东农415稻瘟病抗性遗传分析及基因定位

 粳稻品种东农415自育成以来一直以其早熟、抗病、高产特性而著称,在黑龙江省稻瘟病高发区种植20多年均表现高抗稻瘟病。本研究利用158个采集于黑龙江省不同稻区的稻瘟病菌株对东农415进行接种鉴定,结果表明东农415对黑龙江省稻瘟病菌株有很强的抗性,抗谱高达89.2%。以东农415与丽江新团黑谷(LTH)杂交衍生的F₁和F₂群体为遗传分析试验材料,通过接种鉴定,发现东农415对稻瘟病菌株F-10-11的抗性由一个显性基因控制。进一步采用分子标记结合隐性群体分离分析法,以对菌株F-10-11极端感病的99个F₂单株为作图群体,将东农415的抗病基因定位在第2染色体,距离基因两侧标记RM5300和RM213的遗传距离分别为7.6和3.0 cM,暂命名为Pi-dn(t)。将Pi-dn(t)位点映射到水稻参考基因组图谱上,在抗病位点基因组区段内发现3个编码基因Os02g56010、Os02g55540和Os02g56400具有抗病基因结构域,可作为Pi-dn(t)的候选基因。     

Mapping quantitative trait Loci for resistance to rice blast

Quantitative trait loci (QTLs) conferring resistance to rice blast, caused by Magnaporthe oryzae, have been under-explored. In the present study, composite interval mapping was used to identify the QTLs that condition resistance to the 6 out of the 12 common races (IB1, IB45, IB49, IB54, IC17, and ID1) of M. oryzae using a recombinant inbred line (RIL) population derived from a cross of the moderately susceptible japonica cultivar Lemont with the moderately resistant indica cultivar Jasmine 85. Disease reactions of 227 F(7) RILs were determined using a category scale of ratings from 0, representing the most resistant, to 5, representing the most susceptible. A total of nine QTLs responsive to different degrees of phenotypic variation ranging from 5.17 to 26.53% were mapped on chromosomes 3, 8, 9, 11, and 12: qBLAST3 at 1.9 centimorgans (cM) to simple sequence repeat (SSR) marker RM282 on chromosome 3 to IB45 accounting for 5.17%; qBLAST8.1 co-segregated with SSR marker RM1148 to IB49 accounting for 6.69%, qBLAST8.2 at 0.1 cM to SSR marker RM72 to IC17 on chromosome 8 accounting for 7.22%; qBLAST9.1 at 0.1 cM to SSR marker RM257 to IB54, qBLAST9.2 at 2.1 cM to SSR marker RM108, and qBLAST9.3 at 0.1 cM to SSR marker RM215 to IC17 on chromosome 9 accounting for 4.64, 7.62, and 4.49%; qBLAST11 at 2.2 cM to SSR marker RM244 to IB45 and IB54 on chromosome 11 accounting for 26.53 and 19.60%; qBLAST12.1 at 0.3 cM to SSR marker OSM89 to IB1 on chromosome 12 accounting for 5.44%; and qBLAST12.2 at 0.3 and 0.1 cM to SSR marker OSM89 to IB49 and ID1 on chromosome 12 accounting for 9.7 and 10.18% of phenotypic variation, respectively. This study demonstrates the usefulness of tagging blast QTLs using physiological races by composite interval mapping.     

云南野生稻和地方稻资源抗白叶枯病分析

为鉴定云南稻种质资源对水稻白叶枯病的抗性情况,于孕穗期采用剪叶接种方法,用水稻白叶枯病强致病型代表菌株BD8438、CN9404和X1接种云南野生稻和地方稻种质资源,以病斑长度大于6 cm为感病分界线,对其抗感表现型进行调查分析。结果显示,共鉴定出来源于云南省不同种植生态区的186份地方稻抗性材料和22个野生稻抗病居群。野生稻对水稻白叶枯病的抗病能力较地方稻强,其中疣粒野生稻的抗性最强,抗病等级为0~2;药用野生稻次之,抗病等级为1~2;普通野生稻相对较差,抗病等级为1~5。地方稻抗性资源来自于云南省各个传统水稻种植区,抗性1级的材料占17%,抗性2级的占2%,抗性3级的占81%;按照稻种质资源亚种类型、粘糯性和水旱性分类,各类型地方稻抗性材料所占比例分别为粳稻占61%、籼稻占39%;粘稻占66%、糯稻占34%;水稻占83%、陆稻占17%。从利用抗白叶枯病基因培育新品种的角度评价,这些抗性资源具有潜在的发掘利用价值。     

水稻稻瘟病抗性QTL的定位分析

 稻瘟病是危害水稻最严重的病害之一。以抗稻瘟病的云南省地方品种魔王谷（MWG）和感稻瘟病的湖北省审定品种鄂晚8号（EW8）为亲本材料,构建双单倍体分离群体（DH）。利用22个菌株对亲本材料MWG/EW8进行致病性鉴定,从中筛选到5个毒性不同的鉴别菌株用于考察DH群体的稻瘟病抗性,构建包含120对SSR标记的分子遗传连锁图进行数量性状位点（QTL）的分析,鉴定出3个抗性QTL,均位于第6染色体长臂RM541附近, 3个QTL对抗病表型的贡献率介于7.7%~15.2 %之间,3个QTL的抗病等位基因均源自亲本MWG。     

Haplotype diversity and molecular evolution of the rice <Emphasis Type="Italic">Pikm</Emphasis> locus for blast resistance

Pikm-specific rice blast resistance is conferred by a combination of two genes that have a nucleotide-binding site and a leucine-rich repeat (LRR), Pikm1-TS and Pikm2-TS. To study the evolution of these genes, we investigated the allele diversity of their LRR regions in 16 elite rice cultivars and 35 landraces. Both phylogenetic trees were characterized by a deep bifurcation that separated two major clades of alleles. A high level of polymorphism was detected exclusively between these clades and not within each clade. This two-clade structure commonly observed for the two genes suggests that these genes have evolved together through bidirectional differentiation.     

Genetic distribution of the avirulence gene AVRPiz-t in Thai rice blast isolates and their pathogenicity to the broad-spectrum resistant rice variety Toride 1

Rice blast disease, caused by the filamentous fungus Pyricularia oryzae, is one of the most destructive diseases in rice worldwide. Breeding of resistant rice cultivars remains a cost-effective and environment-friendly means for controlling blast disease, but the resistance tends to break down over time because of the pathogen's rapid adaptation. In this study, AVRPiz-t gene sequences of 46 rice blast isolates were evaluated using a Southern blot analysis. The AVRPiz-t gene was present in 24 of 46 (52.2%) rice blast isolates. The pathogenicity assay showed that all blast isolates were avirulent against Japanese rice cv. Toride 1, which carries several rice blast resistance genes including Piz-t, Pii, Pi37, and Pi-ta. Screening for the Piz-t gene in Thai rice germplasm revealed that less than 20% of rice varieties harbour the Piz-t gene. Therefore, the Toride 1 rice variety could serve as an effective donor of rice blast resistance to be used in rice breeding programmes in Thailand. This study provides evidence for co-evolution between the rice blast resistance gene Piz-t and the rice blast fungal avirulence gene AVRPiz-t. Understanding this relationship will facilitate the sustainable development of breeding for rice blast resistance in the future.     

Molecular mapping of the new blast resistance genes Pi47 and Pi48 in the durably resistant local rice cultivar Xiangzi 3150

The indica rice cultivar Xiangzi 3150 (XZ3150) confers a high level of resistance to 95% of the isolates of Magnaporthe oryzae (the agent of rice blast disease) collected in Hunan Province, China. To identify the resistance (R) gene(s) controlling the high level of resistance in this cultivar, we developed 286 F(9) recombinant inbred lines (RILs) from a cross between XZ3150 and the highly susceptible cultivar CO39. Inoculation of the RILs and an F(2) population from a cross between the two cultivars with the avirulent isolate 193-1-1 in the growth chamber indicated the presence of two dominant R genes in XZ3150. A linkage map with 134 polymorphic simple sequence repeat and single feature polymorphism markers was constructed with the genotype data of the 286 RILs. Composite interval mapping (CIM) using the results of 193-1-1 inoculation showed that two major R genes, designated Pi47 and Pi48, were located between RM206 and RM224 on chromosome 11, and between RM5364 and RM7102 on chromosome 12, respectively. Interestingly, the CIM analysis of the four resistant components of the RILs to the field blast population revealed that Pi47 and Pi48 were also the major genetic factors responsible for the field resistance in XZ3150. The DNA markers linked to the new R genes identified in this study should be useful for further fine mapping, gene cloning, and marker-aided breeding of blast-resistant rice cultivars.     

137份云南省地方稻抗白叶枯病多菌系多基因鉴定

为选育水稻抗性品种和发掘抗白叶枯病基因,以来自中国、日本和菲律宾的9株革兰氏阴性菌黄单胞菌水稻变种Xanthomonas oryzae pv. oryzae菌株为供试菌株评价137份云南省地方稻种质资源对白叶枯病菌的抗性,同时检测供试材料中含14个已克隆抗白叶枯病基因的情况。结果显示,供试材料中至少对1株黄单胞菌水稻变种菌株有抗性的材料有104份,抗1、2、3、4、5、6、7、8和9株黄单胞菌水稻变种菌株的材料分别有25、19、14、19、11、6、5、2和3份;其中白杂一号、珍白18、麻旱谷、老来红、晋糯、香谷、毫冷薅、齐头蚂蚱谷、九谷和来之不易10份材料抗谱较广。大部分抗性材料携带1～7个白叶枯病抗性基因或其同源基因,未检测到含有Xa7、xa13、Xa21和Xa45(t)基因或其同源基因的材料。供试材料携带的抗性基因数量与抗性水平均呈极显著相关。     

Pathogenic Variability of Pyricularia grisea from the High- and Mid-Elevation Zones of Bhutan

ABSTRACT Thirty isolates of P. griseacollected from rice during a blast epidemic in 1995 in the high (1,800 to 2,600 m) and middle (1,200 to 1,800 m) elevations of Bhutan and 80 isolates collected from one rice cultivar from two high- and two mid-elevation sites in 1996 were analyzed for virulence. Differential varieties were indica CO39, with five near-isogenic lines (NILs) for resistance genes in the genetic background of CO39, and japonica Lijiangxintuanheigu (LTH), with five NILs for LTH. Twelve selected Bhutanese landraces also were studied. In addition, 10 blast nurseries consisting of the NIL sets, important local landraces, and representatives of international differential groups were established in the 1996 and 1997 growing seasons in the mid- and high-elevation agroecological zones. The 110 isolates were differentiated into 53 pathotypes based on the 2 NIL sets. Thirteen isolates were avirulent on all of the NILs but were compatible with some landraces. Several isolates were able to attack one of the NILs of CO39 but not CO39. These results strongly suggest that both CO39 and LTH possess previously unidentified resistance. The landraces were not uniform in their reactions to the isolates. When a reaction index taking into account all individual plant reactions was used, isolates that had been assigned to the same pathotype could be further differentiated, indicating that the NIL sets could not completely discriminate virulences in Bhutanese P. grisea populations. In the trap nurseries, disease was always present in the middle elevations, but disease was very low during July 1996 in the high elevations and only present during August and September 1997. Almost all varietal groups were more frequently attacked in the middle than in the high elevations, indicating that the virulence spectrum is wider and the conduciveness of the environment is greater in the middle elevations. Landraces from the high elevations were most susceptible, followed by international differential groups 7 and 8. The results suggest that selection has yielded landraces with more complete and complex resistance in the more disease-conducive mid-elevation environment. At the same time, the pathogen population also possesses a wider virulence spectrum in that environment.     

Identification of blast resistance genes in 358 rice germplasms (<Emphasis Type="Italic">Oryza sativa</Emphasis> L.) using functional molecular markers

Rice blast, caused by the fungus Magnaporthe oryzae, is one of the most devastating diseases of rice (Oryza sativa L.), and neck blast is the most destructive phase of this disease. Although neck blast causes tremendous yield loss, little is known about the molecular mechanisms underlying the neck blast resistance. To address this issue, we collected 358 rice varieties from different ecotypes in China and assessed them for the neck blast resistance under natural conditions favoring disease development in Jining, Shandong Province. Our results showed that 124 (34.6%) and 234 (65.4%) varieties were resistant and susceptible to M. oryzae under natural field conditions, respectively. Among the 358 rice varieties that were screened for the presence of 13 major blast resistance (R) genes against M. oryzae by using functional markers, 259 varieties contained one to seven R genes. In addition, the relationship between the presence of R genes and the disease reactions was also investigated by integrative analysis of phenotyping and genotyping based on functional markers. Our results showed that the rice blast resistance gene Pi2 was significantly correlated with neck blast resistance. Furthermore, any of the 13 major blast R genes was absent from 32 rice varieties exhibited obvious neck blast resistance, which would be the potential materials for identifying novel neck blast R genes. Taken together, our findings provide insight into the distribution of the 13 major blast R genes in the tested Chinese rice germplasm resources, which will serve as a basis for developing rice blast resistant. Furthermore, 32 rice varieties exhibited neck blast resistance, but they did not harbor any of the 13 major blast R genes. In the future, these varieties may be used to identify novel neck blast R genes.     

Molecular and Pathotype Analysis of the Rice Blast Fungus in North Vietnam

Rice blast caused by Pyricularia oryzae is a devastating disease worldwide. In Vietnam, rice blast is especially severe in the Red River Delta in the North. The genetic diversity of 114 P. oryzae isolates collected from rice in 2001 in the Red River Delta and nine additional Vietnamese P. oryzae isolates was analysed using Amplified Fragment Length Polymorphism (AFLP). DNA similarity and cluster analysis based on 160 polymorphic AFLP markers showed twelve different AFLP genetic groups among the 123 field isolates. Isolates collected from japonica hosts clustered separately from indica host isolates with at least 60% dissimilarity with little evidence for gene flow between the two populations. In the 2001 population originating from indica hosts, three genetic groups were predominant and represented 99% of the isolates sampled. One predominant clonal lineage represented 59% of the 2001 indica host population and was found in eleven provinces in the Red River Delta of North Vietnam. Significant genotype flow could be demonstrated between the indica population south of Red river and the indica population north of Red river. There was significant linkage disequilibrium between the AFLP loci within the indica population, indicating that this is not a random mating population. Pathogenicity tests of 25 isolates selected from the 12 AFLP groups on a set of 29 differential rice lines revealed two avirulent isolates and 23 pathotypes. Different combinations of known resistance genes were found to have potential for blast resistance breeding for North Vietnam. First two authors contributed equally     

广西筛选稻瘟病抗源品种的初步研究

 1973~1979年间,在广西稻瘟病圃试验了11198个水稻品种。有一些抗病品种曾以人工接种测定.其中23个品种呈现稳定的高抗至中抗反应。
试验结果表明,红脚占、砦糖、金围矮是广谱抗稻瘟病的品种。
初步观测表明,叶瘟与穗瘟的抗病反应是相关的。
文中讨论了对稻瘟病抗病性的类型,未发现范德普兰克(1968年)描述的对病原的所有小种反应一致的水平抗性品种。     

水稻品种抗稻瘟病特性分类的初步研究

 选用7个稻瘟病菌稳定菌株,将588个水稻品种划分为66个品种类群,为今后开展品种抗病性的基因分析提供了必要的条件。通过测试,提出15个抗多个生理小种的广谱抗性品种,这些良好抗源可作杂交亲本利用。     

Pathotypic and genetic diversity in the population of Rhizoctonia solani AG1‐IA causing rice sheath blight in China

Sheath blight, caused by Rhizoctonia solani AG1‐IA, is one of the most serious diseases of rice. In this study, a total of 175 isolates of R. solani AG1‐IA were collected from five rice‐growing regions in China. Pathogenicity tests revealed that all isolates were virulent to five cultivars with different levels of resistance at the rice seedling stage in the greenhouse. There was considerable variation in aggressiveness, and the isolates were classified into three pathotypes based on disease severity, with moderately virulent isolates prevalent in the population. Forty‐three haplotypes were identified based on ITS sequencing, and 39 haplotypes were distinct among isolates. There were high levels of haplotype diversity and nucleotide diversity within the populations of R. solani AG1‐IA. High gene flow (Nm = 1·63–5·22) was detected, consistent with relatively low differentiation between pairs of populations. Five populations were divided into two distinct clusters by the unweighted pair group method with arithmetic mean (UPGMA), and no spatial population differentiation was discernible. The majority (97·8%) of genetic diversity was distributed among isolates within populations, with only 2·2% of the genetic diversity attributed to differences among populations. The star‐like shape of the haplotype network provided evidence of signatures of population expansion in recent history. No significant relationships were found between the genetic diversity and aggressiveness or geographic origin among populations of R. solani AG1‐IA. These results highlight that the population characteristics of R. solani AG1‐IA should be taken into account in evaluating the germplasm resistance of rice cultivars to sheath blight.     

吉林省主栽水稻品种抗瘟性评价及抗瘟基因型鉴定

 为明确吉林省主栽水稻品种的抗瘟性和抗瘟基因型,选用了一套已明确其无毒基因组成的稻瘟病菌标准菌株对吉林省68个主栽水稻品种进行喷雾和离体划伤接种试验。结果表明,68个品种的抗瘟能力存在较大差异,中抗以上抗病品种达到44个,占供试品种的64.7％;抗瘟基因推导结果结合特异性分子标记检测表明,Pita、Pia、Pish、Pita2、Pib和Pi9等6个基因为吉林省主栽水稻品种中主要的抗瘟基因,并且品种中含有的抗瘟基因数量与抗瘟能力呈正相关;根据结果推测,测试的所有品种中可能不含有Pi2、Pikh、Pikm、Pi11和Pi12基因。本研究揭示了吉林省近年主栽水稻品种的抗瘟能力和抗瘟基因组成,明确了吉林稻区主效抗瘟基因和基因聚合对稻瘟病的抗性贡献,为进一步培育广谱持久抗瘟品种和合理布局抗病品种提供了重要参考。     

Molecular mapping of the blast resistance genes Pi2-1 and Pi51(t) in the durably resistant rice 'Tianjingyeshengdao'

Tianjingyeshengdao' (TY) is a rice cultivar with durable resistance to populations of Magnaporthe oryzae (the causal agent of blast) in China. To understand the genetic basis of its resistance to blast, we developed a population of recombinant inbred lines from a cross between TY and the highly susceptible 'CO39' for gene mapping analysis. In total, 22 quantitative trait loci (QTLs) controlling rice blast resistance were identified on chromosomes 1, 3, 4, 5, 6, 9, 11, and 12 from the evaluation of four disease parameters in both greenhouse and blast nursery conditions. Among these QTLs, 19 were contributed by TY and three by CO39. Two QTL clusters on chromosome 6 and 12 were named Pi2-1 and Pi51(t), respectively. Pi2-1 was detected under both growth chamber and natural blast nursery conditions, and explained 31.24 to 59.73% of the phenotypic variation. Pi51(t) was only detected in the natural blast nursery and explained 3.67 to 10.37% of the phenotypic variation. Our results demonstrate that the durable resistance in TY is controlled by two major and seven minor genes. Identification of the markers linked to both Pi2-1 and Pi51(t) in this study should be useful for marker-aided selection in rice breeding programs as well as for molecular cloning of the identified resistance genes.     

Monitoring of Brazilian wheat blast field populations reveals resistance to QoI,DMI, and SDHI fungicides

Wheat blast is one of the most important and devastating fungal diseases of wheat in South America, South-east Asia, and now in southern Africa. The disease can reduce grain yield by up to 70% and is best controlled using integrated disease management strategies. The difficulty in disease management is compounded by the lack of durable host resistance and the ineffectiveness of fungicide sprays. New succinate dehydrogenase inhibitor (SDHI) fungicides were recently introduced for the management of wheat diseases. Brazilian field populations of the wheat blast pathogen Pyricularia oryzae Triticum lineage (PoTl) sampled from different geographical regions in 2012 and 2018 were shown to be resistant to both QoI (strobilurin) and DMI (azole) fungicides. The main objective of the current study was to determine the SDHI baseline sensitivity in these populations. Moderate levels of SDHI resistance were detected in five out of the six field populations sampled in 2012 and in most of the strains isolated in 2018. No association was found between target site mutations in the sdhB, sdhC, and sdhD genes and the levels of SDHI resistance, indicating that a pre-existing resistance mechanism not associated with target site mutations is probably present in Brazilian wheat blast populations.     

Races of <Emphasis Type="Italic">Magnaporthe oryzae</Emphasis> in Australia and genes with resistance to these races revealed through host resistance screening in monogenic lines of <Emphasis Type="Italic">Oryza sativa</Emphasis>

Rice blast is the most serious disease threat to rice production worldwide. It is difficult to control due to the complex diversity and wide geographic distribution of the causal pathogen Magnaporthe oryzae. In Australia, rice blast occurs in northern Australia but remains exotic to the main south-eastern rice growing area; however, there is the potential for rice blast to threaten this area; in addition, rice production is currently expanding from south-eastern Australia into northern Australia, which makes rice blast a major concern and challenge to rice industry in Australia. Prior to this study, there was lack of information on the race status of M. oryzae present in Australia and on how to manage the disease through host resistance. The races of rice blast isolates collected in northern Australia was characterised based on the disease reactions of eight standard rice differentials used in an international race differential system. The following studies revealed genes conferring resistance to these races through investigating the responses of 25 monogenic rice lines with targeted resistance gene against different races. The rice blast isolates were characterised into five races: IA-1, IA-3, IA-63, IB-3 and IB-59. Genes Pi40, Piz-t, Pi9, Pi5(t) and Pi12(t) exhibited resistance to all the isolates belonging to five races. In addition, two genes showed complete resistance to multiple races, viz. Pi9 that showed complete resistance to races IA-1, IA-3, IA-63 and IB-3 and Pita2 that had complete resistance to races IA-3, IB-3 and IB-59. This study provides information about the races of M. oryzae in Australia. Genes identified conferring resistance to multiple races will not only streamline the identification via molecular markers of imported rice varieties with resistance to rice blast in Australia, but will also allow the Australian rice breeding program to develop new varieties with broad-spectrum resistance to rice blast and pyramid multi-gene resistance into Australian rice varieties.     

禾谷丝核菌(Rhizoctonia cerealis)原生质体制备与再生的研究

禾谷丝核菌(Rhizoctonia cerealis)是引起我国小麦纹枯病的主要致病菌。为了建立高效稳定的禾谷丝核菌遗传转化体系,本试验比较研究了不同细胞壁降解酶、酶液浓度、酶处理温度和时间等因素对禾谷丝核菌原生质体制备的影响,利用正交设计试验优化了原生质体再生条件。结果表明,液体培养6d的菌丝,采用15mg/mL溶壁酶+10mg/mL蜗牛酶组成的混合酶液,30℃下酶解4h,可以获得较高的原生质体释放量,可达到3.0×106个/mL;禾谷丝核菌原生质体再生的最佳条件是以SuTC缓冲液作为渗透压稳定剂悬浮原生质体,采用单层混菌法接种于TB3再生培养基,原生质体再生率可达到58.6%。禾谷丝核菌原生质体制备和原生质体再生条件的优化,为深入研究禾谷丝核菌生长发育的分子遗传学基础和进一步探索小麦纹枯病的致病机理奠定了基础。