Location: Arthropod-borne Animal Diseases Research
2022 Annual Report
Objectives
Objective 1: Ascertain the viral ecology and factors mediating the introduction and expansion of VSV in the U.S.
Objective 1A. Identify viral genetic determinants mediating emergence of epidemic VSV in the US as well as adaptation to insect and animal hosts.
Objective 1B. Characterize epidemiological, biotic and abiotic factors
associated with vectorial capacity, emergence, incursion, and expansion of VSV from endemic areas into the U.S.
Objective 2. Develop intervention strategies to minimize the impact of VSV disease outbreaks.
Objective 2A. Develop model-based early warning systems to predict future incursions of VSV from Mexico to the U.S.
Objective 2B. Identify vector transmission control strategies based on our understanding of virus-vector-host interactions.
Approach
Vesicular stomatitis (VS) is a vector-borne, zoonotic disease caused by the RNA virus, vesicular stomatitis virus (VSV). Disease in cattle and pigs is clinically indistinguishable from foot-and-mouth disease (FMD), one of the most devastating exotic diseases in the US which was eradicated in 1929. For the past 100 years, incursions of VS have occurred in the US at 8–10-year intervals. Viral incursions originating in endemic regions of southern Mexico start in western border states (NM, TX, AZ) and expand northward with outbreaks often covering over a million square kilometers. Recent outbreaks occurred in 2004-05, 2014-15 and 2019-20, causing thousands of cases across 12 states, and suggesting shorter intervals (5-10 y) may be the new normal. This Project Plan is proposed by two ARS Units, with complementary VSV expertise, to conduct research under two overarching objectives or goals: 1) to identify ecological and virus-vector-host factors that mediate incursion and expansion of VS in the US; and 2) to develop countermeasures including rapid assessment, early warning models and vector control strategies, to reduce the impact of VS disease to US agriculture. This project integrates molecular biology, virology, pathology, entomology, phylogeography, and ecology to better understand the viral, vector, host, and environmental drivers of VS epidemiology across its spatiotemporal domain. Our multidisciplinary approach spans from basic research to applied, and from molecular and organismal (biotic) levels to environmental (abiotic) levels. The proposed project also involves mutually beneficial collaborations with the ARS VSV-Grand Challenge project “Vesicular Stomatitis as a Model for a Predictive Disease Ecology” and three other CRIS Project Plans across three National Programs.
Progress Report
This is the first report for this project started during fiscal year (FY) 2022. For additional progress, please refer to project 8064-32000-062-000D. See the final reports for the project that was replaced under 2021 3020-32000-013-000D and 8064-32000-059-000D, “Ecology of Vesicular stomatitis virus” for additional information.
Progress was made toward understanding insect and environmental factors that affect the introduction and expansion of vesicular stomatitis virus (VSV) in the U.S. Specifically, progress was made toward understanding how the genetics of vesicular stomatitis virus (VSV) relate to its ability to spread outside of an endemic region by how it interacts with the insects that transmit it. Previous sequencing of a strain that escaped the endemic region to cause an epidemic in the U.S. (epidemic VSV 1.1) and a closely related strain that did not escape the endemic region (endemic VSV 1.2) revealed significant differences in just seven amino acids. When observed in swine, VSV 1.1 was more virulent than VSV 1.2. We compared the efficiency of these two viral lineages to infect the vector Culicoides sonorensis and disseminate to salivary glands for subsequent bite transmission. Midges infected with epidemic VSV 1.1 strain had significantly higher infection dissemination rates compared to the endemic VSV 1.2 strain. Thus, in addition to being more virulent in pigs, small genetic changes may also affect a strain’s ability to replicate in the insect vectors that transmit them. This research suggests that both virus-animal interactions and virus-vector interactions contribute to the ability of specific viral lineages to escape endemic regions in Mexico to mover northward and cause epidemic outbreaks in the U.S.
Progress was made towards understanding the effects of increasing environmental temperatures on the physiology of biting midges and their vector competence for VSV. Midges held at different constant temperatures (20, 25, 30, and 35 degrees Celsius) were evaluated for their preference of resting temperature after ingesting a blood meal, reproduction rates, VSV vector competence, and overall lifespan. The preferred temperature of resting sites is important as it subsequently mediates the rates at which blood is digested, eggs are laid, and VSV particles are replicated inside the midge. Higher temperatures correlated with shorter lifespans, shorter egg-laying cycles, and higher rates of viral replication including dissemination to the midge’s salivary glands. Lower temperatures correlated to longer lifespans, more days in their egg-laying cycles, and lower viral replication in the midge. Most midges preferred to rest in areas that fall within their preferred physiological range regardless of the environmental temperatures at which they were being maintained. These preferred temperatures maximized their survival, the number of egg-laying cycles, and the likelihood of VSV transmission. This shows that for this vector-virus system, higher global temperatures may mean higher virus transmission rates and regardless of the temperature of the environment in which they are living, midges find their preferred resting temperatures which optimizes the conditions for both the insect and the virus.
Progress was made towards understanding how virus infection may affect sensory gene expression in the insect vector. VSV infects Culicoides sonorensis biting midges which readily transmit them to livestock resulting a clinical vesicular disease similar to foot and mouth in cattle and swine. Previously we showed that this virus infects sensory organs in midges following infectious blood feeding exposure; specifically the eye, cerebral neural node, and the antennae. Based on these results, it has been hypothesized that VSV infection of midges may lead to changes in gene and protein expression in sensory organs, changes in sensory function, and ultimately, changes in behavior such as photo-attraction, host seeking, and circadian rhythm. In collaboration with Kansas State University, progress was made on assessing the effect of infection on sensory gene expression in female C. sonorensis midges. Initial blood feeding trials with subsequent transcriptome sequencing experiments have been conducted. A high degree of variation in changes in gene expression between midges fed on infected versus uninfected blood were seen. Methodology has been improved to minimize this variation in future experimental trials. A new analysis pipeline has also been developed for examining the transcriptome data which is integral in determining what the effects of VSV infection are on gene and protein expression in Culicoides midges.
Progress was also made toward assessing changes in circadian rhythm of VSV-infected midges. If virus infection changes when midges are actively feeding, it will inform a change to the timing of bite mitigation efforts for horses. A Drosophila activity monitoring (DAM) system was adapted to Culicoides midges and is being used to establish baseline circadian rhythms to compare with midges infected with VSV. Several replicate trials with VSV-infected midges have been completed. Results will be used to assess the effects of VSV infection on daily and seasonal activity of midges and inform how that may affect the risk of viral transmission to cattle, horses, and swine.
Progress was made toward establishing an embryonated chicken egg (ECE) model to study the transmission of VSV by Culicoides midges. VSV inoculation methods and minimum infectious doses for positive ECE controls have been established. Results showed that virus inoculation of the microvasculature to result in a productive infection. VSV was successfully isolated from embryonic livers 24 hours following needle inoculation. Initial trials are being conducted by feeding VSV-injected, positive control midges on exposed microvasculature of ECEs to demonstrate VSV transmission.
Progress was made toward understanding how VSV infection may alter the ability of biting midges to perceive and react to light. This is critical for developing efficient trapping strategies. Three LED light arenas were constructed, and several experiment trials have been conducted to test the functionality of the new light arenas on uninfected midges. These baseline behavioral results of uninfected midges will be compared with VSV-infected midges. The first replicate trial of VSV-infected midges has been conducted. These studies are leading to the final goal of customized light traps to specifically attract VSV-infected midges horse owners could employ and that could be placed near cattle operations to enhance surveillance and reduce risk of transmission.
Accomplishments
1. Small genetic changes may determine whether a virus will spread from Mexico to the U.S. by biting midges. Although vesicular stomatitis virus (VSV) is endemic to Mexico, every 5-10 years specific strains escape and spread north to the U.S. to cause disease in cattle, horses, and swine. Significant economic losses to livestock owners result from lengthy quarantines imposed on their premises. The virus, insect, and host aspects needed for viruses to sporadically move out of Mexico are not well understood, making it very difficult to predict when outbreaks will affect U.S. livestock. Previously, ARS researchers at Plum Island, New York, looked at the genes of two VSV strains: an epidemic strain that escaped Mexico to cause outbreaks in the U.S., and an endemic strain that did not. Although very few genetic differences were found between them, they caused quite different levels of disease in pigs. ARS researchers in Manhattan, Kansas, have now shown that the epidemic strain that caused more disease in pigs, also causes more infection in biting midges and is more likely to be transmitted than the endemic strain. Therefore, in addition to affecting how the virus interacts with animals, small genetic changes may also affect the way the virus interacts with the insects that transmit the virus, allowing it to escape endemic regions of Mexico and spread north into the U.S. Researchers can now use this genetic information to better predict when viruses will be more likely to spread north into the U.S. by biting midges.
2. A hotter environment may mean faster virus transmission to livestock by biting midges. Culicoides midges transmit vesicular stomatitis virus (VSV) to cattle, horses, and swine. Midges feed on blood from these animals to obtain protein for laying their eggs. After ingesting the blood meal, midges find a location to rest at a preferred temperature which affects the rate at which the blood is digested, eggs mature, and if there’s virus in the bloodmeal, the rate at which that virus multiplies inside the midge. In collaboration with Kansas State University, ARS researchers in Manhattan, Kansas, evaluated increasing environmental temperatures on the midge’s resting temperature preferences, on their reproduction cycles, on the rate of VSV replication, and on the midge’s overall lifespan. Regardless of the increased environmental temperatures, most midges found areas with preferred climates that maximized their survival, the number of egg-laying cycles during their lifespan, and the likelihood of VSV transmission. Higher temperatures resulted in more virus and shorter blood feeding/egg-laying cycles. This suggests that as global temperatures rise, not only would more frequent contact be expected between midges and animals to accommodate that shorter feeding/egg laying cycle, but more virus would be present in those feeding midges as well. This combination would increase bite rates on animals and increase the viral doses midges deliver to those animals, resulting in an increased frequency of VSV transmission to U.S. cattle, horses, and swine.
Review Publications
Rozo-Lopez, P., Pauszek, S.J., Velazquez Salinas, L., Rodriguez, L.L., Park, Y., Drolet, B.S. 2022. Comparison of endemic and epidemic vesicular stomatitis virus lineages in Culicoides sonorensis midges. Viruses. 14(6):1221-1233. https://doi.org/10.3390/v14061221.
Rozo-Lopez, P., Park, Y., Drolet, B.S. 2022. Effect of constant temperatures on Culicoides sonorensis midge physiology and vesicular stomatitis virus infection. Insects. 13(4):372-386. https://doi.org/10.3390/insects13040372.
