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1.
Int J Mol Sci ; 22(15)2021 Jul 27.
Artigo em Inglês | MEDLINE | ID: mdl-34360756

RESUMO

This study focuses on a commercial plant elicitor based on chitooligosaccharides (BIG®), which aids in rice plant growth and disease resistance to bacterial leaf blight (BLB). When the pathogen (Xoo) vigorously attacks rice that has suffered yield losses, it can cause damage in up to 20% of the plant. Furthermore, Xoo is a seed-borne pathogen that can survive in rice seeds for an extended period. In this study, when rice seeds were soaked and sprayed with BIG®, there was a significant increase in shoot and root length, as well as plant biomass. Furthermore, BIG®-treated rice plants showed a significant reduction in BLB severity of more than 33%. Synchrotron radiation-based Fourier transform infrared (SR-FTIR) analysis was used to characterize BIG®'s mechanism in the chemical structure of rice leaves. The SR-FTIR results at 1650, 1735, and 1114 cm-1 indicated changes in biochemical components such as pectins, lignins, proteins, and celluloses. These findings demonstrated that commercial BIG® not only increased rice growth but also induced resistance to BLB. The drug's target enzyme, Xoo 1075 from Xanthomonas oryzae (PDB ID: 5CY8), was analyzed for its interactions with polymer ingredients, specifically chitooligosaccharides, to gain molecular insights down to the atomic level. The results are intriguing, with a strong binding of the chitooligosaccharide polymer with the drug target, revealing 10 hydrogen bonds between the protein and polymer. Overall, the computational analysis supported the experimentally demonstrated strong binding of chitooligosaccharides to the drug target.


Assuntos
Quitina/análogos & derivados , Resistência à Doença/efeitos dos fármacos , Oryza/microbiologia , Doenças das Plantas/microbiologia , Xanthomonas/crescimento & desenvolvimento , Quitina/química , Quitina/farmacologia
2.
Artigo em Inglês | MEDLINE | ID: mdl-34232119

RESUMO

A novel Gram-stain-negative, aerobic, asporogenous, catalase-positive and oxidase-negative, non-motile, golden-yellow pigmented, rod-shaped bacterium with casein-degrading ability, designated strain GCR10T, was isolated from roots of rice plants collected from a paddy field near Dongguk University, Republic of Korea. The results of subsequent 16S rRNA gene sequence analysis indicated that GCR10T shares the highest sequence identity with Chryseobacterium piscicola VQ-6316sT (98.3%). Strain GCR10T grew at 2-32 °C (optimum, 25 °C), at pH 6.0-8.0 (optimum, pH 7.0) and in the presence of 0-2.0% (w/v) NaCl (optimum in the absence of NaCl). The novel strain was able to produce carotenoid and flexirubin-type pigments. The predominant menaquinone was MK-6 and the major fatty acids were identified as iso-C15 : 0, iso-C17 : 0 3-OH and iso-C17 : 1ω9c. The polar lipids were phosphatidylethanolamine, four unidentified aminoglycolipids, two unidentified aminolipids and two unidentified glycolipids. The genome of GCR10T is 4.3 Mb in length with a DNA G+C content of 36.5 mol%. Average nucleotide identity, digital DNA-DNA hybridization and average amino acid identity values between GCR10T and Chryseobacterium piscicola VQ-6316sT were 82.1, 25.2 and 84.3 %, respectively, which clearly indicates that the novel strain is distinct from its closest relative. The demand for natural biodegradable pigments isolated frominsects, plants or microorganisms is increasing day by day because of their beneficial pharmacological properties. Here, we describe a novel strain that produces two types of pigment, carotenoid and flexirubin. On the basis of the results from phenotypic, genotypic and chemotaxonomic analyses, strain GCR10T represents a novel species of the genus Chryseobacterium, and the name Chryseobacterium caseinilyticum sp. nov. is proposed. The type strain is GCR10T (=KACC 21707T=NBRC 114715T).


Assuntos
Chryseobacterium/classificação , Oryza/microbiologia , Filogenia , Técnicas de Tipagem Bacteriana , Composição de Bases , Caseínas/metabolismo , Chryseobacterium/isolamento & purificação , DNA Bacteriano/genética , Ácidos Graxos/química , Glicolipídeos/química , Hibridização de Ácido Nucleico , Fosfatidiletanolaminas , Pigmentação , Raízes de Plantas/microbiologia , RNA Ribossômico 16S/genética , República da Coreia , Análise de Sequência de DNA , Vitamina K 2/análogos & derivados , Vitamina K 2/química
3.
Molecules ; 26(13)2021 Jun 25.
Artigo em Inglês | MEDLINE | ID: mdl-34202405

RESUMO

The bacterial leaf blight (BLB) caused by Xanthomonas oryzae pv. oryzae (Xoo) is one of the most serious rice diseases, causing huge yield losses worldwide. Several technologies and approaches have been opted to reduce the damage; however, these have had limited success. Recently, scientists have been focusing their efforts on developing efficient and environmentally friendly nanobactericides for controlling bacterial diseases in rice fields. In the present study, a scanning electron microscope (SEM), transmission electron microscope (TEM), and a confocal laser scanning microscope (CLSM) were utilized to investigate the mode of actions of ginger EOs on the cell structure of Xoo. The ginger EOs caused the cells to grow abnormally, resulting in an irregular form with hollow layers, whereas the dimethylsulfoxide (DMSO) treatment showed a typical rod shape for the Xoo cell. Ginger EOs restricted the growth and production of biofilms by reducing the number of biofilms generated as indicated by CLSM. Due to the instability, poor solubility, and durability of ginger EOs, a nanoemulsions approach was used, and a glasshouse trial was performed to assess their efficacy on BLB disease control. The in vitro antibacterial activity of the developed nanobactericides was promising at different concentration (50-125 µL/mL) tested. The efficacy was concentration-dependent. There was significant antibacterial activity recorded at higher concentrations. A glasshouse trial revealed that developed nanobactericides managed to suppress BLB disease severity effectively. Treatment at a concentration of 125 µL/mL was the best based on the suppression of disease severity index, AUDPC value, disease reduction (DR), and protection index (PI). Furthermore, findings on plant growth, physiological features, and yield parameters were significantly enhanced compared to the positive control treatment. In conclusion, the results indicated that ginger essential oils loaded-nanoemulsions are a promising alternative to synthetic antibiotics in suppressing Xoo growth, regulating the BLB disease, and enhancing rice yield under a glasshouse trial.


Assuntos
Gengibre/química , Óleos Voláteis , Oryza , Doenças das Plantas/microbiologia , Xanthomonas/crescimento & desenvolvimento , Óleos Voláteis/química , Óleos Voláteis/farmacologia , Oryza/química , Oryza/crescimento & desenvolvimento , Oryza/microbiologia , Oryza/ultraestrutura , Xanthomonas/ultraestrutura
4.
Artigo em Inglês | MEDLINE | ID: mdl-34319223

RESUMO

Two Gram-stain-negative, non-motile, rod-shaped bacterial strains were isolated from the surfaces of rice roots. They were designated as strains 1303T and 1310. Their colonies were circular, entire, opaque, convex and yellow. They were chitinase- and catalase-positive, reduced nitrate and grew at 16-37 °C (optimum, 30 °C), pH 5.0-10.0 (optimum, pH 7.0) and 0-2.0% NaCl (optimum, 1.0 %). Based on the 16S rRNA gene sequence analysis, they were classified as members of the genus Chitinophaga. Results of phylogenetic and phylogenomic analyses indicated that they formed a cluster with Chitinophaga eiseniae YC6729T, Chitinophaga qingshengii JN246T, Chitinophaga varians 10-7 W-9003T and Chitinophaga fulva G-6-1-13T. When the genomic sequences of strains 1303T and 1310 were compared with their close relatives, the average nucleotide identity and digital DNA-DNA hybridization values were below the cut-off levels. Phosphatidylethanolamine was the major polar lipid. MK-7 was the major respiratory quinone. iso-C15 : 0, C16 : 1 ω5c, iso-C17 : 0 3-OH and summed feature 3 (C16 : 1 ω7c/C16 : 1 ω6c) were the predominant fatty acids. Differential characteristics between both strains and their close relatives were also observed. Based on the distinctions in genotypic, phenotypic and chemotypic features, strains 1303T and 1310 represent members of a novel species of the genus Chitinophaga, for which the name Chitinophaga oryzae sp. nov. is proposed. The type strain is 1303T (=KACC 22075T=TBRC 12926T).


Assuntos
Bacteroidetes/classificação , Oryza/microbiologia , Filogenia , Raízes de Plantas/microbiologia , Microbiologia do Solo , Técnicas de Tipagem Bacteriana , Bacteroidetes/isolamento & purificação , Composição de Bases , DNA Bacteriano/genética , Ácidos Graxos/química , Hibridização de Ácido Nucleico , Fosfatidiletanolaminas/química , Pigmentação , RNA Ribossômico 16S/genética , Análise de Sequência de DNA , Tailândia , Vitamina K 2/análogos & derivados , Vitamina K 2/química
5.
Int J Mol Sci ; 22(11)2021 Jun 07.
Artigo em Inglês | MEDLINE | ID: mdl-34200345

RESUMO

Efficient accumulation of flavonoids is important for increased tolerance to biotic stress. Although several plant defense mechanisms are known, the roles of many pathways, proteins, and secondary metabolites in stress tolerance are unknown. We generated a flavanone 3-hydroxylase (F3H) overexpressor rice line and inoculated Xanthomonas Oryzae pv. oryzae and compared the control and wildtype inoculated plants. In addition to promoting plant growth and developmental maintenance, the overexpression of F3H increased the accumulation of flavonoids and increased tolerance to bacterial leaf blight (BLB) stress. Moreover, leaf lesion length was higher in the infected wildtype plants compared with infected transgenics. Kaempferol and quercetin, which scavenge reactive oxygen species, overaccumulated in transgenic lines compared with wildtypes in response to pathogenic infection, detected by scanning electron microscopy and spectrophotometry. The induction of F3H altered the antioxidant system and reduced the levels of glutathione peroxidase activity and malondialdehyde (MDA) contents in the transgenic lines compared with the wildtypes. Downstream gene regulation analysis showed that the expression of F3H increased the regulation of flavonol synthase (FLS), dihydroflavonol 4-reductase (DFR), and slender rice mutant (SLR1) during BLB stress. The analysis of SA and JA signaling revealed an antagonistic interaction between both hormones and that F3H induction significantly promoted SA and inhibited JA accumulation in the transgenic lines. SA-dependent nonexpressor pathogenesis-related (NPR1) and Xa1 showed significant upregulation in the infected transgenic lines compared with the infected control and wildtype lines. Thus, the overexpression of F3H was essential for increasing BLB stress tolerance.


Assuntos
Antioxidantes/metabolismo , Resistência à Doença/imunologia , Flavonoides/metabolismo , Hormônios/metabolismo , Oxigenases de Função Mista/metabolismo , Oryza/imunologia , Doenças das Plantas/imunologia , Resistência à Doença/genética , Regulação da Expressão Gênica de Plantas , Oxigenases de Função Mista/genética , Oryza/genética , Oryza/metabolismo , Oryza/microbiologia , Doenças das Plantas/genética , Doenças das Plantas/microbiologia , Folhas de Planta/genética , Folhas de Planta/imunologia , Folhas de Planta/metabolismo , Folhas de Planta/microbiologia , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Plantas Geneticamente Modificadas/genética , Plantas Geneticamente Modificadas/imunologia , Plantas Geneticamente Modificadas/metabolismo , Plantas Geneticamente Modificadas/microbiologia , Estresse Fisiológico , Xanthomonas/fisiologia
6.
Arch Microbiol ; 203(7): 4189-4199, 2021 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-34076737

RESUMO

Bacillus velezensis is widely known for its inherent biosynthetic potential to produce a wide range of bio-macromolecules and secondary metabolites, including polyketides (PKs) and siderophores, as well as ribosomally and non-ribosomally synthesized peptides. In the present study, we aimed to investigate the bio-macromolecules, such as proteins and peptides of Bacillus velezensis strains, namely A6 and P42 by whole-cell sequencing and highlighted the potential application in controlling phytopathogens. The bioactive compounds, specifically secondary metabolites, were characterized by whole-cell protein profiling, Thin-Layer Chromatography, Infra-Red Spectroscopy, Nuclear Magnetic Resonance, Gas Chromatograph and Electro Spray Liquid Chromatography. Gas Chromatography analysis revealed that the A6 and P42 strains exert different functional groups of compounds, such as aromatic ring, aliphatic, alkene, ketone, amine groups and carboxylic acid. Whole-cell protein profiling of A6 and P42 strains of B. velezensis by nano-ESI LC-MS/MS revealed the presence of 945 and 5303 proteins, respectively. The in vitro evaluation of crude extracts (10%) of A6 and P42 significantly inhibited the rice pathogen, Magnaporthe oryzae (MG01), whereas the cell-free culture filtrate (75%) of strain P42 showed 58.97% inhibition. Similarly, in vitro evaluation of crude extract (10%) of P42 strain inhibited bacterial blight of pomegranate pathogen, Xanthomonas axonopodis pv. punicae, which eventually resulted in a higher inhibition zone of 3 cm, whereas the cell-free extract (75%) of the same strain significantly suppressed the growth of the pathogen with an inhibition zone of 1.48 cm. From the results obtained, the crude secondary metabolites and cell-free filtrates (containing bio-macromolecules) of the strains A6 and P42 of B. velezensis can be employed for controlling the bacterial and fungal pathogens of crop plants.


Assuntos
Ascomicetos , Bacillus , Doenças das Plantas , Xanthomonas axonopodis , Antifúngicos/química , Antifúngicos/isolamento & purificação , Antifúngicos/farmacologia , Ascomicetos/efeitos dos fármacos , Bacillus/química , Cromatografia Líquida , Oryza/microbiologia , Controle Biológico de Vetores , Doenças das Plantas/microbiologia , Doenças das Plantas/prevenção & controle , Romã (Fruta)/microbiologia , Espectrometria de Massas em Tandem , Xanthomonas axonopodis/efeitos dos fármacos
7.
Ecotoxicol Environ Saf ; 220: 112390, 2021 Sep 01.
Artigo em Inglês | MEDLINE | ID: mdl-34098428

RESUMO

Due to its immense capability to concentrate in rice grain and ultimately in food chain, cadmium (Cd) has become the cause of an elevated concern among agriculturists, scientists and the environmental activists. Symbiotic association of Piriformospora indica (P. indica) has been characterized as a potential aid in combating heavy metal stress in plants for sustainable crop production but our scant knowledge regarding ameliorative tendency of P. indica against Cd, specifically in rice, necessitates an in-depth investigation. This study aimed at elaborating the underlying mechanisms involved in P. indica-mediated tolerance against Cd stress in two rice genotypes, IR8 and ZX1H, varying in Cd accumulation pattern. Either colonized or un-inoculated with P. indica, seedlings of both genotypes were subjected to Cd stress. The results showed that P. indica colonization significantly supported plant biomass, photosynthetic attributes and chlorophyll contents in Cd stressed plants. P. indica colonization sustained chloroplast integrity and reduced Cd translocation (46% and 64%), significantly lowering malondialdehyde (MDA) content (11.3% and 50.4%) compared to uninoculated roots under Cd stress in IR8 and ZX1H, respectively. A genotypic difference was evident when a 2-fold enhancement in root peroxidase (POD) activity was recorded in P. indica colonized IR8 plants as compared to ZX1H. The root proteomic analysis was performed using isobaric tags for relative and absolute quantification (iTRAQ) and the results showed that P. indica alleviates Cd stress in rice via down-regulation of key glycolysis cycle enzymes in a bid to reduce energy consumption by the plants and possibly re-directing it to Cd defense response pathways; and up-regulation of glutamine synthetase, a key enzyme in the L-Arg-dependent pathway for nitric oxide (NO) production, which acts as a stress signaling molecule, thus conferring tolerance by reduction of NO-mediated modification of essential proteins in response to Cd stress. Conclusively, both the tested genotypes benefited from P. indica symbiosis at varying levels by an enhanced detoxification capacity and signaling efficiency in response to stress. Hence, a step forward towards the employment of an environmentally sound and self-renewing approach holding the hope for a healthy future.


Assuntos
Basidiomycota/fisiologia , Cádmio/toxicidade , Oryza/efeitos dos fármacos , Oryza/microbiologia , Raízes de Plantas/efeitos dos fármacos , Antioxidantes/metabolismo , Biomassa , Clorofila/metabolismo , Poluentes Ambientais/toxicidade , Malondialdeído/metabolismo , Oryza/metabolismo , Fotossíntese , Raízes de Plantas/metabolismo , Raízes de Plantas/microbiologia , Proteômica , Plântula/metabolismo , Simbiose
8.
Int J Mol Sci ; 22(10)2021 May 13.
Artigo em Inglês | MEDLINE | ID: mdl-34068366

RESUMO

Magnaporthe oryzae (M. oryzae) is a typical cause of rice blast in agricultural production. Isobavachalcone (IBC), an active ingredient of Psoralea corylifolia L. extract, is an effective fungicide against rice blast. To determine the mechanism of IBC against M. oryzae, the effect of IBC on the metabolic pathway of M. oryzae was explored by transcriptome profiling. In M. oryzae, the expression of pyruvate dehydrogenase E1 (PDHE1), part of the tricarboxylic acid (TCA cycle), was significantly decreased in response to treatment with IBC, which was verified by qPCR and testing of enzyme activity. To further elucidate the interactions between IBC and PDHE1, the 3D structure model of the PDHE1 from M. oryzae was established based on homology modeling. The model was utilized to analyze the molecular interactions through molecular docking and molecular dynamics simulation, revealing that IBC has π-π stacking interactions with residue TYR139 and undergoes hydrogen bonding with residue ASP217 of PDHE1. Additionally, the nonpolar residues PHE111, MET174, ILE 187, VAL188, and MET250 form strong hydrophobic interactions with IBC. The above results reveal that PDHE1 is a potential target for antifungal agents, which will be of great significance for guiding the design of new fungicides. This research clarified the mechanism of IBC against M. oryzae at the molecular level, which will underpin further studies of the inhibitory mechanism of flavonoids and the discovery of new targets. It also provides theoretical guidance for the field application of IBC.


Assuntos
Chalconas/farmacologia , Proteínas Fúngicas/metabolismo , Magnaporthe/efeitos dos fármacos , Oryza/enzimologia , Doenças das Plantas/imunologia , Piruvato Desidrogenase (Lipoamida)/antagonistas & inibidores , Transcriptoma/efeitos dos fármacos , Proteínas Fúngicas/genética , Fungicidas Industriais/farmacologia , Perfilação da Expressão Gênica , Regulação Fúngica da Expressão Gênica , Magnaporthe/fisiologia , Simulação de Acoplamento Molecular , Oryza/efeitos dos fármacos , Oryza/microbiologia , Doenças das Plantas/microbiologia , Conformação Proteica , Piruvato Desidrogenase (Lipoamida)/genética , Piruvato Desidrogenase (Lipoamida)/metabolismo
9.
Mol Genet Genomics ; 296(4): 939-952, 2021 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-33966102

RESUMO

Plant major resistance (R) genes are effective in detecting pathogen signal molecules and triggering robust defense responses. Investigating the natural variation in R genes will allow identification of the critical amino acid residues determining recognition specificity in R protein and the discovery of novel R alleles. The rice blast resistance gene Pike, comprising of two adjacent CC-NBS-LRR genes, namely, Pike-1 and Pike-2, confers broad-spectrum resistance to Magnaporthe oryzae. Here, we demonstrated that Pike-1 determined Pike-specific resistance through direct interaction with the pathogen signal molecule AvrPik. Analysis of natural variation in 79 Pike-1 variants in the Asian cultivated rice Oryza sativa and its wild relatives revealed that the CC and NBS regions, particularly the CC region of the Pike-1 protein were the most diversified. We also found that balancing selection had occurred in O. sativa and O. rufipogon to maintain the genetic diversity of the Pike-1 alleles. By analysis of amino acid sequences, we identified 40 Pike-1 variants in these rice germplasms. These variants were divided into three major groups that corresponded to their respective clades. A new Pike allele, designated Pikg, that differed from Pike by a single amino acid substitution (D229E) in the Pike-1 CC region of the Pike protein was identified from wild rice relatives. Pathogen assays of Pikg transgenic plants revealed a unique reaction pattern that was different from that of the previously identified Pike alleles, namely, Pik, Pikh, Pikm, Pikp, Piks and Pi1. These findings suggest that minor amino acid residues in Pike-1/Pikg-1 determine pathogen recognition specificity and plant resistance. As a new blast R gene derived from rice wild relatives, Pikg has potential applications in rice breeding.


Assuntos
Ascomicetos/patogenicidade , Resistência à Doença/genética , Oryza , Proteínas de Plantas/genética , Alelos , Sequência de Aminoácidos , Análise Mutacional de DNA , Genes de Plantas/genética , Genes de Plantas/fisiologia , Estudos de Associação Genética , Variação Genética/fisiologia , Interações Hospedeiro-Patógeno/genética , Interações Hospedeiro-Patógeno/imunologia , Oryza/genética , Oryza/microbiologia , Melhoramento Vegetal , Doenças das Plantas/genética , Doenças das Plantas/imunologia , Plantas Geneticamente Modificadas
10.
Arch Microbiol ; 203(6): 3533-3540, 2021 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-33938971

RESUMO

Plant growth-promoting rhizobacteria (PGPR) are known to stimulate plant growth because of their versatility in nutrient transformation. However, the success of PGPR inoculation depends not only on their ability to promote plant growth but also on their capacity to metabolize substrates that can be used as energy for the development and survival of the crops. Given the important influence of seed germination and vigor on crop yield, this study investigated the biochemical characteristics and effectiveness of multi-trait PGPR isolates in enhancing upland rice seedling growth and vigor. Biochemical identification was done using Biolog GEN III Microbial Identification System. Isolates were characterized based on their ability to metabolize all major classes of biochemicals in the carbon source utilization and chemical sensitivity assays. Identified rhizobacterial isolates were tested in vitro to evaluate their inoculation effects on the growth of PSB Rc23 upland rice seedlings. Biochemical identification results showed that rhizobacterial isolates have extensive metabolic activities in a wide range of carbon sources. Inoculation effects revealed that isolate IBBw1a was the most effective in enhancing root length and vigor index of rice seedlings in vitro, yielding a significant increase of 60% and 53%, respectively, over the uninoculated control. This study suggests that rhizobacterial isolates from upland rice may have commercial significance to improve seedling growth and vigor. These isolates will undergo a further assessment of their effectiveness in actual upland rice field conditions as they were already proven effective growth promoters in laboratory and screenhouse conditions. Such future activity can uncover their efficacy as potential biofertilizers in the actual soil environment.


Assuntos
Oryza , Rhizobiaceae , Plântula , Microbiologia do Solo , Oryza/microbiologia , Desenvolvimento Vegetal , Rhizobiaceae/fisiologia , Plântula/microbiologia , Solo
11.
Nat Commun ; 12(1): 3148, 2021 05 25.
Artigo em Inglês | MEDLINE | ID: mdl-34035309

RESUMO

Structured Illumination Microscopy enables live imaging with sub-diffraction resolution. Unfortunately, optical aberrations can lead to loss of resolution and artifacts in Structured Illumination Microscopy rendering the technique unusable in samples thicker than a single cell. Here we report on the combination of Adaptive Optics and Structured Illumination Microscopy enabling imaging with 150 nm lateral and 570 nm axial resolution at a depth of 80 µm through Caenorhabditis elegans. We demonstrate that Adaptive Optics improves the three-dimensional resolution, especially along the axial direction, and reduces artifacts, successfully realizing 3D-Structured Illumination Microscopy in a variety of biological samples.


Assuntos
Imageamento Tridimensional/métodos , Microscopia Intravital/métodos , Iluminação/instrumentação , Animais , Artefatos , Ascomicetos , Caenorhabditis elegans , Linhagem Celular , Imageamento Tridimensional/instrumentação , Microscopia Intravital/instrumentação , Camundongos , Microscopia Confocal/instrumentação , Microscopia Confocal/métodos , Microscopia de Fluorescência/instrumentação , Microscopia de Fluorescência/métodos , Oryza/microbiologia , Reprodutibilidade dos Testes
12.
Arch Microbiol ; 203(6): 3257-3268, 2021 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-33837802

RESUMO

This study aimed to identify four isolates of Trichoderma sp. (Ufra.T06, Ufra.T09, Ufra.T12, and Ufra.T52) and characterize their interaction with Magnaporthe oryzae in vitro and in vivo conditions. The four isolates of Trichoderma sp. were sequenced, investigated as an antagonist against M. oryzae in five Petri plate assays, and as an inhibitor of conidial germination appressoria formation. Finally, were quantified the lytic activity of chitinase (CHI), glucanase (GLU), and protease (PRO) during co-cultivation of Trichoderma sp. and M. oryzae. In vivo, leaf blast suppression was evaluated in two assays: simultaneous and curative application. Both in vitro and in vivo assays were scanned by electron microscopy (SEM). All isolates were identified as Trichoderma asperellum. All in vitro Petri plates assays reduced M. oryzae colony growth (paired-91.18% by Ufra.T09, volatile metabolites-all isolates equally reduced, non-volatile-68.33% by Ufra.T06, thermostability-99.77% by Ufra.T52 and co-cultivate-64.25% by Ufra.T52). The filtrates and conidia suspensions for T. asperellum isolates inhibited the conidia germination and appressoria formation significantly. In co-cultivate (mycelial or cell wall), all enzymes (GLU, CHI, and PRO) and times (24, 48, and 72 h) showed increased activity. In vivo, reduced leaf blast severity until 94.64% (Ufra.T52cs) in a simultaneous and until 85% (Ufra.T09 24 and 48 hasi) in a curative application. T. asperellum isolates showed efficient control of M. oryzae by mycoparasitism, and antibiosis mechanisms were interfered with by the M. oryzae infection process.


Assuntos
Antibiose , Ascomicetos , Hypocreales , Oryza , Ascomicetos/fisiologia , Ascomicetos/ultraestrutura , Hypocreales/genética , Hypocreales/ultraestrutura , Oryza/microbiologia , Doenças das Plantas/microbiologia
13.
Viruses ; 13(4)2021 03 31.
Artigo em Inglês | MEDLINE | ID: mdl-33807245

RESUMO

Jumbo phages have DNA genomes larger than 200 kbp in large virions composed of an icosahedral head, tail, and other adsorption structures, and they are known to be abundant biological substances in nature. In this study, phages in leaf litter compost were screened for their potential to suppress rice seedling rot disease caused by the bacterium Burkholderia glumae, and a novel phage was identified in a filtrate-enriched suspension of leaf litter compost. The phage particles consisted of a rigid tailed icosahedral head and contained a DNA genome of 227,105 bp. The phage could lyse five strains of B. glumae and six strains of Burkholderia plantarii. The phage was named jumbo Burkholderia phage FLC6. Proteomic tree analysis revealed that phage FLC6 belongs to the same clade as two jumbo Ralstonia phages, namely RSF1 and RSL2, which are members of the genus Chiangmaivirus (family: Myoviridae; order: Caudovirales). Interestingly, FLC6 could also lyse two strains of Ralstonia pseudosolanacearum, the causal agent of bacterial wilt, suggesting that FLC6 has a broad host range that may make it especially advantageous as a bio-control agent for several bacterial diseases in economically important crops. The novel jumbo phage FLC6 may enable leaf litter compost to suppress several bacterial diseases and may itself be useful for controlling plant diseases in crop cultivation.


Assuntos
Bacteriófagos/genética , Bacteriófagos/isolamento & purificação , Agentes de Controle Biológico/isolamento & purificação , Burkholderia/virologia , Compostagem , Folhas de Planta/virologia , Plântula/microbiologia , Bacteriófagos/química , Agentes de Controle Biológico/farmacologia , Burkholderia/patogenicidade , Genoma Viral/genética , Especificidade de Hospedeiro , Oryza/microbiologia , Terapia por Fagos , Doenças das Plantas/terapia , Folhas de Planta/microbiologia , Proteômica , Ralstonia/patogenicidade , Ralstonia/virologia
14.
BMC Plant Biol ; 21(1): 197, 2021 Apr 24.
Artigo em Inglês | MEDLINE | ID: mdl-33894749

RESUMO

BACKGROUND: Rice leaf blight, which is a devastating disease worldwide, is caused by the bacterium Xanthomonas oryzae pv. oryzae (Xoo). The upregulated by transcription activator-like 1 (UPT) effector box in the promoter region of the rice Xa13 gene plays a key role in Xoo pathogenicity. Mutation of a key bacterial protein-binding site in the UPT box of Xa13 to abolish PXO99-induced Xa13 expression is a way to improve rice resistance to bacteria. Highly efficient generation and selection of transgene-free edited plants are helpful to shorten and simplify the gene editing-based breeding process. Selective elimination of transgenic pollen of T0 plants can enrich the proportion of T1 transgene-free offspring, and expression of a color marker gene in seeds makes the selection of T2 plants very convenient and efficient. In this study, a genome editing and multiplexed selection system was used to generate bacterial leaf blight-resistant and transgene-free rice plants. RESULTS: We introduced site-specific mutations into the UPT box using CRISPR/Cas12a technology to hamper with transcription-activator-like effector (TAL) protein binding and gene activation and generated genome-edited rice with improved bacterial blight resistance. Transgenic pollen of T0 plants was eliminated by pollen-specific expression of the α-amylase gene Zmaa1, and the proportion of transgene-free plants increased from 25 to 50% among single T-DNA insertion events in the T1 generation. Transgenic seeds were visually identified and discarded by specific aleuronic expression of DsRed, which reduced the cost by 50% and led to up to 98.64% accuracy for the selection of transgene-free edited plants. CONCLUSION: We demonstrated that core nucleotide deletion in the UPT box of the Xa13 promoter conferred resistance to rice blight, and selection of transgene-free plants was boosted by introducing multiplexed selection. The combination of genome editing and transgene-free selection is an efficient strategy to accelerate functional genomic research and plant breeding.


Assuntos
Resistência à Doença , Edição de Genes/métodos , Genoma de Planta , Oryza/genética , Doenças das Plantas/genética , Xanthomonas/fisiologia , Oryza/microbiologia , Melhoramento Vegetal , Doenças das Plantas/microbiologia , Transgenes
15.
Toxins (Basel) ; 13(3)2021 03 11.
Artigo em Inglês | MEDLINE | ID: mdl-33799626

RESUMO

Multiple mycotoxins were tested in milled rice samples (n = 200) from traders at different milling points within the Mwea Irrigation Scheme in Kenya. Traders provided the names of the cultivar, village where paddy was cultivated, sampling locality, miller, and month of paddy harvest between 2018 and 2019. Aflatoxin, citrinin, fumonisin, ochratoxin A, diacetoxyscirpenol, T2, HT2, and sterigmatocystin were analyzed using ultra-high-performance liquid chromatography-tandem mass spectrometry (UHPLC-MS/MS). Deoxynivalenol was tested using enzyme-linked immunosorbent assay (ELISA). Mycotoxins occurred in ranges and frequencies in the following order: sterigmatocystin (0-7 ppb; 74.5%), aflatoxin (0-993 ppb; 55.5%), citrinin (0-9 ppb; 55.5%), ochratoxin A (0-110 ppb; 30%), fumonisin (0-76 ppb; 26%), diacetoxyscirpenol (0-24 ppb; 20.5%), and combined HT2 + T2 (0-62 ppb; 14.5%), and deoxynivalenol was detected in only one sample at 510 ppb. Overall, low amounts of toxins were observed in rice with a low frequency of samples above the regulatory limits for aflatoxin, 13.5%; ochratoxin A, 6%; and HT2 + T2, 0.5%. The maximum co-contamination was for 3.5% samples with six toxins in different combinations. The rice cultivar, paddy environment, time of harvest, and millers influenced the occurrence of different mycotoxins. There is a need to establish integrated approaches for the mitigation of mycotoxin accumulation in the Kenyan rice.


Assuntos
Ração Animal/microbiologia , Produtos Agrícolas/microbiologia , Microbiologia de Alimentos , Fungos/metabolismo , Micotoxinas/análise , Oryza/microbiologia , Animais , Cromatografia Líquida de Alta Pressão , Produtos Agrícolas/crescimento & desenvolvimento , Estudos Transversais , Manipulação de Alimentos , Humanos , Quênia , Micotoxinas/efeitos adversos , Oryza/crescimento & desenvolvimento , Medição de Risco , Espectrometria de Massas por Ionização por Electrospray , Espectrometria de Massas em Tandem
16.
Toxins (Basel) ; 13(3)2021 03 19.
Artigo em Inglês | MEDLINE | ID: mdl-33808596

RESUMO

Ochratoxin A (OTA) is a potential human carcinogen that poses a significant concern in food safety and public health. OTA has been found in a wide variety of agricultural commodities, including cereal grains. This study investigated the reduction of OTA during the preparation of rice- and oat-based porridge by a simulated indirect steam process. The effects of sodium bicarbonate (NaHCO3) and fructose on the reduction of OTA were also investigated. During the processing, OTA in rice- and oat-porridge was decreased by 59% and 14%, respectively, from initial OTA artificially added at 20 µg/kg (dry weight basis). When 0.5% and 1% of sodium bicarbonate were added to rice porridge, increased reduction of OTA was observed as 78% and 68%, respectively. The same amounts of added sodium bicarbonate also further reduced OTA in oat porridge to 58% and 72%, respectively. In addition, increased reduction of OTA in the presence of fructose was observed. A combination of the two, i.e., 0.5% sodium bicarbonate and 0.5% fructose, resulted in a 79% and 67% reduction in rice porridge and oat porridge, respectively. These results indicate that indirect steaming may effectively reduce OTA in preparation of porridge-type products, particularly when sodium bicarbonate and/or fructose are added.


Assuntos
Avena/microbiologia , Culinária , Microbiologia de Alimentos , Frutose/química , Ocratoxinas/análise , Oryza/microbiologia , Bicarbonato de Sódio/química , Análise de Alimentos , Temperatura Alta , Vapor
17.
Toxins (Basel) ; 13(4)2021 04 15.
Artigo em Inglês | MEDLINE | ID: mdl-33920815

RESUMO

Rice bran, a by-product of the rice milling process, has emerged as a functional food and being used in formulation of healthy food and drinks. However, rice bran is often contaminated with numerous mycotoxins. In this study, a method to simultaneous detection of aflatoxins (AFB1, AFB2, AFG1, and AFG2), ochratoxin A (OTA), deoxynivalenol (DON), fumonisins (FB1 and FB2), sterigmatocystin (STG), T-2 toxin, HT-2 toxin, diacetoxyscirpenol (DAS) and zearalenone (ZEA) in rice bran was developed, optimized and validated using dispersive liquid-liquid microextraction (DLLME) and liquid chromatography-tandem mass spectrometry (LC-MS/MS). In DLLME, using a solvent mixture of methanol/water (80:20, v/v) as the dispersive solvent and chloroform as the extraction solvent with the addition of 5% salt improved the extraction recoveries (63-120%). The developed method was further optimized using the response surface methodology (RSM) combined with Box-Behnken Design (BBD). Under the optimized experimental conditions, good linearity was obtained with a correlation coefficient (r2) ≥ 0.990 and a limit of detection (LOD) between 0.5 to 50 ng g-1. The recoveries ranged from 70.2% to 99.4% with an RSD below 1.28%. The proposed method was successfully applied to analyze multi-mycotoxin in 24 rice bran samples.


Assuntos
Cromatografia Líquida de Alta Pressão , Análise de Alimentos , Microbiologia de Alimentos , Microextração em Fase Líquida , Micotoxinas/análise , Oryza/microbiologia , Espectrometria de Massas em Tandem , Manipulação de Alimentos , Química Verde , Reprodutibilidade dos Testes
18.
Molecules ; 26(6)2021 Mar 17.
Artigo em Inglês | MEDLINE | ID: mdl-33802719

RESUMO

Plant diseases reduce crop yield and quality, hampering the development of agriculture. Fungicides, which restrict chemical synthesis in fungi, are the strongest controls for plant diseases. However, the harmful effects on the environment due to continued and uncontrolled utilization of fungicides have become a major challenge in recent years. Plant-sourced fungicides are a class of plant antibacterial substances or compounds that induce plant defenses. They can kill or inhibit the growth of target pathogens efficiently with no or low toxicity, they degrade readily, and do not prompt development of resistance, which has led to their widespread use. In this study, the growth inhibition effect of 24 plant-sourced ethanol extracts on rice sprigs was studied. Ethanol extract of gallnuts and cloves inhibited the growth of bacteria by up to 100%. Indoor toxicity measurement results showed that the gallnut and glove constituents inhibition reached 39.23 µg/mL and 18.82 µg/mL, respectively. Extract treated rice sprigs were dry and wrinkled. Gallnut caused intracellular swelling and breakage of mitochondria, disintegration of nuclei, aggregation of protoplasts, and complete degradation of organelles in hyphae and aggregation of cellular contents. Protection of Rhizoctonia solani viability reached 46.8% for gallnut and 37.88% for clove in water emulsions of 1000 µg/mL gallnut and clove in the presence of 0.1% Tween 80. The protection by gallnut was significantly stronger than that of clove. The data could inform the choice of plant-sourced fungicides for the comprehensive treatment of rice sprig disease. The studied extract effectively protected rice sprigs and could be a suitable alternative to commercially available chemical fungicides. Further optimized field trials are needed to effectively sterilize rice paddies.


Assuntos
Misturas Complexas/farmacologia , Oryza/efeitos dos fármacos , Extratos Vegetais/farmacologia , Rhizoctonia/efeitos dos fármacos , Rhus/química , Syzygium/química , Cromatografia por Troca Iônica , Misturas Complexas/toxicidade , Etanol/química , Eugenol/análise , Fungicidas Industriais/farmacologia , Ácidos Láuricos/análise , Espectrometria de Massas , Microscopia Eletrônica de Varredura , Microscopia Eletrônica de Transmissão , Mitocôndrias/efeitos dos fármacos , Mitocôndrias/ultraestrutura , Micélio/efeitos dos fármacos , Micélio/ultraestrutura , Oryza/microbiologia , Doenças das Plantas/microbiologia , Extratos Vegetais/toxicidade
19.
Int J Mol Sci ; 22(8)2021 Apr 20.
Artigo em Inglês | MEDLINE | ID: mdl-33924035

RESUMO

The nucleotide-binding site-leucine-rich repeat (NBS-LRR) gene family is the largest group of plant disease resistance (R) genes widespread in response to viruses, bacteria, and fungi usually involved in effector triggered immunity (ETI). Forty members of the Chinese cabbage CC type NBS-LRR family were investigated in this study. Gene and protein characteristics, such as distributed locations on chromosomes and gene structures, were explored through comprehensive analysis. CC-NBS-LRR proteins were classified according to their conserved domains, and the phylogenetic relationships of CC-NBS-LRR proteins in Brassica rapa, Arabidopsis thaliana, and Oryza sativa were compared. Moreover, the roles of BrCC-NBS-LRR genes involved in pathogenesis-related defense were studied and analyzed. First, the expression profiles of BrCC-NBS-LRR genes were detected by inoculating with downy mildew and black rot pathogens. Second, sensitive and resistant Chinese cabbage inbred lines were screened by downy mildew and black rot. Finally, the differential expression levels of BrCC-NBS-LRR genes were monitored at 0, 1, 3, 6, 12 and 24 h for short and 0, 3, 5, 7, 10 and 14 days for long inoculation time. Our study provides information on BrCC-NBS-LRR genes for the investigation of the functions and mechanisms of CC-NBS-LRR genes in Chinese cabbage.


Assuntos
Brassica/metabolismo , Brassica/microbiologia , Doenças das Plantas/microbiologia , Arabidopsis/microbiologia , Resistência à Doença , Regulação da Expressão Gênica de Plantas , Oryza/microbiologia , Filogenia , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo
20.
Molecules ; 26(7)2021 Mar 24.
Artigo em Inglês | MEDLINE | ID: mdl-33805102

RESUMO

Three new helvolic acid derivatives (named sarocladilactone A (1), sarocladilactone B (2) and sarocladic acid A (3a)), together with five known compounds (6,16-diacetoxy-25-hy- droxy-3,7-dioxy-29-nordammara-1,17(20)-dien-21-oic acid (3b), helvolic acid (4), helvolinic acid (5), 6-desacetoxy-helvolic acid (6) and 1,2-dihydrohelvolic acid (7)), were isolated from the endophytic fungus DX-THL3, obtained from the leaf of Dongxiang wild rice (Oryza rufipogon Griff.). The structures of the new compounds were elucidated via HR-MS, extensive 1D and 2D NMR analysis and comparison with reported data. Compounds 1, 2, 4, 5, 6 and 7 exhibited potent antibacterial activities. In particular, sarocladilactone B (2), helvolinic acid (5) and 6-desacetoxy-helvolic acid (6) exhibited strongly Staphylococcus aureus inhibitory activity with minimum inhibitory concentration (MIC) values of 4, 1 and 4 µg/mL, respectively. The structure-activity relationship (SAR) of these compounds was primarily summarized.


Assuntos
Antibacterianos , Ácido Fusídico/análogos & derivados , Hypocreales/química , Oryza/microbiologia , Staphylococcus aureus/crescimento & desenvolvimento , Antibacterianos/química , Antibacterianos/isolamento & purificação , Antibacterianos/farmacologia , Ácido Fusídico/química , Ácido Fusídico/isolamento & purificação , Ácido Fusídico/farmacologia
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