Animals and blood collection
The blood samples of BLV-infected and BLV-uninfected cattle, which were used in this study, were obtained from several farmers and veterinarians in Japan. BLV infection was confirmed by detection of the provirus using nested polymerase chain reaction (PCR) that targets the viral LTR and by detection of the anti-BLV antibody using a commercial ELISA kit (JNC, Tokyo, Japan), as described previously [19]. The blood samples of BLV-infected pregnant cattle (animals #1–#5, Table 1) were collected at two points before delivery, on the day of delivery (day 0), and on three points after delivery (days 1, 7, and 14). Blood collection on day 0 was performed before parturition.
For estradiol administration to BLV-infected cattle, 5 BLV-infected cattle (animals #6–#10, Table 1) were kept in an animal facility at the Faculty of Veterinary Medicine, Hokkaido University (Sapporo, Hokkaido, Japan, animals #6 and #7), and a biosafety level I animal facility at the Animal Research Center, Agricultural Research Department, Hokkaido Research Organization (Shintoku, Hokkaido, Japan, animals #8–#10). These animal experiments were approved by the Ethics Committee of the Faculty of Veterinary Medicine, Hokkaido University, and the Ethics Committee of the Animal Research Center, Agricultural Research Department, Hokkaido Research Organization.
Blood samples were shipped immediately after collection and stored at 4°C until the experiment. After 24 hours from collection, blood samples were used for the following analyses.