Biology subjects

Fournier, E.

Publications and source records attributed to Fournier, E..

12 recordsLinked to original sources

bioRxiv2026

MIRA: an open source and user-friendly software to automate counting and sizing of fungal spores

BackgroundThe quantification of fungal spores constitutes a fundamental metric in phytopathology, serving as the primary variable for inoculum standardization and being used as a proxy for disease severity. Historically, spore quantification has relied on manual hemocytometry, which remains the most precise counting process to date, where chambers such as the Malassez slide are used to count a subsample of the inoculum. However, this method applied manually is highly labor-intensive, time-consuming, and can be prone to operator-dependent variability. To overcome these limitations, we introduce MIRA (Microscopy Image Recognition & Analysis), a novel open-source software integrating You Only Look Once (YOLO) deep learning algorithms. Featuring a user-friendly graphical interface, MIRA is adaptable to multiple camera systems and supports advanced object detection models, including YOLOv11 and YOLOv26. ResultsWe demonstrate that MIRA can be used to accurately detect and count spores from several phytopathogenic fungi, automatically measure spore surface area, and to differentiate spores across different genera. In an exhaustive comparative analysis using Pyricularia oryzae spores as an example, MIRA was benchmarked against manual gold-standard counting slides (Malassez and Kova) and indirect spectrophotometric methods (SPARK). The P. oryzae model loaded via MIRA achieved a strong correlation (R = 0.96) with manual gold standards while reducing processing time by over 90% for high-concentration samples (10 spores/mL). Beyond this benchmark, we also successfully tested specific YOLO models designed to recognize macro- and microconidia of Fusarium oxysporum f. sp. cubense, a model for Pseudocercospora fijiensis, and a single multiclass model capable of identifying six different rice pathogenic fungi. We provide comprehensive tutorials for operating the software and training custom detection models for free using Roboflow and Google Colab. MIRA is available both as open-source Python code and as standalone executables for Windows and Linux. ConclusionsMIRA provides a rapid, accurate, and highly reproducible alternative to manual spore counting, effectively removing a major bottleneck in phytopathology workflows. By combining advanced YOLO-based deep learning with an accessible interface and comprehensive training resources, MIRA makes accessible automated image analysis for researchers without programming expertise. Moreover, MIRA drastically improves the efficiency of high-throughput disease phenotyping and can be adapted for a wide range of microscopic quantification tasks across various biological disciplines.

plant biology↗

bioRxiv2026

Population structure of the rice blast fungus Pyricularia oryzae in the context of elevated host heterogeneity

In a context where pathogen emergence threatens global food security, crop resistance to pathogens is crucial for preventing and controlling epidemics. Agrosystems with high intraspecific cultivated diversity are expected to better mitigate infectious diseases compared with homogeneous ones. However, the long-term impact of cultivated genotypic heterogeneity on pathogen populations is still under debate. The 1,300-year-old traditional agrosystem of the Yuanyang Terraces (YYT) in China exhibits an outstanding level of rice diversity. The rice blast fungus Pyricularia oryzae has been recorded in the area, making this area ideal for studying the impact of long term heterogeneous cultivation on plant-pathogen coevolution. In this study, we analysed 94 P. oryzae genomes from the YYT and compared them to 198 genomes representative of the worldwide diversity. We report elevated levels of genomic diversity of P. oryzae in the YYT. Whereas the worldwide diversity of this species is organized in four lineages, we detected seven lineages within the YYT, four of which are restricted to this area. The sampling dates of the YYT isolates (2009 to 2017) provided sufficient temporal signal to date nodes of the phylogenetic tree of isolates. Endemic lineages appeared to have arisen several centuries ago but later than the origin of the YYT themselves. We also detected recent introductions of worldwide lineages. Linkage disequilibrium analyses and in vitro cross experiments suggest that P. oryzae reproduces asexually in the YYT. These results suggest that long-term intraspecific crop diversity in the YYT has promoted the emergence and maintenance of a highly diverse, locally adapted pathogen population.

evolutionary biology↗

bioRxiv2026

Monoclonal anti-dsRNA antibody-based metagenomics (MADAM) reveal Pyricularia oryzae mycovirome

This study introduces MADAM (Monoclonal Anti-dsRNA Antibody-Based Metagenomics), a novel approach that integrates multiple technical modules previously used independently in other protocols. MADAM combines monoclonal antibody-mediated double-stranded RNA (dsRNA) enrichment, sequence-independent RT-PCR, and Oxford Nanopore Technologies (ONT) sequencing. Applied to Pyricularia oryzae, the causal agent of rice blast disease, MADAM enabled the comprehensive characterization of mycovirus genomes from four fungal isolates collected in Yunnan, China. The approach achieved high viral read recovery rates (46.9-72.7%) and identified 18 P. oryzae-associated RNA viruses spanning seven families: Botourmiaviridae, Deltaormycoviridae, Mymonaviridae, Partitiviridae, Polymycoviridae, Splipalmiviridae, and Ambiguiviridae. Seventeen nearly complete to complete viral genomes (1,226-6,085 nucleotides) were recovered, with sequence coverage ranging from 88% to 100%. Co-infections were detected in three of the four isolates, with notable discoveries including the first deltaormycovirus reported in P. oryzae, a putative novel member of Botourmiaviridae, and an additional genomic segment of a polymycovirus. MADAM successfully detected positive-sense, negative-sense ssRNA, and dsRNA viruses, demonstrating its broad applicability. By uncovering novel viruses and resolving complex co-infections, this method proves invaluable for fungal virology, with potential applications in diagnostics, surveillance, and biological control. Ultimately, MADAM advances our understanding of fungal viral diversity and paves the way for further exploration of mycovirus ecology and evolution.

microbiology↗

bioRxiv2024

Identification of genetic bases of male fertility in Pyricularia oryzae by Genome Wide Association Study (GWAS).

The reproductive system of an organism impacts the emergence and evolution of adaptive variants in response to selective constraints. The understanding of the sexual mode of reproduction in pathogens helps to understand their life history. In filamentous Ascomycete fungi, mating type system and the production of gametes are required to reproduce sexually. In the phytopathogenic Ascomycete Pyricularia oryzae, studies of the genetic determinants of sexual reproduction are still limited to the mating type system. This study focuses on identifying the genes involved in male fertility through the production of male gametes known as microconidia. We performed a GWAS analysis coupled with a local score approach on a wild recombinant population of Pyricularia oryzae phenotyped for microconidia production. We identified one genomic region significantly associated with the quantity of microconidia produced. This region contained nine candidate genes, some of them annotated with functions associated to sexual reproduction in model fungi such as Neurospora crassa, Podospora anserina and Sordaria macrospora. The most promising candidate gene contains a Jumonji domain. Proteins belonging to the Jumonji family are conserved among Eukaryotes and are known to be involved in chromatin regulation.

genetics↗

bioRxiv2023

Deep population structure linked to host vernalization requirement in the barley net blotch fungal pathogen

Invasive fungal pathogens pose a substantial threat to widely cultivated crop species, owing to their capacity to adapt to new hosts and new environmental conditions. Gaining insights into the demographic history of these pathogens and unraveling the mechanisms driving coevolutionary processes are crucial for the development of durably effective disease management programs. Pyrenophora teres is a significant fungal pathogen of barley, consisting of two lineages, Ptt and Ptm, with global distributions and demographic histories reflecting barley domestication and spread. However, the factors influencing the population structure of P. teres remain poorly understood, despite the varietal and environmental heterogeneity of barley agrosystems. Here, we report on the population genomic structure of P. teres in France and globally. We used genotyping-by-sequencing to show that Ptt and Ptm can coexist in the same area in France, with Ptt predominating. Furthermore, we showed that differences in the vernalization requirement of barley varieties were associated with population differentiation in France and at a global scale, with one population cluster found on spring barley and another population cluster found on winter barley. Our results demonstrate how cultivation conditions, possibly associated with genetic differences between host populations, can be associated with the maintenance of divergent invasive pathogen populations coexisting over large geographic areas. This study not only advances our understanding of the coevolutionary dynamics of the Pt-barley pathosystem but also prompts further research on the relative contributions of adaptation to the host versus adaptation to abiotic conditions in shaping Ptt populations. Impact statementMany invasive fungal pathogens have successfully followed major crop species throughout their intercontinental range, but continue to represent dynamic biotic threats. During their geographic expansion, invasive fungal populations were subjected to heterogeneous environmental conditions, or different populations of hosts, which could result in adaptation processes. Understanding this history of colonization can allow us to better prevent the emergence of infectious diseases of crops, and to better control them. One such fungus, Pyrenophora teres, negatively impacts barley production globally by causing net blotch disease. In this study, we characterized the genetic makeup of P. teres in France and how it compares with what can be sampled in other regions of the world. We found that both the net and spot forms of Pyrenophora teres can be in the same area in France, but the spot form is more common. We also discovered that the net form populations associated with winter and spring barley are different, which was not known until now. This study opens up numerous experimental perspectives aimed at evaluating whether the two populations of net form are adapted to their hosts or to the conditions of cultivation of their hosts, with the goal of implementing measures that force the pathogen to maladaptation. Data summaryGBS data are available under BioProject PRJEB66440. Single nucleotide polymorphism and reference genome assembly are available under doi: https://doi.org/10.5281/zenodo.10021844. Reads used for genome assembly are available under the BioProject PRJEB66476. The authors confirm all supporting data, code, and protocols have been provided within the article or through supplementary data files.

evolutionary biology↗

bioRxiv2022

In vitro modelling of oral microbial invasion in the human colon

Recent advances in the human microbiome characterization have revealed significant oral microbial detection in stools of dysbiotic patients. However, little is known about the potential interactions of these invasive oral microorganisms with commensal intestinal microbiota and host. In this proof of concept study, we propose a new model of oral to gut invasion by the combined use of an in vitro model simulating both the physicochemical and microbial (lumen and mucus-associated microbes) parameters of the human colon (M-ARCOL), a salivary enrichment protocol and whole metagenome shotgun sequencing. Oral invasion of the intestinal microbiota was simulated by injection of enriched saliva in the in vitro colon model inoculated with faecal sample from the same healthy adult donor. The mucosal compartment of M-ARCOL was able to retain the highest species richness levels over time, whilst it decreased in the luminal compartment. This study also showed that oral microorganisms preferably colonized the mucosal microenvironment, suggesting potential oral-to-intestinal mucosal competitions. This new model of oral-to-gut invasion can provide useful mechanistic insights into the role of oral microbiome in various disease processes.

microbiology↗

bioRxiv2021

SARS-CoV-2 Omicron Spike recognition by plasma from individuals receiving BNT162b2 mRNA vaccination with a 16-weeks interval between doses

Continuous emergence of SARS-CoV-2 variants of concern (VOC) is fueling the COVID-19 pandemic. Omicron (B.1.1.529), is rapidly spreading worldwide. The large number of mutations in its Spike raised concerns about a major antigenic drift that could significantly decrease vaccine efficacy and infection-induced immunity. A long interval between BNT162b2 mRNA doses was shown to elicit antibodies that efficiently recognize Spikes from different VOCs. Here we evaluated the recognition of Omicron Spike by plasma from a cohort of SARS-CoV-2 naive and previously-infected individuals that received their BNT162b2 mRNA vaccine 16-weeks apart. Omicron Spike was recognized less efficiently than D614G, Alpha, Beta, Gamma and Delta Spikes. We compared to plasma activity from participants receiving a short (4-weeks) interval regimen. Plasma from individuals of the long interval cohort recognized and neutralized better the Omicron Spike compared to those that received a short interval. Whether this difference confers any clinical benefit against Omicron remains unknown.

microbiology↗

bioRxiv2021

COEVOLUTION WITH SPATIALLY STRUCTURED RICE LANDRACES MAINTAINS MULTIPLE GENERALIST LINEAGES IN THE RICE BLAST PATHOGEN

Traditional agrosystems, where humans, crops and microbes have coevolved over long periods, can serve as models to understand the eco-evolutionary determinants of disease dynamics and help the engineering of durably resistant agrosystems. Here, we investigated the genetic and phenotypic relationship between rice (Oryza sativa) landraces and their rice blast pathogen (Magnaporthe oryzae) in the traditional Yuanyang terraces of flooded rice paddies in China, where rice landraces have been grown and bred over centuries without significant disease outbreaks. Analyses of genetic subdivision revealed that indica rice plants clustered according to landrace names. Three new diverse lineages of rice blast specific to the Yuanyang terraces coexisted with lineages previously detected at the worldwide scale. Population subdivision in the pathogen population did not mirror pattern of population subdivision in the host. Measuring the pathogenicity of rice blast isolates on landraces revealed generalist life histories. Our results suggest that the implementation of disease control strategies based on the emergence or maintenance of a generalist lifestyle in pathogens may sustainably reduce the burden of disease in crops.

evolutionary biology↗

bioRxiv2020

Ecological differentiation and incipient speciation in the fungal pathogen causing rice blast

Many invasive fungal species coexist as multiple lineages on the same host, but the factors underlying the origin and maintenance of population structure remain largely unknown. Here, we analyzed genetic and phenotypic diversity in isolates of the rice blast fungus (Pyricularia oryzae) covering a broad geographical range. We showed that the four lineages of P. oryzae were found in areas with different prevailing environmental conditions and types of rice grown, indicating niche separation. Pathogenicity tests revealed that specialization to rice subspecies contributed to niche separation between lineages, and differences in repertoires of putative virulence effectors were consistent with differences in host range. Experimental crosses revealed that female sterility and early post-mating genetic incompatibilities acted as strong barriers to gene flow between these lineages. Our results demonstrate that the spread of a pathogen across heterogeneous habitats and divergent populations of a crop species can lead to niche separation and reproductive isolation between distinct invasive lineages.

evolutionary biology↗

bioRxiv2020

Congruent population genetic structures and divergence histories in anther-smut fungi and their host plants Silene italica and the S. nutans species complex

The study of population genetic structure congruence between hosts and pathogens gives important insights into their shared phylogeographic and coevolutionary histories. We studied the population genetic structure of castrating anther-smut fungi (Microbotryum genus) and of their host plants, the Silene nutans species complex, and the morphologically and genetically close S. italica, which can be found in sympatry. Phylogeographic population genetic structure related to persistence in separate glacial refugia has been recently revealed in the S. nutans plant species complex across Western Europe, identifying several distinct lineages. We genotyped 171 associated plant-pathogen pairs of anther-smut fungi and their host plant individuals using microsatellite markers and plant chloroplastic SNPs. We found clear differentiation between fungal populations parasitizing S. nutans and S. italica plants. The population genetic structure of fungal strains parasitizing the S. nutans plant species complex mirrored the host plant genetic structure, suggesting that the pathogen was isolated in glacial refugia together with its host and/or that it has specialized on the plant genetic lineages. Using random forest approximate Bayesian computation (ABC-RF), we found that the divergence history of the fungal lineages on S. nutans was congruent with the one previously inferred for the host plant and likely occurred with ancient but no recent gene flow. Genome sequences confirmed the genetic structure and the absence of recent gene flow between fungal genetic lineages. Our analyses of host-pathogen individual pairs contribute to a better understanding of co-evolutionary histories between hosts and pathogens in natural ecosystems, in which such studies are still scarce.

evolutionary biology↗

bioRxiv2017

Coexistence of multiple endemic and pandemic lineages of the rice blast pathogen

The rice blast fungus Magnaporthe oryzae (syn. Pyricularia oryzae) is both a threat to global food security and a model for plant pathology. Molecular pathologists need an accurate understanding of the origins and line of descent of M. oryzae populations, to identify the genetic and functional bases of pathogen adaptation, and to guide the development of more effective control strategies. We used a whole-genome sequence analysis of samples from different times and places to infer details about the genetic makeup of M. oryzae from a global collection of isolates. Analyses of population structure identified six lineages within M. oryzae, including two pandemic on japonica and indica rice, respectively, and four lineages with more restricted distributions. Tip-dating calibration indicated that M. oryzae lineages separated about a millenium ago, long after the initial domestication of rice. The major lineage endemic to continental Southeast Asia displayed signatures of sexual recombination and evidence of DNA acquisition from multiple lineages. Tests for weak natural selection revealed that the pandemic spread of clonal lineages entailed an evolutionary cost, in terms of the accumulation of deleterious mutations. Our findings reveal the coexistence of multiple endemic and pandemic lineages with contrasting population and genetic characteristics within a widely distributed pathogen.\n\nImportanceThe rice blast fungus Magnaporthe oryzae (syn. Pyricularia oryzae) is a textbook example of a rapidly adapting pathogen, and is responsible for one of the most damaging diseases of rice. Improvements in our understanding of Magnaporthe oryzae diversity and evolution are required, to guide the development of more effective control strategies. We used genome sequencing data for samples from around the world to infer the evolutionary history of M. oryzae. We found that M. oryzae diversified about a thousand years ago ago, separating into six main lineages: two pandemic on japonica and indica rice, respectively, and four with more restricted distributions. We also found that a lineage endemic to continental Southeast Asia displayed signatures of sexual recombination and the acquisition of genetic material from multiple lineages. This work provides a population-level genomic framework for defining molecular markers for the control of rice blast and investigations of the molecular basis of differences in pathogenicity between M. oryzae lineages.

evolutionary biology↗

bioRxiv2017

Gene flow between divergent cereal- and grass-specific lineages of the rice blast fungus Magnaporthe oryzae

Delineating species and epidemic lineages in fungal plant pathogens is critical to our understanding of disease emergence and the structure of fungal biodiversity, and also informs international regulatory decisions. Pyricularia oryzae (syn. Magnaporthe oryzae) is a multi-host pathogen that infects multiple grasses and cereals, is responsible for the most damaging rice disease (rice blast), and of growing concern due to the recent introduction of wheat blast to Bangladesh from South America. However, the genetic structure and evolutionary history of M. oryzae, including the possible existence of cryptic phylogenetic species, remain poorly defined. Here, we use whole-genome sequence information for 76 M. oryzae isolates sampled from 12 grass and cereal genera to infer the population structure of M. oryzae, and to reassess the species status of wheat-infecting populations of the fungus. Species recognition based on genealogical concordance, using published data or extracting previously-used loci from genome assemblies, failed to confirm a prior assignment of wheat blast isolates to a new species (Pyricularia graminis tritici). Inference of population subdivisions revealed multiple divergent lineages within M. oryzae, each preferentially associated with one host genus, suggesting incipient speciation following host shift or host range expansion. Analyses of gene flow, taking into account the possibility of incomplete lineage sorting, revealed that genetic exchanges have contributed to the makeup of multiple lineages within M. oryzae. These findings provide greater understanding of the eco-evolutionary factors that underlie the diversification of M. oryzae and highlight the practicality of genomic data for epidemiological surveillance in this important multi-host pathogen.\n\nImportanceInfection of novel hosts is a major route for disease emergence by pathogenic micro-organisms. Understanding the evolutionary history of multi-host pathogens is therefore important to better predict the likely spread and emergence of new diseases. Magnaporthe oryzae is a multi-host fungus that causes serious cereal diseases, including the devastating rice blast disease, and wheat blast, a cause of growing concern due to its recent spread from South America to Asia. Using whole genome analysis of 76 fungal strains from different hosts, we have documented the divergence of M. oryzae into numerous lineages, each infecting a limited number of host species. Our analyses provide evidence that inter-lineage gene flow has contributed to the genetic makeup of multiple M. oryzae lineages within the same species. Plant health surveillance is therefore warranted to safeguard against disease emergence in regions where multiple lineages of the fungus are in contact with one another.

microbiology↗