Recently, begomovirus/betasatellite disease complexes were found to be associated with alphasatellites, and their presence modulated disease symptoms and/or viral DNA accumulation in infected plants. However, the biological functions of alphasatellites during begomovirus/betasatellite infections remain unclear. Tomato yellow leaf curl China virus (TYLCCNV) associated with a betasatellite (TYLCCNB) is a widespread monopartite begomovirus in China. In the Yunnan province of China, the TYLCCNV/TYLCCNB disease complex is found in association with an alphasatellite (TYLCCNA). In this study, in order to explain the mechanisms underlying TYLCCNV/TYLCCNB infection and reductions in viral DNA accumulation caused by TYLCCNA, we analyzed the transcriptome profiles of Nicotiana benthamiana seedlings challenged by TYLCCNV/TYLCCNB or TYLCCNV/TYLCCNB/TYLCCNA using RNA sequencing. In total, 2272 and 1207 differentially expressed genes (DEGs) were identified to respond to TYLCCNV/TYLCCNB and TYLCCNV/TYLCCNB/TYLCCNA infections, respectively. Compared with the DEGs in the TYLCCNV/TYLCCNB-infected N. benthamiana seedlings, the number of DEGs in plants co-infected with TYLCCNV/TYLCCNB + TYLCCNA was significantly reduced. Additionally, 36 DEGs were identified to be regulated by TYLCCNA, six of which were further analyzed using the virus-induced gene silencing (VIGS) approach. Silencing of these six TYLCCNA responsive DEGs caused more severe disease symptoms and higher viral DNA accumulation levels, suggesting that TYLCCNA responsive DEGs may attenuate TYLCCNV/TYLCCNB infection.

Monopartite begomoviruses and their associated satellites form unique disease complexes that have emerged as a serious threat to agriculture worldwide. It is well known that frequent recombination contributes to the diversification and evolution of geminiviruses. In this study, we identified a novel defective satellite molecule (RecSat) in association with Tobacco leaf curl Yunnan virus (TbLCYNV) in a naturally infected tobacco plant. Sequence analysis showed that Recsat comprises 754 nucleotides in size and is a chimera involving alphasatellite and betasatellite sequences, containing both betasatellite-conserved region and alphasatellite stem-loop structure. Recombination analysis revealed that RecSat has arisen from three independent recombination events likely involving Tomato yellow leaf curl China betasatellite, Ageratum yellow vein China betasatellite and Tobacco curly shoot alphasatellite. Co-inoculation of RecSat with TbLCYNV induced symptoms indistinguishable from those induced by TbLCYNV alone in Nicotiana benthamiana. Southern blot hybridization showed that RecSat could be trans-replicated stably in N. benthamiana plants by TbLCYNV, and impaired the accumulation of helper virus and co-inoculated alphasatellite. Our results provide the first evidence for recombination between two distinct types of satellites among geminivirus complex and highlight recombination as a driving force for geminivirus evolution.

Alphasatellites, which encode only a replication-associated protein (alpha-Rep), are frequently found to be non-essential satellite components associated with begomovirus/betasatellite complexes, and their presence can modulate disease symptoms and/or viral DNA accumulation during infection. Our previous study has shown that there are three types of alphasatellites associated with begomovirus/betasatellite complexes in Yunnan province in China and they encode three corresponding types of alpha-Rep proteins. However, the biological functions of alpha-Reps remain poorly understood. In this study, we investigated the biological functions of alpha-Reps in post-transcriptional gene silencing (PTGS) and transcriptional gene silencing (TGS) using 16c and 16-TGS transgenic Nicotiana benthamiana plants. Results showed that all the three types of alpha-Rep proteins were capable of suppressing the PTGS and reversing the TGS. Among them, the alpha-Rep of Y10DNA1 has the strongest PTGS and TGS suppressor activities. We also found that the alpha-Rep proteins were able to increase the accumulation of their helper virus during coinfection. These results suggest that the alpha-Reps may have a role in overcoming host defense, which provides a possible explanation for the selective advantage provided by the association of alphasatellites with begomovirus/betasatellite complexes.

The genus Begomovirus constitutes a group of devastating plant viruses causing significant economic losses in the production of agricultural crops. In the present study, conducted in Yunnan province, China, a novel monopartite begomovirus was identified from Bidens pilosa, Crassocephalum crepidioides, tomato, pepper, and common bean, showing severe disease symptoms. The complete genome of the virus shows the typical organization of monopartite begomoviruses and shares the highest nucleotide sequence identity (88.73–88.95%) with crassocephalum yellow vein virus (CraYVV). According to the species criteria of the genus Begomovirus, this virus is a novel Begomovirus species, which has since been named “bidens pilosa leaf crumple virus (BpLCrV)”. Recombination analysis revealed that the novel species is a potential recombinant begomovirus derived from CraYVV and ageratum leaf curl virus (ALCuV), and phylogenetic analysis showed that BpLCrV was clustered with tomato yellow leaf curl Thailand virus (TYLCTHV) from China. We successfully developed a BpLCrV infectious clone. Agrobacterium-mediated inoculation of the BpLCrV infectious clone could effectively infect Nicotiana benthamiana, Nicotiana glutinosa, and Datura stramonium and cause disease symptoms. Thus, in this study, BpLCrV was, for the first time, identified and characterized as a novel begomovirus that infects not only weeds but also different important crops, potentially threatening agricultural production.

The genus Begomovirus constitute a group of devastating plant viruses causing significant economic losses in the production of agricultural crops. In the present study, a novel monopartite begomovirus was identified from Bidens Pilosa, Crassocephalum crepidioides, tomato, pepper and common bean showing severe disease symptoms in the Yunnan province of China. The complete genome of the virus shows the typical organization of monopartite reoviruses and shares the highest nucleotide sequence identity (88.73-88.95%) with crassocephalum yellow vein virus (CraYVV). According to the species criteria of the genus Begomovirus, this virus is a novel Begomovirus specie which was then named as “bidens pilosa leaf crumple virus (BpLCrV)”. Recombination analysis revealed that the novel species is a potential recombinant begomovirus derived from CraYVV and ageratum leaf curl virus (ALCuV), and phylogenetic analysis showed that BpLCrV was clustered with tomato yellow leaf curl Thailand virus (TYLCTHV) from China. We successfully developed the BpLCrV infectious clone. Agrobacterium-mediated inoculation of the BpLCrV infectious clone could effectively infect Nicotiana benthamiana, Nicotiana glutinosa and Datura stramonium and caused disease symptoms. Thus, in this study BpLCrV was firstly identified and characterized as a novel begomovirus, it infects not only weeds but also different important crops, with potential threat to agricultural production.