The glassy-winged sharpshooter (Homalodisca vitripennis) is a highly vagile and polyphagous insect, which feeds on more than 100 plant species throughout the southwestern United States. Sharpshooters are the predominant vector of Xylella fastidiosa (Xf), a xylem-limited bacterium that is the causal agent of Pierce's disease (PD) of grapevine. Infected H. vitripennis transmit the bacterium while feeding. The rise of PD has been economically damaging for agriculture and H. vitripennis have become a target for disease control. A dicistrovirus identified as Homalodisca coagulata virus-01 (HoCV-01) has been associated with an increase in mortality rates within infected H. vitripennis populations. A host is required for HoCV-01 replication and cell culture provides the logistically and economically valuable means for producing a virus biopesticide. In this study, we developed a system for large-scale propagation of H. vitripennis cells via tissue culture, providing viral replication machinery. Cells were inoculated with low levels of HoCV-1, medium was removed every 24H for 168H, RNA extracted using TRIzol and analyzed with qRT-PCR. Cells were also trypan blue stained and counted to determine cell survivability. Whole virus particles were extracted within 72-96H after infection before total cell culture collapse occurred. This study shows that H. vitripennis cells are capable of being cultured and used for virus mass production, suitable to produce a biopesticide.
Date of publication
Biesbrock, Anna, "Propagation of Homalodisca Coagulata Virus-01 via Homalodisca vitripennis Cell Culture" (2013). Biology Theses. Paper 2.