Objective 11 - Kim Hammond-Kosack (RRes)

Introgression of extreme resistance to Septoria tritici from the diploid wheat Triticum monococcum into hexaploid wheat (RRes)

Background

For the past decade, Septoria leaf blotch disease has been the No1 disease of UK wheat crops causing losses in excess of 30 million annually despite the best practice applications of fungicides. Wheat crops routinely receive 2-3 fungicide applications per annum to control foliar diseases. In 2002 Septoria tritici strains resistant to strobulurin fungicides emerged in the UK, and these are now widespread. As a consequence older triazole chemistries are now routinely applied to reduce disease levels. However, this practice has already led to the emergence of multiple drug resistant strains across the UK. Overall, the necrotic leaf lesion disease symptom cause a dramatic reduction to grain yield and quality as well as a large residual pool of unused nitrogen fertiliser in the soil. On the HGCA recommended list most of the cultivars listed only have a Septoria resistance rating of 4-6 which is inadequate. Also the increasing incidence of wild and wet winters favours this disease because the fungus is infective as soon as air temperatures exceed +6 C (Pietraville et al., 2003).
The potential introduction of the EU directive 91/414 to reduce the diversity of pesticides available will have long term implications to durable Septoria control. If the older triazoles which still remain effective against Septoria tritici were removed by directive 91/414 the ability to control this disease would be extremely difficult.
The diploid einkorn wheat Triticium monococcum was identified in WGIN 1 to be an excellent source of extreme resistance to Septoria (Jing et al., 2008). Genetic analyses revealed that resistance to multiple isolates was controlled by a single locus on chromosome 7A which has been designated TmStb1. So far, the closest markers (SSR and DArT) identified flank the locus at 1.7 and 4.9 cM. F1 plants generated between hexaploid wheat and Tm have been shown to display good resistant to Septoria, even though the Tm allele is present only in the heterozygous state.

Activities

1. To complete the introgression of the TmStb1 locus into different elite hexaploid wheats.
2. To test the efficacy of this novel resistance source under both glasshouse (single isolate tests) and field conditions (natural inoculum).
3. To provide closer linked markers to TmStb1 locus that can be used by the breeders in marker assisted selections.
4. To screen for other alleles of TmStb1 locus in T. monococcum germplasm using defined flanking molecular markers and compare their efficacy.
5. Introgression of Septoria resistance using hexaploid wheat-T. monococcum chromosomal substitution lines

Workplan

1. Backcross of hexaploid wheat and T. monococcum F1 plants.
Through WGIN1 support, several F1 progeny has been generated by crossing two T. monococcum accessions with three hexaploid wheat varieties. The F1 plants are in the process of being backcrossing with corresponding hexaploid wheat genotypes. Genomic DNA samples will be extracted from F1 and BCF1 plant and genotyped using representative SSR markers to validate the present of T. monococcum chromosomes, particularly the 7Am region spanning the TmStb1 locus. The resultant BCF1 will be allowed to self and the F2 progeny genotyped to identify BCF1:2 recombinants containing the TmStb1 region. The recombinants will allow for seed bulk production. The activity to generate BCF1:2 population will be finished by the end of the first year. The introgression will then continue to more advanced backcrosses during the remaining of the project.

2. Phenotyping of recombinants containing the TmStb1 region for resistance to Septoria tritici under glasshouse and field conditions.
For glasshouse assay, the previously established young seedling attached-leaf assay will be adopted (Jing et al., 2008). The Mycosphaerella graminicola isolate IPO323 will be infiltrated into the leaves for resistance/susceptibility screen. For field studies, standard replicated plot trials will be used to examine the resistance of the introgression lines against UK field isolates. This activity will be carried out from the 2nd year onwards.

3. Development of molecular markers closely linked to TmStb1 for breeding selection.
The current markers linked to TmStb1 may not provide high enough resolution for accurate marker assistant selection. To provide useful markers, DArT marker technology will be further exploited. Two interrelated activities are planned. First of all, a second F2 population will be used to define DArT markers more closely linked to TmStb1. Secondly, the DArT marker clones will be ordered from Triticarte and clone-end sequencing performed. Subsequently, the DArT markers will be converted into PCR-based markers, for use in the back-crossing programme and provided to breeders. This activity will be carried out in the first two years.

4. To screen for other alleles of TmStb1 locus in T. monococcum germplasm using defined flanking molecular markers and compare their efficacy.
Considering the existence of large genetic diversity in T. monococcum (Jing et al., 2007), it is desirable to explore the allelic variation of the TmStb1 locus and correlate the variation with the efficacy of resistance to Septoria tritici. Once closely linked markers from Activity 3 are generated, the markers will be used to screen about 300 T. monococcum accessions in the RRes collection assembled during WGIN1 for variation in TmStb1 region. Accessions with variant alleles will then be screened for resistance to differential isolates of M. graminicola. This activity will start in the third year.

5. Introgression of Septoria resistance using hexaploid wheat-T. monococcum chromosomal substitution lines
Over the past decades, many useful amphiploids of wheat and its wild relatives have been generated for various agronomical traits. These resources may prove to be invaluable for the identification of additional sources of novel resistance to Septoria tritici. As a start, we will test the wheat T. monococcum chromosomal substitution lines. For this, Chinese spring chromosome 1A, 3A and 5A substituted by T. monococcum 1Am, 3Am and 5Am will be used. Both glasshouse and field experiments will be carried out. If the resistance to UK field M. graminicola populations is confirmed, these lines will be crossed to elite UK wheat varieties. Considering the hexaploid wheat nature of these substitution lines, if additional novel Septoria resistance loci exit in this germplasm then this source could be easily and rapidly used for trait introgression. However, ph1 mutants will be used to increase recombination rates and to allow for refining the chromosomal segments responsible for the resistance. Molecular markers combined with cytogenetic tools such as FISH and GISH will be used to identify the physical locations of the resistance loci. This activity will be carried out during the whole project (i.e. years 1 to 5).

6. Archiving and storage of genetic materials
All information on the seed stocks generated during this introgression project will be entered into the JICs Genetic Resource Unit database, even though the seed will remain at Rothamsted Research.