Genome scans for selection signatures identify candidate virulence genes for adaptation of the soybean cyst nematode to host resistance.

Plant pathogens are constantly under selection pressure for host resistance adaptation. Soybean cyst nematode (SCN, Heterodera glycines) is a major pest of soybean primarily managed through resistant cultivars; however, SCN populations have evolved virulence in response to selection pressures driven by repeated monoculture of the same genetic resistance. Resistance to SCN is mediated by multiple epistatic interactions between Rhg (for resistance to H. glycines) genes. However, the identity of SCN virulence genes that confer the ability to overcome resistance remains unknown. To identify candidate genomic regions showing signatures of selection for increased virulence, we conducted whole genome resequencing of pooled individuals (Pool-Seq) from two pairs of SCN populations adapted on soybeans with Peking-type (rhg1-a, rhg2, and Rhg4) resistance. Population differentiation and principal component analysis-based approaches identified approximately 0.72-0.79 million SNPs, the frequency of which showed potential selection signatures across multiple genomic regions. Chromosomes 3 and 6 between population pairs showed the greatest density of outlier SNPs with high population differentiation. Conducting multiple outlier detection tests to identify overlapping SNPs resulted in a total of 966 significantly differentiated SNPs, of which 285 exon SNPs were mapped to 97 genes. Of these, six genes encoded members of known stylet-secreted effector protein families potentially involved in host defence modulation including venom-allergen-like, annexin, glutathione synthetase, SPRYSEC, chitinase, and CLE effector proteins. Further functional analysis of identified candidate genes will provide new insights into the genetic mechanisms by which SCN overcomes soybean resistance and inform the development of molecular markers for rapidly screening the virulence profile of an SCN-infested field.

MiR398b Targets Superoxide Dismutase Genes in Soybean in Defense Against Heterodera glycines via Modulating Reactive Oxygen Species Homeostasis.

MicroRNAs play crucial roles in plant defense responses. However, the underlying mechanism by which miR398b contributes to soybean responses to soybean cyst nematode (Heterodera glycines) remains elusive. In this study, by using Agrobacterium rhizogenes-mediated transformation of soybean hairy roots, we observed that miR398b and target genes GmCCS and GmCSD1b played vital functions in soybean-H. glycines interaction. The study revealed that the abundance of miR398b was downregulated by H. glycines infection, and overexpression of miR398b enhanced the susceptibility of soybean to H. glycines. Conversely, silencing of miR398b improved soybean resistance to H. glycines. Detection assays revealed that miR398b rapidly senses stress-induced reactive oxygen species, leading to the repression of target genes GmCCS and GmCSD1b and regulating the accumulation of plant defense genes against nematode infection. Moreover, exogenous synthetic ds-miR398b enhanced soybean sensitivity to H. glycines by modulating H2O2 and O2- levels. Functional analysis demonstrated that overexpression of GmCCS and GmCSD1b in soybean enhanced resistance to H. glycines. RNA interference-mediated repression of GmCCS and GmCSD1b in soybean increased susceptibility to H. glycines. RNA sequencing revealed that a majority of differentially expressed genes in overexpressed GmCCS were associated with oxidative stress. Overall, the results indicate that miR398b targets superoxide dismutase genes, which negatively regulate soybean resistance to H. glycines via modulating reactive oxygen species levels and defense signals.

N6-Methyladenosine (m6A) Sequencing Reveals Heterodera glycines-Induced Dynamic Methylation Promoting Soybean Defense.

Unraveling the intricacies of soybean cyst nematode (Heterodera glycines) race 4 resistance and susceptibility in soybean breeding lines-11-452 (highly resistant) and Dongsheng1 (DS1, highly susceptible)-was the focal point of this study. Employing cutting-edge N6-methyladenosine (m6A) and RNA sequencing techniques, we delved into the impact of m6A modification on gene expression and plant defense responses. Through the evaluation of nematode development in both resistant and susceptible roots, a pivotal time point (3 days postinoculation) for m6A methylation sequencing was identified. Our sequencing data exhibited robust statistics, successful soybean genome mapping, and prevalent m6A peak distributions, primarily in the 3' untranslated region and stop codon regions. Analysis of differential methylation peaks and differentially expressed genes revealed distinctive patterns between resistant and susceptible genotypes. In the highly resistant line (11-452), key resistance and defense-associated genes displayed increased expression coupled with inhibited methylation, encompassing crucial players such as R genes, receptor kinases, and transcription factors. Conversely, the highly susceptible DS1 line exhibited heightened expression correlated with decreased methylation in genes linked to susceptibility pathways, including Mildew Locus O-like proteins and regulatory elements affecting defense mechanisms. Genome-wide assessments, Gene Ontology and Kyoto Encyclopedia of Genes and Genomes analyses, and differential methylation peak/differentially expressed gene overlap emphasized the intricate interplay of m6A modifications, alternative splicing, microRNA, and gene regulation in plant defense. Protein-protein interaction networks illuminated defense-pivotal genes, delineating divergent mechanisms in resistant and susceptible responses. This study sheds light on the dynamic correlation between methylation, splicing, and gene expression, providing profound insights into plant responses to nematode infection.

Identification and Biological Characterization of a New Cyst Nematode, Heterodera glycines sbsp.n. tabacum, Parasitizing Tobacco in China.

In an investigation of diseases from plant-parasitizing nematodes in Henan Province, a cyst nematode was found on tobacco roots and in rhizosphere soil. We identified this strain as a new cyst nematode subspecies, Heterodera glycines sbsp.n. tabacum. The cysts and second-stage juveniles (J2s) parasitizing Henan tobacco were larger than those of H. glycines. A single 345-bp fragment was amplified from H. glycines sbsp.n. tabacum, whereas the 345- and 181-bp fragments were amplified from the soybean cyst nematode. Thus, H. glycines sbsp.n. tabacum was distinct from H. glycines. There were base transversions at 504 sites and base transitions at 560, 858, 920, and 921 sites in the rDNA-ITS sequences of H. glycines sbsp.n. tabacum compared with H. glycines, and there were base transitions at 41, 275, 278, and 380 sites in the mtDNA-COI sequences. In the phylogenetic tree based on the rDNA-ITS and mtDNA-COI regions, H. glycines sbsp.n. tabacum was clustered on a single branch. Based on the randomly amplified polymorphic DNA (RAPD) technique, sequence characterized amplified region (SCAR)-PCR primers were designed. A single 1,113-bp fragment was amplified by specific primers (HtF1/HtR1) from H. glycines sbsp.n. tabacum, while no fragments were obtained from H. glycines. The H. glycines sbsp.n. tabacum can infect soybean plants but cannot complete its life cycle on soybean. Eleven tested tobacco cultivars were infected, with an average reproduction factor (Rf) of 9.74 and a maximum of 64.2 in 'K326'. The cumulative egg hatching rate of H. glycines sbsp.n. tabacum in the presence of tobacco root exudates was 42.6% at 32 days posthatching, which was significantly greater than that in the presence of soybean root exudates (30.3%) or sterile water (33.1%). In summary, the cyst nematode population parasitizing Henan tobacco was identified as a new subspecies, H. glycines sbsp.n. tabacum.

Characterization of microRNAs in the cyst nematode Heterodera glycines identifies possible candidates involved in cross-kingdom interactions with its host Glycine max.

The soybean cyst nematode (SCN - Heterodera glycines) is one of the most damaging pests to the cultivated soybean worldwide. Using a wide array of stylet-secreted effector proteins, this nematode can restructure its host cells into a complex and highly active feeding structure called the syncytium. Tight regulation of these proteins is thought to be essential to the successful formation of this syncytium. To date, multiple mechanisms have been proposed to regulate the expression of these proteins including through post-transcriptional regulation. MicroRNAs (miRNAs) are a class of small, roughly 22-nucleotide-long, non-coding RNA shown to regulate gene expression through its interaction with the 3' untranslated region of genes. These same small RNAs have also been hypothesized to be able to cross over kingdom barriers and regulate genes in other species in a process called cross-kingdom interactions. In this study, we characterized the miRNome of the SCN via sequencing of small-RNAs isolated from whole nematodes and exosomes representing all developmental stages. We identified 121 miRNA loci encoding 96 distinct miRNA families including multiple lineage- and species-specific candidates. Using a combination of plant- and animal-specific miRNA target predictors, we generated a unique repertoire of miRNA:mRNA interacting partners in the nematode and its host plant leading to the identification of a set of nine probable cross-kingdom miRNA candidates.

Soybean Cyst Nematode Management Is Improved by Combining Native and Transgenic Resistance.

Field trials were conducted to assess the benefit of combining a transgenic soybean cyst nematode (SCN) resistance trait, Cry14Ab-1 expressed by the event GMB151, with the native resistance allele rhg1b from PI 88788. The GMB151 event and rhg1b were crossed into common genetic backgrounds and segregated out to create four genetically related lines within each background. The lines created contained both native and transgenic resistance (rhg1b + GMB151), only native resistance (rhg1b alone), only transgenic resistance (GMB151 alone), or neither resistance type (susceptible). The benefit of GMB151 and rhg1b for SCN management was evaluated by measuring SCN control and yield protection. Soybean cyst nematode control was assessed by counting the number of females and cysts on roots early in the season and measuring the change in SCN egg population density over the entire season. The GMB151 transgenic event and the native resistance allele rhg1b both reduced early season SCN reproduction and contributed to significantly higher soybean yield. Compared to susceptible lines, the rhg1b allele improved yield by 33%, while GMB151 improved yield by 13%. Combining the GMB151 event and rhg1b allele resulted in greater SCN control and yield improvement than either provided alone. The combination of GMB151 and rhg1b reduced season-long SCN reproduction by 50% and resulted in 44% greater yield than the susceptible lines. Soybean cyst nematode virulence to rhg1b continues to increase due to the continuous planting of PI 88788-derived resistant cultivars. Pyramiding GMB151 with rhg1b provides a new management option to improve SCN control and soybean yield.

Interactions Among Heterodera glycines, Macrophomina phaseolina, and Soybean Genotype.

Heterodera glycines, the soybean cyst nematode (SCN), and fungal pathogen Macrophomina phaseolina are economically important soybean pathogens that may coinfest fields. Resistance remains the most effective management tactic for SCN, and the rhg1-b resistance allele derived from plant introduction 88788 is most commonly deployed in the northern United States. The concomitant effects of SCN and M. phaseolina on soybean performance, as well as the effect of the rhg1-b allele in two different genetic backgrounds, were evaluated in three environments (during 2013 to 2015) and a greenhouse bioassay. Within two soybean populations, half of the lines had the rhg1-b allele, and the other half had the susceptible allele in the backgrounds of the cultivars IA3023 and LD00-3309. Significant interactions between soybean rhg1-b allele and M. phaseolina-infested plots were observed in 2014. In all experiments, initial SCN populations (Pi) and M. phaseolina in roots were associated with reduced soybean yield. SCN reproduction factor (RF = final population/Pi) was affected by SCN Pi, rhg1-b, and genetic background. A background-by-genotype interaction on yield was observed only in 2015, with a stronger rhg1-b effect in the LD00-3309 background, which suggested that the susceptible parent 'IA3023' is tolerant to SCN. SCN female index from greenhouse experiments was compared with field RF, and Lin's concordance and Pearson's correlation coefficients decreased with increasing field SCN Pi in soil. In this study, both SCN and M. phaseolina reduced soybean yield asymptomatically, and the impact of SCN rhg1-b resistance was dependent on SCN virulence but also population density.

Single Nematode Transcriptomic Analysis, Using Long-Read Technology, Reveals Two Novel Virulence Gene Candidates in the Soybean Cyst Nematode, Heterodera glycines.

The soybean cyst nematode (Heterodera glycines, SCN), is the most damaging disease of soybean in North America. While management of this pest using resistant soybean is generally still effective, prolonged exposure to cultivars derived from the same source of resistance (PI 88788) has led to the emergence of virulence. Currently, the underlying mechanisms responsible for resistance breakdown remain unknown. In this study, we combined a single nematode transcriptomic profiling approach with long-read sequencing to reannotate the SCN genome. This resulted in the annotation of 1932 novel transcripts and 281 novel gene features. Using a transcript-level quantification approach, we identified eight novel effector candidates overexpressed in PI 88788 virulent nematodes in the late infection stage. Among these were the novel gene Hg-CPZ-1 and a pioneer effector transcript generated through the alternative splicing of the non-effector gene Hetgly21698. While our results demonstrate that alternative splicing in effectors does occur, we found limited evidence of direct involvement in the breakdown of resistance. However, our analysis highlighted a distinct pattern of effector upregulation in response to PI 88788 resistance indicative of a possible adaptation process by SCN to host resistance.

Prevalence of pest nematodes associated with soybean (Glycine max) in Wisconsin from 1998 to 2021.

The prevalence of Heterodera glycines and other cyst and vermiform genera was determined from 8,009 soil samples over two decades. Prevalence of cyst nematodes for farms increased from 16% in 1998 to 1999, reaching a peak of 40%, with marked differences among Wisconsin's nine agricultural districts in how much the odds of a positive test increased. Estimates at the sample scale also increased over time but peaked at 29%. Assay of all nematodes beginning in 2012 showed Pratylenchus, Helicotylenchus, and Xiphinema to be more prevalent in Wisconsin soybean fields than cyst nematodes. Prevalence estimates for Pratylenchus and Helicotylenchus for soybean and rotation crops ranged from 76 to 89% and 58 to 83%, respectively. Species identification of Pratylenchus from a subset of the samples revealed six species. The majority of cyst-positive samples were infested with Pratylenchus, and count data showed that the number of cyst eggs and juveniles per 100 cm3 soil was 60% lower in samples positive for Pratylenchus. The influence was reciprocal, as Pratylenchus population densities were 41% lower in samples positive for cyst nematodes, suggesting a competitive interaction. The Wisconsin soybean nematode testing program provides a useful model for estimating nematode prevalence using citizen-based surveys.

Seed Treatments for Management of Soybean Cyst Nematode, Heterodera glycines, in Mid-Atlantic Soybean Production.

Soybean Cyst Nematode (SCN), Heterodera glycines Ichinohe, is the most important pathogen of soybean in the Mid-Atlantic region. In recent decades, a decline in the effectiveness of genetic resistance has been observed and additional management approaches are needed. Seed treatments are of rising interest, but no local data on product response exists for the region. In 2020-2021, two experiments were conducted to observe the effects of chemical and biological seed treatment options. In one experiment, chemical seed treatments pydiflumetofen (Saltro®) and fluopyram (ILEVO®) were screened against nontreated plain seed for SCN suppression. In a second experiment, pydiflumetofen, fluopyram and four biological nematode-protectant seed treatments with a standard base insecticide and fungicide treatment were compared to nontreated plain seed and seed with only the standard base treatment to test product efficacy against SCN. Seed treatments increased the percent emergence over plain seed. Nematode reproductive factors and female counts from roots were collected, but did not statistically differ between seed treatments or plain seed. Yield differences were observed in one of the five trials, where pydiflumetofen + base seed treatment yielded the highest (p < 0.001) at 3813.1 kg/ha. Response from seed treatments varied, with no specific seed treatment consistently reducing SCN populations or increasing yield across trials. Seed treatments may have potential as an element of an integrated management approach for SCN.

Chemical Profiles of Heterodera glycines Suppressive Soils in Double Cropping Soybean Production.

We previously reported soybean fields double-cropped with winter wheat having reduced soybean cyst nematode (SCN) (Heterodera glycines) counts compared to fallow. A follow-up metagenomics study identified several fungal and bacterial taxa enriched in wheat fields, and some were reported to parasitize SCN. Knowing that phytocompounds with potential nematicidal activity are released via wheat roots and stubble, we implemented a dichloromethane-based extraction method and a gas chromatography-mass spectrometry (GCMS) system to investigate soil chemical profiles of samples collected from these fields and review the potential nematicidal activity of compounds with higher concentration in double cropping fields. 51 compounds were detected during the GCMS analysis, eight with unknown identification. Several compounds, including multiple fatty acids, had larger relative peak areas when double-cropped, compared to fallow samples. This study, along with our previously published one, provided a better understanding of the mechanisms that govern the effect of wheat on SCN populations. Rather than driven by a single mechanism, the suppression of SCN in soybean fields double-cropped with winter wheat was potentially linked to enriched microbial communities, increased populations of beneficial organisms, and higher concentrations of chemicals with potential nematicidal activity. To our knowledge, this is the first study using GCMS to characterize soil chemical profiles in soybean fields double-cropped with winter wheat regarding the suppression of SCN populations.

Arabidopsis non-host resistance PSS30 gene enhances broad-spectrum disease resistance in the soybean cultivar Williams 82.

Non-host resistance (NHR), which protects all members of a plant species from non-adapted or non-host plant pathogens, is the most common form of plant immunity. NHR provides the most durable and robust form of broad-spectrum immunity against non-adaptive pathogens pathogenic to other crop species. In a mutant screen for loss of Arabidopsis (Arabidopsis thaliana) NHR against the soybean (Glycine max (L.) Merr.) pathogen Phytophthora sojae, the Phytophthora sojae-susceptible 30 (pss30) mutant was identified. The pss30 mutant is also susceptible to the soybean pathogen Fusarium virguliforme. PSS30 encodes a folate transporter, AtFOLT1, which was previously localized to chloroplasts and implicated in the transport of folate from the cytosol to plastids. We show that two Arabidopsis folate biosynthesis mutants with reduced folate levels exhibit a loss of non-host immunity against P. sojae. As compared to the wild-type Col-0 ecotype, the steady-state folate levels are reduced in the pss1, atfolt1 and two folate biosynthesis mutants, suggesting that folate is required for non-host immunity. Overexpression of AtFOLT1 enhances immunity of transgenic soybean lines against two serious soybean pathogens, the fungal pathogen F. virguliforme and the soybean cyst nematode (SCN) Heterodera glycines. Transgenic lines showing enhanced SCN resistance also showed increased levels of folate accumulation. This study thus suggests that folate contributes to non-host plant immunity and that overexpression of a non-host resistance gene could be a suitable strategy for generating broad-spectrum disease resistance in crop plants.

Enhanced resistance to soybean cyst nematode in transgenic soybean via host-induced silencing of vital Heterodera glycines genes.

Soybean cyst nematode (SCN, Heterodera glycines Ichinohe) is the most economically damaging pathogen affecting soybean production worldwide. Host-induced gene silencing provides a promising approach to confer resistance to plant parasitic nematodes. In the present study, we produced stable transgenic soybean plants individually harboring the inverted repeats of three essential H. glycines genes, Hg-rps23, Hg-snb1, and Hg-cpn1, and evaluated their resistance to SCN infection. Molecular characterization confirmed the stable integration of the hairpin double stranded (ds) RNA in host plants. Inoculation assays with SCN race 3 showed significant reduction of female index (FI, 11.84 ~ 17.47%) on the roots of T4 transgenic plants, with 73.29 ~ 81.90% reduction for the three RNA interference (RNAi) constructs, compared to non-transformed plants (NT, 65.43%). Enhanced resistance to SCN race 3 was further confirmed in subsequent generations (T5) of transgenic soybean. Moreover, when inoculated with SCN race 4 which was considered highly virulent to most of soybean germplasms and varieties, transgenic soybean plants also exhibited reduced FIs (9.96 ~ 23.67%) and increased resistance, relative to the NT plants (46.46%). Consistently, significant down-regulation in transcript levels of the Hg-rps23, Hg-snb1, Hg-cpn1 genes were observed in the nematodes feeding on the transgenic roots, suggesting a broad-spectrum resistance mediated by the host-mediated silencing of vital H. glycines genes. There were no significant differences in morphological traits between transgenic and NT soybean plants under conditions with negligible SCN infection. In summary, our results demonstrate the effectiveness of host-induced silencing of essential H. glycines genes to enhance broad-spectrum SCN resistance in stable transgenic soybean plants, without negative consequences on the agronomic performance.

Assessing Direct and Residual Effects of Cover Crops on the Soybean Cyst Nematode, Heterodera glycines.

Greenhouse experiments were conducted to determine if cover crops directly decrease population densities of the soybean cyst nematode (SCN), Heterodera glycines, and/or have residual effects on reproduction of the nematode on soybean (Glycine max). Population densities of SCN were not significantly decreased by nine cover crop plants or three cover crop mixes compared with a non-planted soil control in a repeated 60-day-long greenhouse experiment. When susceptible soybeans were grown in the soils after cover crop growth, fewer SCN females formed after three annual ryegrass (Lolium multiflorum) cultivars (Bounty, King, and RootMax), the Daikon radish (Raphanus sativus var. longipinnatus) cultivar CCS779, Kodiak mustard (Brassica juncea), and a mix containing cereal rye, crimson clover (Trifolium incarnatum), plus Daikon radish (cultivars not stated) compared with following the non-planted control. In another repeated experiment, cover crops were grown for 56 days in SCN-infested soil in the greenhouse then exposed to Iowa winter conditions for 28 days to simulate winter termination of the plants. One treatment, a cover crop mix containing 'Bounty' annual ryegrass plus 'Enricher' Daikon radish, had a decrease in SCN population density greater than the non-planted control at the end of the experiment. Significantly fewer SCN females formed on soybeans grown after several cover crops, including the three annual ryegrass cultivars that had the suppressive residual effects in the first experiment. In summary, there were no cover crop treatments that consistently decreased SCN population densities across experiments, and only one cover crop treatment in one experiment significantly reduced SCN population densities more than a non-planted soil control. However, there was a somewhat consistent, adverse, residual effect of cover crops on reproduction of SCN on susceptible soybeans after growth of multiple cover crops.

Functional Characterization of Ubiquitination Genes in the Interaction of Soybean-Heterodera glycines.

Ubiquitination is a kind of post-translational modification of proteins that plays an important role in plant response to biotic and abiotic stress. The response of soybean GmPUB genes to soybean cyst nematode (SCN, Heterodera glycines) infection is largely unknown. In this study, quantitative real-time PCR (qRT-PCR) was performed to detect the relative expression of 49 GmPUB genes in susceptible cultivar William 82 and resistant cultivar Huipizhi after SCN inoculation. The results show that GmPUB genes responded to cyst nematode infection at 1 day post-inoculation (dpi), 5 dpi, 10 dpi and 15 dpi. The expression levels of GmPUB16A, GmPUB20A, GmCHIPA, GmPUB33A, GmPUB23A and GmPUB24A were dramatically changed during SCN infection. Furthermore, functional analysis of these GmPUB genes by overexpression and RNAi showed that GmPUB20A, GmPUB33A and GmPUB24A negatively regulated soybean resistance under SCN stress. The results from our present study provide insights into the complicated molecular mechanism of the interaction between soybean and SCN.

Soybean Cyst Nematode Population Development and its Effect on Pennycress in a Greenhouse Study.

Midwest crop production is dominated by two summer annual crops grown in rotation, viz., corn (Zea mays L.) and soybean (Glycine max L.). Winter oilseed crops, such as pennycress (Thlaspi arvense L.), can provide ecosystem and economic benefits when added to the corn-soybean rotation. However, adding a new crop adds risks, such as increased pest pressure. The objectives of this study were to (i) evaluate population development of three soybean cyst nematode (SCN; Heterodera glycines) biotypes on three pennycress genotypes and susceptible soybean and (ii) determine whether SCN inoculation level influenced plant biomass. SCN population density and biomass were determined after 60 d in the greenhouse. At the inoculation level of 2,000 eggs/100 cm3 soil, the average egg density for the three pennycress genotypes was 1,959 eggs/100 cm3 soil, lower than that for the susceptible soybean 'Sturdy' (9,601 eggs/100 cm3 soil). At the inoculation level of 20,000 eggs/100 cm3 soil, the average egg density for the three pennycress genotypes was 6,668 eggs/100 cm3 soil, lower than that for 'Sturdy' (40,740 eggs/100 cm3 soil). The inoculation level did not affect plant biomass. Pennycress is an alternative host to SCN under greenhouse conditions but is a less suitable host than soybean.

t-SNAREs bind the Rhg1 α-SNAP and mediate soybean cyst nematode resistance.

Soybean cyst nematode (SCN; Heterodera glycines) is the largest pathogenic cause of soybean yield loss. The Rhg1 locus is the most used and best characterized SCN resistance locus, and contains three genes including one encoding an α-SNAP protein. Although the Rhg1 α-SNAP is known to play an important role in vesicle trafficking and SCN resistance, the protein's binding partners and the molecular mechanisms underpinning SCN resistance remain unclear. In this report, we show that the Rhg1 α-SNAP strongly interacts with two syntaxins of the t-SNARE family (Glyma.12G194800 and Glyma.16G154200) in yeast and plants; importantly, the genes encoding these syntaxins co-localize with SCN resistance quantitative trait loci. Fluorescent visualization revealed that the α-SNAP and the two interacting syntaxins localize to the plasma membrane and perinuclear space in both tobacco epidermal and soybean root cells. The two syntaxins and their two homeologs were mutated, individually and in combination, using the CRISPR-Cas9 system in the SCN-resistant Peking and SCN-susceptible Essex soybean lines. Peking roots with deletions introduced into syntaxin genes exhibited significantly reduced resistance to SCN, confirming that t-SNAREs are critical to resisting SCN infection. The results presented here uncover a key step in the molecular mechanism of SCN resistance, and will be invaluable to soybean breeders aiming to develop highly SCN-resistant soybean varieties.

Characterization of Virulence Phenotypes of Soybean Cyst Nematode (Heterodera glycines) Populations in North Dakota.

Soybean cyst nematode (SCN; Heterodera glycines) continues to be the greatest threat to soybean production in the United States. Because host resistance is the primary strategy used to control SCN, knowledge of SCN virulence phenotypes (HG types) is necessary for choosing sources of resistance for SCN management. To characterize SCN virulence phenotypes in North Dakota, a total of 419 soybean fields across 22 counties were sampled during 2015, 2016, and 2017. SCN was detected in 42% of the fields sampled, and population densities in these samples ranged from 30 to 92,800 eggs and juveniles per 100 cm3 of soil. The SCN populations from some of the infested fields were virulence-phenotyped with seven soybean indicator lines and a susceptible check ('Barnes') using the HG type tests. Overall, 73 SCN field populations were successfully virulence-phenotyped. The HG types detected in North Dakota were HG types 0 (frequency rate: 36%), 7 (27%), 2.5.7 (19%), 5.7 (11%), 1.2.5.7 (4%), and 2.7 (2%). However, before this study only HG type 0 was detected in North Dakota. The designation of each of these HG types detected was also validated by repeating the HG type tests for 33 arbitrarily selected samples. This research for the first time reports several new HG types detected in North Dakota and confirms that the virulence of SCN populations is shifting and overcoming resistance, highlighting the necessity of using different resistance sources, rotating resistance sources, and identifying novel resistance sources for SCN management in North Dakota.

Genomic Profiling of Virulence in the Soybean Cyst Nematode Using Single-Nematode Sequencing.

Soybean cyst nematode (SCN) is one of the most important diseases in soybean. Currently, the main management strategy relies on planting resistant cultivars. However, the overuse of a single resistance source has led to the selection of virulent SCN populations, although the mechanisms by which the nematode overcomes the resistance genes remain unknown. In this study, we used a nematode-adapted single-cell RNA-seq approach to identify SCN genes potentially involved in resistance breakdown in Peking and PI 88788 parental soybean lines. We established for the first time the full transcriptome of single SCN individuals allowing us to identify a list of putative virulence genes against both major SCN resistance sources. Our analysis identified 48 differentially expressed putative effectors (secreted proteins required for infection) alongside 40 effectors showing evidence of novel structural variants, and 11 effector genes containing phenotype-specific sequence polymorphisms. Additionally, a differential expression analysis revealed an interesting phenomenon of coexpressed gene regions with some containing putative effectors. The selection of virulent SCN individuals on Peking resulted in a profoundly altered transcriptome, especially for genes known to be involved in parasitism. Several sequence polymorphisms were also specific to these virulent nematodes and could potentially play a role in the acquisition of nematode virulence. On the other hand, the transcriptome of virulent individuals on PI 88788 was very similar to avirulent ones with the exception of a few genes, which suggest a distinct virulence strategy to Peking.

Occurrence of the White Soybean Cyst Nematode, Heterodera sojae, and the Soybean Cyst Nematode, H. glycines, in Korea.

Soybean cyst nematode (Heterodera glycines, SCN) is the most harmful pathogen of soybean (Glycine max (L.) Merr.) worldwide. In 2016, a new soybean-parasitic cyst nematode, Heterodera sojae (the white soybean cyst nematode) was found parasitizing the roots of soybean plants in Korea. To investigate the distribution and population density of H. sojae, 943 soil samples were collected from soybean fields in all nine provinces in Korea in 2017 to 2018. Cyst nematodes were detected in 343 samples (36.4%) from eight of the nine provinces, except the island of Jeju province. Among the 343 samples, H. glycines was found in 227 samples (66.2%), H. sojae in 95 samples (27.7%), and 21 samples (6.1%) were infested with both H. sojae and H. glycines. Wide distribution of H. sojae in soybean fields indicates that H. sojae is an important cyst nematode species parasitizing soybean together with H. glycines.