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Novel vaccine and diagnostic strategies against Schmallenberg virus

Periodic Reporting for period 1 - SchmaVirusVacDiag (Novel vaccine and diagnostic strategies against Schmallenberg virus)

Reporting period: 2015-06-15 to 2017-06-14

The aim of the project was to enable the fellow to gain further insight into the nature of Schmallenberg virus (SBV), a pathogen that continues to threaten the European livestock industry. The project also focussed on therapeutic measures to combat the virus, including the development and validation of novel DNA vaccines against SBV.

Our interest in SBV was of particular concern to the European livestock industry, as outbreaks of the disease has resulted widespread abortion of sheep, goats and cattle. It should also be noted that in recent years outbreaks continue to be reported from the Benelux region to the UK and Ireland.

The overall objectives could be summarized into three catergories:
1) The elucidation of the structure of SBV: Determination of a high-resolution structure of Schmallenberg virus, as well as determining the structure of the individual protein components of the virus.

Determining the structure of SBV is important, as it would enable for the development of better antiviral therapies in order to help combat this disease. Furthermore, the elucidation of the overall structure of the virus would also help in identifying other structurally related viruses in circulation. While the fellow was not successful in this part during the fellowship, he is continuing this line of research during his year-long incorporation into the host laboratory of Dr. Abrescia.

2) The development of novel DNA vaccines to prevent/reduce SBV infection: We contributed to the development of novel DNA vaccine candidates, confirm their expression in vivo, followed by validating their degree of protection in small animal models. This part of the project proved to be highly successful, with two novel DNA vaccine candidates having shown to protect experimental mice from SBV-induced illness.

Currently, the use of SBV vaccines among European livestock producers is very limited, due to the cost of the vaccine, as well as the need for constant re-vaccination among herds. As DNA vaccines are considerably cheaper to produce, and their effects having been shown to be long-lasting, novel DNA vaccines to combat SBV infection would greatly encourage prophylactic programs to protect European livestock from this disease. With this in mind, the results that we have obtained during this fellowship may contribute to addressing this increasingly pressing agricultural problem.

3) The development of a rapid diagnostic test for the detection of SBV, including the development of monoclonal antibodies against components of the virus.

As Schmallenberg virus is rapidly transmitted through insect vectors, large-scale outbreaks have been shown to occur in a very short period of time. Therefore, the development of a rapid diagnostic test, which could be used by scientists and non-scientists alike, would enable for the rapid identification of the presence of SBV, and would enable for containment measures to enact immediately after SBV detection. While such a diagnostic kit was not developed during the two-year fellowship, the fellow is still continuing this portion of this project, during his year-long incorporation into the host laboratory of Dr. Abrescia.
1) Recombiant Protein Production:
In this Work package, the fellow was to use various protein expression systems to generate recombinant SBV proteins for use in WP2 and WP4. During the past two years, different strategies were used to complete this section; multiple protein designs were used to express proteins in both bacterial and mammalian cell lines.

2) WP2- Crystallography

In this Work package, the recombinant SBV proteins generated from WP1 were to be seeded in various buffer of varying salinity and pH, with the intent of finding the optimal conditions for protein crystallization. The generated crystals would then be subjected to X-ray diffraction, with the resulting data to be used to deduce their respective protein structure.

3) WP3- SBV Vaccine
In this Work package, several DNA vaccine candidates were generated, based on the cloning of several portions of the SBV genome. The vaccine candidates were then assayed in vitro, using HEK 293 cells, to determine their ability to express their respective cDNA. Following this step, IFNAR-/- mice were vaccinated twice (two weeks apart), then challenged with virulent SBV. Clinical signs were monitored for 2 weeks, and vaccine efficacy was determined by which vaccine groups exhibited minimal weight loss. In this Work package, we were able to find two separate DNA vaccine candidates and determine their mode of immunity. The results of this Work package was published in Antiviral Research earlier this year (2017). This publication was made Open Access, and deposited in ResearchGate for broad dissemination.

4) WP4- Monoclonal
In this Work package, the objective was to use the recombinant proteins generated in WP1 to inoculate mice for monoclonal antibody production, for subsequent use in the development of a rapid SBV diagnostic kit. Once the mice were injected multiple times with the recombinant proteins, spleens would be extracted for hybridoma production. The hybridomas would then be screened for monoclonal antibodies specific for the recombinant SBV proteins.
This project may ultimately impact the socio-economic aspects of the livestock industry. Considering the potential losses which may occur due to Schmallenberg disease, entire herds of sheep, goat and cattle may adversely affect the health and well-being of these animals. This, in turn, could severally impact local agrarian economies, as well as inflicting hardship on people employed in this industry. It is our hope that this first step in low-cost DNA vaccination may help contribute in limited such losses, and help protect this important sector of European agriculture.
Prevalence of Schmallenberg Virus (since its discover in 2011)