Asalf B

Bjørnstad Å

Brown JKM

Chen XM

He ZH

Huerta-Espino J

Lillemo M

Singh RP

English

Sydney eScholarship

 1General disease resistance loci against biotrophic pathogens in wheat  Lillemo M1, Asalf B1, Singh RP2, Huerta-Espino J3, Chen XM4, He ZH4,5, Brown JKM6, Bjørnstad Å1 1 Dept. of Plant and Env. Sciences, Norwegian Univ. of Life Sciences, Ås, Norway, 2 CIMMYT, México, 3 Campo Experimental Valle de Mexico-INIFAP, Mexico, 4Institute of Crop Sciences/National Wheat Improvement Center, CAAS, Beijing, China, 5 CIMMYT China Office, c/o CAAS, Beijing, China, 6 Dept. of Disease and Stress Biology, John Innes Centre, Norwich, England.  INTRODUCTION There is increased interest in breeding wheat cultivars for partial and race non-specific resistance against diseases like leaf rust (LR), stripe rust (YR) and powdery mildew (PM) which are caused by biotrophic pathogens with a high evolutionary potential. The CIMMYT bread wheat line Saar was developed in Mexico with a high level of race non-specific resistance to LR and YR based on the adult plant rust resistance loci Lr34/Yr18 and Lr46/Yr29 (Navabi et al. 2003, 2004). International testing showed that it also exhibits good partial resistance to PM, and a genetic study indicated that the resistance was governed by at least three genes (Lillemo et al. 2006). The PM resistance of Saar was furthermore shown to be associated with Leaf Tip Necrosis (LTN), which indicated that the resistance to LR, YR and PM in Saar could be under common genetic control (Lillemo et al. 2007). The objectives of the research presented in this paper were: (i) to elucidate the molecular genetic control of LR, YR and PM resistance in Saar through QTL mapping, and (ii) to characterize the PM resistance associated with Lr34/Yr18 and Lr46/Yr29 through detached leaf-tests with near-isogenic lines (NILs).  MATERIALS AND METHODS A population of 113 RILs from a cross between Avocet and Saar was evaluated for PM resistance over two years at two locations in Norway (Hamar and Ås in 2005 and 2006) and once in Beijing, China, in 2005. LTN and LR were evaluated in Cd. Obregon, Mexico in the 2004-05 cropping season and YR in Toluca, Mexico in 2005. The population was genotyped with SSR markers selected from a bulked-segregant analysis with the five most resistant and susceptible entries based on the PM data, and supplemented with DArT genotyping. Simple and composite interval mapping was conducted with PlabQTL v. 1.2.  NILs with Lr34/Yr18 and Lr46/Yr29 in the genetic background of Avocet and Lr34/Yr18 in the background of Lalbahadur were grown in a spore-free glasshouse compartment and used for sampling leaf segments from plants at different growth stages from the first seedling leaves to flag leaves of adult plants. Leaf segments of 3 cm were laid down in plastic boxes with benzimidazole agar and inoculated with fresh conidia of Blumeria graminis f.sp. tritici with the aid of a settling tower. Boxes with inoculated leaf segments were placed in a growth chamber at 15°C and 16 h photoperiod at low light intensity. The number of sporulating colonies on each leaf segment was counted 10 days after inoculation, and divided by the leaf area to calculate the infection efficiency (colonies/cm2). The sporulation rate (spores/colony) was assessed by tapping spores from individual leaf segments into 0.1% NaCl solution and counting the spore density in a Coulter Counter.  RESULTS   High correlation coefficients among the disease severity scores of the Avocet x Saar population (Table 1) indicated that part of the resistance to LR, YR and PM was under common genetic control, and a likely involvement of Lr34/Yr18 in the resistance to PM was indicated by a strong association with LTN (Table 1).   Table 1. Correlation coefficients among phenotypic traits in the Avocet x Saar population. All correlations were highly significant at the level α=0.0001.  PM China LR Mexico YR Mexico LTN Mexico PM Norway 0.697 0.738 0.762 -0.624 PM China   0.547 0.552 -0.575 LR Mexico   0.762 -0.849 YR Mexico    -0.640  A summary of the QTL mapping is shown in Table 2. Two QTLs were detected with resistance from Saar that had major effects on all three diseases. These correspond to the adult plant rust resistance loci Lr34/Yr18 and Lr46/Yr29 on 7DS and 1BL, respectively.  One further QTL with resistance from Saar was detected on 3AS that affected both PM and YR. Avocet contributed a major QTL for PM resistance on 4BL, while minor QTLs for PM resistance from Saar were detected on 5AL, 5BL and 7BL. More details about the QTL mapping in Avocet x Saar and information about closely linked molecular markers are given by Lillemo et al. (2008).   The PM resistance associated with the Lr34/Yr18 and Lr46/Yr29 loci have recently been named Pm38 and Pm39, respectively (Lillemo et al. 2008).   2 Leaf segments of first seedling leaves of the NILs showed a susceptible infection type to 18 differential isolates (data not shown), which indicated that the resistance genes Pm38 and Pm39 confer race non-specific resistance to PM in a similar manner as the rust resistance genes at the same loci.   A significant reduction in infection efficiency associated with the presence of Pm38 or Pm39 in the genetic background of Avocet was only detected in flag leaves while the effect of Pm38 was also significant in penultimate leaves (Table 3). No significant reduction in sporulation rate was detected at any growth stage, but there was a tendency toward less sporulation associated with Pm38 in penultimate leaves (data not shown). No reliable measurement of sporulation rate was possible on flag leaves due to the very low numbers of colonies.  In the more susceptible genetic background of Lalbahadur there was no reduction in infection efficiency (Table 4) or sporulation rate (Table 5) associated with Pm38 even in the flag leaves. On the contrary, the presence of Pm38 seemed to increase the infection efficiency (Table 4).  To determine whether the leaf age was important for the expression of Pm38 resistance in Lalbahadur, the infection efficiency was measured on flag leaves at three different growth stages. Although there was a tendency toward increased resistance in older flag leaves, there was no overall effect of Pm38 (Table 6).   LTN was expressed in some of the flag leaves and a different grouping of the data according to this parameter revealed a clear effect of Pm38 in the presence of LTN (Table 7).  Table 3. Infection efficiency (colonies/cm2) measured on detached leaves of Avocet NILs at different growth stages.  1st leaves3rd leaves 5th leaves F-1 leaves Flag leaves Avocet 11.0 10.2 2.34 2.43 0.66 Avocet+ Pm38 10.1 11.8 3.73 1.04 0.08 Avocet+ Pm39 12.6 10.8 1.40 1.82 0.06 Table 4.  Infection efficiency (colonies/cm2) measured on detached leaves of Lalbahadur NILs at different growth stages.  1st leaves 3rd leaves 5th leaves F-1 leaves Flag leaves Lalbahadur 31.8 21.0 17.4 15.0   9.1 Lalb+ Pm38 25.3 20.5 26.3 36.3 16.0 Table 5.  Sporulation rate (spores/colony) measured on detached leaves of Lalbahadur NILs at different growth stages.  1st leaves 3rd leaves 5th leaves F-1 leaves Flag leaves Lalbahadur 60.3 46.8 81.3 31.6 20.9Lalb+ Pm38 64.6 40.7 69.2 40.7 20.4 Table 6.  Infection efficiency (colonies/cm2) measured on flag leaves of Lalbahadur NILs at different growth stages.  GS 55 +5d +16d +23d Lalbahadur   6.39 1.25 1.08 0.95 Lalb+ Pm38 10.09 3.95 2.76 1.59 Table 2. Results of composite interval mapping of PM, LR and YR resistance and simple interval mapping of LTN in the Avocet x Saar population. The table shows R2 values obtained from multiple regression. Chrom. arm Marker interval Source1 PM Norway PM ChinaLR MexicoYR Mexico LTN  Mexico 1BL Xwmc719-Xhbe248 S 31.1 9.5 49.1 17.4 12.1 3AS Xstm844tcac-Xbarc310 S 19.5   8.7  4BL XwPt-1505 -Xgwm149 A 40.2 21.0    5AL Xgwm617b- Xwmc327 S 10.2     5BS Xbarc4 -Xgwm274b  S 9.0 9.7    6AL Xbarc3-XwPt-7063 A    14.4  7BL Xwmc581-XwPt-8007 S  4.9    7DS Xgwm1220-Xswm10 S 46.3 27.9 73.1 40.0 76.6 1BL x 7DS     43.1   % of the phenotypic variance explained 72.0 48.8 81.2 52.8  % of the genotypic variance explained 80.4 96.0 82.5 55.3  1 S = Saar, A = Avocet  3Table 7.  The effect of LTN on infection efficiency (colonies/cm2) measured on flag leaves of Lalbahadur NILs at different growth stages.  GS 55 +16d GS 55 +23d Lalbahadur 1.08 0.95 Lalb+ Pm38,  - LTN 6.13 4.86 Lalb+ Pm38, + LTN 0.51 0.31 It was observed in the detached leaf experiments with both Avocet and Lalbahadur NILs that lines with Pm38 and Pm39 had a more frequent occurrence of necrosis associated with PM colonies. Preliminary results from microscopy studies show that both genes are associated with a higher frequency of hypersensitive cell death at the seedling stage (data not shown). DISCUSSION The most significant finding of the QTL study was the major contribution to PM resistance of the LTN-associated rust resistance loci on 7DS and 1BL. The PM resistance of these loci was further confirmed in the detached leaf experiments with NILs, which also showed that the PM resistance of both loci is race non-specific and only expressed in adult plants. These are also common features of the LR and YR resistances of these loci and indicate that the underlying genes most likely work through physiological processes that affect all three diseases in a similar manner. Interestingly, the Lr34/Yr18/Pm38 locus is also associated with resistance to stem rust, but the expression of the resistance depends on the genetic background (Spielmeyer et al. 2008; Vanegas et al. 2008).   While the resistance of Pm38 and Pm39 was expressed in all flag leaves in the moderately susceptible genetic background of Avocet, the resistance of Pm38 could only be detected in the highly susceptible background of Lalbahadur when LTN was clearly expressed. This could be an indication of a direct physiological link between LTN and the disease resistance at this locus.   Further insight into the resistance mechanisms of Lr34/Yr18/Pm38 has come from a recent microarray study that found the resistance gene to be associated with an up-regulation of many genes commonly associated with abiotic stress responses in both rust inoculated and mock inoculated flag leaves (Hulbert et al. 2007). The up-regulation of most of these transcripts was much stronger in the leaf tips than the leaf bases, which corresponds well with the general disease resistance gradient commonly found in adult plant leaves of lines with Lr34/Yr18/Pm38.  The general disease resistance loci Lr34/Yr18/Pm38 and Lr46/Yr29/Pm39 are of major advantage to wheat breeding as they allow for simultaneous selection of race non-specific and potentially durable resistance against three of the most important biotrophic pathogens in wheat. Furthermore, the selection can be aided by closely linked molecular markers (Lillemo et al. 2008). REFERENCES Hulbert SH, Bai J, Fellers JP, Pacheco MG, Bowden RL (2007) Gene expression patterns in near isogenic lines for wheat rust resistance gene Lr34/Yr18. Phytopathology 97:1083-1093 Lillemo M, Asalf B, Singh RP, Huerta-Espino J, Chen XM, He ZH, Bjørnstad Å (2008) The adult plant rust resistance loci Lr34/Yr18 and Lr46/Yr29 are important determinants of partial resistance to powdery mildew in bread wheat line Saar. Theor Appl Genet 116:1155-1166 Lillemo M, Singh RP, Huerta-Espino J, Chen XM, He ZH, Brown JKM (2007) Leaf rust resistance gene Lr34 is involved in powdery mildew resistance of CIMMYT bread wheat line Saar. In: Buck HT, Nisi JE, Salomon N (eds) Developments in Plant Breeding Vol 12 Wheat productions in stressed environments Proceedings of the 7th International Wheat Conference Mar del Plata, Argentina Nov 27-Dec 2, 2005, pp 97-102 Lillemo M, Skinnes H, Singh RP, van Ginkel M (2006) Genetic analysis of partial resistance to powdery mildew in bread wheat line Saar. Plant Dis 90:225-228 Navabi A, Singh RP, Tewari JP, Briggs KG (2003) Genetic analysis of adult-plant resistance to leaf rust in five spring wheat genotypes. Plant Dis 87:1522-1529 Navabi A, Singh RP, Tewari JP, Briggs KG (2004) Inheritance of high levels of adult-plant resistance to stripe rust in five spring wheat genotypes. Crop Sci 44:1156-1162 Spielmeyer W, Singh RP, McFadden H, Wellings CR, Huerta-Espino J, Kong X, Appels R, Lagudah ES (2008) Fine scale genetic and physical mapping using interstitial deletion mutants of Lr34 /Yr18: a disease resistance locus effective against multiple pathogens in wheat. Theor Appl Genet 116:481-490 Vanegas CDG, Garvin DF, Kolmer JA (2008) Genetics of stem rust resistance in the spring wheat cultivar Thatcher and the enhancement of stem rust resistance by Lr34. Euphytica 159:391-401    

General disease resistance loci against biotrophic pathogens in wheat

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General disease resistance loci against biotrophic pathogens in wheat

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