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Rubisco small subunit (RbcS), a core component with crucial effects on the structure and kinetic properties of the Rubisco enzyme, plays an important role in response to plant growth, development, and various stresses. Although Rbcs genes have been characterized in many plants, their muti-functions in soybeans remain elusive. In this study, a total of 11 GmRbcS genes were identified and subsequently divided into three subgroups based on a phylogenetic relationship. The evolutionary analysis revealed that whole-genome duplication has a profound effect on GmRbcSs . The cis-acting elements responsive to plant hormones, development, and stress-related were widely found in the promoter region. Expression patterns based on the RT-qPCR assay exhibited that GmRbcS genes are expressed in multiple tissues, and notably Glyma.19G046600 ( GmRbcS8 ) exhibited the highest expression level compared to other members, especially in leaves. Moreover, differential expressions of GmRbcS genes were found to be significantly regulated by exogenous plant hormones, demonstrating their potential functions in diverse biology processes. Finally, the function of GmRbcS8 in enhancing soybean resistance to soybean mosaic virus (SMV) was further determined through the virus-induced gene silencing (VIGS) assay. All these findings establish a strong basis for further elucidating the biological functions of RbcS genes in soybeans.

Soybean mosaic virus (SMV) is a potyvirus found worldwide in soybean (Glycine max). GmCYB5-4 is a strong candidate interactor of P3. In this study, we comprehensively analyzed the GmCYB5 family in soybeans, including its distribution on chromosomes, promoter analysis, conserved motifs, phylogenetic analysis, and expression patterns. We cloned the full-length GmCYB5-4 and examined its interaction with P3 in yeast, which was later confirmed using bimolecular fluorescence complementation (BiFc). We silenced GmCYB5-4 using a bean pottle mosaic viris (BPMV) based system to generate Sil CYB5-4 tissues, which surprisingly knocked down four isoforms of GmCYB5s for functional characterization. Sil CYB5-4 plants were challenged with the SC3 strain to determine its involvement in SMV infection. Silencing GmCYB5-4 increased SMV accumulation, indicating that GmCYB5-4 inhibited SMV proliferation. However, further experiments are needed to elucidate the mechanism underlying the involvement of GmCYB5-4 in SMV infection., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024. Published by Elsevier Inc.)

Background: Soybean mosaic disease caused by soybean mosaic virus (SMV) is one of the most devastating and widespread diseases in soybean producing areas worldwide. The WRKY transcription factors (TFs) are widely involved in plant development and stress responses. However, the roles of the GmWRKY TFs in resistance to SMV are largely unclear., Results: Here, 185 GmWRKYs were characterized in soybean (Glycine max), among which 60 GmWRKY genes were differentially expressed during SMV infection according to the transcriptome data. The transcriptome data and RT-qPCR results showed that the expression of GmWRKY164 decreased after imidazole treatment and had higher expression levels in the incompatible combination between soybean cultivar variety Jidou 7 and SMV strain N3. Remarkably, the silencing of GmWRKY164 reduced callose deposition and enhanced virus spread during SMV infection. In addition, the transcript levels of the GmGSL7c were dramatically lower upon the silencing of GmWRKY164. Furthermore, EMSA and ChIP-qPCR revealed that GmWRKY164 can directly bind to the promoter of GmGSL7c, which contains the W-box element., Conclusion: Our findings suggest that GmWRKY164 plays a positive role in resistance to SMV infection by regulating the expression of GmGSL7c, resulting in the deposition of callose and the inhibition of viral movement, which provides guidance for future studies in understanding virus-resistance mechanisms in soybean., (© 2024. The Author(s).)

Potyvirus genomes are expressed as polyproteins that are autocatalytically cleaved to produce 10 to 12 multifunctional proteins, among which P1 is the most variable. It has long been hypothesized that P1 plays role(s) in host adaptation and host specificity. We tested this hypothesis using two phylogenetically distinct potyviruses: soybean mosaic virus (SMV), with a narrow host range, and clover yellow vein virus (ClYVV), with a broader host range. When the full-length P1 cistron of SMV-N was replaced with P1 from ClYVV-No.30, the chimera systemically infected only SMV-N-permissive hosts. Hence, there were no changes in the host range or host specificity of the chimeric viruses. Despite sharing only 20.3% amino acid sequence identity, predicted molecular models of P1 proteins from SMV-N and ClYVV-No.30 showed analogous topologies. These observations suggest that P1 of ClYVV-No.30 can functionally replace P1 of SMV-N. However, the P1 proteins of these two potyviruses are not determinants of host specificity and host range., (© 2024. The Author(s), under exclusive licence to Springer-Verlag GmbH Austria, part of Springer Nature.)

RNA silencing is an innate immune mechanism of plants against invasion by viral pathogens. Artificial microRNA (amiRNA) can be engineered to specifically induce RNA silencing against viruses in transgenic plants and has great potential for disease control. Here, we describe the development and application of amiRNA-based technology to induce resistance to soybean mosaic virus (SMV), a plant virus with a positive-sense single-stranded RNA genome. We have shown that the amiRNA targeting the SMV P1 coding region has the highest antiviral activity than those targeting other SMV genes in a transient amiRNA expression assay. We transformed the gene encoding the P1-targeting amiRNA and obtained stable transgenic Nicotiana benthamiana lines (amiR-P1-3-1-2-1 and amiR-P1-4-1-2-1). Our results have demonstrated the efficient suppression of SMV infection in the P1-targeting amiRNA transgenic plants in an expression level-dependent manner. In particular, the amiR-P1-3-1-2-1 transgenic plant showed high expression of amiR-P1 and low SMV accumulation after being challenged with SMV. Thus, a transgenic approach utilizing the amiRNA technology appears to be effective in generating resistance to SMV., (© 2024. The Author(s), under exclusive licence to Springer Nature Switzerland AG.)

Soybean [Glycine max (L.) Merr.] is an important legume crop worldwide, which provides abundant plant protein and oil for human beings. Soybean mosaic virus (SMV) can cause serious damage to the yield and quality of soybean, but it is difficult to control SMV with chemicals, breeding SMV-resistant varieties has become the most effective way to control the disease. Therefore, it is important to identify SMV resistance genes from soybean resources and apply them to soybean breeding. In this study, the disease rates (DRs) of 219 soybean accessions to SMV strain SC7 in two environments were investigated. A high-density NJAU 355 K SoySNP array was used for genome-wide association study (GWAS) of DR. A 274 kb region on chromosome 15 (1,110,567 bp to 1,384,173 bp) was repeatedly detected in two environments. Six new significant single nucleotide polymorphisms (SNPs) on chromosome 15 were identified. Four of these six SNPs were located within two candidate genes, Glyma.15G015700 and Glyma.15G015800. The elite haplotype Glyma.15G015700 Hap I with low DR exhibited strong resistance to SC7. The expression of Glyma.15G015700 in the SMV-resistant accession increased significantly after inoculation with SC7. Furthermore, most of the proteins predicted to interact with Glyma.15G015700 are heat shock proteins, which have been shown to be related to disease resistance. In summary, new SMV resistance loci and a new candidate gene, Glyma.15G015700, were identified and might be utilized in further soybean disease resistance breeding., (© 2024. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.)

Soybean mosaic virus (SMV) is one of the main pathogens that can negatively affect soybean production and quality. To study the gene regulatory network of soybeans in response to SMV SC15, the resistant line X149 and susceptible line X97 were subjected to transcriptome analysis at 0, 2, 8, 12, 24, and 48 h post-inoculation (hpi). Differential expression analysis revealed that 10,190 differentially expressed genes (DEGs) responded to SC15 infection. Weighted gene co-expression network analysis (WGCNA) was performed to identify highly related resistance gene modules; in total, eight modules, including 2256 DEGs, were identified. Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analysis of 2256 DEGs revealed that the genes significantly clustered into resistance-related pathways, such as the plant-pathogen interaction pathway, mitogen-activated protein kinases (MAPK) signaling pathway, and plant hormone signal transduction pathway. Among these pathways, we found that the flg22, Ca 2+ , hydrogen peroxide (H 2 O 2 ), and abscisic acid (ABA) regulatory pathways were fully covered by 36 DEGs. Among the 36 DEGs, the gene Glyma.01G225100 (protein phosphatase 2C, PP2C) in the ABA regulatory pathway, the gene Glyma.16G031900 (WRKY transcription factor 22, WRKY22) in Ca 2+ and H 2 O 2 regulatory pathways, and the gene Glyma.04G175300 (calcium-dependent protein kinase, CDPK) in Ca 2+ regulatory pathways were highly connected hub genes. These results indicate that the resistance of X149 to SC15 may depend on the positive regulation of flg22, Ca 2+ , H 2 O 2 , and ABA regulatory pathways. Our study further showed that superoxide dismutase (SOD) activity, H 2 O 2 content, and catalase (CAT) and peroxidase (POD) activities were significantly up-regulated in the resistant line X149 compared with those in 0 hpi. This finding indicates that the H 2 O 2 regulatory pathway might be dependent on flg22- and Ca 2+ -pathway-induced ROS generation. In addition, two hub genes, Glyma.07G190100 ( encoding F-box protein) and Glyma.12G185400 (encoding calmodulin-like proteins, CMLs), were also identified and they could positively regulate X149 resistance. This study provides pathways for further investigation of SMV resistance mechanisms in soybean.

Soybean (Glycine max ) is an important oilseed, protein and biodiesel crop. It faces significant threats from bacterial, fungal and viral pathogens, which cause economic losses and jeopardises global food security. In this article, we explore the relationship between soybeans and these pathogens, focusing on the molecular responses that are crucial for soybeans defence mechanisms. Molecular responses involve small RNAs and specific genes, including resistance (R) genes that are pivotal in triggering immune responses. Functional genomics, which makes use of cutting-edge technologies, such as CRISPR Cas9 gene editing, allows us to identify genes that provide insights into the defence mechanisms of soybeans with the focus on using genomics to understand the mechanisms involved in host pathogen interactions and ultimately improve the resilience of soybeans. Genes like GmKR3 and GmVQ58 have demonstrated resistance against soybean mosaic virus and common cutworm, respectively. Genetic studies have identified quantitative trait loci (QTLs) including those linked with soybean cyst nematode, root-knot nematode and Phytophthora root and stem rot resistance. Additionally, resistance against Asian soybean rust and soybean cyst nematode involves specific genes and their variations in terms of different copy numbers. To address the challenges posed by evolving pathogens and meet the demands of a growing population, accelerated soybean breeding efforts leveraging functional genomics are imperative. Targeted breeding strategies based on a deeper understanding of soybean gene function and regulation will enhance disease resistance, ensuring sustainable agriculture and global food security. Collaborative research and continued technological advancements are crucial for securing a resilient and productive agricultural future.

Autophagy plays a critical role in nutrient recycling/re-utilizing under nutrient deprivation conditions. However, the role of autophagy in soybeans has not been intensively investigated. In this study, the Autophay-related gene 7 ( ATG7 ) gene in soybeans (referred to as GmATG7 ) was silenced using a virus-induced gene silencing approach mediated by Bean pod mottle virus (BPMV). Our results showed that ATG8 proteins were highly accumulated in the dark-treated leaves of the GmATG7 -silenced plants relative to the vector control leaves (BPMV-0), which is indicative of an impaired autophagy pathway. Consistent with the impaired autophagy, the dark-treated GmATG7 -silenced leaves displayed an accelerated senescence phenotype, which was not seen on the dark-treated BPMV-0 leaves. In addition, the accumulation levels of both H 2 O 2 and salicylic acid (SA) were significantly induced in the GmATG7 -silenced plants compared with the BPMV-0 plants, indicating an activated immunity. Consistently, the GmATG7 -silenced plants were more resistant against both Pseudomonas syringae pv. glycinea ( Psg ) and Soybean mosaic virus (SMV) compared with the BPMV-0 plants. However, the activated immunity in the GmATG7 -silenced plant was not dependent upon the activation of MPK3/MPK6. Collectively, our results demonstrated that the function of Gm ATG7 is indispensable for autophagy in soybeans, and the activated immunity in the GmATG7 -silenced plant is a result of impaired autophagy.

Soybean is a major legume crop that plays an important role in food production, industrial production, and animal husbandry. Here, we characterize a novel soybean-infecting monopartite geminivirus identified in China. Analysis of the contigs de novo assembled from sequenced small interfering RNAs, followed by PCR, cloning, and sequencing, the complete viral genome was determined to be 2782 nucleotides. The genome contains the conserved nonanucleotide sequence, TAATATTAC and other sequence features typical of the family Geminiviridae , and encodes two and four open reading frames in the virion-sense and the complementary-sense strands, respectively. Genome-wide pairwise identity analysis revealed that the novel virus shares less than 65.6% identity with previously characterized geminiviruses. Phylogenetic and recombination analysis indicated that this virus was placed in a unique taxon within the family Geminiviridae and potentially arose from recombination. An infectious clone of this virus was further constructed and its infectivity was tested in different species of plants. Successful infection and characteristic symptoms were observed in Glycine max , Nicotiana benthamiana , N. tabacum , N. glutinosa , and N. tabacum cv. Samsun plants. Taken together, this virus represents a member of an unclassified genus of the family Geminiviridae , for which the name soybean yellow leaf curl virus is proposed.

Soybean mosaic virus (SMV) is a severe soybean (Glycine max) pathogen. Here we characterize a soybean SMV resistance cluster (SRC) that comprises five resistance (R) genes. SRC1 encodes a Toll/interleukin-1 receptor and nucleotide-binding site (TIR-NBS [TN]) protein, SRC4 and SRC6 encode TIR proteins with a short EF-hand domain, while SRC7 and SRC8 encode TNX proteins with a noncanonical basic secretory protein (BSP) domain at their C-termini. We mainly studied SRC7, which contains a noncanonical BSP domain and gave full resistance to SMV. SRC7 possessed broad-spectrum antiviral activity toward several plant viruses including SMV, plum pox virus, potato virus Y, and tobacco mosaic virus. The TIR domain alone was both necessary and sufficient for SRC7 immune signaling, while the NBS domain enhanced its activity. Nuclear oligomerization via the interactions of both TIR and NBS domains was essential for SRC7 function. SRC7 expression was transcriptionally inducible by SMV infection and salicylic acid (SA) treatment, and SA was required for SRC7 triggered virus resistance. SRC7 expression was posttranscriptionally regulated by miR1510a and miR2109, and the SRC7-miR1510a/miR2109 regulatory network appeared to contribute to SMV-soybean interactions in both resistant and susceptible soybean cultivars. In summary, we report a soybean R gene cluster centered by SRC7 that is regulated at both transcriptional and posttranscriptional levels, possesses a yet uncharacterized BSP domain, and has broad-spectrum antiviral activities. The SRC cluster is special as it harbors several functional R genes encoding atypical TIR-NBS-LRR (TNL) type R proteins, highlighting its importance in SMV-soybean interaction and plant immunity., (© American Society of Plant Biologists 2021. All rights reserved. For permissions, please email: journals.permissions@oup.com.)

Bean pod mottle virus (BPMV) is a destructive virus that causes serious economic losses in many countries every year, highlighting the importance of its effective detection. In this study, we developed a fast reverse transcription-cross-priming amplification (RT-CPA) coupled with lateral flow dipstick (LFD) diagnostic method for BPMV detection. The RT-CPA-LFD assay that targets the coat protein gene of BPMV was highly specific against diagnosing four other common viruses transmitted by soybean seeds, i.e., Southern bean mosaic virus (SBMV), Tomato ringspot virus (ToRSV), Arabis mosaic virus (ArMV), and Tobacco ringspot virus (TRSV). The sensitivities of the real-time fluorescent RT-CPA and the RT-CPA-LFD assay were at least 50 pg/μl and 500 pg/μl, respectively. Despite a compromise in the limit of detection of the RT-CPA method compared with TaqMan-MGB real-time RT-PCR, our results demonstrated a notably better performance in the detection of field samples of BPMV-infested soybean seeds. With the advantages of efficiency and convenience by visual determination, the RT-CPA-LFD assay presents a potential application for the rapid and accurate detection of BPMV in routine tests., (© 2022. The Author(s).)

Medicines and agricultural biocides are often discovered using large phenotypic screens across hundreds of compounds, where visible effects of whole organisms are compared to gauge efficacy and possible modes of action. However, such analysis is often limited to human-defined and static features. Here, we introduce a novel framework that can characterize shape changes (morphodynamics) for cell-drug interactions directly from images, and use it to interpret perturbed development of Phakopsora pachyrhizi, the Asian soybean rust crop pathogen. We describe population development over a 2D space of shapes (morphospace) using two models with condition-dependent parameters: a top-down Fokker-Planck model of diffusive development over Waddington-type landscapes, and a bottom-up model of tip growth. We discover a variety of landscapes, describing phenotype transitions during growth, and identify possible perturbations in the tip growth machinery that cause this variation. This demonstrates a widely-applicable integration of unsupervised learning and biophysical modeling., (© 2021. The Author(s).)

Soybean mosaic virus (SMV) causes severe yield losses and seed quality reduction in soybean (Glycine max) production worldwide. Rsc4 from cultivar Dabaima is a dominant genetic locus for SMV resistance, and its mapping interval contains three nucleotide-binding domain leucine-rich repeat-containing (NLR) candidates (Rsc4-1, Rsc4-2, and Rsc4-3). The NLR-type resistant proteins were considered as important intracellular pathogen sensors in the previous studies. In this study, based on transient expression assay in Nicotiana benthamiana leaves, we found that the longest transcript of Rsc4-3 is sufficient to confer resistance to SMV, and CRISPR/Cas9-mediated editing of Rsc4-3 in resistant cultivar Dabaima compromised the resistance. Interestingly, Rsc4-3 encodes a cell-wall-localized NLR-type resistant protein. We found that the internal polypeptide region responsible for apoplastic targeting of Rsc4-3 and the putative palmitoylation sites on the N terminus are essential for the resistance. Furthermore, we showed that viral-encoded cylindrical inclusion (CI) protein partially localizes to the cell wall and can interact with Rsc4-3. Virus-driven or transient expression of CI protein of avirulent SMV strains is enough to induce resistance response in the presence of Rsc4-3, suggesting that CI is the avirulent gene for Rsc4-3-mediated resistance. Taken together, our work identified a unique NLR that recognizes plant virus in the apoplast, and provided a simple and effective method for identifying resistant genes against SMV infection., (Copyright © 2021 The Author. Published by Elsevier Inc. All rights reserved.)

Autophagy plays a critical role in nutrient recycling and stress adaptations. However, the role of autophagy has not been extensively investigated in crop plants. In this study, soybean autophagy-related gene 2 ( GmATG2 ) was silenced, using virus-induced silencing (VIGS) mediated by Bean pod mottle virus (BPMV). An accelerated senescence phenotype was exclusively observed for the GmATG2- silenced plants under dark conditions. In addition, significantly increased accumulation of both ROS and SA as well as a significantly induced expression of the pathogenesis-related gene 1 ( PR1 ) were also observed on the leaves of the GmATG2- silenced plants, indicating an activated immune response. Consistent with this, GmATG2 -silenced plants exhibited a significantly enhanced resistance to Pseudomonas syringae pv. glycinea (Psg) relative to empty vector control plants (BPMV-0). Notably, the activated immunity of the GmATG2- silenced plants was independent of the MAPK signaling pathway. The fact that the accumulation levels of ATG8 protein and poly-ubiquitinated proteins were significantly increased in the dark-treated GmATG2- silenced plants relative to the BPMV-0 plants indicated that the autophagic degradation is compromised in the GmATG2- silenced plants. Together, our results indicated that silencing GmATG2 compromises the autophagy pathway, and the autophagy pathway is conserved in different plant species.

TGA transcription factors (TFs) exhibit basal resistance in Arabidopsis , but susceptibility to a pathogen attack in tomatoes; however, their roles in soybean ( Glycine max ) to Soybean mosaic virus (SMV) are unknown. In this study, 27 TGA genes were isolated from a SMV hyper-susceptible soybean NN1138-2 , designated GmTGA1 ~ GmTGA27 , which were clustered into seven phylogenetic groups. The expression profiles of GmTGAs showed that the highly expressed genes were mainly in Groups I, II, and VII under non-induction conditions, while out of the 27 GmTGAs , 19 responded to SMV-induction. Interestingly, in further transient N. benthamiana -SMV pathosystem assay, all the 19 GmTGAs overexpressed did not promote SMV infection in inoculated leaves, but they exhibited basal resistance except one without function. Among the 18 functional ones, GmTGA8 and GmTGA19, with similar motif distribution, nuclear localization sequence and interaction proteins, showed a rapid response to SMV infection and performed better than the others in inhibiting SMV multiplication. This finding suggested that GmTGA TFs may support basal resistance to SMV even from a hyper-susceptible source. What the mechanism of the genes ( GmTGA8 , GmTGA19 , etc.) with basal resistance to SMV is and what their potential for the future improvement of resistance to SMV in soybeans is, are to be explored.

Most of R (resistance) genes encode the protein containing NBS-LRR (nucleotide binding site and leucine-rich repeat) domains. Here, N. benthamiana plants were used for transient expression assays at 3-4 weeks of age. We identified a TNL (TIR-NBS-LRR) encoding gene GmRUN1 that was resistant to both soybean mosaic virus (SMV) and tobacco mosaic virus (TMV). Truncation analysis indicated the importance of all three canonical domains for GmRUN1-mediated antiviral activity. Promoter-GUS analysis showed that GmRUN1 expression is inducible by both salicylic acid (SA) and a transcription factor GmDREB3 via the cis-elements as-1 and ERE (ethylene response element), which are present in its promoter region. Interestingly, GmRUN1 gDNA (genomic DNA) shows higher viral resistance than its cDNA (complementary DNA), indicating the existence of intron-mediated enhancement (IME) for GmRUN1 regulation. We provided evidence that intron2 of GmRUN1 increased the mRNA level of native gene GmRUN1 , a soybean antiviral gene SRC7 and also a reporter gene Luciferase, indicating the general transcriptional enhancement of intron2 in different genes. In summary, we identified an antiviral TNL type soybean gene GmRUN1 , expression of which was regulated at different layers. The investigation of GmRUN1 gene regulatory network would help to explore the mechanism underlying soybean-SMV interactions.

Background: A key issue for implementation of CRISPR-Cas9 genome editing for plant trait improvement and gene function analysis is to efficiently deliver the components, including guide RNAs (gRNAs) and Cas9, into plants. Plant virus-based gRNA delivery strategy has proven to be an important tool for genome editing. However, its application in soybean which is an important crop has not been reported yet. ALSV (apple latent spherical virus) is highly infectious virus and could be explored for delivering elements for genome editing., Results: To develop a ALSV-based gRNA delivery system, the Cas9-based Csy4-processed ALSV Carry (CCAC) system was developed. In this system, we engineered the soybean-infecting ALSV to carry and deliver gRNA(s). The endoribonuclease Csy4 effectively releases gRNAs that function efficiently in Cas9-mediated genome editing. Genome editing of endogenous phytoene desaturase (PDS) loci and exogenous 5-enolpyruvylshikimate-3-phosphate synthase (EPSPS) sequence in Nicotiana. benthamiana (N. benthamiana) through CCAC was confirmed using Sanger sequencing. Furthermore, CCAC-induced mutagenesis in two soybean endogenous GW2 paralogs was detected., Conclusions: With the aid of the CCAC system, the target-specific gRNA(s) can be easily manipulated and efficiently delivered into soybean plant cells by viral infection. This is the first virus-based gRNA delivery system for soybean for genome editing and can be used for gene function study and trait improvement., (© 2021. The Author(s).)

Key Message: In the soybean variant V94-5152, a BCMV-resistance gene was mapped near to the region of SMV-resistance Rsv4 locus, raising a possibility that V94-5152 may rely on Rsv4 locus to resist against both SMV and BCMV. Both Soybean mosaic virus (SMV) and Bean common mosaic virus (BCMV) can induce soybean mosaic diseases, but few studies have explored soybean resistance against BCMV so far. In this study, V94-5152, a soybean variant resistant to BCMV and SMV, was crossed with a susceptible cultivar, Williams 82 to map the resistance gene. By inoculating 292 F 2 individuals with a BCMV isolate HZZB011, a segregation ratio of 3 resistant: 1 susceptible was observed, suggesting that V94-5152 possesses a single-dominant resistance gene against BCMV-HZZB011. Bulk segregation analysis (BSA) then revealed that the resistance gene is closely linked to BARCSOYSSR_02_0617, a simple sequence repeat (SSR) marker on chromosome 2. Genotyping neighboring SSR markers among the 292 F 2 individuals enabled us to draw a genetic linkage map, which indicated that the BCMV-resistance gene is located 0.2 cM downstream of BARCSOYSSR_02_0617. Amplification and sequencing ten candidate genes (Glyma02g121300 to Glyma02g122200) around this marker then revealed four genes containing nonsynonymous changes or indels. Also, this location is near to the recently cloned SMV-resistance Rsv4 locus from the cultivar Peking. By obtaining ten more sequences of Rsv4 locus from cultivated and wild soybean materials, we further investigated the variation and evolutionary patterns of this virus-resistance locus. It was evident that positive selections had been acting on this locus, with one critical amino acid change (R55P) shared by all resistance soybeans tested., (© 2021. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.)

Soybean mosaic virus (SMV) is one of the most widespread and devastating viral diseases worldwide. The genetic architecture of qualitative resistance to SMV in soybean remains unclear. Here, the Rsvg2 locus was identified as underlying soybean resistance to SMV by genome-wide association and linkage analyses. Fine mapping results showed that soybean resistance to SMV strains G2 and G3 was controlled by a single dominant gene, GmST1, on chromosome 13, encoding a sulfotransferase (SOT). A key variation at position 506 in the coding region of GmST1 associated with the structure of the encoded SOT and changed SOT activity levels between RSVG2-S and RSVG2-R alleles. In RSVG2-S allele carrier "Hefeng25", the overexpression of GmST1 carrying the RSVG2-R allele from the SMV-resistant line "Dongnong93-046" conferred resistance to SMV strains G2 and G3. Compared to Hefeng25, the accumulation of SMV was decreased in transgenic plants carrying the RSVG2-R allele. SMV infection differentiated both the accumulation of jasmonates and expression patterns of genes involved in jasmonic acid (JA) signalling, biosynthesis and catabolism in RSVG2-R and RSVG2-S allele carriers. This characterization of GmST1 suggests a new scenario explaining soybean resistance to SMV., (© 2021 John Wiley & Sons Ltd.)

The host range of previously reported bymoviruses is restricted to plants belonging to the family Poaceae. Soybean leaf rugose mosaic virus (SbLRMV) from non-Poaceae plants is related to bymoviruses based on a partial genome sequence. However, unlike bymoviruses, this virus infects plants of at least four dicotyledonous families, including Fabaceae, and causes disease in soybean. Complete nucleotide sequences of two variants of SbLRMV were determined, and its taxonomic position was clarified. RNA1 is 7109 nucleotides (nt) long with one large open reading frame (ORF), possibly encoding a polyprotein of 257 kDa. This polyprotein is likely processed into eight mature proteins. The entire RNA1 ORF shares 52%-55% nucleotide sequence identity and 27%-43% amino acid sequence identity, and the coat protein shares 49%-54% nucleotide sequence identity and 30%-34% amino acid sequence identity to other bymoviruses. The similarity to other viruses in the family Potyviridae is generally lower. RNA2 is 3413 or 3415 nt long and putatively encodes a polyprotein of 108 kDa. This protein is probably cleaved into two mature proteins. The sequences of these two RNAs are very similar to those of bymoviruses. Phylogenetic analysis of members of the family Potyviridae showed that RNA1 and RNA2 of SbLRMV formed a basal clade with known bymoviruses. Inoculation tests using leaf samples suggested that SbLRMV RNA1 can systemically infect and cause disease in soybean without the presence of RNA2. In conclusion, SbLRMV is an atypical member of the genus Bymovirus that infects soybean (Fabaceae) and other dicots rather than gramineous hosts.

Common bean plants (Phaseolus vulgaris L.) showing different virus-like symptoms were collected in northwestern Argentina. Dot-blot hybridization tests showed that the begomoviruses bean golden mosaic virus and tomato yellow vein streak virus were the most prevalent, but they also revealed the presence of unknown begomoviruses. The complete genome sequence of one of these unknown begomoviruses was determined. Sequence analysis showed that the virus is a typical New World begomovirus, for which the name "bean bushy stunt virus" (BBSV) is proposed. Biological assays based on biolistic inoculations showed that BBSV induced leaf roll and stunting symptoms similar to those observed in the field-collected common bean sample.

In a comparative analysis of genome sequences from isolates of the baculovirus Chrysodeixis includens nucleopolyhedrovirus (ChinNPV) from Brazil and Guatemala, we identified a subset of isolates possessing chimeric genomes. We identified six distinct phylogenetically incongruous regions (PIRs) dispersed in the genomes, of between 279 and 3345 bp in length. The individual PIRs possessed high sequence similarity among the affected ChinNPV isolates but varied in coverage in some instances. The donor for four of the PIRs implicated in horizontal gene transfer (HGT) was identified as Trichoplusia ni single nucleopolyhedrovirus (TnSNPV), an alphabaculovirus closely related to ChinNPV, or another unknown but closely related virus. BLAST searches of the other two PIRs returned only ChinNPV sequences, but HGT from an unknown donor baculovirus cannot be excluded. Although Chrysodeixis includens and Trichoplusia ni are frequently co-collected from soybean fields in Brazil, pathogenicity data suggest that natural coinfection of C. includens larvae with ChinNPV and TnSNPV is probably uncommon. Additionally, since the chimeric ChinNPV genomes with tracts of TnSNPV sequence were restricted to a single monophyletic lineage of closely related isolates, a model of progressive restoration of the native DNA sequence by recombination with ChinNPV possessing a fully or partially non-chimeric genome is reasonable. However, multiple independent HGT from TnSNPV to ChinNPV during the evolution of these isolates cannot be excluded. Mortality data suggest that the ChinNPV isolates with chimeric genomes are not significantly different in pathogenicity towards C. includens when compared to most other ChinNPV isolates. Exclusion of the PIRs prior to phylogenetic analysis had a large impact on the topology of part of the maximum-likelihood tree, revealing a homogenous clade of three isolates (IB, IC and ID) from Paraná state in Brazil collected in 2006, together with an isolate from Guatemala collected in 1972 (IA), comprising the lineage uniquely affected by HGT from TnSNPV. The other 10 Brazilian ChinNPV isolates from Paraná, Mato Grosso, and Minas Gerais states showed higher variability, where only three isolates from Paraná state formed a monophyletic group correlating with geographical origin.

Plant viruses often infect several distinct host species. Sometimes, viruses can systemically infect a specific host whereas, in other cases, only local infections occur in other species. How viral and host factors interact to determine systemic infections among different hosts is largely unknown, particularly for icosahedral positive-stranded RNA viruses. The Tobacco necrosis virus-A Chinese isolate belongs to the genus Alphanecrovirus in the family Tombusviridae . In this study, we investigated variations in systemic infections of tobacco necrosis virus-A C (TNV-A C ) in Nicotiana benthamiana and Glycine max (soybean) by alanine-scanning mutagenesis of the viral coat protein (CP), which is essential for systemic movement of TNV-A C . We found that three amino acids, R 169 , K 177 , and Q 233 , are key residues that mediate varying degrees of systemic infections of N. benthamiana and soybean. Further analysis revealed that variations in systemic trafficking of TNV-A C CP mutants in N. benthamiana and soybean are associated with virion assembly and stability. The CP amino acids K 177 and Q 233 are highly conserved among all TNV-A isolates and are replaced by Q and K in the TNV-D isolates. We demonstrated that systemic infectivity of either TNV-A C K177A and Q233A or K177Q and Q233K mutants are correlated with the binding affinity of the mutated CPs to the host-specific Hsc70-2 protein. These results expand our understanding of host-dependent long-distance movement of icosahedral viruses in plants.[Formula: see text] Copyright © 2021 The Author(s). This is an open access article distributed under the CC BY-NC-ND 4.0 International license.

Soybean thrips ( Neohydatothrips variabilis ) are one of the most efficient vectors of soybean vein necrosis virus, which can cause severe necrotic symptoms in sensitive soybean plants. To determine which other viruses are associated with soybean thrips, the metatranscriptome of soybean thrips, collected by the Midwest Suction Trap Network during 2018, was analyzed. Contigs assembled from the data revealed a remarkable diversity of virus-like sequences. Of the 181 virus-like sequences identified, 155 were novel and associated primarily with taxa of arthropod-infecting viruses, but sequences similar to plant and fungus-infecting viruses were also identified. The novel viruses were predicted to have positive-sense RNA, negative-stranded RNA, double-stranded RNA, and single-stranded DNA genomes. The assembled sequences included 100 contigs that represented at least 95% coverage of a virus genome or genome segment. Sequences represented 12 previously described arthropod viruses including eight viruses reported from Hubei Province in China, and 12 plant virus sequences of which six have been previously described. The presence of diverse populations of plant viruses within soybean thrips suggests they feed on and acquire viruses from multiple host plant species that could be transmitted to soybean. Assessment of the virome of soybean thrips provides, for the first time, information on the diversity of viruses present in thrips.

Plant pathogens deliver virulence effectors into plant cells to modulate plant immunity and facilitate infection. Although species-specific virulence effector screening approaches have been developed for several pathogens, these assays do not apply to pathogens that cannot be cultured and/or transformed outside of their hosts. Here, we established a rapid and parallel screening assay, called the virus-induced virulence effector (VIVE) assay, to identify putative effectors in various plant pathogens, including unculturable pathogens, using a virus-based expression vector. The VIVE assay uses the potato virus X (PVX) vector to transiently express candidate effector genes of various bacterial and fungal pathogens into Nicotiana benthamiana leaves. Using the VIVE assay, we successfully identified Avh148 as a potential virulence effector of Phytophthora sojae. Plants infected with PVX carrying Avh148 showed strong viral symptoms and high-level Avh148 and viral RNA accumulation. Analysis of P. sojae Avh148 deletion mutants and soybean hairy roots overexpressing Avh148 revealed that Avh148 is required for full pathogen virulence. In addition, the VIVE assay was optimized in N. benthamiana plants at different developmental stages across a range of Agrobacterium cell densities. Overall, we identified six novel virulence effectors from seven pathogens, thus demonstrating the broad effectiveness of the VIVE assay in plant pathology research., (© 2020 The Authors. Molecular Plant Pathology published by British Society for Plant Pathology and John Wiley & Sons Ltd.)

Soybean mosaic virus (SMV), which belongs to the Potyviridae , causes significant reductions in soybean yield and seed quality. In this study, both tag-free and reporter gene green fluorescent protein ( GFP )-containing infectious clones for the SMV N1 strain were constructed by Gibson assembly and with the yeast homologous recombination system, respectively. Both infectious clones are suitable for agroinfiltration on the model host N. benthamiana and show strong infectivity for the natural host soybean and several other legume species. Both infectious clones were seed transmitted and caused typical virus symptoms on seeds and progeny plants. We used the SMV-GFP infectious clone to further investigate the role of key amino acids in the silencing suppressor helper component-proteinase (Hc-Pro). Among twelve amino acid substitution mutants, the co-expression of mutant 2-with an Asparagine→Leucine substitution at position 182 of the FRNK (Phe-Arg-Asn-Lys) motif-attenuated viral symptoms and alleviated the host growth retardation caused by SMV. Moreover, the Hc-Prom2 mutant showed stronger oligomerization than wild-type Hc-Pro. Taken together, the SMV infectious clones will be useful for studies of host-SMV interactions and functional gene characterization in soybeans and related legume species, especially in terms of seed transmission properties. Furthermore, the SMV-GFP infectious clone will also facilitate functional studies of both virus and host genes in an N. benthamiana transient expression system.

The Arabidopsis resistance protein RPS5 is activated by proteolytic cleavage of the protein kinase PBS1 by the Pseudomonas syringae effector protease AvrPphB. We have previously shown that replacing seven amino acids at the cleavage site of PBS1 with a motif cleaved by the NIa protease of turnip mosaic virus (TuMV) enables RPS5 activation upon TuMV infection. However, this engineered resistance conferred a trailing necrosis phenotype indicative of a cell-death response too slow to contain the virus. We theorized this could result from a positional mismatch within the cell between PBS1 TuMV , RPS5, and the NIa protease. To test this, we relocalized PBS1 TuMV and RPS5 to cellular sites of NIa accumulation. These experiments revealed that relocation of RPS5 away from the plasma membrane compromised RPS5-dependent cell death in Nicotiana benthamiana , even though PBS1 was efficiently cleaved. As an alternative approach, we tested whether overexpression of plasma membrane-localized PBS1 TuMV could enhance RPS5 activation by TuMV. Significantly, overexpressing the PBS1 TuMV decoy protein conferred complete resistance to TuMV when delivered by either agrobacterium or by aphid transmission, showing that RPS5-mediated defense responses are effective against bacterial and viral pathogens. Lastly, we have now extended this PBS1 decoy approach to soybean by modifying a soybean PBS1 ortholog to be cleaved by the NIa protease of soybean mosaic virus (SMV). Transgenic overexpression of this soybean PBS1 decoy conferred immunity to SMV, demonstrating that we can use endogenous PBS1 proteins in crop plants to engineer economically relevant disease resistant traits.

Soybean mosaic virus (SMV)-disease is one of the most serious and widespread diseases in soybean (Glycine max). In the present study, a DnaJ protein in soybean designated GmCPIP (SMV coat protein-interacting protein) was screened by the QIS-Seq (quantitative interactor screening with next-generation sequencing) method, and the interaction between SMV CP and GmCPIP was confirmed by the yeast two-hybrid (Y2H) system and bimolecular fluorescence complementation (BiFC) assay. Subcellular localization analysis indicated that both proteins are localized in the cytoplasm, cytomembrane and nucleus. Quantitative real-time polymerase chain reaction (qRT-PCR) analysis showed that infection with SMV-SC4 temporarily increased the transcription of GmCPIP. Virus-induced gene silencing (VIGS) down-regulated the GmCPIP gene by 82%, and the accumulation of SMV was decreased by 88.6% in GmCPIP-silenced plants inoculated with SMV-SC4. The interaction of GmCPIP with SMV CP seems to contribute to SMV infection in soybean., (Copyright © 2020 Elsevier B.V. All rights reserved.)

Resistance genes are an effective means for disease control in plants. They predominantly function by inducing a hypersensitive reaction, which results in localized cell death restricting pathogen spread. Some resistance genes elicit an atypical response, termed extreme resistance, where resistance is not associated with a hypersensitive reaction and its standard defense responses. Unlike hypersensitive reaction, the molecular regulatory mechanism(s) underlying extreme resistance is largely unexplored. One of the few known, naturally occurring, instances of extreme resistance is resistance derived from the soybean Rsv3 gene, which confers resistance against the most virulent Soybean mosaic virus strains. To discern the regulatory mechanism underlying Rsv3-mediated extreme resistance, we generated a gene regulatory network using transcriptomic data from time course comparisons of Soybean mosaic virus-G7-inoculated resistant (L29, Rsv3-genotype) and susceptible (Williams82, rsv3-genotype) soybean cultivars. Our results show Rsv3 begins mounting a defense by 6 hpi via a complex phytohormone network, where abscisic acid, cytokinin, jasmonic acid, and salicylic acid pathways are suppressed. We identified putative regulatory interactions between transcription factors and genes in phytohormone regulatory pathways, which is consistent with the demonstrated involvement of these pathways in Rsv3-mediated resistance. One such transcription factor identified as a putative transcriptional regulator was MYC2 encoded by Glyma.07G051500. Known as a master regulator of abscisic acid and jasmonic acid signaling, MYC2 specifically recognizes the G-box motif ("CACGTG"), which was significantly enriched in our data among differentially expressed genes implicated in abscisic acid- and jasmonic acid-related activities. This suggests an important role for Glyma.07G051500 in abscisic acid- and jasmonic acid-derived defense signaling in Rsv3. Resultantly, the findings from our network offer insights into genes and biological pathways underlying the molecular defense mechanism of Rsv3-mediated extreme resistance against Soybean mosaic virus. The computational pipeline used to reconstruct the gene regulatory network in this study is freely available at https://github.com/LiLabAtVT/rsv3-network., Competing Interests: The authors have declared that no competing interests exist.

Soybean vein necrosis virus (SVNV) was first identified in Arkansas and Tennessee in 2008 and is now known to be widespread in the United States and Canada. Multiple species of thrips transmit this and other tospoviruses with Neohydatothrips variabilis (Beach) (soybean thrips) cited as the most efficient vector for SVNV. In this study, 18 soybean, Glycine max (L.) Merr., genotypes were evaluated in four experiments by infesting plants with noninfected and SVNV-infected thrips using choice and no-choice assays. In both choice experiments with noninfected and SVNV-infected thrips, the lowest number of immature soybean thrips occurred on plant introductions (PIs) 229358 and 604464 while cultivars Williams 82 and Williamsfield Illini 3590N supported higher counts of mature thrips. The counts between the two assays (noninfected and SVNV-infected thrips) were positively correlated. In both no-choice experiments with noninfected and SVNV-infected thrips, counts of thrips did not differ by soybean genotypes. Further studies are needed to characterize the inheritance and mechanisms involved in the resistance found in the choice assay., (Published by Oxford University Press on behalf of Entomological Society of America 2019.)

Soybean mosaic virus (SMV), a potyvirus, is the most prevalent and destructive viral pathogen in soybean-planting regions of China. Moreover, other potyviruses, including bean common mosaic virus (BCMV) and watermelon mosaic virus (WMV), also threaten soybean farming. The eukaryotic translation initiation factor 4E (eIF4E) plays a critical role in controlling resistance/susceptibility to potyviruses in plants. In the present study, much higher SMV-induced eIF4E1 expression levels were detected in a susceptible soybean cultivar when compared with a resistant cultivar, suggesting the involvement of eIF4E1 in the response to SMV by the susceptible cultivar. Yeast two-hybrid and bimolecular fluorescence complementation assays showed that soybean eIF4E1 interacted with SMV VPg in the nucleus and with SMV NIa-Pro/NIb in the cytoplasm, revealing the involvement of VPg, NIa-Pro, and NIb in SMV infection and multiplication. Furthermore, transgenic soybeans silenced for eIF4E were produced using an RNA interference approach. Through monitoring for viral symptoms and viral titers, robust and broad-spectrum resistance was confirmed against five SMV strains (SC3/7/15/18 and SMV-R), BCMV, and WMV in the transgenic plants. Our findings represent fresh insights for investigating the mechanism underlying eIF4E-mediated resistance in soybean and also suggest an effective alternative for breeding soybean with broad-spectrum viral resistance., (© 2019 The Authors. Molecular Plant Pathology published by British Society for Plant Pathology and John Wiley & Sons Ltd.)

Soybean mosaic virus (SMV) is a major pathogen causing yield loss. Developing soybean plants tolerant or resistant to SMV is important for mitigating the adverse effects of the viral infection. However, most studies have focused on the resistance to normal SMV strains. Thus, investigations of the resistance or tolerance to the novel recombinant SMV strain have been limited. To address the threat of the recombinant SMV, two soybean parent genotypes with contrasting reactions to the recombinant SMV and 211 F 9:11 recombinant inbred lines were evaluated under artificial inoculation conditions. The JD12 plants are resistant to the recombinant SMV, whereas HT is highly tolerant, but still susceptible. Genetic analyses suggested that the resistance of JD12 is controlled by a single dominant gene and the tolerance is a quantitative trait. The QTL mapping results revealed one QTL (qTsmv-13) for resistance and two QTLs (qTsmv-2 and qTsmv-3) for tolerance. A comparison between known resistance genes and the QTLs identified in this study suggested that qTsmv-13 and qTsmv-2 may correspond to Rsv1 and Rsv4, respectively, whereas qTsmv-3 represents a newly identified QTL for SMV tolerance. We further delimited qTsmv-3 to an interval of approximately 86 kb with a map-based cloning strategy. Only two of five candidate genes, Glyma.03G00550 and Glyma.03G00570, varied between the parents. Additionally, Glyma.03G00550, which is a multidrug and toxic compound extrusion transporter gene, is the likely candidate gene for qTsmv-3. In summary, our research opens a new avenue for formulating strategies to breed soybean varieties tolerant to SMV., Competing Interests: Declaration of Competing Interest The authors declare that there are no conflicts of interest., (Copyright © 2019 Elsevier B.V. All rights reserved.)

Soybean vein necrosis virus (SVNV), the causal agent of the homonymous disease, is a ubiquitous virus in North America. The widespread presence of the virus has led to the hypothesis that mixed infections with other viruses could alter disease symptoms, localization in the plant and even epidemiology. The potential interaction between bean pod mottle virus (BPMV), soybean mosaic virus (SMV), the most economically important soybean viruses in the U.S., and SVNV was assessed in the work presented here. Results revealed that soybean, a local lesion host for SVNV, becomes permissive in the presence of BPMV; whereas there where no obvious interactions observed in mixed infections with SMV.

Orthotospoviruses are acquired by thrips during feeding on infected tissue. Virions travel through the foregut and enter midgut epithelial cells through the interaction between the viral glycoproteins and cellular receptors. Glycoprotein RGD motifs and N-linked glycosylation sites have been predicted to mediate receptor binding or play important roles in virus entry into host cells, yet their function needs to be validated. In this study, peptides derived from the soybean vein necrosis virus N glycoprotein were utilized to identify critical regions in virus-vector interactions. Transmission mediated by single Neohydatothrips variabilis dropped by more than 2/3 when thrips were fed on peptide NASIAAAHEVSQE or the combination of NASIRGDHEVSQE and RLTGECNITKVSLTN when compared to the controls; indicating that this strategy could significantly reduce transmission efficiency, opening new avenues in the control of diseases caused by orthotospoviruses.

Soybean mosaic virus and Clover yellow vein virus are two definite species of the genus Potyvirus within the family Potyviridae . Soybean mosaic virus-N (SMV-N) is well adapted to cultivated soybean ( Glycine max ) genotypes and wild soybean ( G. soja ), whereas it remains undetectable in inoculated broad bean ( Vicia faba ). In contrast, clover yellow vein virus No. 30 (ClYVV-No. 30) is capable of systemic infection in broad bean and wild soybean; however, it infects cultivated soybean genotypes only locally. In this study, SMV-N was shown to also infect broad bean locally; hence, broad bean is a host for SMV-N. Based on these observations, it was hypothesized that lack of systemic infection by SMV-N in broad bean and by ClYVV-No. 30 in cultivated soybean is attributable to the incompatibility of multifunctional helper-component proteinase (HC-Pro) in these hosts. The logic of selecting the HC-Pro cistron as a target is based on its established function in systemic movement and being a relevant factor in host range specificity of potyviruses. To test this hypothesis, chimeras were constructed with precise exchanges of HC-Pro cistrons between SMV-N and ClYVV-No. 30. Upon inoculation, both chimeras were viable in infection, but host range specificity of the recombinant viruses did not differ from those of the parental viruses. These observations suggest that (i) HC-Pro cistrons from SMV-N and ClYVV-No. 30 are functionally compatible in infection despite 55.6 and 48.9% nucleotide and amino acid sequence identity, respectively, and (ii) HC-Pro cistrons from SMV-N and ClYVV-No. 30 are not the determinants of host specificity on cultivated soybean or broad beans, respectively.

Key Message: Genome-wide association analyses identified candidates for genes involved in restricting virus movement into embryonic tissues, suppressing virus-induced seed coat mottling and preserving yield in soybean plants infected with soybean mosaic virus. Soybean mosaic virus (SMV) causes significant reductions in soybean yield and seed quality. Because seedborne infections can serve as primary sources of inoculum for SMV infections, resistance to SMV seed transmission provides a means to limit the impacts of SMV. In this study, two diverse population panels, Pop1 and Pop2, composed of 409 and 199 soybean plant introductions, respectively, were evaluated for SMV seed transmission rate, seed coat mottling, and seed yield from SMV-infected plants. The phenotypic data and genotypic data from the SoySNP50K dataset were analyzed using GAPIT and rrBLUP. For SMV seed transmission rate, a single locus was identified on chromosome 9 in Pop1. For SMV-induced seed coat mottling, loci were identified on chromosome 9 in Pop1 and on chromosome 3 in Pop2. For seed yield from SMV-infected plants, a single locus was identified on chromosome 3 in Pop2 that was within the map interval of a previously described quantitative trait locus for seed number. The high linkage disequilibrium regions surrounding the markers on chromosomes 3 and 9 contained a predicted nonsense-mediated RNA decay gene, multiple pectin methylesterase inhibitor genes (involved in restricting virus movement), two chalcone synthase genes, and a homolog of the yeast Rtf1 gene (involved in RNA-mediated transcriptional gene silencing). The results of this study provided additional insight into the genetic architecture of these three important traits, suggested candidate genes for downstream functional validation, and suggested that genomic prediction would outperform marker-assisted selection for two of the four trait-marker associations.

Key Message: In the soybean cultivar Raiden, both a SMV-resistance gene and a BCMV-resistance gene were fine-mapped to a common region within the Rsv1 complex locus on chromosome 13, in which two CC-NBS-LRR resistance genes (Glyma.13g184800 and Glyma.13g184900) exhibited significant divergence between resistant and susceptible cultivars and were subjected to positive selection. Both Soybean mosaic virus (SMV) and Bean common mosaic virus (BCMV) can induce soybean mosaic diseases. To date, few studies have explored soybean resistance against these two viruses simultaneously. In this work, Raiden, a cultivar resistant to both SMV and BCMV, was crossed with a susceptible cultivar, Williams 82, to fine-map the resistance genes. After inoculating ~ 200 F 2 individuals with either SMV (SC6-N) or BCMV (HZZB011), a segregation ratio of 3 resistant:1 susceptible was observed, indicating that for either virus, a single dominant gene confers resistance. Bulk segregation analysis (BSA) revealed that the BCMV-resistance gene is also linked to the SMV-resistance Rsv1 complex locus. Genotyping the F 2 individuals with 12 simple sequence repeat (SSR) markers across the Rsv1 complex locus then preliminarily mapped the SMV-resistance gene, Rsv1-r, between SSR markers BARCSOYSSR_13_1075 and BARCSOYSSR_13_1161 and the BCMV-resistance gene between BARCSOYSSR_13_1084 and BARCSOYSSR_13_1115. Furthermore, a population of 1009 F 2 individuals was screened with markers BARCSOYSSR_13_1075 and BARCSOYSSR_13_1161, and 32 recombinant F 2 individuals were identified. By determining the genotypes of these F 2 individuals on multiple internal SSR and single nucleotide polymorphism (SNP) markers and assaying the phenotypes of selected recombinant F 2:3 lines, both the SMV- and BCMV-resistance genes were fine-mapped to a common region ( ~ 154.5 kb) between two SNP markers: SNP-38 and SNP-50. Within the mapped region, two CC-NBS-LRR genes exhibited significant divergence between Raiden and Williams 82, and their evolution has been affected by positive selection.

Eukaryotic positive-strand RNA viruses replicate their genomes in membranous compartments formed in a host cell, which sequesters the dsRNA replication intermediate from antiviral immune surveillance. Here, we find that soybean has developed a way to overcome this sequestration. We report the positional cloning of the broad-spectrum soybean mosaic virus resistance gene Rsv4, which encodes an RNase H family protein with dsRNA-degrading activity. An active-site mutant of Rsv4 is incapable of inhibiting virus multiplication and is associated with an active viral RNA polymerase complex in infected cells. These results suggest that Rsv4 enters the viral replication compartment and degrades viral dsRNA. Inspired by this model, we design three plant-gene-derived dsRNases that can inhibit the multiplication of the respective target viruses. These findings suggest a method for developing crops resistant to any target positive-strand RNA virus by fusion of endogenous host genes.

In soybean cultivar L29, the Rsv3 gene is responsible for extreme resistance (ER) against the soybean mosaic virus avirulent strain G5H, but is ineffective against the virulent strain G7H. Part of this ER is attributed to the rapid increase in abscisic acid (ABA) and callose, and to the rapid induction of several genes in the RNA-silencing pathway. Whether these two defense mechanisms are correlated or separated in the ER is unknown. Here, we found that ABA treatment of L29 plants increased the expression of several antiviral RNA-silencing genes as well as the PP2C3a gene, which was previously shown to increase callose accumulation; as a consequence, ABA increased the resistance of L29 plants to G7H. The effect of ABA treatment on these genes was weaker in the rsv3-null cultivar (Somyungkong) than in L29. Besides, G5H-infection of Somyungkong plants subverted the effect of ABA leading to reduced callose accumulation and decreased expression of several RNA-silencing genes, which resulted in increased susceptibility to G5H infection. ABA treatment, however, still induced some resistance to G7H in Somyungkong, but only AGO7b was significantly induced. Our data suggest that Rsv3 modulates the effect of ABA on these two resistance mechanisms, i.e., callose accumulation and the antiviral RNA-silencing pathway, and that in the absence of Rsv3 , some strains can reverse the effect of ABA and thereby facilitate their replication and spread.

Shading in the intercropping system is a major abiotic factor which influences soybean growth and development, while soybean mosaic virus (SMV) is a biotic factor that limits the yield and quality of soybean. However, little is known about the defense response of soybean to SMV in the shade. Thus, in the current study, both intensity and quality (red:far-red, R:FR) of the light were changed to simulate the shaded environment and comparative transcriptome analysis was performed. Morphologically, plant growth was inhibited by SMV, which decreased 35.93% of plant height and 8.97% of stem diameter in the shade. A total of 3548 and 4319 differentially expressed genes (DEGs) were identified in soybean plants infected with SMV under normal light and in the shade. Enrichment analysis showed that the plant defense-related genes were upregulated under normal light but downregulated in the shade. Pathways that were repressed include plant-pathogen interaction, secondary metabolism, sugar metabolism, and vitamin metabolism. In addition, genes associated with signaling pathways such as salicylic acid (SA), jasmonic acid (JA), and ethylene (ETH) were also downregulated in the shade. A qRT-PCR assay of 15 DEGs was performed to confirm transcriptome results. According to our knowledge, this is the first report on soybean response to dual stress factors. These results provide insights into the molecular mechanisms in which soybean plants were infected with SMV in the shade.

Clover yellow vein virus (ClYVV) infects and causes disease in legume plants. However, here, we found that ClYVV isolate No. 30 (ClYVV-No.30) inefficiently multiplied or spread via cell-to-cell movement in mechanically inoculated leaves of a dozen soybean ( Glycine max ) cultivars and resulted in failure to spread systemically. Soybean plants also had a similar resistance phenotype against additional ClYVV isolates. In contrast, all but one of 24 tested accessions of wild soybeans ( G. soja ) were susceptible to ClYVV-No.30. Graft inoculation of cultivated soybean TK780 with ClYVV-No.30-infected wild soybean B01167 scion resulted in systemic infection of the cultivated soybean rootstock. This suggests that, upon mechanical inoculation, the cultivated soybean inhibits ClYVV-No.30, at infection steps prior to the systemic spread of the virus, via vascular systems. Systemic infection of all F1 plants from crossing between TK780 and B01167 and of 68 of 76 F2 plants with ClYVV-No.30 indicated recessive inheritance of the resistance. Further genetic analysis using 64 recombinant inbred lines between TK780 and B01167 detected one major quantitative trait locus, designated d-cv , for the resistance that was positioned in the linkage group D1b (chromosome 2). The mapped region on soybean genome suggests that d-cv is not an allele of the known resistance genes against soybean mosaic virus.

Tobacco Streak Virus (TSV) belongs to the genus Ilarvirus of the family Bromoviridae an emerging pathogen posing threat to the crop species worldwide. Identification of symptoms due to TSV infection by visual observation of plants often results in misdiagnosis as symptoms produced by this virus can match with those reflecting physiological and nutritional disorders affecting cotton. Development of diagnostic tools with rapidity will have immense role to play in detection and management of the emerging virus. The protocol for rapid diagnosis of TSV infected samples by using Reverse Transcription-Loop Mediated Isothermal Amplification (RT-LAMP) was optimised and this is the first report of its use for diagnosis of TSV on cotton and Soybean. The colorimetric detection for diagnostic simplicity of amplified RT-LAMP product by using different dyes lead to enhanced applicability of this technique. The RT-LAMP diagnostic tool can be utilized not only for laboratory research but also for quarantine and field diagnosis of this important emerging pathogen affecting cotton., (Copyright © 2019 Elsevier B.V. All rights reserved.)

Soybean mosaic virus (SMV) is one of the most destructive viral diseases in soybeans (Glycine max). In this study, an interaction between the SMV P3 protein and cytochrome b5 was detected by yeast two-hybrid assay, and bimolecular fluorescence complementation assay showed that the interaction took place at the cell periphery. Further, the interaction was confirmed by co-immunoprecipitation analysis. Quantitative real-time polymerase chain reaction analysis revealed that GmCYB5 gene was differentially expressed in resistant and susceptible soybean plants after inoculation with SMV-SC15 strain. To test the involvement of this gene in SMV resistance, the GmCYB5 was silenced using a bean pod mottle virus (BPMV)-based vector construct. Results showed that GmCYB5-1 was 83% and 99% downregulated in susceptible (NN1138-2) and resistant (RN-9) cultivars, respectively, compared to the empty vector-treated plants. Silencing of GmCYB5 gene promotes SMV replication in soybean plants. Our results suggest that during SMV infection, the host CYB5 protein targets P3 protein to inhibit its proliferation. Taken together, these results suggest that CYB5 is an important factor in SMV infection and replication in soybeans, which could help soybean breeders develop SMV resistant soybean cultivars.

In Arabidopsis , recognition of the AvrPphB effector protease from Pseudomonas syringae is mediated by the disease resistance (R) protein RPS5, which is activated by AvrPphB-induced cleavage of the Arabidopsis protein kinase PBS1. The recognition specificity of RPS5 can be altered by substituting the AvrPphB cleavage site within PBS1 with cleavage sequences for other proteases, including proteases from viruses. AvrPphB also activates defense responses in soybean ( Glycine max ), suggesting that soybean may contain an R protein analogous to RPS5. It was unknown, however, whether this response is mediated by cleavage of a soybean PBS1-like protein. Here, we show that soybean contains three PBS1 orthologs and that their products are cleaved by AvrPphB. Further, transient expression of soybean PBS1 derivatives containing a five-alanine insertion at their AvrPphB cleavage sites activated cell death in soybean protoplasts, demonstrating that soybean likely contains an AvrPphB-specific resistance protein that is activated by a conformational change in soybean PBS1 proteins. Significantly, we show that a soybean PBS1 decoy protein modified to contain a cleavage site for the soybean mosaic virus (SMV) NIa protease triggers cell death in soybean protoplasts when cleaved by this protease, indicating that the PBS1 decoy approach will work in soybean, using endogenous PBS1 genes. Lastly, we show that activation of the AvrPphB-dependent cell death response effectively inhibits systemic spread of SMV in soybean. These data also indicate that decoy engineering may be feasible in other crop plant species that recognize AvrPphB protease activity.

Mungbean yellow mosaic India virus (MYMIV) infecting soybean and other legumes causes yellow mosaic disease (YMD). Evaluation of soybean genotypes for YMD resistance involves field screening at disease hot spots or in a protected environment using infectious clones or viruliferous whiteflies as sources of virus inocula. Development of efficient virus inoculation and quantification protocols to screen soybean genetic stocks against YMD is imperative for breeding resistant varieties. Binary plasmids harbouring complete, tandem dimeric genomic components DNA A and DNA B of MYMIV-soybean isolate were engineered. The infectivity of the clones was demonstrated in soybean genotypes JS335 and UPSM534 that display contrasting YMD resistance. As a follow-up, soybean germplasm lines, breeding lines, and representative cultivars that were initially screened at an YMD hot-spot were then subjected to Agrobacterium-based infection with MYMIV. Quantitative real time polymerase chain reaction (qRT-PCR) based copy number analysis of MYMIV genomic components allowed soybean genotypes to be classified into three discrete categories; resistant, moderately resistant and susceptible to the viral infection. Thus, a soybean germplasm disease screening system based on agro-infection and qRT-PCR based quantification of MYMIV was developed to facilitate breeding YMD resistant soybean. The implications of this study for obtaining YMD resistant soybean cultivars are discussed., (Copyright © 2019 Elsevier B.V. All rights reserved.)

A new virus species, belonging to the family Potyviridae and capable of infecting most of the soybean cultivars grown in Brazil, was collected in Lavras, Minas Gerais, Brazil, and named Soybean yellow shoot virus (SoyYSV). In this study, the complete 9,052-nucleotide genome of SoyYSV was determined and the structural, biological, and molecular properties of the virus were investigated. The SoyYSV genome encoded a single polyprotein that could be subsequently cleaved, generating 11 proteins. The SoyYSV genome shared 49% nucleotide and 36% amino acid sequence identity with Blackberry virus Y . However, the P1 protein of SoyYSV was much smaller and lacked the ALK1 domain characteristic of the genus Brambyvirus . Electron microscopy revealed flexuous filamentous virus particles, 760 to 780 nm in length, and cytoplasmic inclusions typical of those found in plant cells infected with Potyviridae species. In addition to soybean, SoyYSV infected species in the Amaranthaceae, Caricaceae, Fabaceae, and Solanaceae families. Among the most common potyviruses present in Brazil, only SoyYSV induced local necrotic lesions in Carica papaya L. SoyYSV was transmissible by Myzus persicae and Aphis gossypii but lacked the HC-Pro domain required for aphid transmission in other potyviruses. No seed transmission in soybean was observed.

Soybean ( Glycine max ) has become an important crop in Manitoba, Canada, with a 10-fold increase in dedicated acreage over the past decade. Given the rapid increase in production, scarce information about foliar diseases present in the province has been recorded. In order to describe the foliar pathogens affecting this legume, we harnessed next-generation sequencing (NGS) to carry out a comprehensive survey across Manitoba in 2016. Fields were sampled during the V2/3 (33 fields) and R6 (70 fields) growth stages, with at least three symptomatic leaves per field collected and subjected to RNA sequencing. We successfully detected several bacteria, fungi, and viruses known to infect soybean, including Pseudomonas savastanoi pv. glycinea , Septoria glycines , and Peronospora manshurica , as well as pathogens not previously identified in the province (e.g., Pseudomonas syringae pv. tabaci , Cercospora sojina , and Bean yellow mosaic virus ). For some microorganisms, we were able to disentangle the different pathovars present and/or assemble their genome sequence. Since NGS generates data on the entire flora and fauna occupying a leaf sample, we also identified residual pathogens (i.e., pathogens of crops other than soybean) and multiple species of arthropod pests. Finally, the sequence information produced by NGS allowed for the development of polymerase chain reaction-based diagnostics for some of the most widespread and important pathogens. Although there are many benefits of using NGS for large-scale plant pathogen diagnoses, we also discuss some of the limitations of this technology.

Soybean mosaic virus (SMV) is one of the major pathogens causing serious soybean losses. Little is known about the genetic structure and evolutionary biology of the SMV population in southwestern China. In this study, 29 SMV isolates were obtained from Sichuan Province, and the genomic regions encoding the first protein (P1) and coat protein (CP) were sequenced. Combined with SMV isolates from the southeastern and northeastern regions of China, the genetic and molecular evolution of SMV was studied. Recombination analysis revealed that intraspecific and interspecific recombination had occurred in the SMV population. A phylogenetic tree based on the P1 gene reflected the geographic origin of the non-interspecific recombinant SMV (SMV-NI), while a tree based on the CP gene did not. Though frequent gene flow of the SMV-NI populations was found between the southeastern and northeastern populations, the southwestern population was relatively independent. Genetic differentiation was significant between the SMV interspecific recombinant (SMV-RI) and the non-interspecific recombinant (SMV-NI) populations. It was interesting to note that there was an almost identical recombination breakpoint in SMV-RI and Watermelon mosaic virus (WMV). Population dynamics showed that SMV-RI might be in an expanding state, while the SMV-NI population is relatively stable.