Bovine leukaemia virus: rapid detection of proviral DNA by nested PCR in blood and organs of experimentally infected calves

Abstract

The early stage of bovine leukaemia virus (BLV) infection was studied in experimentally infected calves in order to assess the diagnostic applicability of a double polymerase chain reaction (PCR). In addition, the kinetics of infection and virus distribution were evaluated. To simulate the natural route of virus transmission, the calves were infected by transferring two different infectious doses of whole blood from a BLV infected cow. The establishment of infection was determined by the double PCR and syncytia formation assay and by indirect serological methods including indirect ELISA, gp51/p24 ELISA, agar gel immunodiffusion (AGID) and Western blotting. BLV antibodies were first detected in ELISA on post infection (p.i.) day 26. Close agreement was found between the results of the various indirect methods. BLV infection was first detected in peripheral blood lymphocytes (PBL) by the PCR on p.i. day 7. No animal became seropositive to BLV prior to direct detection of BLV infection by the PCR. At slaughter, urine and saliva specimens as well as various organs were collected from the calves and tested by the double PCR. Several of the organs yielded positive results: e.g. spleen, uterus, liver, kidney, abomasum, and lymph nodes. Nine out of eleven spleen suspensions were positive by the PCR, including the spleen from one calf, which otherwise remained negative in all tests throughout the experiment. This phenomenon indicates that an animal may be infected without detectable levels of BLV proviral DNA in PBLs and without circulating antibodies, further emphasizing the diagnostic importance of the PCR. The findings indicate that the PCR is the most rapid method for the early detection of BLV infection in cattle and a valuable tool for studying the tropism of the virus.