Biology subjects

Dodds, P. N.

Publications and source records attributed to Dodds, P. N..

14 recordsLinked to original sources

bioRxiv2026

Haplotype-phased genomes of the barley leaf rust pathogen reveal evidence of repeat element expansion and somatic hybridization

Barley leaf rust disease, caused by Puccinia hordei, leads to substantial yield losses and diminished malting quality of barley across temperate growing regions worldwide. To address the paucity of high-resolution genomic resources for this pathogen, we generated haplotype-phased, chromosome-scale assemblies for ten globally distributed isolates using PacBio HiFi and Hi-C sequencing. Phylogenomic analysis revealed seven distinct lineages of P. hordei, including evidence of nuclear exchange, with a shared nuclear haplotype detected between two US lineages. Nuclear genome sizes ranged from [~]140-147Mbp, with the exception of isolate 90ISR03 from Israel ([~]163Mbp), which also harbored a 6.2Mbp supernumerary scaffold in one nucleus exhibiting chromosomal characteristics. Consistent with its larger genome, P. hordei had a higher repeat content ([~]70%) than related cereal rust fungi, driven primarily by the proliferation of LTR retroelements and DNA transposons. Across the global pan-genome of 13 unique nuclear haplotypes, approximately one-third of all protein orthogroups were conserved across all isolates. Only 18% of predicted effector orthogroups were shared between all haplotypes, reflecting the highly dynamic and variable nature of the effector repertoire. The long-term propagation of clonal P. hordei lineages is apparent both within the US and globally, and nuclear exchange is important for generating novel diversity and virulence profiles. The high degree of genome plasticity is evident in extensive structural variation, including large-scale translocations and inversions as well as a putative accessory chromosome. These chromosome-level, haplotype-resolved genomes provide a foundational resource for exploring the evolution, diversity, and avirulence gene repertoire of P. hordei.

genomics↗

bioRxiv2025

Advancing rust resistance in elite wheat with haplotype mapping and a novel introgression strategy

Wheat production is continually threatened by stripe and leaf rust because virulent races rapidly overcome single race-specific genes. Durable, broad-spectrum resistance is needed. Adult plant resistance (APR) provides partial, stable resistance from multiple minor-effect loci acting additively; pleiotropic loci like Lr34/Yr18 and Lr46/Yr29 add durability. We used an elite Australian panel (OzWheat=589) and a diverse landrace panel (Vavilov=295), genotyped with [~]30K SNPs and phenotyped across environments. Linkage disequilibrium partitioning defined 7,659 genome-wide haploblocks. To prioritise robust signals, we ranked haploblocks by haplotype effect variance and examined the top 100 per trait. For stripe rust, 52/100 were significant, with 32 shared across panels; for leaf rust, 50 were significant, 29 also detected in Vavilov. Several intervals co-localised with APR regions (Lr46/Yr29), and one 7BL interval intersected seedling gene Lr14a. To translate mapping into breeding decisions, we developed an introgression fitness index to quantify the value of resistant haplotypes in elite backgrounds. Using elite cultivar Scepter, we applied a genetic algorithm to select 50 donor parents carrying desirable haplotypes. Simulations showed that pyramiding these haplotypes can enhance resistance while maintaining elite genomic background. This study provides practical breeding tools, including haplotype catalogue and a novel selection index to accelerate rust-resistant wheat development.

genetics↗

bioRxiv2025

Virulence on Pm4 kinase-based resistance is determined by two divergent wheat powdery mildew effectors

The wheat resistance gene Pm4 encodes a kinase fusion protein and has gained particular attention as it confers race-specific resistance against two major wheat pathogens: powdery mildew and blast. Here, we describe the identification of AvrPm4, the mildew avirulence effector recognised by Pm4, using UV- mutagenesis, and its functional validation in wheat protoplasts. We show that AvrPm4 directly interacts with and is phosphorylated by Pm4. Using genetic association and QTL mapping, we furthermore demonstrate that evasion of Pm4 resistance by virulent mildew isolates relies on a second fungal component, SvrPm4, which suppresses AvrPm4-induced cell death. Surprisingly, SvrPm4 was previously described as AvrPm1a. We show that SvrPm4, but not its inactive variant svrPm4, is recognised by the NLR immune receptor Pm1a. These multiple roles of a single effector provide a new perspective on fungal (a)virulence proteins and their combinatorial interactions with different types of immune receptors.

genetics↗

bioRxiv2024

Translocation of effector proteins into plant cells by the flax rust pathogen Melampsora lini

During infection, rust fungi secrete effector proteins into host plant cells from haustoria to aid their colonisation. How rust effectors are secreted from the haustorium and delivered into the cytoplasm of host cells remains poorly understood. We used an Agrobacterium-mediated transformation procedure to generate stable transgenic flax rust strains expressing the effectors AvrM and AvrP123 fused to yellow fluorescent protein (YFP). We showed that both AvrM-YFP and AvrP123-YFP fusion proteins were secreted by the fungus into the extrahaustorial matrix (EHMx); however only AvrM-YFP was delivered into host cells, triggering a typical resistance phenotype in plants carrying the corresponding resistance (R) gene M. The signal peptide of AvrM was sufficient to direct YFP secretion into the EHMx; however, delivery into the host cell required a larger 105 amino acid N-terminal fragment of AvrM. These results indicate that translocation of this protein into the host cell from the EHMx is a separate process from secretion into the EHMx and requires a signal present in AvrM between amino acids 34 and 105. This is in contrast to previous observations of AvrM localisation after transient expression in plants, highlighting the necessity for analysis in the natural infection system.

plant biology↗

bioRxiv2024

A wheat tandem kinase sensor activates an NLR helper to trigger immunity

Most plant resistance genes encode membrane-anchored receptor-like proteins or intracellular nucleotide-binding and leucine-rich repeat (NLR) receptors. In wheat and barley, tandem kinases (TKs) have emerged as a new class of resistance determinants. To understand the modus operandi of the wheat stem rust resistance protein Sr62TK, we identified two genetic interactors-- a host gene required for Sr62TK function and the corresponding fungal AvrSr62 effector. We discovered that the SR62 locus consists of a digenic module encoding Sr62TK and an NLR (Sr62NLR). AvrSr62 binds to the N-terminal kinase of Sr62TK. This triggers displacement of the C-terminal kinase allowing it to recruit Sr62NLR for activation of immune responses. Understanding the mechanism of this two-component resistance complex will help engineering and breeding for durable resistance.

plant biology↗

bioRxiv2023

Genotypic and resistance profile analysis of two oat crown rust differential sets urge coordination and standardisation

Puccinia coronata f. sp. avenae (Pca) is the causal agent of the disease known as crown rust which represents a bottleneck in oat production worldwide. Characterisation of pathogen populations often involves race (pathotype) assignments using differential sets, which are not uniform across countries. This study compared virulence profiles of 25 Pca isolates from Australia using two host differential sets, one from Australia and one from the USA. These differential sets were also genotyped using DArT sequencing technology. Phenotypic and genotypic discrepancies were detected on eight out of 29 common lines between the two sets, indicating that pathogen race assignments based on those lines are not comparable. To further investigate molecular markers that could assist in the stacking of rust resistance genes important for Australia, four published Pc91-linked markers were validated across the differential sets and then screened across a collection of 150 oat cultivars. Drover, Aladdin, and Volta were identified as putative carriers of the Pc91 locus. This is the first report to confirm that the cultivar Volta carries Pc91 and demonstrates the value of implementing molecular markers to characterise materials in breeding pools of oat. Overall, our findings highlight the necessity of examining seed stocks using pedigree and molecular markers to ensure seed uniformity and bring robustness to surveillance methodologies.

plant biology↗

bioRxiv2023

Genome-enabled analysis of population dynamics and virulence associated loci in the oat crown rust fungus Puccinia coronata f. sp. avenae

Puccinia coronata f. sp. avenae (Pca) is an important fungal pathogen causing crown rust that impacts oat production worldwide. Genetic resistance for crop protection against Pca is often overcome by the rapid virulence evolution of the pathogen. This study investigated the factors shaping adaptive evolution of Pca using pathogen populations from distinct geographic regions within the USA and South Africa (SA). Phenotypic and genome-wide sequencing data of these diverse Pca collections, including 217 isolates, uncovered phylogenetic relationships and established distinct genetic composition between populations from northern and southern regions from the USA and SA. The population dynamics of Pca involve a bidirectional movement of inoculum between northern and southern regions of the USA and contributions from clonality and sexuality. The population from SA is solely clonal. A genome-wide association study (GWAS) employing a haplotype-resolved Pca reference genome was used to define eleven virulence-associated loci corresponding to twenty-five oat differential lines. These regions were screened to determine candidate Avr effector genes. Overall, the GWAS results allowed us to identify the underlying genetic traits controlling pathogen recognition in an oat differential set used in the USA to assign pathogen races (pathotypes). Key GWAS findings support complex genetic interactions in several oat lines suggesting allelism among resistance genes or redundancy of genes included in the differential set, multiple resistance genes recognising genetically linked Avr effector genes, or potentially epistatic relationships. A careful evaluation of the composition of the oat differential set accompanied by the development or implementation of molecular markers is recommended.

genomics↗

bioRxiv2023

Plant TIR domains physically interact with EDS1 family proteins to propagate immune signalling.

Plant Toll/interleukin-1 receptor/resistance protein (TIR) type nucleotide-binding and leucine-rich repeat immune receptors (NLRs) require Enhanced Disease Susceptibility 1 (EDS1) family proteins and the helper NLRs NRG1 and ADR1 for immune activation. TIR signalling domains possess NADase activity, producing NAM and v-cADPR from NAD+ in vitro. However, after TIR activation different small non-cyclic signalling molecules have been detected bound to EDS1/SAG101 and EDS1/PAD4 heterodimers. These molecules have not been detected in in vitro assays or as free molecules in planta and it is not clear how they are delivered to the EDS1 complexes. Here we investigate physical and functional interactions between TIR signalling domains, EDS1 family proteins and helper NLRs to clarify these signalling transduction pathways. We show that the NbEDS1-NbSAG101b-NbNRG1 signalling pathway in N. benthamiana is necessary and sufficient for cell death signalling induced by six different TIR-containing NLRs from a range of plant species, suggesting this module is likely a universal requirement for TIR-NLR mediated cell death in N. benthamiana. We also find that TIR domains physically interact with NbEDS1, NbPAD4 and NbSAG101 in planta, independently of each other. We also find evidence for direct interaction of NbNRG1 with NbSAG101b via its C-terminal EP domain, but not with other EDS1 family members. These data suggest a model in which physical interaction between activated TIRs and EDS1 signalling complexes facilitates efficient transfer of low abundance products of TIR catalytic activity directly to EDS1 heterocomplexes. The interaction could also alter TIR catalytic activity to favor production of the ligands recognised by EDS1/SAG101 and EDS1/PAD4.

plant biology↗

bioRxiv2023

Pooled effector library screening in protoplasts rapidly identifies novel Avr genes

Crop breeding for durable disease resistance is challenging due to the rapid evolution of pathogen virulence. While progress in resistance (R) gene cloning and stacking has accelerated in recent years 1-3, the identification of corresponding avirulence (Avr) genes in many pathogens is hampered by the lack of high-throughput screening options. To address this technology gap, we developed a platform for pooled library screening in plant protoplasts for rapid identification of interacting R/Avr pairs. We validated this platform by isolating known and novel Avr genes from wheat stem rust (Puccinia graminis f. sp. tritici) by screening a designed library of putative effectors against individual R genes. Rapid Avr gene identification provides molecular tools to understand and track pathogen virulence evolution by genotype surveillance and optimise R gene deployment and stacking strategies. This screening platform is broadly applicable to many crop pathogens, whilst also adaptable for screening genes involved in other protoplast-selectable traits.

plant biology↗

bioRxiv2022

The largest haplotype-phased and chromosome-scale genome assembly of the fungal pathogen, Puccinia polysora, by HiFi reads and Hi-C data

Rust fungi are characterized by large genomes with high repeat content and have two haploid nuclei in most life stages, which makes achieving high-quality genome assemblies challenging. Here, we described a pipeline using HiFi reads and Hi-C data to assemble a gigabase-sized fungal pathogen, Puccinia polysora f.sp. zeae, to haplotype-phased and chromosome-scale. The final assembled genome is 1.71 Gbp, with ~850 Mbp and 18 chromosomes in each haplotype, being currently one of the two giga-scale fungi assembled to chromosome level. Transcript-based annotation identified 47,512 genes for dikaryotic genome with a similar number for each haplotype. A high level of interhaplotype variation was found with 10% haplotype-specific BUSCO genes, 5.8 SNPs/kbp and structural variation accounting for 3% of the genome size. The P. polysora genome displayed over 85% repeat contents, with genome-size expansion and copy number increasing of species-specific orthogroups. Interestingly, these features did not affect overall synteny with other Puccinia species having smaller genomes. Fine-time-point transcriptomics revealed seven clusters of co-expressed secreted proteins that are conserved between two haplotypes. The fact that candidate effectors interspersed with all genes indicated the absence of a "two-speed genome" evolution in P. polysora. Genome resequencing of 79 additional isolates revealed a clonal population structure of P. polysora in China with low geographic differentiation. Nevertheless, a minor population differentiated from the major population by having mutations on secreted proteins including AvrRppC, indicating the ongoing virulence to evade recognition by RppC, a major resistance gene in Chinese corn cultivars. The high-quality assembly provides valuable genomic resources for future studies on disease management and the evolution of P. polysora.

genomics↗

bioRxiv2022

A chromosome-level, fully phased genome assembly of the oat crown rust fungus Puccinia coronata f. sp. avenae: a resource to enable comparative genomics in the cereal rusts

Advances in sequencing technologies as well as development of algorithms and workflows have made it possible to generate fully phased genome references for organisms with non-haploid genomes such as dikaryotic rust fungi. To enable discovery of pathogen effectors and further our understanding of virulence evolution, we generated a chromosome-scale assembly for each of the two nuclear genomes of the oat crown rust pathogen, Puccinia coronata f. sp. avenae (Pca). This resource complements two previous released partially phased genome references of Pca, which display virulence traits absent in the isolate of historic race 203 (isolate Pca203) which was selected for this genome project. A fully phased, chromosome-level reference for Pca203 was generated using PacBio reads and Hi-C data and a recently developed pipeline named NuclearPhaser for phase assignment of contigs and phase switch correction. With 18 chromosomes in each haplotype and a total size of 208.10 Mbp, Pca203 has the same number of chromosomes as other cereal rust fungi such as Puccinia graminis f. sp. tritici and Puccinia triticina, the causal agents of wheat stem rust and wheat leaf rust, respectively. The Pca203 reference marks the third fully-phased chromosome-level assembly of a cereal rust to date. Here, we demonstrate that the chromosomes of these three Puccinia species are syntenous and that chromosomal size variations are primarily due to differences in repeat element content.

genomics↗

bioRxiv2018

Improved prediction of fungal effector proteins from secretomes with EffectorP 2.0

Plant-pathogenic fungi secrete effector proteins to facilitate infection. We describe extensive improvements to EffectorP, the first machine learning classifier for fungal effector prediction. EffectorP 2.0 is now trained on a larger set of effectors and utilizes a different approach based on an ensemble of classifiers trained on different subsets of negative data, offering different views on classification. EffectorP 2.0 achieves accuracy of 89%, compared to 82% for EffectorP 1.0 and 59.8% for a small size classifier. Important features for effector prediction appear to be protein size, protein net charge as well as the amino acids serine and cysteine. EffectorP 2.0 decreases the number of predicted effectors in secretomes of fungal plant symbionts and saprophytes by 40% when compared to EffectorP 1.0. However, EffectorP 1.0 retains value and combining EffectorP 1.0 and 2.0 results in a stringent classifier with low false positive rate of 9%. EffectorP 2.0 predicts significant enrichments of effectors in 12 out of 13 sets of infection-induced proteins from diverse fungal pathogens, whereas a small cysteine-rich classifier detects enrichment only in 7 out of 13. EffectorP 2.0 will fast-track prioritization of high-confidence effector candidates for functional validation and aid in improving our understanding of effector biology. EffectorP 2.0 is available at http://effectorp.csiro.au.

bioinformatics↗

bioRxiv2017

ApoplastP: prediction of effectors and plant proteins in the apoplast using machine learning

The plant apoplast is integral to intercellular signalling, transport and plant-pathogen interactions. Plant pathogens deliver effectors both into the apoplast and inside host cells, but no computational method currently exists to discriminate between these localizations. We present ApoplastP, the first method for predicting if an effector or plant protein localizes to the apoplast. ApoplastP uncovers features for apoplastic localization common to both effectors and plant proteins, namely an enrichment in small amino acids and cysteines as well as depletion in glutamic acid. ApoplastP predicts apoplastic localization in effectors with sensitivity of 75% and false positive rate of 5%, improving accuracy of cysteine-rich classifiers by over 13%. ApoplastP does not depend on the presence of a signal peptide and correctly predicts the localization of unconventionally secreted plant and effector proteins. The secretomes of fungal saprophytes, necrotrophic pathogens and extracellular pathogens are enriched for predicted apoplastic proteins. Rust pathogen secretomes have the lowest percentage of apoplastic proteins, but these are highly enriched for predicted effectors. ApoplastP pioneers apoplastic localization prediction using machine learning. It will facilitate functional studies and will be valuable for predicting if an effector localizes to the apoplast or if it enters plant cells. ApoplastP is available at http://apoplastp.csiro.au.

bioinformatics↗

bioRxiv2017

De novo assembly and phasing of dikaryotic genomes from two isolates of Puccinia coronata f. sp. avenae, the causal agent of oat crown rust

Oat crown rust, caused by the fungus Puccinia coronata f. sp. avenae (Pca), is a devastating disease that impacts worldwide oat production. For much of its life cycle, Pca is dikaryotic, with two separate haploid nuclei that may vary in virulence genotype, highlighting the importance of understanding haplotype diversity in this species. We generated highly contiguous de novo genome assemblies of two Pca isolates, 12SD80 and 12NC29, from long-read sequences. In total, we assembled 603 primary contigs for a total assembly length of 99.16 Mbp for 12SD80 and 777 primary contigs with a total length of 105.25 Mbp for 12NC29, and approximately 52% of each genome was assembled into alternate haplotypes. This revealed structural variation between haplotypes in each isolate equivalent to more than 2% of the genome size, in addition to about 260,000 and 380,000 heterozygous single-nucleotide polymorphisms in 12SD80 and 12NC29, respectively. Transcript-based annotation identified 26,796 and 28,801 coding sequences for isolates 12SD80 and 12NC29, respectively, including about 7,000 allele pairs in haplotype-phased regions. Furthermore, expression profiling revealed clusters of co-expressed secreted effector candidates, and the majority of orthologous effectors between isolates showed conservation of expression patterns. However, a small subset of orthologs showed divergence in expression, which may contribute to differences in virulence between 12SD80 and 12NC29. This study provides the first haplotype-phased reference genome for a dikaryotic rust fungus as a foundation for future studies into virulence mechanisms in Pca.\n\nImportanceDisease management strategies for oat crown rust are challenged by the rapid evolution of Puccinia coronata f. sp. avenae (Pca), which renders resistance genes in oat varieties ineffective. Despite the economic importance of understanding Pca, resources to study the molecular mechanisms underpinning pathogenicity and emergence of new virulence traits are lacking. Such limitations are partly due to the obligate biotrophic lifestyle of Pca as well as the dikaryotic nature of the genome, features that are also shared with other important rust pathogens. This study reports the first release of a haplotype-phased genome assembly for a dikaryotic fungal species and demonstrates the amenability of using emerging technologies to investigate genetic diversity in populations of Pca.

genomics↗