Genetic dissection of the resistance to Rice stripe virus present in the indica rice cultivar 'IR24'

Rice stripe disease, caused by Rice stripe virus (RSV) and transmitted by the small brown planthopper (Laodelphax striatellus Fallen), is one of the most serious viral diseases of rice in temperate East Asian production regions. Prior quantitative trait loci (QTL) mapping has established that Oryza sativa L. subsp. indica 'IR24' carries positive alleles at the three loci qSTV3, qSTV7, and qSTV11-i. Here, we report an advanced backcross analysis based on three selected chromosome segment substitution lines (CSSLs), each predicted to carry one of these three QTL. Three sets of BC(4)F(2:3) populations were bred from a cross between the critical CSSL and its recurrent parent Oryza sativa L. subsp. japonica 'Asominori'. Both qSTV3 and qSTV11-i were detected in their respective population, but qSTV7 was not. An allelic analysis based on a known carrier of the major RSV resistance gene Stvb-i, which is located on chromosome 11, showed that qSTV11-i was not allelic with Stvb-i. A large mapping population was used to delimit the location of qSTV11-i to a 73.6-kb region. The de novo markers developed for this purpose will be useful as marker-assisted selection tools in efforts to introduce qSTV11-i into breeding programmes aiming to improve the level of RSV resistance.     

Mapping quantitative trait loci conferring resistance to rice black-streaked virus in maize (Zea mays L.)

Maize rough dwarf disease (MRDD) is one of the most serious virus diseases of maize worldwide, and it causes great reduction of maize production. In China, the pathogen was shown to be rice black-streaked virus (RBSDV). Currently, MRDD has spread broadly and leads to significant loss in China. However, there has been little research devoted to this disease. Our aims were to identify the markers and loci underlying resistance to this virus disease. In this study, segregation populations were constructed from two maize elite lines '90110', which is highly resistant to MRDD and 'Ye478', which is highly susceptible to MRDD. The F(2) and BC(1) populations were used for bulk sergeant analysis (BSA) to identify resistance-related markers. One hundred and twenty F(7:9) RILs were used for quantitative trait loci (QTL) mapping through the experiment of multiple environments over 3 years. Natural occurrence and artificial inoculation were both used and combined to determine the phenotype of plants. Five QTL, qMRD2, qMRD6, qMRD7, qMRD8 and qMRD10 were measured in the experiments. The qMRD8 on chromosome 8 was proved to be one major QTL conferring resistance to RBSDV disease in almost all traits and environments, which explained 12.0-28.9 % of the phenotypic variance for disease severity in this present study.     

水稻条纹叶枯病抗性位点的检测和效应分析

利用111个家系组成的热研2号(Oryza sativa subsp. japonica ‘Reyan2’)/ Mi lyang23(Oryza sativa subsp. indica ‘Mi lyang23’)重组自交系(recombinant inbred l ines, RIL)群体(F7), 采用重病区田间自然接种方法, 以病情指数作为条纹叶枯病的表型值, 鉴定了2个亲本及111个RIL家系对条纹叶枯病的抗性。使用QTL Cartographer 软件复合区间作图法, 对水稻(Oryza a sativa)条纹叶枯病抗性基因进行了QTL分析。结果检测到2个抗水稻条纹叶枯病的QTL, 分别位于第2和第11染色体上, 其中第11染色体上的QTL贡献率为19.58%, 表明这是一个主效的QTL, 该QTL及其附近的分子标记, 可以用于水稻条纹叶枯病抗性分子标记辅助育种。     

水稻条纹叶枯病抗性位点的检测和效应分析

利用111个家系组成的热研2号（Oryza sativa subsp．japonica‘Reyan2’）／Milyang23（Oryza sativa subsp．indica‘Milyang23’）重组自交系（recombinant inbred lines,RIL）群体（F7）,采用重病区田间自然接种方法,以病情指数作为条纹叶枯病的表型值,鉴定了2个亲本及111个RIL家系对条纹叶枯病的抗性。使用QTL Cartographer软件复合区间作图法,对水稻（Oryza sativa）条纹叶枯病抗性基因进行了QTL分析。结果检测到2个抗水稻条纹叶枯病的QTL,分别位于第2和第11染色体上,其中第11染色体上的QTL贡献率为19．58％,表明这是一个主效的QTL,该QTL及其附近的分子标记,可以用于水稻条纹叶枯病抗性分子标记辅助育种。     

利用AFLP遗传连锁图定位大麦苗期对叶锈病的部分抗性基因

借助大麦染色体ＡＦＬＰ标记遗传连锁图和ＭａｐＱＴＬＶ３．０作图软件,对大麦叶病的数量抗性基因进行了定位分析,明确了大麦部分抗性品种Ｖａｄａ对叶锈病的潜育期由分别位于染色体１、２、６、７上离短臂末端７９ｃＭ、１８６ｃＭ、５８ｃＭ和１１７ｃＭ处的４个数量抗性基因所控制。     

Identification and fine mapping of qRBSDV-6 MH , a major QTL for resistance to rice black-streaked dwarf virus disease

Rice black streaked-dwarf virus (RBSDV) disease is recently expanding in southern China and poses a serious threat to rice crops. Few studies related to the genetics and breeding of RBSDV resistance have been reported. We have previously mapped a number of quantitative trait loci (QTLs) for RBSDV resistance by using a recombinant inbred line population of ‘Zhenshan 97’ (ZS97, susceptible)/‘Minghui 63’ (MH63, resistant) with natural infection data in two locations. In the present study, we confirmed the presence of a number of resistant QTLs on chromosomes 6, 7, and 9 from MH63 by using the same population in four different locations. We then focused on a major QTL, qRBSDV-6 ^MH , on chromosome 6 and introduced it into a highly susceptible japonica rice variety, ‘Huaidao 5’, using MH63 as the donor via marker-assisted selection, to generate seven backcross inbred lines (BILs). Natural infection and artificial inoculation-based tests revealed that all of the BILs had a significantly higher resistance to RBSDV than the recurrent parent. These results demonstrate that qRBSDV-6 ^MH is a stable major resistance QTL of high breeding value. We also constructed a set of chromosome segment substitution lines (CSSLs) specific to the qRBSDV-6 ^MH region and these used as fine mapping population. Combining the genotypes of CSSLs with the phenotypes from natural infection data in a highly RBSDV epidemic area during two different sowing seasons, we were able to precisely map qRBSDV-6 ^MH to the markers S18 and S23 at a physical distance of 627.6 kb on the Nipponbare reference genome.     

Identification of quantitative trait loci controlling resistance to maize chlorotic dwarf virus

Ineffective screening methods and low levels of disease resistance have hampered genetic analysis of maize (Zea mays L.) resistance to disease caused by maize chlorotic dwarf virus (MCDV). Progeny from a cross between the highly resistant maize inbred line Oh1VI and the susceptible inbred line Va35 were evaluated for MCDV symptoms after multiple virus inoculations, using the viral vector Graminella nigrifrons. Symptom severity scores from three rating dates were used to calculate area under the disease progress curve (AUDPC) scores for vein banding, leaf twist and tear, and whorl chlorosis. AUDPC scores for the F₂ population indicated that MCDV resistance was quantitatively inherited. Genotypic and phenotypic analyses of 314 F₂ individuals were compared using composite interval mapping (CIM) and analysis of variance. CIM identified two major quantitative trait loci (QTL) on chromosomes 3 and 10 and two minor QTL on chromosomes 4 and 6. Resistance was additive, with alleles from Oh1VI at the loci on chromosomes 3 and 10 contributing equally to resistance.     

Quantitative resistance to late blight from Solanum berthaultii cosegregates with R Pi-ber : insights in stability through isolates and environment

Genetic resistance is a valuable tool in the fight against late blight of potatoes but little is known about the stability and specificity of quantitative resistance including the effect of defeated major resistance genes. In the present study we investigated the effect of different isolates of Phytophthora infestans on the mode of action of R _Pi-ber , an R-gene originating from Solanum berthaultii. The experiments were conducted on progenies derived from two reciprocal inter-specific backcrosses of Solanum tuberosum and S. berthaultii. The plant–pathogen interaction was tested in diverse environments including field, greenhouse and growth chamber conditions. The R _Pi-ber gene provided complete resistance against a US8 isolate of P. infestans in all trials. When isolates compatible with R _Pi-ber were used for inoculation, a smaller, but significant resistance effect was consistently detected in the same map position as the R-gene. This indicates that this R-gene provides a residual resistance effect, and/or that additional resistance loci are located in this genomic region of chromosome X. Additional quantitative resistance loci (QRL) were identified in the analyzed progenies. While some of the QRL (such as those near TG130 on chromosome III) were effective against several isolates of the pathogen, others were isolate specific. With a single exception, the S. berthaultii alleles were associated with a decrease in disease severity. Resistance loci reported in the present study co-locate with previously reported R-genes and QRL to P. infestans and other pathogens.     

Mapping multiple disease resistance genes using a barley mapping population evaluated in Peru, Mexico, and the USA

We used a well-characterized barley mapping population (BCD 47 × Baronesse) to determine if barley stripe rust (BSR) resistance quantitative trait loci (QTL) mapped in Mexico and the USA were effective against a reported new race in Peru. Essentially the same resistance QTL were detected using data from each of the three environments, indicating that these resistance alleles are effective against the spectrum of naturally occurring races at these sites. In addition to the mapping population, we evaluated a germplasm array consisting of lines with different numbers of mapped BSR resistance alleles. A higher BSR disease severity on CI10587, which has a single qualitative resistance gene, in Peru versus Mexico suggests there are differences in pathogen virulence between the two locations. Confirmation of a new race in Peru will require characterization using a standard set of differentials, an experiment that is underway. The highest levels of resistance in Peru were observed when the qualitative resistance gene was pyramided with quantitative resistance alleles. We also used the mapping population to locate QTL conferring resistance to barley leaf rust and barley powdery mildew. For mildew, we identified resistance QTL under field conditions in Peru that are distinct from the Mla resistance that we mapped using specific isolates under controlled conditions. These results demonstrate the long-term utility of a reference mapping population and a well-characterized germplasm array for locating and validating genes conferring quantitative and qualitative resistance to multiple pathogens.     

Genome-Wide Association Study Reveals Novel Quantitative Trait Loci Associated with Resistance to Multiple Leaf Spot Diseases of Spring Wheat

Accelerated wheat development and deployment of high-yielding, climate resilient, and disease resistant cultivars can contribute to enhanced food security and sustainable intensification. To facilitate gene discovery, we assembled an association mapping panel of 528 spring wheat landraces of diverse geographic origin for a genome-wide association study (GWAS). All accessions were genotyped using an Illumina Infinium 9K wheat single nucleotide polymorphism (SNP) chip and 4781 polymorphic SNPs were used for analysis. To identify loci underlying resistance to the major leaf spot diseases and to better understand the genomic patterns, we quantified population structure, allelic diversity, and linkage disequilibrium. Our results showed 32 loci were significantly associated with resistance to the major leaf spot diseases. Further analysis identified QTL effective against major leaf spot diseases of wheat which appeared to be novel and others that were previously identified by association analysis using Diversity Arrays Technology (DArT) and bi-parental mapping. In addition, several identified SNPs co-localized with genes that have been implicated in plant disease resistance. Future work could aim to select the putative novel loci and pyramid them in locally adapted wheat cultivars to develop broad-spectrum resistance to multiple leaf spot diseases of wheat via marker-assisted selection (MAS).     

Targeted discovery of quantitative trait loci for resistance to northern leaf blight and other diseases of maize

To capture diverse alleles at a set of loci associated with disease resistance in maize, heterogeneous inbred family (HIF) analysis was applied for targeted QTL mapping and near-isogenic line (NIL) development. Tropical maize lines CML52 and DK888 were chosen as donors of alleles based on their known resistance to multiple diseases. Chromosomal regions (“bins”; n = 39) associated with multiple disease resistance (MDR) were targeted based on a consensus map of disease QTLs in maize. We generated HIFs segregating for the targeted loci but isogenic at ~97% of the genome. To test the hypothesis that CML52 and DK888 alleles at MDR hotspots condition broad-spectrum resistance, HIFs and derived NILs were tested for resistance to northern leaf blight (NLB), southern leaf blight (SLB), gray leaf spot (GLS), anthracnose leaf blight (ALB), anthracnose stalk rot (ASR), common rust, common smut, and Stewart’s wilt. Four NLB QTLs, two ASR QTLs, and one Stewart’s wilt QTL were identified. In parallel, a population of 196 recombinant inbred lines (RILs) derived from B73 × CML52 was evaluated for resistance to NLB, GLS, SLB, and ASR. The QTLs mapped (four for NLB, five for SLB, two for GLS, and two for ASR) mostly corresponded to those found using the NILs. Combining HIF- and RIL-based analyses, we discovered two disease QTLs at which CML52 alleles were favorable for more than one disease. A QTL in bin 1.06–1.07 conferred resistance to NLB and Stewart’s wilt, and a QTL in 6.05 conferred resistance to NLB and ASR.     

Detection and fine mapping of two quantitative trait loci for partial resistance to stripe virus in rice (Oryza sativa L.)

Rice stripe virus (RSV) is one of the most destructive pathogens of rice (Oryza sativa L.) in East Asia. Development of resistant varieties offers a more economical and efficient way to control this disease. In the present study, tests using four inoculation methods were used on 85 backcross inbred lines of Sasanishiki (japonica)/Habataki (indica) to map quantitative trait loci (QTL) conferring resistance to RSV. One QTL on chromosome 3 and two on chromosome 11 were detected, jointly explaining 18?C47?% of the trait variance. The QTL (qSTV11 ^HAB -1 and qSTV11 ^HAB -2) on chromosome 11 were closely linked, and mapped in the intervals G257-RM457 and RM457-RM187, respectively. The stabilities of qSTV11 ^HAB -1 and qSTV11 ^HAB -2 were validated using a set of 38 established chromosome segmental substitution lines. The two QTL, when combined, showed higher resistance than either of them alone in both field and mass inoculation tests, indicating additivity. Fine mapping of the two genes was carried out using 147 recombined F2:3 lines selected from 2,750 secondary F2 plants of the cross Sasanishiki/SL437. Four SSR (simple sequence repeat) and eight InDel (insertion?Cdeletion) markers newly developed to fine-map the two loci. According to the Nipponbare genomic sequence, qSTV11 ^HAB -1 was localized to a 333.2-kb interval which was about 230?kb from the well-known Stvb-i. The other locus, qSTV11 ^HAB -2, which appears to be a new QTL for RSV resistance, was delimited to a 203.9-kb region. Four flanking markers (R15, RM209, R69 and R73) can be used in marker-assisted selection. These results provide an opportunity for map-based cloning of qSTV11 ^HAB -1 and qSTV11 ^HAB -2, thereby promoting the breeding program of RSV resistance.     

Characterization of Rice Black-Streaked Dwarf Virus- and Rice Stripe Virus-Derived siRNAs in Singly and Doubly Infected Insect Vector Laodelphax striatellus

Replication of RNA viruses in insect cells triggers an antiviral defense that is mediated by RNA interference (RNAi) which generates viral-derived small interfering RNAs (siRNAs). However, it is not known whether an antiviral RNAi response is also induced in insects by reoviruses, whose double-stranded RNA genome replication is thought to occur within core particles. Deep sequencing of small RNAs showed that when the small brown planthopper (Laodelphax striatellus) was infected by Rice black-streaked dwarf virus (RBSDV) (Reoviridae; Fijivirus), more viral-derived siRNAs accumulated than when the vector insect was infected by Rice stripe virus (RSV), a negative single-stranded RNA virus. RBSDV siRNAs were predominantly 21 and 22 nucleotides long and there were almost equal numbers of positive and negative sense. RBSDV siRNAs were frequently generated from hotspots in the 5′- and 3′-terminal regions of viral genome segments but these hotspots were not associated with any predicted RNA secondary structures. Under laboratory condition, L. striatellus can be infected simultaneously with RBSDV and RSV. Double infection enhanced the accumulation of particular genome segments but not viral coat protein of RBSDV and correlated with an increase in the abundance of siRNAs derived from RBSDV. The results of this study suggest that reovirus replication in its insect vector potentially induces an RNAi-mediated antiviral response.     

小麦全基因组抗赤霉病QTL关联位点特异性SSR标记的筛选、等位变异及效应解析

赤霉病是小麦主要的流行病害之一。借助标记辅助选择将不同数量性状基因座（quantitative trait loci，QTL）聚合是防治赤霉病有效且环保的方法，可以从源头上控制赤霉病并降低籽粒中毒素含量。抗赤霉病QTL在小麦全基因组均有分布，但除了Fhb1、Fhb2等少数位点有比较可靠的鉴别标记，绝大部分位点缺乏有效的位点特异性鉴别标记。简单重复序列（simple sequence repeat，SSR）标记多态性丰富，可以区分自然群体中不同等位变异，方便用于标记辅助育种。基于此，搜集了不同文献中报道的与赤霉病关联的SSR标记386个，并用这些标记构建全基因组赤霉病抗性QTL一致性图谱，接着对这些关联标记进行拷贝数分析，进而选择位点内的单拷贝SSR标记，将这些单拷贝标记在156个品种组成的自然群体中进行扩增，并与三季大田和三季温室环境下赤霉病抗性进行关联，筛选与赤霉病抗性关联的单拷贝SSR标记，明确这些标记在自然群体中的有效等位变异和效应。结果表明，共8个单拷贝SSR标记至少在两季试验中与表型显著关联(P<0.05)，涉及2B、2D、3B、5A、5B、6A、6D、7A染色体，有5个单拷贝标记位点存在有效等位变异。中国地方品种和日本品种携带更多的有利变异，且有利等位变异数目越多的品种赤霉病抗性越好。研究分析的QTL位点及其关联的单拷贝SSR标记可用于赤霉病抗病育种，有利于提高品种赤霉病抗性水平和育种效率。     

Genomic regions conferring resistance to multiple fungal pathogens in synthetic hexaploid wheat

Fungal diseases are among the most devastating biotic stresses and often cause significant losses in wheat production worldwide. A set of 173 synthetic hexaploid wheat (SHW) characterized for resistance against fungal pathogens that cause leaf, stem and yellow rusts, yellow leaf spot, Septoria nodorum and crown rot were used in genome-wide association study (GWAS). Diversity Arrays Technology (DArT) and DArTSeq markers were employed for marker–trait association in which 74 markers associated with 35 quantitative trait loci (QTL) were found to be significantly linked with disease resistances using a unified mixed model (P = 10^?3 to 10^?5); Of these 15 QTL originated from D genome. Six markers on 1BL, 3BS, 4BL, 6B, and 6D conferred resistance to two diseases representing 10 of the 35 QTL. A further set of 147 SHW genotyped with DArT only markers validated 11 QTL detected in the previous 173 SHW. We also confirmed the presence of the gene Lr46/Yr29/Sr58/Pm39/Ltn2 on 1BL in the SHW germplasm. In addition, gene–gene interactions between significantly associated loci and all loci across the genome revealed five significant interactions at FDR <0.05. Two significant leaf rust and one stem rust interactions were thought to be synergistic, while another two QTL for yellow leaf spot involved antagonistic relations. To the best of our knowledge, this is the first GWAS for six fungal diseases using SHW. Identification of markers associated with disease resistance to one or more diseases represents an important resource for pyramiding favorable alleles and introducing multiple disease resistance from SHW accessions into current elite wheat cultivars.     

RNA N6-methyladenosine modification suppresses replication of rice black streaked dwarf virus and is associated with virus persistence in its insect vector

N⁶ methylation of adenosine (m⁶A) was recently discovered to play a role in regulating the life cycle of various viruses by modifying viral and host RNAs. However, different studies on m⁶A effects on the same or different viruses have revealed contradictory roles for m⁶A in the viral life cycle. In this study, we sought to define the role of m⁶A on infection by rice black streaked dwarf virus (RBSDV), a double-stranded RNA virus, of its vector small brown planthopper (SBPH). Infection by RBSDV decreased the level of m⁶A in midgut cells of SBPHs. We then cloned two genes (LsMETTL3 and LsMETTL14) that encode m⁶A RNA methyltransferase in SBPHs. After interference with expression of the two genes, the titre of RBSDV in the midgut cells of SBPHs increased significantly, suggesting that m⁶A levels were negatively correlated with virus replication. More importantly, our results revealed that m⁶A modification might be the epigenetic mechanism that regulates RBSDV replication in its insect vector and maintains a certain virus threshold required for persistent transmission.     

Joint linkage QTL analyses for partial resistance to Phytophthora sojae in soybean using six nested inbred populations with heterogeneous conditions

Partial resistance to Phytophthora sojae in soybean is controlled by multiple quantitative trait loci (QTL). With traditional QTL mapping approaches, power to detect such QTL, frequently of small effect, can be limited by population size. Joint linkage QTL analysis of nested recombinant inbred line (RIL) populations provides improved power to detect QTL through increased population size, recombination, and allelic diversity. However, uniform development and phenotyping of multiple RIL populations can prove difficult. In this study, the effectiveness of joint linkage QTL analysis was evaluated on combinations of two to six nested RIL populations differing in inbreeding generation, phenotypic assay method, and/or marker set used in genotyping. In comparison to linkage analysis in a single population, identification of QTL by joint linkage analysis was only minimally affected by different phenotypic methods used among populations once phenotypic data were standardized. In contrast, genotyping of populations with only partially overlapping sets of markers had a marked negative effect on QTL detection by joint linkage analysis. In total, 16 genetic regions with QTL for partial resistance against P. sojae were identified, including four novel QTL on chromosomes 4, 9, 12, and 16, as well as significant genotype-by-isolate interactions. Resistance alleles from PI 427106 or PI 427105B contributed to a major QTL on chromosome 18, explaining 10–45 % of the phenotypic variance. This case study provides guidance on the application of joint linkage QTL analysis of data collected from populations with heterogeneous assay conditions and a genetic framework for partial resistance to P. sojae.     

Quantitative trait locus mapping of pyrethroid resistance in Colorado potato beetle, Leptinotarsa decemlineata (Say) (Coleoptera: Chrysomelidae)

Quantitative trait locus (QTL) mapping is a valuable new tool for locating genomic regions that underlie variation in important traits such as insecticide resistance. Because QTL mapping complements a candidate gene strategy for understanding the genetic architecture of important traits, it may also facilitate the identification of genes causing important variation. After mapping the QTL locations, markers closely linked to QTL can be used for genetic analysis of population structure and to measure the spread and increase of resistance-causing QTL alleles. In this study, QTL influencing resistance to the pyrethroid insecticide esfenvalerate were mapped in the Colorado potato beetle Leptinotarsa decemlineata (Say) (CPB). Three QTL contributing to esfenvalerate resistance were identified from a mapping population of 79 individuals analyzed at 90 marker loci. One QTL had a large effect and two QTL had smaller effects. The major QTL occurs on the X chromosome, overlapping the position of a candidate gene (Leptinotarsa decemlineata Voltage sensitive sodium channel [LdVssc1]) previously implicated in pyrethroid resistance. Resistance-increasing alleles at the two minor-effect QTL originated with the susceptible parent, suggesting that alleles of small effect may be segregating in susceptible populations. Comparison of the New York population from which the susceptible parent originated with a more-susceptible population from North Carolina suggests that the minor-effect loci identified here may explain some of the variation in tolerance observed among susceptible populations. DNA sequencing of a portion of LdVssc1 shows that the resistance-conferring allele from the resistant parent does not contain the kdr mutation previously found in CPB and typically observed in other insects that are resistant to pyrethroid insecticides because of changes in this gene.