Biology subjects

Idnurm, A.

Publications and source records attributed to Idnurm, A..

10 recordsLinked to original sources

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↗

bioRxiv2022

pGG-PIP: A GreenGate (GG) entry vector collection with Plant Immune system Promoters (PIP)

The regulatory sequences controlling the expression of a gene (i.e., the promoter) are essential to properly understand a genes function. From their use in mutant complementation assays, to studying their responsiveness to different stimuli via transcriptional reporter lines or using them as proxy for the activation of certain pathways, assays using promoter sequences are valuable tools for insight into the genetic architecture underlying plant life. The GreenGate (GG) system is a plant-specific variant of the Golden Gate assembly method, a modular cloning system that allows the hierarchical assembly of individual donor DNA fragments into one expression clone via a single reaction step. Here, we present a collection of 75 GG entry vectors carrying putative regulatory sequences for Arabidopsis thaliana genes involved in many different pathways of the plant immune system, designated Plant Immune system Promoters (PIP). This pGG-PIP entry vector set enables the rapid assembly of expression vectors to be used for transcriptional reporters of plant immune system components, mutant complementation assays when coupled with coding sequences, mis-expression experiments for genes of interest, or the targeted use of CRISPR/Cas9 genome editing. We used pGG-PIP vectors to create fluorescent transcriptional reporters in A. thaliana and demonstrated the potential of these reporters to image the responsiveness of specific plant immunity genes to infection and colonization by the fungal pathogen Fusarium oxysporum. Using the PLANT ELICITOR PEPTIDE (PEP) pathway as an example, we show that several components of this pathway are locally activated in response to colonization by the fungus.

plant biology↗

bioRxiv2022

Spatially distinct phytohormone responses of individual Arabidopsis thaliana root cells to infection and colonization by Fusarium oxysporum

Jasmonic acid (JA), ethylene (ET) and salicylic acid (SA) are the three major phytohormones coordinating a plants defense response to pathogenic attack. While JA and ET are assumed to primarily control the defense against necrotrophic pathogens, SA-induced defense responses target mainly biotrophic microbes, and can include drastic measures such as programmed cell death as part of the plants hypersensitive response (HR). Fusarium oxysporum is a hemibiotrophic fungal pathogen of several plant species, including many important food crops, and the model plant species Arabidopsis thaliana. Colonization of the plants root vascular tissue by the fungus eventually results in wilting and plant death. A general role for JA, ET and SA in combating infection and colonization of the plant by F. oxysporum has been demonstrated, but their distinct roles and modes of action have so far not been described. Here, using high resolution microscopy with fluorescent marker lines of A. thaliana roots infected with F. oxysporum we show that SA acts spatially separate from JA, in a distinct set of root cells immediately neighboring the fungal colonization site. There, SA induces HR to stop the spread of colonization. JA acts in a different, but also clearly defined set of cells, slightly removed from the colonization site, where it initiates a defense response to actively resist the invader. ET is activated in a stretch of cells that covers both, the cells with activated SA and JA signaling, and may be involved in creating these two distinct zones. These results show how the three phytohormones act together, but spatially and functionally separate from each other, to fight this hemibiotrophic pathogen. Such a high-resolution analysis to resolve the plants immune response to pathogenic infection on an individual cell level and in intact tissue has so far been lacking. Graphical AbstractO_LIColonization of the A. thaliana root tip by F. oxysporum strain Fo5176 leads to immediate cell death of the colonized and surrounding tissue. C_LIO_LIAs the colonization front progresses through the vasculature, the cell death front moves along with it through not only the vasculature, but also the surrounding tissues. C_LIO_LIWRKY70 positively regulates salicylic acid (SA) biosynthesis in cells immediately adjacent to the colonized tissue, inducing a hypersensitive response (HR), thereby killing off the cells deemed lost to the intruder, establishing the cell death front. C_LIO_LISlightly further removed from the HR zone, WRKY11 induces jasmonate (JA) biosynthesis in cells of the vasculature to launch a defense response aimed at actively repelling the fungus. C_LI O_FIG O_LINKSMALLFIG WIDTH=122 HEIGHT=200 SRC="FIGDIR/small/521292v2_ufig1.gif" ALT="Figure 1"> View larger version (50K): org.highwire.dtl.DTLVardef@1bd280forg.highwire.dtl.DTLVardef@191052aorg.highwire.dtl.DTLVardef@1ea43acorg.highwire.dtl.DTLVardef@100ecdb_HPS_FORMAT_FIGEXP M_FIG C_FIG

plant biology↗

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↗

bioRxiv2022

Effects of chitin and chitosan on root growth, biochemical defense response and exudate proteome of Cannabis sativa

Fungal pathogens pose a major threat to Cannabis sativa production, requiring safe and effective management procedures to control disease. Chitin and chitosan are natural molecules that elicit plant defense responses. Investigation of their effects on C. sativa will advance understanding of plant responses towards elicitors and provide a potential pathway to enhance plant resistance against diseases. Plants were grown in the in vitro Root-TRAPR system and treated with colloidal chitin and chitosan. Plant morphology was monitored, then plant tissues and exudates were collected for enzymatic activity assays, phytohormone quantification, qPCR analysis and proteomics profiling. Chitosan treatments showed increased total chitinase activity and expression of pathogenesis-related (PR) genes by 3-5 times in the root tissues. In the exudates, total peroxidase and chitinase activities and levels of defense proteins such as PR protein 1 and endochitinase 2 were increased. Shoot development was unaffected, but root development was inhibited after chitosan exposure. No significant effects on plant defense were observed upon chitin treatment. These results indicate that colloidal chitosan significantly promoted production and secretion of plant defense proteins in C. sativa root system and could be used as a potential elicitor, particularly in hydroponic scenarios to manage crop diseases. HighlightChitosan induces defense protein productions and secretions in the root tissues and exudates of C. sativa, offering a potential pathway to enhance plant resistance against fungal attack.

plant biology↗

bioRxiv2022

Starships are active eukaryotic transposable elements mobilized by a new family of tyrosine recombinases

Transposable elements in eukaryotic organisms have historically been considered selfish, at best conferring indirect benefits to their host organisms. The Starships are a recently discovered feature in fungal genomes that are predicted to confer beneficial traits to their hosts and also have hallmarks of being transposable elements. Here, we provide experimental evidence that Starships are indeed autonomous transposons, using the model Paecilomyces variotii, and identify the HhpA Captain tyrosine recombinase as essential for their mobilization into genomic sites with a specific target site consensus sequence. Furthermore, we identify multiple recent horizontal gene transfer of Starships, implying that they frequency jump between species. Fungal genomes have mechanisms to defend against mobile elements, which are frequently detrimental to the host. We discover that Starships are also vulnerable to repeat-induced point mutation defence, thereby having implications on the evolutionary stability of such elements.

microbiology↗

bioRxiv2020

A new family of structurally conserved fungal effectors displays epistatic interactions with plant resistance proteins

Recognition of a pathogen avirulence (AVR) effector protein by a cognate plant resistance (R) protein triggers a set of immune responses that render the plant resistant. Pathogens can escape this so-called Effector-Triggered Immunity (ETI) by different mechanisms including the deletion or loss-of-function mutation of the AVR gene, the incorporation of point mutations that allow recognition to be evaded while maintaining virulence function, and the acquisition of new effectors that suppress AVR recognition. The Dothideomycete Leptosphaeria maculans, causal agent of oilseed rape stem canker, is one of the few fungal pathogens where suppression of ETI by an AVR effector has been demonstrated. Indeed, AvrLm4-7 suppresses Rlm3- and Rlm9- mediated resistance triggered by AvrLm3 and AvrLm5-9, respectively. The presence of AvrLm4-7 does not impede AvrLm3 and AvrLm5-9 expression, and the three AVR proteins do not appear to physically interact. To decipher the epistatic interaction between these L. maculans AVR effectors, we determined the crystal structure of AvrLm5-9 and obtained a 3D model of AvrLm3, based on the crystal structure of Ecp11-1, a homologous AVR effector candidate from Fulvia fulva. Despite a lack of sequence similarity, AvrLm5-9 and AvrLm3 are structural analogues of AvrLm4-7 (structure previously characterized). Structure-informed sequence database searches identified a larger number of putative structural analogues among L. maculans effector candidates, including the AVR effector AvrLmS-Lep2, all produced during the early stages of oilseed rape infection, as well as among effector candidates from other phytopathogenic fungi. These structural analogues are named LARS (for Leptosphaeria AviRulence and Suppressing) effectors. Remarkably, transformants of L. maculans expressing one of these structural analogues, Ecp11-1, triggered oilseed rape immunity in several genotypes carrying Rlm3. Furthermore, this resistance could be suppressed by AvrLm4-7. These results suggest that Ecp11-1 shares a common activity with AvrLm3 within the host plant which is detected by Rlm3, or that the Ecp11-1 structure is sufficiently close to that of AvrLm3 to be recognized by Rlm3. Author summaryAn efficient strategy to control fungal diseases in the field is genetic control using resistant crop cultivars. Crop resistance mainly relies on gene-for-gene relationships between plant resistance (R) genes and pathogen avirulence (AVR) genes, as defined by Flor in the 1940s. However, such gene-for-gene relationships can increase in complexity over the course of plant-pathogen co-evolution. Resistance against the plant-pathogenic fungus Leptosphaeria maculans by Brassica napus and other Brassica species relies on the recognition of effector (AVR) proteins by R proteins; however, L. maculans produces an effector that suppresses a subset of these specific resistances. Using a protein structure approach, we revealed structural analogy between several of the resistance-triggering effectors, the resistance-suppressing effector, and effectors from other plant-pathogenic species in the Dothideomycetes and Sordariomycetes classes, defining a new family of effectors called LARS. Notably, cross-species expression of one LARS effector from Fulvia fulva, a pathogen of tomato, in L. maculans resulted in recognition by several resistant cultivars of oilseed rape. These results highlight the need to integrate knowledge on effector structures to improve resistance management and to develop broad-spectrum resistances for multi-pathogen control of diseases.

plant biology↗

bioRxiv2020

Eukaryotic transposable elements as "cargo carriers": the forging of metal resistance in the fungus Paecilomyces variotii

The horizontal transfer of large gene clusters by mobile elements is a key driver of prokaryotic adaptation in response to environmental stresses. Eukaryotic microbes face similar environmental stresses yet a parallel role for mobile elements has not yet been established. A stress faced by all microorganisms is the prevalence of toxic metals in their environment. In fungi, identified mechanisms for protection against metals generally rely on genes that are dispersed within an organisms genome. Here we have discovered a large ([~]85 kb) region that confers resistance to several metals in the genomes of some, but not all, strains of a fungus, Paecilomyces variotii. We name this region HEPHAESTUS (H{phi}) and present evidence that this region is mobile within the P. variotii genome with features highly characteristic of a transposable element. While large gene clusters including those for the synthesis of secondary metabolites have been widely reported in fungi, these are not mobile within fungal genomes. HEPHAESTUS contains the greatest complement of host-beneficial genes carried by a transposable element in eukaryotes. This suggests that eukaryotic transposable elements might play a role analogous to their bacterial counterparts in the horizontal transfer of large regions of host-beneficial DNA. Genes within HEPHAESTUS responsible for individual metal resistances include those encoding a P-type ATPase transporter, PcaA, required for cadmium and lead resistance, a transporter, ZrcA, providing resistance to zinc, and a multicopper oxidase, McoA, conferring resistance to copper. Additionally, a subregion of H{phi} conferring resistance to arsenate was identified. The presence of a strikingly similar cluster in the genome of another fungus, Penicillium fuscoglaucum, suggests that HEPHAESTUS arrived in P. variotii via horizontal gene transfer.

molecular biology↗

bioRxiv2019

The fungal gene cluster for biosynthesis of the antibacterial agent viriditoxin

BackgroundViriditoxin is one of the classical secondary metabolites produced by fungi and that has antibacterial and other activities; however, the mechanism of its biosynthesis has remained unknown.\n\nResultsHere, a gene cluster responsible for its synthesis was identified, using bioinformatic approaches from two species that produce viriditoxin and then through gene disruption and metabolite profiling. All eight genes in the cluster in Paecilomyces variotii were mutated, revealing their roles in the synthesis of this molecule and establishing its biosynthetic pathway which includes an interesting Baeyer-Villiger monooxygenase catalyzed reaction. Additionally, a candidate catalytically-inactive hydrolase was identified as being required for the stereoselective biosynthesis of (M)-viriditoxin. The localization of two proteins were assessed by fusing these proteins to green fluorescent protein, revealing that at least two intracellular structures are involved in the compartmentalization of the synthesis steps of this metabolite.\n\nConclusionsThe full pathway for synthesis of viriditoxin was established by a combination of genomics, bioinformatics, gene disruption and chemical analysis processes. Hence, this work reveals the basis for the synthesis of an understudied class of fungal secondary metabolites and provides a new model species for understanding the synthesis of biaryl compounds with a chiral axis.

microbiology↗