Detection and Characterization of Carboxylic Acid Amide-Resistant Plasmopara viticola in China Using a TaqMan-MGB Real-Time PCR

Grape production is increasing globally and so are problems with downy mildew, one of the main constraints in grape production. Downy mildew on grape is caused by Plasmopara viticola, an obligate biotrophic pathogen belonging to the oomycetes. Control of the disease is usually performed by fungicide applications, of which carboxylic acid amide (CAA) fungicides represent one of the most widely used groups of fungicides. Our previous research showed that the extensive application of CAA fungicides can result in fungicide resistance and in China, CAA-resistant isolates of P. viticola were collected from the field in 2014. To monitor the distribution and spread of CAA fungicide resistance, we developed a TaqMan-minor groove binder (MGB) real-time PCR-based method designed on a functional mutation in the PvCesA3 gene that allows efficient identification of CAA fungicide resistant and sensitive genotypes. The assay was validated on 50 isolates using Sanger sequencing and fungicide bioassays and exploited in a comprehensive survey comprising 2,227 single-sporangiophore isolates from eight major grapevine regions in China. We demonstrate that CAA fungicide resistance in P. viticola is widespread in China. On average, 53.3% of the isolates were found to be resistant, but marked differences were found between locations with percentages of resistant isolates varying from 0.3 to 96.6%. Furthermore, the frequency of CAA-resistant isolates was found to be significantly correlated with the exposure to CAA fungicides (P < 0.05). We further discussed the possibilities to apply the TaqMan-MGB real-time PCR assay to assess the frequency of fungicide-resistant P. viticola isolates in each region or vineyard, which would facilitate the correct choice of fungicide for grape downy mildew and resistance management strategies.

First Report of Carboxylic Acid Amide Fungicide Resistance in Plasmopara viticola (Grapevine Downy Mildew) in North America

This report documents the first known occurrence in North America of resistance in grape downy mildew (Plasmopara viticola) to the carboxylic acid amide (CAA) fungicides mandipropamid and dimethomorph. These fungicides (FRAC group 40) have been an important component of downy mildew management programs for the past decade. Resistant isolates were obtained at three locations in Virginia and one in North Carolina, at considerable distances from each other. Resistance was documented by bioassay and the presence of the G1105S mutation, which has been associated with CAA resistance of P. viticola in other areas. Further survey is needed to determine the geographic extent of this resistance.

Emergence of Single Point Mutation in PvCesA3, Conferring Resistance to CAA Fungicides, in Plasmopara viticola Populations in Japan

As far as we know, there is no report of fungicide-resistant Plasmopara viticola in Japan. In the present study, we detected a single point mutation in the PvCesA3 of P. viticola sampled from a Japanese vineyard, which conferred resistance to mandipropamid. The emergence of the mandipropamid-resistant PvCesA3 allele may lead to the spread of mandipropamid-resistant P. viticola in vineyards all over Japan in the near future. Accepted for publication 23 April 2012. Published 29 July 2013.