Biology subjects

Walker, A.-S.

Publications and source records attributed to Walker, A.-S..

14 recordsLinked to original sources

bioRxiv2026

Role of transposons in the specialization of Botrytis cinerea to grapevine: Insights into small RNAs and a new Starship

While transposable elements (TEs) are recognized as major drivers of fungal genome structure, evidence of their direct involvement in the interaction with host plants and the environment is only beginning to emerge. Retrotransposons can generate small RNA (sRNA) that act through cross-kingdom RNA interference, while giant DNA TEs called Starships carry dozens of cargo genes that enrich the accessory gene compartment of fungal genomes. In the polyphagous pathogen Botrytis cinerea, the Vv3 strain and other strains specialized on grapevine display a specific repertoire of TEs, including the retrotransposons BcCopia4, BcGypsy6, and BcGypsy7. This study first explored the putative role of sRNA generated from these retrotransposons in the interaction between the Vv3 strain and its host of origin, grapevine. Putative targets were identified among the host mRNAs, but predicted cleavage sites could not be experimentally validated. Moreover, Dicer mutants unable to produce retrotransposons-derived sRNA remained fully pathogenic on grapevine, indicating that these sRNAs do not act as virulence factors on this host. In parallel, this study provides an updated RNA-seq-based annotation of the accessory genes of the Vv3 strain, which revealed a new 93 kb-Starship harboring 43 cargo genes, some of which are related to arsenic resistance. A formal genetic approach confirmed that this locus confers resistance to this metalloid. This giant TE, named Ariane, was also detected in additional grapevine-specialized strains resistant to arsenic but not in strains isolated from other hosts such as tomato. In conclusion, this study highlights how a Starship giant transposon shaped the accessory genes compartment of the polyphagous fungus B. cinerea and may have contributed to its adaptation to vine cultivation by conferring resistance to arsenic, a compound widely used in vineyards during the last century. IMPACT STATEMENTFungal genomes contain many families of transposons whose functional role in adaptation to the environment and in biotic interactions remained hidden for a long time. In the grey mold fungus Botrytis cinerea, strains specialized on grapevine, such as Vv3, carry a specific repertoire of transposons which provides a valuable opportunity to investigate their role in niche adaptation. In this study, we first investigated retrotransposon-derived small RNA, previously described as effectors capable of manipulating the immunity of the model plant Arabidopsis thaliana. Although in silico analysis of the specific repertoire of small RNAs of the Vv3 strain suggested that some could target the expression of grapevine genes, a genetic approach demonstrated that they do not play a significant role in virulence on this host. In contrast, this study identified a new transposon, named Ariane, that carries 43 cargo genes and confers a selective advantage to the Vv3 strain. Ariane belongs to a family of giant transposons called Starships, recently discovered in fungi and considered to be responsible for horizontal genes transfers between unrelated species. Ariane was detected only in some B. cinerea strains isolated from grapevine, and a genetic cross showed that it provides these strains with the ability to grow in presence of arsenic. Arsenic was used in vineyards until the beginning of the 21st century to control fungal trunk diseases and insect pests. Therefore, Ariane appears to have played an important role in the adaptation of B. cinerea strains to cultivated grapevine. Overall, these results underline the importance of considering Starships when predicting emergence of resistance to antifungal compounds. DATA SUMMARYThe novel data described in this study, i.e., RNA-Seq data and the Starship element are accessible under NCBI GEO accession GSE327899 and at https://doi.org/10.57745/HYWRNM, respectively. All information related to Botrytis cinerea genomes used in this study are centralized and kept up to date at the Bioinfo Bioger genomic web portal: https://bioinfo.bioger.inrae.fr/portal/genome-portal/. Direct links to individual portals are respectively https://bioinfo.bioger.inrae.fr/portal/genome-portal/3/ for B. cinerea Vv3 genome, https://bioinfo.bioger.inrae.fr/portal/genome-portal/2/ for B. cinerea Sl3 genome, and https://bioinfo.bioger.inrae.fr/portal/genome-portal/4/ for B. cinerea populations isolated on tomato or grapevine. Each portal provides: (i) a centralized access to public genomic resources, including the genome, transposon, and RNA repositories; (ii) a data browser to download the genomic files; (iii) a genome browser that enables visualization of features within their genomic context, along with associated expression data. As a summary, the prior main public B. cinerea genomic accessions and resources used in this study are: GCA_039644125 for VV3 genome, GCA_022560135 for Sl3 genome, GCA_000143535 for B05.10 genome, PRJNA624742 for populations, https://doi.org/10.57745/HYWRNM for transposons, and GSE181592 for small RNAs. Furthermore, table S1 summarizes the list and characteristics of the 64 B. cinerea genomes publicly available to date. The genomic data for Vitis vinifera genome PN40024.v4 used in this study are available at: https://integrape.eu/resources/genes-genomes/genome-accessions/.

genomics↗

bioRxiv2023

Plasticity of the MFS1 promotor is not the only driver of Multidrug resistance in Zymoseptoria tritici

Multidrug resistance (MDR) in fungal pathogens poses a growing threat to fungicide efficacy and sustainable agriculture. In the wheat pathogen Zymoseptoria tritici, MDR is primarily linked to overexpression of the MFS1 transporter gene, driven by transposable element (TE) insertions in its promoter or their remnants. Here, we provide a comprehensive analysis of MFS1 promoter polymorphism and its phenotypic impact on MDR across 374 field isolates collected in Europe between 2020 and 2021. We identify six novel structural variants derived from diverse TE types, confirming that the MFS1 promoter undergoes recurrent and independent insertion events. While previously characterized inserts consistently confer resistance, other variants show limited or variable phenotypic effects. Genetic crosses and quantitative phenotyping further reveal that MDR behaves as a quantitative trait and genome-wide association study confirmed MFS1 as the major resistance locus but also identified additional candidate genes involved in xenobiotic detoxification and membrane transport, supporting a polygenic basis for MDR. Our findings highlight the interplay between TE-driven structural variation and background polygenic architecture in shaping resistance phenotypes. In parallel, this study highlights the need to improve MDR monitoring beyond single-locus genotyping and re-evaluate current resistance management strategies that may inadvertently select for broad-spectrum resistance.

molecular biology↗

bioRxiv2022

Preventing multiple resistance above all: new insights for managing fungal adaptation

Sustainable crop protection is crucial for food security, but is threatened by the adaptation of diverse, evolving pathogen population. Resistance can be managed by maximizing selection pressure diversity, by dose variation and the spatial and temporal combination of active ingredients. We explored the interplay between operational drivers for maximizing management strategy sustainability relative to the resistance status of fungal populations. We applied an experimental evolution approach to three artificial populations of an economically important wheat pathogen, Zymoseptoria tritici, differing in initial resistance status. We revealed that diversified selection pressure limited the selection of resistance in naive populations and those with low frequencies of single. Increasing the number of modes of action delayed resistance development most effectively -- ahead of increasing the number of fungicides, fungicide choice based on resistance risk and temporal variation in fungicide exposure -- but favored generalism in the evolved populations. However, the prior presence of multiple resistant resistant isolates and their subsequent selection in populations overrode the effects of diversity in management strategies, incidentally invalidating any universal ranking. Initial resistance composition must therefore be considered specifically in sustainable resistance management, to address real-world field situation. Abbreviated summaryExperimental evolution is a relevant tool for exploring the determinants of antifungal adaptation in fungi. Here, using the model species Z. tritici and two fungicidal modes of action associated with contrasting resistance risks, we have demonstrated that initial population composition, and in particular the presence of multiple resistance, represents the main determinant of adaptive trajectories. Originality-Significance StatementPesticides are part of microbe environment in agricultural systems and may select for resistance. This adaptation of pathogens is a burden for plant health. Using an original multicriteria assessment and experimental evolution, we revealed that multi-fungicide use, selecting for multiple resistance, trumped any other driver of selection, invalidating any universal ranking of management strategies, a dispute extensively illustrated in literature and still inconclusive, especially in agriculture. This outcome refocuses the debate on population diversity and evolution rather than on the intrinsic properties of strategies, as mostly acclaimed in literature. From a practical point of view, our results highlight the importance of considering local population composition when recommending spraying programs. This practice is currently not generalized in agriculture but may be timely to effectively delay resistance evolution and reduce pesticide load in agricultural systems, a growing social demand, since resistance monitoring at relatively fine spatial scales and at low frequency may become commonplace in a near future with the rise of new molecular biology technologies

evolutionary biology↗

bioRxiv2022

Botrytis cinerea strains infecting grapevine and tomato display contrasted repertoires of accessory chromosomes, transposons and small RNAs

AO_SCPLOWBSTRACTC_SCPLOWThe fungus Botrytis cinerea is a polyphagous pathogen that encompasses multiple host-specialized lineages. While several secreted proteins, secondary metabolites and retrotransposons-derived small RNAs have been characterized as virulence factors, their roles in host specialization remain unknown. The aim of this study was to identify the genomic correlates of host-specialization in populations of B. cinerea associated with grapevine and tomato. Using PacBio sequencing, we produced complete assemblies of the genomes of strains Sl3 and Vv3 that represent the French populations T and G1 of B. cinerea, specialized on tomato and grapevine, respectively. Both assemblies revealed 16 core chromosomes that were highly syntenic with chromosomes of the reference strain B05.10. The main sources of variation in gene content were the subtelomeric regions and the accessory chromosomes, especially the chromosome BCIN19 of Vv3 that was absent in Sl3 and B05.10. The repertoires and density of transposable elements were clearly different between the genomes of Sl3 and Vv3 with a larger number of subfamilies (26) and a greater genome coverage in Vv3 (7.7%) than in Sl3 (14 subfamilies, 4.5% coverage). An Helitron-like element was found in almost all subtelomeric regions of the Vv3 genome, in particular in the flanking regions of a highly duplicated gene encoding a Telomere-Linked Helicase, while both features were absent from the Sl3 and B05.10 genomes. Different retrotransposons in the Sl3 and the Vv3 strains resulted in the synthesis of distinct sets of small RNAs. Finally, extending the study to additional strains indicated that the accessory chromosome BCIN19 and the small RNAs producing retrotransposons Copia_4 and Gypsy_7 are common features of the G1 population that are scarcely if ever found in strains isolated from other populations. This research reveals that accessory chromosomes, repertoires of transposons and their derived small RNAs differ between populations of B. cinerea specialized on different hosts. The genomic data characterized in our study pave the way for further studies aiming at investigating the molecular mechanisms underpinning host specialization in a polyphagous pathogen.

genomics↗

bioRxiv2022

NGS data reveals the phyllosphere microbiome of wheat plants infected by the fungal pathogen Zymoseptoria tritici

The fungal pathogen Zymoseptoria tritici is the causal agent of Septoria tritici blotch (STB), a major wheat disease in Western Europe. Microorganisms inhabiting wheat leaves might act as beneficial, biocontrol or facilitating agents that could limit or stimulate the development of Z. tritici. Improving our understanding of microbial communities in the wheat phyllosphere would lead to new insights into STB management. This resource announcement provides fungal and bacterial metabarcoding datasets obtained by sampling wheat leaves differing in the presence of symptoms caused by Z. tritici. Tissues were sampled from three wheat commercial varieties on three sampling dates during a cropping season. Weeds around wheat fields were sampled as well. In total, more than 450 leaf samples were collected. The pathogen Z. tritici was quantified using qPCR. We provide the raw metabarcoding datasets, the Amplicon Sequence Variant (ASV) tables obtained after bioinformatic processing, the metadata associated to each sample (sampling date, wheat variety and tissue health condition), a preliminary descriptive analysis of the data, and the code used for bioinformatic and descriptive statistical analysis.

microbiology↗

bioRxiv2021

Why do fungicide mixtures delay the evolution of resistance? An experimental evolutionary approach

Pesticide resistance poses a critical threat to agriculture, human health and biodiversity. Mixtures of fungicides are recommended and widely used in resistance management strategies. However, the components of the efficiency of such mixtures remain unclear. We performed an experimental evolution study on the fungal pathogen Z. tritici, to determine how mixtures managed resistance. We compared the effect of the continuous use of single active ingredients to that of mixtures, at the minimal dose providing full control of the disease, which we refer to as the "efficient" dose. We found that the performance of efficient-dose mixtures against an initially susceptible population depended strongly on the components of the mixture. Such mixtures were either as durable as the best mixture component used alone, or worse than all components used alone. Moreover, efficient-dose mixture regimes probably select for generalist resistance profiles as a result of the combination of selection pressures exerted by the various components and their lower doses. Our results indicate that mixtures should not be considered a universal strategy. Experimental evaluations of specificities for the pathogens targeted, their interactions with fungicides and the interactions between fungicides are crucial for the design of sustainable resistance management strategies.

evolutionary biology↗

bioRxiv2021

Directed evolution predicts cytochrome b G37V target site modification as probable adaptive mechanism towards the QiI fungicide fenpicoxamid in Zymoseptoria tritici.

Acquired resistance is a threat for antifungal efficacy in medicine and agriculture. The diversity of possible resistance mechanisms, as well as the highly adaptive traits of pathogens make it difficult to predict evolutionary outcomes of treatments. We used directed evolution as an approach to assess the risk of resistance to the new fungicide fenpicoxamid in the wheat pathogenic fungus Zymoseptoria tritici. Fenpicoxamid inhibits complexIII of the respiratory chain at the ubiquinone reduction site (Qi site) of the mitochondrially encoded cytochrome b, a different site than the widely-used strobilurins which the respiratory complex by binding to the ubiquinol oxidation site (Qo site). We identified the G37V change, within the cytochrome b Qi site, as the most likely resistance mechanism to be selected in Z. tritici. This change triggered high fenpicoxamid resistance and halved the enzymatic activity of cytochrome b, despite no significant penalty for in vitro growth. In addition, we identified a negative cross-resistance between isolates harboring G37V or G143A, a Qo site change previously selected by strobilurins. Moreover, double mutants were less resistant to both QiIs and QoIs compared to single mutants. This work is a proof of concept that experimental evolution can be used to predict adaptation to fungicides, and provides new perspectives for the management of QiIs. Originality-Significance StatementO_LIThe highly adaptive traits of pathogens render evolutionary outcomes of antifungal treatments difficult to predict. C_LIO_LIWe used directed evolution to assess the risk of resistance to the new fungicide fenpicoxamid in the wheat pathogenic fungus Zymoseptoria tritici. C_LIO_LIWe identified a target modification as the most likely resistance mechanism to be selected. C_LIO_LIThis change triggered high fenpicoxamid resistance and halved the activity of the target enzyme despite no significant penalty for in vitro growth. C_LIO_LIThis work supports the use of experimental evolution as a method to predict adaptation to fungicides and provides important information for the management of QiIs. C_LI

microbiology↗

bioRxiv2021

I alternate therefore I generalize: how the intrinsic resistance risk of fungicides counterbalances their durability

The evolution of resistance to pesticides is a major burden in agriculture. Resistance management involves maximizing selection pressure heterogeneity, particularly by combining active ingredients with different modes of action. We tested the hypothesis that alternation may delay the build-up of resistance not only by spreading selection pressure over longer periods, but also by decreasing the rate of evolution of resistance to alternated fungicides, by applying an experimental evolution approach to the economically important crop pathogen Zymoseptoria tritici. Our results show that alternation is either neutral or slows the overall resistance evolution rate, relative to continuous fungicide use, but results in higher levels of generalism in evolved lines. We demonstrate that the nature of the fungicides, and therefore their relative intrinsic risk of resistance may underly this trade-off, more so than the number of fungicides and the rhythm of alternation. This trade-off is also dynamic over the course of resistance evolution. These findings open up new possibilities for tailoring resistance management effectively while optimizing interplay between alternation components.

evolutionary biology↗

bioRxiv2020

Monitoring systems for resistance to plant protection products across the world: Between redundancy and complementarity

BACKGROUNDMonitoring resistance to Plant Protection Products (PPPs) is crucial for understanding the evolution of resistances in bioagressors, thereby allowing scientists to design sound bioagressor management strategies. Globally, resistance monitoring is implemented by a wide range of actors that fall into three distinct categories: academic, governmental, and private. The purpose of this study was to investigate worldwide diversity in PPP resistance monitoring systems, and to shed light on their different facets. RESULTSA large survey involving 162 experts from 48 countries made it possible to identify and analyze 250 resistance monitoring systems. Through an in-depth analysis, the features of the different monitoring systems were identified. The main factor differentiating monitoring systems was essentially the capabilities (funding, manpower, technology, etc.) of the actors involved in each system. In most countries, and especially in those with a high Human Development Index, academic, governmental, and private monitoring systems coexist. Overall, systems focus far more on monitoring established resistances than on the detection of emerging resistances. Governmental and private resistance monitoring systems generally have considerable capacities to generate data, whereas academic resistance monitoring systems are more specialized. Governmental actors federate and enroll a wider variety of stakeholders. CONCLUSIONThe results show functional complementarities between the coexisting actors in countries where they coexist. We suggest PPP resistance monitoring might be enhanced if the different actors focus more on detecting emerging resistances (and associated benefits) and increase collaborative and collective efforts and transparency.

plant biology↗

bioRxiv2020

POPULATION GENOMICS REVEALS MOLECULAR DETERMINANTS OF SPECIALIZATION TO TOMATO IN THE POLYPHAGOUS FUNGAL PATHOGEN BOTRYTIS CINEREA

Many fungal plant pathogens encompass multiple populations specialized on different plant species. Understanding the factors underlying pathogen adaptation to their hosts is a major challenge of evolutionary microbiology, and it should help preventing the emergence of new specialized pathogens on novel hosts. Previous studies have shown that French populations of the grey mould pathogen Botrytis cinerea parasitizing tomato and grapevine are differentiated from each other, and have higher aggressiveness on their host-of-origin than on other hosts, indicating some degree of host specialization in this polyphagous pathogen. Here, we aimed at identifying the genomic features underlying the specialization of B. cinerea populations to tomato and grapevine. Based on whole genome sequences of 32 isolates, we confirmed the subdivision of B. cinerea pathogens into two genetic clusters on grapevine and another, single cluster on tomato. Levels of genetic variation in the different clusters were similar, suggesting that the tomato-specific cluster has not recently emerged following a bottleneck. Using genome scans for selective sweeps and divergent selection, tests of positive selection based on polymorphism and divergence at synonymous and non-synonymous sites and analyses of presence/absence variation, we identified several candidate genes that represent possible determinants of host specialization in the tomato-associated population. This work deepens our understanding of the genomic changes underlying the specialization of fungal pathogen populations.

microbiology↗

bioRxiv2020

Large-scale study reveals regional fungicide applications as a major determinant of resistance evolution in the wheat pathogen Zymoseptoria tritici in France.

O_LIResearch rationale: In modern cropping systems, the near-universal use of plant protection products selects for resistance in pest populations. The emergence and evolution of this adaptive trait threaten treatment efficacy. We identified determinants of fungicide resistance evolution and quantified their effects at a large spatiotemporal scale. C_LIO_LIMethods: We focused on Zymoseptoria tritici, which causes leaf blotch in wheat. Phenotypes of qualitative or quantitative resistance to various fungicides were monitored annually, from 2004 to 2017, at about 70 sites throughout 22 regions of France (territorial units of 25 000km2 on average). We modelled changes in resistance frequency with regional anti-Septoria fungicide use, yield losses due to the disease and the regional area under organic wheat. C_LIO_LIKey results: The major driver of resistance dynamics was fungicide use at the regional scale. We estimated its effect on the increase in resistance and relative apparent fitness of each resistance phenotype. The predictions of the model replicated the spatiotemporal patterns of resistance observed in field populations (R2 from 0.56 to 0.82). C_LIO_LIMain conclusion: The evolution of fungicide resistance is mainly determined at the regional scale. This study therefore showed that collective management at the regional scale could effectively complete local actions. C_LI

evolutionary biology↗

bioRxiv2019

The polyphagous plant pathogenic fungus Botrytis cinerea encompasses host-specialized and generalist populations

The host plant is often the main variable explaining population structure in fungal plant pathogens, because specialization contributes to reduce gene flow between populations associated with different hosts. Previous population genetic analysis revealed that French populations of the grey mould pathogen Botrytis cinerea were structured by hosts tomato and grapevine, suggesting host specialization in this highly polyphagous pathogen. However, these findings raised questions about the magnitude of this specialization and the possibility of specialization to other hosts. Here we report specialization of B. cinerea populations to tomato and grapevine hosts but not to other tested plants. Population genetic analysis revealed two pathogen clusters associated with tomato and grapevine, while the other clusters co-occurred on hydrangea, strawberry and bramble. Measurements of quantitative pathogenicity were consistent with host specialization of populations found on tomato, and to a lesser extent, populations found on grapevine. Pathogen populations from hydrangea and strawberry appeared to be generalist, while populations from bramble may be weakly specialized. Our results suggest that the polyphagous B. cinerea is more accurately described as a collection of generalist and specialist individuals in populations. This work opens new perspectives for grey mold management, while suggesting spatial optimization of crop organization within agricultural landscapes.

microbiology↗

bioRxiv2017

Stability in the genetic structure of a Zymoseptoria tritici population from epidemic to interepidemic stages at a small spatial scale

Subpopulations of the wheat pathogen Zymoseptoria tritici (26 sample groups composed of 794 strains) were collected in two nearby wheat fields in the Paris basin, during both epidemic and inter-epidemic periods of three successive years (2009-2013). In addition to the type of inoculum (ascospores vs. pycnidiospores), the alternative presence of wheat debris allowed taking into account its putative origin (local vs. distant). We used a molecular epidemiology approach, based on population genetic indices derived from SSR marker analysis, to describe putative changes in the structure and genotypic diversity of these subpopulations over three years, at a spatiotemporal scale consistent with epidemiological observations. Genetic structure was stable over time (within and between years) and between fields. All subpopulations displayed very high levels of gene and genotypic diversity. The low levels of linkage disequilibrium and the very low clonal fraction at all stages were consistent with the regular occurrence of sexual reproduction in the two fields. A significant increase of the MAT1-1/MAT1-2 ratio was observed over the course of the epidemics, suggesting a competitive advantage of MAT1-1 strains consistently with their greater pathogenicity reported in the literature. Finally, we found that the period, the type of inoculum and its putative origin had little effect on the short term evolution of the local population of Z. tritici. Fungal population size and diversity are apparently large enough to prevent genetic drift at this fine spatiotemporal scale, and more likely short distance migration contributes strongly to the stabilization of genetic diversity among and within plots.

epidemiology↗

bioRxiv2017

Plasticity of the MFS1 promoter leads to multi drug resistance in the wheat pathogen Zymoseptoria tritici

The ascomycete Zymoseptoria tritici is the causal agent of septoria leaf blotch on wheat. Disease control relies mainly on resistant wheat cultivars and on fungicide applications. The fungus displays a high potential to circumvent both methods. Resistance against all unisite fungicides has been observed over decades. A different type of resistance has emerged among wild populations with multi-drug resistant (MDR) strains. Active fungicide efflux through overexpression of the major-facilitator gene, MFS1, explains this emerging resistance mechanism. In this study, we identified as responsible mutations three types of inserts in the MFS1 promoter, two of which harboring potential transcription factor binding sites. We show, that type I insertion leads to MFS1 overexpression and consequently to MDR. Interestingly, all three inserts correspond to repeated elements of the Z. tritici genome.\n\nThese results underline the plasticity of repeated elements leading to fungicide resistance in Z. tritici and which contribute to its adaptive potential.

microbiology↗