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Background: For Corynespora cassiicola (Berk. & M.A. Curtis) C.T. Wei, a necrotrophic pathogen with a broad host range and a worldwide distribution, resistance to fluopyram has been attributed to mutations in SdhB/C/D subunit of the succinate dehydrogenase (SDH) complex. In our previous study, two point mutations in SdhB from isoleucine to valine at position 280 (I280V) and histidine to tyrosine at position 278 (H278Y) showed different resistance phenotypes to fluopyram and boscalid. This research was conducted to explore the correlation between the mutation of SdhB-I280V or SdhB-H278Y and resistance to fluopyram or boscalid and its effect on the fitness characteristics of C. cassiicola., Results: The sdhB gene in a succinate dehydrogenase inhibitor (SDHI)-sensitive C. cassiicola strain (wild type) was successfully replaced with the mutant sdhB gene (GTT at position 280, SdhB-I280V) or with the mutant sdhB gene (TAC at position 278, SdhB-H278Y,). Compared with the wild-type strain, the replacement mutants exhibited significantly different resistance phenotypes, with SdhB-V280 demonstrating moderate resistance to fluopyram and low resistance to boscalid, while SdhB-Y278 was supersensitive to fluopyram and very highly resistant to boscalid. Both of the mutants exhibited decreased sensitivity to salt stress and increased SDH activity. These two mutations had no effect on the mycelial growth rate, sporulation ability, pathogenicity in vivo, sensitivity to osmotic stress and oxidative stress, cell wall damaging agents, or SHAM., Conclusion: Two adjacent mutations in the SdhB gene conferred different resistance phenotypes to SDHIs in C. cassiicola, which is important for the development of alternative antifungal fungicides and fluopyram resistance management. © 2021 Society of Chemical Industry., (© 2021 Society of Chemical Industry.)

Background: Net blotch caused by Pyrenophora teres is an important disease of barley worldwide. In addition to strobilurins (quinone ouside inhibitors) and azoles (demethylation inhibitors), succinate dehydrogenase inhibitors (SDHIs) are very effective fungicides for net blotch control. Recently, SDHI-resistant isolates have been found in the field. Intensive sensitivity monitoring programmes across Europe were carried out to investigate the situation concerning SDHI resistance in P. teres., Results: The first isolates with a lower sensitivity to SDHIs registered in barley were found in Germany in 2012 and carried the B-H277Y substitution in the succinate dehydrogenase enzyme. In 2013 and 2014, a significant increase in isolates with lower SDHI sensitivity was detected mainly in France and Germany, and the range of target-site mutations increased. Most of the resistant isolates carried the C-G79R substitution, which exhibits a strong impact on all SDHIs in microtitre tests. All SDHIs tested were shown to be cross-resistant. Other substitutions are gaining in importance, e.g. C-N75S in France and D-D145G in Germany. So far, no double mutants in SDH genes have been detected. Glasshouse tests showed that SDHI-resistant isolates were still controlled by the SDHI fluxapyroxad when applied preventively. To date, most isolates with C-G79R substitution have not simultaneously carried the F129L change in cytochrome b, which causes resistance towards QoI fungicides at low to moderate levels., Conclusion: Several target-site mutations in the genes of subunits SDH-B, SDH-C and SDH-D with different impact on SDHI fungicides were detected. The pattern of mutations varied from year to year and between different regions. Strict resistance management strategies are recommended to maintain SDHIs as effective tools for net blotch control, especially in areas with low frequencies of resistant isolates. © 2016 Society of Chemical Industry., (© 2016 Society of Chemical Industry.)

Research has established that mutations in highly conserved amino acids of the succinate dehydrogenase (SDH) complex in various fungi confer SDH inhibitor (SDHI) resistance. For Sclerotinia sclerotiorum (Lib.) de Bary, a necrotrophic fungus with a broad host range and a worldwide distribution, boscalid resistance has been attributed to the mutation H132R in the highly conserved SdhD subunit protein of the SDH complex. In our previous study, however, only one point mutation, A11V in SdhB (GCA to GTA change in SdhB), was detected in S. sclerotiorum boscalid-resistant (BR) mutants. In the current study, replacement of the SdhB gene in a boscalid-sensitive (BS) S. sclerotiorum strain with the mutant SdhB gene conferred resistance. Compared with wild-type strains, BR and GSM (SdhB gene in the wild-type strain replaced by the mutant SdhB gene) mutants were more sensitive to osmotic stress, lacked the ability to produce sclerotia and exhibited lower expression of the pac1 gene. Importantly, the point mutation was not located in the highly conserved sequence of the iron-sulfur subunit of SDH. These results suggest that resistance based on non-conserved vs. conserved protein domains differs in mechanism. In addition to increasing our understanding of boscalid resistance in S. sclerotiorum, the new information will be useful for the development of alternative antifungal drugs., (© 2014 BSPP AND JOHN WILEY & SONS LTD.)

The increasing global reliance on pesticides for agricultural pest control has raised significant environmental concerns, particularly due to inadequate monitoring of emerging chemicals in surface waters. This study addresses the potential contamination of aquatic ecosystems by developing and validating a method for detecting trace amounts of four recently registered fungicides: three succinate dehydrogenase inhibitors (fluopyram, penthiopyrad, pydiflumetofen) and fluopicolide, a structurally related fungicide. Employing QuEChERS-based sample extraction combined with ultra-high-performance liquid chromatography (UHPLC-MS-MS), this method achieves detection limits of 0.1 to 0.2 μg/L, with recovery rates between 90% and 110%, and intra-day relative standard deviation values well within the acceptable range of less than 20%. Applied to surface grab water samples from the greater Melbourne area, Australia, the method successfully identified all four fungicides at trace levels, including a notable high concentration of fluopyram (7.3 μg/L) during autumn, with the others intermittently detected at lower concentrations. This study represents the first documented instance of quantifiable detections of these four fungicides in Australian surface water systems. Given their high toxicity to several organisms and the limited global data on these substances, our findings underscore the critical need for continuous monitoring to inform strategies to safeguard aquatic ecosystems from these chemicals. [ABSTRACT FROM AUTHOR]

Succinate dehydrogenase inhibitor fungicides (SDHIs) are frequently detected in the marine environment. However, studies on the toxicity of SDHIs to marine organisms, Mytilus coruscus (M. coruscus), are poorly reported. Therefore, the antioxidant activities and metabolomic response of four SDHIs, namely, boscalid (BC), thifluzamide (TF), fluopyram (FO), and bixafen (BIX), to (M. coruscus), were comprehensively investigated. The antioxidant activity of BC and TF was significantly increased (p<0.05), whereas those of FO and BIX were significantly decreased. Furthermore, metabolite discriminations among M. coruscus to four SDHIs were illustrated by an untargeted metabolomics approach. A total of 52, 50, 93, and 129 differential metabolites were obtained for BC, TF, FO, and BIX. KEGG of the different metabolites show that the four SDHIs had differential effects on the metabolic pathways of M. coruscus. The current study demonstrated four SDHIs triggered glucose metabolism, lipid metabolism, tricarboxylic acid cycle, and oxidative phosphorylation processes and caused the disruption of nutrient and energy conversion processes in mussels. Finally, five biomarkers were screened by analyzing common differential metabolites that emerged from the four SDHI exposures, which could be used for risk assessment of marine ecosystem exposure to SDHIs. Our results demonstrated the use of metabolomics to understand the potential mechanisms of toxicity of four SDHIs to mussels and to identify potential targets for future targeted risk assessment. [ABSTRACT FROM AUTHOR]

Rice sheath blight caused by Rhizoctonia solani occurs worldwide and is mainly controlled by fungicides. SYP-32497 is a novel succinate dehydrogenase inhibitor, which interferes with the succinate ubiquinone reductase in the mitochondrial respiratory chain of fungi. This study aimed to evaluate the baseline sensitivity of R. solani from 13 major rice producing areas in China to SYP-32497 and fluxapyroxad. The study also explored the cause for the activity discrepancy between SYP-32497 and fluxapyroxad via an enzyme activity inhibition test and molecular docking. A total of 360 R. solani isolates were sensitive to SYP-32497 and fluxapyroxad. Baseline sensitivities were unimodally distributed with mean values of 0.00667 ± 0.00475 and 0.0657 ± 0.0250 μg mL−1, respectively, for SYP-32497 and fluxapyroxad. Enzyme activity assays and molecular docking results revealed that SYP-32497 exhibited a much higher SDH inhibition (IC50 = 0.300 μg mL−1) than to fluxapyroxad (IC50 = 1.266 μg mL−1) because of its excellent SDH binding ability via hydrogen bonding, π-cation, and hydrophobic interactions. These results suggest that SYP-32497 is a good suitable control agent for alternative rice sheath blight. [ABSTRACT FROM AUTHOR]

The soybean cyst nematode (SCN) (Heterodera glycines Ichinohe) is a significant yield-limiting factor in soybean production in the Midwestern US. Several management practices are implemented to mitigate yield losses caused by SCN, including using SDHI (succinate dehydrogenase inhibitors) fungicides delivered as seed treatments. A set of studies was conducted to evaluate the effect of two seed-applied succinate dehydrogenase inhibitors (SDHI) compounds, fluopyram and pydiflumetofen, on SCN population densities, plant injury, and plant growth. Cyst counts in untreated control and pydiflumetofen treated plants were 3.44 and 3.59 times higher than fluopyram, respectively, while egg counts were 8.25 and 7.06 times higher in control and pydiflumetofen. Next-generation sequencing was later employed to identify transcriptomic shifts in gene expression profiles in fluopyram and pydiflumetofen -treated seedlings. RNA expression patterns of seed treatments clustered by sampling time (5 DAP vs. 10 DAP); therefore, downstream analysis was conducted by timepoint. At 5 DAP, 10,870 and 325 differentially expressed genes (DEG) were identified in fluopyram and pydiflumetofen, respectively. These same treatments generated 219 and 2 DEGs at 10 DAP. Multiple DEGs identified in soybean seedlings treated with fluopyram are linked to systemic resistance, suggesting a potential role of systemic resistance in the suppression of SCN by fluopyram, in addition to the known nematicidal activity. The non-target inhibition of soybean succinate dehydrogenase genes by fluopyram may be the origin of the phytotoxicity symptoms observed and potentially the source of the systemic resistance activation reported in the current study. This work helps to elucidate the mechanisms of suppression of SCN by fluopyram. [ABSTRACT FROM AUTHOR]

Herein, a High-Throughput Semi-automated Emulsive Liquid-Liquid Microextraction (HTSA-ELLME) method was developed to detect Succinate Dehydrogenase Inhibitor (SDHI) fungicides in food samples via UHPLC-MS/MS. The Oil-in-Water (O/W) emulsion comprising a hydrophobic extractant and water was dilutable with the aqueous sample solution. Upon injecting the primary emulsion into the sample solution, a secondary O/W emulsion was formed, allowing SDHI fungicides to be extracted. Subsequently, a NaCl-saturated solution was injected in the secondary O/W emulsion as a demulsifier to rapidly separate the extractant, eliminating the need for centrifugation. A 12-channel electronic micropipette was used to achieve a high-throughput semi-automation of the novel sample pretreatment. The linear range was 0.003–0.3 μg L−1 with R 2 > 0.998. The limit of detection was 0.001 μg L−1. The HTSA-ELLME method successfully detected SDHI fungicides in water, juice, and alcoholic beverage samples, with recoveries and relative standard deviations of 82.6–106.9% and 0.8–5.8%, respectively. Unlike previously reported liquid-liquid microextraction approaches, the HTSA-ELLME method is the first to be both high-throughput and semi-automated and may aid in designing pesticide pretreatment processes in food samples. Schematic diagram of the HTSA-ELLME procedure. [Display omitted] • A novel Emulsive Liquid-Liquid Microextraction (ELLME) method was developed. • High-throughput semi-automated operation was achieved by an electronic micropipette. • The method only required a small amount of green solvent and no adsorption materials. • The NaCl solution was used as a demulsifier instead of centrifugation. • The simple, fast, green, and sensitive method was applied to five SDHI fungicides. [ABSTRACT FROM AUTHOR]

The evolution and spread of pan-azole resistance alleles in clinical and environmental isolates of Aspergillus fumigatus is a global human health concern. The identification of hotspots for azole resistance development in the wider environment can inform optimal measures to counteract further spread by minimizing exposure to azole fungicides and reducing inoculum build-up and pathogen dispersal. We investigated the fungicide sensitivity status of soil populations sampled from arable crops and the wider environment and compared these with urban airborne populations. Low levels of azole resistance were observed for isolates carrying the CYP51A variant F46Y/M172V/E427K, all belonging to a cluster of related cell surface protein (CSP) types which included t07, t08, t13, t15, t19, and t02B, a new allele. High levels of resistance were found in soil isolates carrying CYP51A variants TR34/L98H and TR46/Y121F/T289A, all belonging to CSP types t01, t02, t04B, or t11. TR46/Y121F/M172V/T289A/G448S (CSP t01) and TR46/Y121F/T289A/S363P/I364V/G448S (CSP t01), a new haplotype associated with high levels of resistance, were isolated from Dutch urban air samples, indicating azole resistance evolution is ongoing. Based on low numbers of pan-azole resistant isolates and lack of new genotypes in soils of fungicide-treated commercial and experimental wheat crops, we consider arable crop production as a coldspot for azole resistance development, in contrast to previously reported flower bulb waste heaps. This study also shows that, in addition to azole resistance, several lineages of A. fumigatus carrying TR-based CYP51A variants have also developed acquired resistance to methyl benzimidazole carbamate, quinone outside inhibitor and succinate dehydrogenase (Sdh) inhibitor fungicides through target-site alterations in the corresponding fungicide target proteins; beta-tubulin (F200Y), cytochrome b (G143A), and Sdh subunit B (H270Y and H270R), respectively. Molecular typing showed that several multi-fungicide resistant strains found in agricultural soils in this study were clonal as identical isolates have been found earlier in the environment and/or in patients. Further research on the spread of different fungicide-resistant alleles from the wider environment to patients and vice versa can inform optimal practices to tackle the further spread of antifungal resistance in A. fumigatus populations and to safeguard the efficacy of azoles for future treatment of invasive aspergillosis. [ABSTRACT FROM AUTHOR]

BACKGROUND A new generation of succinate dehydrogenase inhibitors (SDHIs) with high efficiency and broad‐spectrum antifungal activity has been frequently used in crop production. Sclerotinia stem rot is a major disease of various plants and crops caused by Sclerotinia sclerotiorum. Although benzovindiflupyr and isopyrazam reportedly have high activity against S. sclerotiorum, little is known about the bioactivity of different SDHIs classes against S. sclerotiorum or the mechanism of their differential antifungal activity. RESULTS: The in vitro tests revealed that the pyrazole‐4‐carboxamides of SDHIs (benzovindiflupyr, isopyrazam, fluxapyroxad, pydiflumetofen) had the highest activity against S. sclerotiorum followed by pyridine carboxamides (boscalid), pyridinyl‐ethyl benzamides (fluopyram) and thiazole carboxamides (thifluzamide), and of these thifluzamide showed poor antifungal activity with EC50 values greater than 6.01 mg L−1. The pyrazole‐4‐carboxamides of SDHIs showed satisfactory protective and curative activity against Sclerotinia stem rot. After treatment with the pyrazole‐4‐carboxamides of SDHIs, mitochondrial function in S. sclerotiorum decreased significantly. The enzyme activity assays revealed a lower affinity between thifluzamide and the Sc‐Sdh complex than was observed for the other six fungicides, with IC50 values ranging from 0.0036 to 1.2088 μmol L−1. Additionally, the docking positions of fungicides were similar, yet binding energies were different in the docking study with the Sdh complex. The correspondingly weaker hydrogen bonds may be responsible for the poor activity of thifluzamide against S. sclerotiorum. CONCLUSION: Understanding different binding features of various SDHIs classes with the Sc‐Sdh complex might be beneficial for the design and development of highly effective broad‐spectrum fungicides to ensure high yield and quality in crops by reducing fungicide use. © 2020 Society of Chemical Industry [ABSTRACT FROM AUTHOR]

Zymoseptoria tritici is the causal agent of septoria tritici blotch ( STB), a foliar wheat disease important worldwide. Succinate dehydrogenase inhibitors ( SDHIs) have been used in cereals for effective control of STB for several years, but resistance towards SDHIs has been reported in several phytopathogenic fungi. Resistance mechanisms are target-site mutations in the genes coding for subunits B, C and D of the succinate dehydrogenase ( SDH) enzyme. Previous monitoring data in Europe indicated the presence of single isolates of Z. tritici with reduced SDHI sensitivity. These isolates carried mutations leading to amino acid exchanges: C-T79N, C-W80S in 2012; C-N86S in 2013; B-N225T and C-T79N in 2014; and C-V166M, B-T268I, C-N86S, C-T79N and C-H152R in 2015. The current study provides results from microtitre and greenhouse experiments to give an insight into the impact of different mutations in field isolates on various SDHIs. In microtitre tests, the highest EC50 values for all tested SDHIs were obtained with mutants carrying C-H152R. Curative greenhouse tests with various SDHIs confirmed the findings of microtitre tests that isolates with C-H152R are, in general, controlled with lower efficacy than isolates carrying B-T268I, C-T79N and C-N86S. SDHI-resistant isolates of Z. tritici found in the field were shown to have cross-resistance towards all SDHIs tested. So far, SDHI-resistant isolates of Z. tritici have been found in low frequencies in Europe. Therefore, FRAC recommendations for resistance management in cereals, including a limited number of applications, alternation and combination with other MOAs, should be followed to prolong SDHI field efficacy. [ABSTRACT FROM AUTHOR]

ABSTRACT Succinate dehydrogenase inhibitor ( SDHI) fungicides constitute a relatively recent fungicide class registered for the treatment of grey mould on grapevine in Italy. The sensitivity profile to a novel compound fluopyram was established for a set of 203 Botrytis cinerea isolates collected from Sicilian vineyards within 2009-2012 prior its introduction into market. In addition, its performances were compared in in vitro and in vivo assays with other registered SDHI fungicide boscalid, to evaluate their frequency distributions EC50 values and cross-resistance patterns. Results of the article showed that EC50 values for fluopyram ranged from 0.05 to 1.98 µg mL−1. Although EC50 values of boscalid ranged from 0.01 to 89.52 µg mL−1, no cross-resistance relationship was observed between the two fungicides ( r = 0.003; P = 0.964) within our B. cinerea population. On further confirming these data, boscalid failed in controlling grey mould infections when boscalid-resistant isolates were inoculated on grape berries whereas fluopyram exhibited a good efficacy against the same isolates. This study represents the first report on the baseline sensitivity to fluopyram within B. cinerea population from Sicilian table grape vineyards in Italy, and it clearly shows the lack of cross-resistance in vitro and in vivo between fluopyram and boscalid for the field pathogen isolates. These results provided useful information for managing of fungicide resistance suggesting that fluopyram could be a valid alternative to boscalid for the control of grey mould of table grape. [ABSTRACT FROM AUTHOR]

Thifluzamide is a SDHI (succinate dehydrogenase inhibitor) fungicide, which interferes with succinate ubiquinone reductase in the mitochondrial electron transport chain of fungi. Presently, jinggangmycin is the major fungicide extensively used for the control of rice sheath blight caused by Rhizoctonia solani and resistance to jinggangmycin was first reported to occur in China. A total of 128 isolates of R. solani from Anhui Province of China were characterised for the baseline sensitivity to thifluzamide. The isolates were very sensitive to thifluzamide and the baseline sensitivity curve was unimodal with an average EC50 value of 0.058 ± 0.012 µg mL−1. However, EC50 values of boscalid (another SDHI fungicide) for inhibition of mycelial growth of 22 arbitrarily selected R. solani isolates ranged from 1.89 to 2.68 µg mL−1. Thifluzamide applied at 110 µg mL−1 exhibited excellent protective and curative activity against rice sheath blight and provided 81.1-91.0% protective or curative control efficacy. In field trials in 2010 and 2011, control efficacies of thifluzamide at 82 g.a.i ha−1 15 and 30 days after second application were 84.2% and 86.7%, respectively, suggesting excellent activity against sheath blight. There was a statistically significant difference in the efficacy between thifluzamide and boscalid or jinggangmycin. These results suggested that thifluzamide should be a good alternative fungicide to jinggangmycin for the control of rice sheath blight. [ABSTRACT FROM AUTHOR]