Gray mold in strawberries in the Paraná state of Brazil is caused by Botrytis cinerea and its isolates exhibit multiple-fungicide resistance

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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Characterization of Botrytis cinerea From Commercial Cut Flower Roses

Plant Disease ◽

10.1094/pdis-09-18-1623-re ◽

2019 ◽

Vol 103 (7) ◽

pp. 1577-1583 ◽

Cited By ~ 7

Author(s):

M. Muñoz ◽

J. E. Faust ◽

G. Schnabel

Keyword(s):

Botrytis Cinerea ◽

Fungicide Resistance ◽

Management Practices ◽

Cultural Practices ◽

Disease Incidence ◽

Resistance Management ◽

Gray Mold ◽

Economic Losses ◽

Cut Flower ◽

Aspergillus Brasiliensis

Botrytis cinerea Pers. infects cut flower roses (Rosa × hybrida L.) during greenhouse production and gray mold symptoms are often expressed in the postharvest environment, resulting in significant economic losses. Disease management is based on cultural practices and preventative chemical treatments; however, gray mold outbreaks continue to occur. Rose tissues from six commercial shipments from two greenhouses in Colombia were evaluated to determine the Botrytis species composition as well as identify other pathogens present, gray mold incidence and severity, and fungicide resistance profiles. Botrytis isolates (49 total) were grouped into six morphological phenotypes, and all were identified to be B. cinerea sensu stricto. Disease incidence was higher in the petals than in the stem, stamen, ovary, sepal, or leaf tissues. Other fungi were isolated infrequently and included Alternaria alternata, Cladosporium cladosporioides, Epicoccum nigrum, Penicillium citrinum, Aspergillus brasiliensis, and Diplodia sp. Fungicide resistance profiles were determined using previously established discriminatory doses. Isolates resistant to thiophanate-methyl, iprodione, boscalid, and cyprodinil were found frequently in all shipments and in both greenhouses. The frequency of resistance to penthiopyrad, fenhexamid, fluopyram, isofetamid, and fludioxonil varied between shipments and greenhouses. No resistance to pydiflumetofen was observed at the discriminatory doses tested. Isolates with resistance to multiple chemical classes were commonly found. These results indicate that fungicide resistance management practices may improve preharvest and postharvest gray mold control of cut flower roses.

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Population Genetics and Fungicide Resistance of Botrytis cinerea on Vitis and Prunus spp. in California

Phytopathology ◽

10.1094/phyto-09-19-0362-r ◽

2020 ◽

Vol 110 (3) ◽

pp. 694-702 ◽

Cited By ~ 2

Author(s):

Jeffery A. DeLong ◽

Seiya Saito ◽

Chang-Lin Xiao ◽

Rachel P. Naegele

Keyword(s):

Botrytis Cinerea ◽

Fungicide Resistance ◽

Gray Mold ◽

Broad Host Range ◽

Multilocus Genotype ◽

Postharvest Diseases ◽

High Genetic Diversity ◽

Action Committee ◽

Vitis Sp ◽

Prunus Spp

Botrytis cinerea, the causal agent of gray mold, has high genetic diversity and a broad host range. In Vitis sp. and Prunus spp., B. cinerea causes pre- and postharvest diseases, and fungicides are routinely applied to prevent yield loss. In total, 535 isolates of B. cinerea collected from Vitis sp. and Prunus spp. in 2012, 2016, and 2017 were genotyped using 18 microsatellite markers and the transposable elements (TEs) Boty and Flipper. Only nine of the polymorphic markers and the two TEs were considered informative and retained for the final analyses. Of the 532 isolates, 297 were tested for resistance to seven fungicides representing six Fungicide Resistance Action Committee classes. After clone correction, 295 multilocus genotype groups were retained across the 3 years in 326 individuals, and four genetic subpopulations were detected. High levels of clonality were observed across the dataset. Significant pairwise differentiation was detected among years, locations, and TE composition. However, most of the diversity observed was within a subpopulation and not among subpopulations. No genetic differentiation was detected among resistant and sensitive isolates for individual fungicide classes. When resistance to the total number of fungicides was compared, regardless of the fungicide class, significant differentiation was detected among isolates that are resistant to two fungicide classes and those resistant to three or four fungicide groups. Fungicide resistance frequencies were stable for most chemistries evaluated with the exception of fluopyram, which increased from 2012 to 2016/2017.

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Fungicide Resistance Profiling in Botrytis cinerea Populations from Blueberry in California and Washington and Their Impact on Control of Gray Mold

Plant Disease ◽

10.1094/pdis-02-16-0229-re ◽

2016 ◽

Vol 100 (10) ◽

pp. 2087-2093 ◽

Cited By ~ 25

Author(s):

S. Saito ◽

T. J. Michailides ◽

C. L. Xiao

Keyword(s):

Botrytis Cinerea ◽

Fungicide Resistance ◽

Resistance Management ◽

Central Valley ◽

Gray Mold ◽

Postharvest Disease ◽

Quinone Outside Inhibitor ◽

Development Of Resistance ◽

Western Washington ◽

Moderately Resistant

Gray mold caused by Botrytis cinerea is a major postharvest disease of blueberry grown in the Central Valley of California and western Washington State. Sensitivities to boscalid, cyprodinil, fenhexamid, fludioxonil, and pyraclostrobin, representing five different fungicide classes, were examined for 249 (California) and 106 (Washington) B. cinerea isolates recovered from decayed blueberry fruit or flowers. In California and Washington, 7 and 17 fungicide-resistant phenotypes, respectively, were detected: 66 and 49% of the isolates were resistant to boscalid, 20 and 29% were moderately resistant to cyprodinil, 29 and 29% were resistant to fenhexamid, and 66 and 55% were resistant to pyraclostrobin. All isolates from California were sensitive to fludioxonil, whereas 70% of the isolates from Washington showed reduced sensitivity to fludioxonil. In California, 26 and 30% of the isolates were resistant to two and three classes of fungicides, respectively. In Washington, 31, 14, 16, and 9% of the isolates were resistant to two, three, four, and five classes of fungicides, respectively. Inherent risk of the development of resistance to quinone outside inhibitor (QoI) fungicides was assessed by detecting the presence of the Bcbi-143/144 intron in gene cytb. The intron was detected in 11.8 and 40% of the isolates in California and Washington, respectively, suggesting that the risk of QoI resistance is higher in California than in Washington. On detached blueberry fruit inoculated with 11 isolates exhibiting different fungicide-resistant phenotypes, most fungicides failed to control gray mold on fruit inoculated with the respective resistant phenotypes but the mixture of cyprodinil and fludioxonil was effective against all fungicide-resistant phenotypes tested. Our findings would be useful in designing and implementing fungicide resistance management spray programs for control of gray mold in blueberry.

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

Plant Disease ◽

10.1094/pdis-01-17-0067-re ◽

2017 ◽

Vol 101 (7) ◽

pp. 1306-1313 ◽

Cited By ~ 23

Author(s):

Dolores Fernández-Ortuño ◽

Alejandro Pérez-García ◽

Manuel Chamorro ◽

Eduardo de la Peña ◽

Antonio de Vicente ◽

...

Keyword(s):

Amino Acid ◽

Botrytis Cinerea ◽

Fungicide Resistance ◽

Mycelial Growth ◽

Growing Season ◽

Gray Mold ◽

Effective Concentration ◽

Site Specific ◽

Resistance Patterns ◽

Succinate Dehydrogenase Inhibitors

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.

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Botrytis fragariae, a New Species Causing Gray Mold on Strawberries, Shows High Frequencies of Specific and Efflux-Based Fungicide Resistance

Applied and Environmental Microbiology ◽

10.1128/aem.00269-17 ◽

2017 ◽

Vol 83 (9) ◽

Cited By ~ 24

Author(s):

Sabrina Rupp ◽

Cecilia Plesken ◽

Sibylle Rumsey ◽

Madeline Dowling ◽

Guido Schnabel ◽

...

Keyword(s):

United States ◽

New Species ◽

Multidrug Resistance ◽

Botrytis Cinerea ◽

Fungicide Resistance ◽

Southeastern United States ◽

Gray Mold ◽

Diagnostic Pcr ◽

Content Type ◽

A New Species

ABSTRACT Botrytis cinerea causes pre- and postharvest decay of many fruit and vegetable crops. A survey of German strawberry fields revealed Botrytis strains that differed from B. cinerea in diagnostic PCR markers and growth appearance. Phylogenetic analyses showed that these strains belong to an undescribed species in Botrytis clade 2, named Botrytis fragariae sp. nov. Isolates of B. fragariae were detected in strawberry fields throughout Germany, sometimes at frequencies similar to those of B. cinerea, and in the southeastern United States. B. fragariae was isolated from overwintering strawberry tissue but not from freshly infected fruit. B. fragariae invaded strawberry tissues with an efficiency similar to or lower than that of B. cinerea but showed poor colonization of inoculated nonhost plant tissues. These data and the exclusive occurrence of this fungus on strawberry plants indicate that B. fragariae is host specific and has a tissue preference different from that of B. cinerea. Various fungicide resistance patterns were observed in B. fragariae populations. Many B. fragariae strains showed resistance to one or several chemical classes of fungicides and an efflux-based multidrug resistance (MDR1) phenotype previously described in B. cinerea. Resistance-related mutations in B. fragariae were identical or similar to those of B. cinerea for carbendazim (E198A mutation in tubA), azoxystrobin (G143A in cytB), iprodione (G367A+V368F in bos1), and MDR1 (gain-of-function mutations in the transcription factor mrr1 gene and overexpression of the drug efflux transporter gene atrB). The widespread occurrence of B. fragariae indicates that this species is adapted to fungicide-treated strawberry fields and may be of local importance as a gray mold pathogen alongside B. cinerea. IMPORTANCE Gray mold is the most important fruit rot on strawberries worldwide and requires fungicide treatments for control. For a long time, it was believed to be caused only by Botrytis cinerea, a ubiquitous pathogen with a broad host range that quickly develops fungicide resistance. We report the discovery and description of a new species, named Botrytis fragariae, that is widely distributed in commercial strawberry fields in Germany and the southeastern United States. It was observed on overwintering tissue but not on freshly infected fruit and seems host specific on the basis of its occurrence and artificial infection tests. B. fragariae has also developed resistance to several fungicides that is caused by mutations similar to those known in B. cinerea, including an efflux-based multidrug resistance. Our data indicate that B. fragariae could be of practical importance as a strawberry pathogen in some regions where its abundance is similar to that of B. cinerea.

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Sources of Primary Inoculum of Botrytis cinerea and Their Impact on Fungicide Resistance Development in Commercial Strawberry Fields

Plant Disease ◽

10.1094/pdis-02-17-0203-re ◽

2017 ◽

Vol 101 (10) ◽

pp. 1761-1768 ◽

Cited By ~ 17

Author(s):

Michelle Souza Oliveira ◽

Achour Amiri ◽

Adrian I. Zuniga ◽

Natalia A. Peres

Keyword(s):

United States ◽

Botrytis Cinerea ◽

Fungicide Resistance ◽

Integrated Approach ◽

The United States ◽

Gray Mold ◽

Resistance Development ◽

Primary Inoculum ◽

Potential Source ◽

Northern United States

Strawberry transplants produced in nurseries across Canada, northern United States, and California are shipped annually to other strawberry-growing regions, including Florida. Botrytis cinerea, the causal agent of gray mold, causes latent infections on transplants which are suggested as a potential source of primary inoculum in strawberry fields. In this study, we investigated the survival of B. cinerea isolates over the summer in Florida, the presence of B. cinerea in transplants from 14 nurseries from Canada and the United States in 2011, 2012, and 2013, and the sensitivity of nursery population to several botryticides. Botrytis cinerea was detected on dead strawberry plants sampled from commercial strawberry fields between March and June but not in July and August, suggesting that the fungus does not over-summer in strawberry fields in Florida. Nursery transplants surveyed in 2011, 2012, and 2013 showed B. cinerea incidences of 20 to 37, 20 to 83, and 2.5 to 92.5%, respectively. In total, 409 isolates were tested for sensitivity to pyraclostrobin, boscalid, pyrimethanil, fenhexamid, iprodione, penthiopyrad, fluopyram, and fludioxonil. Overall, respective resistance frequencies were 91.7, 79.3, 33.2, 20.7, 2.4, 0.2, 0.2, and 0.0%. A majority of isolates tested were resistant to either 3 or 4 fungicides simultaneously. These findings reinforce the need for an integrated approach between strawberry nurseries and production fields to improve gray mold management and mitigate future risks of resistance development in B. cinerea.

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Characterization of the Field Fludioxonil Resistance and Its Molecular Basis in Botrytis cinerea from Shanghai Province in China

Microorganisms ◽

10.3390/microorganisms9020266 ◽

2021 ◽

Vol 9 (2) ◽

pp. 266

Author(s):

Weizhen Wang ◽

Yuan Fang ◽

Muhammad Imran ◽

Zhihong Hu ◽

Sicong Zhang ◽

...

Keyword(s):

Amino Acid ◽

Botrytis Cinerea ◽

Fungicide Resistance ◽

Molecular Basis ◽

Resistance Mechanisms ◽

Gray Mold ◽

Amino Acid Sequences ◽

Necrotrophic Pathogen ◽

Moderate Resistance

Botrytis cinerea is a destructive necrotrophic pathogen that can infect many plant species. The control of gray mold mainly relies on the application of fungicides, and the fungicide fludioxonil is widely used in China. However, the field fungicide resistance of B. cinerea to this compound is largely unknown. In this study, B. cinerea isolates were collected from different districts of Shanghai province in 2015–2017, and their sensitivity to fludioxonil was determined. A total of 65 out of 187 field isolates (34.76%) were found to be resistant to fludioxonil, with 36 (19.25%) showing high resistance and 29 (15.51%) showing moderate resistance. Most of these resistant isolates also showed resistance to iprodione, and some developed resistance to fungicides of other modes of action. AtrB gene expression, an indicator of MDR1 and MDR1h phenotypes, was not dramatically increased in the tested resistant isolates. Biological characteristics and osmotic sensitivity investigations showed that the fitness of resistant isolates was lower than that of sensitive ones. To investigate the molecular resistance mechanisms of B. cinerea to fludioxonil, the Bos1 amino acid sequences were compared between resistant and sensitive isolates. Resistant isolates revealed either no amino acid variations or the mutations I365S, I365N, Q369P/N373S, and N373S.

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Advances in understanding fungicide resistance in Botrytis cinerea in China

Phytopathology ◽

10.1094/phyto-07-20-0313-ia ◽

2020 ◽

Author(s):

Wenyong Shao ◽

Youfu Zhao ◽

Zhonghua Ma

Keyword(s):

Botrytis Cinerea ◽

Fungicide Resistance ◽

Target Genes ◽

Agricultural Practices ◽

Resistance Mechanisms ◽

Gray Mold ◽

Yield Losses ◽

Quinone Outside Inhibitor ◽

Significant Yield ◽

Fungicide Target

Gray mold, caused by Botrytis cinerea, is a devastating disease causing significant yield losses in various economic plants. Fungicide application is one of the main strategies for management of gray mold; however, B. cinerea has developed resistance to various groups of fungicide. In China, benzimidazole-, dicarboximide-, and quinone outside inhibitor (QoI)-resistant populations of B. cinerea have become dominant. Substitute mutations in fungicide target genes are responsible for resistance in B. cinerea. Based on known resistance mechanisms, molecular methods including loop-mediated isothermal amplification (LAMP) have been developed for rapid detection of resistant isolates of B. cinerea. Since B. cinerea is able to quickly develop resistance to various fungicides, various integrated strategies have been developed, including biological and agricultural practices, to manage fungicide resistance in B. cinerea in the last decade.

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