湖北麻鸭乙型肝炎病毒体内感染模型的建立

Establishment of an in vivo model for duck hepatitis B virus infection using Hubei duckling

目的建立标准化的湖北麻鸭乙型肝炎病毒（DHBV）动物模型，并初步观察拉米呋啶（3TC）体内抗DHBV的作用。方法将收集的转染细胞培养上清液纯化后作为体内实验感染的标准毒种，经腹腔接种2日龄雏鸭，连续观察50d，用Real-TimePCR检测血清中DHBVDNA出现和持续时间。药物组则于接种3d后给予3TC，检测观察期间感染鸭血清中病毒含量的变化情况。结果雏鸭实验感染率达到87．5％；雏鸭在接种后第7天出现病毒血症，血清病毒含量于第11天达到高峰值，后逐步降低，但在观察期内仍维持较高水平。在3TC治疗期间，感染鸭体内病毒血症处于较低水平，被感染肝细胞的数量也明显减少，且无明显的毒性作用；在停药3d后，病毒血症即出现反跳现象，此后又有上升趋势。结论通过接种转染细胞上清液成功建立湖北麻鸭乙型肝炎病毒动物模型，药物评估实验证明其良好的稳定性和实用性；该模型也为研究DHBV生物学特性提供了有力的手段。

Objective To develop a standard duck hepatitis B virus （DHBV） animal model using a local Hubei species of duck, Ma Ya, and use it as an in vivo experimental system to study antiviral strategies against hepatitis B. Methods Two-day-old Ma Ya ducklings were experimentally infected via intraperitoneal injection with the DHBV inocula, which was collected from the transfected culture superuatant of 1.5-fold-overlength genome recombinant plasmid. Blood samples were taken twice or thrice a week during post-inoculation for 50 days. Viremia was quantified by serum real-time PCR to show the peak. Antiviral treatment of the DHBV-infected ducklings was started 3 days post-inoculation. The animals received oral administration of lamivudine （3TC） at a dose of 25 mg·kg^-1·d^-1 for 5 days, followed by a maintenance therapy thrice weekly for 3 more weeks. Serum was quantified to show the viremia peak and liver biopsy specimens were analysed by Southern blotting and in-situ hybridization at the end of antiviral drug treatment, Results The experimental infection rate of 2-day-old ducklings was 87.5 %, Viremia started to be detectable on day 7 and reached a peak on day 11 post-inoculation, followed by a decrease and fluctuations. Four weeks of oral administration of 3TC led to a significant decrease in viremia peak during drug treatment. This effect was not sustained, as a rebound in viremia was observed after drug withdrawal, Similarly, the analysis of liver biopsies at the end of 3TC treatment showed a marked decrease in DHBV DNA, However, after drug withdrawal a rebound of intrahepatic DHBV DNA was observed in duck livers, Conclusion The Hubei duck model with experimental DHBV infection of transfected supernatant is more suitable for the hepadnavirus biologic research due to its stability and practicability.