Characteristics of fluopicolide-resistance mutants in Phytophthora nicotianae, the pathogen causing black shank disease in tobacco.

Black shank, a devastating disease in tobacco production worldwide, is caused by the oomycete plant pathogen Phytophthora nicotianae. Fluopicolide is a pyridinylmethyl-benzamides fungicide with a unique mechanism of action and has been widely used for controlling a variety of oomycetes such as Plasmopara viticola, Phytophthora infestans, Pseudoperonospora cubensis, P. nicotianae and Bremia lactucae. However, the fluopicolide-resistance risk and molecular basis in P. nicotianae have not been reported. In this study, the sensitivity profile of 141 P. nicotianae strains to fluopicolide was determined, with a mean median effective concentration (EC50) value of 0.12 ± 0.06µg/mL. Five stable fluopicolide-resistant mutants of P. nicotianae were obtained by fungicide adaptation, and the compound fitness index of these resistant mutants were lower than that of their parental isolates. Additionally, cross-resistance tests indicated that the sensitivity of fluopicolide did not correlate with other oomycete fungicides, apart from fluopimomide. DNA sequencing revealed two point mutations, G765E and N769Y, in the PpVHA-a protein in the fluopicolide-resistant mutants. Transformation and expression of PpVHA-a genes carrying G765E and N769Y in the sensitive wild-type isolate confirmed that it was responsible for fluopicolide resistance. These results suggest that P. nicotianae has a low to medium resistance risk to fluopicolide in laboratory and that point mutations, G765E and N769Y, in PpVHA-a are associated with the observed fluopicolide resistance.

Sensitivity analysis and point mutations in BcSDHB confer cyclobutrifluram resistance in Botrytis cinerea from China.

Botrytis cinerea is one of the most destructive pathogens worldwide. It can damage over 200 crops, resulting in significant yield and quality losses. Cyclobutrifluram, a new generation of succinate dehydrogenase inhibitors, exhibits excellent inhibitory activity against B. cinerea. However, the baseline sensitivity and resistance of B. cinerea to cyclobutrifluram remains poorly understood. This study was designed to monitor the sensitivity frequency distribution, assess the resistance risk, and clarify the resistance mechanism of B. cinerea to cyclobutrifluram. The baseline sensitivity of B. cinerea isolates to cyclobutrifluram was 0.89 µg/mL. Cyclobutrifluram-resistant B. cinerea populations are present in the field. Six resistant B. cinerea isolates investigated in this study possessed enhanced compound fitness index compared to the sensitive isolates according to mycelial growth, mycelial dry weight, conidiation, conidial germination rate, and pathogenicity. Cyclobutrifluram exhibited no cross-resistance with tebuconazole, fludioxonil, cyprodinil, or iprodione. Sequence alignment revealed that BcSDHB from cyclobutrifluram-resistant B. cinerea isolates had three single substitutions (P225F, N230I, or H272R). Molecular docking verified that these mutations in BcSDHB conferred cyclobutrifluram resistance in B. cinerea. In conclusion, the resistance risk of B. cinerea to cyclobutrifluram is high, and the point mutations in BcSDHB (P225F, N230I, or H272R) confer cyclobutrifluram resistance in B. cinerea. This study provided important insights into cyclobutrifluram resistance in B. cinerea and offered valuable information for monitoring and managing cyclobutrifluram resistance in the future.

PsAF5 functions as an essential adapter for PsPHB2-mediated mitophagy under ROS stress in Phytophthora sojae.

Host-derived reactive oxygen species (ROS) are an important defense means to protect against pathogens. Although mitochondria are the main intracellular targets of ROS, how pathogens regulate mitochondrial physiology in response to oxidative stress remains elusive. Prohibitin 2 (PHB2) is an inner mitochondrial membrane (IMM) protein, recognized as a mitophagy receptor in animals and fungi. Here, we find that an ANK and FYVE domain-containing protein PsAF5, is an adapter of PsPHB2, interacting with PsATG8 under ROS stress. Unlike animal PHB2 that can recruit ATG8 directly to mitochondria, PsPHB2 in Phytophthora sojae cannot recruit PsATG8 to stressed mitochondria without PsAF5. PsAF5 deletion impairs mitophagy under ROS stress and increases the pathogen's sensitivity to H2O2, resulting in the attenuation of P. sojae virulence. This discovery of a PsPHB2-PsATG8 adapter (PsAF5) in plant-pathogenic oomycetes reveals that mitophagy induction by IMM proteins is conserved in eukaryotes, but with differences in the details of ATG8 recruitment.

Analysis of resistance risk and mechanism of the 14α-demethylation inhibitor ipconazole in Fusarium pseudograminearum.

Ipconazole is a broad-spectrum triazole fungicide that is highly effective against Fusarium pseudograminearum. However, its risk of developing resistance and mechanism are not well understood in F. pseudograminearum. Here, the sensitivities of 101 F. pseudograminearum isolates to ipconazole were investigated, and the average EC50 value was 0.1072 µg/mL. Seven mutants resistant to ipconazole were obtained by fungicide adaption, with all but one showing reduced fitness relative to the parental isolates. Cross-resistance was found between ipconazole and mefentrifluconazole and tebuconazole, but none between ipconazole and pydiflumetofen, carbendazim, fludioxonil, or phenamacril. In summary, these findings suggest that there is a low risk of F. pseudograminearum developing resistance to ipconazole. Additionally, a point mutation, G464S, was seen in FpCYP51B and overexpression of FpCYP51A, FpCYP51B and FpCYP51C was observed in ipconazole-resistant mutants. Assays, including transformation and molecular docking, indicated that G464S conferred ipconazole resistance in F. pseudograminearum.

Resistance risk, resistance mechanism and the effect on DON production of a new SDHI fungicide cyclobutrifluram in Fusarium graminearum.

Fusarium head blight in wheat is caused by Fusarium graminearum, resulting in significant yield losses and grain contamination with deoxynivalenol (DON), which poses a potential threat to animal health. Cyclobutrifluram, a newly developed succinate dehydrogenase inhibitor, has shown excellent inhibition of Fusarium spp. However, the resistance risk of F. graminearum to cyclobutrifluram and the molecular mechanism of resistance have not been determined. In this study, we established the average EC50 of a range of F. graminearum isolates to cyclobutrifluram to be 0.0110 µg/mL. Six cyclobutrifluram-resistant mutants were obtained using fungicide adaptation. All mutants exhibited impaired fitness relative to their parental isolates. This was evident from measurements of mycelial growth, conidiation, conidial germination, virulence, and DON production. Interestingly, cyclobutrifluram did not seem to affect the DON production of either the sensitive isolates or the resistant mutants. Furthermore, a positive cross-resistance was observed between cyclobutrifluram and pydiflumetofen. These findings suggest that F. graminearum carries a moderate to high risk of developing resistance to cyclobutrifluram. Additionally, point mutations H248Y in FgSdhB and A73V in FgSdhC1 of F. graminearum were observed in the cyclobutrifluram-resistant mutants. Finally, an overexpression transformation assay and molecular docking indicated that FgSdhBH248Y or FgSdhC1A73V could confer resistance of F. graminearum to cyclobutrifluram.

Ametoctradin resistance risk and its resistance-related point mutation in PsCytb of Phytophthora sojae confirmed using ectopic overexpression.

Ametoctradin is mainly used to treat plant oomycetes diseases, but the mechanism and resistance risk of ametoctradin in Phytophthora sojae remain unknown. This study determined the ametoctradin sensitivity of 106 P. sojae isolates and found that the frequency distribution of the median effective concentration (EC50) of ametoctradin was unimodal with a mean value of 0.1743 ± 0.0901 µg/mL. Furthermore, ametoctradin-resistant mutants had a substantially lower fitness index compared with that of wild-type isolates. Although ametoctradin did not show cross-resistance to other fungicides, negative cross-resistance to amisulbrom was found. In comparison to sensitive isolates, the control efficacy of ametoctradin to resistant mutants was lower, implying a low to moderate ametoctradin resistance risk in P. sojae. All ametoctradin-resistant mutants contained a S33L point mutation in PsCytb. A system with overexpression of PsCytb in the nucleus was established. When we ectopically overexpressed S33L-harboring PsCytb, P. sojae developed ametoctradin resistance. We hypothesized that the observed negative resistance between ametoctradin and amisulbrom could be attributed to conformational changes in the binding cavity of PsCytb at residues 33 and 220.

Unveiling Vacuolar H+-ATPase Subunit a as the Primary Target of the Pyridinylmethyl-Benzamide Fungicide, Fluopicolide.

An estimated 240 fungicides are presently in use, but the direct targets for the majority remain elusive, constraining fungicide development and efficient resistance monitoring. In this study, we found that Pcα-actinin knockout did not influence the sensitivity of Phytophthora capsici to fluopicolide, which is a notable oomycete inhibitor. Using a combination of Bulk Segregant Analysis Sequencing and Drug Affinity Responsive Target Stability (DARTS) assays, the vacuolar H+-ATPase subunit a (PcVHA-a) was pinpointed as the target protein of fluopicolide. We also confirmed four distinct point mutations in PcVHA-a responsible for fluopicolide resistance in P. capsici through site-directed mutagenesis. Molecular docking, ATPase activity assays, and a DARTS assay suggested a fluopicolide-PcVHA-a interaction. Sequence analysis and further molecular docking validated the specificity of fluopicolide for oomycetes or fish. These findings support the claim that PcVHA-a is the target of fluopicolide, proposing vacuolar H+-ATPase as a promising target for novel fungicide development.

Mefentrifluconazole-Resistant Risk and Resistance-Related Point Mutation in FpCYP51B of Fusarium pseudograminearum.

Mefentrifluconazole, a triazole fungicide, exhibits remarkable efficacy in combating Fusarium spp. The mean EC50 value of mefentrifluconazole against 124 isolates of Fusarium pseudograminearum was determined to be 1.06 µg/mL in this study. Fungicide taming produced five mefentrifluconazole-resistant mutants with resistance factors ranging from 19.21 to 111.34. Compared to the original parental isolates, the fitness of three resistant mutants was much lower, while the remaining two mutants displayed enhanced survival fitness. There was evidence of positive cross-resistance between tebuconazole and mefentrifluconazole. Mefentrifluconazole resistance in F. pseudograminearum can be conferred by FpCYP51BL144F, which was identified in four mutants according to molecular docking and site-directed transformation experiments. Overexpression of FpCYP51s was also detected in the resistant mutants. In conclusion, mefentrifluconazole has a low-to-medium resistance risk in F. pseudograminearum, and the L144F mutation in FpCYP51B and the increased expression level of FpCYP51s may be responsible for mefentrifluconazole resistance in F. pseudograminearum.

Inhibitory activity to Fusarium spp. and control potential for wheat Fusarium crown rot of a novel succinate dehydrogenase inhibitor cyclobutrifluram.

BACKGROUND: Wheat Fusarium crown rot (FCR) is a serious problem primarily caused by Fusarium pseudograminearum, a pathogenic agent known to produce mycotoxins, including deoxynivalenol (DON). Cyclobutrifluram, a novel succinate dehydrogenase inhibitor devised by Syngenta, has immense potential to control both nematodes and Fusarium diseases. However, its efficacy in combating Fusarium species, its ability to prevent and reverse the detrimental effects of FCR, and its impact on the production of DON by F. pseudograminearum are yet to be fully ascertained. RESULTS: Cyclobutrifluram exhibited substantial inhibitory activity against Fusarium species, with half-maximal effective concentration values ranging from 0.0021-0.0647 µg mL-1 . It demonstrated significant inhibitory activity toward three developmental stages of F. pseudograminearum, F. graminearum and F. asiaticum. Furthermore, cyclobutrifluram showed both protective and curative activities against FCR and was rapidly absorbed by roots and transported to wheat stems and leaves. Cyclobutrifluram could also decrease DON production by F. pseudograminearum. CONCLUSION: This investigation has revealed the potential of cyclobutrifluram as a formidable candidate fungicide, particularly in its ability to effectively combat FCR and other Fusarium-related ailments. This novel compound has exceptional pathogen-fighting capabilities, coupled with remarkable systemic translocation properties and a notable ability to reduce the production of DON. © 2023 Society of Chemical Industry.

Evaluation of SYP-34773's resistance risk and its impact on the activity of mitochondrial respiratory electron transport chain complex I in Phytophthora litchii.

BACKGROUND: SYP-34773 is a low-toxicity pyrimidine amine compound, which was synthesized by modifying the lead compound diflumetorim. Previous literature has shown that it can strongly inhibit the mycelial growth of several important plant pathogens, including Phytophthora litchii. However, the resistance risk of SYP-34773 has not been reported for P. litchii. RESULTS: The mean effective concentration (EC50 ) value of SYP-34773 against the mycelial growth of 111 P. litchii isolates was 0.108 ± 0.008 µg mL-1 , which can be used as the baseline sensitivity for SYP-34773 resistance detection in the future. Six mutants were obtained from two parental strain through fungicide induction, whose resistance factors fell between 194- and 687-fold, with stability. Results regarding mycelial growth, sporangial production, sporangial germination, zoospore release, cystspore germination, and pathogenicity showed that the mutants' compound fitness index values were significantly lower than those of their parental isolate. Furthermore, there was no cross-resistance between SYP-34773 and diflumetorim in P. litchii. Significant inhibition of the mitochondrial complex I enzyme activity in two wild-type P. litchii isolates, but not in mutants, was observed upon treatment with SYP-34773. CONCLUSION: The resistance risk of SYP-34773 in P. litchii is moderate, and resistance management strategies should be adopted in field use. SYP-34773 is a mitochondrial complex I inhibitor, and SYP-34773-resistant P. litchii isolates did not show cross-resistance against diflumetorim. © 2023 Society of Chemical Industry.

Resistance to the DMI fungicide mefentrifluconazole in Monilinia fructicola: risk assessment and resistance basis analysis.

BACKGROUND: Brown rot disease, caused by Monilinia fructicola, poses a significant challenge to peach production in China. The efficacy of mefentrifluconazole, a new triazole fungicide, in controlling brown rot in peaches has been remarkable. However, the resistance risk and mechanism associated with this fungicide remain unclear. This study was designed to assess the resistance risk of M. fructicola to mefentrifluconazole and reveal the potential resistance mechanism. RESULTS: The mean median effective concentration (EC50 ) of 101 M. fructicola isolates to mefentrifluconazole was 0.003 µg mL-1 , and the sensitivity exhibited a unimodal distribution. Seven mefentrifluconazole-resistant mutants were generated from three parental isolates in the laboratory through fungicide adaption. The biological characteristics of the resistant mutants revealed that three of them exhibited enhanced survival fitness compared to the parental isolates, whereas the remaining four mutants displayed reduced survival fitness. Mefentrifluconazole showed strong positive cross-resistance with fenbuconazole, whereas no cross-resistance was observed with pyrimethanil, procymidone or pydiflumetofen. No overexpression of MfCYP51 gene was detected in the resistant mutants. Multiple sequence alignment revealed that three resistant mutants (MXSB2-2, Mf12-1 and Mf12-2) had a point mutation (G461S) in MfCYP51 protein. Molecular docking techniques confirmed the contribution of this point mutation to mefentrifluconazole resistance. CONCLUSION: The risk of M. fructicola developing resistance to mefentrifluconazole is relatively low-to-medium and point mutation G461S in MfCYP51 could confer mefentrifluconazole resistance in M. fructicola. This study provided essential data for monitoring the emergence of resistance and developing resistance management strategies for mefentrifluconazole. © 2023 Society of Chemical Industry.

Characteristics of famoxadone-resistant mutants of Phytophthora litchii and their effect on lychee fruit quality.

Lychee downy blight (LDB), a common disease caused by the oomycete Phytophthora litchii, poses a significant threat to both pre- and post-harvest stages, leading to substantial economic losses. Famoxadone, a quinone outside inhibitor fungicide, was registered for controlling LDB in China in 2002. However, limited information is available regarding the risk, mechanism, and impact on lychee fruit quality associated with famoxadone resistance. In this study, we determined the sensitivity of 133 P. litchii isolates to famoxadone, yielding a mean EC50 value of 0.46 ± 0.21 µg/mL. Through fungicide adaption, we derived resistant mutants with M124I and Y131C substitutions in PlCyt b (Cytochrome b in P. litchii) from wild-type isolates. In vitro assessments revealed that the fitness of the resistant mutants was significantly lower compared to the parental isolates. These laboratory findings demonstrate a moderate resistance risk of P. litchii to famoxadone. Molecular docking analyses indicated that the M124I and Y131C alterations disrupted hydrogen bonds and weakened the binding energy between famoxadone and PlCyt b. This indicates that the M124I and Y131C changes do indeed confer famoxadone resistance in P. litchii. Infection caused by famoxadone-resistant mutants exhibited a decreased or comparable impact on the characteristic traits of lychee fruit compared to the sensitive isolate. For future detection of famoxadone-resistant strains, AS-PCR primers were designed based on the M124I substitution.

Attributes of Cyazofamid-Resistant Phytophthora litchii Mutants and Its Impact on Quality of Litchi Fruits.

In this study, we determined the sensitivity of 148 Phytophthora litchii isolates to cyazofamid, yielding a mean EC50 value of 0.0091 ± 0.0028 µg/mL. Through fungicide adaptation, resistant mutants (RMs) carrying the F220L substitution in PlCyt b were derived from wild-type isolates. Notably, these RMs exhibited a lower fitness compared with the parental isolates. Molecular docking analysis further revealed that the F220L change contributed to a decrease in the binding energy between cyazofamid and PlCyt b. The total phenol and flavonoid contents in the litchi pericarp treated with cyazofamid on day 5 were significantly higher than in other treatments. Overall, the laboratory assessment indicated a moderate risk of cyazofamid resistance in P. litchii, but the emergence of the F220L change could lead to a high level of resistance. Thus, cyazofamid represents a promising agrochemical for controlling postharvest litchi downy blight and extending the shelf life of litchi fruits.

Three point mutations in AaCYP51 combined with induced overexpression of AaCYP51 conferred low-level resistance to mefentrifluconazole in Alternaria alternata.

Tomato early blight is a significant disease that causes substantial losses to tomato yield and quality. Mefentrifluconazole, an isopropanol-azole subgroup of triazole fungicides, has been registered in China for controlling various plant diseases, including tomato early blight, grape anthracnose, and apple brown spot. However, limited information is available on the mefentrifluconazole resistance risk and mechanism in plant pathogens. The sensitivity to mefentrifluconazole of 122 isolates of Alternaria alternata, one of the causal agents of tomato early blight, collected from different provinces in China, was evaluated. The results showed a unimodal curve for the sensitivity frequency, with an average EC50 of 0.306 µg/mL. Through fungicide adaption, six resistant mutants (N4, N5, T4, T5, NG1, and NG10) were obtained from three parental isolates, with a mutation frequency of 3.28 × 10-4 and resistance factors ranging between 19 and 147. The survival fitness of the resistant mutants, except for NG1, was significantly lower than that of their parental isolates. Positive cross-resistance was observed between mefentrifluconazole and difenoconazole or fenbuconazole, whereas no cross-resistance was found with three non-DMI fungicides. Furthermore, three distinct point mutations were detected in the AaCYP51 protein of the resistant mutants: I300S in T4 and T5; A303T in N4, NG1, and NG10; and A303V in N5. Compared to the parental isolates, the AaCYP51 gene was overexpressed in all six resistant mutants when treated with mefentrifluconazole. In summary, the resistance risk of A. alternata to mefentrifluconazole was low, and point mutations and overexpression of the AaCYP51 gene were identified as contributing factors to mefentrifluconazole resistance in A. alternata.

A Novel FYVE Domain-Containing Protein Kinase, PsZFPK1, Plays a Critical Role in Vegetative Growth, Sporangium Formation, Oospore Production, and Virulence in Phytophthora sojae.

Proteins containing both FYVE and serine/threonine kinase catalytic (STKc) domains are exclusive to protists. However, the biological function of these proteins in oomycetes has rarely been reported. In the Phytophthora sojae genome database, we identified five proteins containing FYVE and STKc domains, which we named PsZFPK1, PsZFPK2, PsZFPK3, PsZFPK4, and PsZFPK5. In this study, we characterized the biological function of PsZFPK1 using a CRISPR/Cas9-mediated gene replacement system. Compared with the wild-type strain, P6497, the PsZFPK1-knockout mutants exhibited significantly reduced growth on a nutrient-rich V8 medium, while a more pronounced defect was observed on a nutrient-poor Plich medium. The PsZFPK1-knockout mutants also showed a significant increase in sporangium production. Furthermore, PsZFPK1 was found to be essential for oospore production and complete virulence but dispensable for the stress response in P. sojae. The N-terminal region, FYVE and STKc domains, and T602 phosphorylation site were found to be vital for the function of PsZFPK1. Conversely, these domains were not required for the localization of PsZFPK1 protein in the cytoplasm. Our results demonstrate that PsZFPK1 plays a critical role in vegetative growth, sporangium formation, oospore production, and virulence in P. sojae.

Phytophthora RxLR effector PcSnel4B promotes degradation of resistance protein AtRPS2.

Phytophthora capsici deploys effector proteins to manipulate host immunity and facilitate its colonization. However, the underlying mechanisms remain largely unclear. In this study, we demonstrated that a Sne-like (Snel) RxLR effector gene PcSnel4 is highly expressed at the early stages of P. capsici infection in Nicotiana benthamiana. Knocking out both alleles of PcSnel4 attenuated the virulence of P. capsici, while expression of PcSnel4 promoted its colonization in N. benthamiana. PcSnel4B could suppress the hypersensitive reaction (HR) induced by Avr3a-R3a and RESISTANCE TO PSEUDOMONAS SYRINGAE 2 (AtRPS2), but it did not suppress cell death elicited by Phytophthora infestin 1 (INF1) and Crinkler 4 (CRN4). COP9 signalosome 5 (CSN5) in N. benthamiana was identified as a host target of PcSnel4. Silencing NbCSN5 compromised the cell death induced by AtRPS2. PcSnel4B impaired the interaction and colocalization of Cullin1 (CUL1) and CSN5 in vivo. Expression of AtCUL1 promoted the degradation of AtRPS2 and disrupted HR, while AtCSN5a stabilized AtRPS2 and promoted HR, regardless of the expression of AtCUL1. PcSnel4 counteracted the effect of AtCSN5 and enhanced the degradation of AtRPS2, resulting in HR suppression. This study deciphered the underlying mechanism of PcSnel4-mediated suppression of HR induced by AtRPS2.

Resistant risk and resistance-related point mutation in SdhC1 of pydiflumetofen in Fusarium pseudograminearum.

BACKGROUND: Fusarium pseudograminearum is one of the dominant pathogens of Fusarium crown rot (FCR) worldwide. Unfortunately, no fungicides have yet been registered for the control of FCR in wheat in China. Pydiflumetofen, a new-generation succinate dehydrogenase inhibitor, exhibits excellent inhibitory activity to Fusarium spp. A resistance risk assessment of F. pseudograminearum to pydiflumetofen and the resistance mechanism involved have not yet been investigated. RESULTS: The median effective concentration (EC50 ) value of 103 F. pseudograminearum isolates to pydiflumetofen was 0.0162 µg mL-1 , and the sensitivity exhibited a unimodal distribution. Four resistant mutants were generated by fungicide adaption, which possessed similar or impaired fitness compared to corresponding parental isolates based on the results of mycelial growth, conidiation, conidium germination rate, and virulence determination. Pydiflumetofen showed strong positive cross-resistance with cyclobutrifluram and fluopyram but no cross-resistance with carbendazim, phenamacril, tebuconazole, fludioxonil, or pyraclostrobin. Sequence alignment revealed that pydiflumetofen-resistant F. pseudograminearum mutants had two single-point mutations of A83V or R86K in FpSdhC1 . Molecular docking further confirmed that point mutation of A83V or R86K in FpSdhC1 could confer resistance of F. pseudograminearum to pydiflumetofen. CONCLUSION: Fusarium pseudograminearum shows an overall moderate risk of developing resistance to pydiflumetofen, and point mutation FpSdhC1 A83V or FpSdhC1 R86K could confer pydiflumetofen resistance in F. pseudograminearum. This study provided vital data for monitoring the emergence of resistance and developing resistance management strategies for pydiflumetofen. © 2023 Society of Chemical Industry.

Resistance Risk and Resistance-Related Point Mutations in Target Protein Cyt b of the Quinone Inside Inhibitor Amisulbrom in Phytophthora litchii.

Amisulbrom is a novel quinone inside inhibitor, which exhibits excellent inhibitory activity against phytopathogenic oomycetes. However, the resistance risk and mechanism of amisulbrom in Phytophthora litchii are rarely reported. In this study, the sensitivity of 147 P. litchii isolates to amisulbrom was determined, with an average EC50 of 0.24 ± 0.11 µg/mL. The fitness of resistant mutants, obtained by fungicide adaption, was significantly lower than that of the parental isolates in vitro. Cross-resistance was detected between amisulbrom and cyazofamid. Amisulbrom could not inhibit the cytochrome bc1 complex activity with H15Y and G30E + F220L point mutations in cytochrome b (Cyt b) in vitro. Molecular docking indicated that the H15Y or G30E point mutation can decrease the binding energy between amisulbrom and P. litchii Cyt b. In conclusion, P. litchii might have a medium resistance risk to amisulbrom, and a novel point mutation H15Y or G30E in Cyt b could cause high amisulbrom resistance in P. litchii.

Resistance Risk Assessment for the New OSBP Inhibitor Y18501 in Pseudoperonospora cubensis and Point Mutations (G705V, L798W, and I812F) in PscORP1 that Confer Resistance.

Y18501 is a new oxysterol-binding protein inhibitor (OSBPI) that shows strong inhibitory activity against Pseudoperonospora cubensis. In this study, the sensitivities of 159 Ps. cubensis isolates to Y18501 were determined, with EC50 values ranging from 0.001 to 11.785 µg/mL, indicating that a Y18501-resistant subpopulation has appeared in the field. Ten Y18501-resistant mutants were obtained by fungicide adaptation and displayed fitness equal to or stronger than their parental isolates, which suggests that the resistance risk of Ps. cubensis to Y18501 is high. The consecutive applications of Y18501 in the field resulted in the rapid resistance of Ps. cubensis and decreased control efficacy of cucumber downy mildew (CDM), which could be alleviated by compounding with mancozeb. A positive cross-resistance was detected between Y18501 and oxathiapiprolin. The amino acid substitutions G705V, L798W, and I812F in PscORP1 conferred resistance to Y18501 in Ps. cubensis, which was validated by molecular docking and molecular dynamics simulations.

Resistance Risk and Resistance-Related Point Mutation in SdhB and SdhC1 of Cyclobutrifluram in Fusarium pseudograminearum.

Cyclobutrifluram is a novel succinate dehydrogenase inhibitor (SDHI) developed by Syngenta and helps to inhibit Fusarium pseudograminearum. Here, the potential for cyclobutrifluram resistance in F. pseudograminearum and the resistance mechanism involved were evaluated. Baseline sensitivity of F. pseudograminearum to cyclobutrifluram was determined with a mean EC50 value of 0.0248 µg/mL. Fungicide adaption generated five resistant mutants, which possess a comparable or a slightly impaired fitness compared to corresponding parental isolates. This indicates that the resistance risk of F. pseudograminearum to cyclobutrifluram might be moderate. Cyclobutrifluram-resistant isolates also demonstrated resistance to pydiflumetofen but sensitivity to carbendazim, phenamacril, tebuconazole, fludioxonil, or pyraclostrobin. Additionally, point mutations H248Y in FpSdhB and A83V or R86K in FpSdhC1 were found in cyclobutrifluram-resistant F. pseudograminearum mutants. Molecular docking and overexpression transformation assay revealed that FpSdhBH248Y and FpSdhC1A83V or FpSdhC1R86K confer the resistance of F. pseudograminearum to cyclobutrifluram.