Aggressiveness of Small-Spored Alternaria spp. and Their Sensitivity to Succinate Dehydrogenase Inhibitor Fungicides

Brown leaf spot of potato is caused by a number of small-spored Alternaria spp. Alternaria alternata sensu stricto, Alternaria arborescens, and Alternaria tenuissima have been reported with increasing frequency in commercial potato fields. Potato cultivars with resistance to small-spored Alternaria spp. have yet to be developed; therefore, the application of foliar fungicides is a primary management strategy. Greenhouse inoculation assays demonstrated that isolates of these three small-spored Alternaria spp. were pathogenic to potato. Significant differences in aggressiveness were observed across isolates; however, there was no trend in aggressiveness based on species. Significant fungicide by isolate interactions in in vitro fungicide sensitivity and significant differences between baseline and non-baseline isolates were observed in all three small-spored Alternaria spp. The ranges of in vitro sensitivity of A. alternata baseline isolates to boscalid (EC50 <0.010 to 0.89 µg/ml), fluopyram (<0.010 to 1.14 µg/ml) and solatenol (<0.010 to 1.14 µg/ml) were relatively wide when compared to adepidyn (<0.010 to 0.023 µg/ml). The baseline sensitivity of A. arborescens and A. tenuissima isolates to all four fungicides were less than 0.065 µg/ml. Between 10 and 21% of non-baseline A. alternata isolates fell outside the baseline range established for the four SDHI fungicides evaluated. In A. arborescens, 10 to 80% of non-baseline isolates had higher sensitivities than the baseline. A. tenuissima isolates fell outside the baseline for boscalid (55%), fluopyram (14%), and solatenol (14%) but none fell outside the baseline range for adepidyn. Evaluations of in vivo fungicide efficacy demonstrated that most isolates were equally controlled by the four SDHI fungicides. However, reduced boscalid efficacy was observed for four isolates (two each of A. arborescens and A. tenuissima) and reduced fluopyram control was observed in one A. alternata isolate. Results of these studies demonstrate that isolates of all three species could be contributing to the brown leaf spot pathogen complex and that monitoring both species diversity and fungicide sensitivity could be advantageous for the management of brown leaf spot in potatoes with SDHI fungicides.

Epidemiological Studies of Pan-Azole Resistant Aspergillus fumigatus Populations Sampled during Tulip Cultivation Show Clonal Expansion with Acquisition of Multi-Fungicide Resistance as Potential Driver

Pan-azole resistant isolates are found in clinical and environmental Aspergillus fumigatus (Af) populations. Azole resistance can evolve in both settings, with Af directly targeted by antifungals in patients and, in the environment, Af unintendedly exposed to fungicides used for material preservation and plant disease control. Resistance to non-azole fungicides, including methyl benzimidazole carbamates (MBCs), quinone outside inhibitors (QoIs) and succinate dehydrogenase inhibitors (SDHIs), has recently been reported. These fungicide groups are not used in medicine but can play an important role in the further spread of pan-azole resistant genotypes. We investigated the multi-fungicide resistance status and the genetic diversity of Af populations sampled from tulip field soils, tulip peel waste and flower compost heaps using fungicide sensitivity testing and a range of genotyping tools, including STRAf typing and sequencing of fungicide resistant alleles. Two major clones were present in the tulip bulb population. Comparisons with clinical isolates and literature data revealed that several common clonal lineages of TR34/L98H and TR46/Y121F/T289A strains that have expanded successfully in the environment have also acquired resistance to MBC, QoI and/or SDHI fungicides. Strains carrying multiple fungicide resistant alleles have a competitive advantage in environments where residues of multiple fungicides belonging to different modes of action are present.

Global Distributions of Clarireedia Species and their In Vitro Sensitivity Profiles to Fungicides

Dollar spot is reported to be caused by multiple Clarireedia species and is a serious problem on many turfgrasses around the world. To our knowledge, the distribution of different Clarireedia species and their sensitivity profiles to fungicides remains unknown. In this study, a total of 275 isolates were characterized by ITS sequence. Amounts of 124, 59 and 75 isolates were identified as C. jacksonii, C. monteithiana and C. paspali, respectively, while each species of C. homoeocarpa and C. bennettii had only five isolates. Four and three isolates were identified as two potential new species, which remained to be further characterized. C. jacksonii and C. monteithiana were distributed worldwide, while C. paspali was restricted to China. Of the isolates with host information, 81% (93/115) and 19% (22/115) of C. jacksonii isolates were collected from C3 and C4 plants, respectively, 97% (56/58) of the C. monteithiana isolates were collected from C4 plants and all C. paspali isolates were collected from C4 plants. The coexistence of different Clarireedia species on the same C4 host type in the same locales was found in Shanghai (Paspalum vaginatum), Jiangsu (Paspalum vaginatum) and Florida (Cynodon dactylon). The study revealed that differential fungicide sensitivity patterns were observed in different species in Clarireedia for the first time. Similar differential sensitivity profiles were also found in the locales with coexistence of at least two species. The findings from this study suggest that the adjacent coexistence of different Clarireedia species and the differential fungicide sensitivity profiles of different species will complicate dollar spot disease control.

Comparing the Fungicide Sensitivity of Sclerotinia sclerotiorum Using Mycelial Growth and Ascospore Germination Assays

The infection of the floral tissues of snap bean and other crops by Sclerotinia sclerotiorum, the causative agent of white mold, is by ascospores. Irrespective of the fungicide mode of action being evaluated, in vitro fungicide sensitivity tests are conducted almost exclusively using mycelial growth assays. This is likely due to difficulties and time involved in sclerotial conditioning required to produce apothecia and ascospores. The objective of this research was to compare estimates of fungicide sensitivity between mycelial growth and ascospore germination assays for S. sclerotiorum. Sensitivity assays were conducted using serial doses of three fungicides commonly used to control white mold: boscalid, fluazinam, and thiophanate-methyl. A total of 27 isolates were evaluated in replicated trials conducted for each fungicide and assay type. The effective concentration to reduce mycelial growth or ascospore germination by 50% (EC50) was estimated for each isolate, fungicide, assay type, and trial. The median EC50 values obtained from ascospore germination assays were 52.7, 10.0, and 2.7 times higher than those estimated from the mycelial growth for boscalid, fluazinam, and thiophanate-methyl, respectively. No significant correlation was found between EC50 values estimated by the two methods. These findings highlight differences that may be important in evaluating the sensitivity of S. sclerotiorum given the fungicide mode of action and how they will be used in the field.

Evidence for the Role of CYP51A and Xenobiotic Detoxification in Differential Sensitivity to Azole Fungicides in Boxwood Blight Pathogens

Boxwood blight, a fungal disease of ornamental plants (Buxus spp.), is caused by two sister species, Calonectria pseudonaviculata (Cps) and C. henricotiae (Che). Compared to Cps, Che is documented to display reduced sensitivity to fungicides, including the azole class of antifungals, which block synthesis of a key fungal membrane component, ergosterol. A previous study reported an ergosterol biosynthesis gene in Cps, CYP51A, to be a pseudogene, and RNA-Seq data confirm that a functional CYP51A is expressed only in Che. The lack of additional ergosterol biosynthesis genes showing significant differential expression suggests that the functional CYP51A in Che could contribute to reduced azole sensitivity when compared to Cps. RNA-Seq and bioinformatic analyses found that following azole treatment, 55 genes in Cps, belonging to diverse pathways, displayed a significant decrease in expression. Putative xenobiotic detoxification genes overexpressed in tetraconazole-treated Che encoded predicted monooxygenase and oxidoreductase enzymes. In summary, expression of a functional CYP51A gene and overexpression of predicted xenobiotic detoxification genes appear likely to contribute to differential fungicide sensitivity in these two sister taxa.

Identification of metalaxyl and ethaboxam insensitive Pythium sylvaticum pathogenic to pulse crops in Montana, USA.

Pythium root rot and damping-off caused by Pythium spp. are important diseases of pulse crops. In a 2016 pathogen survey from dry pea growing fields in Montana, along with commonly known causal agents P. ultimum and P. irregulare, an isolate identified as P. sylvaticum (LPPY17) was isolated from the rhizosphere of a diseased pea plant collected from Valley County, MT. Root rots and damping-off caused by P. sylvaticum have not previously been reported for chickpea, pea, and lentil crops. The isolate LPPY17 was tested for fungicide resistance in vitro, and results indicated a reduced sensitivity to metalaxyl and ethaboxam containing fungicides. LPPY17 was also tested for pathogenicity on chickpea, pea, and lentil seedlings in the greenhouse, and the results from the study revealed LPPY17 is capable of causing both root rots and damping off. Due to the potential pathogenicity and reduced fungicide sensitivity of this species, in the future it will be important to monitor for P. sylvaticum in pulse root rot surveys and diagnostics, as management options may be different from other common Pythium spp.

Review of Pythium species Causing Damping-off in Corn

Seedling blights and root rots caused by Pythium species account for almost twenty-five million dollars in annual losses to corn (Zea mays) production in the United States and Ontario. Variations in annual rainfall and increasing use of no-till agriculture can favor soilborne pathogens like Pythium. To date, forty-four species have been reported as pathogenic to corn in the United States. The average annual corn planting date in the United States has shifted approximately one week earlier in the past decade, exposing young corn plants to longer germination periods of generally cooler temperatures, favoring attack by Pythium. Optimal temperatures, aggressiveness, and response to chemical and biological treatment options vary by species. This review consolidates the species of Pythium reported as corn pathogens in literature to date. It also provides an insight into management strategies and discusses variations in fungicide sensitivity observed in corn-related Pythium species.

Assessing Missouri soybean fields for azoxystrobin resistant cercospora sojina

Successful management of Cercospora sojina, the causal agent of frogeye leaf spot, can be achieved through utilizing resistant varieties and fungicide applications. Fungicides in the quinone outside inhibitor (QoI) class are most effective in controlling C. sojina in the field. Unfortunately, azoxystrobin fungicide-resistant isolates of C. sojina have been recovered in many soybean growing areas of the U.S. Fungicide-resistant isolates of C. sojina were first detected in 2011 and 2012 in two counties in southeast Missouri, but no further assessment was conducted. In this study, 121 isolates were collected from 15 surveyed counties between 2019 and 2020 in an effort to understand the geographical distribution of fungicide-resistant C. sojina. Isolates were collected from fields based on the presence of frogeye leaf spot symptoms. Samples were brought to the laboratory and isolates were recovered from individual lesions. A fungicide sensitivity bioassay was conducted to determine which isolates were resistant to the QoI class of fungicides. The fungicide sensitivity bioassay consisted of full-strength PDA amended with technical grade azoxystrobin at five different concentrations and a no fungicide control. Out of 121 isolates, 81 were fungicide-resistant representing 13 of the 15 Missouri counties included in the survey. Notably, the northwest corner of Missouri had the highest concentration of fungicide-resistant isolates, consistent with the recent recovery of fungicide-resistant C. sojina isolates in Iowa and Nebraska. The widespread recovery of fungicide-resistant C. sojina in multiple counties throughout Missouri provides new insight into disease management in the state.