Identification of candidate virulence loci in Striga hermonthica, a devastating parasite of African cereal crops

Parasites have evolved suites of proteins, Virulence Factors (VFs), that are delivered into host plants to facilitate colonization. Whilst VFs mediating plant-microbe and plant-nematode interactions have been characterised extensively, less is known about VFs mediating parasitic plant interactions with their hosts. Striga hermonthica is an obligate, root-parasitic plant capable of parasitizing multiple cereal hosts in sub-Saharan Africa, causing devastating losses in yields. An understanding of the molecular nature and allelic variation of VFs in S. hermonthica is essential for breeding durable resistance and delaying the evolution of parasite virulence. To address this issue, we assembled a genome for Striga hermonthica and identified candidate VFs by combining in silico prediction of secreted proteins with pooled sequencing of parasites growing on a susceptible and a strongly resistant rice host. Consistent with predictions for parasites, like S. hermonthica, that can interact with multiple hosts, we identified multiple loci, potentially with a wide range of functions, implicated in overcoming host resistance. Extremely different allele frequencies were observed at 152 non-secreted and 38 putatively secreted VFs between S. hermonthica parasitising the resistant and susceptible rice varieties. Our candidate, secreted VFs encompassed functions such as host cell wall modification, protease inhibitors, oxidoreductase and kinase activities, as well as several with unknown functions. Consistent with maintenance of variation at virulence loci by balancing selection the candidate loci had significantly higher Tajima’s D on average than the genomic background. Our results show that diverse strategies are used by S. hermonthica to overcome different layers of host resistance. Understanding the maintenance of variation at virulence loci by balancing selection will be critical to managing the evolution of virulence as a part of a sustainable control strategy.

Two genomic regions of a sodium azide induced rice mutant confer broad-spectrum and durable resistance to blast disease

Rice blast, one of the most destructive epidemic diseases, annually causes severe losses in grain yield worldwide. To manage blast disease, breeding resistant varieties is considered a more economic and environment-friendly strategy than chemical control. For breeding new resistant varieties, natural germplasms with broad-spectrum resistance are valuable resistant donors, but the number is limited. Therefore, artificially induced mutants are an important resource for identifying new broad-spectrum resistant (R) genes/loci. To pursue this approach, we focused on a broad-spectrum blast resistant rice mutant line SA0169, which was previously selected from a sodium azide induced mutation pool of TNG67, an elite japonica variety. We found that SA0169 was completely resistant against the 187 recently collected blast isolates and displayed durable resistance for almost 20 years. Linkage mapping and QTL-seq analysis indicated that a 1.16-Mb region on chromosome 6 (Pi169-6(t)) and a 2.37-Mb region on chromosome 11 (Pi169-11(t)) conferred the blast resistance in SA0169. Sequence analysis and genomic editing study revealed 2 and 7 candidate R genes in Pi169-6(t) and Pi169-11(t), respectively. With the assistance of mapping results, six blast and bacterial blight double resistant lines, which carried Pi169-6(t) and/or Pi169-11(t), were established. The complementation of Pi169-6(t) and Pi169-11(t), like SA0169, showed complete resistance to all tested isolates, suggesting that the combined effects of these two genomic regions largely confer the broad-spectrum resistance of SA0169. The sodium azide induced mutant SA0169 showed broad-spectrum and durable blast resistance. The broad resistance spectrum of SA0169 is contributed by the combined effects of two R regions, Pi169-6(t) and Pi169-11(t). Our study increases the understanding of the genetic basis of the broad-spectrum blast resistance induced by sodium azide mutagenesis, and lays a foundation for breeding new rice varieties with durable resistance against the blast pathogen.

A Combination of Leaf Rust Resistance Genes, Including Lr34 and Lr46, Is the Key to the Durable Resistance of the Canadian Wheat Cultivar, Carberry

The hexaploid spring wheat cultivar, Carberry, was registered in Canada in 2009, and has since been grown over an extensive area on the Canadian Prairies. Carberry has maintained a very high level of leaf rust (Puccinia triticina Eriks.) resistance since its release. To understand the genetic basis of Carberry’s leaf rust resistance, Carberry was crossed with the susceptible cultivar, Thatcher, and a doubled haploid (DH) population of 297 lines was generated. The DH population was evaluated for leaf rust in seven field environments at the adult plant stage. Seedling and adult plant resistance (APR) to multiple virulence phenotypes of P. triticina was evaluated on the parents and the progeny population in controlled greenhouse studies. The population was genotyped with the wheat 90 K iSelect single nucleotide polymorphism (SNP) array, and quantitative trait loci (QTL) analysis was performed. The analysis using field leaf rust response indicated that Carberry contributed nine QTL located on chromosomes 1B, 2B (2 loci), 2D, 4A, 4B, 5A, 5B, and 7D. The QTL located on 1B, 2B, 5B, and 7D chromosomes were observed in two or more environments, whereas the remainder were detected in single environments. The resistance on 1B, detected in five environments, was attributed to Lr46 and on 7D, detected in seven environments to Lr34. The first 2B QTL corresponded with the adult plant gene, Lr13, while the second QTL corresponded with Lr16. The seedling analysis showed that Carberry carries Lr2a, Lr16, and Lr23. Five epistatic effects were identified in the population, with synergistic interactions being observed for Lr34 with Lr46, Lr16, and Lr2a. The durable rust resistance of Carberry is attributed to Lr34 and Lr46 in combination with these other resistance genes, because the resistance has remained effective even though the P. triticina population has evolved virulent to Lr2a, Lr13, Lr16, and Lr23.

Quantitative genetic analysis of interactions in the pepper-Phytophthora capsici pathosystem

ABSTRACTThe development of pepper cultivars with durable resistance to the oomycete Phytophthora capsici has been challenging due to differential interactions between the species that allow certain pathogen isolates to cause disease on otherwise resistant host genotypes. Currently, little is known about the pathogen genes that are involved in these interactions. To investigate the genetic basis of P. capsici virulence on individual pepper genotypes, we inoculated sixteen pepper accessions – representing commercial varieties, sources of resistance, and host differentials – with 117 isolates of P. capsici, for a total of 1,864 host-pathogen combinations. Analysis of disease outcomes revealed a significant effect of inter-species genotype-by-genotype interactions, although these interactions were quantitative rather than qualitative in scale. Isolates were classified into five pathogen subpopulations, as determined by their genotypes at over 60,000 single-nucleotide polymorphisms (SNPs). While absolute virulence levels on certain pepper accessions significantly differed between subpopulations, a multivariate phenotype reflecting relative virulence levels on certain pepper genotypes compared to others showed the strongest association with pathogen subpopulation. A genome-wide association study (GWAS) identified four pathogen loci significantly associated with virulence, two of which colocalized with putative RXLR effector genes and another with a polygalacturonase gene cluster. All four loci appeared to represent broad-spectrum virulence genes, as significant SNPs demonstrated consistent effects regardless of the host genotype tested. Host genotype-specific virulence variants in P. capsici may be difficult to map via GWAS, perhaps controlled by many genes of small effect or by multiple alleles that have arisen independently at the same loci.

Genetic Dissection of the Breakdown of Durable Resistance in Indica Rice Variety PTB33 to Brown Planthoppers Nilaparvata Lugens (Stål)

BackgroundTo avoid and delay the resistance breakdown of varieties against pathogens and insect pests, broad-spectrum and durable resistance by multiple genes pyramiding are expected to be one of the practical approaches. The indica rice variety PTB33 (Oryza sativa L.) shows high and durable resistance to the brown planthopper (BPH, Nilaparvata lugens Stål). However, this variety gradually lost its resistance against the recent virulence development of BPH. However, breakdown processes are not fully elucidated by individual genetic loci. ResultsEffective QTLs were explored across the whole genomic region against four BPH populations collected in Japan in 1988, 1989, 1999, and 2013 using high-density single-nucleotide polymorphism (SNP) markers obtained by genotyping-by-sequencing. Among seven genomic regions of PTB33 likely conferring BPH resistance, four QTLs, qFSA4a, qFSA6, qFSA11, and qFSA12 on chromosomes 4, 6, 11, and 12, respectively, were validated as BPH resistance QTLs. The PTB33 alleles at the four QTLs positively contributed to BPH resistance. Infestation of monogenic segregating lines showed that the PTB33 alleles at qFSA11, qFSA12, and qFSA4a lost resistance effects at least in 1989, 1999, and 2013, respectively. ConclusionThis study showed breakdown of durable resistance in PTB33 was explained by step-by-step losses of genetic effects at each resistance locus and probably multiple acquisitions of virulence genes in BPH in a gene-by-gene specific manner.

Bulked Segregant RNA-Seq Provides Distinctive Expression Profile Against Powdery Mildew in the Wheat Genotype YD588

Wheat powdery mildew, caused by the fungal pathogen Blumeria graminis f. sp. tritici (Bgt), is a destructive disease leading to huge yield losses in production. Host resistance can greatly contribute to the control of the disease. To explore potential genes related to the powdery mildew (Pm) resistance, in this study, we used a resistant genotype YD588 to investigate the potential resistance components and profiled its expression in response to powdery mildew infection. Genetic analysis showed that a single dominant gene, tentatively designated PmYD588, conferred resistance to powdery mildew in YD588. Using bulked segregant RNA-Seq (BSR-Seq) and single nucleotide polymorphism (SNP) association analysis, two high-confidence candidate regions were detected in the chromosome arm 2B, spanning 453,752,054-506,356,791 and 584,117,809-664,221,850 bp, respectively. To confirm the candidate region, molecular markers were developed using the BSR-Seq data and mapped PmYD588 to an interval of 4.2 cM by using the markers YTU588-004 and YTU588-008. The physical position was subsequently locked into the interval of 647.1–656.0 Mb, which was different from those of Pm6, Pm33, Pm51, Pm52, Pm63, Pm64, PmQ, PmKN0816, MlZec1, and MlAB10 on the same chromosome arm in its position, suggesting that it is most likely a new Pm gene. To explore the potential regulatory genes of the R gene, 2,973 differentially expressed genes (DEGs) between the parents and bulks were analyzed using gene ontology (GO), clusters of orthologous group (COG), and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analysis. Based on the data, we selected 23 potential regulated genes in the enriched pathway of plant-pathogen interaction and detected their temporal expression patterns using an additional set of wheat samples and time-course analysis postinoculation with Bgt. As a result, six disease-related genes showed distinctive expression profiles after Bgt invasion and can serve as key candidates for the dissection of resistance mechanisms and improvement of durable resistance to wheat powdery mildew.

The influence of endophytes on rice fitness under environmental stresses

Key Message Endophytes are crucial for the promotion of rice growth and stress tolerance and can be used to increase rice crop yield. Endophytes can thus be exploited in biotechnology and genetic engineering as eco-friendly and cost-effective means for the development of high-yielding and stress-tolerant rice plants. Abstract Rice (Oryza sativa) crop is continuously subjected to biotic and abiotic stresses, compromising growth and consequently yield. The situation is exacerbated by climate change impacting on ecosystems and biodiversity. Genetic engineering has been used to develop stress-tolerant rice, alongside physical and chemical methods to mitigate the effect of these stresses. However, the success of these strategies has been hindered by short-lived field success and public concern on adverse effects associated. The limited success in the field of stress-tolerant cultivars developed through breeding or transgenic approaches is due to the complex nature of stress tolerance as well as to the resistance breakdown caused by accelerated evolution of pathogens. It is therefore necessary to develop novel and acceptable strategies to enhance rice stress tolerance and durable resistance and consequently improve yield. In the last decade, plant growth promoting (PGP) microbes, especially endophytes, have drawn the attention of agricultural scientists worldwide, due to their ability to mitigate environmental stresses in crops, without causing adverse effects. Increasing evidence indicates that endophytes effectively confer fitness benefits also to rice under biotic and abiotic stress conditions. Endophyte-produced metabolites can control the expression of stress-responsive genes and improve the physiological performance and growth of rice plants. This review highlights the current evidence available for PGP microbe-promoted tolerance of rice to abiotic stresses such as salinity and drought and to biotic ones, with special emphasis on endophytes. Associated molecular mechanisms are illustrated, and prospects for sustainable rice production also in the light of the impending climate change, discussed.

Wheat genetic loci conferring resistance to stripe rust in the face of genetically diverse races of the fungus Puccinia striiformis f. sp. tritici

Key message Analysis of a wheat multi-founder population identified 14 yellow rust resistance QTL. For three of the four most significant QTL, haplotype analysis indicated resistance alleles were rare in European wheat. Abstract Stripe rust, or yellow rust (YR), is a major fungal disease of wheat (Triticum aestivum) caused by Puccinia striiformis Westend f. sp. tritici (Pst). Since 2011, the historically clonal European Pst races have been superseded by the rapid incursion of genetically diverse lineages, reducing the resistance of varieties previously showing durable resistance. Identification of sources of genetic resistance to such races is a high priority for wheat breeding. Here we use a wheat eight-founder multi-parent population genotyped with a 90,000 feature single nucleotide polymorphism array to genetically map YR resistance to such new Pst races. Genetic analysis of five field trials at three UK sites identified 14 quantitative trait loci (QTL) conferring resistance. Of these, four highly significant loci were consistently identified across all test environments, located on chromosomes 1A (QYr.niab-1A.1), 2A (QYr.niab-2A.1), 2B (QYr.niab-2B.1) and 2D (QYr.niab-2D.1), together explaining ~ 50% of the phenotypic variation. Analysis of these four QTL in two-way and three-way combinations showed combinations conferred greater resistance than single QTL, and genetic markers were developed that distinguished resistant and susceptible alleles. Haplotype analysis in a collection of wheat varieties found that the haplotypes associated with YR resistance at three of these four major loci were rare (≤ 7%) in European wheat, highlighting their potential utility for future targeted improvement of disease resistance. Notably, the physical interval for QTL QYr.niab-2B.1 contained five nucleotide-binding leucine-rich repeat candidate genes with integrated BED domains, of which two corresponded to the cloned resistance genes Yr7 and Yr5/YrSp. Graphical abstract

Genetic Resistance of Lactuca spp. against Fusarium oxysporum f. sp. lactucae Race 1

Fusarium wilt of lettuce is caused by the pathogen Fusarium oxysporum f. sp. lactucae (Fol) and is a growing threat to global lettuce production. Fol was first detected in Florida in 2017 and was subsequently confirmed as race 1. Management strategies for this long-persisting soil pathogen are limited, time-consuming and expensive, and they may lack efficacy. Identifying diverse sources of genetic resistance is imperative for breeding adapted cultivars with durable resistance. The objectives of this study were to identify sources of resistance against a race 1 isolate of Fol in Florida, delineate the relationship between foliar and taproot symptoms, and investigate the inheritance of resistance and partial resistance in two F2 populations. Thirteen experiments were conducted in greenhouse and field locations to characterize the diversity of genetic resistance in the genus Lactuca. Leaf cultivars Dark Lollo Rossa and Galactic; romaine breeding lines 43007, 60182, and C1145; and iceberg breeding line 47083 consistently exhibited low foliar and taproot disease symptoms. Resistance was not identified among the wildtype Lactuca or primitive plant introductions (PI) in this study based on taproot symptoms. An additional test was conducted to study the segregation pattern of Fol resistance between one resistant and one susceptible accession (R × S) and one partial resistant and one susceptible accession (PR × S). The F2 population from ‘60182 × PI 358001-1’ fit the expected segregation ratio for a single recessive locus model, whereas the ratio for ‘Dark Lollo Rossa × PI 358001-1’ did not fit either recessive or dominant single locus models. These sources of resistance are potential candidates for developing commercial cultivars with multiple resistance loci against Fol race 1, especially for the Florida lettuce production system.