RESUMO
Early blight caused by Alternaria solani is a common foliar disease of potato around the world, and serious infections result in reduced yields and marketability due to infected tubers. The major aim of this study is to figure out the synergistic effect between microorganism and fungicides and to evaluate the effectiveness of Bacillus subtilis NM4 in the control of early blight in potato. Based on its colonial morphology and a 16S rRNA analysis, a bacterial antagonist isolated from kimchi was identified as B. subtilis NM4 and it has strong antifungal and anti-oomycete activity against several phytopathogenic fungi and oomycetes. The culture filtrate of strain NM4 with the fungicide effectively suppressed the mycelial growth of A. solani, with the highest growth inhibition rate of 83.48%. Although exposure to culture filtrate prompted hyphal alterations in A. solani, including bulging, combining it with the fungicide caused more severe hyphal damage with continuous bulging. Surfactins and fengycins, two lipopeptide groups, were isolated and identified as the main compounds in two fractions using LC-ESI-MS. Although the surfactin-containing fraction failed to inhibit growth, the fengycin-containing fraction, alone and in combination with chlorothalonil, restricted mycelial development, producing severe hyphal deformations with formation of chlamydospores. A pot experiment combining strain NM4, applied as a broth culture, with fungicide, at half the recommended concentration, resulted in a significant reduction in potato early blight severity. Our results indicate the feasibility of an integrated approach for the management of early blight in potato that can reduce fungicide application rates, promoting a healthy ecosystem in agriculture.
Assuntos
Alternaria , Bacillus subtilis , Fungicidas Industriais , Lipopeptídeos , Nitrilas , Doenças das Plantas , Solanum tuberosum , Solanum tuberosum/microbiologia , Doenças das Plantas/microbiologia , Doenças das Plantas/prevenção & controle , Alternaria/efeitos dos fármacos , Alternaria/crescimento & desenvolvimento , Bacillus subtilis/efeitos dos fármacos , Bacillus subtilis/crescimento & desenvolvimento , Fungicidas Industriais/farmacologia , Nitrilas/farmacologia , Lipopeptídeos/farmacologia , RNA Ribossômico 16S/genética , Hifas/efeitos dos fármacos , Hifas/crescimento & desenvolvimento , Micélio/efeitos dos fármacos , Micélio/crescimento & desenvolvimento , Peptídeos Cíclicos/farmacologiaRESUMO
AIMS: Microbial biocontrol agents have become an effective option to mitigate the harmfulness of chemical pesticides in recent years. This study demonstrates the control efficacy of Bacillus velezensis CE 100 on the anthracnose causal agent, Colletotrichum gloeosporioides. METHODS AND RESULTS: In vitro antifungal assays revealed that the culture filtrate and volatile organic compounds of B. velezensis CE 100 strongly restricted the mycelial development of C. gloeosporioides. Moreover, a bioactive compound, butyl succinate, was isolated from the n-butanol crude extract of B. velezensis CE 100 (bce), and identified by liquid chromatography-electrospray ionization hybrid ion-trap and time-of-flight mass spectrometry (LC-ESI-QTOF-MS) and one-dimensional (1D) and two-dimensional (2D) nuclear magnetic resonance (NMR). Treatment with purified butyl succinate at a concentration of 300 µg mL-1 strongly controlled conidial germination of C. gloeosporioides with an inhibition rate of 98.66%, whereas butyl succinate at a concentration of 400 µg mL-1 showed weak antifungal action on the mycelial growth of C. gloeosporioides with an inhibition rate of 31.25%. Scanning electron microscopy revealed that the morphologies of butyl succinate-treated hyphae and conidia of C. gloeosporioides were severely deformed with shriveled and wrinkled surfaces. Furthermore, butyl succinate was able to control carbendazim-resistant C. gloeosporioides, demonstrating that it could be a promising agent for the suppression of other carbendazim-resistant fungal pathogens. An in vivo biocontrol assay demonstrated that the strain ce 100 broth culture and butyl succinate showed higher control efficacy on apple anthracnose than bce. CONCLUSIONS: Our findings provide insight into the antifungal potential of B. velezensis ce 100 and its butyl succinate for efficient control of phytopathogenic fungi, such as C. gloeosporiodes, in plant disease protection. This is the first study to demonstrate the antifungal potential of bacteria-derived butyl succinate for control of C. gloeosporioides.
Assuntos
Colletotrichum , Malus , Antifúngicos/farmacologia , Antifúngicos/química , Ácido Succínico/farmacologia , Succinatos , Doenças das Plantas/prevenção & controle , Doenças das Plantas/microbiologiaRESUMO
The aim of this study was to determine the plant growth-promoting effect of Bacillus subtilis PE7 on growth of melon plants. B. subtilis PE7 isolated from kimchi was identified based on colonial and microscopic morphology along with analyses of 16S rRNA and pycA gene sequences. Strain PE7 showed different levels of inhibition on phytopathogens and was able to grow at variable temperatures and pH values. Strain PE7 had the ability to produce siderophores, indole-3-acetic acid (IAA), ammonia, exopolysaccharides, and 1-aminocyclopropane-1-carboxylic acid deaminase, as well as solubilize insoluble phosphate and zinc. The IAA secretion of strain PE7 showed a concentration-dependent pattern based on the concentration of l-tryptophan supplemented in the fertilizer-based culture medium. The LC-MS analysis indicates the presence of IAA in the culture filtrate of strain PE7. Treatment of the B. subtilis PE7 culture containing different metabolites, mainly IAA, significantly promoted melon growth in terms of higher growth parameters and greater plant nutrient contents compared to treatments with the culture without IAA, fertilizer, and water. The cells of B. subtilis PE7 attached to and firmly colonized the roots of the bacterized melon plants. Based on our results, B. subtilis PE7 can be utilized as a potential microbial fertilizer to substitute chemical fertilizers in sustainable agriculture.
RESUMO
Pectobacterium carotovorum is a problematic bacterial pathogen causing soft rot in different vegetable crops, resulting in yield losses during pre- and post-harvest periods. In this study, Bacillus velezensis CE 100 showed antibacterial activity against P. carotovorum. Co-inoculation experiment indicated that B. velezensis CE 100 reduced the proliferation rate of P. carotovorum at the early incubation period and that a long incubation time induced a loss of viability of the bacterial pathogen. Agar well diffusion assay revealed that the culture filtrate of strain CE 100 affected the growth of P. carotovorum in a dose-dependent pattern. In time-kill assay, inoculation of P. carotovorum with 50% culture filtrate of strain CE 100 resulted in a complete loss of survival at 4 h incubation period. An antibacterial compound isolated from chloroform extract of B. velezensis CE 100 was identified as macrolactin A based on results of 1H and 13C NMR and mass spectrometry. However, time-kill assay showed that purified macrolactin A at a concentration of 200 µgmL-1 was not highly effective to control the growth of P. carotovorum although reduction in cell number of P. carotovorum was observed. Moreover, in vivo assay revealed that B. velezensis CE 100 effectively controlled bacterial soft rot. As a consequence, it significantly improved cucumber growth. Therefore, B. velezensis CE 100 could be used as an eco-friendly bioagent for effective control of bacterial soft rot to minimize global economic losses in crop production.
Assuntos
Cucumis sativus , Pectobacterium , Pectobacterium carotovorum , Doenças das Plantas/prevenção & controle , Doenças das Plantas/microbiologia , Bactérias , Antibacterianos/farmacologiaRESUMO
Botrytis cinerea, the causal agent of gray mold is one of the major devastating fungal pathogens that occurs in strawberry cultivation and leads to massive losses. Due to the rapid emergence of resistant strains in recent years, an ecofriendly disease management strategy needs to be developed to control this aggressive pathogen. Bacillus velezensis CE 100 exhibited strong antagonistic activity with 53.05% against B. cinerea by dual culture method. In the present study, 50% of culture filtrate supplemented into PDA medium absolutely inhibited mycelial growth of B. cinerea whereas the highest concentration (960 mg/L) of different crude extracts including ethyl acetate, chloroform, and n-butanol crude extracts of B. velezensis CE 100, strongly inhibited mycelial growth of B. cinerea with the highest inhibition of 79.26%, 70.21% and 69.59% respectively, resulting in severe damage to hyphal structures with bulging and swellings. Hence, the antifungal compound responsible was progressively separated from ethyl acetate crude extract using medium pressure liquid chromatography. The purified compound was identified as methyl hippurate by nuclear magnetic resonance and mass spectrometry. The inhibitory effect of methyl hippurate on both spore germination and mycelial growth of B. cinerea was revealed by its dose-dependent pattern. The spore germination rate was completely restricted at a concentration of 3 mg/mL of methyl hippurate whereas no mycelial growth was observed in agar medium supplemented with 4 mg/mL and 6 mg/mL of methyl hippurate by poisoned food method. Microscopic imaging revealed that the morphologies of spores were severely altered by long-time exposure to methyl hippurate at concentrations of 1 mg/mL, 2 mg/mL and 3 mg/mL and hyphae of B. cinerea were severely deformed by exposure to methyl hippurate at concentrations of 2 mg/mL, 4 mg/mL and 6 mg/mL. No significant inhibition on tomato seed germination was observed in treatments with methyl hippurate (2 mg/mL) for both 6 h and 12 h soaking period as compared to the controls. Based on these results, B. velezensis CE 100 could be considered a potential agent for development of environmentally friendly disease control strategies as a consequence of the synergetic interactions of diverse crude metabolites and methyl hippurate.
Assuntos
Bacillus/química , Botrytis/efeitos dos fármacos , Fungicidas Industriais/farmacologia , Hipuratos/farmacologia , Bacillus/metabolismo , Botrytis/crescimento & desenvolvimento , Fungicidas Industriais/química , Fungicidas Industriais/isolamento & purificação , Fungicidas Industriais/metabolismo , Hipuratos/química , Hipuratos/isolamento & purificação , Hipuratos/metabolismo , Hifas/efeitos dos fármacos , Hifas/crescimento & desenvolvimento , Solanum lycopersicum/microbiologia , Doenças das Plantas/microbiologia , Esporos Fúngicos/efeitos dos fármacos , Esporos Fúngicos/crescimento & desenvolvimentoRESUMO
Composting is a microbiological process that converts organic waste into organic soil amendment. We reveal enhanced biodiversity and microbial population with subsequent enhancement of composting efficiency of lignocellulosic waste using thermoacidophilic effective microorganisms (tEM). Composting with tEM + shading (tEMA) or tEM without shading (tEMB) increased the average microbial population by 12.0% or 6.7%, respectively compared to non-tEM composting without shading/control (C). The biodiversity in tEMA or tEMB treated groups was increased by 34.7% or 43.7%, respectively, compared to C. The highest increase in population (31.7% and 9.4%) and diversity (91.2% and 91.6%) were observed in tEMA and tEMB at 30 d, respectively. Regarding microbial structure, the most dominant phylum shifted from Proteobacteria to Bacteroidetes during composting. From 60 to 120 d, tEM notably improved the average abundance of Firmicutes (mainly Bacillus) by 166.7% and 75.8% in tEMA and tEMB groups, respectively. The overall gradation rate of large compost granules (<2 mm) increased by 36.4% and 24.7%, following tEMA and tEMB treatment, respectively. The average rate of increase in bulk density was 42.6% or 33.3% by tEMA or tEMB, respectively, compared to C. We reveal the major differences in microbial structure, including a higher abundance of beneficial microbes like Bacillus in tEM treated composts. The study revealed that tEM could improve biodiversity and population of microbes, especially during thermophilic phase (above 45 °C), with a subsequent increase in composting rate, mineralization, and product quality. The results of this study are particularly invaluable in the areas of environmental conservation and organic agriculture.
Assuntos
Compostagem , Biodiversidade , Lignina , SoloRESUMO
Lysobacter antibioticus HS124 inhibited mycelial growth of Fusarium graminearum (74.66%) under the dual culture method. Microscopic investigation clearly showed that amendment with different concentrations (10%, 30%, and 50%) of HS124 bacterial culture filtrate on potato dextrose agar plates caused abnormal hyphal structures, including swelling and distortion. Its inhibition toward mycelial growth of F. graminearum was increased with increasing concentration of n-butanol crude extract of HS124. The highest inhibition (43.14%) was detected at a crude concentration of 10 mg/disc, whereas the lowest inhibition (21.57%) was observed at 2 mg/disc. Although mycelial growth of F. graminearum was promoted by volatile organic compounds (VOCs) produced by HS124 as compared with the control, these VOCs clearly decreased fungal pigmentation resulting in a reduction of fungal sporulation. Microscopic investigation revealed hyphal deformation of F. graminearum due to VOCs. These compounds also had a negative effect on spore germination of F. graminearum. In vivo evaluations demonstrated that HS124 inoculation of wheat plants reduced crown rot disease incidence by 73.70% as compared with the control. HS124 inoculation of wheat plants also promoted most of the growth characteristics compared with the control or fungicide-treated plants. Our results provide strong evidence that HS124 could control F. graminearum infections and promote growth of wheat plants as part of management strategies for crown rot disease.
Assuntos
Agentes de Controle Biológico/farmacologia , Fusarium/efeitos dos fármacos , Lysobacter/fisiologia , Doenças das Plantas/prevenção & controle , Triticum/crescimento & desenvolvimento , Triticum/microbiologia , Fusarium/crescimento & desenvolvimento , Fusarium/fisiologia , Hifas/efeitos dos fármacos , Hifas/crescimento & desenvolvimento , Hifas/fisiologia , Lysobacter/metabolismo , Doenças das Plantas/microbiologia , Esporos Fúngicos/efeitos dos fármacos , Esporos Fúngicos/crescimento & desenvolvimento , Esporos Fúngicos/fisiologia , Compostos Orgânicos Voláteis/metabolismo , Compostos Orgânicos Voláteis/farmacologiaRESUMO
The aim of the present study is to describe the purification and identification of methyl 2,3-dihydroxybenzoate (M2,3DB), isolated for the first time from Paenibacillus elgii HOA73, and to subsequently investigate its antifungal activity against important plant pathogens. The results show that M2,3DB can be purified by many different chromatographic techniques and is identified as methyl 2,3-dihydroxybenzoate based on nuclear magnetic resonance (NMR) and liquid chromatography-mass spectrometry (LC-MS) spectra analyses. M2,3DB was firstly evaluated for its antifungal activity, where the growth of Botrytis cinerea and Rhizoctonia solani was almost completely inhibited at an M2,3DB concentration of 50 µg/mL. Growth inhibition of Phytophthora capsici and Fusarium oxysporum f.sp lycopersici was found at the same M2,3DB concentration by 48.8% and 36.6%, respectively. Minimum inhibitory concentrations (MICs) of M2,3DB that inhibited any visible mycelial growth of B. cinerea, R. solani, and F. oxysporum f.sp lycopersici were defined as 32, 32, and 64 µg/mL, respectively. The broad antifungal activity of M2,3DB against various plant pathogens suggests its scope as a biofungicide in the management of plant disease.
Assuntos
Antifúngicos/isolamento & purificação , Antifúngicos/farmacologia , Hidroxibenzoatos/isolamento & purificação , Hidroxibenzoatos/farmacologia , Paenibacillus/química , Antifúngicos/administração & dosagem , Antifúngicos/química , Botrytis/efeitos dos fármacos , Botrytis/crescimento & desenvolvimento , Cromatografia Líquida/métodos , Fusarium/efeitos dos fármacos , Fusarium/crescimento & desenvolvimento , Hidroxibenzoatos/administração & dosagem , Hidroxibenzoatos/química , Espectroscopia de Ressonância Magnética/métodos , Espectrometria de Massas/métodos , Testes de Sensibilidade Microbiana , Phytophthora/efeitos dos fármacos , Phytophthora/crescimento & desenvolvimento , Doenças das Plantas/microbiologia , Plantas/microbiologia , Rhizoctonia/efeitos dos fármacos , Rhizoctonia/crescimento & desenvolvimentoRESUMO
In this study, the expression patterns of extracellular chitinase and ß-1,3-glucanase from cultured Wickerhamomyces anomalus EG2 treated with chitin, glucan, and chemical chitinase inhibitors (kinetin, caffeine, and acetazolamide) were investigated using SDS-PAGE. Relationship between enzyme expression and antifungal activity from yeast plays a very important role for biocontrol of phytopathoges. To determine antifungal activity against phytopathogens, W. anomalus EG2 was shown to strongly inhibit hyphal growth of Fusarium oxysporum KACC 40032 and Rhizoctonia solani KACC 40111. Slight chitinase activity was observed 12 h after incubation in both PDB and YPD medium without colloidal chitin. The molecular weight of chitinase was approximately 124 kDa ß-1,3-Glucanase isoenzyme (GN1 and GN2) was observed distinctly on SDS-PAGE gels when laminarin was used as a substrate. ß-1,3-Glucanase isoenzyme was not observed when using glucan-containing high polymer complex (GHPC) as a substrate. Production of chitinase from W. anomalus EG2 was inhibited slightly by acetazolamide. Abnormal and cluster-shaped cells of W. anomalus EG2 were observed in both PDB and YPD medium treated with colloidal chitin. These results indicated that W. anomalus EG2 could be applied commercially as a biological control agent of phytopathogens and as a bioinhibitor of yeast cell growth.
Assuntos
Antifúngicos/farmacologia , Candida/efeitos dos fármacos , Candida/enzimologia , Celulases/metabolismo , Quitina/metabolismo , Quitinases/metabolismo , Glucanos/metabolismo , Acetazolamida/antagonistas & inibidores , Agentes de Controle Biológico , Cafeína/antagonistas & inibidores , Candida/classificação , Candida/isolamento & purificação , Quitinases/química , Quitinases/efeitos dos fármacos , Fusarium/crescimento & desenvolvimento , Hifas/crescimento & desenvolvimento , Cinetina/antagonistas & inibidores , Peso Molecular , Filogenia , Patologia Vegetal , RNA Ribossômico 18S/genética , Rhizoctonia/crescimento & desenvolvimentoRESUMO
In this study, we isolated Bacillus licheniformis MH48 from rhizosphere soil and demonstrated that this strain shows significant antifungal activity against Rhizoctonia solani, Colletotrichum gloeosporioides, and Phytophthora capsici. Our results showed that a 50% concentration of bacterial cell-free culture filtrate of B. licheniformis MH48 shows strong activity against fungal pathogens. Benzoic acid produced by B. licheniformis MH48 was purified by various chromatographic techniques and identified by nuclear magnetic resonance and gas chromatography-mass spectrometry analysis. Benzoic acid displayed antifungal activity against R. solani and C. gloeosporides with minimum inhibitory concentration of 128 µg/mL against mycelial growth. Microscopic examination revealed that benzoic acid (50 µg/mL and 100 µg/mL) transformed C. gloeosporioides conidial morphology and inhibited conidial germination. In addition, benzoic acid (100 µg/mL and 200 µg/mL) degraded R. solani mycelia. Therefore, our results demonstrate that B. licheniformis MH48 strain shows potential for utility as a biological agent for the control of various fungal pathogens of plants.
Assuntos
Antifúngicos/farmacologia , Bacillus licheniformis/química , Ácido Benzoico/farmacologia , Fatores Biológicos/farmacologia , Colletotrichum/efeitos dos fármacos , Phytophthora/efeitos dos fármacos , Rhizoctonia/efeitos dos fármacos , Antifúngicos/isolamento & purificação , Bacillus licheniformis/isolamento & purificação , Ácido Benzoico/isolamento & purificação , Fatores Biológicos/isolamento & purificação , Cromatografia , Colletotrichum/crescimento & desenvolvimento , Cromatografia Gasosa-Espectrometria de Massas , Espectroscopia de Ressonância Magnética , Testes de Sensibilidade Microbiana , Phytophthora/crescimento & desenvolvimento , Rhizoctonia/crescimento & desenvolvimento , Microbiologia do Solo , Esporos Fúngicos/efeitos dos fármacos , Esporos Fúngicos/crescimento & desenvolvimentoRESUMO
Rhizoctonia solani is the cause of substantial economic loss in many crops. The aim of this study is to investigate biocontrol potential of Bacillus sp. L60 against R. solani and to purify an antifungal compound. In this study, Bacillus sp. L60 demonstrated significant antagonism toward R. solani with the dual culture assay. The antifungal compound was extracted from Bacillus sp. L60 culture supernatant with n-butanol, and identified as N-butyl-tetrahydro-5-oxofuran-2-carboxamide (BT-5O-2C) having molecular weights of 185.1052 Da with the formula C9 H15 NO3 using NMR and HR-ESI-MS analysis. The minimum inhibitory concentration (MIC) value of the antifungal compound was 256 µg ml-1 against R. solani. Therefore, our results clearly demonstrated BT-5O-2C as well as Bacillus sp. L60 as potential biological control agents for the management of R. solani.
Assuntos
4-Butirolactona/análogos & derivados , Antibiose , Antifúngicos/isolamento & purificação , Bacillus/metabolismo , Rhizoctonia/fisiologia , 1-Butanol/farmacologia , 4-Butirolactona/isolamento & purificação , 4-Butirolactona/farmacologia , Antifúngicos/farmacologia , Bacillus/genética , Bacillus/isolamento & purificação , Meios de Cultura/química , Espectroscopia de Ressonância Magnética , Testes de Sensibilidade Microbiana , República da Coreia , Rhizoctonia/efeitos dos fármacos , Microbiologia do SoloRESUMO
The aim of the current study was to describe the role and mechanism of Bacillus amyloliquefaciens Y1 against the root-knot nematode, Meloidogyne incognita, under in vitro and in vivo conditions. Initially, the exposure of the bacterial culture supernatant and crude extract of Y1 to M. incognita significantly inhibited the hatching of eggs and caused the mortality of second-stage juveniles (J2), with these inhibitory effects depending on the length of incubation time and concentration of the treatment. The dipeptide cyclo(d-Pro-l-Leu) was identified in B. amyloliquefaciens culture for the first time using chromatographic techniques and nuclear magnetic resonance (NMR ¹H, 13C, H-H COSY, HSQC, and HMBC) and recognized to have nematocidal activity. Various concentrations of cyclo(d-Pro-l-Leu) were investigated for their effect on the hatching of eggs and J2 mortality. Moreover, the in vivo nematocidal activity of the Y1 strain was investigated by conducting pot experiments in which tomato plants were inoculated with M. incognita. Each and every pot was amended 50 mL of fertilizer media (F), or Y1 culture, or nematicide (N) (only once), or fertilizer media with N (FN) at 1, 2, 3, 4 and 5 weeks after transplantation. The results of the pot experiments demonstrated the antagonistic effect of B. amyloliquefaciens Y1 against M. incognita as it significantly decreases the count of eggs and galls per root of the tomato plant as well as the population of J2 in the soil. Besides, the investigation into the growth parameters, such as the length of shoot, shoot fresh and dry weights of the tomato plants, showed that they were significantly higher in the Y1 strain Y1-treated plants compared to F-, FN- and N-treated plants. Therefore, the biocontrol repertoire of this bacterium opens a new insight into the applications in crop pest control.
Assuntos
Antinematódeos/farmacologia , Bacillus amyloliquefaciens/metabolismo , Peptídeos Cíclicos/biossíntese , Peptídeos Cíclicos/farmacologia , Tylenchoidea/efeitos dos fármacos , Animais , Antinematódeos/isolamento & purificação , Cromatografia Líquida de Alta Pressão , Espectroscopia de Ressonância Magnética , Peptídeos Cíclicos/isolamento & purificaçãoRESUMO
To investigate the expression patterns of chitinase on SDS-PAGE gel, Paenibacillus ehimensis MA2012 was incubated in gelatin-chitin medium (GCM) at 30 °C for 7 days. Six major bands (Ch3, Ch4, Ch5, Ch6, Ch7, and Ch8) of chitinase isozymes in GC medium appeared on SDS-PAGE gel during the incubation period. Chitinase activity staining of P. ehimensis MA2012 was detected on 2-DE with different pI values (4-11). After DEAE-Sephadex chromatography, eight bands (Ch1 to Ch8) of chitinase isozymes were stained strongly with Calcofluor white M2R at fraction 45. After Sephadex G-75 gel filtration, six bands (Ch3 to Ch8) of chitinase isozymes were stained with Calcofluor white M2R at fractions of 11-12. The specific activity of the purified chitinase was 3.8 units mg(-1) protein with a purification factor of 0.27. Inhibition rate of the conidial germination of Colletotrichum gloeosporioides was 87% in partial purified chitinase treatment compared with control.
Assuntos
Antifúngicos/metabolismo , Quitinases/metabolismo , Colletotrichum/efeitos dos fármacos , Paenibacillus/enzimologia , Esporos Fúngicos/efeitos dos fármacos , Antifúngicos/isolamento & purificação , Quitinases/isolamento & purificação , Cromatografia em Gel , Cromatografia por Troca Iônica , Meios de Cultura/química , Eletroforese em Gel Bidimensional , Eletroforese em Gel de Poliacrilamida , Perfilação da Expressão Gênica , Isoenzimas/isolamento & purificação , Isoenzimas/metabolismo , Paenibacillus/crescimento & desenvolvimento , TemperaturaRESUMO
In this study, evidences for antagonism were established by production of antifungal metabolites from Streptomyces griseus H7602, which were active to inhibit mycelial growth of Phytophthora capsici in the in vitro assays. Mycelial growth and zoosporangia formation of P. capsici was strongly inhibited in the medium containing the cell free culture filtrate of S. griseus H7602. Antifungal metabolites from the cell free culture filtrate of S. griseus H7602 showed substantial antagonistic effects on P. capsici. In addition, a purified antifungal compound was separated from the antifungal metabolites of S. griseus H7602 and identified to be 1H-pyrrole-2-carboxylic acid (PCA) by spectra analyses. PCA showed strong antifungal activity and was evaluated for the first time for its antagonism against P. capsici under in vitro conditions. Minimum inhibitory concentration (MIC) value of PCA was low (4 µg ml(-1)), and the mycelial growth of P. capsici was almost inhibited at concentration of 64 µg ml(-1). This study suggests that the PCA may be useful as biofungicides against P. capsici, and the prominent antagonism of antifungal metabolites from S. griseus H7602 highlights it as a candidate for biocontrol of P. capsici.
Assuntos
Antibiose , Antifúngicos/isolamento & purificação , Antifúngicos/farmacologia , Phytophthora/efeitos dos fármacos , Phytophthora/crescimento & desenvolvimento , Prolina/análogos & derivados , Streptomyces griseus/metabolismo , Antifúngicos/metabolismo , Cromatografia Líquida de Alta Pressão , Espectroscopia de Ressonância Magnética , Espectrometria de Massas , Testes de Sensibilidade Microbiana , Micélio/efeitos dos fármacos , Micélio/crescimento & desenvolvimento , Prolina/isolamento & purificação , Prolina/metabolismo , Prolina/farmacologia , Microbiologia do SoloRESUMO
This study was undertaken to describe purification, identification, and characteristics of protocatechuic acid (PCA) isolated for the first time from Paenibacillus elgii HOA73 against Botrytis cinerea (the cause of gray mold disease on strawberry fruit). PCA was purified by different chromatographic techniques and identified as PCA (3,4-dihydroxybenzoic acid) by nuclear magnetic resonance and liquid chromatography-mass spectrometry analyses. PCA displayed potent antifungal activity against B. cinerea and Rhizoctonia solani. However, the antifungal activities were not sufficient to inhibit mycelial growth of Phytophthora capsici and Fusarium oxysporum. The minimum inhibitory concentration of PCA to inhibit any visible mycelial growth of both B. cinerea and R. solani was 64 µg ml(-1) . Most B. cinerea conidia displayed altered shape and absence of germination, or were degraded after treatment with 50 and 100 µg ml(-1) PCA, respectively. Moreover, gray mold formation on strawberry fruit was almost or completely inhibited by these PCA concentrations 7 days following infection with B. cinerea conidia, respectively. PCA may be a promising alternative to chemical fungicides as a potential biofungicide to prevent growth of B. cinerea in strawberry fruit disease management.
Assuntos
Antifúngicos/isolamento & purificação , Antifúngicos/farmacologia , Botrytis/efeitos dos fármacos , Fragaria/microbiologia , Hidroxibenzoatos/isolamento & purificação , Hidroxibenzoatos/farmacologia , Paenibacillus/química , Botrytis/crescimento & desenvolvimento , Cromatografia Líquida , Fusarium/efeitos dos fármacos , Fusarium/crescimento & desenvolvimento , Espectroscopia de Ressonância Magnética , Espectrometria de Massas , Testes de Sensibilidade Microbiana , Micélio/efeitos dos fármacos , Micélio/crescimento & desenvolvimento , Phytophthora/efeitos dos fármacos , Phytophthora/crescimento & desenvolvimento , Doenças das Plantas/microbiologia , Doenças das Plantas/prevenção & controle , Rhizoctonia/efeitos dos fármacos , Rhizoctonia/crescimento & desenvolvimento , Esporos Fúngicos/citologia , Esporos Fúngicos/efeitos dos fármacosRESUMO
In this study, a novel lipopeptide antibiotic was isolated from the culture supernatant of Paenibacillus ehimensis strain MA2012. After analyses by mass spectrometry (MS), nuclear magnetic resonance (NMR), and high resolution mass spectrometry (HR-MS/MS) the compound was identified to be polypeptin C consisting of 3-hydroxy-4-methyl-hexanoic acid moiety and nine amino acids as peptide body. It has the same molecular mass (1115 Da) with that of polypeptin A and B but the amino acid positions differ. A relatively low concentration (125 ppm) of polypeptin C lowered the surface tension of water from 72.2 to 36.4 mN/m. It showed antimicrobial activity against several plant pathogenic bacteria and fungi. When the polypeptin C was applied to the ripe pepper fruits previously inoculated with conidia of Colletotrichum gloeosporioides, the hyphal growth on the fruit was significantly suppressed. Moreover, the hyphal morphology of C. gloeosporioides was greatly affected by the purified compound. All these data suggest the great potential of P. ehimensis MA2012 to control plant fungal and bacterial diseases.
Assuntos
Anti-Infecciosos/isolamento & purificação , Anti-Infecciosos/farmacologia , Paenibacillus/metabolismo , Polimixinas/isolamento & purificação , Polimixinas/farmacologia , Anti-Infecciosos/química , Bactérias/efeitos dos fármacos , Agentes de Controle Biológico , Colletotrichum/efeitos dos fármacos , DNA Ribossômico/química , Fungos/efeitos dos fármacos , Hifas/efeitos dos fármacos , Hifas/crescimento & desenvolvimento , Testes de Sensibilidade Microbiana , Peso Molecular , Polimixinas/biossíntese , Polimixinas/química , RNA Ribossômico 16S/metabolismo , Tensão Superficial/efeitos dos fármacos , Espectrometria de Massas em Tandem , ÁguaRESUMO
This study was conducted to investigate biocontrol potential of Paenibacillus ehimensis KWN38 against Fusarium oxysporum f.sp. lycopersici causing Fusarium wilt disease in tomato. Our result showed that P. ehimensis KWN38 produced extracellular organic compounds and crude enzyme to inhibit F. oxysporum f.sp. lycopersici conidial germination in in vitro assays. Tomato seedlings were treated with water (W), grass medium (G), G with P. ehimensis KWN38 inoculation (GP) and G along with synthetic fungicide (GSf). Disease symptoms were was first observed in G and W at 12 days after infection (DAI) while symptoms were noticeable in the GP and GSf treatments at 20 and 24 DAI, respectively. Tomato plants treated with P. ehimensis KWN38 or fungicide significantly reduced Fusarium wilt disease incidence and severity as compared to control tomato plants treated with water and grass medium. The similar results were also found in the root mortality of tomato plants. At 25 DAI, most plants in control treatments (W and G) wilted and the brown vascular systems of infected plants was clearly differentiable from normal green vascular system of healthy plants from GP and GSf. Plants in the GP showed higher fresh and dry weights of both root and shoots than those in W and G treatments. Leaf peroxidase and polyphenol oxidase activities of tomato plants in G and W were higher than those in GP and GSf. Root enzyme activities showed a similar pattern but the values were higher than leaf enzyme. The results clearly demonstrated that P. ehimensis KWN38 may be considered as biocontrol agent of Fusarium wilt disease in tomato.
Assuntos
Antibiose , Fusarium/crescimento & desenvolvimento , Paenibacillus/fisiologia , Controle Biológico de Vetores/métodos , Doenças das Plantas/prevenção & controle , Solanum lycopersicum/fisiologia , Antifúngicos/metabolismo , Antifúngicos/farmacologia , Biomassa , Solanum lycopersicum/crescimento & desenvolvimento , Solanum lycopersicum/microbiologia , Paenibacillus/crescimento & desenvolvimento , Paenibacillus/metabolismo , Raízes de Plantas/crescimento & desenvolvimento , Raízes de Plantas/microbiologia , Raízes de Plantas/fisiologiaRESUMO
The aim of this study was to characterize the roles of sulphur (S) nutrition in modulating the responses to iron (Fe) deficiency in the photosynthetic organelles of oilseed rape. Eight-week-old plants grown hydroponically were fed with S-sufficient or S-deprived solution with or without Fe(III)-EDTA. Responses to four S and Fe combined treatments were analysed after 5 and 10 days. Leaf chlorosis was generated by either S- or Fe-deprivation, with a decrease in chlorophyll and carotenoid content. These negative effects were more severe in the absence of S. The expression of Fe²âº transporter (IRT1) and Fe(III) chelate reductase (FRO1) gene was induced for the first 5 days and decreased after 10 days in the S-deprived roots, but largely improved by S supply even in the absence of Fe. Lack of ferric chelate reducing activity in the Fe-deprived roots in the absence of S was largely improved by S supply. The activity of photosynthesis, RuBisCO and sucrose synthase was closely related to S status in leaves. Electron microscopic observation showed that the Fe-deficiency in the absence of S greatly resulted in a severe disorganisation of thylakoid lamellae with loss of grana. However, these impacts of Fe-deficiency were largely restored in the presence of S. The present results indicate that S nutrition has significant role in ameliorating the damages in photosynthetic apparatus caused by Fe-deficiency.
Assuntos
Brassica napus/metabolismo , Ferro/metabolismo , Fotossíntese , Enxofre/metabolismo , Brassica napus/efeitos dos fármacos , Brassica napus/fisiologia , Carotenoides/metabolismo , Proteínas de Transporte de Cátions/genética , Clorofila/metabolismo , Cloroplastos/metabolismo , Cloroplastos/ultraestrutura , FMN Redutase/genética , Regulação da Expressão Gênica de Plantas , Glucosiltransferases/metabolismo , Ferro/farmacologia , Organelas/metabolismo , Folhas de Planta/metabolismo , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Raízes de Plantas/fisiologia , Estômatos de Plantas/metabolismo , Ribulose-Bifosfato Carboxilase/metabolismo , Enxofre/farmacocinética , Enxofre/farmacologiaRESUMO
The antifungal activities of methanolic extracts from Terminalia nigrovenulosa bark (TNB) was investigated for effects on the initial growth of mycelia against Fusarium solani. The ethyl acetate fraction separated from TNB demonstrated the highest antifungal activity against F. solani. The antifungal compound was isolated from TNB using silica gel column and Sephadex LH-20 chromatography combined with thin-layer chromatography and high performance liquid chromatography. Structural identification of the antifungal compound was conducted using (1)H NMR, (13)C NMR, and liquid chromatography-tandem mass spectrometry. The purified antifungal compound was gallic acid (GA) or 3,4,5-trihydroxy benzoic acid. Purified-GA possesses the high antifungal activity against F. solani, and that antifungal activity was dosage-dependent. The hyphae became collapsed and shrunken after 24 h incubation with GA (500 ppm). In pot experiments, the application of TNB crude extract was found to be effective in controlling the cucumber Fusarium root rot disease by enhancing activities of chitinase, peroxidase thereby promoting the growth of plants. The applied TNB extract significantly suppressed root rot disease compared to control. It resulted in 33, 75 and 81% disease suppression with 100, 500 and 1000 ppm of TNB crude extract, respectively. The study effectively demonstrated biological activities of the TNB extract, therefore suggesting the application of TNB for the control of soil-borne diseases of cucumber plants.
Assuntos
Antifúngicos/farmacologia , Fusarium/efeitos dos fármacos , Ácido Gálico/farmacologia , Terminalia/química , Antifúngicos/isolamento & purificação , Cromatografia , Cucumis sativus/microbiologia , Relação Dose-Resposta a Droga , Fusarium/crescimento & desenvolvimento , Ácido Gálico/isolamento & purificação , Espectroscopia de Ressonância Magnética , Micélio/efeitos dos fármacos , Micélio/crescimento & desenvolvimento , Casca de Planta/química , Doenças das Plantas/microbiologia , Raízes de Plantas/microbiologia , Espectrometria de Massas em TandemRESUMO
In this study, the 3,4-dihydroxybenzoic acid (3,4-DHBA) from Terminalia nigrovenulosa bark (TNB) was purified and its in vitro nematicidal activity was investigated against Meloidogyne incognita. The purification of 3,4-DHBA used a silica gel column and Sephadex LH-20 chromatography combined with thin-layer chromatography and high performance liquid chromatography. Structural identification of the 3,4-DHBA was conducted using (1)H nuclear magnetic resonance (NMR), (13)C NMR, and liquid chromatography time-of-flight mass spectrometry. Nematicidal activity bioassays revealed that 3,4-DHBA treatment resulted in 33.3, 47.5, 72.5 and 94.2% J2 mortality at 0.125, 0.25, 0.5 and 1.0 mg/ml, respectively after 12 h incubation. J2 mortality was increased significantly (P < 0.0001) with increasing incubation time in the range of 54.2-94.2% from 3 to 9 h after incubation with 3,4-DHBA (1.0 mg/ml), but with no significant difference observed where the incubation time was increased from 9 to 12 h. The 3,4-DHBA treatment resulted in 33.3, 65.0, 76.7 and 85.0% hatch inhibition at 0.125, 0.25, 0.5 and 1.0 mg/ml, respectively, 3 days after incubation. Changes in the shape of the eggs were determined after incubation for 1 day with a 3,4-DHBA concentration of 1.0 mg/ml.