Constraining Evolution of Alternaria alternata Resistance to a Demethylation Inhibitor (DMI) Fungicide Difenoconazole.

Meng-Han He,Tian Wang,Yan-Ping Wang,Wen Zhu,Jiasui Zhan,E-Jiao Wu,Li-Ping Shang,Li-Na Yang,Lin-Lin Shen

doi:10.3389/fmicb.2019.01609

Abstract

Evolution of fungicide resistance in plant pathogens is one of major concerns in sustainable plant disease management. In this study, the genetics and potential of developing resistance to a demethylation inhibitor (DMI) fungicide, difenoconazole, in the fungal pathogen Alternaria alternata was investigated using a comparative analysis of genetic variation in molecular (Single Sequence Repeats, SSR) and phenotypic (fungicide tolerance) markers. No difenoconazole resistance was found in the 215 A. alternata isolates sampled from seven different ecological zones in China despite the widespread use of the fungicide for more than 20 years. This result suggests that the risk of developing resistance to difenoconazole in A. alternata is low and we hypothesize that the low risk is likely caused by fitness penalties incurred by resistant mutants and the multiple mechanisms involving in developing resistance. Heritability and plasticity account for ∼24 and 3% of phenotypic variation, respectively, indicating that genetic adaptation by sequence variation plays a more important role in the evolution of difenoconazole resistance than physiological adaptation by altering gene expression. Constraining selection in the evolution of A. alternata resistance to difenoconazole was documented by different patterns of population differentiation and isolate-by-distance between SSR markers and difenoconazole tolerance. Though the risk of developing resistance is low, the findings of significant differences in difenoconazole tolerance among isolates and populations, and a skewing distribution toward higher tolerance suggests that a stepwise accumulation of tolerance to the fungicide might be occurring in the pathogen populations. As a consequence, dynamic management programs guided by evolutionary principles such as spatiotemporal rotations of fungicides with different modes of action are critical to prevent the continued accumulation of tolerance or the evolution of resistance to difenoconazole and other DMI fungicides.

Highlights

Plant diseases caused by pathogenic fungi have been and continue to be one of the major factors threatening global food security and social stability and development (Brent and Hollomon, 2007; Savary et al, 2012; Chourasiya et al, 2013; Hahn, 2014)
The main objectives of the present study were to: (i) evaluate the potential for developing difenoconazole resistance in A. alternata, by quantifying genetic and environmental factors contributing to phenotypic variation of difenoconazole tolerance using a common garden experiment and (ii) determine the role of natural selection in the evolution of difenoconazole resistance by comparing the spatial distribution of genetic variation in SSR markers and difenoconazole tolerance
They were collected from plants separated by >100 cm and The A. alternata isolates were molecularly assayed with eight microsatellite markers (PAS1, PAS2, PAS3, PAS4, PAS5, PAS6, PAS7, and Ad8) previously and genotypes of the isolates were determined by GenClone 2.0 (Arnaud-Haond et al, 2007) using the allele information at each of the eight SSR loci

Summary

Introduction

Plant diseases caused by pathogenic fungi have been and continue to be one of the major factors threatening global food security and social stability and development (Brent and Hollomon, 2007; Savary et al, 2012; Chourasiya et al, 2013; Hahn, 2014). The application of fungicide benefits society directly and immediately by reducing food losses and indirectly and in the long term, by improving the quality and longevity of human life and supporting economic development (Cooper and Dobson, 2007). Due to their short generation time and large population size (Angelini et al, 2015; Delmas et al, 2017), fungal pathogens can rapidly evolve and adapt to meet changing environments including those imposed by the introduction of new fungicides in agricultural systems. Resistant pathotypes usually originate from mutation and/or recombination (sexual and asexual) in a single population, increase frequency through natural selection and migrate to other populations directly or via intermediary stepping-stone populations (Gisi et al, 2002; Angelini et al, 2015)

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Results