Updated April 6, 2022:
A description of relevant progress for principle and co-principle investigators is below for each objective and sub objective in our proposal.
Objective 1.1: The Baum group has accomplished an analysis of gene expansions and contractions, as well as the presence and absence of gene families in H. glycines compared to 13 related species in the Tylenchomorpha. We are functionally characterizing these gene families and their association with previously published H. glycines effectors, as well as their secretory status, expression, nuclear localization, and impact of their variability across 15 populations of H. glycines. We have shown that 551 gene expansions in sedentary nematodes differentiate them from migratory nematodes, 124 gene expansions in cyst nematodes differentiate them from root-knot nematodes, and 1,100 gene expansions in H. glycines differentiates it from Globodera species. We have also shown a number of gene gain and loss events that have directly impacted the H. glycines genome, many of which have shown an atypical inheritance pattern across related species. In one instance, we show 175 gene families that are only shared between H. glycines and Meloidogyne species, 15 of which are targeted to the secretory pathway, including two predicted H. glycines effectors. Using effectors from relatively closely related species, we have identified 405 putative effector families present in H. glycines, though only 137 of these families contain a gene that produces a protein with a signal peptide for secretion. Of these 137 putative effectors, 11 were associated with gene expansions in sedentary nematodes compared to migratory nematodes, 6 were associated with the expansions of gene families in cyst nematodes versus root-knot nematodes, and 24 were associated with the expansion of Heterodera gene families in comparison to Globodera. Using population variation from 15 distinct populations of H. glycines, we were able to show which genes typically produce secreted proteins across the assayed SCN lines. In this respect, we show that 59 gene expansion families have consistent signals for secretion across multiple populations, even though they were not predicted to be secreted in the previously published TN10 genome. These results warrant further research into gene evolution. We also have developed plans to update SCNBase with new RNAseq from the recently published J2 gland RNAseq and will integrate gene family criteria. To complement the methods of identifying genes involved in the parasitism of H. glycines, we have undertaken a genome assembly and annotation project of male and female specimens. Males and females dramatically change their parasitic behavior with the onset of adulthood and, thus, studying their genomes and transcriptomes will reveal insights into gene functions, particularly for effectors. We sequenced male and female genomes using nanopore long reads and added male and female specific RNA-seq to this analysis to better predict the gene variation among the sexes. Since the last report we have scaffolded these male and female genomes with HiC to obtain 9 pseudomolecules for each genome. Interestingly, we found large differences in genome size between these nanopore genomes (male: 115.5Mb, female: 112.6Mb) and our previously published TN10 genome (158Mb), which is likely attributable to the differences in sequencing technology and assembly software. Subsequently, we used male and female RNAseq with Braker to annotate genes in each genome, finding that the disparity continues between these nanopore genomes and TN10 genome at the genic level. In the male and female gene annotations, we found 16,421 genes and 16,530 genes, respectively. These gene annotation totals are substantially reduced from our previous annotation of the TN10 genome at 22,465 genes, though the gene copy number reduction is in proportion with the genome size reduction. We have begun to compare the genomes at the level of gene structure and expression using Orthofinder and differential expression analyses. Thus far we have found that 9-10,000 of differentially expressed genes cluster to the same orthologue family, leaving slightly more than half of the genes with significant divergence between the sexes. The Mitchum group submitted the following manuscript during the past quarter - Verma A, Lin M, Smith D, Lee C, Walker JC, Hewezi T, Davis EL, Hussey RS, Baum TJ, Mitchum MG. A novel cyst nematode effector (2D01) targets the Arabidopsis HAESA receptor-like kinase. Mol. Plant-Microbe Interact.
Objective 1.2: Phase III of this project sees the Baum group developing further resources to expand the toolbox that will aid us in understanding SCN virulence. Focusing once again on the three gland cells that SCN uses to produce the tools (effectors) required for establishment of successful infection, as well as defense suppression, we have improved on the technology used in phase II. Taking advantage of recent developments toward single cell sequencing, and specifically new technology available for the generation of single cell RNA-seq libraries, we have applied these technologies to our work with gland cell isolation and transcriptomics in SCN. We successfully generated single cell RNA-seq libraries for our avirulent (PA3) and virulent (MM10) populations. These libraries represented four biological replications of 5 dorsal and subventral gland cells per rep. This was our group’s first attempt at applying single cell sequencing technologies to SCN transcriptomics and required a fair amount of optimization. Through this process, we developed a technique for live gland cell collection, versus fixed tissue collection, which seems to have improved the quality of RNA collected and allowed us to collect picogram quantities of RNA from individual gland cells. The analysis of the libraries generated via the single cell sequencing approach show that these libraries are as good, if not slightly better, than the libraries previously generated for this project and previously in our lab group. The overall coverage of genes identified within these new J3 single cell libraries, with reference to the SCN TN10 genome is slightly greater than the coverage of genes identified with our previous pooled gland cell parasitic J2 libraries. We identified a total of 14,667 and 14,000 genes at a normalized read count of at least 5 counts from the SCN PA3 J3 and MM10 J3 libraries, respectively. This is compared to a total of 12,495 and 12,289 genes at the same read count cutoff for the respective SCN populations with our older technology. While we cannot rule out that this could be due to life stage differences, it certainly points to the fact that this new technology can yield similar, if not better numbers of identified genes from lesser amounts of material. We are currently working on the transcriptomics to generate both broad and specific comparisons of both the life stages and virulence differences that exist in our two SCN reference populations. This will provide an extensive and novel look at SCN virulence at this level.
All seven of the genomes of the additional Hg types are now assembled, and the assemblies are completed and frozen. They are ready to distribute or submit to NCBI, however we need to annotate and analyze them before publishing a paper on the genomes, and to make them useful to more people in the group.
The Hudson group has been proceeding with analysis in several ways. Firstly, genome synteny for the 7 newly assembled SCN strains was compared with mummer (dotplots) and circos (BLAST hits) to the chromosome level SCN assembly TN10 (Masonbrink et al 2021). These data showed that the assembled 9 chromosomes were in similar structure with the previously reported TN10, although not identical. We started the genome functional and structural annotation. Repetitive elements modeled with RepeatModeler and quantified repetitive content with RepeatMasker. The repeat analysis showed repeat content was similar across strains with respect to repeat class and quantity. Subsequently, we began gene prediction using RNAseq evidence (previously reported Gardner et al 2018 & Lian et al 2019) and protein models (from previously reported TN10 Masonbrink et al 2021) and other cyst nematodes including Globodera pallida (Cotton et al 2014) and Globodera rostochiensis (Eves-van der Akker 2016). Preliminary gene models show similar structure to the previous assemblies gene models. Our future work will include refinement of gene models and subsequent functional annotation of the gene models.
Objective 1.3: The Baum group has indicated that characterization of the function of 28B03 effector family has been advanced to a natural stopping point to publish the first extensive report on this effector’s function during parasitism. A complete manuscript has been written and all data compiled. We are waiting on last additions and reviews from co-authors and then will submit the manuscript to start the publication process. Given the thorough study of this effector, this manuscript is a very extensive and thorough functional assessment of this effector and its ability to interfere with a plant signal transduction pathway. In short, 28B03 targets a novel plant kinase protein, which in turn cooperatively with another plant kinase leads to the initiation of signal transduction processes that initiate a subset of plant defense responses. We have shown that the 28B03 effector interferes with this signal transduction, thus, compromising plant defenses and leading to increased host susceptibility. Through our confocal experiments utilizing co-localization studies of 28B03 and the identified kinases, we can infer that our proposed cascade model is valid, as the effector and kinases are co-localized together in these assays. This work identifies a potential plant target to increase plant resistance (i.e., one can now devise mechanisms to interfere with the inhibitory function of 28B03 to prevent the nematode from inactivating a plant defense signal transduction kinase).
The Mitchum lab collected sufficient genetic material of two pairs of SCN populations (unadapted or adapted to reproduce on resistant soybeans) and optimized their DNA extraction procedure to meet the stringent requirements necessary for the Pool-Seq strategy. We have completed the sequencing and bioinformatic analysis is currently underway. The Pool-Seq approach should help guide us toward the candidate virulence regions important for breaking the Peking-type (Rhg4-mediated) resistance.
Objective 2: The Diers group is preparing and distributing seed for all collaborators for the 2022 SCN resistance source rotation study. This will be the fourth year of the rotation study and we will be rotating the plots back to what was grown in them during 2020. We now have egg numbers from the plots grown in 2019-2021 and HG type values from 2019-2020. These results show that continuous planting of PI 90763 had the lowest egg number increases in plots across the three years. However, the continuous production of this source of resistance is selecting nematodes that can overcome this resistance and the female index (FI) in plots grown with PI 90763 was 41 after 2020, which indicates that the nematode population in these plots may start increasing. The rotation that had the lowest increase in egg numbers over the three years is rhg1b+soja+ch10 in 2019 followed by PI 90763 in 2020 and then rhg1b+soja+ch10 in 2021. This rotation also showed a low increase in egg numbers in other states and did not increase the FI on PI 88788 or Peking. The study will be repeated in 2022 to provide further insight into these resistance sources.
The Scaboo group processed soil samples for determining egg density and HG types for each microplot. The egg density results showed that there is an increasing trend in SCN population density in the third year of rotation except for plots with continuous PI 90763 and rhg1-b+G. soja+10 rotated with PI 90763. There was also a significant reduction in the percentage change in the egg count for these two treatments. The HG type data also showed that the continuous rhg1-a + rhg4 and continuous PI 90763 treatments facilitated the development of more virulent nematode populations with HG type (HG type: 1.2.3-; race 4), where nematode had adapted on the Peking type resistance sources (PI 90763, Peking and Pickett). Similarly, nematode populations from treatments involving rotations of rhg1-b (and/or stacked resistances) with PI 90763 showed to have reduced female index on PI 88788, PI 90763, and Peking indicator lines (Figure 2B). We look forward to the upcoming planting season where we plan on rotating an additional cycle of continuous and rotated schemes as conducted previously in 2020.
The Mitchum group was sent SCN material recovered from the continuous and rotation microplots and they were increased, processed for eggs, and archived as part of a wormplasm collection for future sequencing efforts to pinpoint virulence genes.
At the conclusion of the 2021 growing season, The Tylka group collected two separate multi-core soil samples from each microplot in the experiments conducted in central Iowa and north central Iowa. One set of soil samples from each experiment were processed at Iowa State University to determine the end-of-season SCN egg population density in each microplot. The second set of soil samples were sent to the University of Missouri for HG Type testing to determine how the soybean genotypes grown in the microplots in 2021 have affected or shifted the virulence profiles (HG types) of the SCN populations from the 2020 growing season and from the initial SCN populations that were added to the microplots in the spring of 2019.
Preliminary data analysis show some trends in changes SCN population densities. In both experiments, the greatest SCN population densities occurred in microplots in which the susceptible soybean variety was grown. Most of the microplots that had continuous cropping of the same resistance in 2019, 2020, and 2021 had greater SCN population densities than microplots in which resistant genotypes were rotated in 2019, 2020, and 2021. The lowest population densities occurred in microplots where soybeans with SCN resistance from PI 90763 were grown. The microplots in which soybeans with rhg1-b, rhg1-b + soja, and rhg1-b + soja + ch10 SCN resistance were grown (collectively referred to as genotypes having “PI88788-type” resistance) in 2019, rotated to soybeans with PI 90763 SCN resistance in 2020, and then rotated back to the same resistance as in 2019 had increased SCN population densities in 2021 compared to population densities at the end of both previous years. Even though SCN population densities declined after rotating from genotypes with PI88788-type resistance in 2019 to rhg1-a + rhg4 (or “Peking-type”) resistance in 2020, population densities increased to levels greater than in 2019 and 2020 when genotypes with PI88788-type resistance were again grown in the microplots in 2021. Similarly, plots in which rhg1-a + rhg4 or Peking-type resistance was grown in 2019 then rotated to rhg1-b, rhg1-b + soja, or rhg1-b + soja + ch10 SCN (PI88788-type resistance) resistance in 2020, and then back to rhg1-a + rhg4 resistance in 2021 had greater SCN population densities at the end of the 2021 growing season than in the previous two years.
The results of the HG Type tests on the SCN populations in the microplots at harvest in 2020 revealed that almost all of the SCN populations at both experimental locations had increased virulence from 2019, with the SCN populations in each plot having elevated female indices (FI) on Peking and most having increased FIs on PI 88788. SCN populations in the two experiments were HG Type 1.2 or 1.2.3 and the FIs ranged from 9-33% on Peking in Kanawha, 18-53% on Peking in Ames, 40-64% on PI 88788 in Kanawha and 33-64% on PI88788 in Ames. In both experiments, the SCN populations in microplots in which PI 90763 or rhg1-a + rhg4 (Peking-type) resistance was grown had elevated FIs on PI 90763, with a range of 0%- 23% in Kanawha and 2%-32% in Ames. The SCN populations in microplots that were rotated from rhg1-a + rhg4 resistance to the three different genotypes containing rhg1-b (namely rhg1-b, rhg1-b + soja, and rhg1-b + soja + ch10) had decreased FIs on PI 90763 from 2019 to 2020. Changes in virulence (FIs) on PI437654 were not detected in SCN populations in any of the microplots with any of the cropping sequences. HG Type test results of the SCN populations in soil samples collected from the microplots at harvest in 2021 are not yet available.
Objective 3: Greg Tylka conducted 19 interviews with radio and newspaper/magazine journalists and gave 12 presentations (in person and virtual) from October 2021 through March 2022. The loss of effectiveness of PI88788 SCN resistance was discussed in every interview and presentation, and this current NCSRP-funded research project was mentioned and described whenever time/space permitted.
Melissa Mitchum is serving as the Chair of the organizing committee for the 2022 National Soybean Nematode Conference (NSNC). The location, venue, and draft schedule was developed during this reporting period. Save the date flyers and the scientific program will be developed and announced in the next reporting period.
Objective 4: The Diers group has continued to organize these tests. The results from the 2021 SCN Regional Test were received from cooperators and summarized in a report. The initial version of the report was sent to cooperators on December 16th and the final version was delivered on January 13th. These timely deliveries of results are important so cooperators can make decisions on selections in time for winter crosses and nurseries. Plans have been made for the 2022 SCN Regional Test. This test will include 225 entries that range from MG 0 to IV. The tests have been organized, the seed has been received from cooperators, repackaged, and is being shipped to cooperators. Arrangements also have been made to test the lines for SCN resistance in a greenhouse at the University of Missouri. We are moving the tests to the University of Missouri because the nematology lab at the University of Illinois has been slow in providing test results.
Objective 5: The Diers and Scaboo groups have continued to advance breeding efforts towards the development of cultivars with novel SCN resistance. For this reporting period, we are excited to report that we have now completed successful crossing attempts (3 back-crosses) using PI 90763 as a donor parent, and LD11-2170 and SA13-1385 as recurrent parents, for three major genes associated with resistance to virulent nematode populations (rhg1-a, rhg2, and Rhg4). For each crossing attempt, we have identified desirable F1 plants using marker assisted selection, and we have sped up the process by utilizing our winter nurseries in Hawaii and Puerto Rico for the last two years. As I type this report, our staff and student are in Kekaha Kauai Hawaii tissue sampling BC3F2 plants to identify homozygous individuals with desired combinations of our target genes. During the summer of 2022, we will grow plant rows derived from selected plants, and our first yield trials of this material will be in the summer of 2023.
Updated November 2, 2022:
A description of relevant progress for principle and co-principal investigators is below for each objective and sub objective in our proposal. Our team has made tremendous progress in accomplishing our research goals, conducting field experiments, publishing refereed journal articles, and communicating our results to scientists and soybean producers. We had a group meeting in March of 2022 to discuss current research progress and goals and we are on track to continue our cutting-edge research in soybean cyst nematode biology, management, and breeding for novel resistance.
Objective 1: Identify SCN virulence genes to better understand how the nematode adapts to reproduce on resistant varieties.
Sub-objective 1.1: Combine, compare, and catalogue the genomes that compromise the SCN pan-genome. (Hudson, Baum, Mitchum)
Previously the Baum group reported on gene family expansion and contraction across 13 plant-parasitic species of the Tylenchomorpha. We now have annotated these genes including secretory status, effector homology, nuclear localization, gene variability across 15 populations, and expression across various stages of the H. glycines lifecycle. We found 551 gene expansions in sedentary nematodes differentiate them from migratory nematodes, 124 gene expansions in cyst nematodes differentiate them from root-knot nematodes, and 1,100 gene expansions differentiates H. glycines from Globodera species. One interesting finding from this analysis lies with the inability of nematodes to produce their own cholesterol. To obtain this vital resource, plant-parasitic nematodes must acquire cholesterol from their hosts. We found that a gene family of expanded and secreted genes in cyst nematodes may be involved with this process. SCNBase has undergone many updates since our last report. We have modified the naming schemes of the predicted proteomes, transcriptomes, and genomes so that they are consistent across genomics tools and more intuitive for users to access. To further disseminate H. glycines genomics resources, we collaborated with Wormbase to host the most current H. glycines genome assembly and annotation. To better understand the biology of male and female H. glycines nematodes, we previously reported a genome assembly and annotation project for each sex. We now assessed differences in expression between the sexes. In this analysis there were numerous on/off differences in expression with one sex having high expression and the other sex completely lacking expression: 512 genes in females and 744 genes in males were silenced. A comparison of expressed genes revealed 6,543 genes were upregulated in females and 6,920 were upregulated in males. Many expression differences were tied to sex-related gene functions, though few have been reported in the literature for nematodes in the Tylenchomorpha.
Previously, the Mitchum lab employed a dual effector prediction strategy that coupled the traditional secreted protein prediction strategy with a newly developed nematode effector prediction tool, N-Preffector, to identify novel effector candidates in a de novo transcriptome assembly of the pre-parasitic and parasitic life stages of H. glycines with potential roles in virulence. From this analysis, 1,383 SignalP positive, N-Preffector positive candidates were identified, of which 210 were upregulated in parasitic juvenile life stages (Gardner et al., 2018). This transcriptome analysis, which preceded the release of the pseudomolecule genome assembly generated through this project (Masonbrink et al., 2021), represented sequences from the whole-nematode. Since then, in collaboration with the Baum lab, we also generated a gland cell-specific RNA-seq resource for H. glycines representing an avirulent and virulent population (Maier et al., 2021). Thus, to further narrow the 210 candidates to those that may be expressed in the nematode esophageal gland cells and likely function in virulence, we carried out an in silico in situ analysis by cross comparing this list with the gland RNA-seq dataset. Effectors upregulated in the transcriptomic data but missing from the gland data were eliminated. This in silico comparison narrowed down the list of candidate effectors which were Signal P, N-Preffector positive with some evidence of expression in nematode gland cells to 123 candidates. The predicted candidate effector protein sequences were further analyzed for nuclear localization signals (NLSs). One or more NLSs were predicted in 32 putative effectors. Sequences hitting to known effector sequences of plant-parasitic nematodes or housekeeping genes were rejected, reducing the list to eight novel candidate effectors with high to moderate expression in the gland RNA-seq dataset. We mined the SCN pseudomolecule genome assembly to determine the gene structures and genomic organization of these sequences. These were selected for further analysis in sub-objective 1.3 below.
Sub-objective 1.2: Resequencing of the genomes and transcriptomes of virulent SCN populations and conduct comparative analyses. (Hudson, Mitchum, Baum)
Building upon our prior success with generating novel SCN gland cell-specific libraries, we are developing additional SCN life-stage specific libraries which will provide important transcriptomic data on development stages not previously available. Currently underway in the Baum lab are SCN gland cell-specific libraries for the pre-infective J2 life stage, for both the avirulent (PA3) and virulent (MM10) populations previously used. This will provide valuable data on the early stages of nematode development and effector activation, which can then be compared to later parasitic stages. Additionally, we are now able to separate, identify and collect subventral glands, specifically. We can then generate subventral gland cell-specific RNA-seq libraries, which will identify which transcripts are specific to the subventral gland. We can then use that as a “subtraction” for the genes that are transcribed in our libraries generated from both types of gland cells, at the same life stage, and can infer from that which genes are transcribed in the dorsal glands. Having this gland specific transcriptome will be immensely powerful in elucidating effector timing and function, given what we already know about the SCN parasitic lifestyle. Effectors involved in host invasion and initiation of the syncytium are typically thought of as being produced in the subventral glands and active early in the SCN life cycle. Effectors that are involved in syncytial maintenance and host defense suppression are thought of as being produced in the dorsal gland, which becomes active later in the SCN life cycle. By finally having a well-defined transcriptome for each cell type, we can once-and-for-all confirm these observations and potentially find additional novel subventral and dorsal-specific effectors. Additionally, by also having these cell-specific transcriptomes that vary by virulence, we can identify temporally variant effectors that also may exist in different configurations based on virulence.
To perform single SCN-J2 genome sequencing, The Hudson group tested different DNA extraction methods and kits to find a method that gave sufficient DNA quality and quantity to get whole genome data on a single nematode. Genomic DNA samples from multiple kits were then sent to the Roy J. Carver Biotechnology Center on campus to assess first DNA quality, then quality of the library construction process, and finally trial sequencing to select the proper kit. We are now collecting 200 DNA samples from “MM1” SCN population and expect to receive a second population after finishing the first one. We will likely be able to send the whole samples (400/two populations) for sequencing around mid-November. The annotations and assemblies for the seven SCN Hg types have been finalized and are ready to submit once collaborators have completed analysis.
The Mitchum lab completed the preliminary bioinformatics analysis of the Pool-Seq data derived from sequencing two pairs of SCN populations unadapted or adapted to reproduce on resistant soybeans. We have been running multiple software packages to identify genomic regions and candidate genes potentially involved in overcoming resistance. By calculating population differentiation estimated from single nucleotide polymorphism (SNP) data, we identified five genomic regions spanning four chromosomes which contained distinct peaks formed by clusters of SNPs, indicating strong signatures of selection. Some candidate regions were detected in both pairs of SCN populations, while others were unique genomic regions under selection in each contrast. Interestingly and as expected, some of these include genes known to be involved in plant defense suppression. Therefore, we hypothesize that these genomic regions and their genes may have undergone selection pressure to overcome soybean resistance to SCN. We are currently using other tools to confirm extra evidence of selection in these candidate regions.
Sub-objective 1.3: Validate and characterize genes associated with SCN virulence and evaluate their utility as novel resistance targets. (Mitchum, Baum)
The Baum group has reported that characterization of the function of 28B03 effector family has been advanced to a natural stopping point to publish the first extensive report on this effector’s function during parasitism. A complete manuscript is being finalized. Furthermore, we are exploring how the knowledge gained for 28B03 can be used to develop novel management tools against SCN. In addition, we are adapting new techniques for use in our laboratory to functionally study SCN virulence determinants identified in our genomic assessments described above. For this purpose, we have streamlined the use of soybean hairy roots as parts of composite plants as a powerful tool to study SCN genes. Also, we are in the process of constructing a set of cloning and expression vectors to be used in the composite hairy-root plants. These vectors will allow ease of cloning and gene transfers between the different vectors which will aid the high throughput study of their functions in soybean roots. Also, we are establishing different approaches to determine and study the interacting soybean proteins for SCN effectors. Finally, we are establishing a reliable methodology to routinely silence SCN genes by soaking them in double-stranded RNA. All these advances will aid the further functional characterization of the genes identified in our genomic studies described above and will unravel the mechanisms that determine SCN virulence. Such knowledge is critical when designing novel SCN control tools.
The manuscript submitted by the Mitchum lab during the past quarter - Verma A, Lin M, Smith D, Lee C, Walker JC, Hewezi T, Davis EL, Hussey RS, Baum TJ, Mitchum MG. A novel cyst nematode effector (2D01) targets the Arabidopsis HAESA receptor-like kinase was revised and accepted for publication in the journal Molecular Plant Pathology this quarter. It was also selected by the Editor in Chief for a highlight on the British Society of Plant Pathology website emphasizing the scientific/societal significance and impact of this study and images from the paper will be featured on the cover of the December 2022 issue. Further characterization of the 8 novel effector candidates identified under objective 1.1 is underway. We have profiled the expression of these genes in SCN throughout the life cycle and initiated studies to confirm where they effectors potentially localize within host cells after secretion by the nematode.
Objective 2: Complete the evaluation of how rotations of various resistance gene combinations impact SCN field population densities and virulence profiles. (Diers, Scaboo, Tylka, Mitchum)
To start the 4th year of the project, the Tylka group planted each microplot with the 4th year planting scheme (same as year 2) in May 2022 and collected multi-core soil samples from each microplot in the experiments conducted in central Iowa and north central Iowa. After 30 days, plants in each microplot were thinned to 30 plants per row, monitored throughout the rest of the growing season, and hand weeded to control weed populations. Soybeans at both locations were harvested in late October and two soil samples were collected from each microplot. One set of soil samples from each experiment will be processed at Iowa State University to determine the end-of-season SCN egg population density in each microplot, and the other samples were sent to the University of Missouri for HG type testing to determine how the virulence profiles (or HG types) of the SCN population in each microplot may have been affected (shifted) by the soybean genotypes grown in the microplots in 2022 and in previous years. HG Type test results for samples taken at harvest in 2021 were received in mid-June 2022. Preliminary results revealed increased virulence in many of the SCN populations at both locations from 2020, although those grown in rotation with rhg1-a + rhg4 (Peking-type) showed a reduced female index (FI). SCN populations in the two experiments were HG Type 1.2 or 1.2.3 and the FIs ranged from 5-51% on Peking in Kanawha, 3-68% on Peking in Ames, 12-66% on PI 88788 in Kanawha and 10-65% on PI 88788 in Ames. The average FI on PI 88788 was reduced from 2020 in every treatment in Kanawha, while at Ames the female indices of all treatments were reduced except for those grown in rotation with the Rhg1-b pyramided alleles and PI 90763 combination.
The fourth year of tests with the different gene combinations were planted by the Diers group in the rotation field this spring and the plots were maintained through the growing season. Soil samples will be taken soon to study the impact of the rotations on the nematode population levels and HG types.
The fourth year of the field experiment was planted by the Scaboo group this spring to the continuous and rotated schemes as conducted previously in 2020. Soybeans were recently harvested and soil samples will be collected in the near future for SCN egg count and HG type testing. We will begin processing these samples directly after they are collected. SCN egg count data will be available shortly after processing to quantify egg density from each microplot. HG type testing will take several months to increase and characterize the SCN population from each microplot.
Objective 3: Translate the results of objectives 1-3 to the SCN Coalition to increase the profitability of soybean for producers and inform growers on effective rotation schemes designed to protect our resistant sources. (Tylka, Mitchum)
Melissa Mitchum continued to chair the planning committee of the 2022 National Soybean Nematode Conference, and Tylka is a member of the committee as well. The conference will be held in mid-December 2022. The planning committee has identified and invited speakers on important soybean nematode-related topics, which include the loss of effectiveness of SCN resistance in the US. One ISU graduate student working on this NCSRP-funded project will present results from all three states participating in the field microplot studies of Objective 2 of this project at the conference. One UGA graduate student working this NCSRP-funded project will present results from mapping SCN virulence genes of Objective 1 of this project at the conference.
Between April 1 and September 30, 2022 Tylka gave 15 interviews with ag media personnel. In each interview, the loss of effectiveness of PI88788 SCN resistance was mentioned or discussed in detail, and the NCSRP-funded research also was mentioned when time allowed.
Objective 4: Organize tests of experimental lines developed by public breeders in the north central US states and Ontario. (Diers)
The Diers group sent seed to cooperators for the 2022 SCN Regional Test. This test includes 225 entries that range from MG 0-IV. The regional test cooperators grew the tests over the summer and are in the process of harvesting them. We forwarded data sheets to the cooperators and they will use them to send us their test data and we will analyze the data across environment and summarize this information in a report that will be sent out in December.
Objective 5: Diversify the genetic base of SCN resistance in soybean by developing and evaluating germplasm and varieties with new combinations of resistance genes in high-yielding backgrounds. (Diers, Scaboo)
The Scaboo group has now completed successful crossing attempts (3 backcrosses) using PI 90763 as a donor parent, and LD11-2170 and SA13-1385 as recurrent parents, for three major genes associated with resistance to virulent nematode populations (rhg1-a, rhg2, and Rhg4). For each crossing attempt, we have identified desirable F1 plants using marker assisted selection, and we have sped up the process by utilizing our winter nurseries in Hawaii and Puerto Rico for the last two years. During the summer of 2022, we grew over 100 plant rows derived from selected plants, and our first yield trials of this material will be in the summer of 2023. Additionally, we tested over 85 new and advanced SCN resistant lines in state, regional, and national yield trials during 2022. Harvest is currently over halfway competed, and analysis and advancements will commence during the next quarter.
The Diers group has continued to work to diversify SCN resistance away from PI 88788 based resistance in Midwestern adapted cultivars. To do this, we have continued to select for the major SCN resistance genes rhg1-a and Rhg4. During the past summer, we tested over 5000 plants to select these genes and selected over 950. These selected plants will be advanced to plant rows during next growing season. In addition, in advanced yield tests we evaluated 35 experimental lines that carried both rhg1-a and Rhg4 and 22 lines with two SCN resistance genes from G. soja.
View uploaded report 