An NBS-LRR protein in the Rpp1 locus negates the dominance of Rpp1-mediated resistance against Phakopsora pachyrhizi in soybean.

The soybean Rpp1 locus confers resistance to Phakopsora pachyrhizi, causal agent of rust, and resistance is usually dominant over susceptibility. However, dominance of Rpp1-mediated resistance is lost when a resistant genotype (Rpp1 or Rpp1b) is crossed with susceptible line TMG06_0011, and the mechanism of this dominant susceptibility (DS) is unknown. Sequencing the Rpp1 region reveals that the TMG06_0011 Rpp1 locus has a single nucleotide-binding site leucine-rich repeat (NBS-LRR) gene (DS-R), whereas resistant PI 594760B (Rpp1b) is similar to PI 200492 (Rpp1) and has three NBS-LRR resistance gene candidates. Evidence that DS-R is the cause of DS was reflected in virus-induced gene silencing of DS-R in Rpp1b/DS-R or Rpp1/DS-R heterozygous plants with resistance partially restored. In heterozygous Rpp1b/DS-R plants, expression of Rpp1b candidate genes was not significantly altered, indicating no effect of DS-R on transcription. Physical interaction of the DS-R protein with candidate Rpp1b resistance proteins was supported by yeast two-hybrid studies and in silico modeling. Thus, we conclude that suppression of resistance most likely does not occur at the transcript level, but instead probably at the protein level, possibly with Rpp1 function inhibited by binding to the DS-R protein. The DS-R gene was found in other soybean lines, with an estimated allele frequency of 6% in a diverse population, and also found in wild soybean (Glycine soja). The identification of a dominant susceptible NBS-LRR gene provides insight into the behavior of NBS-LRR proteins and serves as a reminder to breeders that the dominance of an R gene can be influenced by a susceptibility allele.

Precipitation change accentuates or reverses temperature effects on aphid dispersal.

Global temperatures are generally increasing, and this is leading to a well documented advancement and extension of seasonal activity of many pest insects. Effects of changing precipitation have received less attention, but might be complex because rain and snow are increasing in some places but decreasing in others. This raises the possibility that altered precipitation could accentuate, or even reverse, the effects of rising temperatures on pest outbreaks. We used >592 K aphid suction-trap captures over 15 years, in the heavily farmed central USA, to examine how the activity of Aphis glycines (soybean aphid), Rhopalosiphum maidis (corn aphid), and Rhopalosiphum padi (bird cherry-oat aphid) changed with variation in both temperature and precipitation. Increasing precipitation caused late-season flight activity of A. glycines and early-season activity of R. padi to shift earlier, while increasing temperature did the same for early-season activity of A. glycines and R. maidis. In these cases, precipitation and temperature exhibited directionally similar, but independent, effects. However, precipitation sometimes mediated temperature effects in complex ways. At relatively low temperatures, greater precipitation generally caused late-season flights of R. maidis to occur earlier. However, this pattern was reversed at higher temperatures with precipitation delaying late-season activity. In contrast, greater precipitation delayed peak flights of R. padi at lower temperatures, but caused them to occur earlier at higher temperatures. So, in these two cases the interactive effects of precipitation on temperature were mirror images of one another. When projecting future aphid flight phenology, models that excluded precipitation covariates consistently underpredicted the degree of phenological advance for A. glycines and R. padi, and underpredicted the degree of phenological delay for R. maidis under expected future climates. Overall, we found broad evidence that changing patterns of aphid flight phenology could only be understood by considering both temperature and precipitation changes. In our study region, temperature and precipitation are expected to increase in tandem, but these correlations will be reversed elsewhere. This reinforces the need to include both main and interactive effects of precipitation and temperature when seeking to accurately predict how pest pressure will change with a changing climate.

Evaluation of Soybean Entries in the Pan-African Trials for Response to Coniothyrium glycines, the Cause of Red Leaf Blotch.

Red leaf blotch (RLB), caused by the fungus Coniothyrium glycines, is an important disease of soybean known to cause yield losses across soybean-growing regions in Africa. Fungicides are one option to manage this disease, but utilization of host resistance may be a better option suited for smallholder soybean farmers in Africa. Fifty-nine soybean entries were evaluated for RLB severity in nine field locations in Ethiopia, Kenya, Uganda, and Zambia. Disease incidence was 100% and disease severity differed (P < 0.01) among entries at eight of the nine locations. Mean severity ratings ranged from 1.4 to 3.2 based on a 0-to-5 scale, with higher disease severities recorded in Ethiopia followed by Zambia. Eight of the 59 entries were common to all nine locations and had severity ratings ranging from 1.6 to 2.9. The cultivar SC Signal had the lowest RLB severity ratings in the combined analysis over locations. Based on correlations of weather variables to RLB severity, mean rainfall from planting to 30 days before assessment date had a positive correlation (r = 0.70; P = 0.035), as did mean morning maximum wind speed (r = 0.88; P = 0.016). Other variables, such as temperature and relative humidity, did not correlate to RLB severity. This is the most comprehensive report to date on the occurrence of RLB in the region, which for the first time demonstrates an association between rainfall and wind speed with RLB severity. It also represents the first extensive report evaluating soybean genotypes for resistance against RLB in multiple environments.

Evaluation of perennial Glycine species for response to Meloidogyne incognita, Rotylenchulus reniformis, and Pratylenchus penetrans.

Root-knot (Meloidogyne incognita (Kofoid & White) Chitwood), reniform (Rotylenchulus reniformis Lindford & Oliveira), and lesion nematodes (Pratylenchus penetrans (Cobb) Filipjev & Schuurmans Stekhoven) are plant-parasitic nematodes that feed on soybean (Glycine max (L.) Merr.) roots, limiting seed production. The availability of resistance in soybeans to these nematodes is limited. However, new sources of resistance can be discovered in wild relatives of agronomic crops. Perennial Glycine species, wild relatives to soybean, are a source of valuable genetic resources with the potential to improve disease resistance in soybean. To determine if these perennials have resistance against nematodes, 18 accessions of 10 perennial Glycine species were evaluated for their response to M. incognita and R. reniformis, and eight accessions of six perennial Glycine species were evaluated for their response to P. penetrans. Pot experiments were conducted for M. incognita and R. reniformis in a growth chamber and in vitro experiments were conducted for P. penetrans. We found both shared and distinct interactions along the resistance-susceptible continuum in response to the three plant-parasitic nematode species. Ten and 15 accessions were classified as resistant to M. incognita based on eggs per gram of root and gall index, respectively. Among them, G. tomentella plant introductions (PIs) 446983 and 339655 had a significantly lower gall index than the resistant soybean check cv. Forrest. Of three R. reniformis resistant accessions identified in this study, G. tomentella PI 441001 showed significantly greater resistance to R. reniformis than the resistant check cv. Forrest based on nematodes per gram of root. In contrast, no resistance to P. penetrans was recorded in any perennial Glycine species.

Complex life histories predispose aphids to recent abundance declines.

Many animals change feeding habits as they progress through life stages, exploiting resources that vary in space and time. However, complex life histories may bring new risks if rapid environmental change disrupts the timing of these switches. Here, we use abundance times series for a diverse group of herbivorous insects, aphids, to search for trait and environmental characteristics associated with declines. Our meta dataset spanned three world regions and >300 aphid species, tracked at 75 individual sites for 10-50 years. Abundances were generally falling, with median changes of -8.3%, -5.6%, and -0.1% per year in the central USA, northwestern USA, and United Kingdom, respectively. Aphids that obligately alternated between host plants annually and those that were agricultural pests exhibited the steepest declines, relative to species able to persist on the same host plant year-round or those in natural areas. This suggests that host alternation might expose aphids to climate-induced phenology mismatches with one or more of their host plant species, with additional risks from exposure to insecticides and other management efforts. Warming temperatures through time were associated with milder aphid declines or even abundance increases, particularly at higher latitudes. Altogether, while a warming world appeared to benefit some aphid species in some places, most aphid species that had time-sensitive movements among multiple host plants seemed to face greater risk of decline. More generally, this suggests that recent human-induced rapid environmental change is rebalancing the risks and rewards associated with complex life histories.

Aphis glycines virus 1, a new bicistronic virus with two functional internal ribosome entry sites, is related to a group of unclassified viruses in the Picornavirales.

A novel picorna-like virus, provisionally named Aphis glycines virus 1 (ApGlV1) was discovered by high-throughput sequencing of soybean total RNAs and detected in suction trap-collected Aphis glycines. The ApGlV1 genome contains two large ORFs organized similar to those of dicipiviruses in the Picornaviridae where ORFs 1 and 2 encode structural and nonstructural proteins, respectively. Both ORFs are preceded by internal ribosome entry site (IRES) elements. The 5' IRES was more active in dual luciferase activity assays than the IRES in the intergenic region. The ApGlV1 genome was predicted to encode a serine protease instead of a cysteine protease and showed very low aa sequence identities to recognized members of the Picornavirales. In phylogenetic analyses based on capsid protein and RNA-dependent RNA polymerase sequences, ApGlV1 consistently clustered with a group of unclassified bicistronic picorna-like viruses discovered from arthropods and plants that may represent a novel family in the order Picornavirales.

Whole-genome resequencing identifies quantitative trait loci associated with mycorrhizal colonization of soybean.

KEY MESSAGE: A whole-genome resequencing-derived SNP dataset identified six quantitative trait loci (QTL) significantly associated with colonization of soybean by an arbuscular mycorrhizal fungus (Rhizophagus intraradices). Candidate genes identified in these QTL regions include homologs to known nodulin protein families and other symbiosis-specific genes. Arbuscular mycorrhizal fungi (AMF) form associations with over 80% of all terrestrial plant species and assist their host plants by increasing their nutrient uptake, drought tolerance, and resilience against pathogens and pests. Genotypic variation of crop plants to AMF colonization has been identified in crops, including soybean; however, the genetics controlling levels of AMF colonization in soybean are unknown. The overall goal of our study was to identify genomic regions associated with mycorrhizal colonization in soybean using genome-wide association analysis. A diverse panel of 350 exotic soybean genotypes inoculated with Rhizophagus intraradices were microscopically evaluated for root colonization using a modified gridline intersect method. Root colonization differed significantly (P < 0.001) among genotypes and ranged from 11 to 70%. A whole-genome resequencing-derived SNP dataset identified six quantitative trait loci (QTL) significantly associated with R. intraradices colonization that explained 24% of the phenotypic variance. Candidate genes identified in these QTL regions include homologs to known nodulin protein families and other symbiosis-specific genes. The results showed there was a significant genetic component to the level of colonization by R. intraradices in soybean. This information may be useful in the development of AMF-sensitive soybean cultivars to enhance nutrient uptake, drought tolerance, and disease resistance in the crop.

Reduction of Sudden Death Syndrome Foliar Symptoms and Fusarium virguliforme DNA in Roots Inoculated With Rhizophagus intraradices.

There is increasing interest in incorporating arbuscular mycorrhizal fungi (AMF) into agricultural production because of the benefits they provide, including protection against pathogens and pests. Sudden death syndrome (SDS) of soybean is a devastating disease caused by the soilborne pathogen Fusarium virguliforme. Multiple management methods are needed to control SDS. The relationship between F. virguliforme and AMF is not well documented. The goal of this study was to determine whether soybean plants co-inoculated with F. virguliforme and the AMF species Rhizophagus intraradices showed reduced SDS foliar symptom severity and reduced relative F. virguliforme DNA quantities in soybean roots. Six soybean genotypes were inoculated with F. virguliforme alone or with R. intraradices in a greenhouse experiment. Averaged over the six soybean genotypes, area under the disease progress curve values and relative F. virguliforme DNA quantities were 45 and 28% lower (P < 0.05), respectively, in roots co-inoculated with R. intraradices compared with roots of control plants inoculated with F. virguliforme only. Weight of roots co-inoculated with R. intraradices were 58% higher (P < 0.05) compared with roots of plants not inoculated with R. intraradices. Nutrient analysis showed higher boron, phosphorus, potassium, sodium, and sulfur concentrations in root tissues of plants co-inoculated with R. intraradices compared with plants inoculated with F. virguliforme (P < 0.05). Overall, this study showed that R. intraradices reduced SDS severity and relative F. virguliforme DNA quantities while simultaneously increasing growth and nutrient uptake of plants. Further testing of AMF inoculants in the field will indicate whether incorporating them into soybean SDS management practices will reduce the impact of SDS on soybean production.

Evaluation of Foliar Diseases for Soybean Entries in the Pan-African Trials in Malawi and Zambia.

Soybean production has expanded worldwide including countries in sub-Saharan Africa. Several national and international agencies and research groups have partnered to improve overall performance of soybean breeding stocks and have introduced new germplasm from Brazil and the United States with the goal of developing new high-yielding cultivars. Part of this effort has been to test improved soybean lines/cultivars accumulated from private and public sources in multilocational trials in sub-Saharan Africa. These trials are known as the Pan-African Soybean Variety Trials, and the entries come from both private and public breeding programs. The objective of this research was to evaluate entries in the trials that include commercial cultivars or advanced experimental lines for the incidence and severity of foliar diseases. All trials were planted in December 2018 with six located in Zambia and one in Malawi. Plants were evaluated during the reproductive growth stages using a visual pretransformed severity rating scale. Foliar disease ratings were recorded for three bacterial diseases, six fungal diseases, one oomycete, and viruses. The overall occurrence of most of the diseases was high except for soybean rust and target spot, which were only found at two and one location, respectively. However, disease severity was generally low, although there were differences in disease severity ratings among the entries at some of the locations for brown spot, downy mildew, frogeye leaf spot, red leaf blotch, and soybean rust.

Assembly and annotation of a draft genome sequence for Glycine latifolia, a perennial wild relative of soybean.

Glycine latifolia (Benth.) Newell & Hymowitz (2n = 40), one of the 27 wild perennial relatives of soybean, possesses genetic diversity and agronomically favorable traits that are lacking in soybean. Here, we report the 939-Mb draft genome assembly of G. latifolia (PI 559298) using exclusively linked-reads sequenced from a single Chromium library. We organized scaffolds into 20 chromosome-scale pseudomolecules utilizing two genetic maps and the Glycine max (L.) Merr. genome sequence. High copy numbers of putative 91-bp centromere-specific tandem repeats were observed in consecutive blocks within predicted pericentromeric regions on several pseudomolecules. No 92-bp putative centromeric repeats, which are abundant in G. max, were detected in G. latifolia or Glycine tomentella. Annotation of the assembled genome and subsequent filtering yielded a high confidence gene set of 54 475 protein-coding loci. In comparative analysis with five legume species, genes related to defense responses were significantly overrepresented in Glycine-specific orthologous gene families. A total of 304 putative nucleotide-binding site (NBS)-leucine-rich-repeat (LRR) genes were identified in this genome assembly. Different from other legume species, we observed a scarcity of TIR-NBS-LRR genes in G. latifolia. The G. latifolia genome was also predicted to contain genes encoding 367 LRR-receptor-like kinases, a family of proteins involved in basal defense responses and responses to abiotic stress. The genome sequence and annotation of G. latifolia provides a valuable source of alternative alleles and novel genes to facilitate soybean improvement. This study also highlights the efficacy and cost-effectiveness of the application of Chromium linked-reads in diploid plant genome de novo assembly.

Trichothecene-Producing Fusarium Species Isolated from Soybean Roots in Ethiopia and Ghana and their Pathogenicity on Soybean.

Numerous pathogen surveys have reported that diverse Fusarium spp. threaten soybean production in North and South America. However, little research has been conducted to characterize Fusarium pathogens of soybean in sub-Saharan Africa. Our objectives were to (i) identify Fusarium spp. isolated from discolored root segments of soybean grown in Ethiopia and Ghana using DNA sequence data, (ii) determine whether isolates nested in the Fusarium incarnatum-equiseti and F. sambucinum species complexes (FIESC and FSAMSC, respectively) produced trichothecene mycotoxins in vitro, and (iii) test these isolates for pathogenicity on soybean. Molecular phylogenetic analyses revealed that the trichothecene mycotoxin-producing isolates comprised three undescribed species within the FIESC and FSAMSC. Mycotoxin type B trichothecene 4,15-diacetylnivalenol or T-2 toxin and related type A neosolaniol trichothecenes were produced by 18 of the 21 isolates. Of the 12 isolates from Ethiopia and Ghana tested for their impact on seed germination, 5, comprising two undescribed phylospecies (i.e., Fusarium sp. number 3 and Fusarium sp. FIESC 2,) completely inhibited germination, whereas 4 caused no reduction in germination. Root lesions induced by all 12 isolates were greater than the uninoculated negative control. Additional variation among the isolates was reflected in differences (α = 0.05) in lesion lengths, which ranged from 34 to 67% of total root length. This is the first report characterizing FIESC and FSAMSC isolates from soybean roots in Ethiopia and Ghana.

Integration of sudden death syndrome resistance loci in the soybean genome.

KEY MESSAGE: Complexity and inconsistencies in resistance mapping publications of soybean sudden death syndrome (SDS) result in interpretation difficulty. This review integrates SDS mapping literature and proposes a new nomenclature system for reproducible SDS resistance loci. Soybean resistance to sudden death syndrome (SDS) is composed of foliar resistance to phytotoxins and root resistance to pathogen invasion. There are more than 80 quantitative trait loci (QTL) and dozens of single nucleotide polymorphisms (SNPs) associated with soybean resistance to SDS. The validity of these QTL and SNPs is questionable because of the complexity in phenotyping methodologies, the disease synergism between SDS and soybean cyst nematode (SCN), the variability from the interactions between soybean genotypes and environments, and the inconsistencies in the QTL nomenclature. This review organizes SDS mapping results and proposes the Rfv (resistance to Fusarium virguliforme) nomenclature based on supporting criteria described in the text. Among ten reproducible loci receiving our Rfv nomenclature, Rfv18-01 is mostly supported by field studies and it co-localizes to the SCN resistance locus rhg1. The possibility that Rfv18-01 is a pleiotropic resistance locus and the concern about Rfv18-01 being confounded with Rhg1 is discussed. On the other hand, Rfv06-01, Rfv06-02, Rfv09-01, Rfv13-01, and Rfv16-01 were identified both by screening soybean leaves against phytotoxic culture filtrates and by evaluating SDS severity in fields. Future phenotyping using leaf- and root-specific resistance screening methodologies may improve the precision of SDS resistance, and advanced genetic studies may further clarify the interactions among soybean genotypes, F. virguliforme, SCN, and environments. The review provides a summary of the SDS resistance literature and proposes a framework for communicating SDS resistance loci for future research considering molecular interactions and genetic breeding for soybean SDS resistance.

Field evaluation of three sources of genetic resistance to sudden death syndrome of soybean.

KEY MESSAGE: Despite numerous challenges, field testing of three sources of genetic resistance to sudden death syndrome of soybean provides information to more effectively improve resistance to this disease in cultivars. Sudden death syndrome (SDS) of soybean [Glycine max (L.) Merrill] is a disease that causes yield loss in soybean growing regions across the USA and worldwide. While several quantitative trait loci (QTL) for SDS resistance have been mapped, studies to further evaluate these QTL are limited. The objective of our research was to map SDS resistance QTL and to test the effect of mapped resistance QTL on foliar symptoms when incorporated into elite soybean backgrounds. We mapped a QTL from Ripley to chromosome 10 (CHR10) and a QTL from PI507531 to chromosomes 1 and 18 (CHR1 and 18). Six populations were then developed to test the following QTL: cqSDS-001, with resistance originating from PI567374, CHR10, CHR1, and CHR18. The populations which segregated for resistant and susceptible QTL alleles were field tested in multiple environments and evaluated for SDS foliar symptoms. While foliar disease development was variable across environments and populations, a significant effect of each QTL on disease was detected within at least one environment. This includes the detection of cqSDS-001 in three genetic backgrounds. The QTL allele from the resistant parents was associated with greater resistance than the susceptible alleles for all QTL and backgrounds with the exception of the allele for CHR18, where the opposite occurred. This study highlights the importance and difficulties of evaluating QTL and the need for multi-year SDS field testing. The information presented in this study can aid breeders in making decisions to improve resistance to SDS.

Identification of Diverse Mycoviruses through Metatranscriptomics Characterization of the Viromes of Five Major Fungal Plant Pathogens.

UNLABELLED: Mycoviruses can have a marked effect on natural fungal communities and influence plant health and productivity. However, a comprehensive picture of mycoviral diversity is still lacking. To characterize the viromes of five widely dispersed plant-pathogenic fungi, Colletotrichum truncatum, Macrophomina phaseolina, Diaporthe longicolla, Rhizoctonia solani, and Sclerotinia sclerotiorum, a high-throughput sequencing-based metatranscriptomic approach was used to detect viral sequences. Total RNA and double-stranded RNA (dsRNA) from mycelia and RNA from samples enriched for virus particles were sequenced. Sequence data were assembled de novo, and contigs with predicted amino acid sequence similarities to viruses in the nonredundant protein database were selected. The analysis identified 72 partial or complete genome segments representing 66 previously undescribed mycoviruses. Using primers specific for each viral contig, at least one fungal isolate was identified that contained each virus. The novel mycoviruses showed affinity with 15 distinct lineages: Barnaviridae, Benyviridae, Chrysoviridae, Endornaviridae, Fusariviridae, Hypoviridae, Mononegavirales, Narnaviridae, Ophioviridae, Ourmiavirus, Partitiviridae, Tombusviridae, Totiviridae, Tymoviridae, and Virgaviridae More than half of the viral sequences were predicted to be members of the Mitovirus genus in the family Narnaviridae, which replicate within mitochondria. Five viral sequences showed strong affinity with three families (Benyviridae, Ophioviridae, and Virgaviridae) that previously contained no mycovirus species. The genomic information provides insight into the diversity and taxonomy of mycoviruses and coevolution of mycoviruses and their fungal hosts. IMPORTANCE: Plant-pathogenic fungi reduce crop yields, which affects food security worldwide. Plant host resistance is considered a sustainable disease management option but may often be incomplete or lacking for some crops to certain fungal pathogens or strains. In addition, the rising issues of fungicide resistance demand alternative strategies to reduce the negative impacts of fungal pathogens. Those fungus-infecting viruses (mycoviruses) that attenuate fungal virulence may be welcome additions for mitigation of plant diseases. By high-throughput sequencing of the RNAs from 275 isolates of five fungal plant pathogens, 66 previously undescribed mycoviruses were identified. In addition to identifying new potential biological control agents, these results expand the grand view of the diversity of mycoviruses and provide possible insights into the importance of intracellular and extracellular transmission in fungus-virus coevolution.

Characterization and genetics of multiple soybean aphid biotype resistance in five soybean plant introductions.

KEY MESSAGE: Five soybean plant introductions expressed antibiosis resistance to multiple soybean aphid biotypes. Two introductions had resistance genes located in the Rag1, Rag2, and Rag3 regions; one introduction had resistance genes located in the Rag1, Rag2, and rag4 regions; one introduction had resistance genes located in the Rag1 and Rag2 regions; and one introduction had a resistance gene located in the Rag2 region. Soybean aphid (Aphis glycines Matsumura) is the most important soybean [Glycine max (L.) Merr.] insect pest in the USA. The objectives of this study were to characterize the resistance expressed in five plant introductions (PIs) to four soybean aphid biotypes, determine the mode of resistance inheritance, and identify markers associated with genes controlling resistance in these accessions. Five soybean PIs, from an initial set of 3000 PIs, were tested for resistance against soybean aphid biotypes 1, 2, 3, and 4 in choice and no-choice tests. Of these five PIs, PI 587663, PI 587677, and PI 587685 expressed antibiosis against all four biotypes, while PI 587972 and PI 594592 expressed antibiosis against biotypes 1, 2, and 3. F2 populations derived from PI 587663 and PI 587972 were evaluated for resistance against soybean aphid biotype 1, and populations derived from PIs 587677, 587685, and 594592 were tested against biotype 3. In addition, F2:3 plants were tested against biotypes 2 and 3. Genomic DNA from F2 plants was screened with markers linked to Rag1, Rag2, Rag3, and rag4 soybean aphid-resistance genes. Results showed that PI 587663 and PI 594592 each had three genes with variable gene action located in the Rag1, Rag2, and Rag3 regions. PI 587677 had three genes with variable gene action located in the Rag1, Rag2 and rag4 regions. PI 587685 had one dominant gene located in the Rag1 region and an additive gene in the Rag2 region. PI 587972 had one dominant gene located in the Rag2 region controlling antixenosis- or antibiosis-type resistance to soybean aphid biotypes 1, 2, or 3. PIs 587663, 587677, and 587685 also showed antibiosis-type resistance against biotype 4. Information on multi-biotype aphid resistance and resistance gene markers will be useful for improving soybean aphid resistance in commercial soybean cultivars.

Identification of Multiple Phytotoxins Produced by Fusarium virguliforme Including a Phytotoxic Effector (FvNIS1) Associated With Sudden Death Syndrome Foliar Symptoms.

Sudden death syndrome (SDS) of soybean is caused by a soilborne pathogen, Fusarium virguliforme. Phytotoxins produced by F. virguliforme are translocated from infected roots to leaves, in which they cause SDS foliar symptoms. In this study, additional putative phytotoxins of F. virguliforme were identified, including three secondary metabolites and 11 effectors. While citrinin, fusaric acid, and radicicol induced foliar chlorosis and wilting, Soybean mosaic virus (SMV)-mediated overexpression of F. virguliforme necrosis-inducing secreted protein 1 (FvNIS1) induced SDS foliar symptoms that mimicked the development of foliar symptoms in the field. The expression level of fvnis1 remained steady over time, although foliar symptoms were delayed compared with the expression levels. SMV::FvNIS1 also displayed genotype-specific toxicity to which 75 of 80 soybean cultivars were susceptible. Genome-wide association mapping further identified three single nucleotide polymorphisms at two loci, where three leucine-rich repeat receptor-like protein kinase (LRR-RLK) genes were found. Culture filtrates of fvnis1 knockout mutants displayed a mild reduction in phytotoxicity, indicating that FvNIS1 is one of the phytotoxins responsible for SDS foliar symptoms and may contribute to the quantitative susceptibility of soybean by interacting with the LRR-RLK genes.

Genome-wide association and genomic prediction identifies associated loci and predicts the sensitivity of Tobacco ringspot virus in soybean plant introductions.

BACKGROUND: Genome-wide association study (GWAS) is a useful tool for detecting and characterizing traits of interest including those associated with disease resistance in soybean. The availability of 50,000 single nucleotide polymorphism (SNP) markers (SoySNP50K iSelect BeadChip; www.soybase.org ) on 19,652 soybean and wild soybean plant introductions (PIs) in the USDA Soybean Germplasm Collection allows for fast and robust identification of loci associated with a desired phenotype. By using a genome-wide marker set to predict phenotypic values, genomic prediction for phenotype-unknown but genotype-determined PIs has become possible. The goal of this study was to describe the genetic architecture associated with sensitivity to Tobacco ringspot virus (TRSV) infection in the USDA Soybean Germplasm Collection. RESULTS: TRSV-induced disease sensitivities of the 697 soybean PIs were rated on a one to five scale with plants rated as one exhibiting mild symptoms and plants rated as five displaying terminal bud necrosis (i.e., bud blight). The GWAS identified a single locus on soybean chromosome 2 strongly associated with TRSV sensitivity. Cross-validation showed a correlation of 0.55 (P < 0.01) to TRSV sensitivity without including the most significant SNP marker from the GWAS as a covariate, which was a better estimation compared to the mean separation by using significant SNPs. The genomic estimated breeding values for the remaining 18,955 unscreened soybean PIs in the USDA Soybean Germplasm Collection were obtained using the GAPIT R package. To evaluate the prediction accuracy, an additional 55 soybean accessions were evaluated for sensitivity to TRSV, which resulted in a correlation of 0.67 (P < 0.01) between actual and predicted severities. CONCLUSION: A single locus responsible for TRSV sensitivity in soybean was identified on chromosome 2. Two leucine-rich repeat receptor-like kinase genes were located near the locus and may control sensitivity of soybean to TRSV infection. Furthermore, a comprehensive genomic prediction for TRSV sensitivity for all accessions in the USDA Soybean Germplasm Collection was completed.

Transfection of Sclerotinia sclerotiorum with in vitro transcripts of a naturally occurring interspecific recombinant of Sclerotinia sclerotiorum hypovirus 2 significantly reduces virulence of the fungus.

UNLABELLED: A recombinant strain of Sclerotinia sclerotiorum hypovirus 2 (SsHV2) was identified from a North American Sclerotinia sclerotiorum isolate (328) from lettuce (Lactuca sativa L.) by high-throughput sequencing of total RNA. The 5'- and 3'-terminal regions of the genome were determined by rapid amplification of cDNA ends. The assembled nucleotide sequence was up to 92% identical to two recently reported SsHV2 strains but contained a deletion near its 5' terminus of more than 1.2 kb relative to the other SsHV2 strains and an insertion of 524 nucleotides (nt) that was distantly related to Valsa ceratosperma hypovirus 1. This suggests that the new isolate is a heterologous recombinant of SsHV2 with a yet-uncharacterized hypovirus. We named the new strain Sclerotinia sclerotiorum hypovirus 2 Lactuca (SsHV2L) and deposited the sequence in GenBank with accession number KF898354. Sclerotinia sclerotiorum isolate 328 was coinfected with a strain of Sclerotinia sclerotiorum endornavirus 1 and was debilitated compared to cultures of the same isolate that had been cured of virus infection by cycloheximide treatment and hyphal tipping. To determine whether SsHV2L alone could induce hypovirulence in S. sclerotiorum, a full-length cDNA of the 14,538-nt viral genome was cloned. Transcripts corresponding to the viral RNA were synthesized in vitro and transfected into a virus-free isolate of S. sclerotiorum, DK3. Isolate DK3 transfected with SsHV2L was hypovirulent on soybean and lettuce and exhibited delayed maturation of sclerotia relative to virus-free DK3, completing Koch's postulates for the association of hypovirulence with SsHV2L. IMPORTANCE: A cosmopolitan fungus, Sclerotinia sclerotiorum infects more than 400 plant species and causes a plant disease known as white mold that produces significant yield losses in major crops annually. Mycoviruses have been used successfully to reduce losses caused by fungal plant pathogens, but definitive relationships between hypovirus infections and hypovirulence in S. sclerotiorum were lacking. By establishing a cause-and-effect relationship between Sclerotinia sclerotiorum hypovirus Lactuca (SsHV2L) infection and the reduction in host virulence, we showed direct evidence that hypoviruses have the potential to reduce the severity of white mold disease. In addition to intraspecific recombination, this study showed that recent interspecific recombination is an important factor shaping viral genomes. The construction of an infectious clone of SsHV2L allows future exploration of the interactions between SsHV2L and S. sclerotiorum, a widespread fungal pathogen of plants.

Genomic characterization of plant cell wall degrading enzymes and in silico analysis of xylanases and polygalacturonases of Fusarium virguliforme.

BACKGROUND: Plant cell wall degrading enzymes (PCWDEs) are a subset of carbohydrate-active enzymes (CAZy) produced by plant pathogens to degrade plant cell walls. To counteract PCWDEs, plants release PCWDEs inhibitor proteins (PIPs) to reduce their impact. Several transgenic plants expressing exogenous PIPs that interact with fungal glycoside hydrolase (GH)11-type xylanases or GH28-type polygalacturonase (PG) have been shown to enhance disease resistance. However, many plant pathogenic Fusarium species were reported to escape PIPs inhibition. Fusarium virguliforme is a soilborne pathogen that causes soybean sudden death syndrome (SDS). Although the genome of F. virguliforme was sequenced, there were limited studies focused on the PCWDEs of F. virguliforme. Our goal was to understand the genomic CAZy structure of F. viguliforme, and determine if exogenous PIPs could be theoretically used in soybean to enhance resistance against F. virguliforme. RESULTS: F. virguliforme produces diverse CAZy to degrade cellulose and pectin, similar to other necrotorphic and hemibiotrophic plant pathogenic fungi. However, some common CAZy of plant pathogenic fungi that catalyze hemicellulose, such as GH29, GH30, GH44, GH54, GH62, and GH67, were deficient in F. virguliforme. While the absence of these CAZy families might be complemented by other hemicellulases, F. virguliforme contained unique families including GH131, polysaccharide lyase (PL) 9, PL20, and PL22 that were not reported in other plant pathogenic fungi or oomycetes. Sequence analysis revealed two GH11 xylanases of F. virguliforme, FvXyn11A and FvXyn11B, have conserved residues that allow xylanase inhibitor protein I (XIP-I) binding. Structural modeling suggested that FvXyn11A and FvXyn11B could be blocked by XIP-I that serves as good candidate for developing transgenic soybeans. In contrast, one GH28 PG, FvPG2, contains an amino acid substitution that is potentially incompatible with the bean polygalacturonase-inhibitor protein II (PvPGIP2). CONCLUSIONS: Identification and annotation of CAZy provided advanced understanding of genomic composition of PCWDEs in F. virguliforme. Sequence and structural analyses of FvXyn11A and FvXyn11B suggested both xylanases were conserved in residues that allow XIP-I inhibition, and expression of both xylanases were detected during soybean roots infection. We postulate that a transgenic soybean expressing wheat XIP-I may be useful for developing root rot resistance to F. virguliforme.

Characterization of Disease Resistance Loci in the USDA Soybean Germplasm Collection Using Genome-Wide Association Studies.

Genetic resistance is a key strategy for disease management in soybean. Over the last 50 years, soybean germplasm has been phenotyped for resistance to many pathogens, resulting in the development of disease-resistant elite breeding lines and commercial cultivars. While biparental linkage mapping has been used to identify disease resistance loci, genome-wide association studies (GWAS) using high-density and high-quality markers such as single nucleotide polymorphisms (SNPs) has become a powerful tool to associate molecular markers and phenotypes. The objective of our study was to provide a comprehensive understanding of disease resistance in the United States Department of Agriculture Agricultural Research Service Soybean Germplasm Collection by using phenotypic data in the public Germplasm Resources Information Network and public SNP data (SoySNP50K). We identified SNPs significantly associated with disease ratings from one bacterial disease, five fungal diseases, two diseases caused by nematodes, and three viral diseases. We show that leucine-rich repeat (LRR) receptor-like kinases and nucleotide-binding site-LRR candidate resistance genes were enriched within the linkage disequilibrium regions of the significant SNPs. We review and present a global view of soybean resistance loci against multiple diseases and discuss the power and the challenges of using GWAS to discover disease resistance in soybean.