Journal of Fungi

Background: Invasive pulmonary aspergillosis (IPA) is a serious infection in critically ill patients. Galactomannan detection in endotracheal aspirates (ETA) has emerged as a promising non-invasive diagnostic method. This study evaluates the supportive diagnostic value of ETA galactomannan in ICU patients suspected to have IPA. Methods: We conducted a prospective observational cohort study over two years, enrolling 120 patients in the medicine ICU at a tertiary care centre in India (January 2022 to October 2023). Patients aged over 14 years on mechanical ventilation for >48 hours meeting the entry criteria of the BM-AspICU algorithm were included. ETA galactomannan was measured and correlated with IPA classification. Results: Of 120 patients, 37% (n=44) had probable IPA and 63% (n=76) were classified as colonisers or possible IPA. The optimal ETA galactomannan cut-off was 1.097, yielding sensitivity 72.73% (95% CI 57.2 - 85.0%), specificity 84.2% (95% CI 74.4 - 90.7%), PLR 4.86, NLR 0.35, and AUC 0.844 Conclusion: ETA galactomannan supports IPA diagnosis with favourable sensitivity and specificity. However, given the limitations of clinical scoring-based reference standards and the potential plateau in colonizer reduction at higher cut-offs, it should be integrated into a comprehensive diagnostic approach incorporating clinical, radiological, and microbiological criteria.

IntroductionLomentospora prolificans is a pathogenic filamentous fungus that causes disease primarily in people with severely compromised immune systems. It is pan-resistant to antifungal drugs, but the mechanism of its resistance to amphotericin B (AMB) is unknown. ObjectivesWe aimed to investigate the mechanism of resistance to AMB of L. prolificans. MethodsThe AMB susceptibility of L. prolificans protoplasts was measured using broth microdilution. L. prolificans, either intact, homogenized or fractionated was incubated with AMB in broth. The same activity was carried out with Aspergillus fumigatus as a control. This broth was then used to prepare microdilution plates with Saccharomyces cerevisiae to determine the activity of the conditioned AMB. ResultsAMB was 16-fold more effective in inhibiting the growth of L. prolificans protoplasts than conidia, but only two-fold more effective against A. fumigatus protoplasts than conidia. Incubation of L. prolificans hyphae with AMB in media diminished drug activity to a much greater extent than A. fumigatus, with 8-fold greater fungal mass of the latter required to achieve the effect of the former. Homogenization and fractionization of L. prolificans revealed that the factor inhibiting AMB activity was soluble with a mass >100 kda. DNase, trypsin, proteinase K, amyloglucosidase, SDS and 0.22 m had no effect on the AMB resistance factor, while treatment with urea, acetonitrile inactivated it. ConclusionWe report a different mechanism for AMB resistance based on the existence of a substance residing in the L. prolificans cell wall that can eliminate the antifungal activity of AMB.

The pathogenic fungus Trichosporon asahii causes severe mycoses in immunocompromised hosts, such as neutropenic patients. In Cryptococcus neoformans, the unfolded protein response (UPR) sensor Ire1 induces hxl1 mRNA splicing and contributes to stress responses and virulence. The function of Ire1-triggered hxl1 mRNA splicing in stress tolerance and virulence of T. asahii, however, remains unclear. Here, we demonstrated that ire1- and hxl1 gene-deficient T. asahii mutants are sensitive to dithiothreitol (DTT), an inducer of endoplasmic reticulum stress, and exhibit reduced virulence in a silkworm infection model. DTT treatment induced hxl1 mRNA splicing in the wild-type strain, whereas ire1 gene-deficient mutants did not undergo hxl1 mRNA splicing. The ire1 gene-deficient mutants were more sensitive than the parent strain to DTT, H2O2, Congo red, and SDS, and showed reduced virulence in silkworms. Similarly, hxl1 gene-deficient mutants exhibited increased sensitivity to these stressors and reduced virulence. Both the ire1 gene-deficient and hxl1 gene-deficient mutants showed decreased expression of reactive oxygen species-detoxifying related genes CAT2, SOD1, and SOD2, compared with the parent strain. Together, these findings suggest that Ire1-triggered hxl1 mRNA splicing contributes to stress resistance and virulence in T. asahii.

During mycological surveys in Yunnan Province, China, specimens of a fungus producing massive, upright stromata up to 50 cm high and individually 2.2 Kg in weight were sampled. Through an integrative taxonomic approach combining detailed morphology, multilocus phylogeny (ITS, LSU, RPB2, TUB2), and phylogenomic analyses, this fungus is proposed as the new species Dianjunus rex gen. et sp. nov., the type of the new family Dianjunaceae (Xylariales). Phylogenetic analyses robustly place Dianjunaceae as a distinct sister clade to Graphostromataceae. Divergence time estimation dates the origin of this family to the early Paleocene (~65 Mya), coinciding with the post-K-Pg extinction period, when an estimated 75% of all plant and animal species went extinct, and a significant ecological reorganization of life on earth happened. The stromata of D. rex represent the largest fructifications documented within the Ascomycota, significantly expanding the known morphological range of the Xylariales. The study provides a comprehensive description, including a nodulisporium-like anamorph with periconiella-like branching patterns, and discusses the taxon's phylogenetic placement, and distinctive morphology. This discovery highlights the unexplored fungal diversity in East Asian forests.

Purpose: Central nervous system tuberculosis (CNS TB) presenting with vasculitis or arachnoiditis causes significant morbidity and mortality. The purpose of this study was to characterise the clinical spectrum and outcomes of patients diagnosed with TB arachnoiditis and TB-associated CNS vasculitis Methods A retrospective study was conducted between October 2020 and September 2023, screening patients admitted with suspected CNS TB to a tertiary care hospital. Patients diagnosed with proliferative arachnoiditis, and TB-associated CNS vasculitis were recruited. Their clinical details, follow-up records, and outcomes were assessed. Results Among 318 patients admitted with suspected CNS TB, 87 patients had complications, with follow-up data available for 69 patients. Vasculitis, spinal arachnoiditis (SA), and optochiasmatic arachnoiditis (OCA) was diagnosed in 66 (76%), 41 (47%), and 26 (30%) patients respectively. Median duration of follow-up was 490 days. Median mRS at discharge was 4. 18 (69%) OCA patients and 14 (35%) SA patients received pulse methylprednisolone. Intrathecal hyaluronidase was administered in 14 patients and thalidomide was given to 9 patients. 30 (46%) patients with vasculitis were treated with aspirin. 69 patients completed follow-up, 49% died. Among the remaining, 88.6% had improvement with treatment with a median mRS of 2 (1-3). Among patients with OCA, 3(23.1%) showed complete improvement with a median improvement of 3 points on Likert scale. In the SA patients, 19 (55.9%, 34) patients were alive on follow-up, with a median mRS of 1. Aspirin use was not associated with better mRS or survival in patients with vasculitis. A multivariable Cox proportional model showed age at diagnosis to be the only predictor of mortality (HR 1.04, 95% CI (1.01- 1.08), p =0.012). Conclusions TB arachnoiditis and CNS vasculitis are severe complications of CNS TB, and management remains a challenge. The poor therapeutic response to intrathecal hyaluronidase, thalidomide, and aspirin highlights need for further larger prospective trials and search for alternative agents.

Biological control strategies are increasingly being explored as sustainable alternatives for managing rice blast disease caused by Magnaporthe oryzae. In this study, we characterized three Bacillus pumilus isolates (DC01, DC09, and DC13) and evaluated their antifungal and plant-beneficial properties against M. oryzae. Whole genome sequencing revealed multiple biosynthetic gene clusters associated with the production of antimicrobial metabolites. All three isolates inhibited fungal growth in dual-culture assays, whereas heat-stable diffusible antifungal activity was primarily associated with the cell-free supernatants of DC09 and DC13. Exposure to bacterial supernatants disrupted fungal development, inducing abnormal hyphal morphology characterized by bulbous swelling, altered polarity, and increased branching in M. oryzae. Volatile organic compound assays further revealed that the DC isolates suppress fungal growth in the absence of physical contact. The isolates additionally inhibited the growth of other phytopathogenic fungi and selected human bacterial pathogens. All strains exhibited plant growth-promoting traits, including indole-3-acetic acid production and osmotic stress tolerance, whereas DC09 also displayed phosphate-solubilizing activity. Importantly, root inoculation with the DC isolates significantly reduced rice blast disease severity and induced expression of defense-associated genes involved in jasmonic acid/ethylene signaling and immune priming. Collectively, these findings identify the DC isolates, particularly DC09 and DC13, as promising multi-mechanistic biological control agents for sustainable rice blast management.

Nakaseomyces glabratus (Candida glabrata) is a WHO high-priority fungal pathogen associated with fungal antimicrobial resistance (fAMR). Given nosocomial transmission occurs sporadically, resistant strains could be transmitted, a concern for critically ill patients. We conducted a genomic investigation and retrospective observational study of N. glabratus to identify any nosocomial transmission of fAMR and understand resistance mechanisms and clinical and demiological factors among patients at a quaternary hospital in Melbourne, Australia. We selected stored N. glabratus with and without fAMR associated with similar patient clinical characteristics and performed whole genome sequencing. Clinical and epidemiological data were extracted from medical records. Phylogenetic, mutational, copy-number variation (CNV) and mitochondrial genomic analyses were performed, with a focus on the fAMR gene PDR1. Of 54 isolates collected over seven years, 20 (37%) were fluconazole-resistant and four (7%) had elevated flucytosine minimum inhibitory concentrations (MICs) (range 2-32 g/ml). There were no significant clinical differences between patients with and without fluconazole resistance. Most (55%) fluconazole-resistant isolates carried PDR1 mutations. Resistance was distributed throughout the phylogeny suggesting predominantly independent acquisition. However, a cluster of four resistant isolates with the same PDR1 mutation suggested nosocomial transmission. One probable ERG11 gene duplication, and two petite variants with apparent mitochondrial genomic deletions, were seen in association with fluconazole resistance. In this study, we identified a small probable nosocomial fAMR transmission cluster, and novel variants in PDR1, ERG11 and FCY2 associated with fAMR phenotypes. Future study should confirm functional impacts and systematically investigate for nosocomial transmission of resistance, including colonisation states.

BackgroundThe corn smut fungus Ustilago maydis is an important microbial model organism representing a genetically amenable and readily cultivable basidiomycete. Research in this fungus addresses a broad range of fundamental questions and its biotechnological exploitation is on the rise. Although genetic engineering in principle is well established, efficient methodology for synthetic biology approaches such as metabolic engineering or pathway transplantation has remained limited. ResultsHere, we present a comprehensive toolbox for U. maydis based on modular cloning and the characterization of more than 20 promoters. Careful comparative evaluation of insertion loci and terminator as well as reporter effects was conducted and a novel color-based strategy for straightforward genome integration was implemented. Moreover, the cloning and subsequent one-step integration of four transcriptional units into U. maydis was demonstrated by creating a "rainbow" strain producing four fluorescent proteins. ConclusionOverall, this next generation toolkit strongly advances genetic engineering and systems biology approaches in U. maydis, fostering its development into a valuable and competitive fungal chassis and prime model, particularly in applied research.

Rhizosphere microbial communities contribute to the growth and health of their host but may be altered by the incidence of soil-borne pathogens. In avocado, the oomycete Phytophthora cinnamomi, causal agent of Phytophthora root rot (PRR), has been shown to alter rhizosphere bacterial communities, although its effect on fungal communities has seldom been explored. Our objective was thus to determine whether P. cinnamomi induced shifts in diversity, composition and co-occurrence networks of fungal communities in the rhizosphere of avocado trees, and to identify potential antagonists of P. cinnamomi that could be further considered for disease management. Fungal communities associated with the rhizosphere of asymptomatic and PRR-symptomatic avocado trees were studied through ITS metabarcoding. Although -diversity metrics were not significantly different between asymptomatic and PRR-symptomatic trees, differences in {beta}-diversity of rhizosphere fungal communities were detected. Moreover, PRR led to the enrichment of saprotrophic taxa and opportunistic pathogens such as Fusarium, Cladosporium or Plectosphaerella in the avocado rhizosphere, which were possibly attracted by the release of resources from necrosed roots. Co-occurrence network analysis revealed that fungal networks in the rhizosphere of PRR-symptomatic trees were more complex and connected than those from asymptomatic trees, suggesting a response of fungal communities to the disturbance caused by the pathogen. Some connector taxa from the PRR-symptomatic networks (Gibellulopsis, Cladorrhinum or Mycenella) were also identified as members of the P. cinnamomi pathobiome. Their negative correlations with the pathogen indicate they may act as potential antagonists, which calls for further isolation efforts to confirm their biocontrol activity of PRR.

Parastagonospora nodorum (Berk.) Quaedvlieg, Verkley & Crousis, a necrotrophic fungal pathogen, is the causal agent for septoria nodorum blotch, a major constraint on global wheat production. Pathogen-produced necrotrophic effectors (NEs) that interact with host-sensitivity genes in an inverse gene-for-gene manner, collectively leading to effector-triggered susceptibility (ETS). Here, we investigated the transcriptional responses of two Triticum aestivum L. genotypes, Mace and Lancer, following infiltration with a novel NE, SnTox8. A total of 12,679 unique differentially expressed genes in Mace and 149 in Lancer were detected from transcriptomic analysis. In the SnTox8-sensitive cultivar, Mace, numerous defence-related genes were induced, including protein phosphorylation cascades, reactive oxygen species bursts, calcium signalling, phytohormone modulation, and suppression of photosynthesis, consistent with findings from other ETS models, in which necrotrophic fungal pathogens hijack host defence systems to proliferate. The interaction also activated genes involved in signal transduction, metabolism, membrane modification, and molecular transport, reflecting a coordinated host reprogramming that promotes cellular dysfunction and cell death, thereby facilitating necrotrophic pathogenesis. In contrast, Lancer, an SnTox8-insensitive cultivar, exhibited minimal transcriptional changes with no evidence of effector recognition or downstream defence-related activities. Overall, this study exhibited that SnTox8 manipulates kinase-mediated immune signalling and metabolic reprogramming to convert defence activation into host cell death, revealing a mechanistic basis for ETS in wheat. The identified SnTox8-Snn8-triggered processes were confirmed through additional transcriptome analysis of Mace mutants. Outcomes from this study establish a foundation for identifying, functionally characterising and validating the corresponding host susceptibility gene Snn8.

Endophytic biocontrol agents may contribute to grapevine health, but their ability to establish, persist, and move within woody tissues remains poorly understood. In this study, a stem injection method was developed to introduce Pseudomonas chlororaphis M71 and Trichoderma atroviride SC1 into rooted and grafted grapevine plants, and their spatial and temporal colonization patterns were compared with the movement of a dye tracer. The dye tracer moved rapidly through xylem tissues, whereas both microorganisms showed more restricted early distribution. Over time, M71 and SC1 displayed distinct colonization patterns. M71 persisted after injection, but remained localized near the inoculation site, with limited movement toward roots or distal aerial tissues. In grafted plants, M71 recovery depended on the injection site and declined more markedly after rootstock injection than after scion injection. In contrast, SC1 showed broader and more persistent colonization. In rooted cuttings, SC1 was recovered from stem and root tissues up to 56 days post-injection, and in grafted plants it was recovered across the graft union, particularly after scion injection. Microscopy supported internal localization of both microorganisms. GFP-labelled M71 and SC1 hyphae were observed mainly within xylem vessels, and viable microorganisms were recovered from corresponding wood tissues. No contamination was observed in control plants. These results show that beneficial microorganisms can be introduced into grapevine tissues by stem injection and that bacterial and fungal biocontrol agents differ markedly in their internal movement and persistence. IMPORTANCEIntroducing beneficial microorganisms directly into plant tissues could help in establishing protective endophytic populations, but little is known about how such microorganisms move and persist inside grapevine. This study shows that stem injection can deliver Pseudomonas chlororaphis M71 and Trichoderma atroviride SC1 into grapevine tissues without visible phytotoxicity. The two microorganisms followed different colonization patterns. M71 remained mostly localized near the injection site, whereas SC1 spread more broadly and persisted in both rooted and grafted plants. These findings provide a basis for developing targeted endophytic biocontrol strategies in grapevine propagation and early plant establishment. This approach may be particularly relevant for grapevine trunk diseases and other vascular disorders, in which pathogens colonize internal woody tissues.

Due to climate change, extreme weather events such as droughts are becoming more frequent and intense. This has a profound impact on plant performance and ecological interactions, including those involving herbivorous insects. The combined impact of drought stress and insect herbivory on plant metabolism has rarely been studied, particularly in woody plants. In this study, we investigated the influence of varying degrees of drought, both alone and in combination with herbivory by the leaf beetle Chrysomela tremulae, on the morphological and chemical characteristics of black poplar (Populus nigra) trees using a full factorial experimental design. We quantified morphological traits, volatile organic compound (VOC) emissions, phytohormone and amino acid concentrations, and phenolic profiles. Drought conditions increased the concentrations of salicylic acid (SA) and abscisic acid (ABA), while feeding induced ABA and SA. Amino acid profiles shifted significantly under drought conditions, particularly in beetle-infested plants. In contrast, salicinoids, which are the most important phenolic defense compounds in poplars, remained relatively stable. We also observed significant compound-specific effects on both constitutive and herbivore-induced VOC emissions. Our results demonstrate that drought and insect herbivory exert a joint influence on the chemical responses of P. nigra across multiple metabolic pathways. These findings highlight how the interaction between abiotic and biotic stresses can influence the defense chemistry of trees, which will consequently affect ecological interactions in forest ecosystems in the face of climate change.

The contribution of accessory or conditionally dispensable chromosomes to host-specific virulence was first demonstrated in members of the Fusarium solani species complex (FSSC) that are pathogens of garden pea, Pisum sativum L. The phenomenon has since been shown to exist in many fungal plant pathogens, including the closely related F. oxysporum species complex (FOSC). Genome analysis of members of the FSSC and FOSC pathogenic on pea revealed a diverse size range of the accessory genome of these fungi. Despite the ~65 million years of diverging time, regions on a chromosome known to carry host-specific virulence factors for pea, including the cytochrome P450 pisatin demethylase (PDA) and other pea pathogenicity (PEP) genes, were present in all genomes of these pea pathogens. Genes directly involved in virulence on pea - PEP2, PDA, and PEP5- were the most frequently clustered together. Transcriptome analysis of fungal mycelia treated with the pea phytoalexin pisatin, identified 1,155 differentially expressed genes where many were involved in cellular stress responses. As wilt pathogens that invade host xylem, members of the FOSC encode more putative effectors, when compared to those in the FSSC, and several FOSC effectors were identified to confer race specificity. The conservation of part of the accessory genomes across two evolutionarily diverged species complexes suggests a common origin. Horizontal transfer of accessory chromosomes containing genetic loci involved in pathogenesis for garden pea offers a parsimonious explanation of the polyphyletic origin of host specificity.

Accessory genes are thought to contribute to fungal adaptation and pathogenicity by modulating host immunity, while core genes play crucial roles in maintaining fundamental biological processes. Rust fungi (order Pucciniales) are obligate biotrophic plant-pathogens and infect economically relevant crops. Here, we characterize core and accessory gene repertoires across rust fungi, with a particular focus on Phakopsora pachyrhizi, the causal agent of Asian soybean rust. Across Pucciniales genomes, accessory genes represented the largest fraction of gene content (~44.6% on average), whereas core genes accounted for a smaller proportion (~18-35%). Notably, variations in accessory gene content among rust fungi are perhaps attributed to lineage-specific gene expansions and losses. Core gene content was positively correlated with total gene number across Pucciniales genomes, suggesting retention after gene duplication events, consistent with their essential biological functions. Among P. pachyrhizi genes expressed during soybean infection, core effectors were associated with cysteine-rich proteins, pectin-degrading enzymes, and SPFH/Band 7 family, while accessory effectors included phosphatidylethanolamine-binding proteins, trehalose phosphatases, and CFEM domain-containing proteins. The in-plant induced core and accessory genes in P. pachyrhizi also comprised multiple families of CAZymes (GH5/GH7 cellulases, CE5 cutinases, CE8 pectinesterases, CE4/GH18 chitin-modifying enzymes); proteases (aspartyl proteases, serine carboxypeptidases, alpha/beta hydrolases); transporters (amino acid permeases, ferric reductase-like transmembrane proteins, and OPT oligopeptide transporter), and transcription factors (bZIP, GATA zinc finger, STE-like, and homeobox KN). Our study highlights that core and accessory gene families have shaped P. pachyrhizi-soybean interactions, identifying promising targets for functional studies aimed at elucidating host-adaptation mechanisms in rust fungi.

The Eurasian spruce bark beetle Ips typographus, a major forest pest on Norway spruce (Picea abies), forms intimate associations with several types of microbial symbionts. While previous research has focused primarily on filamentous fungi, yeasts have remained largely unexplored. Here, we show that yeasts associated with I. typographus may contribute to host tree colonization by providing defensive benefits. Dominant yeasts (Yamadazyma, Kuraishia, Nakazawaea, and Wickerhamomyces), which are phylogenetically related to other insect-associated Saccharomycotina, significantly attract adult beetles. Moreover, several yeasts inhibit the growth of the pathogenic fungus Trichoderma harzianum in vitro, and beetle eggs benefit from the presence of Kuraishia capsulata by reduced fungal infection under semi-natural conditions. Strikingly, these effects are mediated by the yeasts' transformation of the tree's defensive stilbene glycosides into antimicrobial aglycones and phenolic acids that accumulate in beetle galleries. These findings reveal a previously unrecognized role of symbiotic yeasts in converting spruce defensive stilbene glycosides into antimicrobial aglycones and oxidative cleavage products that accumulate in beetle galleries, and might contribute to the survival of their bark beetle host.

Background: Human activities such as mining generally lead to increased heavy metal concentrations in the environment. While traditional remediation techniques are often costly, the use of fungi as bioremediators, known as mycoremediation, is increasingly gaining attention as a sustainable approach for removal of heavy metals. Here, we evaluated heavy metal levels inside the Kiirunavaara iron ore mine in Northern Sweden and analysed fungal responses to various metal concentrations by comparing growth and metal uptake in mine-derived isolates and closely related control isolates. Results: Sediments inside the mine were enriched in heavy metals compared to those from the outlet of the mine to natural lakes. Six Fusarium isolates were recovered from contaminated mining environments: five isolates from inside the mine were identified as Fusarium oxysporum, and one isolate from the outlet was identified as Fusarium tricinctum. Isolates from the mine and outlet showed overall higher survival and biomass production in presence of copper, iron, and zinc across a range of concentrations (up to 1000 mg/L) compared to control isolates. At the same time, these isolates often exhibited reduced relative metal uptake. As a result, mycoremediation potential, assessed as total uptake in the grown mycelium, was isolate-dependent. Conclusions: Based on these results, we conclude that Fusarium isolates from the Kiirunavaara mine show increased growth in media enriched with heavy metals compared to closely related control isolates. We additionally show that mycoremediation potential is not necessarily associated with environmental origin. Instead, mycoremediation potential should be evaluated on a case-by-case basis for each isolate and based on specific needs for mycoremediation.

The receptor kinase FERONIA (FER) perceives endogenous RAPID ALKALINIZATION FACTOR (RALF) peptides and regulates a plethora of plant physiological processes, including immunity. RALF peptides also bind to LEUCINE-RICH REPEAT EXTENSIN (LRX) proteins as structural components of the cell wall. We recently showed that the FER-RALF-LRX pathway supports colonization by the obligate biotrophic fungal pathogen Erysiphe cruciferarum (Ecr), a member of the powdery mildew species complex that infects Arabidopsis. Genetic disruption of the pathway primarily affects conidiation of the fungus, raising the question of effects on fungal nutrition. To get further insight into the underlying mechanisms, we performed RNA sequencing (RNAseq) to identify differential transcriptional responses of FER-RALF-LRX pathway mutants upon Ecr infection. Surprisingly, our results revealed that pathway disruption has a limited impact on the overall transcriptional changes upon fungal infection. However, consistent with previous reports, FER-RALF-LRX pathway mutants show changes in basal expression of a plethora of genes, mainly associated with cell wall metabolism, jasmonic acid signalling, amino acid biosynthesis and secondary metabolism. Many of these genes are regulated by Ecr infection across genotypes, too. This raises the question whether these are relevant pathway components for powdery mildew host establishment downstream of the FER-RALF-LRX module. In summary, our data reveals new insights into FER-RALF-LRX-dependent responses that may support host susceptibility to biotrophic plant pathogens.

Bacterial leaf blight (BB), caused by Xanthomonas oryzae pv. oryzae (Xoo), causes yield losses exceeding 50% in affected areas, including the Bagre rice plain in Burkina Faso. Genome-edited (GEd) rice lines have been successful in tackling BB. Modifications in the Xoo virulence protein target site upstream of three SWEET susceptibility genes in two elite rice varieties, IR64 and Ciherang-Sub1, have been demonstrated to confer broad-spectrum resistance to Asian and East African Xoo strains. Here, we evaluate the potential of the GEd lines as a solution for BB management in Burkina Faso. We challenged the GEd lines against five locally collected Burkinabe Xoo strains under controlled green-house conditions and assessed their agro-morphological performance under field conditions representative of local agroecological conditions. Greenhouse pathogen assays demonstrated that GEd IR64 and Ciherang-Sub1 lines were resistant to all tested local Xoo strains across three successive generations. We identified TalC as the primary disease-causing effector in the local Xoo populations. Irrigated field trials conducted over two seasons in the Kou Valley, Burkina Faso, revealed absence of agro-morphological penalties in GEd lines compared to their parental wild-type lines. Observed trait variation was attributable to environmental fluctuations rather than genomic modifications. Collectively, our findings demonstrate that genome editing of the rice lines does not impose growth penalties, and support the suitability of GEd IR64 and Ciherang-Sub1 for large-scale adoption in Burkina Faso, pending multi-location validation and introgression into locally adapted varieties.

Light leaf spot (LLS), caused by subcuticular hemibiotrophic ascomycete fungus Pyrenopeziza brassicae, is a major constraint on oilseed rape (Brassica napus) production, yet the genetic and biochemical mechanisms of quantitative disease resistance (QDR) remain poorly defined. Here, disease phenotyping, pathogen quantification, microscopy, gene expression profiling and glucosinolate (GSL) analysis were integrated to dissect resistance mechanisms in B. napus. Disease assays of 19 diverse lines revealed clear contrasts between susceptible and resistant genotypes, with the commercial cultivar Ambassador showing a phenotype inconsistent with the UK Recommended List rating. Microscopy demonstrated that resistance within doubled haploid line Cubs Root does not inhibit spore germination or penetration but restricts hyphal branching and subcuticular colonisation from 4 to 8 days post-inoculation. Expression profiling of seven candidate gene expression markers (GEMs) and pathogenesis-related PR1 showed that cinnamate-4-hydroxylase, phospholipase C4, {beta}-adaptin, universal stress protein and the 40S ribosomal subunit protein S24 were strongly pathogen-induced in resistant lines, whereas a BAHD acyltransferase, a putative susceptibility factor, was induced only in susceptible cultivars. GSL profiling identified negative correlations between disease severity and total GSLs, particularly aliphatic and aromatic GSLs, with 2{square}phenylethyl and 7-methylsulfinyl heptyl GSLs showing the strongest associations with resistance. Together, these results highlight coordinated transcriptional and metabolic responses that limit pathogen proliferation and provide targets for breeding durable LLS resistance in B. napus.

The basidiomycete Ustilago maydis is a well-characterized model organism for studying pathogen-host interactions and of great interest for a broad spectrum of biotechnological applications. We set here to develop light inducible molecular tools to enable dynamic studies on signaling networks and fungi-host communication, and for metabolic engineering approaches. In particular, light-controlled, optogenetic switches provide quantitative, spatio-temporal control capabilities, are minimal invasive and reversible. We engineered two blue light-inducible LOV-domain-based gene expression switches, to up- (Blue-ON) and down-regulate (Blue-OFF) gene expression, and performed a functional characterization in sporidia and hyphae of U. maydis. Profiting from the dynamic control ranges and rapid kinetics, we implemented the optoswitches to control cell morphology by initiating the transition from a haploid sporidial cellular morphotype to filaments upon regulation of the levels of the polarity factor Rac1 and its constitutive active mutant Q61L. In addition to showing how expression level of effectors can be precisely regulated as an approach to understand fungi-plants interaction, we show in two proof-of-principle applications targeted control over U. maydis filamentous fungal invasion of plant tissue and the mechanisms of tumor formation. For this we placed under Blue-ON and Blue-OFF control two U. maydis effectors, See1 (Seedling efficient effector 1) and TIN2 (Tumor inducing 2), and tumor formation was assayed on maize leaves. Taken together, this study established blue-light switches as effective tools to control morphogenesis and pathogenesis in U. maydis.