First Report of QoI Resistance in Plasmopara viticola from Vineyards of Maharashtra, India.

Abstract

Grape is an important fruit crop of India with about 1,235,000 MT of production a year from 111,000 ha. Maharashtra is the major grape-growing state, producing about 85% of the total production. Downy mildew caused by Plasmopara viticola (Berk. & Curt.) Berl. & de Toni is one of the most important diseases of grapes in the world and also in this area. QoI fungicides have been widely used in Maharashtra for downy mildew management for about a decade. Recently, grape growers reported failure of QoI fungicides in downy mildew control. Therefore, the present study was undertaken to check for the presence of QoI resistance in field isolates of P. viticola. During October to December 2014, fifteen downy mildew-affected samples were collected from seven vineyards in Maharashtra. Of these, seven samples were from Sangli, and five and three samples were from Nashik and Pune regions, respectively. One sample was also collected from Mizoram State in north India, where there is no history of QoI fungicide use. Sensitivity to QoI fungicide was determined using a modified 24-well leaf-disc bioassay (FRAC 2003). Healthy leaves were taken from the 6th node from the apex of a growing shoot of the susceptible cultivar Thomson Seedless and 15-mm disks were cut. The leaf disks were placed upside-down in wells containing 1 ml of 0.5% water agar amended with 0, 1, 10, 50, 100, and 1,000 µg/ml of azoxystrobin 23% SC (Amistar 23 SC Syngenta) or kresoxim methyl technical grade 94% w/w (Rallis India Ltd., Mumbai). Each treatment was replicated four times. Leaf disks were inoculated with 10 µl of a suspension containing 50,000 sporangia/ml of P. viticola collected from a single lesion. Plates were incubated at 22°C with alternating periods of 12 h light and dark. After six days, lesion area was measured and EC50 value was calculated by regression analysis of percent area of infection versus log10 fungicide concentration. A nested PCR-RFLP method was used to detect possible G143A mutation in the cytochrome b gene of these P. viticola isolates (Furuya et al. 2009). The EC50 of 5 isolates collected from vineyards with no QoI application during the season was ≤0.13 µg/ml to azoxystrobin and 0.03 µg/ml to kresoxim methyl, whereas the EC50 of 11 isolates from vineyards where 7 to 11 applications of QoI fungicides were made during the season was ≥110 µg/ml. In the QoI-insensitive isolates, the nested PCR product of 332 bp was digested by restriction enzyme Fnu4HI in two products of 134 and 198 bp, confirming resistance due to G143A point mutation (Gisi et al. 2002). This is first report of presence of QoI resistance in P. viticola in Maharashtra, India. Further monitoring of resistance to QoI fungicides in P. viticola population in this region is required.