Host-dependent differences during synergistic infection by Potyviruses with potato virus X.

SUMMARY A comparative analysis of the synergistic interaction between PVX and either PVY or TEV potyviruses was performed in Nicotiana benthamiana and N. tabacum plants. In each PVX/potyvirus combination, doubly infected plants developed much more severe symptoms than singly infected ones. However, while PVX accumulation increased in doubly infected N. tabacum plants compared with singly infected plants, the accumulation of PVX did not vary drastically in doubly infected N. benthamiana plants with respect to single infected ones. These findings suggest that the relationship between viral titre enhancement and synergism in PVX/potyvirus infections is host dependent. Since PVX and potyviruses contain suppressors of a plant antiviral defence system mediated by gene silencing, differences observed in the response of these two related hosts to PVX/potyvirus interactions might reflect the effect of these viruses on host specific antiviral defences.

Transient expression of homologous hairpin RNA causes interference with plant virus infection and is overcome by a virus encoded suppressor of gene silencing.

Specific post-transcriptional gene silencing (PTGS) of target genes can be induced in a variety of organisms by providing homologous double-stranded RNA (dsRNA) molecules. In plants, PTGS is part of a defense mechanism against virus infection. We have previously shown and patented that direct delivery to nontransgenic plants of dsRNA derived from viral sequences specifically interfere with virus infection. Here, we show that transient expression of constructs encoding hairpin RNA homologous to a rapidly replicating plant tobamovirus also interferes with virus multiplication in a sequence-dependent manner. A three-day lag period between delivery of hairpin RNA and virus into the same tissues completely block virus infectivity. Several hallmarks characteristic of PTGS were associated with viral interference mediated by hairpin RNA: high level of sequence identity between the hairpin RNA and the target RNA, presence of siRNAs in extracts derived from leaves infiltrated with hairpin RNA, and helper component-proteinase (HC-Pro) of potyviruses, a suppressor of PTGS, overcame interference. No evidence for a mobile silencing suppression signal induced by transient expression of HC-Pro was observed. The approach described here has the potential to be used as a versatile tool for studying the onset of PTGS in cases involving virus infection, in opposition to dsRNA-transgenic plants, which allow primarily for the study of PTGS maintenance.

Detection of Both Subgroups I and II of Cucumber Mosaic Cucumovirus and Their Satellite RNAs on Pepper in Argentina.

Pepper plants (Capsicum annuum L.) showing stunting, mild to severe mosaic, leaf narrowing and deformation, prominent veins, fruit atrophy or malformation, and necrotic patterns on leaves and fruits were collected from commercial and experimental farms from seven districts in the central western neighboring provinces of Mendoza and San Juan. Cucumber mosaic cucumovirus (CMV) was identified directly from field samples from crude RNA extracts by reverse transcription-polymerase chain reaction (RT-PCR) (1). The PCR-amplified products were analyzed by restriction enzyme digestion with PvuII and VspI to distinguish the CMV subgroups. Serological assays (double immunodiffusion tests and indirect triple antibody sandwich enzyme-linked immunosorbent assay) with subgroup-specific monoclonal and polyclonal antisera also differentiated between the subgroups, although with lower sensitivity than found with PCR. Both subgroups I and II were found, no mixed infections were detected, and subgroup membership was related to geographical distribution: subgroup-I samples were detected in San Juan sites (north), and samples collected from the southern area (Mendoza) were of subgroup II. Some samples were sap-transmitted and passed repeatedly through tobacco and pepper. Satellite RNAs (satRNAs) were detected from total RNA extracts by RT-PCR with specific primers (5'-end oligonucleotide: 5'GGGTTATATCTGCGTGAG3' and 5'CACGGAGATCAGCATAGC3' as 3'-end primer). One satRNA isolate from each area was amplified and cloned. Three and five clones, respectively, were sequenced, obtaining 313/315 nucleotide and 334 nucleotide consensus sequences that appear to be similar (98.2 and 98.9%, respectively) to CMV-Y satRNA. This is the first report of CMV subgrouping and detection of CMV satRNAs in Argentina. Reference: (1) H. Rizos et al. J. Gen. Virol. 73:2099, 1992.