Nodorum Pathosystem Research Articles

Genetics of resistance to septoria nodorum blotch in wheat.

Septoria nodorum blotch (SNB) is a foliar disease of wheat caused by the necrotrophic fungal pathogen Parastagonospora nodorum. Research over the last two decades has shown that the wheat-P. nodorum pathosystem mostly follows an inverse gene-for-gene model. The fungus produces necrotrophic effectors (NEs) that interact with specific host gene products encoded by dominant sensitivity (S) genes. When a compatible interaction occurs, a 'defense response' in the host leads to programmed cell death thereby provided dead/dying cells from which the pathogen, being a necrotroph, can acquire nutrients allowing it to grow and sporulate. To date, nine S gene-NE interactions have been characterized in this pathosystem. Five NE-encoding genes, SnTox1, SnTox3, SnToxA, SnTox5, and SnTox267, have been cloned along with three host S genes, Tsn1, Snn1, and Snn3-D1. Studies have shown that P. nodorum hijacks multiple and diverse host targets to cause disease. SNB resistance is often quantitative in nature because multiple compatible interactions usually occur concomitantly. NE gene expression plays a key role in disease severity, and the effect of each compatible interaction can vary depending on the other existing compatible interactions. Numerous SNB-resistance QTL have been identified in addition to the known S genes, and more research is needed to understand the nature of these resistance loci. Marker-assisted elimination of S genes through conventional breeding practices and disruption of S genes using gene editing techniques are both effective strategies for the development of SNB-resistant wheat cultivars, which will become necessary as the global demand for sustenance grows.

Genetics of Variable Disease Expression Conferred by Inverse Gene-For-Gene Interactions in the Wheat-Parastagonospora nodorum Pathosystem.

The wheat-Parastagonospora nodorum pathosystem involves the recognition of pathogen-secreted necrotrophic effectors (NEs) by corresponding wheat NE sensitivity genes. This inverse gene-for-gene recognition leads to necrotrophic effector-triggered susceptibility and ultimately septoria nodorum blotch disease. Here, we used multiple pathogen isolates to individually evaluate the effects of the host gene-NE interactions Tan spot necrosis1-Stagonospora nodorum ToxinA (Tsn1-SnToxA), Stagonospora nodorum necrosis1-Stagonospora nodorum Toxin1 (Snn1-SnTox1), and Stagonospora nodorum necrosis3-B genome homeolog1-Stagonospora nodorum Toxin3 (Snn3-B1-SnTox3), alone and in various combinations, to determine the relative importance of these interactions in causing disease. Genetic analysis of a recombinant inbred wheat population inoculated separately with three P. nodorum isolates, all of which produce all three NEs, indicated that the Tsn1-SnToxA and Snn3-B1-SnTox3 interactions contributed to disease caused by all four isolates, but their effects varied and ranged from epistatic to additive. The Snn1-SnTox1 interaction was associated with increased disease for one isolate, but for other isolates, there was evidence that this interaction inhibited the expression of other host gene-NE interactions. RNA sequencing analysis in planta showed that SnTox1 was differentially expressed between these three isolates after infection. Further analysis of NE gene-knockout isolates showed that the effect of some interactions could be masked or inhibited by other compatible interactions, and the regulation of this occurs at the level of NE gene transcription. Collectively, these results show that the inverse gene-for-gene interactions leading to necrotrophic effector-triggered susceptibility in the wheat-P. nodorum pathosystem vary in their effects depending on the genetic backgrounds of the pathogen and host, and interplay among the interactions is complex and intricately regulated.

The Wheat Snn7 Gene Confers Susceptibility on Recognition of the Parastagonospora nodorum Necrotrophic Effector SnTox7.

Parastagonospora (syn. ana, Stagonospora; teleo, Phaeosphaeria) nodorum (Berk.) Quaedvleig, Verkley & Crous is a necrotrophic fungal pathogen that causes the disease Septoria nodorum blotch (SNB) on wheat (Triticum aestivum L. subsp. aestivum). The fungus produces necrotrophic effectors (NEs) that cause cell death when recognized by corresponding host genes, which ultimately leads to disease. To date, eight host gene-NE interactions have been described in the wheat-P. nodorum system. Here, we report the identification and partial characterization of a ninth interaction involving a P. nodorum-produced NE designated SnTox7 and a wheat gene designated Snn7. SnTox7 is a small protein with an estimated size less than 30 kDa and largely resistant to heat and chemical treatment. The Snn7 gene governs sensitivity to SnTox7 and was delineated to a 2.7-cM interval on the long arm of wheat chromosome 2D. The Snn7-SnTox7 interaction explained 33% of the variation in disease among a segregating population, indicating that the interaction plays a prominent role in the development of SNB. The Snn7 sensitivity allele was identified in the hexaploid wheat cultivar Timstein, but evaluation of a set of 52 hexaploid lines of diverse origin indicated that few genotypes harbored a functional Snn7 allele, thus indicating that Snn7 is relatively rare. The identification of the Snn7-SnTox7 interaction adds to our knowledge of the wheat-P. nodorum pathosystem, which has become a model for necrotrophic specialist fungal pathogens and their interactions with plants leading to necrotrophic effector-triggered susceptibility.

Physiological specialization in the western Canadian population of Phaeosphaeria nodorum

Stagonospora nodorum blotch (SNB) caused by Phaeosphaeria nodorum is an important foliar disease of wheat. Efficient breeding for SNB resistance requires detailed knowledge of the pathosystem so that targeted improvements can be made. The objective of this study was to examine the wheat–P. nodorum pathosystem for evidence of host specificity amongst Canadian isolates. To this end, 49 P. nodorum isolates were inoculated on 16 wheat lines in indoor seedling inoculation experiments. Biologically significant host × pathogen interactions were identified, indicating that host specificity exists in the Canadian isolates tested. This observation is consistent with an inverse gene-for-gene pathosystem based upon host-selective toxins. Unweighted Pair Group Method with Arithmetic Mean (UPGMA) analysis of disease reaction data suggested nine virulence clusters; however, additional sub-groupings could be argued based upon the reaction of specific wheat lines to isolates within virulence clusters. The common wheat lines 86ISMN 2137, RL5407 and W7984 had good resistance to the P. nodorum isolates in seedling tests, which will be useful for improvements in common wheat and possibly durum wheat. The development of a SNB differential set is discussed. P. nodorum isolates characterized in this study will be useful for future studies in the wheat–P. nodorum pathosystem.

Whole-Genome QTL Analysis of Stagonospora nodorum Blotch Resistance and Validation of the SnTox4–Snn4 Interaction in Hexaploid Wheat

Necrotrophic effectors (also known as host-selective toxins) are important determinants of disease in the wheat-Stagonospora nodorum pathosystem. To date, five necrotrophic effector-host gene interactions have been identified in this system. Most of these interactions have additive effects while some are epistatic. The Snn4-SnTox4 interaction was originally identified in a recombinant-inbred population derived from a cross between the Swiss winter wheat cultivars 'Arina' and 'Forno' using the S. nodorum isolate Sn99CH 1A7a. Here, we used a recombinant-inbred population consisting of 121 lines developed from a cross between the hexaploid land race Salamouni and the hexaploid wheat 'Katepwa' (SK population). The SK population was used for the construction of linkage maps and quantitative trait loci (QTL) detection using the Swiss S. nodorum isolate Sn99CH 1A7a. The linkage maps developed in the SK population spanned 3,228 centimorgans (cM) and consisted of 441 simple-sequence repeats, 9 restriction fragment length polymorphisms, 29 expressed sequence tag sequence-tagged site markers, and 5 phenotypic markers. The average marker density was 6.7 cM/marker. Two QTL, designated QSnb.fcu-1A and QSnb.fcu-7A on chromosome arms 1AS and 7AS, respectively, were associated with disease caused by the S. nodorum isolate Sn99CH 1A7a. The effects of QSnb.fcu-1A were determined by the Snn4-SnTox4 interaction and accounted for 23.5% of the phenotypic variation in this population, whereas QSnb.fcu-7A accounted for 16.4% of the phenotypic variation for disease but was not associated with any known effector sensitivity locus. The effects of both QTL were largely additive and collectively accounted for 35.7% of the total phenotypic variation. The results of this research validate the effects of a compatible Snn4-SnTox4 interaction in a different genetic background, and it provides knowledge regarding genomic regions and molecular markers that can be used to improve Stagonospora nodorum blotch resistance in wheat germplasm.

Variable Expression of the Stagonospora nodorum Effector SnToxA Among Isolates Is Correlated with Levels of Disease in Wheat

Most research on host?pathogen interactions is focused on mechanisms of resistance, but less is known regarding mechanisms of susceptibility. The wheat?Stagonospora nodorum pathosystem involves pathogen-produced effectors, also known as host-selective toxins, that interact with corresponding dominant host genes to cause disease. Recognition of the S. nodorum effectors SnToxA and SnTox2 is mediated by the wheat genes Tsn1 and Snn2, respectively. Here, we inoculated a population of wheat recombinant inbred lines that segregates for Tsn1 and Snn2 with conidia from two S. nodorum isolates, Sn4 and Sn5, which both produce SnToxA and SnTox2 to compare the effects of compatible Tsn1?SnToxA and Snn2?SnTox2 interactions between the two isolates. Genetic analysis revealed that the two interactions contribute equally to disease caused by isolate Sn4 but the Tsn1?SnToxA interaction contributed substantially more to disease conferred by Sn5 than did the Snn2?SnTox2 interaction. Sequence analysis of the SnToxA locus from Sn4 and Sn5 indicated that they were 99.5% identical, with no polymorphisms in the coding region or the predicted promoters. Analysis of transcription levels showed that expression levels of SnToxA peaked at 26 h postinoculation for both isolates but SnToxA expression in Sn5 was more than twice that of Sn4. This work demonstrates that necrotrophic effectors of different isolates can be expressed at different levels in planta, and that higher levels of expression lead to increased levels of disease in the wheat?S. nodorum pathosystem.

Genetic analysis of disease susceptibility contributed by the compatible Tsn1–SnToxA and Snn1–SnTox1 interactions in the wheat-Stagonospora nodorum pathosystem

Stagonospora nodorum is a foliar pathogen of wheat that produces several host-selective toxins (HSTs) and causes the disease Stagonospora nodorum blotch (SNB). The wheat genes Snn1 and Tsn1 confer sensitivity to the HSTs SnTox1 and SnToxA, respectively. The objectives of this study were to dissect, quantify, and compare the effects of compatible Snn1-SnTox1 and Tsn1-SnToxA interactions on susceptibility in the wheat-S. nodorum pathosystem. Inoculation of a wheat doubled haploid population that segregates for both Snn1 and Tsn1 with an S. nodorum isolate that produces both SnTox1 and SnToxA indicated that both interactions were strongly associated with SNB susceptibility. The Snn1-SnTox1 and Tsn1-SnToxA interactions explained 22 and 28% of the variation in disease, respectively, and together they explained 48% indicating that their effects are largely additive. The Snn1-SnTox1 interaction accounted for 50% of the variation when the population was inoculated with an S. nodorum strain where the SnToxA gene had been mutated, eliminating the Tsn1-SnToxA interaction. These results support the theory that the wheat-S. nodorum pathosystem is largely based on multiple host-toxin interactions that follow an inverse gene-for-gene scenario at the host-toxin interface, but disease exhibits quantitative variation due to the mainly additive nature of compatible interactions. The elimination of either Snn1 or Tsn1 toxin sensitivity alleles resulted in decreased susceptibility, but the elimination of both interactions was required to obtain high levels of resistance. We propose the use of molecular markers to select against Snn1, Tsn1, and other toxin sensitivity alleles to develop wheat varieties with high levels of SNB resistance.

Identification and characterization of a novel host–toxin interaction in the wheat–Stagonospora nodorum pathosystem

Stagonospora nodorum, casual agent of Stagonospora nodorum blotch (SNB) of wheat, produces a number of host-selective toxins (HSTs) known to be important in disease. To date, four HSTs and corresponding host sensitivity genes have been reported, and all four host-toxin interactions are significant factors in the development of disease. Here, we describe the identification and partial characterization of a fifth S. nodorum produced HST designated SnTox4. The toxin, estimated to be 10-30 kDa in size, was found to be proteinaceous in nature. Sensitivity to SnTox4 is governed by a single dominant gene, designated Snn4, which mapped to the short arm of wheat chromosome 1A in a recombinant inbred (RI) population. The compatible Snn4-SnTox4 interaction is light dependent and results in a mottled necrotic reaction, which is different from the severe necrosis that results from other host-toxin interactions in the wheat-S. nodorum pathosystem. QTL analysis in a population of 200 RI lines derived from the Swiss winter wheat varieties Arina and Forno revealed a major QTL for SNB susceptibility that coincided with the Snn4 locus. This QTL, designated QSnb.fcu-1A, explained 41.0% of the variation in disease on leaves of seedlings indicating that a compatible Snn4-SnTox4 interaction plays a major role in the development of SNB in this population. Additional minor QTL detected on the short arms of chromosomes 2A and 3A accounted for 5.4 and 6.0% of the variation, respectively. The effects of the three QTL were largely additive, and together they explained 50% of the total phenotypic variation. These results provide further evidence that host-toxin interactions in the wheat-S. nodorum pathosystem follow an inverse gene-for-gene model.

Host-selective toxins produced by Stagonospora nodorum confer disease susceptibility in adult wheat plants under field conditions

Stagonospora nodorum, causal agent of Stagonospora nodorum blotch (SNB), is a destructive pathogen of wheat worldwide. As is true for many necrotrophic host-pathogen systems, the wheat-S. nodorum system is complex and resistance to SNB is usually quantitatively inherited. We recently showed that S. nodorum produces at least four proteinaceous host-selective toxins that interact with dominant host sensitivity/susceptibility gene products to induce SNB in seedlings. Here, we evaluated a population of wheat recombinant inbred lines that segregates for Tsn1, Snn2, and Snn3, which confer sensitivity to the toxins SnToxA, SnTox2, and SnTox3, respectively, to determine if compatible host-toxin interactions are associated with adult plant susceptibility to SNB foliar disease under field conditions. Artificial inoculation of the population in 2 years and two locations with a fungal isolate known to produce SnToxA and SnTox2 indicated that compatible SnToxA-Tsn1 and SnTox2-Snn2 interactions accounted for as much as 18 and 15% of the variation in disease severity on the flag leaf, respectively. As previously reported for seedlings, the effects of these two interactions in conferring adult plant susceptibility were largely additive. Additional adult plant resistance QTLs were identified on chromosomes 1B, 4B, and 5A, of which, the 1B and 5A QTLs were previously reported to be associated with seedling resistance to SNB. Therefore, in this population, some of the same QTLs are responsible for seedling and adult plant resistance/susceptibility. This is the first report showing that host-selective toxins confer susceptibility of adult plants to SNB, further substantiating the importance of compatible toxin-host interactions in the wheat-S. nodorum pathosystem.

Development, identification, and validation of markers for marker-assisted selection against the Stagonospora nodorum toxin sensitivity genes Tsn1 and Snn2 in wheat

The wheat-Stagonospora nodorum pathosystem involves a number of pathogen-produced host-selective toxins that interact with host genes in an inverse gene-for-gene manner to cause disease. The wheat intervarietal recombinant inbred population derived from BR34 and Grandin (BG population) segregates for the toxin sensitivity genes Tsn1, Snn2, and Snn3, which confer sensitivity to the toxins ToxA, SnTox2, and SnTox3, respectively. Here, we report the addition of 141 molecular markers to the BG population linkage maps, the identification and/or development of markers tightly linked to Tsn1 and Snn2, and the validation of the markers using a set of diverse wheat accessions. The BG population maps now contain 787 markers, and new simple sequence repeat (SSR) markers closely linked to Snn2 on chromosome arm 2DS were identified. In an effort to target more markers to the Snn2 locus, STS markers were developed from 2DS bin-mapped ESTs resulting in the development and mapping of 36 markers mostly to the short arms of group 2 chromosomes. Together, SSR and EST-STS markers delineated Snn2 to a 4.0 cM interval. SSRs developed in related work for Tsn1 were mapped in the BG population and delineated the gene to a 1.0 cM interval. Evaluation of the markers for Tsn1 and Snn2 in a diverse set of wheat genotypes validated their utility for marker-assisted selection, which is particularly efficient for removing toxin sensitivity alleles from elite germplasm and varieties.

Quantitative disease resistance assessment by real-time PCR using the Stagonospora nodorum-wheat pathosystem as a model

Measuring disease resistance accurately and reproducibly is a key requirement for the introgression of partial resistance genes into breeding lines. Here, a qPCR protocol is used to measure fungal biomass, using the wheat-Stagonospora nodorum pathosystem as a model. Seven cultivars of differing reported resistance levels were used. Fungal biomass taken at 220°C thermal days after inoculation accurately predicts the final grain weight loss. It is concluded that a test based on qPCR methods is specific, quantitative, rapid and objective. Such tests could provide useful and economic tools in the development of robustly resistant crop cultivars.

TheTsn1–ToxA interaction in the wheat–Stagonospora nodorum pathosystem parallels that of the wheat–tan spot system

The wheat tan spot fungus (Pyrenophora tritici-repentis) produces a well-characterized host-selective toxin (HST) known as Ptr ToxA, which induces necrosis in genotypes that harbor the Tsn1 gene on chromosome 5B. In previous work, we showed that the Stagonospora nodorum isolate Sn2000 produces at least 2 HSTs (SnTox1 and SnToxA). Sensitivity to SnTox1 is governed by the Snn1 gene on chromosome 1B in wheat. SnToxA is encoded by a gene with a high degree of similarity to the Ptr ToxA gene. Here, we evaluate toxin sensitivity and resistance to S. nodorum blotch (SNB) caused by Sn2000 in a recombinant inbred population that does not segregate for Snn1. Sensitivity to the Sn2000 toxin preparation cosegregated with sensitivity to Ptr ToxA at the Tsn1 locus. Tsn1-disrupted mutants were insensitive to both Ptr ToxA and SnToxA, suggesting that the 2 toxins are functionally similar, because they recognize the same locus in the host to induce necrosis. The locus harboring the tsn1 allele underlies a major quantitative trait locus (QTL) for resistance to SNB caused by Sn2000, and explains 62% of the phenotypic variation, indicating that the toxin is an important virulence factor for this fungus. The Tsn1 locus and several minor QTLs together explained 77% of the phenotypic variation. Therefore, the Tsn1-ToxA interaction in the wheat-S. nodorum pathosystem parallels that of the wheat-tan spot system, and the wheat Tsn1 gene serves as a major determinant for susceptibility to both SNB and tan spot.