Nonstructural protein NS4 of Rice Stripe Virus plays a critical role in viral spread in the body of vector insects

Rice stripe virus (RSV), a tenuivirus, is transmitted by small brown planthopper (SBPH) in a persistent-propagative manner. In this study, sequential infection of RSV in the internal organs of SBPH after ingestion of virus indicated that RSV initially infected the midgut epithelium, and then progressed to the visceral muscle tissues, through which RSV spread to the entire alimentary canal. Finally, RSV spread into the salivary glands and reproductive system. During viral infection, the nonstructural protein NS4 of RSV formed cytoplasmic inclusions in various tissues of viruliferous SBPH. We demonstrated that the ribonucleoprotein particles of RSV were closely associated with NS4-specific inclusions in the body of viruliferous SBPH through a direct interaction between NS4 and nucleoprotein of RSV. Moreover, the knockdown of NS4 expression due to RNA interference induced by dsRNA from NS4 gene significantly prevented the spread of RSV in the bodies of SBPHs without a significant effect on viral replication in continuous cell culture derived from SBPH. All these results suggest that the nonstructural protein NS4 of RSV plays a critical role in viral spread by the vector insects.