Fusarium virguliforme (Akoi, O’Donnell, Homma & Lattanzi), causal agent of soybean (Glycine max L. Merr.) sudden death syndrome (SDS), first caused a significant disease loss in 1987 . F. virguliforme was not prevalent in Asia by 2011 but had spread quickly across the Americas from about 1980–2011. SDS has become a major pest problem for soybean growers and breeders in the Americas . The origins of the disease remain unclear but F. virguliforme may be a new pathogen of soybean since no complete resistance has been reported.
F. virguliforme, like many plant pathogenic Fusaria, were facultative hemi-biotrophic pathogens of plant roots with many host species [1, 3, 4]. However, only soybean among known hosts showed the leaf scorch when infected by F. virguliforme. Soybean cultivars showed a wide range of susceptibility to both leaf scorch and root rot suggesting cultivar-specific partial resistance existed F. virguliforme appeared to be a clonal pathogen [3, 6]. There were some variations in aggressiveness among field isolates and maintained strains but there were no races reported, by 2011.
Soybean resistance to SDS was multi-geneic and had two components; a partial resistance to root infection and rot caused directly at the site of infection by the fungus; and a partial resistance to leaf scorch caused indirectly by translocated fungal toxins [5, 7, 8]. The bases of resistance might include resistances to one or more toxins ; and both local and systemic resistances following pathogen recognition [10–12].
Heterodera glycines I., the soybean cyst nematode (SCN) was probably an ancient pest of soybean since complete resistances to some Hg Types of SCN was found in about 1% of pre-domesticated and early domesticated Plant Introductions (PIs) . Interestingly most of these PIs were also partially resistant to SDS . One locus, the Rfs2/Rhg1 locus on chromosome 18, was shown to underlie coinheritance of resistance to SDS in the roots and also reduce root infestation by SCN [10, 11, 14–17]. Fine map development did not resolve Rfs2 from Rhg1 suggesting the underlying gene(s) were either very closely linked or pleiotropic [11, 16].
Rfs2 underlay partial resistance to the spread of root infections by F. virguliforme [5, 7, 11]. The site of infection did not rot as rapidly when this allele was present and rates of root growth nearly equal to non-infested plants were maintained. Toxin translocation to leaves appeared reduced because leaf scorches did not develop or were less severe. The sudden plant death characteristic of SDS was manifested as both early senescence and an unusual abscission, basal to the leaflets instead of the petiole. Neither occurred if the Rfs2 allele was present.
Equally the Rhg1 locus underlay partial resistance to SCN [13, 18]. H.glycines, like many plant parasitic nematodes, were obligate endoparasites of plant roots. Like F. virguliforme
H. glycines has many alternate hosts. Over the past 50 years, the number of Hg Types (ex. races) has expanded from 4 in the 1960’s to 16–20 [19, 20]. However, the Rhg1 locus was constant, being required for partial resistance to all Hg Types in most PIs and cultivars. Full resistance to SCN required 1–4 loci in addition to Rhg1, the number depending on the nature of the cyst population parasitizing the roots [17, 21–25]. Genetic diversity was found among SCN isolates, even inbred cyst populations like PA3, Hg type 0 [13, 26]. Further, variation among the host plant roots response to SCN has been associated with temperature  such that environmental conditions must be rigorously controlled during assays [13, 16].
The resistance or susceptible interaction(s) between the nematode and soybean affected by Rhg1 was not induced until females stopped moving through the roots and established a feeding site comprising several giant cells [28–30]. Full resistance to SCN, based on the combined action of the genes at Rhg1 and one or more additional Rhg loci, was manifest as; cell wall appositions to surround the feeding site; failure to supply the feeding site a tracheary element; and a necrosis as the feeding site develops. If the resistance allele at Rhg1 was present normal rates of root growth were slightly depressed but the above ground stunting, yellowing and early senescence did not occur.
Inheritance of resistance to SCN was first reported in the PI ‘Peking’ . Three recessive loci (rhg1
rhg3) and a dominant locus (Rhg4) were assigned gene names by parsimony though other dominance models were equally likely. The Peking derived resistance alleles of rhg1 and Rhg4 were introgressed into the cv. ‘Forrest’ [31–33]. In crosses based on Forrest and with SCN isolate PA3 the rhg1-a was shown to be codominant with the susceptibility allele of ‘Essex’ (rhg1-e) and alone capable of providing partial resistance. Consequently, the Forrest, ‘Hartwig’, Peking and ‘PI 437654’ allele was renamed to Rhg1-a ( and hereafter) by the Soybean Genetics Committee.
The Rhg1 locus was located to a sub-telomeric region of the soybean chromosome 18 (molecular linkage group G; Lg G) by many studies [17, 21, 23–25]. All of these segregating populations that were later tested with F. virguliforme also had an Rfs2-like activity against SDS [8, 15, 34]. However, an Rhg1-like locus was found at other locations in a few SCN resistant PIs, including Lg B1 (chromosome 11) , mid LgG [35, 36] and Lg B2 (chromosome 14) [37, 38]. The effects of the Rhg1-like loci found at other locations than chromosome 18 on resistance to SDS were not reported by 2012.
It has been shown the resistance allele of Rhg1-a/Rfs2 (from Peking) was associated with reduced seed yield when SCN is not present in the fields [39–41]. That phenomenon might be related to delayed seedling development and stand formation . How the root reduction contributes to resistance may involve a locus on chromosome 7 (LG M) where Rzd, an interacting allele needed for resistance to zygote death, was strictly co-inherited in phase with Rhg1/Rfs2-a . Therefore, effects on development were predicted for the gene(s) underlying Rhg1-a/Rfs2-a [16, 43–47].
Fine scale genetic maps and BAC based genomic analysis identified a 42 kbp region encompassed in BAC B73p06 as the Rhg1/Rfs2 locus [16, 45, 48]. The region encoded an RLK (Glyma18g02680 named GmRLK18-1 hereafter), a variant laccase (Glyma18g02690) and a predicted Na/H ion antiporter (Glyma18g0270). The GmRLK18-1 and Gmlaccase18-1 were expressed in roots, shoots and flowers but not nodules or seeds. The antiporter was not expressed in any organ tested in Essex or Forrest. However, it might be expressed in flowers and seed at very low abundance . Many nucleotide differences were found in the region encompassing the GmRLK18-1 and Gmlaccase18-1 genes from fragments of sequences from the resistant allele in Forrest compared to susceptible genotypes ‘Asgrow 3244’ and ‘Williams 82’ [16, 29, 48, 50, 51]. However, the GmRLK18-1 was considered the most likely candidate gene underlying Rhg1-a based on fine maps and association analyses.
RLKs are part of the eukaryotic tumor necrosis factor beta receptor super-family [44, 52]. In plants they represent a major class of resistance genes and also a major class of developmental regulators. The GmRLK18-1 gene at Rhg1/Rfs2 had nine alleles recognized among PIs and cultivars [16, 44–48, 50]. Of those four alleles were associated with partial resistance to SCN and five with susceptibility. The second most important resistance allele was from PI88788 and named rhg1-b because it was recessive and discovered second . However, a small scale experiment with RNAi to the RLK (named GmRLK18-1-b) at the rhg1-b in transgenic hairy roots did not show a large effect on cyst numbers . Either dominance or incomplete inhibition of the protein might have occurred. Here, additional experiments were undertaken with the GmRLK18-1 at Rhg1-a in stable transformed soybean lines. Here, a molecular basis for resistance to SDS and SCN was inferred from functional analyses of the Rhg1/Rfs2-a locus in near isogenic lines (NILs) and the Forrest allele of the GmRLK18-1 gene in transgenic plants.