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Colletotrichum acutatum and C. gloeosporioides are fungal plant pathogens that have a global distribution, extensive host range, and convoluted taxonomy. Both species can cause grape ripe rot and are considered endemic to Virginia US. In 2012, C. acutatum and C. gloeosporioides were reclassified into species complexes that consist of 31 and 22 accepted species, respectively. The objectives of this study were to: 1) survey Virginia vineyards for grape ripe rot, and morphologically and phylogenetically identify isolates to the species within the complexes, 2) conduct an in vitro fungicide assay to screen fifteen commercial fungicides and combinations of two fungicides for efficacy to control isolates from seven Colletotrichum species from Virginia vineyards, 3) sequence gene fragments from three subunits of the SDH enzyme in the fungicide-screened isolates to observe potential resistance mutations, 4) investigate the susceptibility of three grapevine tissues to Colletotrichum species, 5) observe potential infection structures before and after the application of fungicides, 6) evaluate the efficacy of commercial fungicide controls of grape ripe rot in the field, and determine the most advantageous timing of applications. In my studies, I identified six Colletotrichum species: C. aenigma, C. conoides, C. fioriniae, C. gloeosporioides, C. kahawae, and C. nymphaeae. I also found two additional groups; an isolate similar to C. limetticola and C. melonis and a group of isolates that are similar to C. alienum, C. fructicola, and C. nupharicola. I also identified captan, and mancozeb as two potential active ingredients for control of grape ripe rot isolates from Virginia via the in vitro fungicide assay. Additionally, I found that combinations of two active ingredients could increase the efficacy of benzovindiflupyr, copper, and polyoxin-D. C. fioriniae germination and production of melanized appressoria was documented on leaves. I observed appressorium formation with isolates of two C. fructicola-like genotypes and C. nymphaeae, as well as secondary conidiation with isolates of C. aenigma, C. fructicola-like genotype 3, and C. nymphaeae on blooms. And finally, benzovindiflupyr, cyprodinil + fludioxonil pre-mix, and potassium phosphite + tebuconazole were identified as candidates for chemical control for grape ripe rot in the field. PHD Colletotrichum acutatum and C. gloeosporioides are two fungal plant pathogens that are found on a wide range of crops around the globe. Both fungal species cause the disease grape ripe rot and have been found in Virginia (VA) USA since the late 1800s. Originally, grape ripe rot was considered a minor disease in VA; however, based on communications with local VA vineyard managers, grape ripe rot was found to cause up to 30% direct crop loss. Further indirect economic loss occurs during wine production due to the production of unpalatable, tobacco-like, off flavors from the infected grapes. Sensory studies found this wine flavor change occurred with as little as 3% of the total crushed grapes being infected. Grape ripe rot appears as a sunburn-like, tan injury on the surface of white-fruited grape berries. As the disease progresses, the dark injury expands across the surface of the berry and rings of salmon-colored spore masses form. On red-fruited grapes, the formation of spore masses is usually the first observable sign. Over time, the infected berries will shrivel down to a soft, pustule-covered raisin. Both C. acutatum and C. gloeosporioides cause the same grape ripe rot symptoms on fruit and overlap in fungal appearance. In addition, investigations of these pathogens using molecular techniques have revealed that each consists of a number of genetically distinct groups that are difficult to distinguish by appearance. Therefore, in 2012, C. acutatum and C. gloeosporioides were reclassified into 31 and 22 newly accepted species, respectively, using molecular techniques. The objectives of this study were to: 1) survey VA vineyards for grape ripe rot, and visually and molecularly identify isolates to the species within the new complexes, 2) conduct a laboratory fungicide assay to screen fifteen commercial fungicides and combinations of two fungicides for control of isolates from VA vineyards, 3) sequence gene fragments from three subunits of the SDH enzyme in the fungicide-screened isolates to observe potential resistance mutations, 4) investigate the susceptibility of three grapevine tissues to Colletotrichum species, 5) observe potential infection structures before and after the application of fungicides, 6) evaluate the efficacy of commercial fungicide controls of grape ripe rot in the field, and determine the most advantageous timing of applications. In my studies, I identified six Colletotrichum species: C. aenigma, C. conoides, C. fioriniae, C. gloeosporioides¸ C. kahawae, and C. nymphaeae. I also found two additional groups; an isolate similar to C. limetticola and C. melonis and a group of isolates that are similar to C. alienum, C. fructicola, and C. nupharicola. Our lab also identified four active ingredients as potential controls of grape ripe rot in the laboratory fungicide assay; captan, mancozeb, tetraconazole and thiophanate-methyl. Additionally, combinations of two compounds can increase the effectiveness of benzovindiflupyr, copper, and polyoxin-D. of C. fructicola-like isolates, and C. nymphaeae formed infection structures on blooms. of C. aenigma, C. fructicola-like genotype 3, and C. nymphaeae formed spores on blooms without producing symptoms. C. fioriniae spores germinated and produced infection structures on leaves without producing symptoms.

Botrytis cinerea is the principal member of the species complex that causes bunch rot of grapes and gray mold disease on other hosts including fruits and ornamental crops. It has developed resistance to many fungicides, and isolates from eastern US strawberry fields have regularly been identified with resistance to several modes of action. During the 2011-2015 growing seasons, 487 isolates were collected from Virginia wine grapes, strawberries, and ornamental crops and evaluated for sensitivity to eight different fungicides by a germ tube elongation method; for a subset of isolates, a 24-well plate mycelial growth assay was also used, and baseline sensitivity to polyoxin-D was evaluated. Resistance to benzimidazoles and quinone outside inhibitors, and low-level resistance to iprodione were common. Boscalid resistance was common in wine grapes and ornamentals. Resistance to the hydroxyanilide fenhexamid during germ tube elongation was found in only 5% of wine grape isolates, but in 33% of isolates from strawberries and ornamentals. All of the fenhexamid-resistant isolates were identified as B. cinerea carrying various mutations in the erg27 gene. An additional subset of isolates was identified with moderate resistance to fenhexamid during mycelial growth, but not germination and germ tube growth. These were identified as B. cinerea HydR2 isolates, which possess an unknown mechanism of resistance towards fenhexamid in mycelial growth. Moderate resistance to cyprodinil was common, but in grape inoculation tests, moderately resistant isolates were controlled by a field rate of cyprodinil. Diminished sensitivity to fludioxonil and fluopyram was rare. Polyoxin-D controlled most isolates in mycelial growth tests at 100 µg/ml. Master of Science in Life Sciences