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OBJECTIVE: Identification and characterization of a novel bacterial carbohydrate esterase (PaCes7) with application potential for lignocellulose and pesticide degradation. RESULTS: PaCes7 was identified from the lignocellulolytic bacterium, Pantoea ananatis Sd-1 as a new carbohydrate esterase. Recombinant PaCes7 heterologously expressed in Escherichia coli showed a clear preference for esters with short-chain fatty acids and exhibited maximum activity towards α-naphthol acetate at 37 °C and pH 7.5. Purified PaCes7 exhibited its catalytic activity under mesophilic conditions and retained more than 40% activity below 30 °C. It displayed a relatively wide pH stability from pH 6-11. Furthermore, the enzyme was strongly resistant to Mg2+, Pb2+, and Co2+ and activated by K+ and Ca2+. Both P. ananatis Sd-1 and PaCes7 could degrade the pesticide carbaryl. Additionally, PaCes7 was shown to work in combination with cellulase and/or xylanase in rice straw degradation. CONCLUSIONS: The data suggest that PaCes7 possesses promising biotechnological potential.
Assuntos
Proteínas de Bactérias , Esterases , Lignina/metabolismo , Pantoea/enzimologia , Praguicidas/metabolismo , Proteínas de Bactérias/química , Proteínas de Bactérias/genética , Proteínas de Bactérias/metabolismo , Biodegradação Ambiental , Carbaril/metabolismo , Estabilidade Enzimática , Esterases/química , Esterases/genética , Esterases/metabolismo , Pantoea/genéticaRESUMO
Microbial antagonists and their bioactive metabolites provide one of the best alternatives to chemical pesticides to control crop disease for sustainable agriculture and global food security. The rice endophyte Streptomyces hygroscopicus OsiSh-2, with remarkable antagonistic activity towards the rice blast fungus Magnaporthe oryzae, was reported in our previous study. The present study deciphered the possible direct interaction mode of OsiSh-2 against M. oryzae. An in vitro antibiotic assay for OsiSh-2 culture filtrate revealed strong suppression of mycelial growth, conidial germination and appressorial formation of M. oryzae. Meanwhile, severe morphological and internal abnormalities in M. oryzae hyphae were observed under a scanning electron microscope and transmission electron microscope. Foliar treatment of rice seedlings by OsiSh-2 culture filtrate in the greenhouse and in the field showed 23.5% and 28.3% disease reduction, respectively. Correspondingly, OsiSh-2 culture filtrate could induce disorganized chitin deposition in the cell wall and lowered ergosterol content in the cell membrane of M. oryzae. Additionally, cell wall integrity pathway activation, large cell electrolytes release, reactive oxygen species accumulation and tricarboxylic acid cycle-related enzyme activity changes were found in M. oryzae. All these results suggested that the direct antagonistic activity of OsiSh-2 against M. oryzae may be attributed to damaging the integrity of the cell wall and membrane and disrupting mitochondrial function in the pathogen.
Assuntos
Antifúngicos/farmacologia , Endófitos/fisiologia , Magnaporthe/efeitos dos fármacos , Oryza/microbiologia , Controle Biológico de Vetores , Streptomyces/químicaRESUMO
This work demonstrates conservation of average orbital angular momentum for high harmonic generation of fractional vortex beams. High harmonics are generated in reflected light beams in a three-dimensional particle-in-cell simulation. The average orbital angular momentum of the beam is calculated when a relativistic linearly polarized fractional vortex beam impinges on a solid foil. The harmonic generation progress can be well explained by using the vortex oscillating mirror model. Both simulation and theoretical analysis show that the average orbital momentum of the nth harmonic is n times that of the fundamental frequency beam. This provides evidence that the average orbital angular momentum obeys momentum conservation during the harmonic generation of fractional vortex beams.
RESUMO
OBJECTIVE: To identify and characterize a novel bacterial pyranose 2-oxidase (P2Ox) and investigate its potential use in lignin degradation applications. RESULTS: A new bacterial P2Ox (PaP2Ox) enzyme was identified in the lignocellulolytic bacterium Pantoea ananatis Sd-1. The PaP2Ox open reading frame was cloned, and the encoded protein was heterologously expressed in an Escherichia coli expression system. Unlike another reported bacterial P2Ox enzyme, the purified PaP2Ox exhibits a homotetrameric spatial conformation that is similar to fungal P2Oxs, with each subunit having a molecular mass of 65 kDa. The recombinant PaP2Ox exhibits maximum activity at 50 °C and pH 6.5 with D-glucose as its preferred substrate. In addition, this enzyme was shown to work in combination with bacterial laccase in lignin degradation. CONCLUSIONS: The bacterial enzyme PaP2Ox has potential use in ligninolytic systems and shows promising value in industrial biotechnological applications.
Assuntos
Desidrogenases de Carboidrato/genética , Desidrogenases de Carboidrato/metabolismo , Pantoea/enzimologia , Proteínas de Bactérias/química , Proteínas de Bactérias/genética , Proteínas de Bactérias/metabolismo , Desidrogenases de Carboidrato/química , Clonagem Molecular , Regulação Bacteriana da Expressão Gênica , Regulação Enzimológica da Expressão Gênica , Temperatura Alta , Concentração de Íons de Hidrogênio , Lacase/metabolismo , Lignina/química , Modelos Moleculares , Peso Molecular , Pantoea/genética , Conformação Proteica , Multimerização Proteica , ProteóliseRESUMO
Secreted common fungal extracellular membrane (CFEM) domain proteins have been implicated in multiple biological functions in fungi. However, it is still largely unknown whether the ferric iron (Fe3+), as an important trace element, was involved with the biological function of CFEM proteins. In this study, a new CFEM protein CgCsa, with high expression levels at the early inoculation stage on peppers by Colletotrichum gloeosporioides was investigated. Deletion of the targeted gene CgCsa revealed multiple biological roles in hyphal growth restriction, highly reduced conidial yield, delayed conidial germination, abnormal appressorium with elongated bud tubes, and significantly reduced virulence of C. gloeosporioides. Moreover, in CgCsa mutants, the expression levels of four cell wall synthesis-related genes were downregulated, and cell membrane permeability and electrical conductivity were increased. Compared to the wild-type, the CgCsa mutants downregulated expressions of iron transport-related genes, in addition, its three-dimensional structure was capable binding with iron. Increase in the Fe3+ concentration in the culture medium partially recovered the functions of ΔCgCsa mutant. This is probably the first report to show the association between CgCsa and iron homeostasis in C. gloeosporioides. The results suggest an alternative pathway for controlling plant fungal diseases by deplete their trace elements.
Assuntos
Colletotrichum , Proteínas Fúngicas , Regulação Fúngica da Expressão Gênica , Homeostase , Ferro , Colletotrichum/patogenicidade , Colletotrichum/genética , Colletotrichum/crescimento & desenvolvimento , Proteínas Fúngicas/genética , Proteínas Fúngicas/metabolismo , Ferro/metabolismo , Virulência/genética , Esporos Fúngicos/crescimento & desenvolvimento , Doenças das Plantas/microbiologia , Hifas/crescimento & desenvolvimento , Mutação , Proteínas de Membrana/genética , Proteínas de Membrana/metabolismoRESUMO
Ferroptosis is an iron-dependent cell death that was discovered recently. For beneficial microbes to establish mutualistic relationships with hosts, precisely controlled cell death in plant cells is necessary. However, whether ferroptosis is involved in the endophyteâplant system is poorly understood. Here, we reported that endophytic Streptomyces hygroscopicus OsiSh-2, which established a sophisticated and beneficial interaction with host rice plants, caused ferroptotic cell death in rice characterized by ferroptosis- and immune-related markers. Treatments with ferroptosis inhibitors and inducers, different doses of OsiSh-2, and the siderophore synthesis-deficient mutant ΔcchH revealed that only moderate ferroptosis induced by endophytes is essential for the establishment of an optimal symbiont to enhance plant growth. Additionally, ferroptosis involved in a defence-primed state in rice, which contributed to improved resistance against rice blast disease. Overall, our study provides new insights into the mechanisms of endophyteâplant interactions mediated by ferroptosis and suggests new directions for crop yield promotion.
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Resistência à Doença , Endófitos , Ferroptose , Oryza , Doenças das Plantas , Streptomyces , Simbiose , Oryza/microbiologia , Oryza/genética , Oryza/imunologia , Ferroptose/genética , Endófitos/fisiologia , Resistência à Doença/genética , Doenças das Plantas/microbiologia , Streptomyces/genética , Streptomyces/fisiologia , Sideróforos/metabolismo , Ferro/metabolismoRESUMO
Soil arsenic (As) phytoremediation has long faced the challenge of efficiently absorbing As by plant accumulators while maintaining their health and fast growth. Even at low doses, arsenic is highly toxic to plants. Therefore, plant growth-promoting microorganisms that can mediate As accumulation in plants are of great interest. In this study, the endophyte Enterobacter sp. YG-14 (YG-14) was found to have soil mobilization activity. By constructing a siderophore synthesis gene deletion mutant (ΔentD) of YG-14, the endophyte was confirmed to effectively mobilize Fe-As complexes in mining soil by secreting enterobactin, releasing bioavailable Fe and As to the rhizosphere. YG-14 also enhances As accumulation in host plants via extracellular polymer adsorption and specific phosphatase transfer protein (PitA) absorption. The root accumulation of As was positively correlated with YG-14 root colonization. In addition, YG-14 promoted plant growth and alleviated oxidative damage in R. pseudoacacia L. under arsenic stress. This is the first study, from phenotype, physiology, and molecular perspectives, to determine the role of endophyte in promoting As phytostabilization and maintaining the growth of the host plant. This demonstrated the feasibility of using endophytes with high siderophore production to assist host plants in As phytoremediation.
Assuntos
Arsênio , Poluentes do Solo , Arsênio/metabolismo , Enterobacter/metabolismo , Sideróforos/metabolismo , Endófitos , Plantas/metabolismo , Solo , Biodegradação Ambiental , Poluentes do Solo/metabolismoRESUMO
Endophytes play an important role in shaping plant growth and immunity. However, the mechanisms for endophyte-induced disease resistance in host plants remain unclear. Here, we screened and isolated the immunity inducer ShAM1 from the endophyte Streptomyces hygroscopicus OsiSh-2, which strongly antagonizes the pathogen Magnaporthe oryzae. Recombinant ShAM1 can trigger rice immune responses and induce hypersensitive responses in various plant species. After infection with M. oryzae, blast resistance was dramatically improved in ShAM1-inoculated rice. In addition, the enhanced disease resistance by ShAM1 was found to occur through a priming strategy and was mainly regulated through the jasmonic acid-ethylene (JA/ET)-dependent signaling pathway. ShAM1 was identified as a novel α-mannosidase, and its induction of immunity is dependent on its enzyme activity. When we incubated ShAM1 with isolated rice cell walls, the release of oligosaccharides was observed. Notably, extracts from the ShAM1-digested cell wall can enhance the disease resistance of the host rice. These results indicated that ShAM1 triggered immune defense against pathogens by damage-associated molecular pattern (DAMP)-related mechanisms. Our work provides a representative example of endophyte-mediated modulation of disease resistance in host plants. The effects of ShAM1 indicate the promise of using active components from endophytes as plant defense elicitors for the management of plant disease. IMPORTANCE The specific biological niche inside host plants allows endophytes to regulate plant disease resistance effectively. However, there have been few reports on the role of active metabolites from endophytes in inducing host disease resistance. In this study, we demonstrated that an identified α-mannosidase protein, ShAM1, secreted by the endophyte S. hygroscopicus OsiSh-2 could activate typical plant immunity responses and induce a timely and cost-efficient priming defense against the pathogen M. oryzae in rice. Importantly, we revealed that ShAM1 enhanced plant disease resistance through its hydrolytic enzyme (HE) activity to digest the rice cell wall and release damage-associated molecular patterns. Taken together, these findings provide an example of the interaction mode of endophyte-plant symbionts and suggest that HEs derived from endophytes can be used as environmentally friendly and safe prevention agent for plant disease control.
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Magnaporthe , Oryza , Resistência à Doença , Endófitos/fisiologia , alfa-Manosidase/metabolismo , alfa-Manosidase/farmacologia , Magnaporthe/metabolismo , Doenças das Plantas , Parede CelularRESUMO
Endophytes can benefit the growth and stress resistance of host plants by secreting bioactive components. Thiamine is an essential vitamin involved in many metabolic pathways and can only be synthesized by microbes and plants. In this study, we found that thiamine could inhibit the development of the phytopathogen Magnaporthe oryzae and decrease the rice blast index under field conditions. In the thiamine biosynthesis pathway, the key enzyme ShTHIC of an endophyte Streptomyces hygroscopicus OsiSh-2 and OsTHIC of rice (Oryza sativa) were highly homologous. Gene overexpression or knockout approaches revealed that both THIC contributed to thiamine synthesis and resistance to M. oryzae. Furthermore, S. hygroscopicus OsiSh-2 colonization led to a decrease in the thiamine synthesis level of rice but still maintained thiamine homeostasis in rice. However, inoculation with the ShTHIC knockout strain ΔTHIC reduced the thiamine content in rice, although the thiamine synthesis level of rice was increased. After infection with M. oryzae, blast resistance was dramatically improved in OsiSh-2-inoculated rice but decreased in ΔTHIC-inoculated rice compared with non-inoculated rice. This result demonstrated that ShTHIC could regulate thiamine biosynthesis and consequently assist blast resistance in the OsiSh-2-rice symbiont. Our results revealed a novel blast-resistance mechanism mediated by a key thiamine biosynthetic enzyme from an endophyte OsiSh-2.
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Magnaporthe , Oryza , Resistência à Doença/genética , Endófitos/genética , Oryza/genética , Oryza/metabolismo , Doenças das Plantas/genética , Tiamina/metabolismoRESUMO
The limited availability of nutrient Fe severely impairs the health of almost all organisms. Endophytic actinobacteria can benefit the host plant in different ways. We previously inferred that the rice (Oryza) endophytic Streptomyces hygroscopicus OsiSh-2 possesses a highly efficient Fe-acquisition system. In this work, we first evaluated the effects of OsiSh-2 on the Fe-deficiency resilience of the host rice. The results demonstrated that the inoculation of OsiSh-2 considerably increased the plant biomass, Fe concentration and translocation factor, and chlorophyll content, and net leaf photosynthetic rate under Fe limiting condition. The expression of genes involved with Fe3+-reduction-related strategy in rice was up-regulated, while that involved with Fe3+-chelation-related strategy was down-regulated by OsiSh-2 treatment. Meanwhile, the OsiSh-2-rice symbiont showed enhancement of Fe3+-chelate reductase activity, total siderophore production, and acidification trend in the rhizosphere under Fe deficiency compared to plants without this endophyte. In conclusion, endophytic OsiSh-2 could protect plants against Fe-deficient stress by a sophisticated interaction with the host, including modulating Fe chelation, solubilization, reduction and translocation, ultimately leading to enhanced fitness of plant.
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Deficiências de Ferro , Oryza/microbiologia , Streptomyces/fisiologia , Endófitos/fisiologia , Oryza/fisiologia , SideróforosRESUMO
Plants fine-tune the growth-defense trade-off to survive when facing pathogens. Meanwhile, plant-associated microbes, such as the endophytes inside plant tissues, can benefit plant growth and stress resilience. However, the mechanisms for the beneficial microbes to increase stress resistance with little yield penalty in host plants remain poorly understood. In the present study, we report that endophytic Streptomyces hygroscopicus OsiSh-2 can form a sophisticated interaction with host rice, maintaining cellular homeostasis under pathogen-infection stress, and optimize plant growth and disease resistance in rice. Four-year field trials consistently showed that OsiSh-2 could boost host resistance to rice blast pathogen Magnaporthe oryzae while still maintaining a high yield. The integration of the proteomic, physiological, and transcriptional profiling analysis revealed that OsiSh-2 induced rice defense priming and controlled the expression of energy-consuming defense-related proteins, thus increasing the defense capability with the minimized costs of plant immunity. Meanwhile, OsiSh-2 improved the chloroplast development and optimally maintained the expression of proteins related to plant growth under pathogen stress, thus promoting the crop yield. Our results provided a representative example of an endophyte-mediated modulation of disease resistance and fitness in the host plant. The multilayer effects of OsiSh-2 implicate a promising future of using endophytic actinobacteria for disease control and crop yield promotion. IMPORTANCE Under disease stress, activation of defense response in plants often comes with the cost of a reduction in growth and yield, which is referred as the growth-defense trade-off. The microorganisms which can be recruited by plants to mitigate the growth-defense trade-off are of great value in crop breeding. Here, we reported a rice endophytic actinomycetes Streptomyces hygroscopicus OsiSh-2, which can improve host performances on resistance to rice blast while still sustaining high yield in the 4-year field trials. The proteomic, physiological, and transcriptional profiling data offer insights into the molecular basis underlying the balancing between defense and growth in OsiSh-2-rice symbiont. The findings provide an example for the endophyte-mediated modulation of growth-defense trade-offs in plants and indicated the promising application of endophytic actinobacterial strains in agriculture to breed "microbe-optimized crops."