Fungicides Boscalid Research Articles

Agricultural fungicides inadvertently influence the fitness of Colorado potato beetles, Leptinotarsa decemlineata, and their susceptibility to insecticides

The Colorado potato beetle (CPB), Leptinotarsa decemlineata (Say), is an agricultural pest of solanaceous crops which has developed insecticide resistance at an alarming rate. Up to this point, little consideration has been given to unintended, or inadvertent effects that non-insecticide xenobiotics may have on insecticide susceptibility in L. decemlineata. Fungicides, such as chlorothalonil and boscalid, are often used to control fungal pathogens in potato fields and are applied at regular intervals when L. decemlineata populations are present in the crop. In order to determine whether fungicide use may be associated with elevated levels of insecticide resistance in L. decemlineata, we examined phenotypic responses in L. decemlineata to the fungicides chlorothalonil and boscalid. Using enzymatic and transcript abundance investigations, we also examined modes of molecular detoxification in response to both insecticide (imidacloprid) and fungicide (boscalid and chlorothalonil) application to more specifically determine if fungicides and insecticides induce similar metabolic detoxification mechanisms. Both chlorothalonil and boscalid exposure induced a phenotypic, enzymatic and transcript response in L. decemlineata which correlates with known mechanisms of insecticide resistance.

Is pollinator visitation of Helianthus annuus (sunflower) influenced by cultivar or pesticide treatment?

Beneficial insects, such as Apis mellifera (honey bees) and Bombus sp. (bumble bees), play important roles in the environment as pollinators, and they are directly affected by agronomical practices, such as pesticide spraying or the selection of crop cultivar. Sunflower is an important resource for these pollinators, and its yield is highly dependent on pollination. Six sunflower hybrid cultivars (ES Biba, Gonzalo, Drake, Vellox, NK Neoma, and P63LE10) were used to compare the visitation rates (VR) of pollinators during flowering in three years (2015, 2016, and 2017). From this set, cultivar P63LE10 was selected to evaluate how pesticide treatments (mixture of two insecticide products containing acetamiprid and pirimicarb, respectively, and two fungicide products containing propiconazole plus prochloraz and dimoxystrobin plus boscalid, respectively) affect pollinator visitation. The amount of pesticide residue in sunflower inflorescences was analyzed to determine the content over the duration of the experiments. The sunflower cultivar P63LE10 was significantly the most attractive to both groups of pollinators in 2017. This cultivar was visited by the highest number of honey bees also in the other years. For bumble bees, cultivar P63LE10, along with cultivars NK Neoma and Vellox, had the highest visitation rates in 2015 and 2016. In 2017, plants treated by the fungicide dimoxystrobin plus boscalid had the highest visitation rates for both groups of pollinators. For bumble bees plants treated with this fungicide were significantly more visited (VR 23.1 ± 2.1%) than these treated with insecticide with pirimicarb (16.4 ± 2%) and the untreated control (17.9 ± 1.7%), and for honey bees plants treated with dimoxystrobin plus boscalid were significantly more visited (VR 24 ± 4.5%) than all other treatments except with acetamiprid (20.5 ± 3.5%). The residues of tested pesticides were found in blooms for the whole time of the experiment (range of values for 2016 and 2017 [ng/g]: acetamiprid 144–6, 3360–56; pirimicarb 177–12, 223–10; prochloraz 380–6, 1970–912; propiconazole 324–17, 1024–1018; boscalid 216–154, 9600–1319; dimoxystrobin 452–137, 6560–911). The influence of cultivar was more important than pesticide treatment in the selection of a preferred food source by pollinators.

Thioredoxin and Glutaredoxin Systems Required for Oxidative Stress Resistance, Fungicide Sensitivity, and Virulence of Alternaria alternata.

This study determined the function of thioredoxin and glutaredoxin systems in the phytopathogenic fungus Alternaria alternata via analyzing mutants obtained from the targeted deletion of genes encoding thioredoxin peroxidase (Tsa1), thioredoxin reductase (Trr1), and glutathione reductase (Glr1). Trr1 and Glr1, but not Tsa1, are required for growth and conidiation. The reduced growth and conidiation seen in the Trr1 or Glr1 deletion mutant can be restored by glutathione. Deletion mutants showing growth inhibition by oxidants are defective for H2O2 detoxification and induce smaller lesions on citrus leaves. Trr1 and Glr1, but not Tsa1, also contribute to NaCl resistance. Glr1 is required for sorbitol resistance and is responsible for resistance to mancozeb and boscalid but not chlorothalonil fungicides, a novel phenotype that has not been reported in fungi. Trr1 is required for resistance to boscalid and chlorothalonil fungicides but confers susceptibility to mancozeb. The Tsa1 deletion mutant displays wild-type sensitivity to the tested fungicides. The expression of Tsa1 and Trr1 is regulated by the oxidative stress responsive regulators Yap1, Hog1, and Skn7. The expression of Tsa1, but not Trr1, is also regulated indirectly by the NADPH oxidase. The results indicate that the capability to resist oxidative stress is required for virulence of A. alternataIMPORTANCE The thioredoxin and glutaredoxin systems are important thiol antioxidant systems in cells, and knowledge of these two systems in the plant-pathogenic fungus A. alternata is useful for finding new strategies to reduce the virulence of this pathogen. In this study, we demonstrated that thiol antioxidant system-related genes (Tsa1, Trr1, and Glr1) are required for H2O2 detoxification and virulence in A. alternata Moreover, deletion of Trr1 results in hypersensitivity to the fungicides chlorothalonil and boscalid, and Glr1 deletion mutants are highly sensitive to mancozeb, which is the fungicide mostly used in citrus fields. Therefore, our findings demonstrate that the ability to detoxify reactive oxygen species (ROS) plays a critical role in pathogenesis on citrus and provide novel insights into the physiological functions of thiol-containing systems in fungicide sensitivity for A. alternata.

Baseline sensitivity and toxic actions of boscalid against Sclerotinia sclerotiorum

Baseline sensitivity of Sclerotinia sclerotiorum to the succinate dehydrogenase inhibitor (SDHI) fungicide boscalid was established by determining the effective concentration causing mycelial growth inhibition by 50% (EC50) values. The mean EC50 values of 76 isolates collected in 2008 and 77 isolates collected in 2014 were 0.0383 and 0.0395 μg/mL, respectively. There was no significant difference (P = 0.695) in boscalid sensitivity between the two years, and the frequency distribution of EC50 values was unimodal, but right-skewed. Correlation analysis showed that there was a statistically significant positive correlation (R2 = 0.856, P<0.001) of sensitivity between boscalid and thifluzamide, but no significant correlation (R2 = 0.01, P = 0.650) was detected between boscalid and the demethylation inhibitor (DMI) fungicide prochloraz. Both preventive and curative efficacies of boscalid were significantly higher (P<0.05) than those of the reference fungicide carbendazim. Boscalid in potato dextrose agar (PDA) at 0.04–0.16 μg/mL significantly reduced the number of sclerotia per plate, but the weight of sclerotia increased slightly. Light microscopic observations showed that boscalid in PDA caused hyphae to branch more at the tip. Transmission electron microscopy (TEM) observations showed that mitochondria were deformed for the mycelia grown on boscalid-amended PDA. Boscalid at 0.01 or 0.05 μg/mL had no significant effect on mycelial cell membrane permeability of S. sclerotiorum. These results will deepen our understanding of the toxic action of boscalid, and the baseline sensitivity can be used as a reference point in future boscalid resistance monitoring programs.

Multiple nucleobase transporters contribute to boscalid sensitivity in Aspergillus nidulans

The development of fungicide-resistant fungal populations represents a major challenge for the agrochemical and agri-food sectors, which threatens food supply and security. The issue becomes complex for fungi that cause quantitative and qualitative losses due to mycotoxin biosynthesis. Nonetheless, currently, the molecular details underlying fungicide action and fungal resistance mechanisms are partially known. Here, we have investigated whether plasma membrane transporters contribute to specific fungicide uptake in the model fungus Aspergillus nidulans. Independent physiological tests and toxicity screening of selected fungicides provided evidence that the antifungal activity of Succinate Dehydrogenase Inhibitors (SDHIs) is associated with the expression of several nucleobase-related transporters. In particular, it was shown that a strain genetically inactivated in all seven nucleobase-related transporters is resistant to the fungicide boscalid, whereas none of the single null mutants exhibited significant resistance level. By constructing and testing isogenic strains that over-express each one of the seven transporters, we confirmed that five of them, namely, UapC, AzgA, FycB, CntA, and FurA, contribute to boscalid uptake. Additionally, by employing metabolomics we have examined the effect of boscalid on the metabolism of isogenic strains expressing or genetically lacking boscalid-related nucleobase transporters. The results confirmed the involvement of specific nucleobase transporters in fungicide uptake, leading to the discovery of corresponding metabolites-biomarkers. This work is the first report on the involvement of specific transporters in fungicide uptake and toxicity and their impact on fungal metabolism regulation and results might be further exploited towards the deeper understanding of fungal resistance to fungicides.

Biological Control of Botrytis cinerea: Interactions with Native Vineyard Yeasts from Washington State.

Native yeasts are of increasing interest to researchers, grape growers, and vintners because of their potential for biocontrol activity and their contributions to the aroma, flavor, and mouthfeel qualities of wines. To assess biocontrol activity, we tested 11 yeasts from Washington vineyards, representing isolates of Candida saitoana, Curvibasidium pallidicorallinum, Metschnikowia chrysoperlae, M. pulcherrima, Meyerozyma guilliermondii, Saccharomyces cerevisiae, and Wickerhamomyces anomalus, for ability to colonize Thompson Seedless grape berries, inhibit the growth of Botrytis cinerea in vitro, and suppress disease symptoms on isolated berries. The yeast-like fungus Aureobasidium pullulans was also included based on its known biocontrol activity against B. cinerea in studies on apple and grape. All yeast strains multiplied rapidly in grape berries and reached densities of over log 6 cells per wound as early as 2 days after inoculation with 200 cells. One of the Botrytis isolates used in this study was much less virulent than the others and was provisionally identified as B. prunorum based on multilocus sequence analysis. Suppression of the growth of B. cinerea isolates 111bb, 207a, 207cb, and 407cb occurred on berries treated with A. pullulans P01A006, Metschnikowia chrysoperlae P34A004 and P40A002, M. pulcherrima P01A016 and P01C004, Meyerozyma guilliermondii P34D003, and S. cerevisiae HNN11516. Inhibition of Botrytis isolates by the yeast strains was more common on berries than in vitro, suggesting the possibility that niche competition was a more likely biocontrol mechanism than antibiosis in planta. Metabolic profiling of yeast strains and B. cinerea isolates using Biolog YT plates revealed seven distinct metabolic groups. Furthermore, the yeast strains showed partial to complete tolerance to the commonly used fungicides fluopyram, triflumizole, metrafenone, pyraclostrobin, and boscalid. Implications of these findings for field deployment of native Washington yeasts as biocontrol agents against B. cinerea are discussed.

Sensitivity of Phacidiopycnis spp. Isolates from Pome Fruit to Six Pre- and Postharvest Fungicides.

Phacidiopycnis washingtonensis and P. pyri cause speck rot and Phacidiopycnis rot on apple and pear, respectively. Infection occurs in the orchard and remains latent, and symptoms appear after months of storage. Decay management relies on orchard sanitation and pre- and postharvest fungicides. In a 2017 survey, speck rot accounted for 6.4% of apple decay in central Washington, whereas Phacidiopycnis rot accounted for 3.9 and 6.7% of total pear decay in Washington and Oregon, respectively. Sensitivities of baseline populations of 110 P. washingtonensis and 76 P. pyri isolates collected between 2003 and 2005 to preharvest fungicides pyraclostrobin (PYRA) and boscalid (BOSC) and to postharvest fungicides thiabendazole (TBZ), fludioxonil (FDL), pyrimethanil (PYRI), and difenoconazole (DFC) were evaluated using a mycelial growth inhibition assay. Mean effective concentrations necessary to inhibit 50% growth (EC50) of P. washingtonensis were 0.1, 0.3, 0.8, 1.8, 2.1, and 4.8 µg/ml for FDL, PYRI, TBZ, DFC, PYRA, and BOSC, respectively. Respective mean EC50 values for P. pyri were 0.2, 0.6, 1.6, 1.1, 0.4, and 1.8 µg/ml. The sensitivity of exposed P. washingtonensis and P. pyri populations collected in 2017 revealed potential shifts toward BOSC and PYRA resistance. The efficacy of the six fungicides to control isolates of each pathogen with different in vitro sensitivity levels was evaluated on apple and pear fruit. FDL, DFC, and PYRI controlled both Phacidiopycnis spp. regardless of their EC50 values after 5 months of storage at 0°C in a regular atmosphere. The consistent occurrence of Phacidiopycnis spp. will require continuous monitoring and development of disease management strategies based on fungicide phenotypes and efficacy of existing fungicides assessed herein.

Synthesis of Boscalidviaa three-step telescoped continuous flow process implemented on a MJOD reactor platform

A three step continuous/semi-flow process leading to the fungicide Boscalid® is disclosed.

Behavioral responses of honey bees (Apis mellifera) to natural and synthetic xenobiotics in food

While the natural foods of the western honey bee (Apis mellifera) contain diverse phytochemicals, in contemporary agroecosystems honey bees also encounter pesticides as floral tissue contaminants. Whereas some ubiquitous phytochemicals in bee foods up-regulate detoxification and immunity genes, thereby benefiting nestmates, many agrochemical pesticides adversely affect bee health even at sublethal levels. How honey bees assess xenobiotic risk to nestmates as they forage is poorly understood. Accordingly, we tested nine phytochemicals ubiquitous in nectar, pollen, or propolis, as well as five synthetic xenobiotics that frequently contaminate hives—two herbicides (atrazine and glyphosate) and three fungicides (boscalid, chlorothalonil, and prochloraz). In semi-field free-flight experiments, bees were offered a choice between paired sugar water feeders amended with either a xenobiotic or solvent only (control). Among the phytochemicals, foragers consistently preferred quercetin at all five concentrations tested, as evidenced by both visitation frequency and consumption rates. This preference may reflect the long evolutionary association between honey bees and floral tissues. Of pesticides eliciting a response, bees displayed a preference at specific concentrations for glyphosate and chlorothalonil. This paradoxical preference may account for the frequency with which these pesticides occur as hive contaminants and suggests that they present a greater risk factor for honey bee health than previously suspected.

Effects of the fungicides azoxystrobin, pyraclostrobin and boscalid on the physiology of Japanese cucumber

Abstract Strobilurins are fungicides with beneficial physiological effects on crop yield due to promotion of net carbon assimilation, nitrate reductase enzyme activity, stress tolerance and hormonal balance. The carboxamides complement the action of the strobilurins when applied alternately or together. This study aimed to evaluate the effect of application of pyraclostrobin, azoxystrobin and boscalid on grafted and ungrafted Japanese cucumber (Cucumis sativus L.) in order to analyze the effects of these fungicides on plant physiology and increased fruit production. The experimental design was completely randomized in a 2 × 5 factorial scheme, with grafted and ungrafted cucumber plants and 5 fungicide treatments: control; azoxystrobin 60 g a.i ha−1; boscalid 50 g a.i ha−1; pyraclostrobin 50 g a.i ha−1; and boscalid 100 g a.i ha−1 + pyraclostrobin 50 g a.i ha−1, applied 18 days after transplanting and then at intervals of seven days. The effect of the treatments was evaluated based on the average yield and fruit mass, in addition to observations of gas exchange, lipid peroxidation, and the activities of peroxidase, superoxide dismutase, catalase and nitrate reductase. Based on the results obtained, it was concluded that the fungicide treatments resulted in different responses between the grafted and ungrafted plants. The physiological benefits were more evident in the grafted plants treated with pyraclostrobin or boscalid alone, and these benefits manifested in terms of fruit production and increased the activity of the antioxidative system, thereby reducing stress. The higher productivity probably occurred due to the better physiological performance of these plants, mainly at the beginning of development, presenting greater activity of the enzyme nitrate reductase, in addition to the higher net CO2 assimilation and carboxylation efficiency.

Sensitivity of &lt;i&gt;Botrytis cinerea&lt;/i&gt; to fungicides used in New Zealand wine grape spray programmes

Fungicide resistance development in Botrytis cinerea threatens the efficacy of anilinopyrimidine (AP) fungicides, which have been used for botrytis bunch rot control in New Zealand vineyards for about 20 years. A 2016 survey of 33 vineyards in Gisborne, Hawke's Bay and Marlborough showed that the majority of 670 isolates tested in agar-based assays were sensitive to the AP cyprodinil (EC50 &lt; 1 mg/litre), although 18% showed low-resistance (EC50 1-10 mg/litre) and 10% showed medium-resistance (EC50 &gt;10 mg/litre). There was high cross-resistance to the APs cyprodinil and pyrimethanil. The survey also established baseline sensitivity to the succinate dehydrogenase inhibitor (SDHI) fungicide boscalid and &gt;90% of isolates were sensitive (EC50 &lt; 5 mg/litre). There was a low degree of cross-resistance between the SDHIs boscalid and fluopyram. There was also high sensitivity to fludioxonil (0.009 to 0.018 mg/litre) and to fenhexamid (0.49 to 0.76 mg/litre).

Investigating Selection of Demethylation Inhibitor Fungicide‐Insensitive Sclerotinia homoeocarpa Isolates by Boscalid, Flurprimidol, and Paclobutrazol

Dollar spot, caused by Sclerotinia homoeocarpa F.T. Bennett, is the most economically important disease of golf course turfgrass in the northern United States. Fungicide resistance, especially to demethylation inhibitor (DMI) fungicides, is common for S. homoeocarpa. Plant growth regulators (PGRs) are commonly applied on golf courses and have a similar chemistry to DMIs. Previous research has suggested multiple resistance among the PGRs flurprimidol and paclobutrazol, the succinase dehydrogenase inhibitor fungicide boscalid, and the DMI propiconazole, with evidence for the involvement of the multidrug efflux transporter gene, ShPDR1. The objective of this research was to investigate if flurprimidol, paclobutrazol, and boscalid select for DMI insensitivity in the field. We tested eight treatments: flurprimidol alone, tank‐mixed with propiconazole, and rotated with propiconazole; paclobutrazol alone, tank‐mixed with propiconazole, and rotated with propiconazole; propiconazole alone; and boscalid alone. S. homoeocarpa was isolated from infected turfgrass three times per year in 2010 and 2011 following treatment applications. A significant shift in DMI insensitivity was observed from the initial sampling in 2010 to the final sampling in 2011 for all treatments, except the PGRs alone and the untreated control. Both PGRs induced overexpression of ShPDR1, and we confirmed the functional involvement of ShPDR1 in reducing S. homoeocarpa sensitivity to PGRs using the ShPDR1 overexpressing yeast mutant. These results have important implications for managing DMI resistance, further investigating the fungistatic activity of PGRs, and developing new fungicides for S. homoeocarpa.

Characterizing the binding interaction of fungicide boscalid with bovine serum albumin (BSA): A spectroscopic study in combination with molecular docking approach

Boscalid, a carboxamide fungicide, is used in the treatment of grey mould and powdery mildew, widely applied to a variety of crops and fruits such as rice, wheat, grapes and pears. It will become a potential risk for health due to its widely application and residue in crops and fruits. In this study, the binding interaction between boscalid and bovine serum albumin (BSA) was characterized using steady-state fluorescence spectroscopy, ultraviolet spectroscopy (UV), synchronous fluorescence spectroscopy, 3D fluorescence spectroscopy, Fourier transform infrared spectroscopy (FT-IR) and molecular docking to ascertain the store, transport and distribution of boscalid in vivo. The experimental results indicated that the fluorescence of BSA was quenched due to the forming the static boscalid–BSA complex with the binding constant of 4.57×103M−1 at 298 K and boscalid bound on the subdomain III A (site II) of BSA through van der Waals force and hydrogen bonding interaction. The binding process of boscalid with BSA was spontaneous and enthalpy-driven process based on ΔG0<0 and |ΔH0|>T|ΔS0| over the studied temperature range. Meanwhile, the obvious change in the conformation of boscalid was observed while the slight change in the conformation of BSA when binding boscalid to the BSA, implying that the flexibility of boscalid contributes to increasing the stability of the boscalid–BSA complex.

The challenges of predicting pesticide exposure of honey bees at landscape level

To evaluate the risks of pesticides for pollinators, we must not only evaluate their toxicity but also understand how pollinators are exposed to these xenobiotics in the field. We focused on this last point and modeled honey bee exposure to pesticides at the landscape level. Pollen pellet samples (n = 60) from 40 Belgian apiaries were collected from late July to October 2011 and underwent palynological and pesticide residue analyses. Areas of various crops around each apiary were measured at 4 spatial scales. The most frequently detected pesticides were the fungicides boscalid (n = 19, 31.7%) and pyrimethanil (n = 10, 16.7%) and the insecticide dimethoate (n = 10, 16.7%). We were able to predict exposure probability for boscalid and dimethoate by using broad indicators of cropping intensity, but it remained difficult to identify the precise source of contamination (e.g. specific crops in which the use of the pesticide is authorized). For pyrimethanil, we were not able to build any convincing landscape model that could explain the contamination. Our results, combined with the late sampling period, strongly suggest that pesticides applied to crops unattractive to pollinators, and therefore considered of no risk for them, may be sources of exposure through weeds, drift to neighboring plants, or succeeding crops.

The Exposure of Honey Bees to Pesticide Residues in the Hive Environment with Regard to Winter Colony Losses

Abstract The present studies are the second part of the research project dedicated to finding the causes for increased winter mortality of honey bee colonies. The aim of this task was to investigate incidents of overwintered colonies′ death with regard to the potential interrelation to the exposure to pesticides. The samples of winter stores of bee bread and sugar food (honey or syrup processed by bees), beeswax and bees collected from apiaries with low and high rates of winter colony mortality were searched for acaricides used to control V. destructor and plant protection pesticides. The presence of acaricides used in apiculture has been detected in the 51% beeswax samples. The most abundant acaricide was tau-fluvalinate. The stores of bee bread and sugar food had a similar frequency of plant protection pesticide occurrence, ranging between 50-60%, but the number of active substances and their concentrations were substantially lower in sugar food samples. The most prevalent pesticides in pollen were fungicides (carbendazim and boscalid) and insecticides (acetamiprid and thiacloprid). Only a few pesticides were found in the several dead honey bees. The level of pesticide contamination (frequency, concentration, toxicity) of hive products and bees originating from apiaries with both a high and low winter colony survival rates, was similar, which created a similar extent of risk. Although the multiple varroacides and pesticides were present in the hive environment we not found unequivocal links between their residues and high winter colony mortality.

Host Resistance and Chemical Control for Management of Sclerotinia Stem Rot of Soybean in Ohio

Recent outbreaks of Sclerotinia stem rot (SSR) of soybean in Ohio, along with new fungicides and cultivars with resistance to this disease, have led to a renewed interest in studies to update disease management guidelines. The effect of host resistance (in moderately resistant [MR] and moderately susceptible [MS] cultivars) and chemical control on SSR and yield was evaluated in 12 environments from 2014 to 2016. The chemical treatments evaluated were an untreated check, four fungicides (boscalid, picoxystrobin, pyraclostrobin, and thiophanate-methyl), and one herbicide (lactofen) applied at soybean growth stage R1 (early flowering) alone or at R1 followed by a second application at R2 (full flowering). SSR developed in 6 of 12 environments, with mean disease incidence in the untreated check of 2.5 to 41%. The three environments with high levels of SSR (disease incidence in the untreated check >20%) were used for further statistical analysis. There were significant effects (P < 0.05) of soybean cultivar and chemical treatment on SSR levels. Significantly lower levels of SSR were observed in MR cultivars. Both boscalid and lactofen reduced SSR but did not increase yield. Pyraclostrobin increased SSR compared with the untreated check in the three environments with high levels of disease. In the six fields where SSR did not develop, chemical treatment did not increase yield, nor was the yield from the MR cultivar significantly different from the MS cultivar. For Ohio, MR cultivars alone were effective for management of SSR in soybean fields where this disease has historically occurred.

Synthesis of novel fenfuram-diarylether hybrids as potent succinate dehydrogenase inhibitors

Twelve novel fenfuram-diarylether hybrids were designed, synthesized and characterized by 1H NMR and MS. Their in vitro antifungal activities were evaluated against five phytopathogenic fungi by mycelial growth inhibition method. Most compounds showed significant antifungal effect on Rhizoctonia solani and Sclerotinia sclerotiorum. Compound 1c exhibited the most potent antifungal effect on R. solani with an EC50 value of 0.242mg/L, superior to the commercial fungicide boscalid (EC50=1.758mg/L) and the lead fungicide fenfuram (EC50=7.691mg/L). Molecular docking revealed that compound 1c featured a higher affinity for succinate dehydrogenase (SDH) than fenfuram. Furthermore, it was shown that the 2-chlorophenyl group of compound 1c formed a π-π stacking with D/Tyr-128 and a Cl-π interaction with B/His-249, which made compound 1c more active than fenfuram against SDH.

Exposure assessment of honeybees through study of hive matrices: analysis of selected pesticide residues in honeybees, beebread, and beeswax from French beehives by LC-MS/MS.

Apiculture and pollination services are seriously threatened by bee weakening and losses phenomena. In this context, a survey was performed on samples from beehives across French areas during the 2012-2016 growing seasons, primarily taken from symptomatic colonies. A total of 488 honeybees, beebread, and wax were analyzed for the presence of pesticide residues. A total of 13 analytes including neonicotinoids and pyrethroids insecticides together with some of their metabolites and the fungicide boscalid were screened within samples. Methodologies based on efficient modified quick, easy, cheap, effective, rugged, and safe extractions followed by an LC-MS/MS quantification were implemented for each matrix. Thirty-eight percent of the 125 bee samples, 61% of the 87 wax samples, and 77% of the 276 beebread samples contained at least one of the targeted pesticides. Beebread was the most contaminated matrix with an average of two pesticide detections by positive sample and a maximum of seven compounds for a sample. Neonicotinoids and boscalid were the most often detected pesticides, whatever the matrix. The comparison of neonicotinoid detections in samples collected before and after the partial neonicotinoid ban in France displays a decrease in the frequency of detections for contamination levels lower than 1ng/g in beebread. For higher levels and other matrices, no tendency can be drawn.

Resistance to the SDHI Fungicides Boscalid, Fluopyram, Fluxapyroxad, and Penthiopyrad in Botrytis cinerea from Commercial Strawberry Fields in Spain.

Gray mold, caused by the necrotrophic fungus Botrytis cinerea., is one of the most economically important diseases of strawberry. Gray mold control involves the application of fungicides throughout the strawberry growing season; however, B. cinerea isolates resistant to multiple classes of site-specific fungicides have been recently reported in the Spanish gray mold population. Succinate dehydrogenase inhibitors (SDHI) constitute a relatively novel class of fungicides registered for gray mold control representing new alternatives for strawberry growers. In the present study, 37 B. cinerea isolates previously characterized for their sensitivity to boscalid and amino acid changes in the SdhB protein were used to determine the effective concentration that reduces mycelial growth by 50% (EC50) to fluopyram, fluxapyroxad, and penthiopyrad. The present study was also conducted to obtain discriminatory doses to monitor SDHI fungicide resistance in 580 B. cinerea isolates collected from 27 commercial fields in Spain during 2014, 2015, and 2016. The EC50 values ranged from 0.01 to >100 μg/ml for fluopyram, <0.01 to 4.19 μg/ml for fluxapyroxad, and, finally, <0.01 to 59.65 μg/ml for penthiopyrad. Based on these results, as well as findings from a previous publication, the discriminatory doses chosen to examine sensitivities to boscalid, fluopyram, fluxapyroxad, and penthiopyrad were 100, 15, 1, and 6 μg/ml, respectively. Over the course of the 3-year monitoring period, the overall frequencies of resistance to the four SDHI were 56.9, 6.9, 12.9, and 24.6%, respectively. The frequency of boscalid-resistant isolates decreased from 73 to 41% over the years; however, the fluopyram-resistant isolates increased from 5 to 10% after 1 year of registration. Four SDHI resistance patterns were observed in our population, which included patterns I (30%; resistance to boscalid), II (13.8%; resistance to boscalid and penthiopyrad), III (5.7%; boscalid, fluxapyroxad, and penthiopyrad), and IV (7.9%; resistance to boscalid, fluopyram, fluxapyroxad, and penthiopyrad). Patterns I and II were associated with the amino acid substitutions H272R and H272Y; pattern III was associated only with the H272Y mutation; and, finally, pattern IV was associated with the N230I allele in the SdhB subunit. For gray mold management, it is suggested that the simultaneous use of boscalid and penthiopyrad should be limited to one application per season; however, fluxapyroxad and, especially, fluopyram could be used as valid SDHI alternatives for gray mold control, although they should be applied with caution.

대추 중 살균제 boscalid와 pyraclostrobin의 잔류 소실 특성

대추 중 살균제 boscalid와 pyraclostrobin의 잔류 소실 특성