Rapid Generation of Attenuated Infectious Bursal Disease Virus from Dual-Promoter Plasmids by Reduction of Viral Ribonucleoprotein Activity
Rapid Generation of Attenuated Infectious Bursal Disease Virus from Dual-Promoter Plasmids by Reduction of Viral Ribonucleoprotein Activity
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通过降低病毒核糖核蛋白活性从双启动子质粒快速产生减毒传染性法氏囊病病毒
DOI：
10.1128/jvi.01569-19
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发表时间：
2020-01
影响因子：
5.4
通讯作者：
Ye Chengjin
中科院分区：
文献类型：
--
作者：
Yang Hui;Wang Yu;Ye Chengjin
传染性法氏囊病病毒（IBDV）感染引起的鸡的免疫抑制每年在世界范围内的家禽业中导致显著的经济损失。目前，疫苗接种仍是预防IBDV流行的最佳方法。然而，随着IBDV变异株数量的增加，研制抗原匹配的IBDV减毒活疫苗具有挑战性。在此，我们首先开发了一个双启动子反向遗传系统，用于快速产生IBDV。利用该系统，拯救了表现出降低的病毒RNP活性的表达VP 1-FLAG的减毒IBDV/1FLAG。此外，IBDV/1FLAG接种诱导的中和抗体水平与其亲本相似，保护鸡免受致死性攻击。我们的研究，第一次，描述了一种双启动子反向遗传方法，用于快速产生减毒IBDV，同时保持整个亲本抗原性，这表明一种潜在的新方法，以减毒流行性变异株IBDV疫苗开发。摘要传染性法氏囊病病毒（IBDV）是双RNA病毒科的一种病毒，它通过破坏法氏囊（BF）中的B细胞而导致雏鸡的免疫抑制。鉴于IBDV变异株数量的增加，我们迫切需要一种生产减毒病毒用于疫苗开发的方法。为了实现这一目标，开发了双启动子质粒用于快速产生IBDV，其中RNA聚合酶II和RNA聚合酶I（Pol I）启动子位于IBDV基因组序列的上游，随后是小鼠Pol I终止子和合成的聚腺苷酸化信号。这种方法不需要反式补充质粒来表达IBDV核糖核蛋白（RNP）的主要组分VP 1和VP 3。基于本研究发现VP 1-FLAG部分保留了IBDV RNP活性，我们成功地拯救了表达VP 1-FLAG的有复制能力的IBDV/1FLAG。与亲本病毒相比，IBDV/1FLAG在体外培养细胞中形成的空斑较小，但在体内诱导的免疫保护率相同。而IBDV/1FLAG免疫鸡既未观察到发育迟缓，也未观察到严重的BF病变。总的来说，这是第一份报告，病毒RNP活性的影响，通过添加一个表位标签上的组成病毒蛋白。此外，这项工作证明了从双启动子质粒通过在VP 1的C末端融合FLAG标签降低病毒RNP活性来快速产生减毒IBDV。这将是一种方便的策略，以减毒流行性变异IBDV毒株，用于快速和有效的疫苗开发。传染性法氏囊病病毒（IBDV）感染导致的鸡免疫抑制每年在全球家禽业中导致重大经济损失。目前，疫苗接种仍是预防IBDV流行的最佳方法。然而，随着IBDV变异株数量的增加，研制抗原匹配的IBDV减毒活疫苗具有挑战性。在此，我们首先开发了一个双启动子反向遗传系统，用于快速产生IBDV。利用该系统，拯救了表现出降低的病毒RNP活性的表达VP 1-FLAG的减毒IBDV/1FLAG。此外，IBDV/1FLAG接种诱导的中和抗体水平与其亲本相似，保护鸡免受致死性攻击。我们的研究，第一次，描述了一种双启动子反向遗传方法，用于快速产生减毒IBDV，同时保持整个亲本抗原性，这表明一种潜在的新方法，以减毒流行性变异株IBDV疫苗开发。
Immunosuppression in chickens as a result of infectious bursal disease virus (IBDV) infection leads to significant economic losses in the poultry industry worldwide every year. Currently, vaccination is still the best way to prevent the prevalence of IBDV. However, with the occurrence of increasing numbers of variant IBDV strains, it is challenging to develop antigen-matched live attenuated vaccine. Here, we first developed a dual-promoter reverse-genetic system for the rapid generation of IBDV. Using this system, the attenuated IBDV/1FLAG expressing VP1-FLAG, which displays the decreased viral RNP activity, was rescued. Moreover, IBDV/1FLAG inoculation induced a similar level of neutralizing antibodies to that of its parental counterpart, protecting chickens against lethal challenge. Our study, for the first time, describes a dual-promoter reverse-genetic approach for the rapid generation of attenuated IBDV while maintaining entire parental antigenicity, suggesting a potential new method to attenuate epidemic variant IBDV strains for vaccine development. ABSTRACT Infectious bursal disease virus (IBDV) of the Birnaviridae family leads to immunosuppression of young chickens by destroying B cells in the bursa of Fabricius (BFs). Given the increasing number of variant IBDV strains, we urgently require a method to produce attenuated virus for vaccine development. To accomplish this goal, the dual-promoter plasmids in which the RNA polymerase II and RNA polymerase I (Pol I) promoters were placed upstream of the IBDV genomic sequence, which was followed by mouse Pol I terminator and a synthetic polyadenylation signal, were developed for rapid generation of IBDV. This approach did not require trans-supplementation of plasmids for the expression of VP1 and VP3, the main components of IBDV ribonucleoprotein (RNP). Based on the finding in this study that the IBDV RNP activity was partially retained by VP1-FLAG, we successfully rescued the replication-competent IBDV/1FLAG expressing VP1-FLAG. Compared with its parental counterpart, IBDV/1FLAG formed smaller size plaques in cultured cells and induced the same 100% immune protection in vivo. However, neither retarded development nor severe BFs lesion was observed in the IBDV/1FLAG-inoculated chickens. Collectively, this is the first report that viral RNP activity was affected by the addition of an epitope tag on the componential viral proteins. Furthermore, this work demonstrates the rapid generation of attenuated IBDV from dual-promoter plasmids via reducing viral RNP activity by a fused FLAG tag on the C terminus of VP1. This would be a convenient strategy to attenuate epidemic variant IBDV strains for rapid and efficient vaccine development. IMPORTANCE Immunosuppression in chickens as a result of infectious bursal disease virus (IBDV) infection leads to significant economic losses in the poultry industry worldwide every year. Currently, vaccination is still the best way to prevent the prevalence of IBDV. However, with the occurrence of increasing numbers of variant IBDV strains, it is challenging to develop antigen-matched live attenuated vaccine. Here, we first developed a dual-promoter reverse-genetic system for the rapid generation of IBDV. Using this system, the attenuated IBDV/1FLAG expressing VP1-FLAG, which displays the decreased viral RNP activity, was rescued. Moreover, IBDV/1FLAG inoculation induced a similar level of neutralizing antibodies to that of its parental counterpart, protecting chickens against lethal challenge. Our study, for the first time, describes a dual-promoter reverse-genetic approach for the rapid generation of attenuated IBDV while maintaining entire parental antigenicity, suggesting a potential new method to attenuate epidemic variant IBDV strains for vaccine development.
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影响因子：
3.7
作者：
Méndez F;de Garay T;Rodríguez D;Rodríguez JF
通讯作者：
Rodríguez JF
影响因子：
3.7
作者：
Nouën CL;Toquin D;Müller H;Raue R;Kean KM;Langlois P;Cherbonnel M;Eterradossi N
通讯作者：
Eterradossi N
影响因子：
5.4
作者：
Huang, ZH;Elankumaran, S;Samal, SK
通讯作者：
Samal, SK
影响因子：
5.3
作者：
Zuo-ren Wang;N. Day;P. Trifillis;M. Kiledjian
通讯作者：
Zuo-ren Wang;N. Day;P. Trifillis;M. Kiledjian
糖皮质激素诱导的亮氨酸拉链在传染性法氏囊病病毒 (IBDV) 诱导的 I 型干扰素表达抑制和通过与 VP4 相互作用增强宿主细胞中 IBDV 生长中的关键作用
影响因子：
5.4
作者：
Li, Zhonghua;Wang, Yongqiang;Zheng, Shijun J.
通讯作者：
Zheng, Shijun J.