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- Laeverenz Schlogelhofer, Hannah, et al.
(författare)
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Combining SIMS and mechanistic modelling to reveal nutrient kinetics in an algal-bacterial mutualism
- 2021
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Ingår i: PLOS ONE. - : Public Library of Science (PLoS). - 1932-6203. ; 16:5
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Tidskriftsartikel (refereegranskat)abstract
- Microbial communities are of considerable significance for biogeochemical processes, for the health of both animals and plants, and for biotechnological purposes. A key feature of microbial interactions is the exchange of nutrients between cells. Isotope labelling followed by analysis with secondary ion mass spectrometry (SIMS) can identify nutrient fluxes and heterogeneity of substrate utilisation on a single cell level. Here we present a novel approach that combines SIMS experiments with mechanistic modelling to reveal otherwise inaccessible nutrient kinetics. The method is applied to study the onset of a synthetic mutualistic partnership between a vitamin B-12-dependent mutant of the alga Chlamydomonas reinhardtii and the B-12-producing, heterotrophic bacterium Mesorhizobium japonicum, which is supported by algal photosynthesis. Results suggest that an initial pool of fixed carbon delays the onset of mutualistic cross-feeding; significantly, our approach allows the first quantification of this expected delay. Our method is widely applicable to other microbial systems, and will contribute to furthering a mechanistic understanding of microbial interactions.
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| 2. |
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| 3. |
- Nieves-Morión, Mercedes, et al.
(författare)
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Single-Cell Measurements of Fixation and Intercellular Exchange of C and N in the Filaments of the HeterocystForming Cyanobacterium Anabaena sp. Strain PCC 7120
- 2021
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Ingår i: Mbio. - 2150-7511 .- 2161-2129. ; 12:4
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Tidskriftsartikel (refereegranskat)abstract
- Under diazotrophic conditions, the model filamentous, heterocystforming cyanobacterium Anabaena sp. strain PCC 7120 develops a metabolic strategy based on the physical separation of the processes of oxygenic photosynthesis, in vegetative cells, and N-2 fixation, in heterocysts. This strategy requires the exchange of carbon and nitrogen metabolites and their distribution along the filaments, which takes place through molecular diffusion via septal junctions involving FraCD proteins. Here, Anabaena was incubated in a time course (up to 20 h) with [C-13]bicarbonate and N-15(2) and analyzed by secondary ion mass spectrometry imaging (SIMS) (large-geometry SIMS [LG- SIMS] and NanoSIMS) to quantify C and N assimilation and distribution in the filaments. The C-13/C-12 and N-15/N-14 ratios measured in wild-type filaments showed a general increase with time. The enrichment was relatively homogeneous in vegetative cells along individual filaments, while it was reduced in heterocysts. Heterocysts, however, accumulated recently fixed N at their poles, in which the cyanophycin plug [multi-L-arginyl-poly(L-aspartic acid)] is located. In contrast to the rather homogeneous label found along stretches of vegetative cells, C-13/C-12 and N-15/N-14 ratios were significantly different between filaments both at the same and different time points, showing high variability in metabolic states. A fraC fraD mutant did not fix N-2, and the C-13/C-12 ratio was homogeneous along the filament, including the heterocyst in contrast to the wild type. Our results show the consumption of reduced C in the heterocysts associated with the fixation and export of fixed N and present an unpredicted heterogeneity of cellular metabolic activity in different filaments of an Anabaena culture under controlled conditions. IMPORTANCE Filamentous, heterocyst- forming cyanobacteria represent a paradigm of multicellularity in the prokaryotic world. Physiological studies at the cellular level in model organisms are crucial to understand metabolic activities and qualify specific aspects related to multicellularity. Here, we used stable isotopes (C-13 and N-15) coupled to LG-SIMS and NanoSIMS imaging to follow single-cell C and N-2 fixation and metabolic dynamics along the filaments in the model heterocyst-forming cyanobacterium Anabaena sp. strain PCC 7120. Our results show a close relationship between C and N fixation and distribution in the filaments and indicate that wild-type filaments in a culture can exhibit a substantial variability of metabolic states. This illustrates how some novel properties can be appreciated by studying microbial cultures at the single-cell level.
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| 4. |
- Pierella Karlusich, Juan José, et al.
(författare)
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Global distribution patterns of marine nitrogen-fixers by imaging and molecular methods
- 2021
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Ingår i: Nature Communications. - : Springer Science and Business Media LLC. - 2041-1723. ; 12:1
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Tidskriftsartikel (refereegranskat)abstract
- Nitrogen fixation has a critical role in marine primary production, yet our understanding of marine nitrogen-fixers (diazotrophs) is hindered by limited observations. Here, we report a quantitative image analysis pipeline combined with mapping of molecular markers for mining >2,000,000 images and >1300 metagenomes from surface, deep chlorophyll maximum and mesopelagic seawater samples across 6 size fractions (<0.2-2000m). We use this approach to characterise the diversity, abundance, biovolume and distribution of symbiotic, colony-forming and particle-associated diazotrophs at a global scale. We show that imaging and PCR-free molecular data are congruent. Sequence reads indicate diazotrophs are detected from the ultrasmall bacterioplankton (<0.2m) to mesoplankton (180-2000 mu m) communities, while images predict numerous symbiotic and colony-forming diazotrophs (>20 mu m). Using imaging and molecular data, we estimate that polyploidy can substantially affect gene abundances of symbiotic versus colony-forming diazotrophs. Our results support the canonical view that larger diazotrophs (>10 mu m) dominate the tropical belts, while unicellular cyanobacterial and non-cyanobacterial diazotrophs are globally distributed in surface and mesopelagic layers. We describe co-occurring diazotrophic lineages of different lifestyles and identify high-density regions of diazotrophs in the global ocean. Overall, we provide an update of marine diazotroph biogeographical diversity and present a new bioimaging-bioinformatic workflow. Nitrogen fixation by diazotrophs is critical for marine primary production. Using Tara Oceans datasets, this study combines a quantitative image analysis pipeline with metagenomic mining to provide an improved global overview of diazotroph abundance, diversity and distribution.
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