Diagnostic value of molecular markers for <Emphasis Type="Italic">Lr</Emphasis> genes and characterization of leaf rust resistance of German winter wheat cultivars with regard to the stability of vertical resistance

Breeding for resistance is an efficient strategy to manage wheat leaf rust caused by Puccinia triticina f. sp. tritici. However, a prerequisite for the directed use of Lr genes in breeding and the detection of new races virulent to these Lr genes is a detailed knowledge on Lr genes present in wheat cultivars. Therefore, respective molecular markers for 18 Lr genes were tested for specificity and used to determine Lr genes in 115 wheat cultivars. Results obtained were compared to available pedigree data. Using respective molecular markers, genes Lr1, Lr10, Lr26, Lr34 and Lr37 were detected, but data were not always in accordance with pedigree data. However, leaf rust scoring data of field trials confirmed the reliability of DNA markers. These reliable marker data facilitated the analyses of the development of virulent leaf rust races from 2002 to 2009 based on released cultivars. A sudden change from low infection rates to susceptibility was observed for Lr1, Lr3, Lr10, Lr13, Lr14, Lr16, Lr26 and Lr37 since 2006. Cultivars carrying several leaf rust resistance genes showed no significant shift to susceptibility except one cultivar which revealed an increasing infection rate at a low level. In summary, it turned out that pedigree data are often not reliable and a detection of Lr genes by diagnostic markers is fundamental to combine Lr genes in cultivars for a durable resistance against leaf rust, and to conduct reliable surveys based on released cultivars, instead of ‘Thatcher’ NILs.     

Virulence of Puccinia triticina in Turkey and leaf rust resistance in Turkish wheat cultivars

Leaf rust caused by Puccinia triticina is a common disease on wheat in the coastal regions of Turkey. Collections of P. triticina from infected wheat leaves were obtained from the main wheat production zones of Turkey in 2009 and 2010. A total of 104 single uredinial isolates were tested for virulence on 20 lines of Thatcher wheat that differ for single leaf rust resistance genes. Forty-four different virulence phenotypes were identified over both years. Four phenotypes were found in both years. Phenotype FHPTQ found in 2009, with virulence to genes Lr2c, Lr3, Lr16, Lr26, Lr3ka, Lr17a, Lr30, LrB, Lr10, Lr14a, Lr18, Lr3bg, and Lr14b, was the most common phenotype at 15.4 % of the total isolates. Forty-three winter and spring wheat cultivars from Turkey were tested as seedlings with 13 different P. triticina virulence phenotypes from Canada, the US and Turkey. The infection types on the cultivars were compared with infection types on the Thatcher near isogenic lines to postulate the presence of seedling leaf rust resistance genes in the cultivars. Resistance genes Lr1, Lr3a, Lr10, Lr14a, Lr17a, Lr20, Lr23, and Lr26 were postulated to be present in the Turkish wheat cultivars. DNA of the wheat cultivars was tested with PCR markers to determine the presence of the adult plant resistance genes Lr34 and Lr37. Marker data indicated the presence of Lr34 in 20 cultivars and Lr37 in three cultivars. Field plot evaluations of the wheat cultivars indicated that no single Lr gene conditioned highly effective leaf rust resistance. Resistant cultivars varied for combinations of seedling and adult plant resistance genes.     

Roles of peroxidases and NADPH oxidases in the oxidative response of wheat (Triticum aestivum) to brown rust (Puccinia triticina) infection

The goal of this work was to establish which enzymes – peroxidases or NADPH oxidases – play the most important role in the resistance‐related oxidative burst response of wheat to infection by brown rust (Puccinia triticina). The expression of four peroxidases and two NADPH oxidases was analysed in the susceptible wheat cv. Thatcher and isogenic lines with different Lr resistance genes after pathogen inoculation. Of the peroxidases, TaPrx118 and TaPrx112 were induced several times more strongly than TaPrx103 and TaPrx107. The induction of peroxidases was more pronounced than that of NADPH oxidases. The patterns of peroxidase expression clearly differentiated moderately resistant from highly resistant lines and corresponded to oxidative response profiles. The possible involvement of peroxidases or NADPH oxidases was verified with enzyme‐specific inhibitors. The oxidative burst in the susceptible cv. Thatcher and in the lines TcLr24, TcLr25, TcLr9 was peroxidase‐dependent, while the response in line TcLr26 was NADPH‐oxidase‐dependent. It is postulated that class III peroxidases play a leading role in the formation of reactive oxygen species molecules during the response of wheat to pathogen infection. The results suggest a high level of redundancy of some peroxidase genes induced in biotic stress. The role of both enzyme systems in wheat response/resistance to brown rust is discussed in relation to the oxidative response, the efficiency of resistance, and the presence and origin of particular Lr resistance genes.     

Low diversity and fast evolution in the population of Puccinia triticina causing durum wheat leaf rust in France from 1999 to 2009, as revealed by an adapted differential set

No internationally agreed differential set is available for characterization of virulences in populations of Puccinia triticina causing wheat leaf rust on durum wheat. In a first step, 73 potentially differential host genotypes were tested with 96 durum leaf rust isolates collected in France. A differential set, adapted to the local epidemiological context and useful for comparison with international studies was selected, including French commercial cultivars, Thatcher lines with Lr genes, and international cultivars. In the second step, a sample of 310 isolates collected in France from 1999 to 2009 was characterized on this set. Diversity was very low, as only five pathotypes were distinguished. Genotyping of a subset of 76 isolates according to 20 SSR markers confirmed this low diversity, with 73 isolates belonging to a single dominant genotype. Population was strongly shaped by cultivars, and the findings explain the successive breakdown of resistance sources deployed in French durum wheat cultivars. The gene Lr14a, suggested to be an efficient source of resistance in several European and American countries, was overcome by pathotypes frequent in France since 2000. Postulation of resistance genes in the commercial cultivars led to a proposed simplified version of the differential set. This study, providing new information about leaf rust resistance genes present in the French durum wheat germplasm, highlights the need to diversify sources of resistance to P. triticina in this germplasm. The results are also discussed in terms of relatedness and intercontinental migration of P. triticina on durum wheat.     

Diversity of virulence phenotypes among annual populations of wheat leaf rust in Israel from 1993 to 2008

Leaf rust, caused by the fungus Puccinia triticina, is the most common rust disease of wheat in wheat‐producing areas worldwide. The Israeli population of wheat leaf rust has been consistently monitored since 1993. A total of 840 single urediniospore isolates from Triticum aestivum (567), T. dicoccoides (119) and T. durum (154) were analysed during 1993–2008. The structure of the pathogen population has changed to a large extent since 1993. The annual populations of P. triticina were separated into two distinct groups: 1993–1999 and 2000–2008. Differentiation among the annual pathogen populations, as well as between the overall populations of the 1990s and 2000s, could be mainly attributed to the following forces: (i) migration of leaf rust urediniospores from neighbouring regions; and (ii) selection pressure of new yellow rust‐resistant wheat cultivars that have been introduced into Israel since 1997. Genetic multiplicity of wild emmer contributes to P. triticina variability in Israel. Leaf rust populations collected from common wheat, wild emmer and durum wheat differed. The population that originated from T. durum was rather stable during the years of the survey, whereas that from T. aestivum changed significantly from the 1990s to the 2000s. Diversity within the annual populations of P. triticina was highest in 1994 when many new pathotypes and associations between virulences were observed. Single‐step derivatives of the new pathotypes became dominant after 2000. Significant changes in virulence frequency to a number of Lr genes (e.g. Lr2a, Lr15, Lr17, Lr21, Lr26) were also registered in 2000–2008.     

春小麦品种沈免2063抗叶锈性遗传分析

为明确春小麦品种沈免2063所含抗叶锈病基因的对数、身份、显隐性和互作关系,以沈免2063为父本,分别与感病品种Thatcher及小麦抗叶锈病近等基因系Lr9、Lr19、Lr24、Lr25、Lr28、Lr42和Lr43的载体品系杂交,获得F1、F2和F3代群体后,分别在苗期和成株期进行抗病性测定。结果表明:沈免2063含有3对显性遗传且相互独立作用的抗叶锈病基因Lr9、Lr19和Lr25,在苗期,沈免2063对致病类型CBG/QQ的抗病性由Lr9和Lr25控制,对PHT/RP的抗病性由上述3对抗叶锈病基因控制;在成株期,沈免2063对优势致病类型PHT/RP和THT/TP等比混合菌种的抗病性由上述3对抗叶锈病基因控制。Lr9、Lr19和Lr25在育成品种中出现频率很低,目前尚很有效,但这3个基因均为典型的垂直抗病性基因,应进行基因布局、基因轮换等科学组配,才能延长其使用寿命。     

Suppression subtractive hybridization and microarray analysis reveal differentially expressed genes in the <Emphasis Type="Italic">Lr39/41</Emphasis>-mediated wheat resistance to <Emphasis Type="Italic">Puccinia triticina</Emphasis>

Wheat leaf rust caused by Puccinia triticina (Pt) is one of the most severe fungal diseases threatening the global wheat production. The use of leaf rust resistance (Lr) genes in wheat breeding programs is the major solution to solve this issue. Wheat isogenic line carrying the Lr39/41 gene has shown a moderate to high resistance to most of the Pt pathotypes detected in China. In the present study, a typical hypersensitive response (HR) was observed using microscopy in leaves of the Lr39/41 isogenic line inoculated with the avirulent Pt pathotype THTT from 48 h-post inoculation. Two Lr39/41 resistance-associated suppression subtractive hybridization (SSH) libraries with a total of 6000 clones were established. Microarray hybridizations were performed on all obtained SSH clones using RNAs extracted from leaves of the Pt-inoculated and non-inoculated Lr39/41 isogenic lines, and leaves of the Pt-inoculated and non-inoculated Thatcher susceptible lines. Differentially expressed clones were analyzed by significance analysis of microarrays (SAM), followed by further sequencing. A total of 36 Lr39/41-resistance-related differentially expressed genes (DEGs) were identified, many of which had been previously reported to be involved in the plant defense response. The expression levels of eight selected DEGs during different stages of the Lr39/41-mediated resistance were further quantified by a qRT-PCR assay. Several pathogenesis-related (PR) and HR-related genes seem to be crucial for the Lr39/41-mediated resistance. In general, a brief profile of DEGs associated with the Lr39/41-mediated wheat resistance to Pt was drafted.     

Comparative microscopic and molecular analysis of Thatcher near‐isogenic lines with wheat leaf rust resistance genes Lr2a,Lr3, LrB or Lr9 upon challenge with different Puccinia triticina races

Thatcher near‐isogenic lines (NILs) of wheat carrying resistance gene Lr2a, Lr3, LrB or Lr9 were inoculated with Puccinia triticina races of virulence phenotype BBBD, MBDS, SBDG and FBDJ. Puccinia triticina infection structures were analysed under the fluorescence microscope over a course of 14 days after inoculation (dai). The relative proportion of P. triticina and wheat genomic DNA in infected leaves was estimated with a semiquantitative multiplex PCR analysis using P. triticina‐ and wheat‐specific primers. The occurrence of a hypersensitive response (HR), cellular lignification and callose deposition in inoculated plants was investigated microscopically. In interactions producing highly resistant infection type (IT) ‘0;’, a maximum of two haustorial mother cells per infection site were produced, and there was no increase in the proportion of P.  triticina genomic DNA in infected leaves, indicating the absence of P. triticina growth. In comparison, sizes of P. triticina colonies increased gradually in interactions producing moderately resistant IT ‘1’ and ‘2’, with the highest proportion of P. triticina genomic DNA found in leaves sampled at 14 dai. In interactions producing susceptible IT ‘3–4’, the highest proportion of P. triticina genomic DNA was found in leaves sampled at 10 dai (45·5–51·5%). HR and cellular lignification were induced in interactions producing IT ‘0;’ and ‘1’ at 1 dai but they were not observed in interactions producing IT ‘2’ until 2 dai. No HR or cellular lignification were induced in interactions producing susceptible IT ‘3–4’. Furthermore, a strong deposition of callose was induced in Lr9 + BBBD and Lr9 + FBDJ (IT ‘0;’), whereas this defence response was not induced in resistant or susceptible interactions involving Lr2a, Lr3 or LrB, indicating that Lr9 mediated resistance was different from that conditioned by Lr2a, Lr3 or LrB.     

Genetics of leaf rust resistance in spring wheat cultivars alsen and norm

ABSTRACT Alsen is a recently released spring wheat cultivar that has been widely grown in the United States because it has resistance to Fusarium head blight and leaf rust caused by Puccinia triticina. Norm is a high yielding wheat cultivar that has been very resistant to leaf rust since it was released. Alsen and Norm were genetically examined to determine the number and identity of the leaf rust resistance genes present in both wheats. The two cultivars were crossed with leaf rust susceptible cv. Thatcher and F(1) plants were backcrossed to Thatcher. Eighty one and seventy three BCF(1) of Thatcher times; Alsen and Thatcher x Norm respectively, were selfed to obtain BCF(2) families. The BCF(2) families were tested as seedlings with different isolates of P. triticina that differed for virulence to specific leaf rust resistance genes. The BCF(2) families that lacked seedling resistance were also tested as adult plants in greenhouse tests and in a field rust nursery plot. Segregation of BCF(2) families indicated that Alsen had seedling genes Lr2a, Lr10, and Lr23 and adult plant genes Lr13 and Lr34. Norm was determined to have seedling genes Lr1, Lr10, Lr16, and Lr23 and adult plant genes Lr13 and Lr34. The characterization of Lr23 in the segregating populations was complicated by the presence of a suppressor gene in Thatcher and the high temperature sensitivity of resistance expression for this gene. The effective leaf rust resistance in Alsen is due to the interaction of Lr13 and Lr23, with Lr34; and the effective leaf rust resistance in Norm is due to the interaction of Lr13, Lr16, and Lr23, with Lr34.     

Is virulence phenotype evolution driven exclusively by Lr gene deployment in French Puccinia triticina populations?

Puccinia triticina is a highly damaging wheat pathogen. The efficacy of leaf rust control by genetic resistance is mitigated by the adaptive capacity of the pathogen, expressed as changes in its virulence combinations (pathotypes). An extensive P. triticina population survey has been carried out in France over the last 30 years, describing the evolutionary dynamics of this pathogen in response to cultivar deployment. We analysed the data set for the 2006–2016 period to determine the relationship between the Lr genes in the cultivars and virulence in the pathotypes. Rust populations were dominated by a small number of pathotypes, with variations in most of the virulence frequencies related to the corresponding Lr gene frequencies in the cultivated landscape. Furthermore, the emergence and spread of a new virulence matched the introduction and use of the corresponding Lr gene (Lr28), confirming that the deployment of qualitative resistance genes is an essential driver of evolution in P. triticina populations. However, principal component analysis (PCA) revealed that certain pathotype–cultivar associations cannot be explained solely by the distribution of Lr genes in the landscape. This conclusion is supported by the predominance of a few pathotypes on some cultivars, with the persistence of several other compatible pathotypes at low frequencies. Specific interactions are not, therefore, sufficient to explain the distribution of virulence in rust populations. The hypothesis that quantitative interactions between P. triticina populations and bread wheat cultivars—based on differences in aggressiveness—is also a driver of changes in pathotype frequencies deserves further investigation.     

Diversity of virulence phenotypes among annual populations of Puccinia triticina originating from common wheat in Israel during the period 2000–15

Leaf rust, caused by the fungus Puccinia triticina, is the most common rust disease of wheat in wheat-producing areas worldwide. The Israeli population of P. triticina has been consistently monitored since 1993. A total of 784 single urediniospore isolates from Triticum aestivum were analysed during 2000–15. The structure of the pathogen population has changed to a large extent since 2000. The annual populations of P. triticina were separated into two distinct groups, 2000–11 and 2012–15, while populations of 2000–5 and 2006–12 were differentiated to a lesser extent. The change in the population originating from T. aestivum during the period 2000–15 is less significant compared to changes in the 1990s described previously. Diversity within the annual populations of P. triticina was rather stable during the period studied. Three new pathotypes, characterized by virulences on Lr3ka and Lr30 genes, became dominant between 2012 and 2015, while all but one prevailing pathotypes in 2000–11 were avirulent on these two genes. Significant changes in virulence frequencies on a number of Lr genes (Lr2c, Lr3ka, Lr15, Lr21, Lr23, Lr26, Lr30) and pairwise associations of virulences (mainly with Lr2c and Lr26) were registered in 2012–15 or earlier. It is postulated that the composition and pathotype structure of the P. triticina population in Israel is determined by wind-disseminated urediniospores from neighbouring regions, where the migration of P. triticina from the eastern part of the Mediterranean Sea and from the Horn of Africa seem to have the greatest influence.     

Virulence Phenotypes of a Worldwide Collection of Puccinia triticina from Durum Wheat

ABSTRACT A total of 78 isolates of Puccinia triticina from durum wheat from Argentina, Chile, Ethiopia, France, Mexico, Spain and the United States and 10 representative isolates of P. triticina from common wheat from the United States were tested for virulence phenotypes on seedling plants of 35 near-isogenic lines of Thatcher wheat. Isolates with virulence on lines with leaf rust resistance genes Lr10, Lr14b, Lr20, Lr22a, Lr23, Lr33, Lr34, Lr41, and Lr44 represented the most frequent phenotype. Cluster analysis showed that P. triticina from durum wheat from South America, North America, and Europe had an average similarity in virulence of 90%, whereas isolates from Ethiopia were <70% similar to the other leaf rust isolates collected from durum wheat. Of the 11 isolates from Ethiopia, 7 were avirulent to Thatcher and all near-isogenic lines of Thatcher. The isolates from common wheat had an average similarity in virulence of 60% to all leaf rust isolates from durum wheat. P. triticina from durum wheat was avirulent to many Lr genes frequently found in common wheat. It is possible that P. triticina currently found on durum wheat worldwide had a single origin, and then spread to cultivated durum wheat in North America, South America, and Europe, whereas P. triticina from Ethiopia evolved on landraces of durum wheat genetically distinct from the cultivated durum lines grown in Europe and the Americas.     

冬小麦品种北京837抗叶锈病基因的染色体定位研究

 1990~1993年间,引用中国春全套单体系列和抗叶锈病小麦近等基因系(或单基因系)为材料,采用单体遗传分析和基因推导相结合的方法,对冬小麦品种北京837抗叶锈病基因进行染色体定位研究,明确其对生理小种叶中1号的抗性系由分别位于染色体1B和6B上的两个显性互补基因所控制。位于1B染色体上的基因可能是Lr26,位于6B上的可能是Lr3a,二者可抵抗我国小麦叶锈菌群体中的部分生理小种(或毒性基因组合)。     

Regional and temporal differentiation of virulence phenotypes of Puccinia triticina from common wheat in Russia during the period 2001–2018

Leaf rust, caused by the fungus Puccinia triticina, is one of the most damaging rust diseases of wheat in Russia. Populations of P. triticina were monitored in seven regions of Russia from 2001 to 2018, with a total of 5,191 single urediniospore isolates from bread wheat (Triticum aestivum) being analysed. Populations have changed significantly in all regions since 2012, after 2 years of drought (2010–2011). Regional collections of P. triticina were also significantly different between the two periods 2001–2009 and 2012–2018, with changes along two geographic gradients from West Siberia to the north-west and south-west (North Caucasia) of the European part of Russia. All tested isolates were avirulent to resistance gene Lr9 in 2001–2009 but, since 2010, virulence to Lr9 has occurred and annually increased in the Asian part of Russia (Ural and West Siberia) due to deployment of cultivars with the Lr9 gene. Virulence to Lr2a and Lr15 was considerably lower in Dagestan (6%–33%) and all European regions (35%–67%) than in Asian regions (84%–96%). During 2001–2009, virulence on Lr1 was also lower in Dagestan (33%) and the European regions (50%–77%) than in Asia (91%–96%); however, by 2012–2018, nearly all isolates were virulent on Lr1. Remarkable changes were observed in frequencies of P. triticina races defined by their virulence/avirulence to Lr1 and Lr2a genes. We postulate the P. triticina population in Dagestan is specific to that area and pathogen populations in European and Asian parts of Russia are distinct.     

Histochemical and chemical evidence for lignin accumulation during the expression of resistance to leaf rust fungi in wheat

Levels of individual phenolic acids were examined in primary leaves of wheat (Triticum aestivum) after inoculation with avirulent and virulent strains of the leaf rust fungus (Puccinia recondita f. sp. tritici) at stages when previous work had shown fungal and host cells to be affected by expression of the Lr20 or Lr28 alleles for resistance. The predominant phenolic acid, ferulic acid, as well as p-coumaric and syringic acids were detected in primary leaves in both unbound and bound forms. They were not detected in germinating urediniospores of either rust strain. Levels of unbound phenolic acids changed little in response to infection. In Lr28-bearing leaves inoculated with an avirulent strain, increased concentrations of bound phenolic acids and three other unidentified compounds were observed about 4 h after many single or small groups of cells had undergone hypersensitive collapse. In an Lr20-bearing cultivar, levels of bound phenolic acids fell in leaves inoculated with either a virulent or avirulent rust strain. Coniferyl aldehyde and coniferyl alcohol were not detected in healthy or inoculated leaves of either wheat cultivar. Attempts to affect expression of resistance by application of inhibitors of phenylalanine ammonia-lyase were not successful and both wheat cultivars remained resistant to avirulent rust strains. The bound phenolic acids which accumulate in cells undergoing a hypersensitive response may play a role in resistance of Lr28-bearing wheat to the leaf rust fungus.     

Diversity in Puccinia triticina detected on wheat from 2008 to 2010 and the impact of new races on South African wheat germplasm

Samples of wheat and triticale infected with leaf rust were collected from 2008 to 2010 in South Africa to identify Puccinia triticina races. Races were identified based on their virulence profile on standard differential lines. Eight races were identified from 362 isolates. The dominant races were 3SA133 (syn. PDRS) in 2008 (78 %) and 2009 (34 %), and 3SA145 (47 %) in 2010. Race 3SA145 (CCPS) identified in 2009 was a new race in South Africa with virulence for the adult plant resistance gene Lr37. Another new race, 3SA146 (MCDS), was identified in 2010. Race 3SA146 is also virulent for Lr37 but unlike 3SA145, it is virulent for Lr1 and Lr23 and avirulent for Lr3ka and Lr30. Microsatellite analysis showed that 3SA145 and 3SA146 shared 70 % genetic similarity with each other, but only 30 % similarity with other races in South Africa, suggesting that both represent foreign introductions. In seedling tests of 98 South African winter and spring cultivars and advanced breeding lines, 27 % were susceptible to 3SA145 and 3SA146 but resistant to 3SA133. In greenhouse studies of 59 spring wheat adult plants, 19 % of breeding lines and 46 % of cultivars were susceptible to 3SA145, whereas 29 % of the lines and 53 % of cultivars were susceptible to 3SA146. The cssfr6 gene-specific DNA marker confirmed the presence of Lr34 gene for leaf rust resistance in a homozygous condition in 28 wheat entries. Five entries were heterogeneous for Lr34. Several entries which were susceptible as seedlings to the new races carried Lr34. These lines are expected to show lower levels of leaf rust as adult plants. Results of these studies indicate a continued vulnerability of South African wheat cultivars to new races and emphasise the importance of regular rust monitoring and the need to incorporate genes for durable resistance.     

Histopathology of durable adult plant resistance to leaf rust in the Brazilian wheat variety Toropi

Leaf rust, caused by the fungus Puccinia triticina is a major disease of wheat (Triticum aestivum) worldwide. This disease is prevalent in southern South America where the environmental conditions and high genetic variability of P. triticina favour epidemics. The primary means of controlling pathogenic P. triticina races has been through using wheat varieties containing race-specific resistance genes. The defence mechanisms involved in durable race non-specific resistance to P. triticina are probably distinct from those involved in non-durable race-specific resistance. We investigated the histological components of resistance to P. triticina present in three wheat genotypes: the race non-specific resistant Brazilian variety Toropi; the race-specific resistant line RL6010 Lr9; and the susceptible Brazilian variety BRS 194. Plants of these three genotypes were inoculated with P. triticina race MFP and tissue samples excised from flag leaves at various times after inoculation to assess the number of infective structures, frequency of cell death and the accumulation of autofluorescent cells and hydrogen peroxide. The genotypes showed different resistance mechanisms active at different times during the infection process. Our results for Toropi indicate that there was a reduction in the extent of formation of stomatal appressoria and all subsequent structures. During attempted penetration we also observed the production of autofluorescent compounds and late cell death, but not peroxide formation. This non-specific resistance to P. triticina involves both pre-haustorial and post-haustorial mechanisms which may be responsible for maintaining the low disease severity observed in this variety even under high inoculum pressure.