Strawberry mottle virus (SMoV) significantly compromises strawberry fruit quality and yield. The Pro2Glu protein of SMoV, an RNA silencing suppressor, is pivotal for viral infection; however, its interactions with host factors remain underexplored. This study identifies a critical interaction between SMoV Pro2Glu and strawberry NADH dehydrogenase (ubiquinone) 1 β subcomplex subunit 9 (FaNDUFB9). Using yeast two-hybrid assays and in vivo protein interaction analyses, we confirm that amino acid residues 31-45 of FaNDUFB9 are essential for binding Pro2Glu. Sequence analysis places FaNDUFB9 within the Complex1_LYR_superfamily, and subcellular localisation reveals its presence in both the mitochondria and nucleus. Silencing of NbNDUFB9 in Nicotiana benthamiana elevated Pro2Glu expression, whereas co-expression of FaNDUFB9 with Pro2Glu suppressed Pro2Glu levels and weakened its silencing activity. Furthermore, overexpression of FaNDUFB9 in transgenic N. benthamiana reduced transient SMoV RNA expression. These findings suggest that FaNDUFB9 inhibits expression of Pro2Glu, affecting its silencing activity, and thereby potentially further influencing viral infection.

N6-methyladenosine (m6A) is a common epigenetic modification found in eukaryotic RNA. Recent studies have increasingly highlighted the importance of m6A modification in plant defense against viruses. In this investigation, we found that prunus necrotic ringspot virus (PNRSV) infection affected the m6A modification process. Plant transcriptomes and epitranscriptomes were sequenced to coanalyze the dynamic changes of m6A modifications after PNRSV infection. Further studies revealed that the silencing of evolutionarily conserved C-terminal region (ECTs), encoding m6A readers, led to increased PNRSV accumulation, indicating that ECTs confer resistance to PNRSV. Additionally, we demonstrated that UPF3 and SMG7, which are involved in non-sense-mediated mRNA decay pathways, as well as phenylalanine ammonia-lyase (PAL), a well-known key enzyme in plant defense and an identified m6A-modified gene following PNRSV infection, play crucial roles in regulating PNRSV infection. These findings provide new insights into understanding PNRSV infection and further elucidate the role of m6A in modulating viral infection in plants.

The coat protein (CP) of plant viruses generally has multiple functions involving infection, replication, movement and patho-genicity. Functions of the CP of prunus necrotic ringspot virus (PNRSV), the causal agent of several threatening diseases of Prunus fruit trees, are poorly studied. Previously, we identified a novel virus in apple, apple necrotic mosaic virus (ApNMV), which is phylogenetically related to PNRSV and probably associated with apple mosaic disease in China. Full-length cDNA clones of PNRSV and ApNMV were constructed, and both are infectious in cucumber (Cucumis sativus L.), an experimental host. PNRSV exhibited higher systemic infection efficiency with more severe symptoms than ApNMV. Reassortment analysis of genomic RNA segments 1-3 found that RNA3 of PNRSV could enhance the long-distance movement of an ApNMV chimaera in cucumber, indicating the association of RNA3 of PNRSV with viral long-distance movement. Deletion mutagenesis of the PNRSV CP showed that the basic motif from amino acids 38 to 47 was crucial for the CP to maintain the systemic movement of PNRSV. Moreover, we found that arginine residues 41, 43 and 47 codetermine viral long-distance movement. The findings demonstrate that the CP of PNRSV is required for long-distance movement in cucumber, which expands the functions of ilarvirus CPs in systemic infection. For the first time, we identified involvement of Ilarvirus CP protein during long-distance movement.

A runner, as an elongated branch, develops from the axillary bud (AXB) in the leaf axil and is crucial for the clonal propagation of cultivated strawberry (Fragaria x ananassa Duch.). Runner formation occurs in at least two steps: AXB initiation and AXB outgrowth. HANABA TARANU (HAN ) encodes a GATA transcription factor that affects AXB initiation in Arabidopsis and promotes branching in grass species, but the underlying mechanism is largely unknown. Here, the function of a strawberry HAN homolog FaHAN in runner formation was characterized. FaHAN transcripts can be detected in the leaf axils. Overexpression (OE) of FaHAN increased the number of runners, mainly by enhancing AXB outgrowth, in strawberry. The expression of the strawberry homolog of BRANCHED1 , a key inhibitor of AXB outgrowth in many plant species, was significantly downregulated in the AXBs of FaHAN -OE lines, whereas the expression of the strawberry homolog of SHOOT MERISTEMLESS, a marker gene for AXB initiation in Arabidopsis, was upregulated. Moreover, several genes of gibberellin biosynthesis and cytokinin signaling pathways were activated, whereas the auxin response pathway genes were repressed. Further assays indicated that FaHAN could be directly activated by FaNAC2, the overexpression of which in strawberry also increased the number of runners. The silencing of FaNAC2 or FaHAN inhibited AXB initiation and led to a higher proportion of dormant AXBs, confirming their roles in the control of runner formation. Taken together, our results revealed a FaNAC2-FaHAN pathway in the control of runner formation and have provided a means to enhance the vegetative propagation of cultivated strawberry.

Green Sichuan pepper (Zanthoxylum armatum) is an important economic fruit crop, which is widely planted in the southwest region of China. Recently, a serious disease, namely flower yellowing disease (FYD), broke out, and the virus of green Sichuan pepper nepovirus (GSPNeV) was identified to be highly correlated with the viral symptoms. Meanwhile, green Sichuan pepper idaeovirus (GSPIV) and green Sichuan pepper enamovirus (GSPEV) were also common viruses infecting green pepper. In our research, specific primers were designed according to the reported sequences of the three viruses, and a multiplex reverse transcription-polymerase chain reaction (RT-PCR) method for the simultaneous detection of GSPNeV, GSPIV, and GSPEV was established. The annealing temperature, extension time, and cycle number affecting the multiplex RT-PCR reaction were adjusted and optimized. Sensitivity analysis showed that the system could detect the three viruses simultaneously from the complementary deoxyribonucleic acid (cDNA) samples diluted by 10−3. The results of the ten samples detected by the multiplex RT-PCR system were consistent with the results of a single PCR, indicating that the method can be successfully used for rapid detection of field samples.

Several members of the genus Ilarvirus infect fruit trees and are distributed worldwide. Prunus necrotic ringspot virus (PNRSV) is one of the most prevalent viruses, causing significant losses. Cucumissativus can be infected by several ilarviruses, leading to obvious symptoms, including PNRSV, which suggests that cucumbers could be good hosts for the study of the pathogenesis of ilarviruses. Real-time quantitative PCR is an optimal choice for studying gene expression because of its simplicity and its fast and high sensitivity, while its accuracy is highly dependent on the stability of the reference genes. In this study, we assessed the stability of eleven reference genes with geNorm, NormFinder, ΔCt method, BestKeeper, and the ranking software, RefFinder. The results indicated that the combined use of EF1α and F-BOX was the most accurate normalization method. In addition, the host genes AGO1, AGO4, and RDR6 were selected to test the reliability of the reference genes. This study provides useful information for gene expression analysis during PNRSV infection and will facilitate gene expression studies associated with ilarvirus infection.

Dendrobium officinale is an important traditional Chinese medicinal herb, and the stem tissue is the main medicinal that is harvested from D. officinale. Recently, the first viroid was identified from D. officinale in China, and it has been named Dendrobium viroid (DVd). Whether DVd interferes with metabolic pathways in dendrobium plants and affects the medicinal value of the host is unknown. In this study, metabolomics data from stem tissues supported by transcriptome studies were used to investigate how metabolism modulate of D. officinale is altered by DVd infection. Our results show that metabolism of D. officinale is reprogrammed in many ways during DVd infection, and this is reflected by significant changes in the levels of flavonoids, alkaloids, and phenolic acids. Furthermore, we found that DVd infection significantly decreased the accumulation of flavonoids and alkaloid metabolites in infected stems, and the decreases in these metabolites appears to affect the medicinal components of the infected plants, weakening the host antiviral immune response as well. Conversely, phenolic acids occupy a larger proportion of the up-regulated metabolites from DVd infection in comparison with the mock-inoculated control, and the increase in the total phenolic acids may reflect the activation of the pathogen defense response in D. officinale. Taken together, our results provide an interesting overview and give a better understanding of the relationship between metabolism and DVd infection in the orchid D. officinale.

为科学防控草莓炭疽根腐病,本研究在调查全国12个草莓主产区的109株草莓根部腐烂样品的基础上,对导致草莓根部腐烂的病原菌进行菌落形态鉴定、分离纯化与分子鉴定、多基因序列(ITS-ACT-GAPDH-CAL-TUB2-CHS)联合鉴定以及致病性鉴定.结果表明:109株草莓根部腐烂样品中有27.5％的样品是由草莓炭疽根腐病病原菌胶孢炭疽菌复合种(Colletotrichium gloeosporioides complex)引起;胶孢炭疽菌复合种可分为3种生理小种,分别为暹罗炭疽菌(C.siamense)、隐秘炭疽菌(C.aenigma)与果生刺盘孢菌(C.fructicola);3种炭疽菌致病力由高到低依次为隐秘炭疽菌、果生刺盘孢菌和暹罗炭疽菌.其中隐秘炭疽菌发病较快,引起的病症较重.综上,本研究可为草莓炭疽根腐病病害防控提供有价值的信息.

Strawberry mottle virus (SMoV) is one of the most common viruses infecting strawberries, causing losses to fruit yield and quality. In this study, 165 strawberry leaf samples were collected from six provinces of China, 46 of which tested positive for SMoV. The complete genome sequences of 11 SMoV isolates were obtained from Liaoning (DGHY3, DGHY16-2, DGHY17, DGHY20-2, DGHY21, DGHY26-2), Shandong (SDHY1, SDHY5, SDHY31-2, SDHY33-2), and Beijing (BJMX7). The RNA1 and RNA2 nucleotide identities between the 11 Chinese isolates were 95.4-99.3% and 96.3-99.6%, respectively, and they shared 78.4-96.6% and 84.8-93.5% identities with the available SMoV isolates in GenBank. Recombination analysis revealed that Chinese isolate SDHY33-2 and Canadian isolates Ontario and Simcoe were recombinants, and recombination events frequently occurred in the 3' UTR of SMoV. Phylogenetic analysis showed that in an RNA1 tree, most Chinese isolates clustered into the same group while isolate DGHY17 clustered into another group together with Czech isolate C and three Canadian isolates. In an RNA2 tree, all Chinese isolates clustered into a single group. The phylogenetic analysis based on nucleotide sequences was consistent with the results based on coat protein (CP) and RNA-dependent RNA polymerase (RdRp). Further evolutionary analysis indicated that negative selection drives SMoV evolution, and gene flow plays a major role in genetic differentiation. Additionally, reassortment and recombination also influence the evolution of SMoV. To our knowledge, this is the first report of the complete genome of SMoV isolates from China and a detailed analysis of the SMoV population structure.

Apple (Malus domestica) fruits exhibiting bright stripe symptoms were identified in Weihai City, Shandong Province, China. To investigate the virome in the apple samples, the method of high throughput sequencing (HTS) was used to identify the viruses. It was found that the sequence of citrus concave gum-associated virus (CCGaV) was involved in the apple transcriptome dataset. The full-length genome of the CCGaV-Weihai isolate contained two segments, the RNA1 was 6674 nt in size containing a conserved RNA-dependent RNA polymerase (RdRp), and the RNA2 was ambisense, 2706 nt in length, encoding a movement protein (MP) and a coat protein (CP). Sequence alignment and phylogenetic analyses indicated that CCGaV-Weihai was more closely related to CCGaV-H2799 isolated from the apple host in the United States and distantly related to CCGaV-CGW2 from Citrus sinensis in Italy, indicating a possibly geographical and host differentiation of CCGaV isolates. This was the first identification and characterization of CCGaV infecting apples in China. Additionally, a rapid and sensitive reverse transcription recombinase polymerase amplification (RT-RPA) assay technique was established for CCGaV detection in apple plants. The RT-RPA of CCGaV was not affected by other common viruses in apple plants and is about 10-fold more sensitive than the conventional reverse transcription polymerase chain reaction method, which can be used in large-scale testing.