The SoyaGen Project: Putting Genomics to Work for Soybean Breeders.

The SoyaGen project was a collaborative endeavor involving Canadian soybean researchers and breeders from academia and the private sector as well as international collaborators. Its aims were to develop genomics-derived solutions to real-world challenges faced by breeders. Based on the needs expressed by the stakeholders, the research efforts were focused on maximizing realized yield through optimization of maturity and improved disease resistance. The main deliverables related to molecular breeding in soybean will be reviewed here. These include: (1) SNP datasets capturing the genetic diversity within cultivated soybean (both within a worldwide collection of > 1,000 soybean accessions and a subset of 102 short-season accessions (MG0 and earlier) directly relevant to this group); (2) SNP markers for selecting favorable alleles at key maturity genes as well as loci associated with increased resistance to key pathogens and pests (Phytophthora sojae, Heterodera glycines, Sclerotinia sclerotiorum); (3) diagnostic tools to facilitate the identification and mapping of specific pathotypes of P. sojae; and (4) a genomic prediction approach to identify the most promising combinations of parents. As a result of this fruitful collaboration, breeders have gained new tools and approaches to implement molecular, genomics-informed breeding strategies. We believe these tools and approaches are broadly applicable to soybean breeding efforts around the world.

RXLR effector gene Avr3a from Phytophthora sojae is recognized by Rps8 in soybean.

The use of resistance genes in elite soybean cultivars is one of the most widely used methods to manage Phytophthora sojae. This method relies on effector-triggered immunity, where a Resistant to P. sojae (Rps) gene product from the plant recognizes a specific effector from the pathogen, encoded by an avirulence (Avr) gene. Many Avr genes from P. sojae have been identified in the last decade, allowing a better exploitation of this type of resistance. The objective of the present study was to identify the Avr gene triggering immunity derived from the soybean resistance gene Rps8. The analysis of a segregating F2 progeny coupled with a genotyping-by-sequencing approach led to the identification of a putative Avr8 locus. The investigation of this locus using whole-genome sequencing data from 31 isolates of P. sojae identified Avr3a as the likely candidate for Avr8. Long-read sequencing also revealed that P. sojae isolates can carry up to five copies of the Avr3a gene, compared to the four previously reported. Haplotype and transcriptional analyses showed that amino acid changes and absence of Avr3a transcripts from P. sojae isolates caused changes in virulence towards Rps8. Functional analyses using CRISPR/Cas9 knockout and constitutive expression demonstrated that Rps8 interacted with Avr3a. We also showed that a specific allele of Avr3a is recognized by Rps3a but not Rps8. While Rps3a and Rps8 have been previously described as closely linked, this is the first report of a clear distinction hitherto undefined between these two resistance genes.

Discriminant haplotypes of avirulence genes of Phytophthora sojae lead to a molecular assay to predict phenotypes.

The soybean-Phytophthora sojae interaction operates on a gene-for-gene relationship, where the product of a resistance gene (Rps) in the host recognizes that of an avirulence gene (Avr) in the pathogen to generate an incompatible reaction. To exploit this form of resistance, one must match with precision the appropriate Rps gene with the corresponding Avr gene. Currently, this association is evaluated by phenotyping assays that are labour-intensive and often imprecise. To circumvent this limitation, we sought to develop a molecular assay that would reveal the avirulence allele of the seven main Avr genes (Avr1a, Avr1b, Avr1c, Avr1d, Avr1k, Avr3a, and Avr6) in order to diagnose with precision the pathotypes of P. sojae isolates. For this purpose, we analysed the genomic regions of these Avr genes in 31 recently sequenced isolates with different virulence profiles and identified discriminant mutations between avirulence and virulence alleles. Specific primers were designed to generate amplicons of a distinct size, and polymerase chain reaction conditions were optimized in a final assay of two parallel runs. When tested on the 31 isolates of known virulence, the assay accurately revealed all avirulence alleles. The test was further assessed and compared to a phenotyping assay on 25 isolates of unknown virulence. The two assays matched in 97% (170/175) of the interactions studied. Interestingly, the sole cases of discrepancy were obtained with Avr3a, which suggests a possible imperfect interaction with Rps3a. This molecular assay offers a powerful and reliable tool to exploit and study with greater precision soybean resistance against P. sojae.

Stable predictive markers for Phytophthora sojae avirulence genes that impair infection of soybean uncovered by whole genome sequencing of 31 isolates.

BACKGROUND: The interaction between oomycete plant pathogen Phytophthora sojae and soybean is characterized by the presence of avirulence (Avr) genes in P. sojae, which encode for effectors that trigger immune responses and resistance in soybean via corresponding resistance genes (Rps). A recent survey highlighted a rapid diversification of P. sojae Avr genes in soybean fields and the need to deploy new Rps genes. However, the full genetic diversity of P. sojae isolates remains complex and dynamic and is mostly characterized on the basis of phenotypic associations with differential soybean lines. RESULTS: We sequenced the genomes of 31 isolates of P. sojae, representing a large spectrum of the pathotypes found in soybean fields, and compared all the genetic variations associated with seven Avr genes (1a, 1b, 1c, 1d, 1k, 3a, 6) and how the derived haplotypes matched reported phenotypes in 217 interactions. We discovered new variants, copy number variations and some discrepancies with the virulence of previously described isolates with Avr genes, notably with Avr1b and Avr1c. In addition, genomic signatures revealed 11.5% potentially erroneous phenotypes. When these interactions were re-phenotyped, and the Avr genes re-sequenced over time and analyzed for expression, our results showed that genomic signatures alone accurately predicted 99.5% of the interactions. CONCLUSIONS: This comprehensive genomic analysis of seven Avr genes of P. sojae in a population of 31 isolates highlights that genomic signatures can be used as accurate predictors of phenotypes for compatibility with Rps genes in soybean. Our findings also show that spontaneous mutations, often speculated as a source of aberrant phenotypes, did not occur within the confines of our experiments and further suggest that epigenesis or gene silencing do not account alone for previous discordance between genotypes and phenotypes. Furthermore, on the basis of newly identified virulence patterns within Avr1c, our results offer an explanation why Rps1c has failed more rapidly in the field than the reported information on virulence pathotypes.

Effect of Silicon Absorption on Soybean Resistance to Phakopsora pachyrhizi in Different Cultivars.

Silicon (Si) is recognized for its prophylactic role in alleviating diseases when absorbed by plants and has been proposed as a possible solution against soybean rust, caused by Phakopsora pachyrhizi. However, little is known about its potential effects on soybean (Glycine max) because the plant's ability to absorb Si is poorly defined. In this work, our objectives were to evaluate and quantify the absorption of Si in leaves of different soybean cultivars and to determine if such absorption was able to enhance resistance to soybean rust. In a first set of experiments with cv. Williams 82, hydroponic plants were supplied or not with Si and inoculated with urediniospores of P. pachyrhizi. Chemical analyses revealed no significant differences in the plants' Si content regardless of the treatment, which translated into no effect on rust incidence. However, in a second set of experiments with different cultivars, plants of Korean cultivar Hikmok sorip absorbed nearly four times more Si than those of Williams 82. At the same time, plants from this cultivar exhibited a near absence of disease symptoms when supplied with Si. This resistance appeared to be the result of hypersensitive (HR) reactions that were triggered when plants were fed with Si. These results support the concept that a plant's innate ability to absorb Si will dictate the benefits conferred by a treatment with Si and provide evidence that Si can protect soybean plants against soybean rust through mediated resistance.