Label-free functional analysis of root-associated microbes with dynamic quantitative oblique back-illumination microscopy
Abstract The increasing global demand for food, coupled with concerns about the environmental impact of synthetic fertilizers, underscores the urgency of developing sustainable agricultural practices. Nitrogen-fixing bacteria, known as diazotrophs, offer a potential solution by converting atmospheri...
| Main Authors: | , , , , , , |
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| Format: | Article |
| Language: | English |
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Nature Portfolio
2024-03-01
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| Series: | Scientific Reports |
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| Online Access: | https://doi.org/10.1038/s41598-024-56443-1 |
| _version_ | 1797266815168020480 |
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| author | Caroline Filan Madison Green Abigail Diering Marcus T. Cicerone Lily S. Cheung Joel E. Kostka Francisco E. Robles |
| author_facet | Caroline Filan Madison Green Abigail Diering Marcus T. Cicerone Lily S. Cheung Joel E. Kostka Francisco E. Robles |
| author_sort | Caroline Filan |
| collection | DOAJ |
| description | Abstract The increasing global demand for food, coupled with concerns about the environmental impact of synthetic fertilizers, underscores the urgency of developing sustainable agricultural practices. Nitrogen-fixing bacteria, known as diazotrophs, offer a potential solution by converting atmospheric nitrogen into bioavailable forms, reducing the reliance on synthetic fertilizers. However, a deeper understanding of their interactions with plants and other microbes is needed. In this study, we introduce a recently developed label-free 3D quantitative phase imaging technology called dynamic quantitative oblique back-illumination microscopy (DqOBM) to assess the functional dynamic activity of diazotrophs in vitro and in situ. Our experiments involved three different diazotrophs (Sinorhizobium meliloti, Azotobacter vinelandii, and Rahnella aquatilis) cultured on media with amendments of carbon and nitrogen sources. Over 5 days, we observed increased dynamics in nutrient-amended media. These results suggest that the observed bacterial dynamics correlate with their metabolic activity. Furthermore, we applied qOBM to visualize microbial dynamics within the root cap and elongation zone of Arabidopsis thaliana primary roots. This allowed us to identify distinct areas of microbial infiltration in plant roots without the need for fluorescent markers. Our findings demonstrate that DqOBM can effectively characterize microbial dynamics and provide insights into plant-microbe interactions in situ, offering a valuable tool for advancing our understanding of sustainable agriculture. |
| first_indexed | 2024-04-25T01:06:41Z |
| format | Article |
| id | doaj.art-023de71c2a4c46429569a51fd98e4786 |
| institution | Directory Open Access Journal |
| issn | 2045-2322 |
| language | English |
| last_indexed | 2024-04-25T01:06:41Z |
| publishDate | 2024-03-01 |
| publisher | Nature Portfolio |
| record_format | Article |
| series | Scientific Reports |
| spelling | doaj.art-023de71c2a4c46429569a51fd98e47862024-03-10T12:11:38ZengNature PortfolioScientific Reports2045-23222024-03-0114111310.1038/s41598-024-56443-1Label-free functional analysis of root-associated microbes with dynamic quantitative oblique back-illumination microscopyCaroline Filan0Madison Green1Abigail Diering2Marcus T. Cicerone3Lily S. Cheung4Joel E. Kostka5Francisco E. Robles6George W. Woodruff School of Mechanical Engineering, Georgia Institute of TechnologySchool of Biological Sciences, Georgia Institute of TechnologySchool of Chemistry and Biochemistry, Georgia Institute of TechnologySchool of Chemistry and Biochemistry, Georgia Institute of TechnologySchool of Chemical and Biomolecular Engineering, Georgia Institute of TechnologySchool of Biological Sciences, Georgia Institute of TechnologyWallace H. Coulter Department of Biomedical Engineering, Georgia Institute of Technology and Emory UniversityAbstract The increasing global demand for food, coupled with concerns about the environmental impact of synthetic fertilizers, underscores the urgency of developing sustainable agricultural practices. Nitrogen-fixing bacteria, known as diazotrophs, offer a potential solution by converting atmospheric nitrogen into bioavailable forms, reducing the reliance on synthetic fertilizers. However, a deeper understanding of their interactions with plants and other microbes is needed. In this study, we introduce a recently developed label-free 3D quantitative phase imaging technology called dynamic quantitative oblique back-illumination microscopy (DqOBM) to assess the functional dynamic activity of diazotrophs in vitro and in situ. Our experiments involved three different diazotrophs (Sinorhizobium meliloti, Azotobacter vinelandii, and Rahnella aquatilis) cultured on media with amendments of carbon and nitrogen sources. Over 5 days, we observed increased dynamics in nutrient-amended media. These results suggest that the observed bacterial dynamics correlate with their metabolic activity. Furthermore, we applied qOBM to visualize microbial dynamics within the root cap and elongation zone of Arabidopsis thaliana primary roots. This allowed us to identify distinct areas of microbial infiltration in plant roots without the need for fluorescent markers. Our findings demonstrate that DqOBM can effectively characterize microbial dynamics and provide insights into plant-microbe interactions in situ, offering a valuable tool for advancing our understanding of sustainable agriculture.https://doi.org/10.1038/s41598-024-56443-1Label-free imagingNitrogen fixationQuantitative phase imaging |
| spellingShingle | Caroline Filan Madison Green Abigail Diering Marcus T. Cicerone Lily S. Cheung Joel E. Kostka Francisco E. Robles Label-free functional analysis of root-associated microbes with dynamic quantitative oblique back-illumination microscopy Scientific Reports Label-free imaging Nitrogen fixation Quantitative phase imaging |
| title | Label-free functional analysis of root-associated microbes with dynamic quantitative oblique back-illumination microscopy |
| title_full | Label-free functional analysis of root-associated microbes with dynamic quantitative oblique back-illumination microscopy |
| title_fullStr | Label-free functional analysis of root-associated microbes with dynamic quantitative oblique back-illumination microscopy |
| title_full_unstemmed | Label-free functional analysis of root-associated microbes with dynamic quantitative oblique back-illumination microscopy |
| title_short | Label-free functional analysis of root-associated microbes with dynamic quantitative oblique back-illumination microscopy |
| title_sort | label free functional analysis of root associated microbes with dynamic quantitative oblique back illumination microscopy |
| topic | Label-free imaging Nitrogen fixation Quantitative phase imaging |
| url | https://doi.org/10.1038/s41598-024-56443-1 |
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