In vitro interaction of bovine herpesvirus 1 with uterine tube epithelial cells and oocytes

The aims of this study were to assess in vitro if bovine oocytes and oviductal epithelial cells from slaughterhouses for in vitro fertilization use may be infected with bovine herpesvirus 1; to analyze whether the treatment with trypsin according to the International Embryo Transfer Society guideline is efficient to inactivate the bovine herpesvirus 1; to morphologically study the virus-oocyte interaction through optical microscopy. In this study, Madin Darby Bovine Kidney (MDBK) cells that were co-cultured with oocytes matured in vitro and exposed to bovine herpesvirus 1 showed a cytopathic effect. The nested polymerase chain reaction for the supernatant was positive for the bovine herpesvirus 1, thus suggesting that the cytopathic effect observed in the MDBK monolayer was seen due to virus replication and not because of any culture toxicity. It was also observed cytopathic effect and positive nested polymerase chain reaction in MDBK cells co-cultured with in vitro maturated oocytes free of virus, but that were co-cultured in uterine epithelial cells pre-infected with bovine herpesvirus 1 and washed or not with trypsin, demonstrating an oocyte contamination by the virus. When trypsin-washing efficacy was evaluated, we could observe that the trypsin treatment was not able to eliminate the bovine herpesvirus 1 of the oocytes, and it was not observed any morphological difference in the infected oocytes.

本研究旨在通过体外实验，评估屠宰场获取的、用于体外受精的牛卵母细胞与输卵管上皮细胞是否可感染牛疱疹病毒1型（bovine herpesvirus 1）；分析按照国际胚胎移植学会（International Embryo Transfer Society）指南采用胰酶处理是否可有效灭活牛疱疹病毒1型；并通过光学显微镜从形态学层面研究病毒与卵母细胞的相互作用。本研究中，与体外成熟卵母细胞共培养并暴露于牛疱疹病毒1型的马-达二氏牛肾（Madin Darby Bovine Kidney, MDBK）细胞出现了细胞病变效应。对培养上清液进行巢式聚合酶链式反应（nested polymerase chain reaction）检测，结果呈牛疱疹病毒1型阳性，这表明MDBK单层细胞中观察到的细胞病变效应源于病毒复制，而非培养体系毒性所致。研究同时发现，将未感染病毒的体外成熟卵母细胞与预先感染牛疱疹病毒1型的子宫上皮细胞共培养（无论是否经胰酶洗涤），其共培养体系中的MDBK细胞同样出现细胞病变效应，且巢式聚合酶链式反应检测结果为阳性，由此证明卵母细胞可被该病毒污染。在评估胰酶洗涤的病毒清除效果时，可观察到胰酶处理无法清除卵母细胞携带的牛疱疹病毒1型，且感染病毒的卵母细胞未出现任何形态学差异。