The fungal genus Colletotrichum includes numerous important plant pathogenic species and species complexes that infect a wide variety of hosts. Its taxonomy is particularly complex because species’ phenotypes and genotypes are difficult to differentiate. Two notable complexes, C. acutatum and C. gloeosporioides, are known for infecting temperate fruit crops worldwide. Even species within these complexes vary in traits such as tissue specificity, aggressiveness, geographic distribution, and fungicide sensitivity. With few effective chemicals available to control these pathogens, and the persistent threat of fungicide resistance, there is a need for greater understanding of this destructive genus and the methods that can be used for disease management. This review summarizes current research on diseases caused by Colletotrichum spp. on major fruit crops in the United States, focusing on the taxonomy of species involved, disease management strategies, and future management outlook.

Resistance in Monilinia fructicola to demethylation inhibitor (DMI) fungicides is beginning to emerge in North America, but its molecular basis is unknown. Two potential genetic determinants of DMI fungicide resistance including the 14α-demethylase gene (Mf CYP51 ) and the ATP-binding cassette transporter gene Mf ABC1 , were investigated in six resistant (DMI-R) and six sensitive (DMI-S) field isolates. No point mutations leading to an amino acid change were found in the Mf CYP51 gene. The constitutive expression of the Mf CYP51 gene in DMI-R isolates was significantly higher compared to DMI-S isolates. Gene expression was not induced in mycelium of DMI-R or DMI-S isolates treated with 0.3 μg of propiconazole/ml. A slightly higher average Mf CYP51 copy number value was detected in DMI-R isolates (1.35) compared to DMI-S isolates (1.13); however, this difference could not be verified in Southern hybridization experiments or explain the up to 11-fold-increased Mf CYP51 mRNA levels in DMI-R isolates. Analysis of the upstream nucleotide sequence of the Mf CYP51 gene revealed a unique 65-bp repetitive element at base pair position −117 from the translational start site in DMI-R isolates but not in DMI-S isolates. This repetitive element contained a putative promoter and was named Mona. The link between Mona and the DMI resistance phenotype became even more apparent after studying the genetic diversity between the isolates. In contrast to DMI-S isolates, DMI-R isolates contained an Mf CYP51 gene of identical nucleotide sequence associated with Mona. Still, DMI-R isolates were not genetically identical as revealed by Microsatellite-PCR analysis. Also, real-time PCR analysis of genomic DNA indicated that the relative copy number of Mona among DMI-S and DMI-R isolates varied, suggesting its potential for mobility. Interestingly, constitutive expression of the Mf ABC1 gene in DMI-R isolates was slightly lower than that of DMI-S isolates, but expression of the Mf ABC1 gene in DMI-R isolates was induced in mycelium after propiconazole treatment. Therefore, the Mf ABC1 gene may play a minor role in DMI fungicide resistance in M. fructicola . Our results strongly suggest that overexpression of the Mf CYP51 gene is an important mechanism in conferring DMI fungicide resistance in M. fructicola field isolates from Georgia and that this overexpression is correlated with Mona located upstream of the Mf CYP51 gene.

Botrytis cinerea, the causal agent of gray mold disease, is one of the most important plant-pathogenic fungi affecting strawberry. During the last decade, control of gray mold disease in the southeastern United States has largely been dependent on captan and the use of at-risk fungicides with single-site modes of action, including a combination of the quinone outside inhibitor (QoI) fungicide pyraclostrobin and succinate dehydrogenase inhibitor (SDHI) fungicide boscalid formulated as Pristine 38WG. Reports about loss of efficacy of Pristine in experimental fields in North Carolina prompted us to collect and examine 216 single-spore isolates from 10 conventional fields and 1 organic field in North Carolina and South Carolina in early summer 2011. Sensitivity to pyraclostrobin or boscalid was determined using a conidial germination assay with previously published discriminatory doses. Pyraclostrobin- and pyraclostrobin+boscalid-resistant isolates were found in all conventional fields (with some populations revealing no sensitive isolates) and in the organic field. Among the isolates collected, 66.7% were resistant to pyraclostrobin and 61.5% were resistant to both pyraclostrobin and boscalid. No isolates were identified that were resistant to boscalid but sensitive to pyraclostrobin, indicating that dual resistance may have derived from a QoI-resistant population. The molecular basis of QoI and SDHI fungicide resistance was determined in a subset of isolates. Polymerase chain reaction–restriction fragment length polymorphism analysis of the partial cytochrome b (CYTB) gene showed that pyraclostrobin-resistant isolates possessed the G143A mutation known to confer high levels of QoI fungicide resistance in fungi. Boscalid-resistant isolates revealed point mutations at codon 272 leading to the substitution of histidine to arginine (H272R) or tyrosine (H272Y), affecting the third Fe-S cluster region of the iron-sulfur protein (SdhB) target of SDHIs. The results of the study show that resistance to QoI fungicides and dual resistance to QoI and SDHI fungicides is common in B. cinerea from strawberry fields in the Carolinas. Resistant strains were more frequent in locations heavily sprayed with QoI and SDHI fungicides. However, resistance to both fungicides was also found in the unsprayed, organic field, indicating that some resistant strains may have been introduced from the nursery.

The sensitivity to seven chemical classes of fungicides was investigated in 1,810 Botrytis cinerea isolates collected from strawberry blossoms and fruit in 181 strawberry fields from seven southern states in the United States across 2 years. Ten isolates were examined from each field. Fungicide sensitivity assays were carried out based on visual assessment of diametrical mycelial growth after 4 days of incubation on media amended with discriminatory doses of fungicides in microtiter plates. Results of visual assessments were verified with selected isolates using a previously published germination assay and by inoculating representative isolates with resistant phenotypes on fungicide-sprayed fruit. The overall resistance frequencies of 750 isolates collected in 2012 for thiophanate-methyl, pyraclostrobin, boscalid, cyprodinil, fenhexamid, iprodione, and fludioxonil were 76, 42, 29, 27, 25, 3, and 1%, respectively. Frequencies of 1,060 isolates collected in 2013 were 85, 59, 5, 17, 26, 2, and 1%, respectively. Resistance to thiophanate-methyl and pyraclostrobin was found in virtually every location in both years, whereas resistance to iprodione and fludioxonil was rarely found. Resistant isolates were resistant to either one (23%), two (18%), three (19%), four (14%), five (3%), or six (0.1%) chemical classes of fungicides in 2012. In 2013, this distribution was 24, 29, 26, 8, 2, and 0.3%, respectively. Multifungicide-resistant isolates of B. cinerea were widespread in southern states and evidence suggests that the frequency of isolates with multifungicide resistance increased from 2012 to 2013. The data also show that fungicide resistance in B. cinerea was already present in blossoms, indicating that resistance management needs to be implemented early in the season.

Single-spore isolates of Monilinia fructicola were collected from commercial orchards in South Carolina and Georgia with prolonged past exposure to demethylation inhibitor (DMI) fungicides and from an orchard with no DMI history (baseline population). Sensitivity to propiconazole was determined using the concentration in agar media required to suppress radial growth of mycelium by 50% (EC₅₀. Mean EC₅₀ values from six South Carolina populations were not different from the baseline population (P < 0.05). Two of five populations from Georgia revealed (significantly higher mean EC₅₀ values compared with the baseline population (P < 0.05). Isolates with high (AP5 and AP6) and low (DL71 and DL72) EC₅₀ values were selected to determine disease incidence on peach fruit after protective or curative applications of propiconazole at 0.15 or 0.3 liter/ha (half and full label rate, respectively). Disease incidence was significantly greater on peaches inoculated with AP5 and AP6 after curative treatment with propiconazole at 0.15 liter/ha (P < 0.05). Following protective or curative treatments at 0.3 liter/ha, disease incidence was significantly greater for AP6 but not for AP5. These results suggest that a shift toward reduced sensitivity has developed in some M. fructicola populations from Georgia, and that isolates with reduced sensitivity to propiconazole are more difficult to control in the field. Field testing of DMI fungicides, captan, Q_oI fungicides, and fenhexamid in experimental orchards) indicated that the DMI fungicides are still among the most efficacious products for brown rot (control, and that new products containing Q_oI fungicides may be viable disease control alternatives or rotation partners.