Recipient Organization
IOWA STATE UNIVERSITY
2229 Lincoln Way
AMES,IA 50011
Performing Department
Veterinary Diagnostic and Production Animal Medicine
Non Technical Summary
Bovine coronavirus (BCoV) is one of the leading causes of enterocolitis in young calves and has been associated with winter dysentery in adult animals. It has also been implicated as a cause of respiratory disease in cattle, and while the virus is routinely detected in the lungs of animals with clinical respiratory disease, its role in the bovine respiratory disease complex remains controversial. This is in large part due to a lack of physical evidence showing BCoV infection within microscopic lesions in the bovine respiratory tract. This proposal will evaluate lungs for microscopic pathology from animals where bovine coronavirus was detected in their lungs. Furthermore, we will then attempt to detect the virus within lesions through the use of immunohistochemistry as well as in situ hybridisation. Results from this research are expected to be the first peer-reviewed physicalevidence of bovine coronavirus infection of bovine lungs. This will change the way that bovine coronavirus is viewed and launch further investigations into pathogenesis and immunity.
Animal Health Component
(N/A)
Research Effort Categories
Basic
90%
Applied
(N/A)
Developmental
10%
Goals / Objectives
Bovine coronavirushasbeen implicated as a cause of respiratory disease in cattle, and while the virus is routinely detected in the lungs of animals with clinical respiratory disease, its role in the bovine respiratory disease complex remains controversial. This is in large part due to an inability to repeatably fulfill Koch's postulates, and more importantly, a dearth of physical evidence showing BCoV infection within microscopic lesions in the bovine respiratory tract.The major goal of this project is to provideevidence of bovine coronavirus infection and associated microscopic pathology within the bovine lung. This will be pursued by completion of the following objectives/aims:Evaluation of BCoV PCR positive lungs for BCoV-associated pathology (bronchitis, bronchiolitis, and bronchiolitis obliterans). Histopathology slides from identified bovine respiratory disease cases where BCoV was detected in the lung via PCR will each be evaluated by three diagnostic pathologists to characterize the pathology within respiratory epithelial cells, the proposed target of BCoV infection in the respiratory system.Evaluation of previously identified cases for evidence of active infection within lesions via immunohistochemistry (IHC) and fluorescence in situ hybridization (FISH). It is not enough to show that there are lesions in the lungs of BCoV PCR positive lungs, as there are several infectious agents capable of causing similar lesions (i.e. BRSV, or IBR). Therefore, we will probe the aforementioned histopathology slides from cases where BCoV was detected in the lung via PCR to identify BCoV antigen via IHC and then confirm the presence of the BCoV nucleic acid within these lesions via (FISH).
Project Methods
While our initial analyses of previous bovine coronavirus cases are promising, there is additional work to be done to test the previously stated hypothesis. First, the evaluation of additional cases both for respiratory pathology and IHC reactivity is necessary to increase the power of the study. Analysis will commence by cases with the lowest BCoV Ct in groups of 2 (<20, 20-22, 22-24, etc.) until a group is reached where cases are uniformly negative for BCoV IHC staining. All BCoV IHC positive cases will be further probed by BCoV FISH to provide an additional level of infection confirmation.The scoring of microscopic lesions will be by the presence of epithelial attenuation and bronchiolitis obliterans (yes/no) within the evaluated sections of lung. Additionally, IHC staining will be reported as positive or negative. Grading will not be applied to any of the metrics, as the sections of lungs for these cases are from cases submitted from the field and are often inconsistent in terms of lung lobe sampled, tissue preservation, and the number of sections of lung on each evaluated slide. Histopathology and IHC evaluation will be performed by diagnostic pathologists Rahe, Magstadt, and Siepker. Pathologists will be blinded to the reported results of the other pathologists and following tabulation of the scores, if there is not a consensus between the pathologists on a given metric, a majority of 2/3 will decide the final result.Considering the initial progress, the histopathology evaluation and IHC results are not expected to take more than 3 months. There is not a currently available FISH for BCoV. However, the development of FISH assays is a common practice at the ISU VDL and between development, validation, and evaluation of identified BCoV positive cases this is not expected to take more than one calendar year.