Scion genetics and improvement: development of new disease-resistance varieties
Abstract
Fungicide applications for the control of the two most important diseases of winegrapes in Australia, powdery mildew and downy mildew, could be significantly reduced, or even eliminated, by the breeding of new winegrape varieties with reduced susceptibility to these pathogens. Using marker assisted selection technology, in combination with the rapid flowering microvine mutant, we have successfully combined powdery and downy mildew resistance loci from the wild North American species Muscadinia rotundifolia and the wild Chinese grape species Vitis romanetii to produce 2nd generation mildew-resistant premium winegrape varieties with stronger and more durable resistance to mildews.
Summary
The Wine Australia Strategic Plan 2015-2020 identified ’Increasing Competitiveness’ as a major priority to increase the prosperity of the Australian Wine Industry. One of the strategies identified to improve the performance of Australian vineyards was through the availability of new grapevine varieties that tolerate biotic stressors, generated through plant breeding programs, informed by molecular methods to identify desirable genotypic traits. Through previous co-investment projects with Wine Australia, CSIRO has successfully bred new elite white and red selections that are resistant to powdery and downy mildew. Selected lines of these 1st generation (gen.) mildew- resistant varieties are currently being evaluated for vine performance and wine quality at three regional sites as part of WA-CSIRO SRA project CSP 1701-1.6.
The 1st gen. mildew-resistant winegrape varieties contain a single gene conferring powdery mildew resistance (Run1) and a single gene conferring downy mildew resistance (Rpv1) introgressed from the wild North American grape species Muscadinia rotundifolia. Although these 1st gen. varieties have been shown to be highly resistant to powdery and downy mildew in unsprayed vineyard trials in Nuriootpa, Irymple and Wagga Wagga, we are aware that mildew isolates have been identified in other grape-growing countries that may break the resistance conferred by these genes. To ensure the durability of resistance in these new varieties, it is necessary to combine at least two different resistance genes (loci) within the same variety because the likelihood of the pathogens overcoming both resistance genes simultaneously is extremely low.
The wild Chinese grape species V. romanetii has been reported to contain a single dominant locus (Ren4) that confers strong resistance to powdery mildew. Our research has confirmed that powdery mildew isolates, that are capable of breaking the resistance conferred by Run1, are unable to overcome Ren4-mediated resistance making it a good candidate for ‘pyramiding’ with Run1 in 2nd gen. mildew-resistant varieties. Furthermore, we have discovered that the Ren4 powdery mildew resistance locus is co-located with a strong downy mildew resistance locus (designated Rpvrom) in the V. romanetii genome. This means that the Ren4 and Rpvrom loci co-segregate at a high frequency, effectively acting as a single locus in breeding populations, as in the case for the Run1/Rpv1 locus from M. rotundifolia.
On the basis of these discoveries, a breeding strategy (Strategy 1) was designed based on crosses between previously generated 1st gen. mildew-resistant varieties containing the Run1/Rpv1 locus and microvine breeding lines into which the Ren4/Rpvrom locus from V. romanetii had been introgressed through a number of rounds of backcrossing. Using marker-assisted selection (MAS) we were able to screen over 11,000 seedlings to identify over 400 new 2nd gen. mildew-resistant varieties that were not only positive for both the Run1/Rpv1 and Ren4/Rpvrom mildew resistance loci but also show four different types of berry characters - red skin, red flesh (tenturier), white skin and white skin with muscat character. All 2nd gen. varieties were successfully established in an unsprayed block of the SARDI vineyard in Nuriootpa for evaluation of viticultural and wine sensory properties.
Simultaneously with Strategy 1, an alternative breeding strategy (Strategy 3) was undertaken based on the use of a CSIRO female breeding line (15C002V0002) which is homozygous for the Run1/Rpv1 locus which was crossed with Ren4/Rpvrom microvine breeding lines. A total of 103 new 2nd gen. mildew-resistant winegrape varieties were selected using MAS from over 2,700 seedlings using breeding Strategy 3. However, approximately one third of the 2nd gen. varieties generated via Strategy 3 was found to exhibit an unusual phenotype in the vineyard, characterised by very large numbers of bunches per vine and flowers per bunch and severely inhibited shoot elongation making them unsuitable for commercial wine production.
To expand the genetic (varietal) diversity of the 2nd gen. mildew-resistant varieties, a revised breeding strategy (Strategy 2) was devised which utilised female breeding lines containing both the Run1/Rpv1 and Ren4/Rpvrom resistance loci that had been generated from Strategy 1 and Strategy 3 crosses. The availability of these new female 2nd gen. breeding lines enabled the use of any premium winegrape variety as a parent in the generation of mildew-resistant 2nd gen. progeny. Using Strategy 2, an additional 172 new 2nd gen. mildew-resistant winegrape varieties with either red skin, red flesh, white skin or white skin with muscat character were identified by MAS from the screening of over 4,000 seedlings. Significantly, these additional 172 varieties derived from crosses with a more diverse range of premium winegrape varieties that were not represented Strategy 1 or Strategy 3. To date, 65 Strategy 2 lines have been established in the SARDI vineyard with the aim to plant the remaining 107 lines in late 2022.
The enhanced resistance of 2nd gen. mildew-resistant varieties to powdery mildew and downy mildew, in comparison to 1st gen. mildew-resistant varieties, was demonstrated under identical conditions in laboratory assays. Furthermore, to the best of our knowledge these are the only winegrape varieties available anywhere in the world that combine the Run1/Rpv1 resistance locus from M. rotundifolia and the Ren4/Rpvrom resistance locus from V. piasezkii.
Fruit was harvested from a limited number of two and three year old 2nd gen. mildew-resistant varieties in season 2021/22. Harvest dates ranged from 25 February to 22 April and varieties displayed a range of bunch architectures from tight to loose. Sensory analysis undertaken on micro- scale ferments of fruit harvested from thirty-three 2nd gen. mildew-resistant varieties compared very favourably to wine made from five premium varieties sampled from the adjacent SARDI germplasm collection and fermented in the same way. Indeed, the average hedonic scale score calculated from a panel of four tasters, for wine from 28 of the 33 2nd gen. varieties was ranked equal or higher than the lowest-ranked premium variety and four 2nd gen. varieties produced wine of equal or higher preference to the highest ranked premium variety.
An additional aim was also initiated in Year 2 of the project to generate female breeding lines for use in future scion breeding projects that contain three different powdery mildew resistance loci (Run1, Ren4 & Ren7), together with the two downy mildew resistance loci (Rpv1 & Rpvrom). The Ren7 powdery mildew resistance locus from V. piasezkii was successfully introgressed into a microvine breeding line and then combined with the Run1/Rpv1 and Ren4/Rpvrom resistance loci. A total of 12 female 3rd gen. (Run1/Ren4/Ren7/Rpv1/Rpvrom) breeding lines were successfully identified by MAS. In addition, all of these 3rd gen. breeding lines are positive for the muscat locus and four lines also contain the red-flesh locus which provide more options for future breeding projects. Four 3rd gen. microvine breeding lines commenced flowering before the end of the current project and were used in crosses with pollen from premium winegrape varieties to generate seven new 3rd gen. mildew- resistant varieties for future field evaluation.
The results of this project demonstrate the power of combining marker-assisted selection technology with the rapid breeding microvine mutant, to generate premium winegrape varieties, in a reasonably short period of time, which are essentially V. vinifera-like in growth and sensory characteristics but contain new highly desirable production and quality traits.