scholarly journals Molecular Characterization and Overexpression of CYP51 gene of Difenoconazole Resistance in Lasiodiplodia Theobromae Field Isolates

2021 ◽  
Author(s):  
Chenguang Wang ◽  
Luxi Xu ◽  
Xiaoyu Liang ◽  
Jing Wang ◽  
Xinwei Xian ◽  
...  
Keyword(s):  
Gene Sequence ◽  
Management Strategies ◽  
Resistance Mechanism ◽  
Cross Resistance ◽  
Pcr Analysis ◽  
Baseline Sensitivity ◽  
Lasiodiplodia Theobromae ◽  
Normal Distribution Curve ◽  
Rate Method ◽  
Important Disease

Abstract Stem-end rot (SER) caused by Lasiodiplodia theobromae is an important disease of mango in China. Demethylation inhibitor (DMI) fungicide are widely used for diseases control in mango orchards. The baseline sensitivity to difenoconazole of 138 isolates collected in the field in 2019 from mango were established by the mycelial growth rate method. The cross-resistance to six site-specific fungicides with different modes of action were investigated using 20 isolates randomly selected. The possible reasons for L. theobromae resistance to difenoconazole were preliminarily determined through gene sequence alignment and quantitative real-time PCR analysis. The results showed that the EC50 values of 138 L. theobromae isolates to difenoconazole ranged from 0.01 to 13.72 µg/ml. The frequency of difenoconazole sensitivity formed a normal distribution curve when the outliers were excluded. Difenoconazole showed positive cross-resistance only with the DMI tebuconazole, but not with non-DMI carbendazim, pyraclostrobin, fludioxonil, bromothalonil, or iprodione. Some multifungicide-resistant isolates of L. theobromae were found. Two amino acid substitutions (E209k and G207A) were found in CYP51 protein, but they were not likely related to the resistance phenotype. There was no alteration in promoter region of the CYP51 gene. However, difenoconazole significantly increased the expression of the CYP51 gene in the resistant isolates when compared to the susceptible isolates. This study is important references to explore resistance mechanism. These results are vital to make effective mango diseases management strategies in order to avoid the development of further resistance.

scholarly journals Molecular characterization and overexpression of the difenoconazole resistance gene CYP51 in Lasiodiplodia theobromae field isolates

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Chenguang Wang ◽  
Luxi Xu ◽  
Xiaoyu Liang ◽  
Jing Wang ◽  
Xinwei Xian ◽  
...  
Keyword(s):  
Gene Sequence ◽  
Management Strategies ◽  
Cross Resistance ◽  
Pcr Analysis ◽  
Baseline Sensitivity ◽  
Lasiodiplodia Theobromae ◽  
Normal Distribution Curve ◽  
Dmi Fungicides ◽  
Rate Method ◽  
Important Disease

AbstractStem-end rot (SER) caused by Lasiodiplodia theobromae is an important disease of mango in China. Demethylation inhibitor (DMI) fungicides are widely used for disease control in mango orchards. The baseline sensitivity to difenoconazole of 138 L. theobromae isolates collected from mango in the field in 2019 was established by the mycelial growth rate method. The cross-resistance to six site-specific fungicides with different modes of action were investigated using 20 isolates randomly selected. The possible mechanism for L. theobromae resistance to difenoconazole was preliminarily determined through gene sequence alignment and quantitative real-time PCR analysis. The results showed that the EC50 values of 138 L. theobromae isolates to difenoconazole ranged from 0.01 to 13.72 µg/mL. The frequency of difenoconazole sensitivity formed a normal distribution curve when the outliers were excluded. Difenoconazole showed positive cross-resistance only with the DMI tebuconazole but not with non-DMI fungicides carbendazim, pyraclostrobin, fludioxonil, bromothalonil, or iprodione. Some multifungicide-resistant isolates of L. theobromae were found. Two amino acid substitutions (E209k and G207A) were found in the CYP51 protein, but they were unlikely to be related to the resistance phenotype. There was no alteration in the promoter region of the CYP51 gene. However, difenoconazole significantly increased the expression of the CYP51 gene in the resistant isolates compared to the susceptible isolates. These results are vital to develop effective mango disease management strategies to avoid the development of further resistance.

scholarly journals Point Mutations in the β-Tubulin of Phytophthora sojae Confer Resistance to Ethaboxam

2019 ◽  
Vol 109 (12) ◽  
pp. 2096-2106 ◽  
Author(s):  
Qin Peng ◽  
Zhiwen Wang ◽  
Yuan Fang ◽  
Weizhen Wang ◽  
Xingkai Cheng ◽  
...  
Keyword(s):  
Fungicide Resistance ◽  
Germination Rate ◽  
Management Strategies ◽  
Resistance Management ◽  
Point Mutations ◽  
Phytophthora Sojae ◽  
Cross Resistance ◽  
Baseline Sensitivity ◽  
Resistant Mutants ◽  
Β Tubulin

Ethaboxam is a β-tubulin inhibitor registered for the control of oomycete pathogens. The current study was established to determine the ethaboxam sensitivity of the plant pathogen Phytophthora sojae and investigate the potential for the emergence of fungicide resistance. The effective concentration for 50% inhibition (EC50) of 112 Phytophthora sojae isolates exhibited a unimodal distribution with a mean EC50 for ethaboxam of 0.033 µg/ml. Establishing this baseline sensitivity provided critical data for monitoring changes in ethaboxam-sensitivity in field populations. The potential for fungicide resistance was investigated using adaptation on ethaboxam-amended V8 agar, which resulted in the isolation of 20 resistant mutants. An assessment of the biological characteristics of the mutants including mycelial growth, sporulation, germination rate and pathogenicity indicated that the resistance risk in Phytophthora sojae was low to medium with no cross-resistance between ethaboxam and cymoxanil, metalaxyl, flumorph, and oxathiapiprolin being detected. However, positive cross-resistance was found between ethaboxam and zoxamide for Q8L and I258V but negative cross-resistance for C165Y. Further investigation revealed that the ethaboxam-resistant mutants had point mutations at amino acids Q8L, C165Y, or I258V of their β-tubulin protein sequences. CRISPR/Cas9-mediated transformation experiments confirmed that the Q8L, C165Y, or I258V mutations could confer ethaboxam resistance in Phytophthora sojae and that the C165Y mutation induces high levels of resistance. Taken together, the results of the study provide essential data for monitoring the emergence of resistance and resistance management strategies for ethaboxam, as well as for improving the design of novel β-tubulin inhibitors for future development.

scholarly journals Baseline Sensitivity and Toxic Action of the Sterol Demethylation Inhibitor Flusilazole Against Botrytis cinerea

Plant Disease ◽  
2020 ◽  
Vol 104 (11) ◽  
pp. 2986-2993
Author(s):  
Yong Wang ◽  
Miaomaio Wang ◽  
Letian Xu ◽  
Yang Sun ◽  
Juntao Feng
Keyword(s):  
Botrytis Cinerea ◽  
Plant Pathogens ◽  
Resistance Mechanism ◽  
Point Mutations ◽  
Gray Mold ◽  
Shaanxi Province ◽  
Cross Resistance ◽  
Baseline Sensitivity ◽  
Single Spore ◽  
Demethylation Inhibitor

In the present study, a total of 95 Botrytis cinerea single-spore strains collected from different hosts in Shaanxi Province of China were characterized for their sensitivity to the sterol demethylation inhibitor fungicide flusilazole. The effective concentration for 50% inhibition of mycelial growth (EC50) of flusilazole ranged from 0.021 to 0.372 µg/ml, with an average value of 0.093 µg/ml. Cross-resistance between flusilazole and commonly used fungicides was not detected, and no flusilazole-resistant mutants were induced. Both on detached strawberry leaves and in greenhouse experiments, flusilazole was more effective than the commonly used fungicide carbendazim at reducing gray mold. After culture on PDA plates or detached strawberry leaves, no difference in sclerotia production or pathogenicity was detected between two strains, WG12 (most sensitive to flusilazole) and MX18 (least sensitive to flusilazole). After treatment with flusilazole, however, the two strains lost the ability to produce sclerotia, and oxalic acid and ergosterol contents in mycelium decreased. Interestingly, the inhibition rate of ergosterol content in MX18 was significantly lower than that in WG12. Expression of Cyp51, BcatrD, and Bcmfs1 genes all increased after treatment with flusilazole, especially the Cyp51 and BcatrD genes. However, the expression of Cyp51 gene or BcatrD gene in WG12 and MX18 were significantly different from each other after treatment with flusilazole. In addition, no point mutations in Cyp51 gene were found in MX18. These data suggest flusilazole is a promising fungicide for resistance management of gray mold and also provided novel insights into understanding the resistance mechanism of flusilazole against plant pathogens.

scholarly journals Fusarium basal rot: profile of an increasingly important disease in Allium spp.

2021 ◽  
Author(s):  
Dung Le ◽  
Kris Audenaert ◽  
Geert Haesaert
Keyword(s):  
Disease Management ◽  
Cultural Practices ◽  
Current Knowledge ◽  
Management Strategies ◽  
Future Directions ◽  
Basal Rot ◽  
Major Constraint ◽  
Fusarium Basal Rot ◽  
Chemical Pesticides ◽  
Important Disease

AbstractFusarium basal rot (FBR) is a soil-borne disease that affects Allium species worldwide. Although FBR has long been recognized as a major constraint to the production of economically important Allium species, information that could support disease management remains scattered. In this review, the current knowledge on the causal agents, symptomology and epidemiology, impact, and management strategies of FBR is synthesized. We highlight that FPR is associated with different complexes of several Fusarium species, of which Fusarium oxysporum and F. proliferatum are the most prevalent. These pathogenic complexes vary in composition and virulence, depending on sites and hosts, which can be challenging for disease management. Research to improve disease management using chemical pesticides, resistance cultivars, biocontrol agents, and cultural practices has achieved both promising results and limitations. Finally, research needs and future directions are proposed for the development of effective FBR management strategies.

scholarly journals SNP and SCAR Markers for Specific Discrimination of Antler-Shaped Ganoderma lucidum

2019 ◽  
Vol 7 (1) ◽  
pp. 12 ◽  
Author(s):  
O-Chul Kwon ◽  
Chang-Soo Lee ◽  
Young-Jin Park
Keyword(s):  
Ganoderma Lucidum ◽  
Gene Sequence ◽  
Rapd Analysis ◽  
Random Amplified Polymorphic Dna ◽  
Morphological Characteristics ◽  
Pcr Analysis ◽  
Scar Markers ◽  
Nucleotide Polymorphism ◽  
Single Nucleotide ◽  
Specific Fragment

In this study we identified single nucleotide polymorphism (SNP) and sequence characteristic amplification region (SCAR) markers for specific identification of antler-shaped Ganoderma lucidum strains. When the partial mitochondrial SSU rDNA gene sequence of various antler- and kidney-shaped G. lucidum strains were analyzed and aligned, an SNP was found only in the antler-shaped G. lucidum strain at position 456 bp. In addition, this SNP of antler-shaped strains was digested by HinfI restriction enzyme. We further analyzed the polymorphism of various G. lucidum strains by random amplified polymorphic DNA (RAPD) analysis. In RAPD analysis, we isolated and sequenced a fragment, specific for antler-shaped G. lucidum strains. Based on this specific fragment sequence, two sets of specific primer pairs for antler-shaped G. lucidum strains were designed. PCR analysis revealed that two specific bands were observed only from antler-shaped strains. These two molecular markers will be helpful for identification of morphological characteristics of G. lucidum.

scholarly journals Sensitivity of Mycosphaerella fijiensis from Banana to Trifloxystrobin

Plant Disease ◽  
2001 ◽  
Vol 85 (12) ◽  
pp. 1264-1270 ◽  
Author(s):  
K. M. Chin ◽  
M. Wirz ◽  
D. Laird
Keyword(s):  
Cross Resistance ◽  
In Vitro Tests ◽  
Mycosphaerella Fijiensis ◽  
In Planta ◽  
Baseline Sensitivity ◽  
Single Spore ◽  
Resistance Factors ◽  
Development Site ◽  
Disease Pressure

An ascospore germination method was developed and validated to assess the sensitivity of bulk samples of Mycosphaerella fijiensis to trifloxystrobin. Using this method, the sensitivity of 142 ascospore samples from banana plantations not treated with strobilurins was analyzed to establish a baseline of pathogen sensitivity. A bulk method was utilized for monitoring purposes because it avoids potential complications due to the isolation and propagation of single-spore isolates and enables the testing of larger samples. Following intensive use of strobilurins (6 to 11 applications per year) over 4 years, under conditions of high disease pressure and the absence of sanitary measures at a development site in Costa Rica, bulk samples with 50% effective concentration (EC50) resistance factors (RFs) in excess of 500 compared with the mean baseline sensitivity were detected. Single-ascospore isolates derived from spores germinating at the discriminatory dose of 3 μg/ml were also resistant, suggesting that the frequency of resistant individuals in bulk samples could be estimated from the relative numbers of ascospores growing at this dose. The resistance of selected isolates was confirmed in planta. In vitro tests with four resistant and two sensitive single-ascospore isolates collected from different locations and times indicated possible cross-resistance of trifloxystrobin to azoxystrobin, famoxadone, and fenamidone, but not to propiconazole.

Sensitivity of C. fructicola and C. siamense of peach in China to multiple classes of fungicides and characterization of pyraclostrobin-resistant isolates

Plant Disease ◽  
2021 ◽  
Author(s):  
Hafiz Muhammad Usman ◽  
Qin Tan ◽  
Mohammad Mazharul Karim ◽  
Muhammad Adnan ◽  
Weixiao Yin ◽  
...  
Keyword(s):  
High Resistance ◽  
Mycelial Growth ◽  
Management Strategies ◽  
Cross Resistance ◽  
Qoi Fungicides ◽  
Quinone Outside Inhibitor ◽  
Dmi Fungicides ◽  
Colletotrichum Gloeosporioides Species Complex ◽  
Cyt B Gene

Anthracnose, mainly caused by Colletotrichum gloeosporioides species complex including C. fructicola and C. siamense, is a devastating disease of peach. The chemical control has been widely used for years and management failures have increased towards commonly used fungicides. Therefore, screening of sensitivity of Colletotrichum spp. to fungicides with different modes of action is needed to make proper management strategies for peach anthracnose. In this study, sensitivity of 80 isolates of C. fructicola and C. siamense was screened for pyraclostrobin, procymidone, prochloraz and fludioxonil based on mycelial growth inhibition at discriminatory doses. Results showed that C. fructicola and C. siamense isolates were highly resistant to procymidone and fludioxonil with 100% resistance frequencies to both fungicides, but sensitive to prochloraz, i.e., no resistant isolates were found. For pyraclostrobin, 74% of C. fructicola isolates showed high resistance and 26 % were low resistant, all of the C. siamense isolates were low resistant. No positive cross-resistance was observed between pyraclostrobin and azoxystrobin, even they are members of the same quinone outside inhibitor (QoI) fungicide group, and between pyraclostrobin and non-QoIs. Resistant isolates to QoI fungicides were evaluated for the fitness penalty. Results showed that no significant differences except for mycelial growth rates were detected between highly resistant and low-resistant isolates of C. fructicola. Molecular characterization of Cyt b gene revealed that the G143A point mutation was the determinant of the high resistance in C. fructicola. This study demonstrated the current resistance status of C. fructicola and C. siamense to different fungicides and their future perspectives. Demethylation inhibitor (DMI) fungicides are the best option among different chemicals to control peach anthracnose in China.

Lack of Cross-Resistance of Paraquat-Resistant Hairy Fleabane (Conyza bonariensis) to Other Toxic Oxygen Generators Indicates Enzymatic Protection is Not the Resistance Mechanism

Weed Science ◽  
1989 ◽  
Vol 37 (1) ◽  
pp. 5-11 ◽  
Author(s):  
Kevin C. Vaughn ◽  
Martin A. Vaughan ◽  
Patrick Camilleri
Keyword(s):  
Electron Microscopy ◽  
Chlorophyll Fluorescence ◽  
Resistance Mechanism ◽  
Cross Resistance ◽  
Oxygen Generator ◽  
Conyza Bonariensis ◽  
Singlet Oxygen Generator ◽  
Chlorophyll Bleaching ◽  
Direct Measures ◽  
Hairy Fleabane

Cross-resistance of the paraquat-resistant (R) hairy fleabane to other compounds that accept electrons from photosystem I (PSI) or produce toxic oxygen species was determined by chlorophyll loss, electron microscopy, and chlorophyll fluorescence suppression. Although the R bioype is approximately 100 x more resistant to paraquat than the susceptible (S) biotype based upon the assays for tissue damage, little or no cross-resistance was observed to a number of other PSI electron acceptors, including the bipyridilium herbicide morfamquat. A low level of resistance (approximately 10-fold) was noted to diquat and the singlet oxygen generator rose bengal. As measured by chlorophyll fluorescence suppression, the R biotype was about 100-fold resistant to paraquat, but only 10-fold resistant to diquat, and exhibited no resistance to morfamquat. Because differences observed with this protocol are direct measures of the ability of the herbicide to reach the active site and the results correlate with the level of resistance observed by chlorophyll bleaching or electron microscopy, these data suggest that compartmentalization is the major factor in paraquat resistance in hairy fleabane.

scholarly journals First Report of Botrytis cinerea Causing Gray Mold Disease on Peach from Pakistan

2018 ◽  
Vol 7 (3) ◽  
pp. 131-131
Author(s):  
Raees Ahmed ◽  
Amjad S. Gondal ◽  
Muhammad Tariq Khan ◽  
Shazia Shahzaman ◽  
Sajjad Hyder
Keyword(s):  
Botrytis Cinerea ◽  
Mycelial Growth ◽  
Agar Medium ◽  
Management Strategies ◽  
Gray Mold ◽  
First Report ◽  
Average Production ◽  
Conducive Environment ◽  
Hyphal Tip ◽  
Important Disease

Gray mold caused by Botrytis cinerea is an important disease that attacks fruits, leaves and twigs of peach. Peach is grown on an area of 18,008 ha with an average production of 72,085 tons per year in Pakistan (FAO, 2017). During May 2017, brown spots on 33% of the peach fruits examined were observed in Swat district of KPK province of Pakistan. Infected fruits were incubated at 25±2 °C in a humid chamber resulted in greyish mycelial growth with light brown lesions. Hyphal growths on infected fruits were cultured on PDA media and purified by hyphal tip method. Morphologically whitish grey growth was observed on PDA and later on dark sclerotia were observed after 6-7 days of incubation. Hyphae were found septate with branched hyaline conidiophores having a bunch of ovoid conidia at their tips. Further confirmations were done by amplifying internal transcribed spacer regions (Andrew et al., 2009) and glyceraldehyde-3-phosphate dehydrogenase (G3PDH) region of the isolates (Li et al., 2012). Amplicons sequenced from Macrogen Korea were blasted and submitted in NCBI showed that ITS sequences (Accessions MH049690 and MH049691) were 99% identical with already reported (MG878388 and MG654661) sequences and the G3PDH gene sequences (Accessions MH560352 and MH560353) were 99 % identical with already reported (Accessions MG204876) sequences of B. cinerea. Pathogenicity was confirmed on healthy peach fruits disinfected with 50% ethanol, inoculated by placing a plug of about 1cm2 taken from the edge of actively growing B. cinerea isolate (BTS-16). Fruits were incubated at 25±2 °C in a humid chamber (Abata et al., 2016). A set of healthy fruits mock-inoculated with a plug of agar medium were used as control. Three days after inoculation, inoculated fruits showed sunken lesions with cottony greyish mycelial growth on their surface. Fungus isolated from these infections was re-confirmed as B. cinerea. Conducive environment for the disease progression in nearby areas can result into a huge loss in peach produce so there is a need to devise management strategies to cope with the pathogen. This is the first report of gray mold disease of peach caused by B. cinerea from Pakistan. 

scholarly journals Fungicide resistance characterised across seven chemical classes in a Botrytis cinerea population isolated from Australian vineyards

2021 ◽  
Author(s):  
Lincoln A. Harper ◽  
Scott Paton ◽  
Barbara Hall ◽  
Suzanne McKay ◽  
Richard P. Oliver ◽  
...  
Keyword(s):  
Adverse Effect ◽  
Botrytis Cinerea ◽  
Fungicide Resistance ◽  
Chemical Control ◽  
Management Strategies ◽  
Resistance Management ◽  
Gray Mold ◽  
Chemical Groups ◽  
Important Disease ◽  
Selection Of

AbstractGray mold, caused by Botrytis cinerea, is an economically important disease of grapes in Australia and across grape growing regions worldwide. Control of this disease relies heavily on canopy management and the application of single site fungicides. Fungicide application can lead to the selection of fungicide resistant B. cinerea populations, which has an adverse effect on the chemical control of the disease. Characterising the distribution and severity of resistant B. cinerea populations is needed to inform resistance management strategies. In this study, 725 isolates were sampled from 75 Australian vineyards during 2013 – 2016 and were screened against seven fungicides with different MOAs. The resistance frequencies for azoxystrobin, boscalid, fenhexamid, fludioxonil, iprodione, pyrimethanil and tebuconazole were 5, 2.8, 2.1, 6.2, 11.6, 7.7 and 2.9% respectively. Nearly half of the resistant isolates (43.7%) were resistant to more than one of the fungicides tested. The frequency of vineyards with at least one isolate simultaneously resistant to 1, 2, 3, 4 or 5 fungicides was 19.5, 7.8, 6.5, 10.4 and 2.6%.Resistance was associated with previously published genotypes in CytB (G143A), SdhB (H272R/Y), Erg27 (F412S), Mrr1 (D354Y), Os1 (I365S, N373S + Q369P, I365S + D757N) and Pos5 (P319A, L412F). Expression analysis was used to characterise fludioxonil resistant isolates exhibiting overexpression (6.3 - 9.6-fold) of the ABC transporter encoded by AtrB (MDR1 phenotype). Novel genotypes were also described in Mrr1 (S611N, D616G) and Cyp51 (P357S). Resistance frequencies were lower when compared to most previously published surveys of both grape and non-grape B. cinerea resistance. Nonetheless, continued monitoring of critical chemical groups used in Australian vineyards is recommended.

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