Abstract
As baculoviruses usually have a narrow insecticidal spectrum, knowing the mechanisms by which they control the host-range is prerequisite for improvement of their applications as pesticides. In this study, from supernatant of culture cells transfected with DNAs of an Autographa californica multiple nucleopolyhedrovirus (AcMNPV) mutant lacking the antiapoptotic gene p35 (vAc∆P35) and a cosmid representing a fragment of Spodoptera exigua nucleopolyhedrovirus (SeMNPV), a viral strain was plaque-purified and named vAcRev. vAcRev had a broader host range than either vAc∆P35 or SeMNPV parental virus, being able to infect not only the permissive hosts of its parental viruses but also a nonpermissive host (Spodoptera litura). Genome sequencing indicated that vAcRev comprises a mixture of two viruses with different circular dsDNA genomes. One virus contains a genome similar to vAc∆P35, while in the other viral genome, a 24.4 kbp-fragment containing 10 essential genesis replaced with a 4 kbp-fragment containing three SeMNPV genes including a truncated Se-iap3 gene. RNA interference and ectopic expression assays found that Se-iap3 is responsible for the host range expansion of vAcRev, suggesting that Se-iap3 inhibits the progression of apoptosis initiated by viral infection and promotes viral propagation in hosts both permissive and non-permissive for AcMNPV and SeMNPV.
Highlights
As baculoviruses usually have a narrow insecticidal spectrum, knowing the mechanisms by which they control the host-range is prerequisite for improvement of their applications as pesticides
Most of the transfected cells underwent apoptosis, polyhedral inclusion bodies (PIBs) were observed in a few cells, indicating that productive infection was established in these cells (Fig. 1Aa, arrowhead)
When Sf9, SpLi-221, and Se301 cells were inoculated with the supernatants from the cotransfected cells, PIBs were produced in a few cells of all three cell lines (Fig. 1B, arrowheads), most cells underwent apoptosis
Summary
Obtaining a viral strain with a broader in vitro host range. To study the potential of the SeMNPV genome sustaining the replication of vAc∆P35, vAc∆P35 DNA and a SeMNPV cosmid library, which consists of 5 cosmids and represents the entire viral genome[28], were cotransfected into Sf9 cells. The above genomic analyses, together with the plaque morphology results, suggested that vAcRev-1 and vAcRev-2 cannot replicate individually in Sf9, Se301 and SpLi-221 cell lines due to the lack of essential genes and lack of ability to inhibit apoptosis, respectively. The silencing of vAcRev-IAP3 expression was confirmed by RT-PCR and western blot analysis (Fig. 5C) These results suggest that vAcRev-IAP3 acts as the primary helper to extend the host range of vAcRev, and Se111 does not appear to be necessary for replication of vAcRev but might increase the survival rate of infected cells. RT-PCR showed that vAcRev-iap[3] transcripts could be detected as early as 3 h p.i. in vAcRev-infected Hi5, Sf9, Se301 and SpLi-221 cells (Fig. 6A), suggesting that vAcRev-iap[3] might be an early gene that is transcribed before viral DNA replication. Our results suggested that Se-IAP3 might facilitate the vAc∆P35 replication
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have