Sustainable crop protection is vital for food security, yet it is under threat due to the adaptation of a diverse and evolving pathogen population. Resistance can be managed by maximising the diversity of selection pressure through dose variation and the spatial and temporal combination of active ingredients. This study explores the interplay between operational drivers for maximising the sustainability of management strategies in relation to the resistance status of fungal populations. We applied an experimental evolution approach to three artificial populations of Zymoseptoria tritici, an economically significant wheat pathogen, each differing in initial resistance status. Our findings reveal that diversified selection pressure curtails the selection of resistance in naïve populations and those with low frequencies of single resistance. Increasing the number of modes of action most effectively delays resistance development, surpassing the increase in the number of fungicides, fungicide choice based on resistance risk, and temporal variation in fungicide exposure. However, this approach favours generalism in the evolved populations. The prior presence of multiple resistant isolates and their subsequent selection in populations override the effects of diversity in management strategies, thereby invalidating any universal ranking. Therefore, the initial resistance composition must be specifically considered in sustainable resistance management to address real-world field situations.

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.

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.

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 evolutionary 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.

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 the temporal alternation of active ingredients 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. Here, we applied an original experimental evolution approach to the economically important crop pathogen Zymoseptoria tritici. We observed the dynamics of Z. tritici resistance in 56 independent lines subjected to 14 continuous or alternation regimes of 3 fungicides contrasting for their mode of action, at their EC95 selection doses. For the first time in a plant pathogen, our results show that alternation is either neutral or slows the evolution of resistance, relative to continuous fungicide use, but results in higher levels of generalism in evolved lines. We demonstrate that the mode of action of resistance of fungicides drivingly underlies this trade-off, more so than the number of fungicides and the frequency of alternation. This trade-off is also dynamic over the course of resistance evolution, as shown by the change in population phenotype structure and the relative impact of selection drivers. These findings open up new possibilities for tailoring resistance management effectively while optimizing smart interplay between alternation components. They also confirm experimental evolution as an untapped but promising approach to dissect adaptation in phytopathogenic fungi. Author summary The efficacy of pesticides has been compromised by the generalization of their use, leading to the rapid and widespread evolution of resistance. This constitutes a major economic and environmental burden in agriculture. The temporal alternation (or cycling) of active ingredients is a management strategy that induces temporal variation of selection of pathogens. Here, we dissected how it can be optimized according to the number and nature of alternated modes of action and to the rhythm of their application and sought to understand how these drivers determine the performance of alternation strategies. We used an approach original in plant pathology, experimental evolution, applied on an economically important fungus, Zymoseptoria tritici. We concluded that alternation can delay the rate of resistance selection with performance depending most likely on the mode of action of alternated antifungals, more so than on other drivers. But we also highlighted that alternation regimes select generalist resistance, that is the ability to resist a large diversity of antifungals. This trade-off was dynamic over time. Our study provides new insights for the informed management of pesticides and the reduction of their side effects.

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 evolution of resistance, relative to continuous fungicide use, but results in higher levels of generalism in evolved lines. We demonstrate that the relative risk of resistance intrinsic to fungicide alternation probably underlies a trade-off between the number of fungicides and the frequency 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.

Evaluating fungicide resistance management strategies by means of experimental evolution : the case of Zymoseptoria tritici, the causal agent of Septoria leaf blotch La résistance aux pesticides impacte économiquement la production agricole et la biodiversité. Sa gestion implique de maximiser l'hétérogénéité des pressions de sélection, notamment en combinant des substances actives (SA) de modes d'action différents. Or, le débat concernant l’efficacité des différentes stratégies pour retarder l'émergence et la sélection de la résistance dans les populations pathogènes reste ouvert. Cette thèse aborde deux questions : (i) Comment améliorer la performance des stratégies anti-résistance et quels sont les facteurs permettant leur optimisation ? (ii) Comment le statut initial de la résistance dans les populations module-t-il la performance des stratégies ? Pour illustrer ce débat, nous avons utilisé Zymoseptoria tritici, agent causal de la principale maladie du blé, résistant à la plupart des fongicides en Europe. Nous avons appliqué une approche innovante pour les champignons, l'évolution expérimentale, qui permet d’imiter et d’accélérer, en conditions contrôlées de laboratoire, la résistance aux fongicides habituellement sélectionnée au champ. En particulier, en soumettant un isolat sensible à des régimes de sélection hétérogènes par comparaison avec l'utilisation en séquence des mêmes SA, nous avons dissocié l'impact relatif de plusieurs facteurs sur la réduction de la sélection au sein de stratégies d'alternance, de mélange et de modulation de dose. Dans une première expérimentation, l'alternance s'est avérée être une stratégie bénéfique ou neutre pour diminuer le taux d'évolution de la résistance par rapport à une utilisation continue, mais a favorisé du généralisme. Nous avons démontré que ce compromis résidait probablement dans le risque de résistance intrinsèque des SA, plutôt que dans le nombre de fongicides utilisés ou le rythme de l'alternance. Dans une deuxième expérimentation, nous avons exploré comment un mélange à dose efficace, c'est-à-dire un mélange avec des doses réduites de ses composants permettant un contrôle de la maladie similaire à celui fourni par ses composants utilisés seuls, peut retarder l'évolution de la résistance d'une population sensible. Nous avons conclu qu’un tel mélange pouvait être, selon ses composants, préjudiciable à bénéfique pour la durabilité. Des résistances généralistes ou multiples ont plus ou moins été favorisées en fonction des combinaisons de SA. La réduction de la dose dans des séquences n'a pas empêché la sélection de souches spécifiques fortement résistantes, ni la résistance généraliste dans certaines lignées. Une dernière expérimentation a été conçue pour valider les résultats précédents en soumettant un isolat sensible et deux populations artificielles, contenant de faibles fréquences de résistances simples ou multiples, à des régimes de sélection contrastés par quatre sources d'hétérogénéité. Les résultats ont conclu que l'augmentation du nombre de SA et la variation des modes d'action étaient efficaces pour retarder la sélection de la résistance, que ce soit en mélange ou alternance. Toutefois cela faciliterait un certain généralisme des isolats sélectionnés, en plus des résistances spécifiques initialement introduites. Aussi, aucune de ces sources d'hétérogénéité ne s'est avérée efficace pour atténuer l'évolution de la résistance lorsque la résistance multiple était présente dans les populations. Cela suggère que la recombinaison, pouvant survenir lors de la reproduction sexuée au champ, est un facteur majeur à considérer. Enfin, la réduction de la valeur sélective associée à certains allèles résistants retarde légèrement leur sélection et peut moduler la structure des populations. L'évolution expérimentale s'avère être un outil puissant pour enrichir le débat sur les stratégies de lutte en analysant les interactions entre ses déterminants. Ces résultats ouvrent de nouvelles perspectives pour optimiser la gestion des résistances et suggèrent que la prévention de l'émergence serait plus durable que de limiter la sélection.