Persistent infection of Aedes albopictus C6/36 cells by Bunyamwera virus

Defective RNAs in mosquito cells persistently infected with Bunyamwera virus

“…2 (b) shows a slot blot analysis specifically designed to detect S mRNA in the caesium chloride-pelleted RNA prepared'from C6/36 cells, and demonstrates that S mRNA was abundant in the cells 48 h post-infection, after N synthesis could no longer be detected. Furthermore, total cellular RNA extracted from mosquito cells at times up to 72 h postinfection could be translated in vitro and the viral N protein was synthesized (data not shown), indicating that the S mRNA detected by slot blot analysis was intact and A persistently infected cell line was established essentially as described previously (Elliott & Wilkie, 1986) and showed similar characteristics in terms of fluctuating titres of released virus at different passages (Fig. 3), the production of plaque morphology variants and resistance to superinfection with Bunyamwera virus (data not shown).…”

“…The ratio of defective L segment R N A to full-length L R N A varied between different passages, and at passage 58 the defective R N A was much more abundant than the full-length L segment. No subgeno- Previously we have reported that the S R N A segment is the major R N A species detected in virus particles released from cells persistently infected with Bunyamwera virus (Elliott & Wilkie, 1986). Here we investigated which R N A s were packaged into nucleocapsids by isolating intracellular nucleocapsids on caesium chloride gradients and analysing the extracted R N A by Northern blotting.…”

“…Aedes albopictus mosquito cells (Kascsak & Lyons, 1978;Newton et al, 1981). A previous report from this laboratory (Elliott & Wilkie, 1986) showed that in persistently infected A. albopictus C6/36 cells there was an over-representation of the S RNA segment. In addition, some features, such as fluctuating titres of released virus, suggested the presence of defective interfering (DI) RNAs (Holland, 1990), and subgenomic RNAs derived from the L segment were noted.…”

“…All clones contained a prominent defective L RNA species; the shorter RNAs in cl and c3 had the same mobility on the gel, and were different from the RNAs in c5 and c8. Superinfection experiments (performed as described by Elliott & Wilkie, 1986) showed that c5 and c8 yielded less virus (3 × 10 4 p.f.u./ml and 1 x 104 p.f.u.l/ml respectively) than cl (2 x 105 p.f.u./ml) and c3 (4 x 107 p.f.u./ml); the yield of virus from normal C6/36 cells was 1 x 109 p.f.u./ml. Hence the resistance of the persistently infected cells to superinfection with Bunyamwera virus appeared to correlate with the amount of full-length L R N A present, and not with the amount or occurrence of defective L RNA.…”

A Bunyamwera Virus Minireplicon System in Mosquito Cells

“…Infection of mammalian cells leads to rapid shutoff of host protein synthesis and apoptosis of infected cells in the late stages of infection. Infection of C6/36 mosquito cells by BUN leads to persistent infection (9,18) without induction of apoptosis (unpublished data), a phenomenon similar to the biology of La Crosse virus in mosquito vectors (3). In order to study the differences in BUN replication between mammalian and mosquito cells, we have established a BUN minireplicon system in A. albopictus C6/36 cells.…”