Identification, biological characteristics, and fungicide screening of pathogen of black spot in Aconitum carmichaelii / 中国中药杂志

In Hezhang county, Guizhou province, black spot tends to occur to Aconitum carmichaelii in the hot rainy summer, with the incidence up to 50%-70%, seriously impacting the yield and quality of the medicinal material. Thus, this study aims to clarify the pathogen and the occurrence characteristics. To be specific, the pathogen was isolated and identified according to Koch's postulates and the pathogenicity and biological characteristics were determined. In addition, the sensitivity of the pathogen to four microbial fungicides, four botanical fungicides, and five chemical fungicides was determined with the mycelium growth rate method for the purpose of screening out optimal fungicides. The pathogen was identified as Alternaria alternate, as evidenced by the similar colony morphology and microscopic characteristics and 99.55%-100% similarity in sequences of rDNA-ITS, LSU, 18S, and TEF of the two. The optimum growth conditions for A. alternata were 28 ℃, pH 8, and continuous darkness. Bacillus subtilis had strong inhibitory effect on the pathogen, and the inhibition rate was more than 90% when the concentration was 1 mg·L~(-1). In addition, difenoconazole and quinoline copper can also control the pathogen, with median effective concentration(EC_(50)) of 2.92 and 9.02 mg·L~(-1), respectively. This study lays a theoretical basis for the field control of black spot in A. carmichaelii.

Effect of different fungicides on efficiency of mycorrhizal symbiosis on Salvia miltiorrhiza / 中国中药杂志

Arbuscular mycorrhizal fungi provided is beneficial to Salvia miltiorrhiza for increasing yield, promoting the accumulation of active ingredients, and alleviating S. miltiorrhiza disease etc. However, the application of fungicides will affect the benefit of arbuscular mycorrhizal fungi and there is little research about it. This article study the effect of four different fungicides: carbendazim, polyoxin, methyl mopazine, and mancozeb on mycorrhiza benefit to S. miltiorrhiza by the infection intensity of arbuscular mycorrhizal fungi, the growth of S. miltiorrhiza, and the content of active ingredients. RESULTS:: showed that different fungicides had different effects. The application of mancozeb had the strongest inhibitory effect on the mycorrhizal benefit to S. miltiorrhiza. Mancozeb significantly reduced the mycorrhizal colonization and the beneficial effect of arbuscular mycorrhizal fungi on the growth and the accumulation of active components of S. miltiorrhiza. The application of polyoxin had no significant effect on mycorrhizal colonization. Instead, it had a synergistic effect with the mycorrhizal benefit to promoting the growth and accumulation of rosmarinic acid of S. miltiorrhiza. The inhibitory strengths of four fungicides are: mancozeb>thiophanate methyl, carbendazim>polyoxin. Therefore, we recommend applying biological fungicides polyoxin and avoid applying chemical fungicides mancozeb for disease control during mycorrhizal cultivation of S. miltiorrhiza.

Identification,biological characteristics and fungicide screening of pathogen of southern blight in Cynanchum stauntonii / 中国中药杂志

During the high-temperature and rainy season from June to October in 2017-2019,serious southern blight broke out in the Cynanchum stauntonii planting area in Tuanfeng county,Hubei province,which had a great impact on the yield and quality of medicinal materials. In this study,the pathogen of C. stauntonii was isolated,purified,and identified,and the pathogenicity was tested according to Koch's postulates. Meanwhile,the biological characteristics of the pathogen were analyzed. On this basis,the effective fungicides were screened in laboratory. Finally,the pathogen( BQ-1) was identified as Athelia rolfsii( Deuteromycotina,Basidiomycota,anamorph: Sclerotium rolfsii). The optimum growth conditions for BQ-1 were 25-30 ℃,p H 5-8,and alternating light and dark.The effective chemical fungicides were lime-sulphur-synthelic-solution( LSSS) and flusilazole,and the effective botanical fungicide was osthole. BQ-1 was highly homologous to the pathogen HS-1 of peanut southern blight,with the similarity of 18 S r DNA and TEF sequences at 99. 09%. The southern blight in C. stauntonii might be resulted from that in peanut. In the production of C. stauntonii,the following measures should be taken: avoiding rotation or neighboring with peanut,draining water from June to October to reduce humidity,and reasonably applying fungicides.

Tolerance of dark septate endophytic fungi (DSE) to 1 agrochemicals in vitro / Tolerancia de hongos endofíticos septados oscuros a agroquímicos in vitro

Abstract Dark septate endophytes (DSE) are a heterogeneous group of fungi, mostly belonging to the Phylum Ascomycota, that are involved in a mutualistic symbiosis with plant roots. The aim of this study is to evaluate the behavior of two strains of DSE isolated from wheat roots of two cropping areas in the province of Buenos Aires, Argentina, against some agrochemicals. Of all the isolates obtained, two strains were identified as Alternaria alternata and Cochliobolus sp. These DSE were found to be tolerant to glyphosate, carbendazim and cypermethrin when evaluated at the recommended agronomic dose (AD), 2 AD and, in some cases, 10 AD. This work contributes to the study of the biology of this group of fungi and their tolerance in the presence of xenobiotics widely used in agriculture.© 2019 Asociaci´on Argentina de Microbiolog´ıa. Published by Elsevier Espa˜na, S.L.U. This is an open access article under the CC BY-NC-ND license (https://creativecommons.org/licenses/bync-nd/4.0/).

Resumen Los endófitos septados oscuros (DSE) son un grupo heterogéneo de hongos que participan de una simbiosis mutualista con raíces de plantas, perteneciendo principalmente al Phylum Ascomycota. El objetivo de este estudio fue aislar DSE de raíces de trigo proveniente de dos áreas de cultivo de la provincia de Buenos Aires y evaluar el comportamiento de dos cepas de DSE aisladas de raíces de trigo frente a algunos agroquímicos en dos áreas de cultivo de la provincia de Buenos Aires. De todos los aislamientos obtenidos se seleccionaron dos cepas que se identificaron como Alternaria alternata y Cochliobolus sp. Se encontró que estos DSE son tolerantes al glifosato, el carbendazim y la cipermetrina, evaluados a las dosis agronómicas recomendadas (AD), a 2x AD y, en algunos casos, a 10x AD. Este trabajo contribuye al conocimiento de la biología de este grupo de hongos y su tolerancia a xenobióticos ampliamente utilizados en la agricultura.

RNA-seq analysis of the response of plant-pathogenic oomycete Phytophthora parasitica to the fungicide dimethomorph / Análisis RNA-seq de la respuesta del oomiceto fitopatógeno Phytophthora parasitica al fungicida dimetomorf

Phytophthora parasitica is an important oomycete that causes disease in a variety of plants, dimethomorph fungicides being specific for oomycetes. The aim of this study was to use RNA-seq to rapidly discover the mechanism by which dimethomorph acts in the treatment of P. parasitica. We found that the expression of 832 genes changed significantly after the dimethomorph treatment, including 365 up-regulated genes and 467 down-regulated genes. According to the Gene Ontology (GO) enrichment analysis, pathway enrichment and verification test results, the following conclusions are obtained: (i) the treatment of P. parasitica with dimethomorph causes changes in the expression levels of genes associated with the cell wall and cell wall synthesis; (ii) dimethomorph treatment results in reduced permeability of the cell membrane and changes in the expression of certain transport-related proteins; (iii) dimethomorph treatment increased reactive oxygen species and reduced the expression of genes related to the control of oxidative stress.

Phytophthora parasitica es un importante oomiceto que origina enfermedades en una variedad de plantas; el fungicida dimetomorf es específico contra oomicetos. El objetivo de este estudio fue utilizar la tecnología de RNA-seq para descubrir rápidamente el mecanismo por el que el dimetomorf actúa en el tratamiento de P. parasitica. Descubrimos que la expresión de 832 genes se modificaba significativamente tras el tratamiento con dimetomorf, incluyendo 365 genes que son sobrerregulados y 467 genes que son subrregulados. El análisis de enriquecimiento de ontología de genes (GO), análisis de enriquecimiento de las vías y pruebas de verificación permitieron extraer las conclusiones siguientes: 1) el tratamiento de P. parasitica con dimetomorf origina cambios en los niveles de expresión de los genes relacionados con la pared celular y su síntesis; 2) el tratamiento con dimetomorf origina una reducción de la permeabilidad de la membrana celular, así como cambios en la expresión de ciertas proteínas relacionadas con el transporte, y 3) el tratamiento con dimetomorf incrementó las especies reactivas del oxígeno y redujo la expresión de los genes relacionados con el control del estrés oxidativo.

The potential of compounds isolated from Xylaria spp. as antifungal agents against anthracnose

ABSTRACT Anthracnose is a crop disease usually caused by fungi in the genus Colletotrichum or Gloeosporium. These are considered one of the main pathogens, causing significant economic losses, such as in peppers and guarana. The current forms of control include the use of resistant cultivars, sanitary pruning and fungicides. However, even with the use of some methods of controlling these cultures, the crops are not free of anthracnose. Additionally, excessive application of fungicides increases the resistance of pathogens to agrochemicals and cause harm to human health and the environment. In order to find natural antifungal agents against guarana anthracnose, endophytic fungi were isolated from Amazon guarana. The compounds piliformic acid and cytochalasin D were isolated by chromatographic techniques from two Xylaria spp., guided by assays with Colletotrichum gloeosporioides. The isolated compounds were identified by spectrometric techniques, as NMR and mass spectrometry. This is the first report that piliformic acid and cytochalasin D have antifungal activity against C. gloeosporioides with MIC 2.92 and 2.46 µmol mL-1 respectively. Captan and difenoconazole were included as positive controls (MIC 16.63 and 0.02 µmol mL-1, respectively). Thus, Xylaria species presented a biotechnological potential and production of different active compounds which might be promising against anthracnose disease.

Antifungal activity of Gallesia integrifolia fruit essential oil

ABSTRACT Gallesia integrifolia (Phytolaccaceae) is native to Brazil and has a strong alliaceous odor. The objective of this study was to identify the chemical composition of G. integrifolia fruit essential oil and evaluate fungicidal activity against the main food-borne diseases and food spoilage fungi. The essential oil was extracted by hydrodistillation and identified by GC-MS. From 35 identified compounds, 68% belonged to the organosulfur class. The major compounds were dimethyl trisulfide (15.49%), 2,8-dithianonane (52.63%) and lenthionine (14.69%). The utilized fungi were Aspergillus fumigatus, Aspergillus niger, Aspergillus ochraceus, Aspergillus versicolor, Penicillium funiculosum, Penicillium ochrochloron, Penicillium verrucosum var. cyclopium, and Trichoderma viride. Minimal fungicidal concentration for the essential oil varied from 0.02 to 0.18 mg/mL and bifonazole and ketoconazole controls ranged from 0.20 to 3.50 mg/mL. The lower concentration of the essential oil was able to control P. ochrochloron, A. fumigatus, A. versicolor, A. ochraceus and T. viride. This study shows a high fungicidal activity of G. integrifolia fruit essential oil and can support future applications by reducing the use of synthetic fungicides.

Chemical products induce resistance to Xanthomonas perforans in tomato

The bacterial spot of tomato, caused by Xanthomonas spp., is a very important disease, especially in the hot and humid periods of the year. The chemical control of the disease has not been very effective for a number of reasons. This study aimed to evaluate, under greenhouse conditions, the efficacy of leaf-spraying chemicals (acibenzolar-S-methyl (ASM) (0.025 g.L⁻¹), fluazinam (0.25 g.L⁻¹), pyraclostrobin (0.08 g.L⁻¹), pyraclostrobin + methiran (0.02 g.L⁻¹ + 2.2 g.L⁻¹), copper oxychloride (1.50 g.L⁻¹), mancozeb + copper oxychloride (0.88 g.L⁻¹ + 0.60 g.L⁻¹), and oxytetracycline (0.40 g.L⁻¹)) on control of bacterial spot. Tomatoes Santa Clara and Gisele cultivars were pulverized 3 days before inoculation with Xanthomonas perforans. The production of enzymes associated with resistance induction (peroxidase, polyphenol oxidase, phenylalanine ammonia-lyase, β-1,3-glucanase, and protease) was quantified from leaf samples collected 24 hours before and 24 hours after chemical spraying and at 1, 2, 4, 6, and 8 days after bacterial inoculation. All products tested controlled bacterial spot, but only ASM, pyraclostrobin, and pyraclostrobin + metiram increased the production of peroxidase in the leaves of the two tomato cultivars, and increased the production of polyphenol oxidase and β-1,3-glucanase in the Santa Clara cultivar.

Effect of fungicide on Fusarium verticillioides mycelial morphology and fumonisin B1 production

The effect of fludioxonil + metalaxyl-M on the mycelial morphology, sporulation and fumonisin B1 production by Fusarium verticillioides 103 F was evaluated. Scanning electron microscopy analysis showed that the fungicide caused inhibition of hyphal growth and defects on hyphae morphology such as cell wall disruption, withered hyphae, and excessive septation. In addition, extracellular material around the hyphae was rarely observed in the presence of fludioxonil + metalaxyl-M. While promoting the reduction of mycelial growth, the fungicide increased sporulation of F. verticillioides compared to the control, and the highest production occurred on the 14th day in the treatments and on the 10th day in the control cultures. Fumonisin B1 production in the culture media containing the fungicide (treatment) was detected from the 7th day incubation, whereas in cultures without fungicide (control) it was detected on the 10th day. The highest fumonisin B1 production occurred on the 14th day, both for the control and for the treatment. Fludioxonil + metalaxyl - M can interfere in F. verticillioides mycelial morphology and sporulation and increase fumonisin B1 levels. These data indicate the importance of understanding the effects of fungicide to minimize the occurrence of toxigenic fungi and fumonisins.

Potential of Microbispora sp. V2 as biocontrol agent against Sclerotium rolfsii, the causative agent of southern blight of Zea mays L (Baby corn)–in vitro studies.

The study was undertaken with the aim of exploring novel and beneficial agro activities of rare actinomycetes like Microbispora sp. V2. The antagonistic activity of Microbispora sp. V2 was evaluated as a biocontrol agents against Sclerotium rolfsii, a soil-borne fungal plant pathogen. The methodology performed for evaluation of biocontrol agent was in vitro evaluation assay which comprised of three tests viz., cellophane overlay technique, seed germination test and Thiram (fungicide) tolerance of Microbispora sp. V2. The isolate was found to inhibit the fungal pathogen Sclerotium rolfsii to 91.43% in cellophane assay. In seed germination assay, Microbispora sp. V2 treated seeds resulted in 25.75% increased germination efficiency, as compared to seeds infected by Sclerotium rolfsii. The isolate Microbispora sp. V2 could tolerate 1000 µg mL-1 of Thiram (fungicide). The in vitro assay studies proved that Microbispora sp. V2 can be used as antifungal antagonist and thus posses’ great potential as biocontrol agent against southern blight caused by Sclerotium rolfsii in Zea mays L (Baby corn) which causes large economical losses.

Chemical management in fungicide sensivity of Mycosphaerella fijiensis collected from banana fields in México

The chemical management of the black leaf streak disease in banana caused by Mycosphaerella fijiensis (Morelet) requires numerous applications of fungicides per year. However this has led to fungicide resistance in the field. The present study evaluated the activities of six fungicides against the mycelial growth by determination of EC50 values of strains collected from fields with different fungicide management programs: Rustic management (RM) without applications and Intensive management (IM) more than 25 fungicide application/year. Results showed a decreased sensitivity to all fungicides in isolates collected from IM. Means of EC50 values in mg L-1 for RM and IM were: 13.25 ± 18.24 and 51.58 ± 46.14 for azoxystrobin, 81.40 ± 56.50 and 1.8575 ± 2.11 for carbendazim, 1.225 ± 0.945 and 10.01 ± 8.55 for propiconazole, 220 ± 67.66 vs. 368 ± 62.76 for vinclozolin, 9.862 ± 3.24 and 54.5 ± 21.08 for fludioxonil, 49.2125 ± 34.11 and 112.25 ± 51.20 for mancozeb. A molecular analysis for β-tubulin revealed a mutation at codon 198 in these strains having an EC50 greater than 10 mg L-1 for carbendazim. Our data indicate a consistency between fungicide resistance and intensive chemical management in banana fields, however indicative values for resistance were also found in strains collected from rustic fields, suggesting that proximity among fields may be causing a fungus interchange, where rustic fields are breeding grounds for development of resistant strains. Urgent actions are required in order to avoid fungicide resistance in Mexican populations of M. fijiensis due to fungicide management practices.

Antagonistic potential of fluorescent pseudomonads and control of charcoal rot of chickpea caused by Macrophomina phaseolina.

The effectiveness of plant growth promoting rhizobacteria especially Pseudomonas fluorescens isolates were tested against charcoal rot of chickpea both in green house as well as in field conditions. Most of the isolates reduced charcoal rot disease and promoted plant growth in green house. A marked increase in shoot and root length was observed in P. fluorescens treated plants. Among all the P. fluorescens isolates Pf4-99, was found most effective in the improvement of chickpea crop in green house as well as in field. Pf4-99 effectively promoted plant growth and produced indole acetic acid in culture medium. This isolate also inhibited the mycelial growth of the M. phaseolina under in vitro conditions and reduced the disease severity Potential isolate (Pf4-99) also significantly increased the biomass of the chickpea plants, shoot length, root length and protein content of the chickpea seeds. A part from these, the total number of seeds per plant and their weight were also enhanced. The colonization of Pf4-99 reduced the incidence of seed mycoflora by which indirectly enhanced the seed germination and vigour index of seedlings. The observations revealed that isolate Pf4-99 is quite effective to reduce the charcoal rot disease both in field and greenhouse, and also increases seed yields significantly Therefore, this isolate appears to be an efficient biocontrol agent against charcoal rot disease as well as yield increasing rhizobacterium.

Transformation of the entomopathogenic fungi, paecilomyces fumosoroseus and paecilomyces lilacinus (deuteromycotina: hyphomycetes) to benomyl resistence

The entomopathogenic fungi Paecilomyces fumosoroseus and P. lilacinus have been transformed to resistance to the fungicide benomyl by a polyethylene glycol (PEG)-mediated procedure using a mutant b-tubulin gene from Neurospora crassa carried on plasmid pBT6. Benomyl-resistant transformants of P. lilacinus were obtained that could tolerate greater than 30 µg/ml benomyl and P. fumosoroseus transformants were obtained that could tolerate 20 µg/ml benomyl. Following 5 serial passages of transformants on benomyl-containing media and 5 serial passages on non-selective media, 100 per cent of P. lilacinus transformants were found to be mitotically stable by a conidial germination test. In contrast, only 4 out of 9 transformants of P. fumosoroseus were mitotically stable. Southern blot analysis of genomic DNA from both species suggested that the mechanism of transformation in all transformants was by gene replacement of the b-tubulin allele. Non-homologous vector sequences were not detectable in the genomes of transformants.

Estudo da influência de agrotóxicos sobre o desenvolvimento do Rhizobium leguminosarum biovar phaseoli / Study of the agrotoxics influence over the development of the Rhizobium leguminosarum biovar phaseoli

As bactérias fixadoras de nitrogênio atmosférico, via processo simbiótico, repassam N2 às leguminosas sob forma de amônia, proporcionando aumento na produçäo de alimentos protéicos, reciclagem biológica de nitrogênio do ar, evitando o alto custo da adubaçäo nitrogenada e o efeito potencialmente poluidor do nitrato lixiviado. Testou-se quatro estirpes de Rhizobium phaseoli frente a quatro fungicidas indicados para o tratamento de sementes de Phaseolus vulgaris L. (feijoeiro), no Estado do Rio Grande o sul (Brasil). Considerando-se as dosagens recomendadas dos fungicidas, houve crescimento bacteriano das quatro estirpes frente a Benomil e inibiçäo total frente a Captan; porém, para PCNB e Thiram, a resistência dependeu do tipo de estirpe. Observou-se que näo há influência de fungicidas sobre o desenvolvimento do Rhizobium phaseoli quando seleciona-se a estirpe adequada ao fungicida corretamente dosado.

Effect of some natural pesticides on entomogenous muscardine fungi.

Five commercial preparations of natural pesticides were tested for in vitro compatibility with muscardine fungi, Beauveria brongniartii and Metarhizium anisopliae. Neemark (azadirachtin) was found compatible with both the fungi. Phytoallexin, the natural fungicide, significantly inhibited the growth of both the fungi, while other natural pesticides showed moderate to severe inhibition.

Influence of mancozeb on mitogenically responsive lipids in rat cerebrum and liver.

Mancozeb, a commonly used fungicide, has been shown to induce tumours in mouse skin and maneb, unit constituent of mancozeb, is reported to induce tumours in rats. The mechanism by which mancozeb induced tumorigenicity is not known. Since the levels of inositol phospholipids and phosphatidic acid have roles in the regulation of cell proliferation, the effects of mancozeb on the levels of these lipids were studied in rats. Daily oral administration of commercial grade mancozeb at a concentration of 50 mg/kg body wt for 30 days (5 days a week) caused no significant change in the levels of inositol phospholipids and phosphatidic acid (PA) in both cerebrum and liver, while at high concentration (250 mg/kg body wt) under the same treatment schedule mancozeb increased the levels of these lipids. In cerebrum, the levels of phosphatidylinositol (PI) and PA were increased by 36 and 43% respectively without affecting the levels of polyphosphoinositides, whereas in liver the levels of not only PI (50%) and PA (49%) but also those of polyphosphoinositides were increased. These results show that mancozeb influences the levels of PA and inositol phospholipids, involved in phospholipase C-pathway of signalling.