Tissue distribution of Bovine Leukemia Virus proviral DNA in infected animals

SUAREZ ARCHILLA GUILLERMO; TRONO KARINA; RUIZ VANESA; ALVAREZ IRENE; TARJI SOPHIE; WILLEMS, L.

Lima

Congreso; 19th International Conference on Human Retrovirology: HTLV and Related Viruses.; 2019

International Association of Retrovirlogy

Introduction: Bovine leukemia virus (BLV) is a B-lymphotropic oncogenic retrovirus that induces a chronic disease in cattle, often causing persistent lymphocytosis (PL), with lymphosarcomas developing in up to 10% of infected animals. Although there is still no vaccine commercially available, our group has been performing animal trials using a live-attenuated BLV strain obtained by genetic modification of the wild-type strain. The vaccine proved to be infectious in cows and replicated at low levels, with the development of a competent immune response, conferring protection from wild-type challenge. Although the attenuated strain was not transmitted to calves, maternal colostrum provided passive immunity. Since biosafety of this kind of vaccine is extremely important, we have carried out additional experiments in order to add knowledge in this regard.Objective: To evaluate the distribution of BLV provirus in various organs and tissues after long term infections.Methods: Three cows were studied. Cow A was naturally infected with a field strain of BLV, cows B and C were experimentally infected with the wild type and the attenuated vaccine strains, respectively. After 10 years of infection, animals were euthanased. Blood and tissues from parenchymal organs, lymph nodes and squeletical muscles, including those for human consumption, were collected under sterile conditions. Genomic DNA was extracted from peripheral blood mononuclear cells (PBMCs) and tissues, and tested by nested PCR for BLV provirus detection and real time PCR (qPCR) for proviral load (PVL) quantification. Results: At the time of euthanasia, PVL in PBMCs was high in cows A and B (452767 and 76888 copies/µg DNA, respectively), and extremely low (less than 100 copies/ µg DNA) in cow C. In animals A and B, BLV provirus was detected in all analyzed tissues, with higher PVL in prescapular, precrural and supramammary lymph nodes, spleen, liver, uterus, bone marrow and heart. The provirus was also found in muscle tissues for human consumption although with lower proviral load values (700-3000 copies/ µg DNA). These cows were pregnant, and although proviral DNA was present in the udder fluid, it was undetectable in both embryos. BLV proviral DNA could not be detected in any tissue obtained from the cow infected with the attenuated strain.Conclusion: The undetectable levels of the attenuated strain after long term infection in organs, lymph nodes and muscle tissues indicates that it is safe and suggests that could be considered as a tool for BLV clearance under a food safety point of view. Even though BLV transmission by consumption is not a key point under discussion, the analysis of safety is a necessary step for deregulation of a modified strain.