Replication and persistence of different strains of bovine viral diarrhea virus in an in vitro embryo production system

Recent studies have shown that exposed, in vitro-derived embryos remain con taminated with bovine viral diarrhea virus (BVDV) after washing. However, i ntroduction of a Genotype II versus Genotype I strain of BVDV into an IVF s ystem was reported to provide greater potential for transmission of disease . The primary objective of this study was to compare the potentials for dif ferent strains of noncytopathic BVDV to replicate in an IVF system, associa te with IVF embryos and infect co-cultured cells via association with washe d embryos. The secondary objective was to compare the effect of different s trains of BVDV on embryonic development. Two Genotype I (SD-I and NY-I) and 2 Genotype II (CD-87 and PA-131) strains of BVDV were evaluated. After IVM and IVF of oocytes, presumptive zygotes were washed and transferred into i n vitro cultures containing uterine tubal cells (UTC) and medium that was f ree of BVDV-neutralizing activity. Immediately before addition of zygotes, the cultures were inoculated with 10(5) cell culture infective doses (50%, CCID50) of a strain of BVDV or maintained as a negative control. Cultures o f zygotes were then incubated for 7 d. Embryonic development was observed o n Days 3 and 7, and attempts were made to isolate BVDV from UTC and medium on Day 7. Also on Day 7, groups of intact, washed blastocysts were either t ransferred into virus-free secondary cultures containing UTC or sonicated w ith sonicate fluid assayed by both virus isolation and single-closed-tube r everse transcription nested polymerase chain reaction (RT-nPCR). After 3 d in secondary culture, hatched embryos were enumerated, and medium from the cultures, washed UTC and embryos were tested for BVDV by virus isolation. I n addition, washed UTC and embryos were tested for BVDV using RT-nPCR. All strains of BVDV persisted and replicated in the embryo culture environment, but cleavage beyond the 4-cell stage, blastocyst development and hatching varied among cultures contaminated with different strains of virus. Further , the quantity of BVDV associated with washed embryos from both initial and secondary cultures varied among strains, but the variation was unrelated t o difference in genotype (SD-1 and PA-131 greater than NY-I and CD-87). Alt hough all strains of BVDV replicated in UTC in the initial in vitro culture s and remained associated with washed blastocysts, susceptible UTC in the s econdary in vitro cultures were seldom infected by any strain of virus. (C) 2000 by Elsevier Science Inc.