To identify the molecular effects of Tobacco bushy top virus (TBTV) evolution on the degeneration of tobacco bushy top disease, three TBTV isolates with mild virulence were compared with wild-type TBTV to assess the translation of p35, which relies on a BYDV-like translation element (BTE) in a cap-independent manner. The in vitro expression of p35 in the mild isolates was only 20% to 40% of the expression observed in wt TBTV. Based on translation data from chimeric TBTV RNA, low-level p35 expression in the three mild isolates was associated with two regions: the 5′ terminal 500 nt region (RI) and the 3′ internal region (RV), which included the BTE. For the RV region, low level p35 expression was mainly associated with structural alterations of the BTE instead of specific sequence mutations within the BTE based on SHAPE structural probing and mutation analysis. Additionally, structural alteration of the TBTV BTE resulted from mutations outside of the BTE, implying structural complexity of the local region surrounding the BTE. This study is the first report on the structural alteration of the 3′ cap-independent translation element among different isolates of a given RNA virus, which is associated with variations in viral virulence.

BackgroundDestructive diseases caused by Tomato spotted wilt virus (TSWV) have been reported associated with many important plants worldwide. Recently, TSWV was reported to infect different hosts in China. It is of value to clone TSWV isolates from different hosts and examine diversity and evolution among different TSWV isolates in China as well as worldwide.MethodsRT-PCR was used to clone the full-length genome (L, M and S segments) of three new isolates of TSWV that infected different hosts (tobacco, red pepper and green pepper) in China. Identity of nucleotide and amino acid sequences among TSWV isolates were analyzed by DNAMAN. MEGA 5.0 was used to construct phylogenetic trees. RDP4 was used to detect recombination events during evolution of these isolates.ResultsWhole-genome sequences of three new TSWV isolates in China were determined. Together with other available isolates, 29 RNA L, 62 RNA M and 66 RNA S of TSWV isolates were analyzed for molecular diversity, phylogenetic and recombination events. This analysis revealed that the entire TSWV genome, especially the M and S RNAs, had major variations in genomic size that mainly involve the A-U rich intergenic region (IGR). Phylogenetic analyses on TSWV isolates worldwide revealed evidence for frequent reassortments in the evolution of tripartite negative-sense RNA genome. Significant numbers of recombination events with apparent 5′ regional preference were detected among TSWV isolates worldwide. Moreover, TSWV isolates with similar recombination events usually had closer relationships in phylogenetic trees.ConclusionsAll five Chinese TSWV isolates including three TSWV isolates of this study and previously reported two isolates can be divided into two groups with different origins based on molecular diversity and phylogenetic analysis. During their evolution, both reassortment and recombination played roles. These results suggest that recombination could be an important mechanism in the evolution of multipartite RNA viruses, even negative-sense RNA viruses.Electronic supplementary materialThe online version of this article (doi:10.1186/s12985-015-0457-3) contains supplementary material, which is available to authorized users.

Plant RNA viruses encode essential viral proteins that depend on the host translation machinery for their expression. However, genomic RNAs of most plant RNA viruses lack the classical characteristics of eukaryotic cellular mRNAs, such as mono-cistron, 5′ cap structure, and 3′ polyadenylation. To adapt and utilize the eukaryotic translation machinery, plant RNA viruses have evolved a variety of translation strategies such as cap-independent translation, translation recoding on initiation and termination sites, and post-translation processes. This review focuses on advances in cap-independent translation and translation recoding in plant viruses.

A barley mutant, 194, was observed to exhibit a leaf spot phenotype over the whole course of its growing period. In this study, the phenotype and antioxidant competence were studied in the lesion mimic mutant 194. Plant height was slightly higher in mutant 194 than in the wild type (WT). In addition, leaf spot per plant in mutant 194 was significantly higher than in WT. Antioxidant competence, as indicated by reactive oxygen species (ROS) accumulation, antioxidant enzyme activity, and the expression of antioxidant enzyme-encoding genes was also assessed in mutant 194. Compared to the WT, mutant 194 displayed a relatively higher accumulation of ROS, accompanied by lower activities of some antioxidant enzymes and downregulation of antioxidant enzyme-encoding genes. This demonstrated reduced antioxidant competence in mutant 194. The results suggested that this lower antioxidant competence of mutant 194 could lead to the accumulation of excessive ROS. This excess of ROS could induce programmed cell death and has the potential to promote disease resistance in mutant 194.

BackgroundDuring the past decade, tobacco bushy top disease, which is mainly caused by a combination of Tobacco bushy top virus (TBTV) and Tobacco vein-distorting virus (TVDV), underwent a sudden appearance, extreme virulence and degeneration of the epidemic in the Yunnan province of China. In addition to integrative control of its aphid vector, it is of interest to examine diversity and evolution among different TBTV isolates.Methods5’ and 3’ RACE, combined with one step full-length RT-PCR, were used to clone the full-length genome of three new isolates of TBTV that exhibited mild pathogenicity in Chinese fields. Nucleotide and amino acid sequences for the TBTV isolates were analyzed by DNAMAN. MEGA 5.0 was used to construct phylogenetic trees. RDP4 was used to detect recombination events during evolution of these isolates.ResultsThe genomes of three isolates, termed TBTV-JC, TBTV-MD-I and TBTV-MD-II, were 4152 nt in length and included one distinctive difference from previously reported TBTV isolates: the first nucleotide of the genome was a guanylate instead of an adenylate. Diversity and phylogenetic analyses among these three new TBTV isolates and five other available isolates suggest that ORFs and 3’UTRs of TBTV may have evolved separately. Moreover, the RdRp-coding region was the most variable. Recombination analysis detected a total of 29 recombination events in the 8 TBTV isolates, in which 24 events are highly likely and 5 events have low-level likelihood based on their correlation with the phylogenetic trees. The three new TBTV isolates have individual recombination patterns with subtle divergences in parents and locations.ConclusionsThe genome sizes of TBTV isolates were constant while different ORF-coding regions and 3’UTRs may have evolved separately. The RdRp-coding region was the most variable. Frequent recombination occurred among TBTV isolates. Three new TBTV isolates have individual recombination patterns and may have different progenitors.Electronic supplementary materialThe online version of this article (doi:10.1186/s12985-015-0340-2) contains supplementary material, which is available to authorized users.