Biology subjects

Vaghefi, N.

Publications and source records attributed to Vaghefi, N..

10 recordsLinked to original sources

bioRxiv2026

Genetic characterisation and aggressiveness of the Fusarium oxysporum Species Complex in tomato plants and irrigation water from Australian processing fields

Fusarium oxysporum Species Complex (FOSC) includes important soilborne pathogens of a range of crops worldwide. In Victoria (VIC) and New South Wales (NSW), Australia, FOSC has been identified as the cause of stunting and poor growth in processing tomato fields, resulting in significant yield losses. A total of 40 FOSC isolates were obtained from symptomatic tomato plants, irrigation water samples and culture collections, which were confirmed to be FOSC by multi-gene phylogenetic analyses based on four genomic loci: beta tubulin, calmodulin, the second largest subunit of nuclear RNA polymerase II, and translation elongation factor one-alpha. There was a high level of genetic diversity among isolates, with multiple phylogenetic lineages detected. Glasshouse bioassays demonstrated that all isolates were pathogenic to processing tomato, resulting in significant reductions in plant growth, with above ground height reduced by 11 to 26% and root dry weight by 44 to 83% compared with the control (p < 0.05). Although aggressiveness varied among isolates, growth reduction occurred irrespective of their phylogenetic placement. Moreover, the shared genetic background of isolates from irrigation water and plant samples highlights the role of irrigation water as a potential source of inoculum in Fusarium epidemics, underscoring its significance for disease management in Australian processing tomato systems.

plant biology↗

bioRxiv2026

Elucidating pathogen interactions in Tanacetum cinerariifolium (pyrethrum) using fluorescently labelled Didymella tanaceti and Stagonosporopsis tanaceti

Australia is the largest producer of Pyrethrum (Tanacetum cinerariifolium) globally. Amongst the constraints on production are the fungal pathogens Didymella tanaceti and Stagonosporopsis tanaceti, which pose a significant threat to the industry, causing substantial yield losses. While the infection biology of S. tanaceti is well characterised, knowledge of D. tanaceti and its potential interaction with S. tanaceti on plants remains limited, hindering disease management. We developed fluorescently labelled strains of both pathogens via Agrobacterium tumefaciens-mediated transformation (ATMT). Binary vectors carrying the mNeonGreen or tdTomato fluorescent protein genes were introduced into D. tanaceti and S. tanaceti, respectively, and expression of the fluorescent proteins was confirmed by microscopy. Genome sequencing revealed single-copy T-DNA insertions in all transformants, with minor genomic rearrangements at insertion sites. Detached leaf assays demonstrated that transformed strains retained pathogenicity, producing disease symptoms indistinguishable from those of the wild type. These fluorescently labelled variants enabled detailed visualisation of D. tanaceti infection biology and its interactions with S. tanaceti, including co-infection dynamics. Co-infection assays using fluorescent strains further facilitated simultaneous visualisation and differentiation of both pathogens within host tissues. Importantly, these tools also allowed the first description of the early stages of infection by D. tanaceti in pyrethrum leaves. This study represents the first successful transformation of D. tanaceti and S. tanaceti, providing valuable resources to investigate their infection processes.

pathology↗

bioRxiv2025

Multi-locus phylogenetic network analysis of Ampelomyces mycoparasites isolated from diverse powdery mildews in Australia and the generation of two de novo genome assemblies

The interactions between powdery mildews (Ascomycota, Erysiphaceae), obligate biotrophic pathogens of many plants, and pycnidial fungi belonging to the genus Ampelomyces, are classic examples of specific mycoparasitic relationships. These interactions are common and finely tuned tritrophic relationships amongst host plants, powdery mildews, and Ampelomyces mycoparasites wherever these organisms co-occur in the field. Selected Ampelomyces strains have already been developed as biocontrol agents of powdery mildew infections of some crops. In Australia, their study has received little attention so far. Only a single Ampelomyces strain, included in a whole-genome sequencing (WGS) project, was known from this continent. Here, we report the isolation of 20 more Ampelomyces strains from eight powdery mildew species in Australia. Multi-locus phylogenetic network analyses of all the 21 Australian Ampelomyces strains carried out in combination with 32 reference strains from overseas revealed that the Australian strains belonged to four molecular taxonomic units (MOTUs). All those MOTUs were delimited earlier based on Ampelomyces strains isolated in Europe, North America, and elsewhere. Analyses indicated possible recombination events amongst phylogenetically different Ampelomyces strains. Based on the phylogenetic analyses, two Australian strains belonging to different MOTUs were selected for WGS. Long-read (minION) and short-read (Illumina) technologies were used to provide genome assemblies with high completeness. Both assemblies reported here had bipartite structure, i.e., consisted of AT-rich, gene-sparse regions interspersed with GC-balanced, gene-rich regions. These new high-quality assemblies and evidence-based annotations are important resources for future analyses of mycoparasitic interactions to disentangle molecular mechanisms underlying mycoparasitism, possible new biocontrol applications, and natural tritrophic relationships.

microbiology↗

bioRxiv2025

Plasmid copy number variation impacts pathogenicity and quantification of Curtobacterium flaccumfaciens pv. flaccumfaciens infecting mung bean

In bacteria, plasmids can confer the ability to cause disease. Although they can potentially vary in copy number, little has been reported on the dynamics of plasmids in plant pathogenic bacteria. Pathogenicity of the bacterium Curtobacterium flaccumfaciens pv. flaccumfaciens (Cff), which causes foliar disease on leguminous crops, including mung bean (Vigna radiata), has previously been linked to a plasmid. This study explored the variation in plasmid copy number among a genetically diverse collection of 25 Cff isolates using purposely designed quantitative PCR assays for chromosomal and plasmid DNA targets. Pathogenicity and virulence of six Cff isolates, including one plasmid-free isolate, were assessed on the susceptible mung bean cultivar Opal-AU using visual symptoms, trifoliate dry weights and Cff DNA quantities following stem inoculation. Plasmid copy numbers varied significantly (p <0.001) among plasmid-carrying isolates, ranging from 1.37 to 2.74. For subsequent quantification in planta, the single-copy chromosomal PCR target gene, gyrase b, was selected. Mung beans inoculated with plasmid-carrying isolates exhibited significant variation in visual symptoms (p <0.001), while trifoliate dry weights and Cff DNA quantities did not significantly differ. The treatment using a plasmid-free isolate was not significantly different from the negative control for each of the disease traits. Knowledge of plasmid dynamics in Cff populations lays a foundation for improved understanding of the inheritance and impact of plasmid-related traits. The quantification assays will be useful for monitoring Cff populations and the demonstrated variation in pathogenicity and virulence can assist efforts to breed host plant germplasm with reduced susceptibility to Cff.

pathology↗

bioRxiv2024

Clonal expansion from standing genetic variation underpins the evolution of an emerging plant pathogen in Australia

Pathogens can evolve rapidly, leading to the emergence of novel strains that can overcome commercially deployed host plant resistance. Understanding the genetic and phenotypic diversity and population dynamics of plant pathogens is crucial to inform breeding programs targeting resistance. Tan spot, otherwise known as bacterial wilt, caused by Curtobacterium flaccumfaciens pv. flaccumfaciens, is a destructive disease of mungbean in Australia. Since the 1990s, several mungbean cultivars with partial resistance to tan spot have been released; however, cultivars initially rated as moderately resistant were later rated as moderately susceptible to tan spot. This study investigated the genetic and phenotypic diversity and temporal evolutionary dynamics of C. flaccumfaciens pv. flaccumfaciens in Australian mungbean fields. Whole-genome sequencing of 119 isolates collected from mungbean and other legumes (1986-2019) enabled analyses of pathogen evolution in Australia and in a global context. Results revealed that clonal expansion from standing genetic variation, rather than introduction of novel genotypes, has driven the evolution of this pathogen in Australia since the 1980s. Glasshouse trials confirmed the role of a plasmid in pathogenicity of C. flaccumfaciens pv. flaccumfaciens but found no significant differences in aggressiveness between clonal lineages. This study underscores the importance of understanding pathogen diversity and aggressiveness to guide breeding programs for effective and durable disease resistance and highlights the need for further studies on the competitive fitness of isolates and characterisation of private alleles linked to dominant clonal lineages. SignificanceTan spot is an increasingly significant pathogen affecting beans and mungbean worldwide, with a broad host range and quarantine importance in many countries underscoring the need for effective breeding strategies to manage the disease. Our research provides insights into genetic and phenotypic diversity of this important plant pathogen and temporal changes in its population structure in Australia, which are crucial for developing informed breeding strategies and managing the spread and impact of the disease. This work also underscores the critical need for robust, validated screening protocols and selection of aggressive, locally representative isolates for resistance breeding to ensure accurate assessment of resistance in crops for effective and durable resistance.

evolutionary biology↗

bioRxiv2024

Population genomics of Macrophomina spp. reveals cryptic host specialization and evidence for meiotic recombination

Knowledge of the factors structuring populations of pathogenic fungi is fundamental to disease management efforts and basic biology. High-quality short-read sequence data were obtained for 463 Macrophomina spp. isolates collected from 91 host plant species and soil in 23 countries. Analyses revealed high diversity, admixture, and equal mating type ratios suggesting on-going recombination. Although most tested isolates could asymptomatically colonize strawberry, only isolates from a single phylogroup caused disease. In addition to strawberry, evidence for host specialization was discovered for soybean, demonstrating this broad host range pathogen contains phylogroups with cryptic specialization. Geography x isolate genotype associations were weak, suggesting these species were frequently trafficked between regions. Re-analysis using genomic data supported current species boundaries, and new molecular markers were designed to specifically identify each species. Contrary to expectations, M. phaseolina should be considered a species with both specialist and generalist populations for which meiosis can increase genetic diversity.

genomics↗

bioRxiv2024

Population-level whole genome sequencing of Ascochyta rabiei identifies genomic loci associated with isolate aggressiveness

Ascochyta blight caused by the ascomycete Ascochyta rabiei poses a major biotic threat to chickpea (Cicer arietinum) industries worldwide and incurs substantial costs to the Australian multimillion-dollar chickpea industry both in disease control and yield loss. The fungus was introduced to Australia in the 1970s from an unknown source population, and within a few decades, successfully established in all Australian agroecological chickpea growing regions. Although genetically highly clonal, a broad range of phenotypic variation in terms of aggressiveness exists among the Australian A. rabiei isolates. More recently, highly aggressive isolates capable of causing severe disease symptoms on moderate to highly resistant chickpea cultivars have increased in frequency. To identify genetic loci potentially associated with A. rabiei aggressiveness on Australian chickpea cultivars, we performed deep genome sequencing of 230 isolates collected from a range of agroecological chickpea growing regions between 2013 and 2020. Population genetic analyses using genome-wide single nucleotide polymorphism data identified three main clusters of genetically closely related isolates in Australia. Phylogenetic analyses showed that highly aggressive phenotypes developed multiple times independently throughout the phylogeny. Results point to minor contribution of multiple genetic regions and most likely epigenomic variations to aggressiveness of A. rabiei isolates on Australian chickpea cultivars. IMPACT STATEMENTThis research introduces new knowledge on the Australian A. rabiei population structure, molecular pathogenicity drivers and evolution as a clonal pathogen through comprehensive whole-genome sequencing approach. The knowledge generated on the structure and origin of the Australian A. rabiei and existence of only one mating type continues to inform researchers, growers, breeders and the broader industry on the importance for continued tight biosecurity measures and inform development of accurate and informed disease management and resistance breeding strategies. This research provides a rare real-life example to the effect of genetic drift on a clonal pathogen population and the importance of biosecurity to protect introduction from non-endemic isolates through seed importation in the current era of international markets. DATA SUMMARYAn online dataset containing the data and code required to reproduce the results found in this publication have been deposited at Zenodo (DOI 10.5281/zenodo.12575659). Isolate aggressiveness and collection metadata are available in the Ascochyta dashboard at http://bit.ly/asco-dashboard. Raw sequencing data used in this study was deposited to the NCBI Short Read Archive (SRA) and is available through BioProject PRJNA1175002. The authors confirm all supporting data, code and protocols have been provided within the article or through supplementary data files.

genomics↗

bioRxiv2022

Isolation of strains and their genome sequencing to analyze the mating system of Ophiocordyceps robertsii

The fungal genus Ophiocordyceps contains a number of insect pathogens. One of the best known of these is Ophiocordyceps sinensis, which is used in Chinese medicine and its overharvesting threatens sustainability; hence, alternative sources are being sought. Ophiocordyceps robertsii, found in Australia and New Zealand, has been proposed to be a close relative to O. sinensis, but little is known about this species despite being also of historical significance. Here, O. robertsii strains were isolated into culture and high coverage draft genome sequences obtained and analyzed. This species has a large genome expansion, as also occurred in O. sinensis. The mating type locus was characterized, indicating a heterothallic arrangement whereby each strain has an idiomorphic region of two (MAT1-2-1, MAT1-2-2) or three (MAT1-1-1, MAT1-1-2, MAT1-1-3) genes flanked by the conserved APN2 and SLA2 genes. These resources provide a new opportunity for understanding the evolution of the expanded genome in the homothallic species O. sinensis, as well as capabilities to explore the pharmaceutical potential in a species endemic to Australia and New Zealand. One sentence summaryOphiocordyceps robertsii is a close relative of O. sinensis and has a large genome but with a heterothallic mating system.

microbiology↗

bioRxiv2020

Genome-wide association studies reveal the complex genetic architecture of DMI fungicide resistance in Cercospora beticola

Cercospora leaf spot is the most important disease of sugar beet worldwide. The disease is caused by the fungus Cercospora beticola and is managed principally by timely application of fungicides including those of the sterol demethylation inhibitor (DMI) class. However, reliance on DMIs has caused an increase in resistance to this class of fungicides in multiple C. beticola populations. To better understand the genetic and evolutionary basis for resistance in C. beticola, a genome-wide association study (GWAS) and selective sweep analysis were conducted for the first time in this fungal plant pathogen. We performed whole genome resequencing of 190 C. beticola isolates predominantly from North Dakota and Minnesota that were phenotyped for sensitivity to tetraconazole, the most widely used DMI fungicide in this region. GWAS identified mutations in genes associated with DMI fungicide resistance including a Regulator of G-protein Signaling (RGS) protein, an ATP-binding cassette (ABC) pleiotropic drug resistance transporter, a dual-specificity tyrosine phosphorylation-regulated kinase (DYRK), and a gene annotated as a hypothetical protein. A SNP upstream of CbCYP51, the gene encoding the target of DMI fungicides, was also identified via GWAS. Haplotype analysis of CbCYP51 identified a synonymous mutation (E170) in high linkage disequilibrium with the upstream SNP, and multiple non-synonymous mutations (L144F, I387M and Y464S) associated with DMI resistance. Additionally, a putative codon bias effect for the L144F substitution was identified that generated different resistance potentials. We also identified a CbCYP51 paralog in C. beticola, CbCYP51-like, with high protein homology to CYP51C found uniquely in Fusarium species but CbCYP51-like does not appear to influence DMI sensitivity. Genome-wide scans of selection showed that several of the GWAS mutations for fungicide resistance resided in regions that have recently undergone a selective sweep. Using radial plate growth on selected media as a fitness proxy, we did not find a trade-off associated with DMI fungicide resistance suggesting that resistance mutations can persist in C. beticola populations. Taken together, we show that population genomic data from a crop pathogen can allow the identification of mutations conferring fungicide resistance and inform about their origins in the pathogen population.

evolutionary biology↗

bioRxiv2019

Genome mining of the citrus pathogen Elsinoë fawcettii; prediction and prioritisation of candidate effectors, cell wall degrading enzymes and secondary metabolite gene clusters

AbstractElsinoe fawcettii, a necrotrophic fungal pathogen, causes citrus scab on numerous citrus varieties around the world. Known pathotypes of E. fawcettii are based on host range; additionally, cryptic pathotypes have been reported and more novel pathotypes are thought to exist. E. fawcettii produces elsinochrome, a non-host selective toxin which contributes to virulence. However, the mechanisms involved in potential pathogen-host interactions occurring prior to the production of elsinochrome are unknown, yet the host-specificity observed among pathotypes suggests a reliance upon such mechanisms. In this study we have generated a whole genome sequencing project for E. fawcettii, producing an annotated draft assembly 26.01 Mb in size, with 10,080 predicted gene models and low (0.37%) coverage of transposable elements. The assembly showed evidence of AT-rich regions, potentially indicating genomic regions with increased plasticity. Using a variety of computational tools, we mined the E. fawcettii genome for potential virulence genes as candidates for future investigation. A total of 1,280 secreted proteins and 203 candidate effectors were predicted and compared to those of other necrotrophic (Botrytis cinerea, Parastagonospora nodorum, Pyrenophora tritici-repentis, Sclerotinia sclerotiorum and Zymoseptoria tritici), hemibiotrophic (Leptosphaeria maculans, Magnaporthe oryzae, Rhynchosporium commune and Verticillium dahliae) and biotrophic (Ustilago maydis) plant pathogens. Genomic and proteomic features of known fungal effectors were analysed and used to guide the prioritisation of 77 candidate effectors of E. fawcettii. Additionally, 378 carbohydrate-active enzymes were predicted and analysed for likely secretion and sequence similarity with known virulence genes. Furthermore, secondary metabolite prediction indicated nine additional genes potentially involved in the elsinochrome biosynthesis gene cluster than previously described. A further 21 secondary metabolite clusters were predicted, some with similarity to known toxin producing gene clusters. The candidate virulence genes predicted in this study provide a comprehensive resource for future experimental investigation into the pathogenesis of E. fawcettii.

genomics↗