Submitted to: Virology
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 6/27/2017
Publication Date: 7/5/2017
Publication URL: http://handle.nal.usda.gov/10113/5801879
Citation: Falkenberg, S.M., Dassanayake, R.P., Neill, J.D., Ridpath, J.F. 2017. Improved detection of bovine viral diarrhea virus in bovine lymphoid cell lines using PrimeFlow RNA assay. Virology. 509(2017):260-265. doi. 10.1016/j.virol.2017.06.032.
Interpretive Summary: Infection with bovine viral diarrhea viruses (BVDV) results in losses for the beef and dairy industry. There are several tests that can identify animals that are infected with BVDV. While these tests are important, the ability to understand the changes observed at a single cell level that is infected with BVDV provides opportunities to better understand the immune dysregulation associated with BVDV. Recently a novel flow cytometry-based procedure was developed and we adapted the technique for detection of BVDV infection in individual lymphoid cells. This technique is a reproducible and reliable method to quantitate the amount of virus between different cells as well as identify uninfected cells. Using three bovine lymphoid cells lines with three different infection statuses, one was not infected with BVDV, one was infected with BVDV and one was dual infected with BVDV and bovine leukemia virus (BLV), we were able to demonstrate differences in the prevalence of cells infected in each cell line and differences in viral load.
Technical Abstract: Bovine viral diarrhea virus (BVDV) infections, whether as acute, persistent or contributing to co-infections, result in significant losses for cattle producers. BVDV can be identified by real-time PCR and ELISA, detection and quantification of viral infection at the single cell level is extremely difficult. Detection at the single lymphoid cell level is important due to the immunomodulation that accompanies BVDV infection. A novel PrimeFlow RNA assay using in-situ detection of BVDV was evaluated. The model used to develop this technique included three BL-3 cells lines with different infection statuses, one not infected with BVDV, one infected with BVDV and one dual infected with BVDV and bovine leukosis virus (BLV). Using RNA probes specific for the BVDV-2a Npro-Erns coding region, BVDV RNA was detected from both contaminated BL-3 cell lines by flow cytometry and fluorescent microscopy. This is the first report on in-situ detection of BVDV at the single-cell level.