Analysis of the Role of Bradysiaimpatiens (Diptera: Sciaridae) as a Vector Transmitting Peanut Stunt Virus on the Model Plant Nicotiana benthamiana

Bradysia species, commonly known as fungus gnats, are ubiquitous in greenhouses, nurseries of horticultural plants, and commercial mushroom houses, causing significant economic losses. Moreover, the insects from the Bradysia genus have a well-documented role in plant pathogenic fungi transmission. Here, a study on the potential of Bradysia impatiens to acquire and transmit the peanut stunt virus (PSV) from plant to plant was undertaken. Four-day-old larvae of B. impatiens were exposed to PSV-P strain by feeding on virus-infected leaves of Nicotiana benthamiana and then transferred to healthy plants in laboratory conditions. Using the reverse transcription-polymerase chain reaction (RT-PCR), real-time PCR (RT-qPCR), and digital droplet PCR (RT-ddPCR), the PSV RNAs in the larva, pupa, and imago of B. impatiens were detected and quantified. The presence of PSV genomic RNA strands as well as viral coat protein in N. benthamiana, on which the viruliferous larvae were feeding, was also confirmed at the molecular level, even though the characteristic symptoms of PSV infection were not observed. The results have shown that larvae of B. impatiens could acquire the virus and transmit it to healthy plants. Moreover, it has been proven that PSV might persist in the insect body transstadially. Although the molecular mechanisms of virion acquisition and retention during insect development need further studies, this is the first report on B. impatiens playing a potential role in plant virus transmission.

Multiple cellular compartments engagement in Nicotiana benthamiana-peanut stunt virus-satRNA interactions revealed by systems biology approach

Key message PSV infection changed the abundance of host plant’s transcripts and proteins associated with various cellular compartments, including ribosomes, chloroplasts, mitochondria, the nucleus and cytosol, affecting photosynthesis, translation, transcription, and splicing. Abstract Virus infection is a process resulting in numerous molecular, cellular, and physiological changes, a wide range of which can be analyzed due to development of many high-throughput techniques. Plant RNA viruses are known to replicate in the cytoplasm; however, the roles of chloroplasts and other cellular structures in the viral replication cycle and in plant antiviral defense have been recently emphasized. Therefore, the aim of this study was to analyze the small RNAs, transcripts, proteins, and phosphoproteins affected during peanut stunt virus strain P (PSV-P)–Nicotiana benthamiana interactions with or without satellite RNA (satRNA) in the context of their cellular localization or functional connections with particular cellular compartments to elucidate the compartments most affected during pathogenesis at the early stages of infection. Moreover, the processes associated with particular cell compartments were determined. The ‘omic’ results were subjected to comparative data analyses. Transcriptomic and small RNA (sRNA)–seq data were obtained to provide new insights into PSV-P–satRNA–plant interactions, whereas previously obtained proteomic and phosphoproteomic data were used to broaden the analysis to terms associated with cellular compartments affected by virus infection. Based on the collected results, infection with PSV-P contributed to changes in the abundance of transcripts and proteins associated with various cellular compartments, including ribosomes, chloroplasts, mitochondria, the nucleus and the cytosol, and the most affected processes were photosynthesis, translation, transcription, and mRNA splicing. Furthermore, sRNA-seq and phosphoproteomic analyses indicated that kinase regulation resulted in decreases in phosphorylation levels. The kinases were associated with the membrane, cytoplasm, and nucleus components.

Peanut Stunt Virus and Its Satellite RNA Trigger Changes in Phosphorylation in N. benthamiana Infected Plants at the Early Stage of the Infection

Signaling in host plants is an integral part of a successful infection by pathogenic RNA viruses. Therefore, identifying early signaling events in host plants that play an important role in establishing the infection process will help our understanding of the disease process. In this context, phosphorylation constitutes one of the most important post-translational protein modifications, regulating many cellular signaling processes. In this study, we aimed to identify the processes affected by infection with Peanut stunt virus (PSV) and its satellite RNA (satRNA) in Nicotiana benthamiana at the early stage of pathogenesis. To achieve this, we performed proteome and phosphoproteome analyses on plants treated with PSV and its satRNA. The analysis of the number of differentially phosphorylated proteins showed strong down-regulation in phosphorylation in virus-treated plants (without satRNA). Moreover, proteome analysis revealed more down-regulated proteins in PSV and satRNA-treated plants, which indicated a complex dependence between proteins and their modifications. Apart from changes in photosynthesis and carbon metabolism, which are usually observed in virus-infected plants, alterations in proteins involved in RNA synthesis, transport, and turnover were observed. As a whole, this is the first community (phospho)proteome resource upon infection of N. benthamiana with a cucumovirus and its satRNA and this resource constitutes a valuable data set for future studies.

Complete Genome Sequences of Seven Peanut Stunt Virus Strains from Japan

We present the complete genomic RNA sequences of seven isolates of Peanut stunt virus discovered in diseased legume plants in various regions in Japan. These sequences and the published viral sequences were compared with respect to nucleotide percentages.

The Defense Response of Nicotiana benthamiana to Peanut Stunt Virus Infection in the Presence of Symptom Exacerbating Satellite RNA

Peanut stunt virus (PSV) is a widespread pathogen infecting legumes. The PSV strains are classified into four subgroups and some are defined by the association of satellite RNAs (satRNAs). In the case of PSV, the presence of satRNAs alters the symptoms of disease in infected plants. In this study, we elucidated the plant response to PSV-G strain, which occurs in natural conditions without satRNA. However, it was found that it might easily acquire satRNA, which exacerbated pathogenesis in Nicotiana benthamiana. To explain the mechanisms underlying PSV infection and symptoms exacerbation caused by satRNA, we carried out transcriptome profiling of N. benthamiana challenged by PSV-G and satRNA using species-specific microarrays. Co-infection of plants with PSV-G + satRNA increased the number of identified differentially expressed genes (DEGs) compared with the number identified in PSV-G-infected plants. In both treatments, the majority of up-regulated DEGs were engaged in translation, ribosome biogenesis, RNA metabolism, and response to stimuli, while the down-regulated DEGs were required for photosynthesis. The presence of satRNA in PSV-G-infected plants caused different trends in expression of DEGs associated with phosphorylation, ATP binding, and plasma membrane.