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1.
A single nucleotide polymorphism (snp) in the cytochrome b gene confers resistance to strobilurin fungicides in Erysiphe graminis DC fsp tritici Marchal. On the basis of this point mutation three different types of molecular markers have been developed. Cleaved amplified polymorphic sequences and allele-specific PCR were used to score resistant and sensitive isolates from specifically selected regional populations across Europe. The results of molecular tests were in total agreement with the resistance phenotypes revealed by in vivo tests. Serial dilutions of mixed samples (resistant/sensitive) delimited the detection for strobilurin-resistant alleles to a range of 10-50% for both marker classes. Due to these detection limits no mixture of mitochondria within individual isolates was found. Denaturing high performance chromatography was used to increase the detection sensitivity for the mutant allele. Although the detection limit was lowered to 5-10%, there was no evidence for the existence of mixed mitochondrial genotypes.  相似文献   

2.
Strobilurin-resistant isolates of Blumeria ( Erysiphe ) graminis f.sp. tritici , the cause of wheat powdery mildew, were more than 10-fold less sensitive to azoxystrobin than sensitive isolates. In all resistant isolates, a mutation resulting in the replacement of a glycine by an alanine residue at codon 143 (G143A) in the mitochondrial cytochrome b gene was found. Allele-specific primers were designed to detect this point mutation in infected wheat leaves. Using quantitative fluorescent allele-specific real-time polymerase chain reaction (PCR) measurements, strobilurin-resistant A143 alleles could be detected amongst strobilurin-sensitive G143 alleles at a frequency of at least 1 in 10 000, depending on the amount of target and nontarget DNA. Most isolates tested were dominant homoplasmic for either the A143 or G143 allele, although mixed populations of alleles could be detected in some isolates. In some of these isolates, strobilurin resistance was not always stable when they were maintained for many generations in the absence of selection. The allele-specific real-time PCR assay was also used to follow the dynamics of A143 alleles in field populations of B . graminis f.sp. tritici before and after application of fungicides. As expected, the A143 allele frequency only increased under selection pressure from a strobilurin fungicide. After three sprays of azoxystrobin, a pronounced selection for the strobilurin-resistant allele, with an increase in average frequency from 2·2 to 58%, was measured. The use of quantitative real-time PCR diagnostics for early detection of fungicide resistance genes at low frequency, coupled with risk evaluation, will be invaluable for further resistance risk assessment and validation of antiresistance strategies.  相似文献   

3.
BACKGROUND: Management of grapevine powdery mildew Erysiphe necator Schw. requires fungicide treatments such as sterol demethylation inhibitors (DMIs) or mitochondrial inhibitors (QoIs). Recently, reduction in the efficacy of DMIs or QoIs was reported in Europe and the United States. The aim of the present study was to develop real‐time qPCR tools to detect and quantify several CYP51 gene variants of E. necator: (i) A versus B groups (G37A) and (ii) sensitive versus resistant to sterol demethylase inhibitor fungicides (Y136F). RESULTS: The efficacy of the qPCR tools developed was better than the CAPS method, with a limit of 2 pg for E necator DNA, 0.06 ng for genetic group A and 1.4 ng for the DMI‐resistant allele. The detection limits of qPCR protocols (LOD) ranged from 0.72 to 0.85%, and the quantification limits (LOQ) ranged from 2.4 to 2.85% for the two alleles G47A and Y136F respectively. The application of qPCR to field isolates from French vineyards showed the presence of DMI‐resistant and/or QoI‐resistant alleles in French pathogen populations, linked to genetic group B. CONCLUSION: The real‐time PCR assay developed in this study provides a potentially useful tool for efficient quantification of different alleles of interest for fungicide monitoring and for population structure of E. necator. Copyright © 2010 Society of Chemical Industry  相似文献   

4.
ABSTRACT Spot blotch, caused by Cochliobolus sativus, is an important disease of barley in many production areas and is best controlled through the deployment of resistant cultivars. Information on the genetics of resistance in various sources can be useful in developing effective breeding strategies. Parents of the doubled haploid mapping population Calicuchima-sib/ Bowman-BC (C/B) exhibit a differential reaction to pathotypes 1 and 2 of C. sativus. To elucidate the genetics of spot blotch resistance in this population, C/B progeny were evaluated with both pathotypes at the seedling stage in the greenhouse and at the adult plant stage in the field. At the seedling stage, progeny segregated 84 resistant to 26 susceptible based on the qualitative analysis of infection response (IR) data to pathotype 1. This fit best to a 3:1 ratio, indicating that two genes were involved in conferring resistance. Quantitative analysis of the raw IR data to pathotype 1 revealed a single quantitative trait locus (QTL) on chromosome 4(4H) explaining 14% of the phenotypic variance. Adult plant resistance to pathotype 1 was conferred by QTL on chromosome 2(2H) and chromosome 3(3H), explaining 21 and 32% of the phenotypic variation, respectively. Bowman contributed the resistance alleles on chromosome 3(3H) and chromosome 4(4H), whereas Calicuchima-sib contributed the resistance allele on chromosome 2(2H). Resistance to pathotype 2 was conferred by a single gene (designated Rcs6) on chromosome 5(1H) based on qualitative analysis of data. Rcs6 was effective at both the seedling and adult plant stages and was contributed by Calicuchima-sib. This result was corroborated in the quantitative analysis of raw IR (seedling stage) and disease severity (adult plant stage) data as a single major effect (r(2) = 0.93 and 0.88, respectively) QTL was identified on chromosome 5(1H). Progeny with resistance to both pathotypes were identified in the C/B population and may be useful in programs breeding for spot blotch resistance.  相似文献   

5.
The mutation G143S has been associated with high-level strobilurin resistance in laboratory mutant strains of Cercospora beticola, one of the most destructive pathogens in sugar beet plants. By using allele specific primers (PASA-PCR) and agarose gel visualization, a molecular diagnostic was developed for the detection of the G143S resistance mutation. This assay is simple and applicable in low tech laboratory settings, with high reliability when a relatively large proportion of mutated mitochondrial alleles are present in the resistant strains. To achieve detection of resistant alleles at low frequencies, a more sensitive Real Time PCR based assay capable of discriminating resistant (S143) genotypes in frequencies as low as 1:10,000 resistant:sensitive alleles was developed. Both diagnostics were successfully validated in laboratory strains. Subsequently, a large number of C. beticola isolates from QoI-treated sugar beet experimental fields in Greece were screened for resistance to Qo fungicides using these diagnostics and classic bioassays. No proportion of the 143S resistant allele was detected in all field isolates tested, which was in agreement with the phenotypes revealed by the biotests confirming that the efficacy of QoIs against C. beticola has been sustained in Greece 7 years after their introduction.  相似文献   

6.
大麦种质对叶斑病的抗性鉴定与评价   总被引：1，自引：0，他引：1  
 由麦根腐平脐蠕胞菌引起的叶斑病在世界各大麦种植区均有发生,严重影响大麦的产量和品质。选育和应用抗性品种是防控该病害最有效的策略,然而可利用的抗源非常有限。在本研究中对中国233份具有代表性的大麦种质资源进行成株期抗叶斑病田间人工接种鉴定,发现只有垦啤麦5号等10份材料对3个供试菌株都表现抗病,仅占供试材料的4.3%。另外对37份国内外重要的叶斑病抗源材料进行苗期及成株期抗叶斑病鉴定,结果显示成株期抗叶斑病材料所占比例为41%~46%,苗期抗性材料所占比例为50%~64%,其中ND17293等11份材料在苗期和成株期对3个菌株均表现为抗病,可作为抗源继续加以利用;基于上述鉴定结果,进一步分析发现供试大麦苗期对三个菌株的抗病比例均高于成株期抗病比例,说明大麦在不同生育期对叶斑病的抗性存在较大差异。另外发现大麦对B. sorokiniana不同致病类型的抗性也存在明显的专化性。  相似文献   

7.
 麦根腐平脐蠕孢菌[Bipolaris sorokiniana(Sacc.)Shoemaker]引起的叶斑病是大麦上重要的病害叶部之一,在世界各大麦主要种植区均能造成严重的经济损失。本研究利用3个致病力不同的蠕孢叶斑病菌株Z 12010、Z 12014和Z 13010分别对来源于中国及国外引进的61份大麦种质资源进行苗期抗蠕孢菌叶斑病鉴定,田间接种Z 12014菌株进行成株期抗病性鉴定。在所鉴定的大麦种质中未发现在苗期或成株期对叶斑病免疫的材料。苗期抗叶斑病分析发现,品种之间对3个菌株的抗性存在明显差异,Varunda、Legacy、Tradition、09GW-01、09GW-07、08PJ-36、09PJ-39、垦啤麦7号、垦啤麦10号、驻大麦3号、S-4和中饲麦1号等12份材料对3个菌株表现为中抗至高抗反应型。7份材料成株期抗Z 12014菌株,其中Legacy、蒙啤麦3号、09PJ-39、垦啤麦9号、垦啤麦11号和龙啤麦3号等6份材料具有全生育期抗性特点,品种10-3苗期对菌株Z 12014感病,而成株期抗该菌株。  相似文献   

8.
 麦根腐平脐蠕孢菌[Bipolaris sorokiniana(Sacc.)Shoemaker]引起的叶斑病是大麦上重要的病害叶部之一,在世界各大麦主要种植区均能造成严重的经济损失。本研究利用3个致病力不同的蠕孢叶斑病菌株Z 12010、Z 12014和Z 13010分别对来源于中国及国外引进的61份大麦种质资源进行苗期抗蠕孢菌叶斑病鉴定,田间接种Z 12014菌株进行成株期抗病性鉴定。在所鉴定的大麦种质中未发现在苗期或成株期对叶斑病免疫的材料。苗期抗叶斑病分析发现,品种之间对3个菌株的抗性存在明显差异,Varunda、Legacy、Tradition、09GW-01、09GW-07、08PJ-36、09PJ-39、垦啤麦7号、垦啤麦10号、驻大麦3号、S-4和中饲麦1号等12份材料对3个菌株表现为中抗至高抗反应型。7份材料成株期抗Z 12014菌株,其中Legacy、蒙啤麦3号、09PJ-39、垦啤麦9号、垦啤麦11号和龙啤麦3号等6份材料具有全生育期抗性特点,品种10-3苗期对菌株Z 12014感病,而成株期抗该菌株。  相似文献   

9.
建立了土壤中草甘膦及其主要代谢物氨甲基膦酸高效液相色谱-串联质谱(HPLC-MS/M S)残留检测方法。样品经0.6 mol/L的氢氧化钾溶液提取、C18固相萃取柱过滤净化、9-芴基甲基三氯甲烷(FMOC-Cl)柱前衍生,以乙腈和5 mmol/L的乙酸铵为流动相,C18反相色谱梯度洗脱,电喷雾负离子多反应监测模式(MRM)HPLC-MS/MS检测。结果表明:草甘膦和氨甲基膦酸在1.6~200μg/L范围内线性关系良好,检出限(以信噪比S/N=3计)分别为0.80和0.94μg/kg,定量限(以S/N=10计)分别为2.6和3.0μg/kg;不同类型土壤样品中两种目标物3个水平的平均添加回收率在84%~104%之间,相对标准偏差(RSD)为2.8%~7.5%。应用本方法对部分供试土壤样品进行分析,结果发现所检样品中草甘膦和氨甲基膦酸的检出率较高,分别为38.9%和61.1%,两种物质土壤残留行为较为普遍。  相似文献   

10.
ABSTRACT Pyrenophora teres, the causal agent of net blotch of barley (Hordeum vulgare L.), induces a combination of necrosis and extensive chlorosis in susceptible barley cultivars. Cell-free filtrates from both net and spot forms of P. teres; P. teres f. sp. teres, and P. teres f. sp. maculata were found to contain phytotoxic low molecular weight compounds (LMWCs) and proteinaceous metabolites which appear to be responsible for different components of the symptoms induced by the two forms of the pathogen in a susceptible cultivar of barley (cv. Sloop). Proteins induced only brown necrotic spots or lesions similar to those induced by the pathogens 72 h after inoculation. In contrast, LMWCs induced general chlorosis seen 240 h after inoculation but not the localized necrosis. Neither hydrolyzed or heat- or protease-treated proteinaceous metabolites induced the symptoms. This is the first report of the involvement of proteins produced by P. teres in symptom development during net blotch disease of barley.  相似文献   

11.
赵森  黄健  王炯 《农药学学报》2020,22(4):685-692
建立了超高效液相色谱-串联质谱 (UPLC-MS/MS) 测定环境样品水基质以及人的血液、尿液和肝脏3种生物样品中矮壮素和野麦枯残留的方法。空白水、空白血、空白尿及空白肝脏样品中加入矮壮素和野麦枯标准溶液，经pH = 8的氨水稀释后，涡旋离心，上清液过混合型弱阳离子交换柱 (WCX) 进行富集提取后，采用UPLC-MS/MS检测。质谱检测采用正离子扫描，多反应离子监测模式 (MRM)。定性和定量均以矮壮素和野麦枯母离子和子离子进行分析。结果表明:试验期间矮壮素和野麦枯标准品在不同条件下的降解率为2.4%~11.9%，稳定性好。在0.010、0.10、0.25、0.50和1.0 mg/kg 5个添加水平下，矮壮素和野麦枯在空白水、空白血、空白尿及空白肝脏样品中的回收率在71%~104%之间，相对标准偏差在1.1%~19%之间，定量限均为0.01 mg/kg，检出限均为0.005 mg/kg。该方法灵敏度高，分析速度快，操作简便。  相似文献   

12.
The effects of aldicarb on populations of root lesion nematodes (primarily Pratylenchus penetrans ) and on grain yields of spring barley and wheat were examined in the field over 3 years, 1981*83. The incidence of barley net blotch ( Pyrenophora teres ), wheat leaf blotch ( Leptosphaeria nodorum ), and common root rot ( Cochliobolus sativus ) was also recorded in 1982 and 1983. Aldicarb treatments reduced the size of root lesion nematode populations in soil and roots in all years, except in the mid-season soil sample in 1983. The severity of leaf disease was decreased only in 1982, but the incidence of root rot was not significantly affected by the nematticide. Although aldicarb increased cereal grain yields by approximately 15% there was no significant relationship between numbers of root lesion nematodes in roots and soil and fungal disease symptoms on barley and wheat.  相似文献   

13.
ABSTRACT In search of new durable disease resistance traits in barley to control leaf spot blotch disease caused by the necrotrophic fungus Bipolaris sorokiniana (teleomorph: Cochliobolus sativus), we developed macroscopic and microscopic scales to judge spot blotch disease development on barley. Infection of barley was associated with cell wall penetration and accumulation of hydrogen peroxide. The latter appeared to take place in cell wall swellings under fungal penetration attempts as well as during cell death provoked by the necrotrophic pathogen. Additionally, we tested the influence of a compromised Mlo pathway that confers broad resistance against powdery mildew fungus (Blumeria graminis f. sp. hordei). Powdery mildew-resistant genotypes with mutations at the Mlo locus (mlo genotypes) showed a higher sensitivity to infiltration of toxic culture filtrate of Bipolaris sorokiniana as compared with wild-type barley. Mutants defective in Ror, a gene required for mlo-specified powdery mildew resistance, were also more sensitive to Bipolaris sorokiniana toxins than wild-type barley but showed less symptoms than mlo5 parents. Fungal culture filtrates induced an H2O2 burst in all mutants, whereas wild-type (Mlo) barley was less sensitive. The results support the hypothesis that the barley Mlo gene product functions as a suppresser of cell death. Therefore, a compromised Mlo pathway is effective for control of biotrophic powdery mildew fungus but not for necrotrophic Bipolaris sorokiniana. We discuss the problem of finding resistance traits that are effective against both biotrophic and necrotrophic pathogens with emphasis on the role of the anti-oxidative system of plant cells.  相似文献   

14.
ABSTRACT Leaf blotch, caused by Rhynchosporium secalis, was studied in a range of winter barley cultivars using a combination of traditional plant pathological techniques and newly developed multiplex and real-time polymerase chain reaction (PCR) assays. Using PCR, symptomless leaf blotch colonization was shown to occur throughout the growing season in the resistant winter barley cv. Leonie. The dynamics of colonization throughout the growing season were similar in both Leonie and Vertige, a susceptible cultivar. However, pathogen DNA levels were approximately 10-fold higher in the susceptible cultivar, which expressed symptoms throughout the growing season. Visual assessments and PCR also were used to determine levels of R. secalis colonization and infection in samples from a field experiment used to test a range of winter barley cultivars with different levels of leaf blotch resistance. The correlation between the PCR and visual assessment data was better at higher infection levels (R(2) = 0.81 for leaf samples with >0.3% disease). Although resistance ratings did not correlate well with levels of disease for all cultivars tested, low levels of infection were observed in the cultivar with the highest resistance rating and high levels of infection in the cultivar with the lowest resistance rating.  相似文献   

15.
Mating-type (MAT) locus-specific single nucleotide polymorphisms (SNPs) have been shown to be sufficient for conventional PCR-based differentiation of Pyrenophora teres f. teres (Ptt) and P. teres f. maculata (Ptm), the cause of the net and spot form, respectively, of barley net blotch. Here, we report the cloning and characterization of the MAT locus from 10 California isolates that cause atypical blotch symptoms on barley. Analysis of the full-length nucleotide sequences of one MAT1-1 (1,993?bp) and nine MAT1-2 (2,149 or 2,161?bp) idiomorphs revealed high (98?C99?%) similarity to those of Ptt isolates. However, distinct SNP patterns were identified in the newly cloned MAT idiomorphs. Two new MAT1-2-specific SNPs were found to be conserved in one Australia and eight California isolates that all cause similar atypical blotch symptoms. Phylogenetic analysis indicated that all 10 California isolates form a separate branch (or clade) within the Ptt group, except for one that appears to be ancestral to both Ptt and Ptm. PCR primers designed based on the identified SNP patterns were used successfully to differentiate each atypical isolate from highly virulent forms. This study extends our previous work and, taken together, the results demonstrate that the genetic variation at the MAT locus correlates with variation in the form-specific disease phenotype, and that MAT-specific SNPs can serve as reliable and convenient markers for subspecies level differentiation in P. teres, an economically important plant-pathogenic ascomycete (http://en.wikipedia.org/wiki/Single_nucleotide_polymorphism).  相似文献   

16.
Nga NT  Inoue Y  Chuma I  Hyon GS  Okada K  Vy TT  Kusaba M  Tosa Y 《Phytopathology》2012,102(7):674-682
Barley cultivars show various patterns of resistance against isolates of Magnaporthe oryzae and M. grisea. Genetic mechanisms of the resistance of five representative barley cultivars were examined using a highly susceptible barley cultivar, 'Nigrate', as a common parent of genetic crosses. The resistance of the five cultivars against Setaria, Oryza, Eleusine, and Triticum isolates of M. oryzae was all attributed to a single locus, designated as Rmo2. Nevertheless, the Rmo2 locus in each cultivar was effective against a different range of isolates. Genetic analyses of pathogenicity suggested that each cultivar carries an allele at the Rmo2 locus that recognizes a different range of avirulence genes. One allele, Rmo2.a, corresponded to PWT1, which conditioned the avirulence of Setaria and Oryza isolates on wheat, in a gene-for-gene manner. The other alleles, Rmo2.b, Rmo2.c, and Rmo2.d, corresponded to more than one avirulence gene. On the other hand, the resistance of those cultivars to another species, M. grisea, was conditioned by another locus, designated as Rmo3. These results suggest that Rmo2 is effective against a broad range of blast isolates but is specific to M. oryzae. Molecular mapping revealed that Rmo2 is located on the 7H chromosome.  相似文献   

17.
A Pyrenophora teres f. teres population in North Dakota was analyzed for virulence variation and genetic diversity using 75 monospore isolates that were collected across a 4-year period (2004 to 2007) from two North Dakota State University agricultural experiment stations at Fargo and Langdon. Pathogenicity tests by inoculation onto 22 barley differential lines at seedling stage revealed 49 pathotypes, indicating a wide range of pathogenic diversity. Two-way analysis of variance of disease ratings revealed a significant difference in the virulence among isolates and in the resistance among barley lines, as well as in the interactions between the two. 'CI5791', 'Algerian', and 'Heartland' were three barley lines showing a high level of seedling resistance to all North Dakota isolates tested; however, many previously reported resistance genes have been overcome. Forty multilocus genotypes were identified from this set of isolates by genotyping at 13 simple-sequence repeat loci. High percentages of clonal cultures were detected in the samplings from 2005 and 2007 in Fargo and 2005 in Langdon. Using a clone-corrected sample set, the mean gene diversity (h) was estimated to be 0.58, approximately the same for both locations. The calculated Wright's F(ST) value is small (0.11) but was significantly >0, indicating a significant differentiation between the Fargo and Langdon populations. In the gametic disequilibrium test, only 3 of 78 possible pairwise comparisons over all isolates showed significant (P < 0.05) nonrandom association, suggesting a random mating mode. Our results suggest that the populations from the two locations are derived from a common source and undergo frequent recombination. This research provides important information for barley breeders regarding development and deployment of cultivars with resistance to net form net blotch in this region.  相似文献   

18.
采用乙腈提取、分散SPE净化的前处理方法以及气相色谱电子捕获检测器,测定了苹果、柑橘和香蕉中腐霉利、抑霉唑、异菌脲和咪鲜胺4种防腐保鲜剂的残留量,利用基质匹配标准校正方法补偿基质效应。结果表明,腐霉利、抑霉唑和咪鲜胺在3个添加水平(0.02、0.05和0.10 mg/kg)下的回收率为85.1% ~108.4%,相对标准偏差为0.5% ~7.9%;异菌脲在3个添加水 平(0.05、0.10 和0.20 mg/kg)下的回收率为82.9% ~98.8%,相对标准偏差为3.6% ~ 8.6%。方法的检出限(LOD)在0.001 ~0.008 mg/kg之间,定量限(LOQ)在0.003 ~0.03 mg/kg之间。  相似文献   

19.
建立了QuEChERS-超高效液相色谱-串联质谱(QuEChERS-UPLC-MS/MS)对叶类蔬菜中极性较弱的氯吡脲、多效唑、烯效唑和丙环唑以及强极性的矮壮素共5种植物生长调节剂多残留的检测方法。样品经QuEChERS法预处理,用含体积分数为1%乙酸的乙腈溶液提取,石墨化碳黑(GCB)和N-丙基乙二胺(PSA)净化,C18色谱柱分离,采用电喷雾离子化正离子扫描和多反应监测模式(MRM)检测,基质匹配标准溶液外标法定量。结果表明:在1~1 000 μg/L范围内,氯吡脲、多效唑、烯效唑、丙环唑和矮壮素5种植物生长调节剂的质量浓度与对应的峰面积间呈良好的线性关系,相关系数均大于0.999 3,检出限(LOD)为0.002~0.04 μg/kg,定量限(LOQ)为0.01 mg/kg。在0.01、0.1和0.5 mg/kg添加水平下,5种植物生长调剂在菜心、芥蓝和普通白菜中的平均回收率为71%~93%,相对标准偏差(RSD)为0.2%~10%。该方法快捷、简便、定量准确,适用于叶类蔬菜中同时检测氯吡脲、多效唑、烯效唑、丙环唑和矮壮素等植物生长调节剂的残留。  相似文献   

20.
The effect of barley yellow dwarf virus (BYDV) on the development of net blotch (Pyrenophora eves) and leaf blotch (Rhynchosporium secalis) was examined on seven barley cuitivars. Seedlings were infected with BYDV at the two-leaf stage (G.S. 12). Their susceptibility to three isolates of p teres and isolates of two races (U.K. 1, U.K. 2) of R. secalis was examined at the four-leaf stsge (G.S. 14) and when plants were more mature (G.S. 33/38). At G.S. 14 numbers of lesions produced by P. teres and R. secalis were reduced, on average, by 37 and 72% respectively, and at G.S. 33/38 by 61 and 74%.  相似文献   

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