Skip to Main Content
Meredith Maloney

Blocking Production of Equine Infectious Anemia Virus Using RNA Interference

Meredith Maloney, Sarah Canterberry, Susan Payne, and Charles Long

Department of Veterinary Physiology and Pharmacology, Texas A&M University, College Station, TX

Meredith Maloney

Purpose – Equine Infectious Anemia Virus (EIAV) causes a persistent and highly contagious infection in Equids that is spread through biting insects. There is no vaccine and no cure, thus the current method of control for an animal testing positive (Coggins test) is either lifelong quarantine or euthanasia. RNA interference (RNAi) is an evolutionarily conserved mechanism for post transcriptional degradation of messenger RNA in response to double stranded RNA of adequate homology. Endogenous expression vectors that produce double stranded RNA as short hairpin RNAs (shRNA) can target a specific gene and block translation of resulting mRNA. We believe that the endogenous expression of shRNAs targeting the EIAV genome will reduce the amount of virus shed by cells infected with EIAV.

Hypothesis – The incorporation of shRNA targeting Equine Infectious Anemia Virus will reduce viral shedding in a persistently infected line of FEA+ cells.

Methods – Feline adenocarcinoma cells (FEA) with a persistent infection of EIAV were used as an in vitro model. The cells were genetically modified using a recombinant lentiviral vector to express a shRNA targeting the EIAV genome and a DsRed florescent marker. After determining the number of genetically modified cells, supernatant was collected and tested for viral shedding using a reverse transcriptase (RT) assay.

Results – Targeting Pol2, GP2, Rev2, and SER2 dramatically reduced the amount of virus shed when compared to non-transgenic (Cntl) or transgenic FEA cells expressing a non-targeting shRNA (Luc). RT levels at day 8 were (Cntl= 1) 1.60, 0.01, 0.01, 0.03, and 0.02; at day 12, 0.89, 0.24, 0.02, 0.09, and 0.03; and at day 17, 0.70, 0.95, 0.11, 1.45, and 0.37, for Luc, Pol2, GP2, Rev2, and SER2 shRNAs, respectively.

Conclusions – Virus replication was effectively blocked, however, by day 17 RT activity increased indicating possible viral mutation from RNAi mediated block.