Motif C in nonstructural protein 5 of duck Tembusu virus is essential for viral proliferation

Abstract

Nonstructural protein 5 (NS5) is required for duck Tembusu virus (DTMUV) genome replication, and the GDD motif in motif C is considered the hallmark of RdRp. However, the role of GDD-adjacent amino acids in motif C in viral proliferation is still unclear. To explore the role of motif C in the virus life cycle, DTMUV infectious clones and replicons were used to study the basic characteristics of rDTMUV through mutation of the amino acids in motif C. The replicon replication capability, virus titer, virus copy number, virulence and viral loads in organs were compared. Our results showed that V671A and V672A in motif C impaired DTMUV RNA replication in the replication system. Using an infectious clone system of DTMUV, we further demonstrated that the mutations of these two sites decreased viral titer and delayed the times of CPE appearance and duck embryo death. An in vivo study suggested that rDTMUV and DTMUV caused no obvious differences in ducklings. Similar clinical signs, including splenomegaly with hyperemia and hemorrhage dots of the thymus, were observed. There was no obvious difference in tissue viral loads between wild-type (rDTMUV-WT) and rDTMUV-NS5-V671A or rDTMUV-NS5-V672A. Determining the role of motif C can help in improving the understanding of the mechanism underlying DTMUV proliferation.