Pelzel-McCluskey A, Mayo C, Rodriguez L. Vesicular stomatitis Indiana virus near-full-length genome sequences reveal low genetic diversity during the 2019 outbreak in Colorado, USA. Front Vet Sci. 2023 Feb 14;10:1110483. doi: 10.3389/fvets.2023.1110483. PMID: 36865437; PMCID: PMC9971496.
- Type:
Journal Articles
Status:
Published
Year Published:
2023
Citation:
Westrich, J, McNulty, E, Carpenter, M, Burton, M, Reed, K, Nalls, A, Sandoval, A, Mayo, C, Mathiason, C, (2023). Monitoring longitudinal immunological responses to bluetongue virus 17 in experimentally infected sheep. Virus Research, https://doi.org/10.1016/j.virusres.2023.199246.
- Type:
Journal Articles
Status:
Published
Year Published:
2023
Citation:
Hudson A, McGregor B, Shults P, England M, Silbermagel C, Mayo C, Carpenter M, Sherman T, Cohnstaedt L, (2023). Culicoides-borne Orbivirus epidemiology in a changing climate. Journal of Medical Entomology, https://doi.org/10.1093/jme/tjad098
- Type:
Journal Articles
Status:
Published
Year Published:
2023
Citation:
Borlee, G, Kinkel, T, Broeckling, B, Borlee, B, Mayo, C, Mehaffy, C, (2023). Upper-level inter-disciplinary microbiology CUREs increase student's scientific self-efficacy, scientific identity, and self-assessed skills. Journal of Microbiology & Biology Education, https://doi.org/10.1128/jmbe.00140-23
- Type:
Conference Papers and Presentations
Status:
Accepted
Year Published:
2023
Citation:
Zook, S, Wolfe, B, Wittemeyer, G, Alder, J, Mayo, C, Epizootic hemorrhagic disease virus (EHDV) prevalence in white-tailed and mule deer in northeastern Colorado. National Veterinary Scholars Symposium, San Juan, Puerto Rico, August, 2023. Poster
- Type:
Conference Papers and Presentations
Status:
Accepted
Year Published:
2023
Citation:
Hilty, S, Burton, M, Simpson, K, Mayo, C, Examining seroprevalence of bluetongue virus and epizootic hemorrhagic disease virus across wild and domestic ruminants in Northern Colorado, National Veterinary Scholars Symposium, San Juan, Puerto Rico, August, 2023. Poster
- Type:
Conference Papers and Presentations
Status:
Accepted
Year Published:
2023
Citation:
Dunham, T, Sherman, T, Lee, J, Wilson, W, Cohnstaedt, L, Anderson, T, Reed, K, Stenglein, M, Mayo, C, Bluetongue virus's search for more space and time in the sequencing archive, Rocky Mountain Virology Conference, Pingree Park, CO, October, 2023. Poster
- Type:
Conference Papers and Presentations
Status:
Accepted
Year Published:
2023
Citation:
Carpenter, M, Kopanke, J, Rodgers, C, Lee, J, Graham, B, Stenglein, M, Mayo, C, Characterizing the effect of coinfection ratios on bluetongue virus reassortment in Culicoides sonorensis, European Congress of Entomology, Crete, Greece, 2023. Oral Presentation
- Type:
Conference Papers and Presentations
Status:
Accepted
Year Published:
2023
Citation:
Burton, M, Reed, K, Korte, C, Wolbers, T, Mayo, C, Prevalence and seasonality of Bluetongue Virus on the Front Range of Colorado in domestic ruminants during 2021, American Association of Veterinary Laboratory Diagnosticians, Washington, DC, October, 2023. Oral Presentation
- Type:
Conference Papers and Presentations
Status:
Accepted
Year Published:
2023
Citation:
Burton, M, Reed, K, Korte, C, Wolbers, T, Mayo, C, Prevalence and seasonality of Bluetongue Virus on the Front Range of Colorado in domestic ruminants during 2021, Rocky Mountain Virology Conference, September, 2023. Oral Presentation
- Type:
Conference Papers and Presentations
Status:
Accepted
Year Published:
2023
Citation:
Westrich, J, McNulty, E, Carpenter, M, Burton, M, Sandoval, A, Mayo, C, Mathiason, C, Longitudinal viral progression and immunological responses to Bluetongue virus in experimentally infected ruminants, Rocky Mountain Virology Conference, September, 2023. Oral Presentation
- Type:
Conference Papers and Presentations
Status:
Accepted
Year Published:
2023
Citation:
Carpenter, M, Kopanke, J, Rodgers, C, Lee, J, Reed, K, Graham, B, Stenglein, M, Mayo, C, Characterizing the effect of coinfection ratios on bluetongue virus reassortment in Culicoides sonorensis, American Society of Virology, Athens, GA, June, 2023, Poster Presentation
- Type:
Conference Papers and Presentations
Status:
Accepted
Year Published:
2023
Citation:
Sparrer, M, Hodges, N, Yamashita, T, Ragan,I, Webb,T, Mayer, T, Carpenter, M, Mathiason, C, Mayo, C, Evaluation of transmission potential of SARS-CoV-2 through surveillance of companion and exotic animals, CVMBS Research Day, Fort Collins, CO, January, 2023. Poster
- Type:
Conference Papers and Presentations
Status:
Accepted
Year Published:
2023
Citation:
Burton, M, Reed, K, Korte, C, Wolbers, T, Mayo, C, Prevalence and seasonality of Bluetongue Virus on the Front Range of Colorado in domestic ruminants during 2021, Conference for research workers in animal disease, Chicago, IL, January, 2023. Oral Presentation
- Type:
Conference Papers and Presentations
Status:
Accepted
Year Published:
2023
Citation:
Carpenter, M, Kopanke, J, Rodgers, C, Lee, J, Graham, B, Stenglein, M, Mayo, C, Characterizing the effect of coinfection ratios on bluetongue virus reassortment in Culicoides sonorensis, Conference for research workers in animal disease, Chicago, IL, January, 2023. Oral Presentation
- Type:
Conference Papers and Presentations
Status:
Accepted
Year Published:
2023
Citation:
Perkins A, Cavany S, Westrich J, Sherman T, Carpenter M, Stenglein M, Mathiason C, Mayo C, Bug or Beast: Where are Vector-transmitted viruses more likely to reassort?, Ecology and evolution of infectious diseases conference, University Park, PA, May, 2023, Poster presentation
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Progress 12/15/21 to 12/14/22
Outputs
Target Audience:The target audience of these studies include policy makers (national and regional), veterinary diagnosticians, livestock producers and veterinarians, and industry stakeholders (dairy, beef cattle, sheep, deer, and wildlife). We are engaging with leaders of 4-H and Future Farmers of America (FFA) programs in our local school district in order to develop interactive field exercises that incorporate aspects of vector-borne disease and ruminant host species. The goal of this exercise is to integrate problem solving for understanding and preventing vector-borne diseases that affect the livestock industry. We are also in discussions with the department microbiology, immunology, and pathology to further develop and offer a course-based undergraduate research experience (CURE) at CSU in 2024. The course offered in 2022 was extremely successful and the primary focus of the course content wasreinforcing the collaboration and depth of projects (i.e. sequencing, fieldwork, modeling, in vivo work) we are currently conducting in the laboratory. We have launched the Mayo laboratory website so that the project and project teamscan publicize research findings and distribute educational materials. Team members will continue to be involved in developing and leading an annual "Genomics of Disease in Wildlife" workshop that was offered for the first time in June 2017 (Co-PI Stenglein is co-developer and co-lead). This workshop, which is aimed at fostering the adoption of genomics approaches by wildlife researchers, leverages existing NSF EEID funding (to PI Sue VandeWoude) and includes participation from many members of the team including the PI (Christie Mayo). Important aspects of reaching our target audience through our broader impact activities include reinforcing the study of a globally emerging arbovirus infections of livestock and wildlife, including the potential impact of climate change thereon. Changes/Problems:There are no significant changes or problems that have been encountered for the project. Due to the SARS-CoV-2 pandemic, research activities at Colorado State University were suspended in March and April of 2020. As principle investigators, we were advised to apply for a return to research review in May. Research activities returned at a level of 50%; however, the pandemic contined to have an impact on our overall performance the last 2 years. In the interest of making the most efficient progress, we were able to identify the critical needs and expertise in order to continue the progress. We are remaining flexible based on the uncertainty of the pandemic and potential subsequent re-closures or reduction in research effort at the department or university level. Due to some restrictions for our in vivo work (objective 4), we have expanded our field surveillance to incorporatedeer and cattle so that we can obtain the parameter estimates needed for modeling efforts. This would represent a substantial change in our in vivo work but expands our field surveillance (objective 6) and the data we gain from this opportunity allows us to understand infection dynamics in natural settings. We plan to apply for a no cost extension (NCE) this year and we have reached out to our fiscal officers and sponsored programs office to initiate this process. What opportunities for training and professional development has the project provided?The project has served as a venue for the involvement and training/education of undergraduate, professional (DVM), producers, extension agents, graduate academic students, and postdoctoral scholars. The breadth of opportunities for these individuals included field, laboratory, and modeling studies. The work has continued to expand from temperature dependent reassortment studies in Culicoides sonorensis to in vivo and field studies this past summer. We have been working with livestock producers and wildlife agencies to expand and include free-ranging species field sites in addition to our domestic beef cattle and sheep sites. Laboratory studies have included virologic investigations, laboratory evaluation of vector midges, and immunologic investigation of data obtained in our in vivo studies. The professional development in our students has been rewarding as they learn from each other and their mentors in this multi-disciplinary program. The combination of skillsets and expertise have allowed us to initiate discussions with the department microbiology, immunology, and pathology to develop and offer a Culicoides midge focused course. Specifically, we offered a course-based undergraduate researchexperience (CURE) at CSU during the Spring of 2022. The primary focus of the course content was reinforcing the collaboration and depth of projects we are currently conducting in this project. Overall, we have also incorporated aspects of diversity, equity, and inclusion as we continue to foster these collaborations and promote members of our team. How have the results been disseminated to communities of interest?We have continued to grow as a team of individuals dedicated to accomplishing our project goals in a collaborative space. We have also participated in several opportunities that support the broader impacts of this grant. An individual made me a customizedCulicoidesmidge costume that I have worn proudly at the CSU Open House. The president of our university(Rick Miranda) appreciated the novelty of a midge giving out blue suckers to emphasize the impact of bluetongue transmission. I have also participated in two 4-H events and participated in some K-12 programs with Dr. Wade Ingle showcasing this work in his outreach program. Results of the laboratory studies presented in Objectives 1 and 2have been published in peer reviewed journals. Results from objectives 1-4 have been presented to relevant professional and commodity groups including regional (County-based) livestock producer groups, the American Association of Veterinary Laboratory Diagnosticians, the Conference for Research Workers in Animal Disease (CRWAD), 4-H groups, genomics workshops, and Society for Vector Ecology (SOVE). We have been able to present in person andshift to the virtual platform for most of these opportunities and have remained actively engaged in discussions to promote website development and additional outreach. PI's have been interviewed for relevant podcasts (VetCAST) in order to explain the effects of climate change on vector-borne disease and graduate students have been engaged in STEM, presenting our materials to elementary school children in Loveland. Graduate students have also expressed their creativity in creating and presenting a music video entitled: "Ram of Constant Sorrow" and "Culicoides by Lady Baba." This platform was well-received by our department and succinctly explained studies outlined in objective 1 in a creative way. What do you plan to do during the next reporting period to accomplish the goals?Administrative: Within the last year, we have built and refined our infrastructure for the primary teams focused on reassortment studies, fieldwork, in vivo studies, and modeling efforts. We have been successful in enhancing our communication by forming a monthly meeting for all members of the team. This allows for us to share updates and data among groups. We have also established and will continue to have a retreat and meeting among Co-PD's. This meeting has allowed us to make decisions with regards to resources management and team support. We will be transitioning our organizational structure and designing a smartsheets platform for notifying individuals about published works, additional grant opportunities, and overall success of each group. We built a similar platform for reporting and communicating about the progress report and this seemed to work nicely for timely reporting in such a large group. One of our Co-PI's from Notre Dame is visiting for a sabbatical this year and we anticipate this will be a good opportunity to exchange data and enhance our modeling approaches. Scientific Metrics: Our goal is to complete the remaining laboratory studies and data analysis outlined in objectives 1-4and finalize manuscripts for publication. Data from the objectives have been used to informmodels for estimating the relationship between temperature and the probability of transmission of reassortant viruses. The postdoctoral scholar on this project has presentedtheir work at the annual EEID meeting. Additionally, we plan to expand the model to other viruses and explore phylogenetic methods for estimating bluetongue virus reassortment rates. The results from the fieldwork in 2020-2022 will help to broaden our knowledge of risk factors of vector-borne diseases and is informing the planning for fieldwork in 2023. In particular, data collection on a diverse set of field sites provides information on the role of the domesticwildlife-human interface in the spread of vector-borne disease. Additionally, fine-scale temperature and humidity data will help shed light on the relationship between climate and vector-borne disease risk, which can help strengthen predictions in the face of climate change. We will continue to expand our in vivo work as proposed in objective 4. We are also closely aligned in evaluating the budget and adequate support for these studies as maintaining the required secured indoor facilities and personnel to conduct in vivo studies in the native host (sheep, deer) are considerable. University per diems have increased since the submission and award of this proposal and we have been notified of another planned increase in 2023. We will continue to submit grants seeking additional funds to support this work and will resubmit an NIH R03 to develop novel technology using BTV and sheep as a model system. PI's at CSU and Notre Dame will continue to ensure laboratory and fieldwork (Aims 1-4) are aligned with models developed in Aims 5&6 and we will use preliminary data from our animal surveillance to plan and expand our surveillance efforts in the coming year. Broader Impacts: We will continue to update ourlaboratory website that allows the project team to publicize research findings and to distribute educational materials. Team members will continue developing and participating in the Genomics of Disease in Wildlife workshop that's held annually at CSU. We learned so much from our experience offering a course-based undergraduate research experience (CURE) at CSU in the Spring of 2022 and areengaged in the process for developing additional CURES courses. This course isan ideal platform to demonstrate collaboration and provide education about the importance of vector-borne diseases in ruminant communities. Additionally, we have engaged the associate director for the department of outreach and engagement at CSU. He is including orbivirus educational materials in his infectious disease unit and will be visiting with K12 programs to help us with outreach in these communities.
Impacts
What was accomplished under these goals?
Objective 1: Our proposed work uses bluetongue virus (BTV), epizootic hemorrhagic disease virus, culicoides cell culture, and colony raised Culicoides sonorensis (also known as midges) as our backbone for understanding these relationships.Recent studies have sought to characterize the frequency of reassortment between 2 different endemic strains (BTV-10 and BTV-17). These strains have been isolated in western United States for decades and we were able to demonstrate infections of C.sonorensis with each individual viral strain. The insects were incubated at the same temperatures of our previous studies (20C, 25C, 30C). Results also indicate that midges maintained at 30C demonstrate productive virogenesis earlier in infection (day 3) than midges held at cooler temperatures. In order to evaluate reassortment, coinfected midges collected and processed for BTV plaque-isolation for future amplicon assays. Preliminary results from a single time point and representative plaques have revealed limited reassortment as most reads align to BTV-17. However, RNA was extracted directly from coinfected midges and results of next generation sequencing revealed reads aligning to both BTV-10 and BTV-17 at 25C. While this does not demonstrate reassortment, we identified there are reads aligning to both virus populations in midges at 25C. Sequencing efforts are currently underway to characterize additional plaques and RNA extracted directly from BTV-10 and BTV-17 coinfected midges on additional time points throughout the study so that we can examine the virus populations thoroughly. To further interrogate determinants of BTV reassortment in the midge, we investigated if different coinfection ratios between BTV-10 and BTV-17. Midges were fed blood containing BTV-10, BTV-17, or a combination of BTV-10: BTV-17 atdifferent ratios. Midges were collected for pan BTV and a housekeeping gene qRT-PCR. Midges werealso processed for BTV plaque-isolation and genotypes ofplaques were determined bynext generation sequencing. Plaque genotyping indicated that most plaques fully aligned with one of the parental strains. However, there was reassortment evident in a pool of midges coinfected with BTV-10: BTV-17 at a particular ratio. Additional individual propagated plaques are currently being sequenced to assess for reassortment patterns and in situ hybridization is being explored to further characterize coinfection at the cellular level. Overall, BTV reassortment patterns and their biological consequences will add an important dimension to the modeling of viral expansion and evolution in the context of climate changeObjective 2: Modeling efforts (outlined in Objective 2) were published earlier this year where we developed three mathematical models for within-midge BTV dynamics. These models allowed us to explore the conditions leading to the emergence of reassortant viruses. Our results suggest viruses need to arrive close together in time and that viruses must have similar replication rates for reassortment to be non-negligible. Examination of the full model, which permits cellular co-infection and reassortment, shows that small differences in fitness of the two infecting strains can have a large impact on the frequency with which reassortant virions are observed. These findings are consistent with experimental co-infection studies with BTV strains of different relative fitnesses that did not produce reassortant progeny. This work is also informing the importance of ongoing studies in Objective 1.Objective 3: Field work was conducted from this summersurveying field sites to gain a better understanding of which environments are suitable for development of the Culicoides vector. Potential larval substrate was collected across a range of field sites in Northern Colorado, including livestock operations, rangeland farms, natural spaces including a state park and a wildlife refuge, and research facilities with multiple host species present. Any moist substrate (e.g., puddles, agricultural waste ponds, streams, ponds, tire ruts, water troughs) was identified as a potential larval habitat, and substrates from particular sites were repeatedly sampled every two weeks for the duration of the summer, provided moisture remained in each spot. Samples were brought back to the insectary and monitored over a period of 11 weeks for emergence of adult midges, which would indicate suitable habitat at that particular sampling location. Emergence was compared between site types (eg livestock operation, natural space, etc) and substrate types (e.g.,puddle, stream, etc.) to identify which locations support vector populations. Formal analyses are underway on emergence numbers between site types using regression models, taking into account substrate characteristics such as pH, moisture content, and organic content, to identify associations between vector abundance and site type and substrate type. We are processing data from the 2021 field collections, aimed at studying the relationship between host species diversity and prevalence of bluetongue. Analysis on data studying the impact of host density on Culicoides abundance has continued, using hierarchical Bayesian models.Objective 4: Our in vivo studies have expanded on our previous findings from our cohorts of sheep inoculated with BTV-17. We have successfully expanded our investigations into the pathogenesis and immunological response in sheep to BTV by evaluating a separate serotype, BTV-10. Interestingly, we have found that the virological response (detectable BTV RNA in the peripheral blood) was detected early and observed for the remainder of the time course, as was observed in BTV-17. Additionally, we observed the pan lymphopenia that occurs roughly 8 DPI and rebounds at about 14 DPI remaining elevated until the end of the study. We also observed increases in novel proinflammatory cytokines, immediately before the rebounding of the immune population. To further our understanding of mechanisms of BTV disease in the native hosts, we also evaluated pathogenesis and immunological response of BTV-10 in muntjac deer, an effective model for disease in cervid species. Given that muntjac have never before been evaluated for BTV disease, we performed studies in this species to ensure disease progression.Interestingly, whereas the sheep-maintained infection throughout the time course, the detection of virus in the peripheral blood of the muntjac began to diminish quickly, with 2 animals showing no detectable virus in the blood at the end of study. Interestingly, the lung and splenic tissues evaluated for BTV viral RNA genomes in muntjac showed robust detection in all infected animals, including the animals that no long had peripheral detection. Lastly, all the immunological parameters showed a more subdued response as compared to the sheep studies. From these studies we have two manuscripts in preparation to report on these studies that will be submitted during this year.Objective 5 & 6: Techniques and data generated from work in Objectives 1-4 will be informative to the models created in Aims 5 & 6. Dr. Alex Perkins (Co-PI) will be taking his sabbatical at CSU during the Spring of 2023 and this was strategically planned to incorporate data into the ecological model development. With regards to Objective 6, field studies have identified a shift in circulating serotypes which included a previously undetected serotype in the area. The work from 2021 was used to inform the 2022 field season (May-December) in which another 8 field sites (4 cattle, 4 sheep) were sampled from on a monthly basis. Additionally, a cross-sectional sampling was conducted utilizing sampling from local producer's animals, as well as those at meat processing facilities, for an additional 15 sites including bison. Model development and data analysis are ongoing for both the 2021 and 2022 field seasons.
Publications
- Type:
Journal Articles
Status:
Published
Year Published:
2022
Citation:
Nehring M, Pugh S, Dihle T, Gallichotte E, Nett T, Weber E, Mayo C, Lynne L, Ebel G, Fosdick B, Vandewoude S, Laboratory based SARS-CoV-2 (2022). Receptor Binding Domain serologic assays perform with equivalent sensitivity and specificity to commercial FDA-EUA approved tests, Viruses 15(1), 106; https://doi.org/10.3390/v15010106
- Type:
Journal Articles
Status:
Published
Year Published:
2022
Citation:
Cavany S, Barbera C, Carpenter M, Rodgers C, Sherman T, Stenglein M, Mayo C, Perkins A, (2022). Modeling cellular co-infection and reassortment of bluetongue virus in Culicoides midges, Virus Evolution, Volume 8, Issue 2, veac094, https://doi.org/10.1093/ve/veac094
- Type:
Journal Articles
Status:
Published
Year Published:
2022
Citation:
Kopanke J.H., Carpenter M, Lee J, Reed K, Rodgers C, Burton M, Lovett K, Westrich J, McNulty E, McDermott E, Barbera C, Cavany S, Rohr J, Perkins T.A., Mathiason C.K., Stenglein M, Mayo C (2022). Bluetongue Research at a Crossroads: Modern Genomics Tools Can Pave the Way to New Insights. Annual Reviews of Animal Biosciences.10(1):303-324. doi: 10.1146/annurev-animal-051721-023724
- Type:
Conference Papers and Presentations
Status:
Published
Year Published:
2022
Citation:
2022: Mayo C, Epidemiology of bluetongue virus in the United States: implications for arbovirology at the wildlife-livestock interface, BSL-3 training/transboundary animal diseases training as part of National Bio and Agro-Defense Facility (NBAF), Manhattan, KS, June. Virtual Oral Presentation.
- Type:
Conference Papers and Presentations
Status:
Published
Year Published:
2022
Citation:
2022: Mayo C, Developing Diagnostics tests, biology of vector-borne disease course, Moscow, ID, June, 2022. Oral Presentatio
- Type:
Conference Papers and Presentations
Status:
Accepted
Year Published:
2022
Citation:
Mayo, C, Sustainable one health surveillance approaches and transmission dynamics in domestic, farmed, and wild animals, IDRRN research retreat, August, 2022. Oral Presentation.
- Type:
Conference Papers and Presentations
Status:
Accepted
Year Published:
2022
Citation:
Carpenter, MJ, Kopanke, JH, Rodgers C., Lee, JS, Stenglein M, Mayo, C, Evaluating Bluetongue virus reassortment in Culicoides sonorensis, CVMBS Research Day, Fort Collins, CO, January, 2022. Poster
- Type:
Conference Papers and Presentations
Status:
Accepted
Year Published:
2022
Citation:
Westrich, J, McNulty, E, Carpenter, M, Burton, M, Sandoval, A, Mayo, C, Mathiason, C, Determining longitudinal viral progression and immunological responses to Bluetongue Virus in experimentally infected ruminants, Rocky Mountain Virology Conference. Oral Presentati
- Type:
Conference Papers and Presentations
Status:
Accepted
Year Published:
2022
Citation:
Sparrer, M, Hodges, N, Yamashita, T, Ragan,I, Webb,T, Mayer, T, Carpenter, M, Mathiason, C, Mayo, C, Evaluation of transmission potential of SARS-CoV-2 through surveillance of companion and exotic animals, National Veterinary Scholars Symposium, August 2022. Poster
- Type:
Conference Papers and Presentations
Status:
Accepted
Year Published:
2022
Citation:
Hodges, N, Ragan, I, Webb, T, Adney, D, Mayer, T, Sherman, T., Carpenter, M, Mayo, C, One Health: Veterinary laboratories uniquely prepared to perform human diagnostic testing during SARS-CoV-2 pandemic, American Association of Veterinary Laboratory Diagnosticians Annual Conference, October 2022. Poster
- Type:
Conference Papers and Presentations
Status:
Accepted
Year Published:
2022
Citation:
Sparrer, M, Hodges, N, Yamashita, T, Ragan,I, Webb,T, Mayer, T, Carpenter, M, Mathiason, C, Mayo, C, Evaluation of transmission potential of SARS-CoV-2 through surveillance of companion and exotic animals, American Association of Veterinary Laboratory Diagnosticians Annual Conference, October 2022. Poster, 1st place poster award
- Type:
Conference Papers and Presentations
Status:
Accepted
Year Published:
2022
Citation:
Carpenter MJ, Kopanke J, Rodgers C, Lee JS, Stenglein M, Mayo CE. Characterizing determinants of reassortment of bluetongue virus in Culicoides sonorensis. Poster presentation at the Association for Clinical and Translational Science, April, 2021. Virtual Poster Presentatio
|
Progress 12/15/20 to 12/14/21
Outputs
Target Audience:The target audience of these studies include policy makers (national and regional), veterinary diagnosticians, livestock producers and veterinarians, and industry stakeholders (dairy, beef cattle, sheep, deer, and wildlife). We are engaging with leaders of Future Farmers of America (FFA) programs in our local school district in order to develop interactive field exercises that incorporate aspects of vector-borne disease and ruminant host species. The goal of this exercise is to integrate problem solving for understanding and preventing vector-borne diseases that affect the livestock industry. We are also in discussions with the department microbiology, immunology, and pathology to develop and offer a course-based undergraduate research experience (CURE) at CSU in the Spring of 2022. The primary focus of the course content will be reinforcing the collaboration and depth of projects (i.e. sequencing, fieldwork, modeling, in vivo work) we are currently conducting. We have identified and assigned an individual to initiate a laboratory website so that the project and project team can publicize research findings and distribute educational materials. Team members will continue to be involved in developing and leading an annual "Genomics of Disease in Wildlife" workshop that was offered for the first time in June 2017 (Co-PI Stenglein is co-developer and co-lead). This workshop, which is aimed at fostering the adoption of genomics approaches by wildlife researchers, leverages existing NSF EEID funding (to PI Sue VandeWoude) and includes participation from many members of the team including the PI (Christie Mayo). Important aspects of reaching our target audience through our broader impact activities include reinforcing the study of a globally emerging arbovirus infections of livestock and wildlife, including the potential impact of climate change thereon. Changes/Problems:There are no significant changes or problems that have been encountered for the project. Due to the SARS-CoV-2 pandemic, research activities at Colorado State University were suspended in March and April of 2020. As principle investigators, we were advised to apply for a return to research review in May. Research activities returned at a level of 50%; however, the pandemic contined to have an impact on our overall performance the last 2 years. In the interest of making the most efficient progress, we were able to identify the critical needs and expertise in order to continue the progress. We are remaining flexible based on the uncertainty of the pandemic and potential subsequent re-closures or reduction in research effort at the department or university level. What opportunities for training and professional development has the project provided?The project has served as a venue for the involvement and training/education of undergraduate, professional (DVM), producers, extension agents, graduate academic students, and postdoctoral scholars. The breadth of opportunities for these individuals included field, laboratory, and modeling studies. The work has continued to expand from temperature dependent reassortment studies in Culicoides sonorensis to in vivo and field studies this past summer. We have been working with livestock producers and wildlife agencies to expand and include free-ranging species field sites in addition to our domestic beef cattle and sheep sites. Laboratory studies have included virologic investigations, laboratory evaluation of vector midges, and immunologic investigation of data obtained in our in vivo studies. The professional development in our students has been rewarding as they learn from each other and their mentors in this multi-disciplinary program. The combination of skillsets and expertise have allowed us to initiate discussions with the department microbiology, immunology, and pathology to develop and offer a Culicoides midge focused course. Specifically, we are offering a course-based undergraduate research experience (CURE) at CSU in the Spring of 2022 (currently underway). The primary focus of the course content will be reinforcing the collaboration and depth of projects we are currently conducting in this project. Overall, we have also incorporated aspects of diversity, equity, and inclusion as we continue to foster these collaborations and promote members of our team. How have the results been disseminated to communities of interest?Results of the laboratory studies presented in Objective 1 have been published in peer reviewed journals. Results from objectives 1-4 have been presented to relevant professional and commodity groups including regional (County-based) livestock producer groups, the American Association of Veterinary Laboratory Diagnosticians, the Conference for Research Workers in Animal Disease (CRWAD), and Society for Vector Ecology (SOVE). We have been able to shift to the virtual platform for most of these opportunities and have remained actively engaged in discussions to promote website development and additional outreach. PI's have been interviewed for relevant podcasts (VetCAST) in order to explain the effects of climate change on vector-borne disease and graduate students have been engaged in STEM, presenting our materials to elementary school children in Boulder. Graduate students have also expressed their creativity in creating and presenting a music video entitled: "Ram of Constant Sorrow." This platform was well-received by our department and succinctly explained studies outlined in objective 1 in a creative way. What do you plan to do during the next reporting period to accomplish the goals?Administrative: Within the last year, we have built and refined our infrastructure for the primary teams focused on reassortment studies, fieldwork, in vivo studies, and modeling efforts. We have been successful in enhancing our communication by forming a monthly meeting for all members of the team. This allows for us to share updates and data among groups. We have also established and will continue to have a meeting among Co-PD's. This meeting has allowed us to make decisions with regards to resources management and team support. We will be transitioning our organizational structure and designing a smartsheets platform for notifying individuals about published works, additional grant opportunities, and overall success of each group. We built a similar platform for reporting and communicating about the progress report and this seemed to work nicely for timely reporting in such a large group. One of our Co-PI's from Notre Dame will be visiting for a sabbatical next year and we anticipate this will be a good opportunity to exchange data and enhance our modeling approaches. Scientific Metrics: Our goal is to complete the remaining laboratory studies and data analysis outlined in objective 1 and finalize manuscripts for publication. Data from objective 1 has been used to inform the within midge model for estimating the relationship between temperature and the probability of transmission of reassortant viruses. The postdoctoral scholar on this project is planning on presenting their work at the annual EEID meeting. Additionally, we plan to expand the model to other viruses and explore phylogenetic methods for estimating bluetongue virus reassortment rates. The results from the fieldwork in 2020 and 2021 will help to broaden our knowledge of risk factors of vector-borne diseases and is informing the planning for fieldwork in 2022. In particular, data collection on a diverse set of field sites provides information on the role of the domestic-wildlife-human interface in the spread of vector-borne disease. Additionally, fine-scale temperature and humidity data will help shed light on the relationship between climate and vector-borne disease risk, which can help strengthen predictions in the face of climate change. We will continue to expand our in vivo work as proposed in objective 4. We are also closely aligned in evaluating the budget and adequate support for these studies as maintaining the required secured indoor facilities and personnel to conduct in vivo studies in the native host (sheep, deer and cattle) are considerable. University per diems have increased since the submission and award of this proposal and we have been notified of another planned increase in 2022. We will continue to submit grants seeking additional funds to support this work and will resubmit an NIH R03 to develop novel technology using BTV and sheep as a model system. PI's at CSU and Notre Dame will continue to ensure laboratory and fieldwork (Aims 1-4) are aligned with models developed in Aims 5&6 and we will use preliminary data from our animal surveillance to plan and expand our surveillance efforts in the coming year. Broader Impacts: We will continue to build a laboratory website that allows the project team to publicize research findings and to distribute educational materials. Team members will continue developing and participating in the Genomics of Disease in Wildlife workshop that's held annually at CSU. We are engaged in the process for offering a course-based undergraduate research experience (CURE) at CSU in the Spring of 2022. This will be an ideal platform to demonstrate collaboration and provide education about the importance of vector-borne diseases in ruminant communities. Additionally, our we have engaged the associate director for the department of outreach and engagement at CSU. He is including orbivirus educational materials in his infectious disease unit and will be visiting with K12 programs to help us with outreach in these communities.
Impacts
What was accomplished under these goals?
Bluetongue (BT) and epizootic hemorrhagic disease (EHD) remain economically important, re-emerging disease threats throughout North America and worldwide. Persistent knowledge gaps regarding its evolution and ecology have hindered our ability to effectively predict and prevent outbreaks. Objective 1: We have successfully completed some of the viral evolution studies applying sequencing approaches to understand potential reassortment between BTV-2/BTV-10 and BTV-17/BTV-10 when midges are co-infected. The initial work with BTV-2/BTV-10 was published this year and has allowed us to expand the work to BTV-17/BTV-10. The objective of our first study was to characterize the effect of temperature on bluetongue virus coinfections inC.sonorensis. Coinfections were established inC.sonorenesisthat were reared at three different temperatures. Assessments of survival, virogenesis dynamics, and genotyping of progeny virus were performed. Results indicated that BTV virogenesis varies among different temperatures. Additionally, C. sonorensiscoinfected with BTV-10 and BTV-17 demonstrated a higher proportion of BTV detected pools than single infections with either BTV-17 or BTV-10. Survival also varied by temperature and might offer tradeoffs in effective viral transmission within different environmental conditions. We performed genotyping of plaques from coinfected midges and particular strains contributed the dominant amount of genetic material. The purpose of our second study was to investigate how different ratios of coinfection affected virogenesis dynamics and reassortment. Initial results indicate that virogenesis dynamics are influenced by the ratio of viral strains introduced during midge coinfection and depending on that ratio, some clustered similarly with single infection groups. Propagated plaques are currently being sequenced to assess for reassortment patterns. The objective of our third study was to evaluate frequency of reassortment in midges coinfected with BTV strains that are genetically closer to each other at a single temperature condition. The strains used in these studies were the same BTV-10 strain referenced in previous studies and a different field isolate of BTV-17. Initial results demonstrate that replication dynamics of BTV-10 and BTV-17 single infections are similar compared to those referenced in the first study. Overall, BTV co-infection studies will add an important dimension to the modeling of viral expansion and evolution in the context of climate change. Objective 2: Modeling efforts outlined in Objective 2 are currently underway. A postdoctoral scholar is utilizing data and parameters from studies in Objective 1 to populate and inform his model on viral reassortment and prediction within the Culicoides midge. We have developed a mathematical model of co-infection with two genotypes of bluetongue virus in the midge. The model was then calibrated to data on single infection viral dynamics and reassortment frequency in co-infected midges. The model includes several barriers within the midgeas this has been documented in Culicoides midges infected with BTV. Our results suggest viruses need to arrive close together in time and that viruses must have similar replication rates for reassortment to be non-negligible. We have drafted a full manuscript and are hoping to submit this in the first three months of 2022. Objective 3: One of our graduate students was able to complete a full field season this year with minimal impacts from the SARS-CoV-2 pandemic. They completed field work targeting a diverse group of vertebrate hosts. This entailed setting traps and collecting Culicoides midge species at 17 field sites every 2 weeks for the duration of the field season (mid May - mid October). Field sites represented a range of habitats and host communities: mostly homogenous domestic sites, mixed species domestic, mixed wild and domestic rangeland, and natural sites with only wildlife hosts. Ongoing activities in determining the impact of diversity on transmission of BTV will involve sorting of collected samples (in progress) and further analysis. Model development and data analysis are currently underway for data yielded during the 2020 field season. Trap catch results combined with trap location habitat variables, as well as data from temperature and humidity loggers, are being used to examine the impact of environmental variables on Culicoides populations to increase our knowledge of how land use practices impact transmission. Our graduate student has successfully presented this information at numerous meetings and a manuscript is in preparation. Future field seasons will include studies that interrogate conditions and habitats suitable for Culicoides in the western US (summer 2022). Objective 4: We were able to complete two in vivo studies this year. The first in vivo study was initiated in August of 2021 and the purpose of this study was to build on the findings found from the pilot BTV-17 study conducted in 2020. We evaluated BTV-17 viremia kinetics in the sheep, identified serological responses, and observed several immune subsets (T-, B-cells, and macrophages) and non-cellular immune factors (cytokines) that were dynamically changed over the course of infection. Midges were fed on the BTV infected animals and intermittent positivity was observed in the midges post feeding. This study has solidified the findings of our original pilot study (2020), allowed us to optimize collection and assay techniques, and establish a greater understanding of the overall kinetics of BTV infection in sheep. Our third in vivo study was initiated December 2021 using a different strain of bluetongue virus (BTV-10). The purpose of this study is to evaluate BTV infection, the kinetics of viremia, its impact on the immune response, and the use of midges to feed on positive sheep. This study is, in effect, a comparison to the data generated from the BTV-17 animal studies and used the same parameters as the previous BTV-17 studies. Taken together, we have established a model of BTV infection in sheep and have successfully monitored the longitudinal vertebrate host immunological response and viral infection progression using a combination of traditional methods and cutting-edge technology. These studies will guide our understanding of natural BTV transmission and the data and parameters from these studies will inform future ecological models. Objective 5 & 6: Techniques and data generated from work in Objectives 1-4 will be informative to the models created in Aims 5 & 6. Specifically, for Objective 6, we initiated a pilot study incorporating animal surveillance during the 2021 field season. We collected blood and serum from animals located on 5 cattle and 5 sheep operations. We discovered a substantial number of positive animals using organized and opportunistic surveillance strategies. Increased activity of orbiviruses circulating among our domestic and wildlife ruminant species in the United States and Canada this year has given rise to the CSU Veterinary Diagnostic Laboratory becoming a central location for sample submission when either BTV or EHDV are considered as a primary differential. This opportunity has allowed integration of the diagnostic and research work. Within this scope, we identified a novel serotype of BTV circulating in the western US and partnered with key stakeholders at the state and national level to offer timely serotyping and whole genome sequencing. Subsequently, we have partnered with additional wildlife agencies, universities, and diagnostic labs. This has allowed us to have representation at the national level so that we can make appropriate, science informed decisions about the classification and distribution of these viruses. Diagnostic and clinical service efforts have been extremely rewarding for enhancing research efforts and orbivirus awareness within the last year.
Publications
- Type:
Journal Articles
Status:
Published
Year Published:
2021
Citation:
Kopanke, J., Lee, J., Stenglein, M., Carpenter, M., Cohnstaedt, L.W., Wilson, W.C., Mayo, C. (2021). Exposure of Culicoides sonorensis to Enzootic Strains of Bluetongue Virus Demonstrates Temperature- and Virus-Specific Effects on Virogenesis. Viruses, 13(6). https://doi.org/10.3390
- Type:
Journal Articles
Status:
Published
Year Published:
2021
Citation:
Mayo, C.E., Weyer, C.T., Carpenter, M.J., Reed, K.J., Rodgers, C.P., Lovett, K.M., Guthrie, A.J., Mullens, B.A., Barker, C.M., Reisen, W.K., MacLachlan, N.J. (2021). Diagnostic applications of molecular and serological assays for bluetongue and African horse sickness. Rev Sci Tech, 40(1), 91104. https://doi.org/10.20506
- Type:
Journal Articles
Status:
Published
Year Published:
2021
Citation:
Kopanke JH, Lee JS, Stenglein, MD, Mayo CE (2021). In vitro reassortment between endemic bluetongue viruses features global shifts in segment frequencies and preferred segment combinations. Microorganisms. 9(2), 405; https://doi.org/10.3390/microorganisms9020405.
- Type:
Journal Articles
Status:
Published
Year Published:
2021
Citation:
Gebbiena MB, Strimbu K, Cecilia H, Lerch A, Moore SM, Tran Q, Perkins AT, TenBosch QA (2021). Over 100 Years of Rift Valley Fever: A Patchwork of Data on Pathogen Spread and Spillover. 10 (6); 708 doi: 10.3390/pathogens10060708.
- Type:
Conference Papers and Presentations
Status:
Other
Year Published:
2021
Citation:
Mayo C, Epidemiology of bluetongue virus in the United States: implications for arbovirology at the wildlife-livestock interface, USDA National Bio and Agro-Defense Facility (NBAF), Scientific Symposium, May, 2021.Virtual Oral Presentation.
- Type:
Conference Papers and Presentations
Status:
Other
Year Published:
2021
Citation:
Mayo C, Updates in diagnostic virology & epidemiology of bluetongue virus, Colorado State University Veterinary Teaching Hospital Annual Conference, food animal program, May, 2021. Virtual Presentation.
- Type:
Other
Status:
Other
Year Published:
2021
Citation:
Ebel G, Kading R, Schountz T, Mayo C, Infectious Disease Panel: International Symposium, Colorado State University 3rd International Symposium, March, 2021, Virtual Panel.
- Type:
Conference Papers and Presentations
Status:
Accepted
Year Published:
2021
Citation:
Barbera C, Perkins TA, Mayo C, Rohr J, Impacts of host density on vector abundance and transmission of bluetongue virus, Society of Vector Ecology, September, 2021. Virtual Oral Presentation.
- Type:
Conference Papers and Presentations
Status:
Accepted
Year Published:
2021
Citation:
Carpenter MJ, Kopanke J, Rodgers C, Lee JS, Stenglein M, Mayo CE. Evaluating bluetongue virus reassortment in Culicoides sonorensis. Poster presentation at the American Association of Veterinary Laboratory Diagnosticians Annual Conference, October 2021. Poster Presentation.
- Type:
Conference Papers and Presentations
Status:
Accepted
Year Published:
2021
Citation:
Mayo C, Reed K, Ragan I, Rodgers C, Hodges N, Anderson T, Sherman T, Walck R, Pabilonia, K, Stenglein M, Novel Approach to Diagnose and Genetically Characterize High Consequence Pathogens Affecting Animal Health, National Animal Health Laboratory Network (NAHLN) 2019 farm bill showcase, November 2021.Virtual Oral Presentation.
- Type:
Conference Papers and Presentations
Status:
Accepted
Year Published:
2021
Citation:
Westrich J, McNulty E, Carpenter M, Burton M, Mayo C, Mathiason CK, Monitoring longitudinal immunological responses to Bluetongue Virus 17 in experimentally infected sheep, Conference for research workers in animal disease, Chicago, IL, December 2021. Virtual Oral Presentation.
- Type:
Conference Papers and Presentations
Status:
Accepted
Year Published:
2021
Citation:
Barbera C, Perkins TA, Mayo C, Rohr J, Impacts of host density on vector abundance and transmission of bluetongue virus, Conference for research workers in animal disease, Chicago, IL, December 2021. Oral Presentation.
- Type:
Conference Papers and Presentations
Status:
Accepted
Year Published:
2021
Citation:
Westrich JA, McNulty EE, Carpenter M, Burton M, Mayo C, Mathiason CK, Determining the longitudinal viral progression and immunological responses to Bluetongue Virus 17 experimentally infected sheep,Rocky Mountain Virology Club October 2021. Poster presentation
- Type:
Other
Status:
Accepted
Year Published:
2021
Citation:
Mayo C, Ebel G, Olson D, Hurrell J, Session 3: Fleas, Ticks, Mosquitoes: Change in vector-borne disease, Climate change and one health workshop, May 2021. Virtual Panel.
- Type:
Conference Papers and Presentations
Status:
Accepted
Year Published:
2021
Citation:
Carpenter MJ, Kopanke J, Rodgers C, Lee JS, Stenglein M, Mayo CE. Characterizing determinants of reassortment of bluetongue virus in Culicoides sonorensis. Poster presentation at the Association for Clinical and Translational Science, April, 2021. Virtual Poster Presentation.
|
Progress 12/15/19 to 12/14/20
Outputs
Target Audience:The target audience of these studies include policy makers (national and regional), veterinary diagnosticians, livestock producers and veterinarians, and industry stakeholders (dairy, beef cattle, sheep, deer, and wildlife). We are engaging with leaders of Future Farmers of America (FFA) programs in our local school district in order to developinteractive field exercises that incorporate aspects of vector-borne disease and ruminant host species. The goal of this exercise is to integrate problem solving for understanding and preventing vector-borne diseases that affect the livestock industry. We are also in discussions with the department microbiology, immunology, and pathology to develop and offer a course-based undergraduate research experience (CURE) at CSU in the Spring of 2022. The primary focus of the course content will be reinforcing the collaboration and depth of projects (i.e. sequencing, fieldwork, modeling, in vivo work) we are currently conducting. We have identified and assigned an individual to initiate a laboratory website so that the project and project team can publicize research findings and distribute educational materials. Team members will continue to be involved in developing and leading an annual "Genomics of Disease in Wildlife" workshop that was offered for the first time in June 2017 (Co-PI Stenglein is co- developer and co-lead). This workshop, which is aimed at fostering the adoption of genomics approaches by wildlife researchers, leverages existing NSF EEID funding (to PI Sue VandeWoude) and includes participation from many members of the team including the PI (Christie Mayo). Unfortunately, the course was canceled this past year due to the impacts of COVID but we will continue to collaborate in this activity at CSU. Important aspects of reaching our target audience through our broader impact activities include reinforcing the study of a globally emerging arbovirus infections of livestock and wildlife, including the potential impact of climate change thereon. Changes/Problems:There are no significant changes or problems that have been encountered for the project. Due to the SARS-CoV-2 pandemic, research activities at Colorado State University were suspended in March and April of 2020. As principle investigators, we were advised to apply for a return to research review in May. Research activities returned at a level of 50%; however, the pandemic continedto have an impact on our overall performance this year. In the interest of making the most efficient progress, we were able to identify the critical needs and expertise in order to continue the progress. We are remaining flexible based on the uncertainty of the pandemic and potential subsequent re-closures or reduction in research effort at the department or university level. What opportunities for training and professional development has the project provided?The project has served as a venue for the involvement and training/education of undergraduate, professional (DVM), producers, extension agents, graduate academic students, and postodctoral scholarsin field, laboratory, modeling studies. The work has expandedfrom temperature dependent reassortment studies in Culicoides sonorensis to in vivo and field studies this past summer. We have been working with livestock producers in order to expand and include free-ranging species, specifically beef cattle and sheep, and engaged professional students to participate in this aspect. Laboratory studies have included virologic investigations, laboratory evaluation of vector midges, and immunologic investigation of data obtined in ourin vivo studies. The professional development in our students has been rewarding as they learn from each other and their mentors in this multi-disciplinary program. The combination of skillsets and expertise have allowed us to initiate discussions with the department microbiology, immunology, and pathology in order to develop and offer a course-based undergraduate research experience (CURE) at CSU in the Spring of 2022. The primary focus of the course content will be reinforcing the collaboration and depth of projects (i.e. sequencing, fieldwork, modeling, in vivo work) we are currently conducting. Overall, we have also incorporatedaspects of diversity, equity, and inclusion as we continue to foster these collaborations and promote members of our team. How have the results been disseminated to communities of interest?Results of the laboratory studies presented in Objective 1 have been submitted to peer reviewed journals, and presented to relevant professional and commodity groups including regional (County-based) livestock producer groups, the dairy industry, the American Association of Veterinary Laboratory Diagnosticians, the Conference for Research Workers in Animal Disease (CRWAD).Due to the SARS-CoV-2 pandemic, most of the meetings we attended this year were virtual and there were limited opportunities to conduct outreach initiatives. We have been able to shift to the virtual platform relativelyquickly and are actively engaged in discussions to promote website development and additional outreach. What do you plan to do during the next reporting period to accomplish the goals?Administrative: Within the last year, we have built and will continue to refine ourinfrastructure for the primary teams. We have been successful in enhancing our communication by forming a monthlymeeting for all members of the team. Thisallows for us to share updates and data among groups. We have also established and will continue to have a meeting among Co-PD's every 6 months and this meeting has allowed us to make decisions with regards to resources management and team support.We will be transitioning ourorganizational structure and designing a google drive for notifying individuals about published works, additional grant opportunities, and overall success of each group. Scientific Metrics: Our goal is to complete the bluetongue virus aspects of objective 1 and continue this work with epizootic hemorrhagic disease virus. An additional graduate student has joined Dr. Mayo's laboratory and will be initiating this work in 2021. Data from objective 1 will be used to inform the within midge model for estimating the relationship between temperature and the probability of transmission of reassortant viruses. Additionally, we plan to expand the model to other viruses and explore phylogenetic methods for estimating bluetongue virus reassortment rates. The results from the fieldwork in 2020 will help to broaden our knowledge of risk factors of vector-borne diseases and inform the planning of fieldwork in 2021. In particular, data collection on cattle operations of varying sizes provides information on the role of the domestic-wildlife-human interface in the spread of vector-borne disease. Additionally, fine-scale temperature and humidity data can help shed light on the relationship between climate and vector-borne disease risk, which can help strengthen predictions in the face of climate change. We will continue to expand our in vivo work as proposed in objective 4. We are also closely aligned in evaluating the budget and adequatesupport forthesestudies as maintaining the required secured indoor facilities and personnel to conduct in vivo studies in the native host (sheep, deer and cattle) are considerable. University per diems have increased since the submission and award of this proposal and we have been notified of another planned increase in 2021. We will continue to submit grants seeking additional funds to support this work and willresubmit an NIH R03 to develop RNA flow using BTV and sheep as a model system. Drs. Perkins and Mayo will continue to ensure laboratory and fieldwork (Aims 1-4) are aligned with models developed in Aims 5&6. We will also be planningfor expanding our surveillance efforts to incorporate animal surveillance in addition to insect collections. Broader Impacts: We will continue to build a laboratory websitethat allows the project team to publicize research findings and to distribute educational materials. While the annual "Genomics of Disease in Wildlife" workshop was cancelled this year due to COVID, team memberswill continue developing and participating in this effort. We are engaging in the planning process for offering a course-based undergraduate research experience (CURE) at CSU in the Spring of 2022. This will be an ideal platform to demonstratecollaboration and provide education about the importance of vector-borne diseasesin ruminant communities.
Impacts
What was accomplished under these goals?
Bluetongue (BT) and epizootic hemorraghic disease (EHD) remain economically important, re-emerging disease threats throughout North America and worldwide. Persistent knowledge gaps regarding its evolution and ecology have hindered our ability to effectively predict and prevent outbreaks. Moreover, climate change is expected to continue to reshape the distribution of these arboviral diseases, likely accelerating the rate of incursions and the introduction of non-endemic serotypes into new regions. Objective 1: We continue to apply a number of novel sequencing technologies and experimental approaches to answer specific questions about orbivirus genetic diversity and viral population structure using both in vitro and in vivo systems. Our proposed work uses bluetongue virus (BTV), epizootic hemorrhagic disease virus, culicoides cell culture, and colony raised Culicoides sonorensis (also known as midges) as our backbone for understanding these relationships. Methods use an in vitro system and novel sequencing approaches to understand reassortment between BTV viral strains and EHDV viral strains. The same viral strains allow us to explore how incubation temperature (20°C, 25°C, 30°C) might affect virogenesis and reassortment in Culicoides sononensis, the predominant BTV vector in North America. Our work on BTV 2 and 10 co-infection has been submitted to Virusesfor peer review. Findings from this work demonstrated low infection rates of BTV-2 in Culicoides sonorensis; therefore, we sought to characterize the frequency of reassortment between 2 different endemic strains (BTV-10 and BTV-17). These strains have been isolated in western United States for decades and we were able to demonstrate infections of C.sonorensis with each individual viral strain. The insects were incubated at the same temperatures as our previous work (20°C, 25°C, 30°C). We have completed the infection studies and evaluated vector survival, virogenesis, and BTV reassortment. Overall, midges survived longer in cooler temperatures and virogenesis was greater in midges maintained at 25°C as compared to those maintained at 20°C or 30°C. Results also indicate that midges maintained at 30°C demonstrate productive virogenesis earlier in infection (day 3) than midges held at cooler temperatures (day 7). In order to evaluate reassortment, coinfected midges collected on days 3,7, 11, 15, and 19 were processed for BTV plaque-isolation for future amplicon assays. Preliminary results from a single time point and representative plaques have revealed limited reassortment as most reads align to BTV17. However, RNA was extracted directly from coinfected midges and results of next generation sequencing revealed reads aligning to both BTV 10 and BTV 17 at 25C. While this does not demonstrate reassortment, we identified there are reads aligning to both virus populations in midges at 25C. Sequencing efforts are currently underway to characterize additional plaques and RNA extracted directly from BTV10&17 coinfected midges on additional time points throughout the study so that we can examine the virus populations thoroughly. Overall, BTV reassortment patterns and their biological consequences will add an important dimension to the modeling of viral expansion and evolution in the context of climate change. Objective 2: Modeling efforts outlined in Objective 2 are currently underway. We are utilizing previously published data to inform the model; however, data collected from Aim 1 will help inform the model. Framework for modeling cellular coinfection and viral reassortment was developed and implemented. Full co-infection model was then calibrated to data from el Hussein (1989) on reassortment frequency in co-infected midges using Markov Chain-Monte Carlo methods. We incorporated a midgut infection barrier as in Fu et al. (1999), in which only a proportion (~30%) of infections establish in the midgut and make it to the secondary tissues. Preliminary results suggest viruses need to arrive close together in time (within a few days) for reassortment to be non-negligible. Objective 3: While the COVID pandemic had an impact on the initial portions of our field season, we were able to use previous field studies to guide our fieldwork and site selection for studies outlined in Objective 3. Our graduate student from Notre Dame was able to travel to CSU and initiate work on July 1st. The field season extended to September 18th and weekly collections of insects were conducted on dairy and feedlot facilities using traps baited with carbon dioxide. Data yielded from the processing of the 2020 field samples will be analyzed to detect a relationship between ruminant density and Culicoides life traits, in order to inform our understanding of the potential for vertebrate host density-dependent transmission of BTV and EHDV. Additionally, trap catch results combined with trap location habitat variables, as well as data from iButton temperature and humidity loggers, will be used to examine the impact of environmental variables and on Culicoides populations to inform our knowledge of how land use practices may impact transmission. We plan to use a series of generalized additive models (GAMs) and compare models using model selection techniques. Analysis will be performed in R. Once our findings from the 2020 field season are completely analyzed, further planning for the 2021 season will begin. Objective 4: We have successfully completed an in vivo study of BTV 17 infection in sheep. Infection was determined by intermittent blood draws and assessment for viral RNA by RT-qPCR from mock- and BTV 17-infected animals. BTV viral RNA levels were detected in BTV 17-infected animals at 8 days pi and remained detectable throughout the study (24 days pi). Mock-infected animals remained negative. A serum serological response against BTV 17 was observed in the infected animals as determined by BTV specific ELISA. Modest but clear clinical symptoms were observed in the BTV-infected animals that were not present in mock-infected animals. To evaluate the infectious capacity of our study animals to transmit BTV to C. sonorensis midges, we established midge feeding on infected animals during this time course. Blood meals were observed in a portion of the midges that were reared through their extrinsic incubation period for further downstream analysis by RT-qPCR and next generation sequencing. To gain additional insights and a greater understanding of other potential host parameters that may impact pathogen transmissibility, we evaluated several key immune parameters known to be critical in BTV infection. Circulating CD3+ T- , CD4+ T-, CD8+ T-, B-cells, and monocytes were quantified and evaluated by flow cytometry using previously validated antibodies. In addition to demonstrating the well establish pan-leukopenia in BTV infections, we also reveal interesting trends in specific cell subtype populations that merit further investigation. In addition to characterizing the cellular immune populations, circulating cytokine (CXCL10, CXCL8, CXCL9, DCN, FRZB, IFNG, IL17A, IL21, TNF, and VEGFA) protein levels are being assessed by cytokine array (Raybiotech). Reagents to explore the immune response established against BTV and EHDV in native ruminant host species (deer, cattle and sheep) are limited. To this end, steps are being taken to establish RNA flow cytometry which will permit extended investigation of the immunological response to these viral infections in the native host and thus the role the immune response plays in BTV/EHDV viral pathogenesis. Objective 5&6: Techniques and data generated from work in Aims 1-4 will be informative to the models created in Aims 5&6. We are planning the initial phases of incorporating animal surveillance into the fieldwork component of project so that we can understand the prevalence and phylogeny of circulating orbiviruses (BTV/EHDV).
Publications
- Type:
Journal Articles
Status:
Published
Year Published:
2020
Citation:
Kopanke JH, Lee JS, Stenglein MD, Mayo CE (2020). The genetic diversification of a single bluetongue virus strain using an in vitro model of alternating-host transmission. Viruses. 12(9):E1038. doi: 10.3390/v12091038.
- Type:
Journal Articles
Status:
Submitted
Year Published:
2020
Citation:
Bron GM, Strimbu K, Cecilia H, Lerch A, Moore S, Tran Q, Perkins TA, ten Bosch QA (2020). Over 100 years of Rift Valley Fever: a patchwork of data on pathogen spread and spillover. Submitted to Pathogens.
- Type:
Journal Articles
Status:
Published
Year Published:
2020
Citation:
Mayo C, McDermott E, Kopanke J, Stenglein M, Lee J, Mathiason C, Carpenter M, Reed K, Perkins TA (2020). Ecological Dynamics Impacting Bluetongue Virus Transmission in North America. Fronteirs in Veterinary Science, 7:186.doi: 10.3389/fvets.2020.00186.
- Type:
Conference Papers and Presentations
Status:
Published
Year Published:
2020
Citation:
Carpenter, M, Kopanke, J., Rogers, C, Lee, J, Stenglein, M, Mayo, C.(2020). Characterizing the effect of temperature on bluetongue virus reassortment in Culicoides sonorensis. Conference for Research Workers in Animal Disease, virtual, October, Poster
- Type:
Conference Papers and Presentations
Status:
Published
Year Published:
2020
Citation:
Carpenter, M, Kopanke, J., Rogers, C, Lee, J, Stenglein, M, Mayo, C.(2020). Characterizing the effect of temperature on bluetongue virus reassortment in Culicoides sonorensis. American Association of Veterinary Laboratory Diagnosticians, virtual, October, Poster, Poster award
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Progress 12/15/18 to 12/14/19
Outputs
Target Audience:The target audience of these studies include policy makers (national and regional), veterinary diagnosticians, livestock producers and veterinarians, and industry stakeholders (dairy,beef cattle, sheep, deer, and wildlife). We are planningtoincorporate our research findings into an interactive field exercise and management scenario for both the local schools and state fairs within Larimer County. This exercise will integrate problem solving for understanding and preventing vector-borne diseases that affect the livestock industry. Websites will be established and managed by the project team to publicize research findings and to distribute educational materials. Team members are involved in developing and leading an annual "Genomics of Disease in Wildlife" workshop that was offered for the first time in June 2017 (Co-PI Stenglein is co-developer and co-lead). This workshop, which is aimed at fostering the adoption of genomics approaches by wildlife researchers, leverages existing NSF EEID funding (to PI Sue VandeWoude) and includes participation from many members of the team including the PI (Christie Mayo).Important aspects of reaching our target audience through our broader impactactivities include reinforcing the study of a globally emerging arbovirus infections of livestock and wildlife, including the potential impact of climate change thereon. Changes/Problems:There are no significant changes or problems that have been encountered for the project. Due to some government delays, we did not receive funding until April, 2019. We have made significant progress with regards to projects presented in Aims 1-2 and will be initiating fieldwork, in vivo projects, and modelingas proposed (Aims 3-6) in 2020. What opportunities for training and professional development has the project provided?The project has served as a venue for the involvement and training/education of undergraduate, professional (DVM), extension agents and graduate academic students in both field and laboratory research. Much of the lab work has centered on temperature dependent reassortment studies in culicoides sonorensis. We have been working with livestock producers in order to expand andinclude free-ranging species, specifically beef cattle and sheep, and engaged professional students to participate in this aspect. Laboratory studies have included virologic investigations, as well as the laboratoryevaluation of vector midges. How have the results been disseminated to communities of interest?Results of the laboratorystudies presented in Aim 1 have been submitted to peer reviewed journals, and presented to relevant professional and commodity groups includingregional (County-based) livestock producer groups, the dairy industry, the American Association of Veterinary Laboratory Diagnosticians, the Conference for Research Workers in Animal Disease (CRWAD). What do you plan to do during the next reporting period to accomplish the goals?Administrative: Within the next year, we plan on building a defined infrastructure for the primary teams. We have invested in a program (Smartsheets) that will allow us to build rolling action item lists (RAILS) for each group. This program will provide a concise infrastructure for notifying individuals about published works, additional grant opportunities, and overall success of each group. Scientific Metrics: Our goal is to have Objective 1 complete within 2020 so that these methods can inform Objectives 2-5. We will initiate fieldwork (Aim 3) during the Spring of 2020. All in vivo studies (Aim 4) will be initiated during Spring, 2020. Drs. Perkins and Mayo will ensure laboratory and fieldwork (Aims 1-4) are aligned with models developed in Aims 5&6. Broader Impacts: We plan on establishing formalized websites about the project team to publicize research findings and to distribute educational materials. Team members are involved in and will continue developing an annual "Genomics of Disease in Wildlife" workshop that was offered for the first time in June 2017 (Co-PI Stenglein is co-developer and co-lead). We plan on building our infrastructure for introducing courses in the undergraduate curriculum. The department of Microbiology, Immunology, and Pathology at Colorado State University offers a programthat brings undergraduates into the laboratory ("undergradsinthelab). This will be an ideal platform to initiate programs in understanding the interface of virus-vector-host transmission.
Impacts
What was accomplished under these goals?
Bluetongue (BT) and epizootic hemorraghic disease (EHD) remains economically important, re-emerging disease threats throughout North America and worldwide. Persistent knowledge gaps regarding its evolution and ecology have hindered our ability to effectively predict and prevent outbreaks. Moreover, climate change is expected to continue to reshape the distribution of these arboviral diseases, likely accelerating the rate of incursions and the introduction of non-endemic serotypes into new regions. Objective 1: Through successful completion of laboratory work outlined in Objective 1, we have applied a number of novel sequencing technologies and experimental approaches to answer specific questions about orbivirus genetic diversity and viral population structure using both in vitro and in vivo systems. Upon receiving funds in April, 2019, we initiated our work using bluetongue virus (BTV), culicoides cell culture, and colony raised Culicoides sonorensis as our backbone. Significant findings from these laboratory studies include the following: 1. We applied an in vitro system and novel sequencing approaches to understand reassortment between BTV-2 and BTV-10. Our results indicated that reassortment occurs readily between these two strains within an in vitro system. However, global shifts in segment frequencies arise across passages, possibly indicating a preferred virus backbone. Despite the segment-specific trends that arose, it is interesting to note that measures of effective diversity (which were based on individual plaque genotypes and thus likely underestimated the true diversity detected via metagenomic sequencing) significantly increased during the course of coinfection. This is in contrast to the findings from previous work conducted in our lab where we evaluated the occurrence of genetic variation within a single virus strain across passage. In contrast to coinfections, the measures of population complexity were essentially unchanged across passages. Although these estimates of diversity are not directly comparable, these findings nevertheless allude to the prominent role that reassortment likely plays in BTV's evolution (manuscript in final preparation). 2. We used these same viruses (BTV-2 and BTV-10) to investigate how incubation temperature (20°C, 25°C, 30°C) might affect virogenesis and reassortment in Culicoides sononensis, the predominant BTV vector in North America. Unexpectedly, we found that BTV-2 generally failed to infect midges unless introduced at very high titers, highlighting the complexities of virus-vector interactions and their role in modulating arbovirus expansion. We also detected temperature- and virus-specific effects on midge survival, which is significant for our understanding of BTV ecology and will help refine forthcoming predictive modeling efforts (manuscript in final preparation). 3. Due to low infection rates of BTV-2 in Culicoides sonorensis, we sought to characterize the frequency of reassortment between 2 different endemic strains (BTV-10 and BTV-17). These strains have been isolated in western United States for decades and we were able to demonstrate infections of C.sonorensis with each individual viral strain. The insects were incubated at the same temperatures as presented above in item 2 (20°C, 25°C, 30°C). We have completed the infection studies and insects will be processed for BTV plaque-isolation. The complete genotypes of isolated plaques will be determined using a novel, amplicon-based sequencing approach. Preliminary results indicate that midges maintained at 30°C demonstrate productive virogenesis earlier in infection (day 3) than midges held at cooler temperatures (day 7). Overall, BTV reassortment patterns and their biological consequences will add an important dimension to the modeling of viral expansion and evolution in the context of climate change. Understanding the multiple factors that drive the emergence of viruses is critical for the development of improved control and prevention measures. Objective 2: Modeling efforts outlined in Objective 2 are currently underway. We are utilizing previously published data to inform the model; however, data collected from Aim 1 will help inform the model. Objective 3: Dr. Mayo's laboratory has been conducting routine sentinel animal (2015) and entomologic surveillance (2018-2019) in Colorado. We will be using this data to guide our fieldwork and site selection for studies outlined in Objective 3. We have recruited a graduate student who will be conducting these field studies and integrating modeling approaches under the supervision of Drs. Christie Mayo and Alex Perkins. We have also established collaborations with Dr. Jason Rohr (ecology & environmental biology) and Bethany McGregor (entomologist) who will be instrumental in advising our studies. During August, 2019, our group evaluated field sites in Colorado, Wyoming, and Montana. The University of Notre Dame Environmental Research Center will offer an ideal location to study the ecology of Culicoides and virus-vector-host interactions for both wild and domestic ruminants. Objective 4: Animal care and use protocols have been approved for in vivo work outlined in Objective 4. We plan on initiating these studies during the Spring, 2020. The laboratory studies outlined Objective 1 have allowed us to optimize techniques for infecting culicoides and feeding on vertebrate hosts. Dr. Mathiason's group is optimizing an assay that will allow us to combine flow cytometry with RNA scope technology. Optimally, this will allow us to build upon proposed work and evaluate reassortment within the vertebrate host. Objective 5&6: Drs. Christie Mayo and Alex Perkins are working closely with groups generating data in Aims 1-4. Techniques and data generated from this work will be informative to the models created in Aims 5&6.
Publications
- Type:
Journal Articles
Status:
Published
Year Published:
2019
Citation:
McDermott EG, Mullens BA, Mayo CE, Roark EB, Maupin CR, Gerry AC, Hamer GL. (2019). Laboratory evaluation of stable isotope labeling of Culicoides (Diptera: Ceratopogonidae) for adult dispersal studies. Parasites and Vectors, 12(1): 411 doi: 10.1186/s13071-019-3671-9
- Type:
Book Chapters
Status:
Awaiting Publication
Year Published:
2020
Citation:
Perkins TA, J Rohr. Theories of Diversity in Disease Ecology. In: Theoretical Ecology: Concepts and Applications, 4th edition. (K. McCann and G. Gellner, eds.) Oxford University Press. In press.
- Type:
Journal Articles
Status:
Under Review
Year Published:
2019
Citation:
Kopanke J, Lee J, Stenglein M, Mayo C. (2019). The genetic diversification of a single bluetongue virus strain using an in vitro model of alternating-host transmission. Journal of Virology, (submitted, under review)
- Type:
Conference Papers and Presentations
Status:
Accepted
Year Published:
2019
Citation:
Carpenter M, Kopanke J, Rogers C, Lee J, Stenglein M, Mayo C, (2019). Characterizing the effect of temperature on bluetongue virus reassortment in Culicoides sonorensis. The Conference of Research Workers in Animal Diseases (CRWAD).
- Type:
Journal Articles
Status:
Under Review
Year Published:
2020
Citation:
Mayo C, Kopanke J, Stenglein M, Lee J, Mathiason C, Carpenter M, Reed K, Perkins A, Ecology and evolution of bluetongue virus at the wildlife-domestic interface, Frontiers in Veterinary Science, (submitted, under review).
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