A Conservative Mutant Version of the Mrr1 Transcription Factor Correlates with Reduced Sensitivity to Fludioxonil in Botrytis cinerea.

Fludioxonil is a highly effective phenylpyrrole fungicide for controlling Botrytis cinerea. Although the field efficacy of fludioxonil remains high, Botrytis cinerea isolates with reduced sensitivity have been reported globally. The molecular target of fludioxonil still remains unknown; however, a mechanism of reduced sensitivity to fludioxonil underlies the overexpression of the ATP binding cassette (ABC) transporter AtrB in a dependent pathway of the Mrr1 transcription factor. Fludioxonil is a key player in controlling B. cinerea infection in table grapes in Chile. However, some isolates with a reduced sensitivity to fludioxonil were detected. This study observed endogenous atrB overexpression in Chilean isolates with reduced sensitivity to fludioxonil (n = 22) compared to the sensitive isolates (n = 10). All isolates increased the expression of atrB in a growth medium supplemented with fludioxonil (0.05 µg/mL). However, sensitive isolates showed lower atrB expression than those with reduced fludioxonil sensitivity. Remarkably, a mutant version of the transcription factor Mrr1 carrying 21 amino acid modifications was identified in all isolates with reduced sensitivity to fludioxonil. These changes alter the protein's transcription factor domain and the C-terminal portion of the protein but not the Zn (2)-C6 fungal-type DNA-binding domain. These results suggest a direct relationship between the conserved and divergent mutant version of mrr1 and sensitivity to fludioxonil. This study provides a new target for developing molecular diagnostic strategies to monitor B. cinerea's sensitivity to fludioxonil in the field.

Chilean Botrytis cinerea Isolates with Reduced Sensitivity to Fludioxonil Exhibit Low to Null Fitness Penalties.

The main phytosanitary problem for table grape production in Chile is gray mold caused by the fungus Botrytis cinerea. To manage this issue, the primary method utilized is chemical control. Fludioxonil, a phenylpyrrole, is highly effective in controlling B. cinerea and other plant pathogens. Consistently, there have been no field reports of reduced efficacy of fludioxonil; however, subpopulations with reduced sensitivity to fludioxonil are on the rise globally, as per increasing reports. Our study involved a large-scale evaluation of B. cinerea's sensitivity to fludioxonil in the Central Valley of Chile's primary table grape production area during the growing seasons from 2015 to 2018. Out of 2,207 isolates, only 1.04% of the isolates (n = 23) exceeded the sensitivity threshold value of 1 µg/ml. Remarkably, 95.7% are concentrated in a geographic region (Valparaíso Region). Isolates with reduced sensitivity to fludioxonil showed growth comparable with sensitive isolates and even more robust growth under nutritional deficit, temperature, or osmotic stress, suggesting greater environmental adaptation. When table grape detached berries were stored at 0°C, isolates less sensitive to fludioxonil caused larger lesions than sensitive isolates (2.82 mm compared with 1.48 mm). However, the lesions generated by both types of isolates were equivalent at room temperature. This study found no cross-resistance between fludioxonil and fenhexamid, an essential fungicide integrated with fludioxonil in Chilean B. cinerea control programs. All the Chilean isolates with reduced sensitivity to fludioxonil were controlled by the fludioxonil/cyprodinil mixture, a commonly employed form of fludioxonil. The cyprodinil sensitivity in the isolates with reduced sensitivity to fludioxonil explains their low field frequency despite their null fitness penalties. However, the emergence of fludioxonil-resistant isolates inside the Chilean B. cinerea population demands a comprehensive analysis of their genetic bases, accompanied by monitoring tools that allow the permanence of field fludioxonil efficacy.

Diversity of fungi in seeds of hymenaea stignocarpa and hymenaea courbaril before and after fungicide treatments / Incidência de fungos no tratamento de sementes de hymenaea stignocarpa e hymenaea courbaril

The recovery of degraded areas led to an increase of native forest seedlings production. Jatobado- cerrado (Hymenaea stignocarpa Mart.) and jatoba-da-mata (Hymenaea courbaril L.), native species of the Brazilian Cerrado, are widely used for the recovery of this biome. Although forest species present a diversity of pathogens disseminated by seeds, few studies have been published and no study was found for jatoba. Because the seed is one of the main forms of pathogen dissemination, the aim of our work was to evaluate the natural incidence of fungi in seed lots of jatoba-do-cerrado and jatoba-da-mata and the performance of different fungicides for seed treatment recommendation. The experiment was carried out at the Genetics and Biotechnology Laboratory of the University Center of Patos de Minas. Two chemical treatments were submitted in the seeds and then divided into seven replicates of 15 seeds for each species. The first treatment was with the active principle thiophanate-methyl and fluazine, commercial name Certeza®, in the dose of 145 mL of the product in 100 kg of seeds. The second treatment was with fludioxonil, commercial name Maxim® XL, in the dose of 100 mL of the product in 100 kg of seeds. There was a third treatment corresponded to the control (without seed treatment). The visual analyzes were performed after eight days on each seed with a magnifier to verify the presence of developing fungal colonies. The growth fungi fructifications were observed using a stereomicroscope. The found fungi species were: Aspergillus sp., Fusarium sp., Mucor sp., Penicillium sp., Pestalotiopsis sp., Phomopsis sp., Rhizoctonia sp., Rhizopus sp. and Trichoderma sp. It occurred interaction between seed treatment and species for Aspergillus sp., Fusarium sp., Mucor sp., Phomopsis sp., Rhizoctonia sp. and Trichoderma sp. For Penicillium sp., Pestalotiopsis sp. and Rhizopus sp., only the isolated factors were significant. Jatoba-do-cerrado seeds had a higher incidence than jatoba-da-mata seeds in all the found fungi. Certeza® and Maxim® XL have potential in seed treatment for the species. The fungicide Certeza® was more efficient to control the pathogenic fungi in seed treatment.

A recuperação de áreas degradas levou ao aumento da produção de mudas de espécies florestais nativas. Jatobá-do-cerrado (Hymenaea stignocarpa Mart.) e jatobá-da-mata (Hymenaea courbaril L.), espécies nativas do Cerrado brasileiro, são amplamente utilizadas na recuperação deste bioma. Apesar da diversidade de fungos encontrada em sementes de espécies florestais, poucos são os estudos publicados, e não nenhum relato para o jatobá. Por a semente ser um dos principais meios de disseminação de patógenos, o objetivo do trabalho foi avaliar a incidência natural de fungos em lotes de semente de jatobá-do-cerrado e jatobá-da-mata, e a performance de diferentes fungicidas para a recomendação do tratamento de sementes. O experimento foi realizado no Laboratório de Genética e Biotecnologia do Centro Universitário de Patos de Minas. Dois tratamentos químicos foram realizados nas sementes, que então foram divididas em sete repetições com 15 sementes cada. O primeiro tratamento foi realizado com o princípio ativo tiofanato-metílico e fluazinam, nome comercial Certeza®, na dose de 145 mL do produto para 100 kg de sementes. O segundo tratamento foi realizado com fludioxonil, nome comercial Maxim® XL, na dose de 100 mL do produto para 100 kg de sementes. O terceiro tratamento correspondeu a testemunha (sem tratamento de sementes). Após oito dias, a análise visual foi realizada com o auxílio de uma lupa para verificar a presença de colônias de fungos. As frutificações dos fungos foram observadas com o auxílio de um microscópio óptico. Após as avaliações, os seguintes fungos foram encontrados: Aspergillus sp., Fusarium sp., Mucor sp., Penicillium sp., Pestalotiopsis sp., Phomopsis sp., Rhizoctonia sp., Rhizopus sp. e Trichoderma sp. Ocorreu interação entre o tratamento de sementes e as espécies para Aspergillus sp., Fusarium sp., Mucor sp., Phomopsis sp., Rhizoctonia sp. e Trichoderma sp. Para Penicillium sp., Pestalotiopsis sp. e Rhizopus sp., apenas os fatores isolados foram significativos. Sementes de jatobá-do-cerrado possuíram maior incidência do que as sementes de jatobá-damata para todos os fungos encontrados. Certeza® e Maxim® XL possuem potencial para o tratamento de sementes de jatobá. O fungicida Certeza® foi mais eficiente para o controle de fungos patogênicos no tratamento de sementes.

Differential sensitivity of developmental stages of the South American toad to a fungicide based on fludioxonil and metalaxyl-M.

Agricultural fungicide application in Argentina has increased twice since 2008, with Maxim® XL (2.5% fludioxonil +1% metalaxyl-M) as one of the most used fungicide formulation. The toxicity of this pesticide on Rhinella arenarum was assessed by means of continuous (from embryo and larval development) and 24-h pulse exposure standardized bioassays. Lethality was concentration- and exposure time-dependent. Maxim® XL caused a progressive lethal effect along the bioassays with higher toxicity on embryos than larvae, obtaining 50% lethal concentrations at 96, 336, and 504 h of 10.85, 2.89, and 1.71 mg/L for embryos, and 43.94, 11.79, and 5.76 mg/L for larvae respectively. Lethal 504-h no observed effect concentration values for embryos and larvae were 1 and 2.5 mg/L respectively. A stage-dependent toxicity of Maxim® XL was also demonstrated within the embryo development, with early stages more sensitive than the later ones, and blastula as the most sensitive developmental stage. The risk quotients obtained for chronic risk assessment determined a potential threat for the survival and continuity of R. arenarum populations under these conditions. The results indicate that the levels of the fungicide reaching amphibian habitats could be risky for the early development of this amphibian species. This study also emphasizes the necessity to evaluate the chronic effects of fungicides in pesticide risk assessment.

Effect of Fungicides and Storage Conditions on Postharvest Development of Citrus Black Spot and Survival of Guignardia citricarpa in Fruit Tissues.

Citrus black spot (CBS) is caused by Guignardia citricarpa, which incites lesions on citrus fruit and can induce fruit drop. Quiescent infections occur during the spring and summer, and symptoms appear at fruit maturity or after harvest. Thus, fruit from citrus areas affected by CBS represent a risk for introduction of this pathogen into new areas. The effects of preventive field fungicide programs, postharvest fungicide drenches, packinghouse fungicide applications, and storage temperatures on postharvest symptom development and viability of G. citricarpa in lesions were evaluated in five experiments on Murcott tangor, Valencia oranges, and lemons. Preventive field treatments and fruit storage at 8°C consistently reduced postharvest CBS development, whereas a postharvest fungicide drench or packinghouse treatment with fungicides had no effect on postharvest symptom development. In a separate experiment, postharvest appearance of symptoms was related to the percentage of fruit with symptoms at harvest. The preventive field fungicide program also consistently reduced the percentage of isolation of G. citricarpa from affected fruit, whereas storage temperature and packinghouse fungicide treatment gave variable results. The viability of the fungus declined with storage time of fruit after harvest, but G. citricarpa could still be readily isolated regardless of treatment. In another experiment, the viability of the fungus in detached fruit or peel was minimally affected by temperature or moisture during storage. The frequency of successful isolation declined with time, but G. citricarpa was still recovered frequently from symptomatic tissue at later times. The most effective means to reduce postharvest development of symptoms is through preventive application of fungicides during the fruit growing season and storage of harvested fruit at cold temperatures. None of the measures evaluated substantially reduced viability of G. citricarpa, and the pathogen would likely be introduced on symptomatic fruit from citrus areas with CBS.