Fungicide Resistance in Powdery Mildew Fungi

Powdery mildew fungi (Erysiphales) are among the most common and important plant fungal pathogens. These fungi are obligate biotrophic parasites that attack nearly 10,000 species of angiosperms, including major crops, such as cereals and grapes. Although cultural and biological practices may reduce the risk of infection by powdery mildew, they do not provide sufficient protection. Therefore, in practice, chemical control, including the use of fungicides from multiple chemical groups, is the most effective tool for managing powdery mildew. Unfortunately, the risk of resistance development is high because typical spray programs include multiple applications per season. In addition, some of the most economically destructive species of powdery mildew fungi are considered to be high-risk pathogens and are able to develop resistance to several chemical classes within a few years. This situation has decreased the efficacy of the major fungicide classes, such as sterol demethylation inhibitors, quinone outside inhibitors and succinate dehydrogenase inhibitors, that are employed against powdery mildews. In this review, we present cases of reduction in sensitivity, development of resistance and failure of control by fungicides that have been or are being used to manage powdery mildew. In addition, the molecular mechanisms underlying resistance to fungicides are also outlined. Finally, a number of recommendations are provided to decrease the probability of resistance development when fungicides are employed.

Draft Genome Sequence Resource for Blumeriella jaapii, the Cherry Leaf Spot Pathogen

Blumeriella jaapii is the causal agent of cherry leaf spot (CLS), the most important disease of tart cherry in the Midwestern United States. Infection of leaves by B. jaapii leads to premature defoliation, which places trees at heightened risk of winter injury and death. Current management of CLS relies primarily on the application of three important fungicide classes, quinone outside inhibitors, sterol demethylation inhibitors, and succinate dehydrogenase inhibitors. Here, we present the first high-quality genome of B. jaapii through a hybrid assembly of PacBio long reads and Illumina short reads. The assembled draft genome of B. jaapii is 47.4 Mb and consists of 95 contigs with a N50 value of 1.5 Mb. The genomic information of B. jaapii, representing the most complete sequenced genome of the family Dermateaceae (Ascomycota) to date, provides a valuable resource for identifying fungicide resistance mechanisms of this pathogen and expands our knowledge of the phytopathogenic fungi in this family.

Paralogous CYP51 Genes of Colletotrichum spp. Mediate Differential Sensitivity to Sterol Demethylation Inhibitors

Colletotrichum spp. isolates contain two paralogous CYP51 genes that encode sterol 14-demethylase enzymes; however, their role in sensitivity to demethylation inhibitor (DMI) fungicides is yet to be determined. In this study, each of the two genes from Colletotrichum fioriniae and C. nymphaeae was able to rescue the function of CYP51 in the yeast Saccharomyces cerevisiae, demonstrating their independent function. Deletion of CYP51A led to increased sensitivity to propiconazole, diniconazole, prothioconazole, cyproconazole, epoxiconazole, flutriafol, prochloraz, and difenoconazole in C. fioriniae, and to the same fungicides and tebuconazole in C. nymphaeae, with the exception of prochloraz. Deletion of CYP51B in C. fioriniae and CYP51B in C. nymphaeae made mutants increasingly sensitive to five of nine DMI fungicides tested. The results suggest species-specific, differential binding of DMI fungicides onto the two CYP51 enzymes. Pairing DMIs with different effects on CYP51A and -B deletion mutants resulted in synergistic effects, as determined in mycelial growth inhibition experiments. Deletion mutants showed no fitness penalty in terms of mycelial growth, sporulation, and virulence. Our study elucidates the effect of CYP51A and CYP51B of Colletotrichum spp. on DMI sensitivity, suggesting that using a mixture of DMIs may improve the efficacy for anthracnose management.

Fitness of Erysiphe necator with G143A-Based Resistance to Quinone Outside Inhibitors

Management of grape powdery mildew (Erysiphe necator) using quinone outside inhibitors (QoIs) has eroded in an increasing number of regions due to resistance development. To determine persistence of resistance when QoIs are withdrawn, competition assays were conducted on unsprayed grape plants (Vitis vinifera ‘Chardonnay’) by cycling mixtures of resistant and sensitive isolates characterized as genetically diverse based on microsatellite analyses. Under laboratory conditions, %G143A, quantified by quantitative polymerase chain reaction (qPCR), increased significantly, indicating competitiveness of the resistant fraction. To confirm competitiveness in the field, trials using potted plants were conducted. Percent G143A tended to decrease in one growing season, probably due to spore migration and mixing of populations with natural background inoculum. In a second season, QoI resistance persisted at high frequency for 4 weeks. Resistant populations were also found to persist in one vineyard without QoI application for four consecutive years. The frequency was still about 25% in the fourth year, with higher frequency (36%) in a hotspot section. QoI-resistant populations with >5% G143A also harbored Y136F in the cyp51 gene that confers some resistance to sterol demethylation inhibitors, another fungicide class for powdery mildew control. Double resistance could have been partly responsible for persistence of QoI resistance at this location.

Emergence of Prochloraz-Resistant Populations of Calonectria pauciramosa and Calonectria polizzii in Ornamental Nurseries of Southern Italy

Management of Calonectria spp. infections in nurseries requires scheduled fungicide applications, particularly with methyl benzimidazole carbamates (MBCs) and sterol demethylation inhibitors (DMIs). Due to rising concerns about the occurrence of MBC resistance in different Calonectria populations and variability in prochloraz efficacy in controlling these pathogens, a detailed study on prochloraz sensitivity distributions of Calonectria isolates belonging to the Calonectria scoparia complex was carried out. In total, 105 isolates collected in two distinct periods (1993 to 1996 and 2005 to 2009) were analyzed for prochloraz sensitivity. Based on DNA sequencing and phylogenetic analyses of β-tubulin, histone H3, and translation elongation factor-1α gene sequences, 69 and 36 isolates were identified as C. pauciramosa and C. polizzii, respectively. The isolates collected more recently (group B) had a reduced prochloraz sensitivity, as indicated by greater values for the effective dose to reduce growth by 50% than those collected earlier (group A). The reduced sensitivity detected in vitro corresponded to partial loss of fungicide efficacy in controlling infections in red clover and feijoa under controlled and semi-field conditions, respectively. Frequent prochloraz application in nurseries for controlling Calonectria spp. infections is discouraged.