Characterisation of major genes (Stb)-mediated resistance to Septoria tritici blotch disease in wheat

Lead Research Organisation: University of Nottingham
Department Name: Sch of Biosciences

Abstract

Wheat is one of the most important staple food crops providing 20% of total daily calories consumed by humans worldwide. Septoria tritici blotch (STB) is a devastating disease encountered in most wheat growing areas of the world and is the primary foliar disease of wheat in the UK responsible for year-on-year wheat losses of 5-10% despite fungicide treatments. The causal agent of STB is the fungus Zymoseptoria tritici. Emergence and spread of fungicide resistance in fungal populations seriously threatens wheat production and compromises food security, therefore resistance to STB is an important target in wheat breeding.
To date, 21 major (Stb) genes and several quantitative trait loci (QTL) that make smaller contributions to the resistance phenotype have been identified and mapped genetically. Only one of these genes has so far been cloned and mechanisms of resistance remain poorly understood. Owing to a lack of well-defined QTLs with additive effects and near absence of diagnostic markers, the current STB resistance breeding strategies rely mostly on phenotypic evaluation of breeding materials rather than targeted genotyping based selection. Consequently, over the past decades, there has been only a modest improvement of STB resistance in the commercial wheat germplasm and the majority of current AHDB recommended UK wheat cultivars show only moderate resistance to STB.
Published genetic maps at Stb loci are mostly low resolution and often involve early generation markers (eg RFLP, SSR), which are considered impractical in modern marker-assisted breeding. It is not known whether any of the Stb genes (apart from Stb6) may be present in the UK commercial wheat, or how effective these may be in controlling STB under UK conditions. The proposed PhD project, aims to fill this significant void in wheat breeding research by enhancing our understanding of genetics and mechanisms of STB resistance controlled by known Stb genes through addressing the following objectives:
1.Identify frequency of virulence/avirulence in the current UK field Z. tritici populations towards each Stb gene, and whether any of these genes confer broad-spectrum resistance
2.Assess whether resistance operates at seedling and/or adult plant stage
3.Fine-map the most interesting and useful resistance gene(s), and develop the diagnostic second generation genetic markers (KASP) for use in breeding
4.Gain insight into how the Stb gene(s) are conferring resistance
The Stb genes will be transferred by crossing from foreign and exotic genotypes into a highly susceptible UK wheat background. Development of mapping populations nearly-isogenic lines is already in progress. These materials will then be phenotyped with current UK field Z. tritici isolates under glasshouse conditions followed by high-density genotyping. We have recently cloned Stb6 and shown that it encodes a Wall-associated receptor-like kinase (WAK) protein that defines a novel class of disease resistance proteins (Nat Genet, accepted). Using the latest IWGSC assembly of wheat genome, we have annotated and manually curated all WAK encoding genes and hypothesised that at least some of these may correspond to Stb genes. Indeed, bioinformatics analyses have indicated that several Stb loci correspond to physical intervals containing one or more WAK genes. The student will further test the above hypothesis by assessing the function of candidate WAK genes using Virus-induced gene silencing (VIGS) approach well established in our laboratory (Plant Physiol 160: 582-90). Mechanisms resistance conferred by the Stb genes will be investigated using a combination of molecular biology and bioimaging approaches, in particular determining whether disease resistance is associated or not with cell death, what step in the fungal lifecycle is being targeted, and what defence pathways are activated.
The knowledge gained will aid breeding for STB resistance and therefore help to enhance crop productivity.

Publications

10 25 50
 
Description Have phenotyped interactions between wheat lines (carrying 20 Stb resistance genes in total) and over 100 recent (sampled 2015-2016) UK Z. tritici field isolates, and have identified wheat lines that could be used for crop improvement.

Have phenotyped and genotyped an F3 mapping population segregating for the Stb7 gene, and confirmed that several KASP markers can differentiate between the parental lines.

KASP markers for tracing 16 Stb genes in breeding materials have been designed as part of this project.
Exploitation Route KASP markers and Stb resistance data will be useful to wheat breeders for increasing the Z. tritici resistance in future wheat lines.
Sectors Agriculture, Food and Drink

 
Description British Society for Plant Pathology (BSPP) Travel Fund
Amount € 200 (EUR)
Organisation The British Society of Plant Pathology 
Sector Charity/Non Profit
Country United Kingdom
Start 03/2019 
End 03/2019
 
Title Fluorescent Z. tritici strains 
Description Produced Z. tritici strains expressing the fluorophores GFP and mCherry under the native Tub-2 promoter and terminators. 
Type Of Material Cell line 
Year Produced 2022 
Provided To Others? No  
Impact These tools were developed to enable the confocal examination of wheat leaves inoculated with both singular Z. tritici strains and co-inoculation of both virulent and avirulent strains. These experiments on wheat NIL lines containing Stb genes of interest will enable us to test for the use of stomatal closure as a resistance mechanism in this pathosystem. Preliminary experiments have been performed to identify viable methodologies for such testing. 
 
Title New KASP markers designed to follow Stb resistance genes 
Description KASP markers have been designed for Stb 2, 3, 4, 5, 8, 10, 11, 12, 13, 14, 16q, 18, 19 and Sm3 using data from RAGT Seeds and Rothamsted Research. Five markers have already been tested for Stb7 and segregate correctly with the parental lines. These markers will be useful for academics interested in cloning the respective disease resistance gene, and for breeders interested in transferring these genes into their elite wheat germplasm. 
Type Of Material Technology assay or reagent 
Year Produced 2020 
Provided To Others? No  
Impact Currently used to follow Stb genes through segregating populations in this award. 
 
Description Functional analysis of candidate Snn3 genes 
Organisation Curtin University
Country Australia 
Sector Academic/University 
PI Contribution We carried out functional analysis of candidate Snn3 genes using VIGS silenced lines and a bioassay.
Collaborator Contribution Partners provided sequence data on candidate genes identified through their own research for analysis.
Impact Data not yet sufficient to contribute to major outcomes for this award.
Start Year 2020
 
Description Functional analysis of candidate Snn3 genes 
Organisation National Institute of Agronomy and Botany (NIAB)
Country United Kingdom 
Sector Academic/University 
PI Contribution We carried out functional analysis of candidate Snn3 genes using VIGS silenced lines and a bioassay.
Collaborator Contribution Partners provided sequence data on candidate genes identified through their own research for analysis.
Impact Data not yet sufficient to contribute to major outcomes for this award.
Start Year 2020
 
Description Functional analysis of candidate Stb19 gene 
Organisation Wagga Wagga Agricultural Institute
Country Australia 
Sector Academic/University 
PI Contribution We worked with collaborators on cloning the Stb19 disease resistance gene by functionally analysing candidate genes using VIGS and a Z. tritici bioassay.
Collaborator Contribution Our collaborators identified candidate Stb19 genes and provided sequence data to us. This involved the analysis of complex sequencing data.
Impact This data is not yet sufficient to contribute full outcomes for this award. This collaboration is not multi-disciplinary.
Start Year 2019
 
Description Presenting stand at Hertfordshire County Show 
Form Of Engagement Activity Participation in an activity, workshop or similar
Part Of Official Scheme? No
Geographic Reach Regional
Primary Audience Public/other audiences
Results and Impact We presented a stand at the Hertfordshire County Show which drew in participants using games and displays including "GM Explored through DNA Strawberry Extractions", the "Millennia of Modification" display, "GM Myth-Buster Boards" and games based around concepts of precision vs. accuracy. This was used by presenters as an opening to discuss our own research and its implications with members of the public, including members of the farming industry attending the County Show who demonstrated a professional interest in this work and its implications for their fields.
Year(s) Of Engagement Activity 2019