Quantitative pathogenicity and host adaptation in a fungal plant pathogen revealed by whole-genome sequencing.

Knowledge of genetic determinism and evolutionary dynamics mediating host-pathogen interactions is essential to manage fungal plant diseases. Studies on the genetic architecture of fungal pathogenicity often focus on large-effect effector genes triggering strong, qualitative resistance. It is not clear how this translates to predominately quantitative interactions. Here, we use the Zymoseptoria tritici-wheat model to elucidate the genetic architecture of quantitative pathogenicity and mechanisms mediating host adaptation. With a multi-host genome-wide association study, we identify 19 high-confidence candidate genes associated with quantitative pathogenicity. Analysis of genetic diversity reveals that sequence polymorphism is the main evolutionary process mediating differences in quantitative pathogenicity, a process that is likely facilitated by genetic recombination and transposable element dynamics. Finally, we use functional approaches to confirm the role of an effector-like gene and a methyltransferase in phenotypic variation. This study highlights the complex genetic architecture of quantitative pathogenicity, extensive diversifying selection and plausible mechanisms facilitating pathogen adaptation.

Epidemiological trade-off between intra- and interannual scales in the evolution of aggressiveness in a local plant pathogen population.

The efficiency of plant resistance to fungal pathogen populations is expected to decrease over time, due to their evolution with an increase in the frequency of virulent or highly aggressive strains. This dynamics may differ depending on the scale investigated (annual or pluriannual), particularly for annual crop pathogens with both sexual and asexual reproduction cycles. We assessed this time-scale effect, by comparing aggressiveness changes in a local Zymoseptoria tritici population over an 8-month cropping season and a 6-year period of wheat monoculture. We collected two pairs of subpopulations to represent the annual and pluriannual scales: from leaf lesions at the beginning and end of a single annual epidemic and from crop debris at the beginning and end of a 6-year period. We assessed two aggressiveness traits-latent period and lesion size-on sympatric and allopatric host varieties. A trend toward decreased latent period concomitant with a significant loss of variability was established during the course of the annual epidemic, but not over the 6-year period. Furthermore, a significant cultivar effect (sympatric vs. allopatric) on the average aggressiveness of the isolates revealed host adaptation, arguing that the observed patterns could result from selection. We thus provide an experimental body of evidence of an epidemiological trade-off between the intra- and interannual scales in the evolution of aggressiveness in a local plant pathogen population. More aggressive isolates were collected from upper leaves, on which disease severity is usually lower than on the lower part of the plants left in the field as crop debris after harvest. We suggest that these isolates play little role in sexual reproduction, due to an Allee effect (difficulty finding mates at low pathogen densities), particularly as the upper parts of the plant are removed from the field, explaining the lack of transmission of increases in aggressiveness between epidemics.

Testing Differences Between Pathogen Compositions with Small Samples and Sparse Data.

The structure of pathogen populations is an important driver of epidemics affecting crops and natural plant communities. Comparing the composition of two pathogen populations consisting of assemblages of genotypes or phenotypes is a crucial, recurrent question encountered in many studies in plant disease epidemiology. Determining whether there is a significant difference between two sets of proportions is also a generic question for numerous biological fields. When samples are small and data are sparse, it is not straightforward to provide an accurate answer to this simple question because routine statistical tests may not be exactly calibrated. To tackle this issue, we built a computationally intensive testing procedure, the generalized Monte Carlo plug-in test with calibration test, which is implemented in an R package available at https://doi.org/10.5281/zenodo.635791 . A simulation study was carried out to assess the performance of the proposed methodology and to make a comparison with standard statistical tests. This study allows us to give advice on how to apply the proposed method, depending on the sample sizes. The proposed methodology was then applied to real datasets and the results of the analyses were discussed from an epidemiological perspective. The applications to real data sets deal with three topics in plant pathology: the reproduction of Magnaporthe oryzae, the spatial structure of Pseudomonas syringae, and the temporal recurrence of Puccinia triticina.

Influence of cultivated landscape composition on variety resistance: an assessment based on wheat leaf rust epidemics.

In plant pathology, the idea of designing variety management strategies at the scale of cultivated landscapes is gaining more and more attention. This requires the identification of effects that take place at large scales on host and pathogen populations. Here, we show how the landscape varietal composition influences the resistance level (as measured in the field) of the most grown wheat varieties by altering the structure of the pathogen populations. For this purpose, we jointly analysed three large datasets describing the wheat leaf rust pathosystem (Puccinia triticina/Triticum aestivum) at the country scale of France with a Bayesian hierarchical model. We showed that among all compatible pathotypes, some were preferentially associated with a variety, that the pathotype frequencies on a variety were affected by the landscape varietal composition, and that the observed resistance level of a variety was linked to the frequency of the most aggressive pathotypes among all compatible pathotypes. This data exploration establishes a link between the observed resistance level of a variety and landscape composition at the national scale. It illustrates that the quantitative aspects of the host-pathogen relationship have to be considered in addition to the major resistance/virulence factors in landscape epidemiology approaches.

Aggressiveness components and adaptation to a host cultivar in wheat leaf rust.

Experimental evidence on the capacity of pathogen populations to quantitatively adapt to their hosts and on the life traits that are involved is lacking at this time. In this article, we identified a situation in which a leaf rust pathotype (P1) was found at a high frequency on a widely grown cultivar (Soissons) and we tested the hypothesis that P1 was more aggressive on Soissons than other virulent pathotypes (P2 and P3). Several components of the pathogen life cycle were measured on adult wheat plants in two different experiments under greenhouse conditions: latent period, spore production per lesion and per unit of sporulating tissue, uredinium size, and lesion life span. Regardless of the component, pathotype P1 was repeatedly found to be more aggressive than at least one of the other two pathotypes, with differences of 5 to 51%. Breaking down spore production per lesion into uredinium size and spore production per square millimeter of sporulating tissue showed that the three pathotypes presented different aggressiveness profiles, suggesting different development constraints for the pathogen, either for its growth capacity into host tissues or its ability to exploit the host resources for spore production. Although leaf rust pathotypes present a clonal structure, quantitative differences were found for aggressiveness traits within a pathotype.

Clonality and host selection in the wheat pathogenic fungus Puccinia triticina.

Clonal reproduction in Puccinia triticina, the cause of wheat leaf rust, has long been hypothesized but has never been demonstrated. Using a population genetics approach and microsatellite markers, we analysed genetic diversity of this fungus at each level of genome organisation. Sampling included isolates from two field populations growing on two cultivars carrying specific resistance genes, completed with isolates representing the main pathotypes identified from a national survey. For the two cultivars, populations differentiated according to the distribution of their genotypes and pathotypes. There was a high proportion of repeated genotypes, combined with a significant linkage disequilibrium and a strong negative value for FIS. These three factors, especially heterozygote excess, strongly support the hypothesis of a high rate of clonal reproduction. Each pathotype matched a unique multilocus genotype, except for a few isolates, which were taken to be mutants of the dominant genotype. We discussed the strong relationship between pathotypes and genotypes as the consequence of clonal reproduction combined with a strong selection exerted by host cultivars.

Distribution of pathotypes with regard to host cultivars in French wheat leaf rust populations.

ABSTRACT Isolates of wheat leaf rust collected from durum and bread wheat cultivars in France during 1999-2002 were analyzed for virulence on 18 Thatcher lines with single genes for leaf rust resistance (Lr genes). Sampling focused on the five most widely grown bread wheat cultivars (two susceptible and three resistant) to allow statistical comparison of diversity indexes between the cultivars. Leaf rust populations from durum and bread wheats were different. The diversity of the bread wheat leaf rust pathotypes, as measured by the Shannon index, ranged from 2.43 to 2.76 over the 4 years. Diversity for wheat leaf rust resistance was limited in the host since we postulated only seven seedling resistance genes in the 35 cultivars most widely grown during 1999-2002. Leaf rust populations were strongly differentiated for virulence within bread wheat cultivars, and diversity was higher on those that were resistant, mainly due to a more even distribution of virulence phenotypes than on susceptible cultivars. The pathogen population on the susceptible cv. Soissons was largely dominated by a single pathotype (073100), whereas all other pathotypes virulent on cv. Soissons either decreased in frequency or remained at a low frequency during the period studied. Several pathotypes including the most complex one were found only on resistant cultivars, even though most of them were virulent on the susceptible cv. Soissons. Specific interactions were necessary, but not always sufficient, to account for pathotype distribution and frequencies on the cultivars, suggesting that selection for virulence to host resistance genes is balanced by other selective forces including selection for aggressiveness.

Identifying leaf rust resistance genes and mapping gene Lr37 on the microsatellite map of wheat.

Based on seedling resistance tests, five resistance genes (Lr10, Lr3, Lr13, Lr14a and Lr37) against leaf rust (Puccinia triticina) were identified in 16 cultivars of European winter wheat. STS and SCAR markers were used to verify the presence of the resistance genes Lr37 and Lr10 against leaf rust in cultivars, near-isogenic lines and segregating populations. The Lr37 gene is present in a small translocation from Triticum ventricosum Ces. (Aegilops ventricosa Tausch) and is tightly linked with resistance genes Yr17 and Sr38. The Lr37 gene was identified in the cultivars Kris, Clever, Slade, Apache, Caphorn, Lorraine, Balthasar, Renan and confirmed by two PCR markers. The F3 progenies of the crosses Kris (Lr37) X Nutka (Lr37 not present) were used for map construction. Two STS/SCAR markers specific for Lr37 were mapped in relation to nine polymorphic microsatellites on chromosome 2AS. The microsatellite marker Xgwm1176 mapped relatively close to the STS and SCAR markers for Lr37 with a linkage distance of 4.1 cM.