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Abstract
The soybean cyst nematode (SCN), Heterodera glycines, is a devastating plant parasite that causes considerable soybean yield loss by injecting effector proteins into the plant root cells to transform them into feeding cells. SCN is most effectively managed by planting resistant soybeans, but the repeated deployment of the same resistance sources has led to the emergence of virulent SCN capable of reproducing on resistant cultivars. In order to identify the genes controlling SCN virulence on resistant soybeans, two different next-generation sequencing-based population genomic approaches, a comparative transcriptome (RNA-seq) study and a pooled whole genome resequencing (Pool-seq) analysis, were implemented on SCN inbred populations adapted on susceptible and resistant hosts with or without resistance genes. RNA-seq on virulent and avirulent parasitic juveniles discovered 47 differentially expressed candidate virulence genes that were uniquely up- or down-regulated in the virulent SCN population adapted on the resistant host. These include genes encoding vitamin B-associated proteins (reduced folate carrier, biotin synthase [HgBioB], and thiamine transporter) and nematode effectors known to play roles in plant defense suppression, suggesting that virulent SCN may exert a heightened transcriptional response to cope with enhanced plant defenses and an altered nutritional status of a resistant soybean host. As a complementary approach, a Pool-seq strategy involving pools of virulent and avirulent virgin females was able to pinpoint the genomic regions demonstrating clear signatures of selection, from which 71 candidate virulence genes were identified. The proteins coded by these genes included nematode effectors potentially involved in host defense modulation, such as members of the CLAVATA3/ESR-related peptide (CLE), C-type lectin (C-LEC), chitinase (CHT), annexin (ANN), glutathione synthetase (GS), venom-allergen-like protein (VAP), and secreted SPRY (SP1a/RYanodine receptor)/Ran-binding protein (RanBP) domain-containing protein (SPRYSEC) families, and unknown predicted secreted proteins. Finally, three candidate genes (HgBioB, ANN, and GS) that contain at least two single nucleotide polymorphisms leading to amino acid changes were tested for their correlation to virulence. Sanger-sequencing of individual virgin females from multiple, unrelated SCN inbred populations with known virulence phenotypes revealed that GS SNPs strongly correlate to virulence, suggesting that this GS may play a significant role in virulence. Future functional studies of GS through RNA interference on (a)virulent nematodes will support or refute this hypothesis.