Location: Animal Disease Research Unit
2024 Annual Report
Objectives
Malignant catarrhal fever (MCF) is an often-fatal viral disease that affects primarily ruminants. The disease is caused by gamma herpesviruses in the MCF group of the Macavirus genus. These viruses are carried asymptomatically by certain animal species that serves as virus reservoirs but can cause disease when transmitted to other non-adapted, susceptible species. Ovine herpesvirus-2 (OvHV-2), transmitted by sheep, is globally distributed and the most frequent cause of MCF worldwide. In North America, sheep-associated MCF is a leading cause of death in American bison, which are highly susceptible to the disease. Overall, MCF results in significant economic impact to agriculture not only due to the loss of animals but also because it imposes restrictions to multispecies operations. With no treatment or vaccines available, currently the only way to control the disease is separating carrier and susceptible animals. Therefore, availability of an effective vaccine to MCF and reliable diagnostic assays are top priorities for the agricultural industry. Using rabbits as a laboratory animal model, we have recently demonstrated that protection to MCF can be achieved when using recombinant viral vectors carrying OvHV-2 genes as potential vaccines. In this project we propose to test the safety, efficacy, and duration of protection of selected vaccine candidates in relevant livestock species. We also plan to improve disease control by developing a point-of-care serological assay for detection of MCF virus reservoirs.
Objective 1: Assess the safety and efficacy of new MCF vaccine platforms and delivery systems.
Sub-objective 1.A: Evaluate the safety of immunogen formulations to deliver OvHV-2 gB as an SA-MCF vaccine in cattle and bison.
Sub-objective 1.B: Determine the efficacy of a vaccine targeting OvHV-2 gB in preventing SA-MCF in bison.
Sub-objective 1.C: Assess the risk of SA-MCF vaccinated bison to develop MCF upon exposure to OvHV-2 infected sheep.
Objective 2: Improve MCF diagnosis by developing new optimized diagnostic assays.
Approach
This project focuses on two important aspects of malignant catarrhal fever (MCF) research: Objective 1 is development of a safe and effective sheep-associated (SA)-MCF vaccine for clinically susceptible species and Objective 2 is improvement of diagnostic capabilities for detection of MCF virus reservoirs. In Sub-objective 1.A, we will test a recombinant viral vectored vaccine expressing ovine herpesvirus-2 (OvHV-2) glycoprotein B (gB) in cattle and bison to determine its safety and immunogenicity. Vaccination will be done with the viral vector in adjuvant alone or following prime immunization with DNA encoding OvHV-2 gB. Experimental animals will be monitored for potential adverse effects from immunizations and development of immune responses to the OvHV-2 antigen. Our hypothesis is that immunization with the vaccine formulations and regimes tested does not cause adverse reactions in cattle or bison. In Sub-objective 1.B, we will determine the efficacy of a vaccine in preventing SA-MCF in bison. The hypothesis for this sub-objective is that immunization with a vaccine targeting OvHV-2 gB protects bison from SA-MCF following challenge with a lethal dose of OvHV-2. Experimental animals will be challenged at different times post-vaccination to evaluate the duration of protection induced by vaccination. In Sub-objective 1.C, we will assess the risk of SA-MCF vaccinated bison to develop MCF upon exposure to OvHV-2 infected sheep. This will be a field trial to test the hypothesis that bison immunized with the SA-MCF vaccine have reduced risk of developing MCF than non-vaccinated animals when exposed to OvHV-2 infected sheep. Vaccinated and non-vaccinated control bison will be exposed to a flock of OvHV-2 infected sheep maintained at close proximity for at least 3 months. Development of disease in both groups will be used to calculate protection rates and risks. In Objective 2 we will improve MCF diagnosis by developing a point-of-care serological assay for detection of MCFV antibodies. The goal is to validate and optimize a lateral flow immunoassay using recombinant OvHV-2 gB coupled to quantum dots as antigen. This is expected to result in a rapid, simple, sensitive, and inexpensive method to identify infected animals that can serve as viral reservoirs.
Progress Report
This report documents FY 2024 progress for project 2090-32000-045-000D, titled, “Development of a Vaccine and Improved Diagnostics for Malignant Catarrhal Fever”, which began October 2021.
Under Sub-objective 1.A, ARS researchers in Pullman, Washington, made considerable progress in evaluating immune responses induced by vaccination with AlHV-1/OvHV-2-gB in cattle. Characterization of humoral responses involved quantification of anti-OvHV-2 gB antibodies in the blood and respiratory tract. For characterization of cellular responses, they identified the phenotype of proliferating cells and cytokine production profiles. These findings indicate robust humoral and cellular immune responses to the vaccine candidate, supporting further evaluation of vaccine efficacy in target species such as cattle and bison.
In support to Sub-objective 1.B, ARS researchers in Pullman, Washington, in collaboration with the University of Wyoming, conducted a vaccine/challenge trial in bison to assess the efficacy of the AlHV-1/OvHV-2-gB vaccine candidate. All vaccinated animals developed antibody responses against OvHV-2 glycoprotein B, including neutralizing antibodies in the respiratory tract. Despite sustained antibody levels in the blood until the end of the experiment at 98 days post-prime immunization, vaccinated animals were not protected from malignant catarrhal fever upon challenge with a lethal dose of OvHV-2. This outcome prompted modification of the vaccine formulation to include additional OvHV-2 proteins (gH and gL) known to induce virus-blocking antibodies. The construction of the updated vaccine vector has been initiated, and subsequent trials in bison to evaluate its efficacy will be performed once the new vaccine candidate is available.
Under Objective 2, which aims to improve diagnostic capabilities for detecting anti-MCF virus antibodies in carrier species, ARS researchers in Pullman, Washington, initially focused on developing a lateral flow assay. Despite numerous efforts, production of a protein suitable for the assay encountered challenges due to biochemical characteristics hindering its purification from host cell components. Consequently, the research shifted towards optimizing a currently available competitive enzyme linked immunosorbent assay (ELISA), to address performance issues that limit its adoption and commercial availability. As a first step in the optimization of the ELISA, researchers are evaluating the use of a recombinant protein coupled to a catcher molecule to replace whole virus preparations currently used as antigen. This strategy is expected to improve the ELISA by enhancing its consistency and reproducibility. Also, progress has been made in characterizing monoclonal antibodies against malignant catarrhal fever (MCF) viruses generated in the laboratory. Modifications in the recombinant protein and better characterization of monoclonal antibodies are expected to result in the identification of valuable reagents to be used in improved diagnostic assays for MCF.
ARS researchers in Pullman, Washington, also made substantial progress in a subordinate project that aims at characterizing immune responses in MCF vaccinated and OvHV-2 challenged animals using nascent transcription start site mapping. In collaboration with Washington State University, Pullman, Washington, researchers obtained RNA sequence libraries from bison vaccinated with AlHV-1/OvHV-2-gB and/or infected with OvHV-2 and are currently analyzing the data to identify genes and transcription factors that are differentially expressed or activated following immunizations and during MCF progression. Upon completion, results from this study are expected to establish a foundation for the identification of expression patterns associated with disease progression, potentially serving as genetic markers to assess vaccine efficacy.
Accomplishments
Review Publications
Xavier, A.A., Queiroz, G.R., Lisbôa, J.A., Cunha, C.W., Headley, S.A. 2023. Immunohistochemical identification of a malignant catarrhal fever virus in cattle with renal diseases from Paraná state, Southern Brazil: A retrospective epidemiological study. Tropical Animal Health and Production. 55. Article 344. https://doi.org/10.1007/s11250-023-03740-y.
Gianopulos, K.A., Makio, A.O., Pritchard, S.M., Cunha, C.W., Hull, M.A., Nicola, A.V. 2024. Herpes simplex virus 1 glycoprotein B from a hyperfusogenic virus mediates enhanced cell–cell fusion.. Viruses. 16(2). Article 251. https://doi.org/10.3390/v16020251.
Wainwright, S.H., Cunha, C.W., Webb, B., McGregor, B.L., Drolet, B.S., Welch, J.B. 2024. Reemerging/notifiable diseases to watch. Veterinary Clinics of North America. 40(2):317-335. https://doi.org/10.1016/j.cvfa.2024.01.007.
Herndon, D.R., Herndon, M.K., Cunha, C.W. 2024. Coding complete genome of a strain of ovine herpesvirus-2 associated with a clinical case of malignant catarrhal fever in a domestic lamb. Microbiology Resource Announcements. 13. Article e01107-23. https://doi.org/10.1128/mra.01107-23.
More, D.D., Baker, K.N., Shringi, S., Bastos, R.G., O'Toole, D., Donofrio, G., Cunha, C.W. 2024. Ovine herpesvirus 2 glycoprotein B complementation restores infectivity of a bovine herpesvirus 4 gB-null mutant. Pathogens. 13(3). Article 219. https://doi.org/10.3390/pathogens13030219.
