Grape breeding programs for resistance to mildew need to screen new varieties against a large and diverse pathogen population to ensure that resistance will be useful. We have assembled a diverse collection of mildew strains that are characterized by gene sequences and by variation in aggressiveness. Our collection has already proved its value for understanding the durability of powdery mildew resistance breeding. Several of the isolates we collected from the southeastern U.S. were able to overcome the resistance in grape varieties that have a resistance gene from muscadine grapes. Our isolates have allowed breeders to determine that this gene acts in a race-specific manner and therefore is not likely to remain durable.
Results from the study on diversity and population structure will also be useful for embarking on a study to find the genes responsible for variation in aggressiveness, host specialization and fungicide resistance. We have been able to determine that the eastern U.S. has a diverse native population that will be suitable for association mapping, using many of the same kinds of methods that have been employed in humans to find markers for various diseases. The difference is that we will be looking for markers in a fungus to understand what genes are causing them to be highly damaging (or not) to grapevines.
We will also be looking for genes involved in host specialization to muscadine grapes. This latter question is relevant to whether resistance derived from muscadine grapes in breeding programs will be stable and durable over time.
impact statement issue
Powdery mildew is economically the most important disease of grapes worldwide. Management of powdery mildew relies heavily on fungicide applications, with their attendant direct and environmental costs. Any knowledge about the biology of a pathogen has the potential to help optimize disease management techniques.
Studies of genetic variation of grape powdery mildew in Europe found that there were two distinct genetic groups that differ in their biology and ecology. One group tends to occur earlier in the season than the other. One produces more spores (for faster reproduction) but grows more slowly than the other. However, nothing was known about the diversity of this fungus in the U.S., where it is thought to be native.
The diversity of a pathogen is important to understand when breeders are trying to produce disease-resistant varieties. The problem often arises that a new variety may be resistant to some pathogen strains and not others. Our collections of powdery mildew isolates from throughout the eastern U.S. enabled grape breeders to test their resistant breeding lines against a diverse pathogen population. This resulted in discovering that grapes have race-specific powdery mildew resistance that is not likely to be durable if used in a large geographic area.
impact statement response
We have made considerable progress on studying genotypic and phenotypic variation in the grape powdery mildew fungus. First, we collected powdery mildew (or DNA from mildew) from across the eastern U.S., (including the following states: Connecticut, Georgia, Kansas, Michigan, North Carolina, New York, New Jersey, Pennsylvania, Ohio, Tennessee, Texas, and Virginia).
We also collected powdery mildew in California and obtained DNA samples from Oregon because these are such important grape-growing regions nationally. Colleagues from France, Italy, and Australia sent us DNA that they extracted from powdery mildew in their countries.
Second, we developed techniques for assessing genetic diversity by sequencing three genes. We also developed microsatellite markers derived from transcriptome sequences that were generated in collaboration with Lance Cadle-Davidson at the Grape Genetic Research Unit of the USDA-ARS in Geneva.
Third, for the powdery mildew that we isolated and can grow in the laboratory (on live grape leaves), we have tested for variation in virulence, i.e., the rate that mildew colonies grow and how long it takes them to produce spores (generation time), under controlled conditions.
We found much greater genetic diversity in mildew collected in the eastern U.S. than in other areas. We found no diversity and nearly identical strains of the fungus on the West Coast (California and Oregon) as in Europe and Australia. We interpret this result to mean that the California population was introduced from Europe on imported grapevines.
Populations in Europe and Australia comprise two very different genetic groups. A subpopulation related to one of these (group A) was also found in the southeastern U.S. Group B, which is found in California and Oregon and is more common in Europe and Australia than group A, was not found in the eastern U.S. sample. The eastern U.S. population is the most diverse and, consistent with previous assumptions, is probably the source population for introductions to Europe and the rest of the world.
We also found significant differences in aggressiveness in this fungus. Furthermore, mildew collected from muscadine grapes growing right next to other cultivated grapes could infect other grape species. However, no isolates collected on other grape species could infect muscadines. This indicates that the mildews may be specialized to different host species.
Finally, we studied the genetics of resistance to demethylation inhibiting fungicides (DMIs) by comparing resistance phenotypes to nucleotide sequences (Cyp51) and gene expression of genes known to be involved in DMI resistance. Our studies are showing novel mutations and correlations of expression to resistance. Additional work will be needed to dissect the genetics of this phenotype further.
impact statement summary
The grape powdery mildew fungus has long been thought to be native to the eastern U.S. This idea is based on historical reports of grape powdery mildew in the U.S. earlier than elsewhere and that the grape species native to the U.S. are more resistant to mildew than those in Europe or Asia.
We made the prediction that if this fungus is native to the U.S., then it would have higher genetic diversity here than in other places. To test this hypothesis, we collected mildew from across the eastern U.S., from New York, New England, Mid-Atlantic states, the Southeast, Midwest and as far south as Texas. We also collected mildew in California because it is such an important grape-growing region. Colleagues from Oregon, France, Italy, and Australia sent us DNA that they extracted from mildew in their areas.
By comparing nucleotide sequences of three different genes and genotyping fungal isolates with 10 other genetic markers, we found very little diversity in mildew from California, Oregon, Europe, and Australia. The California population had individuals almost identical to those found in Europe and Australia, probably because the fungus was introduced there from Europe on imported grapevines.
In contrast, the diversity in the eastern U.S. was quite high, consistent with this area being a native population. We also found significant variation in the aggressiveness of this fungus throughout the eastern U.S. and host specialization on muscadine grapes in the southeastern U.S.
