Washing or washing and trypsin treatment is ineffective for removal of noncytopathic bovine viral diarrhea virus from bovine oocytes or embryos after experimental viral contamination of an in vitro fertilization system

The objectives of this study were to investigate the association of oocytes and embryos with bovine viral diarrhea virus (BVDV) in an IVF system and the possibility of rendering them free of infectious virus by sequential washing (10 washes) or by washing and trypsin (0.25%, 90 sec) treatment. In a series of experiments, a noncytopathic strain of BVDV (New York-1; 10(6) TCID50/ml) was introduced into the maturation, fertilization or embryo culture medium during the IVF procedure. The virus was isolated 1) from all 40 samples containing unfertilized washed or unwashed oocytes (n = 360); 2) from 46% of 52 samples of Day 7 embryos (n = 307) produced by IVF, which were washed 10 times; 3) from 66% of the 44 samples of unwashed embryos (n = 325); and 4) from 50% of the 20 samples of embryos (n = 111) exposed to washing and trypsin treatment. In addition, 5 heifers inoculated intravenously with single unwashed Day 7 embryos developed antibodies to BVDV. Three of 5 other heifers seroconverted following inoculation with single embryos that had been washed 10 times. Two of 4 heifers inoculated intravenously with 2 trypsin-treated embryos seroconverted, while the remaining 2 heifers which received a single embryo did not develop antibodies. In summary, our present study shows that under in vitro conditions, BVDV may be associated with groups of unwashed and washed oocytes and embryos, and may persist through the IVF process. The standard washing procedure with or without trypsin treatment is not fully effective in rendering IVF embryos free of BVDV.