Spontaneous Mutations in the Nitrate Reductase Gene napC Drive the Emergence of Eco-friendly Low-N2O-Emitting Alfalfa Rhizobia in Regions with Different Climates
We have recently shown that commercial alfalfa inoculants (e.g., Sinorhizobium meliloti B399), which are closely related to the denitrifier model strain Sinorhizobium meliloti 1021, have conserved nitrate, nitrite, and nitric oxide reductases associated with the production of the greenhouse gas nitr...
| Main Authors: | , , , , , , , , |
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| Format: | info:ar-repo/semantics/artículo |
| Language: | Inglés |
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Springer
2020
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| Online Access: | http://hdl.handle.net/20.500.12123/7665 https://link.springer.com/article/10.1007/s00248-019-01473-w https://doi.org/10.1007/s00248-019-01473-w |
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| author | Brambilla, Silvina Maricel Soto, Gabriela Odorizzi, Ariel Arolfo, Valeria McCormick, Wayne Primo, Emiliano Giordano, Walter Jozefkowicz, Cintia Ayub, Nicolás Daniel |
| author_browse | Arolfo, Valeria Ayub, Nicolás Daniel Brambilla, Silvina Maricel Giordano, Walter Jozefkowicz, Cintia McCormick, Wayne Odorizzi, Ariel Primo, Emiliano Soto, Gabriela |
| author_facet | Brambilla, Silvina Maricel Soto, Gabriela Odorizzi, Ariel Arolfo, Valeria McCormick, Wayne Primo, Emiliano Giordano, Walter Jozefkowicz, Cintia Ayub, Nicolás Daniel |
| author_sort | Brambilla, Silvina Maricel |
| collection | INTA Digital |
| description | We have recently shown that commercial alfalfa inoculants (e.g., Sinorhizobium meliloti B399), which are closely related to the denitrifier model strain Sinorhizobium meliloti 1021, have conserved nitrate, nitrite, and nitric oxide reductases associated with the production of the greenhouse gas nitrous oxide (N2O) from nitrate but lost the N2O reductase related to the degradation of N2O to gas nitrogen. Here, we screened a library of nitrogen-fixing alfalfa symbionts originating from different ecoregions and containing N2O reductase genes and identified novel rhizobia (Sinorhizobium meliloti INTA1–6) exhibiting exceptionally low N2O emissions. To understand the genetic basis of this novel eco-friendly phenotype, we sequenced and analyzed the genomes of these strains, focusing on their denitrification genes, and found mutations only in the nitrate reductase structural gene napC. The evolutionary analysis supported that, in these natural strains, the denitrification genes were inherited by vertical transfer and that their defective nitrate reductase napC alleles emerged by independent spontaneous mutations. In silico analyses showed that mutations in this gene occurred in ssDNA loop structures with high negative free energy (−ΔG) and that the resulting mutated stem-loop structures exhibited increased stability, suggesting the occurrence of transcription-associated mutation events. In vivo assays supported that at least one of these ssDNA sites is a mutational hot spot under denitrification conditions. Similar benefits from nitrogen fixation were observed when plants were inoculated with the commercial inoculant B399 and strains INTA4–6, suggesting that the low-N2O-emitting rhizobia can be an ecological alternative to the current inoculants without resigning economic profitability. |
| format | info:ar-repo/semantics/artículo |
| id | INTA7665 |
| institution | Instituto Nacional de Tecnología Agropecuaria (INTA -Argentina) |
| language | Inglés |
| publishDate | 2020 |
| publishDateRange | 2020 |
| publishDateSort | 2020 |
| publisher | Springer |
| publisherStr | Springer |
| record_format | dspace |
| spelling | INTA76652020-08-04T11:44:25Z Spontaneous Mutations in the Nitrate Reductase Gene napC Drive the Emergence of Eco-friendly Low-N2O-Emitting Alfalfa Rhizobia in Regions with Different Climates Brambilla, Silvina Maricel Soto, Gabriela Odorizzi, Ariel Arolfo, Valeria McCormick, Wayne Primo, Emiliano Giordano, Walter Jozefkowicz, Cintia Ayub, Nicolás Daniel Medicago sativa Óxido Nitroso Inoculación Nitrato Reductasa Rhizobiaceae Factores Climáticos Nitrous Oxide Inoculation Nitrate Reductase Genes Climatic Factors Alfalfa Sinorhizobium meliloti Lucerne We have recently shown that commercial alfalfa inoculants (e.g., Sinorhizobium meliloti B399), which are closely related to the denitrifier model strain Sinorhizobium meliloti 1021, have conserved nitrate, nitrite, and nitric oxide reductases associated with the production of the greenhouse gas nitrous oxide (N2O) from nitrate but lost the N2O reductase related to the degradation of N2O to gas nitrogen. Here, we screened a library of nitrogen-fixing alfalfa symbionts originating from different ecoregions and containing N2O reductase genes and identified novel rhizobia (Sinorhizobium meliloti INTA1–6) exhibiting exceptionally low N2O emissions. To understand the genetic basis of this novel eco-friendly phenotype, we sequenced and analyzed the genomes of these strains, focusing on their denitrification genes, and found mutations only in the nitrate reductase structural gene napC. The evolutionary analysis supported that, in these natural strains, the denitrification genes were inherited by vertical transfer and that their defective nitrate reductase napC alleles emerged by independent spontaneous mutations. In silico analyses showed that mutations in this gene occurred in ssDNA loop structures with high negative free energy (−ΔG) and that the resulting mutated stem-loop structures exhibited increased stability, suggesting the occurrence of transcription-associated mutation events. In vivo assays supported that at least one of these ssDNA sites is a mutational hot spot under denitrification conditions. Similar benefits from nitrogen fixation were observed when plants were inoculated with the commercial inoculant B399 and strains INTA4–6, suggesting that the low-N2O-emitting rhizobia can be an ecological alternative to the current inoculants without resigning economic profitability. Instituto de Biotecnología Fil: Brambilla, Silvina Maricel. Instituto Nacional de Tecnología Agropecuaria (INTA). Instituto de Agrobiotecnología y Biología Molecular e Instituto de Genética; Argentina. Fil: Soto, Gabriela Cinthia. Instituto Nacional de Tecnología Agropecuaria (INTA). Instituto de Agrobiotecnología y Biología Molecular e Instituto de Genética; Argentina. Fil: Odorizzi, Ariel. Instituto Nacional de Tecnología Agropecuaria (INTA). Estación Experimental Agropecuaria Manfredi; Argentina Fil: Arolfo, Valeria. Instituto Nacional de Tecnología Agropecuaria (INTA). Estación Experimental Agropecuaria Manfredi; Argentina Fil: McCormick, Wayne. Ottawa Research and Development Centre; Canadá Fil: Primo, Emiliano. Universidad Nacional de Río Cuarto. Departamento de Biología Molecular; Argentina Fil: Giordano, Walter. Universidad Nacional de Río Cuarto. Departamento de Biología Molecular; Argentina Fil: Jozefkowicz, Cintia . Instituto Nacional de Tecnología Agropecuaria (INTA). Instituto de Agrobiotecnología y Biología Molecular e Instituto de Genética; Argentina. Fil: Ayub, Nicolás Daniel. Instituto Nacional de Tecnología Agropecuaria (INTA). Instituto de Agrobiotecnología y Biología Molecular e Instituto de Genética; Argentin 2020-08-04T11:39:43Z 2020-08-04T11:39:43Z 2020 info:ar-repo/semantics/artículo info:eu-repo/semantics/article info:eu-repo/semantics/publishedVersion http://hdl.handle.net/20.500.12123/7665 https://link.springer.com/article/10.1007/s00248-019-01473-w 0095-3628 1432-184X https://doi.org/10.1007/s00248-019-01473-w eng info:eu-repo/semantics/restrictedAccess application/pdf Springer Microbial Ecology 79 : 1044–1053 (2020) |
| spellingShingle | Medicago sativa Óxido Nitroso Inoculación Nitrato Reductasa Rhizobiaceae Factores Climáticos Nitrous Oxide Inoculation Nitrate Reductase Genes Climatic Factors Alfalfa Sinorhizobium meliloti Lucerne Brambilla, Silvina Maricel Soto, Gabriela Odorizzi, Ariel Arolfo, Valeria McCormick, Wayne Primo, Emiliano Giordano, Walter Jozefkowicz, Cintia Ayub, Nicolás Daniel Spontaneous Mutations in the Nitrate Reductase Gene napC Drive the Emergence of Eco-friendly Low-N2O-Emitting Alfalfa Rhizobia in Regions with Different Climates |
| title | Spontaneous Mutations in the Nitrate Reductase Gene napC Drive the Emergence of Eco-friendly Low-N2O-Emitting Alfalfa Rhizobia in Regions with Different Climates |
| title_full | Spontaneous Mutations in the Nitrate Reductase Gene napC Drive the Emergence of Eco-friendly Low-N2O-Emitting Alfalfa Rhizobia in Regions with Different Climates |
| title_fullStr | Spontaneous Mutations in the Nitrate Reductase Gene napC Drive the Emergence of Eco-friendly Low-N2O-Emitting Alfalfa Rhizobia in Regions with Different Climates |
| title_full_unstemmed | Spontaneous Mutations in the Nitrate Reductase Gene napC Drive the Emergence of Eco-friendly Low-N2O-Emitting Alfalfa Rhizobia in Regions with Different Climates |
| title_short | Spontaneous Mutations in the Nitrate Reductase Gene napC Drive the Emergence of Eco-friendly Low-N2O-Emitting Alfalfa Rhizobia in Regions with Different Climates |
| title_sort | spontaneous mutations in the nitrate reductase gene napc drive the emergence of eco friendly low n2o emitting alfalfa rhizobia in regions with different climates |
| topic | Medicago sativa Óxido Nitroso Inoculación Nitrato Reductasa Rhizobiaceae Factores Climáticos Nitrous Oxide Inoculation Nitrate Reductase Genes Climatic Factors Alfalfa Sinorhizobium meliloti Lucerne |
| url | http://hdl.handle.net/20.500.12123/7665 https://link.springer.com/article/10.1007/s00248-019-01473-w https://doi.org/10.1007/s00248-019-01473-w |
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