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1.
Nat Commun ; 10(1): 2770, 2019 06 24.
Artigo em Inglês | MEDLINE | ID: mdl-31235780

RESUMO

The ability to link soil microbial diversity to soil processes requires technologies that differentiate active microbes from extracellular DNA and dormant cells. Here, we use BONCAT (bioorthogonal non-canonical amino acid tagging) to measure translationally active cells in soils. We compare the active population of two soil depths from Oak Ridge (Tennessee, USA) and find that a maximum of 25-70% of the extractable cells are active. Analysis of 16S rRNA sequences from BONCAT-positive cells recovered by fluorescence-activated cell sorting (FACS) reveals that the phylogenetic composition of the active fraction is distinct from the total population of extractable cells. Some members of the community are found to be active at both depths independently of their abundance rank, suggesting that the incubation conditions favor the activity of similar organisms. We conclude that BONCAT-FACS is effective for interrogating the active fraction of soil microbiomes in situ and provides a new approach for uncovering the links between soil processes and specific microbial groups.


Assuntos
Bactérias/isolamento & purificação , Citometria de Fluxo/métodos , Microbiota , Microbiologia do Solo , Aminoácidos/análise , Aminoácidos/química , Bactérias/genética , Bactérias/metabolismo , Corantes Fluorescentes/química , Filogenia , RNA Ribossômico 16S/genética , RNA Ribossômico 16S/isolamento & purificação , Coloração e Rotulagem/métodos
2.
PeerJ ; 7: e6902, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31119088

RESUMO

Background: Metagenomics has transformed our understanding of microbial diversity across ecosystems, with recent advances enabling de novo assembly of genomes from metagenomes. These metagenome-assembled genomes are critical to provide ecological, evolutionary, and metabolic context for all the microbes and viruses yet to be cultivated. Metagenomes can now be generated from nanogram to subnanogram amounts of DNA. However, these libraries require several rounds of PCR amplification before sequencing, and recent data suggest these typically yield smaller and more fragmented assemblies than regular metagenomes. Methods: Here we evaluate de novo assembly methods of 169 PCR-amplified metagenomes, including 25 for which an unamplified counterpart is available, to optimize specific assembly approaches for PCR-amplified libraries. We first evaluated coverage bias by mapping reads from PCR-amplified metagenomes onto reference contigs obtained from unamplified metagenomes of the same samples. Then, we compared different assembly pipelines in terms of assembly size (number of bp in contigs ≥ 10 kb) and error rates to evaluate which are the best suited for PCR-amplified metagenomes. Results: Read mapping analyses revealed that the depth of coverage within individual genomes is significantly more uneven in PCR-amplified datasets versus unamplified metagenomes, with regions of high depth of coverage enriched in short inserts. This enrichment scales with the number of PCR cycles performed, and is presumably due to preferential amplification of short inserts. Standard assembly pipelines are confounded by this type of coverage unevenness, so we evaluated other assembly options to mitigate these issues. We found that a pipeline combining read deduplication and an assembly algorithm originally designed to recover genomes from libraries generated after whole genome amplification (single-cell SPAdes) frequently improved assembly of contigs ≥10 kb by 10 to 100-fold for low input metagenomes. Conclusions: PCR-amplified metagenomes have enabled scientists to explore communities traditionally challenging to describe, including some with extremely low biomass or from which DNA is particularly difficult to extract. Here we show that a modified assembly pipeline can lead to an improved de novo genome assembly from PCR-amplified datasets, and enables a better genome recovery from low input metagenomes.

3.
Microbiome ; 7(1): 55, 2019 04 03.
Artigo em Inglês | MEDLINE | ID: mdl-30944036

RESUMO

BACKGROUND: Biological soil crusts (biocrusts) are a key component of arid land ecosystems, where they render critical services such as soil surface stabilization and nutrient fertilization. The bundle-forming, filamentous, non-nitrogen-fixing cyanobacterium Microcoleus vaginatus is a pioneer primary producer, often the dominant member of the biocrust microbiome, and the main source of leaked organic carbon. We hypothesized that, by analogy to the rhizosphere of plant roots, M. vaginatus may shape the microbial populations of heterotrophs around it, forming a specialized cyanosphere. RESULTS: By physically isolating bundles of M. vaginatus from biocrusts, we were able to study the composition of the microbial populations attached to it, in comparison to the bulk soil crust microbiome by means of high-throughput 16S rRNA sequencing. We did this in two M. vaginatus-dominated biocrust from distinct desert biomes. We found that a small, selected subset of OTUs was significantly enriched in close proximity to M. vaginatus. Furthermore, we also found that a majority of bacteria (corresponding to some two thirds of the reads) were significantly more abundant away from this cyanobacterium. Phylogenetic placements suggest that all typical members of the cyanosphere were copiotrophs and that many were diazotrophs (Additional file 1: Tables S2 and S3). Nitrogen fixation genes were in fact orders of magnitude more abundant in this cyanosphere than in the bulk biocrust soil as assessed by qPCR. By contrary, competition for light, CO2, and low organic carbon concentrations defined at least a part of the OTUs segregating from the cyanobacterium. CONCLUSIONS: We showed that M. vaginatus acts as a significant spatial organizer of the biocrust microbiome. On the one hand, it possesses a compositionally differentiated cyanosphere that concentrates the nitrogen-fixing function. We propose that a mutualism based on C for N exchange between M. vaginatus and copiotrophic diazotrophs helps sustains this cyanosphere and that this consortium constitutes the true pioneer community enabling the colonization of nitrogen-poor soils. On the other hand, a large number of biocrust community members segregate away from the vicinity of M. vaginatus, potentially through competition for light or CO2, or because of a preference for oligotrophy.


Assuntos
Cianobactérias/classificação , Fixação de Nitrogênio , RNA Ribossômico 16S/genética , Cianobactérias/genética , Cianobactérias/isolamento & purificação , DNA Ribossômico/genética , Clima Desértico , Filogenia , Microbiologia do Solo , Simbiose
4.
Nat Commun ; 9(1): 2530, 2018 06 28.
Artigo em Inglês | MEDLINE | ID: mdl-29955055

RESUMO

While significant efforts have been invested in reconstructing the early evolution of the Earth's atmosphere-ocean-biosphere biogeochemical nitrogen cycle, the potential role of an early continental contribution by a terrestrial, microbial phototrophic biosphere has been largely overlooked. By transposing to the Archean nitrogen fluxes of modern topsoil communities known as biological soil crusts (terrestrial analogs of microbial mats), whose ancestors might have existed as far back as 3.2 Ga ago, we show that they could have impacted the evolution of the nitrogen cycle early on. We calculate that the net output of inorganic nitrogen reaching the Precambrian hydrogeological system could have been of the same order of magnitude as that of modern continents for a range of inhabited area as small as a few percent of that of present day continents. This contradicts the assumption that before the Great Oxidation Event, marine and continental biogeochemical nitrogen cycles were disconnected.


Assuntos
Consórcios Microbianos/fisiologia , Ciclo do Nitrogênio/fisiologia , Nitrogênio/química , Processos Fototróficos/fisiologia , Ecossistema , História Antiga , Consórcios Microbianos/efeitos da radiação , Nitrogênio/história , Nitrogênio/metabolismo , Isótopos de Nitrogênio , Oceanos e Mares , Origem da Vida , Oxirredução , Oxigênio/química , Oxigênio/história , Oxigênio/metabolismo , Processos Fototróficos/efeitos da radiação , Solo/química , Luz Solar
5.
mBio ; 9(2)2018 03 06.
Artigo em Inglês | MEDLINE | ID: mdl-29511079

RESUMO

Biological soil crusts (biocrusts) account for a substantial portion of primary production in dryland ecosystems. They successionally mature to deliver a suite of ecosystem services, such as carbon sequestration, water retention and nutrient cycling, and climate regulation. Biocrust assemblages are extremely well adapted to survive desiccation and to rapidly take advantage of the periodic precipitation events typical of arid ecosystems. Here we focus on the wetting response of incipient cyanobacterial crusts as they mature from "light" to "dark." We sampled a cyanobacterial biocrust chronosequence before (dry) and temporally following a controlled wetting event and used high-throughput 16S rRNA and rRNA gene sequencing to monitor the dynamics of microbial response. Overall, shorter-term changes in phylogenetic beta diversity attributable to periodic wetting were as large as those attributable to biocrust successional stage. Notably, more mature crusts showed significantly higher resistance to precipitation disturbance. A large bloom of a few taxa within the Firmicutes, primarily in the order Bacillales, emerged 18 h after wetting, while filamentous crust-forming cyanobacteria showed variable responses to wet-up across the successional gradient, with populations collapsing in less-developed light crusts but increasing in later-successional-stage dark crusts. Overall, the consistent Bacillales bloom accompanied by the variable collapse of pioneer cyanobacteria of the Oscillatoriales order across the successional gradient suggests that the strong response of few organisms to a hydration pulse with the mortality of the autotroph might have important implications for carbon (C) balance in semiarid ecosystems.IMPORTANCE Desert biological soil crusts are terrestrial topsoil microbial communities common to arid regions that comprise 40% of Earth's terrestrial surface. They successionally develop over years to decades to deliver a suite of ecosystem services of local and global significance. Ecosystem succession toward maturity has been associated with both resistance and resilience to disturbance. Recent work has shown that the impacts of both climate change and physical disturbance on biocrusts increase the potential for successional resetting. A larger proportion of biocrusts are expected to be at an early developmental stage, hence increasing susceptibility to changes in precipitation frequencies. Therefore, it is essential to characterize how biocrusts respond to wetting across early developmental stages. In this study, we document the wetting response of microbial communities from a biocrust chronosequence. Overall, our results suggest that the cumulative effects of altered precipitation frequencies on the stability of biocrusts will depend on biocrust maturity.


Assuntos
Bacillales/fisiologia , Bacillales/genética , Ecossistema , Firmicutes/genética , Firmicutes/fisiologia , RNA Ribossômico 16S/genética , Microbiologia do Solo
6.
Appl Environ Microbiol ; 84(4)2018 02 15.
Artigo em Inglês | MEDLINE | ID: mdl-29222097

RESUMO

Anoxygenic phototrophic bacteria (APBs) occur in a wide range of aquatic habitats, from hot springs to freshwater lakes and intertidal microbial mats. Here, we report the discovery of a novel niche for APBs: endoliths within marine littoral carbonates. In a study of 40 locations around Isla de Mona, Puerto Rico, and Menorca, Spain, 16S rRNA high-throughput sequencing of endolithic community DNA revealed the presence of abundant phylotypes potentially belonging to well-known APB clades. An ad hoc phylogenetic classification of these sequences enabled us to refine the assignments more stringently. Even then, all locations contained such putative APBs, often reaching a significant proportion of all phototrophic sequences. In fact, in some 20% of samples, their contribution exceeded that of oxygenic phototrophs, previously regarded as the major type of endolithic microbe in carbonates. The communities contained representatives of APBs in the Chloroflexales, various proteobacterial groups, and Chlorobi The most abundant phylotypes varied with geography: on Isla de Mona, Roseiflexus and Chlorothrix-related phylotypes dominated, whereas those related to Erythrobacter were the most common in Menorca. The presence of active populations of APBs was corroborated through an analysis of photopigments: bacteriochlorophylls were detected in all samples, bacteriochlorophyll c and a being most abundant. We discuss the potential metabolism and geomicrobial roles of endolithic APBs. Phylogenetic inference suggests that APBs may be playing a role as photoheterotrophs, adding biogeochemical complexity to our understanding of such communities. Given the global extent of coastal carbonate platforms, they likely represent a very large and unexplored habitat for APBs.IMPORTANCE Endolithic microbial communities from carbonates, which have been explored for over 2 centuries in predominantly naturalistic studies, were thought to be primarily composed of eukaryotic algae and cyanobacteria. Our report represents a paradigm shift in this regard, at least for the marine environment, demonstrating the presence of ubiquitous and abundant populations of APBs in this habitat. It raises questions about the role of these organisms in the geological dynamics of coastal carbonates, including coral reefs.


Assuntos
Bactérias Anaeróbias/metabolismo , Microbiota/genética , Fotossíntese/fisiologia , Processos Fototróficos , Anaerobiose , Bactérias Anaeróbias/genética , Proteínas de Bactérias , Bacterioclorofilas , Carbonatos , Chloroflexi/genética , Clorófitas/genética , Recifes de Corais , Cianobactérias/genética , Cianobactérias/isolamento & purificação , Cianobactérias/metabolismo , Água Doce , Microbiota/fisiologia , Filogenia , RNA Ribossômico 16S
7.
Int J Syst Evol Microbiol ; 67(3): 653-658, 2017 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-27902306

RESUMO

A unicellular cyanobacterium, strain Alchichica-D10, was isolated from microbialites of the alkaline Lake Alchichica, Mexico. The cells were short rods (3.9±0.6 µm in length and 1.1±0.1 µm in width) forming biofilms of intense emerald green colour. They exhibited red autofluorescence under UV light excitation. UV-visible absorption spectra revealed that they contain chlorophyll a and phycocyanin, and electron microscopy showed the presence of thylakoids. The strain grew within a temperature range of 15-30 °C. Genomic DNA G+C content was 52.2 mol%. The most remarkable feature of this species was its granular cytoplasm, due to the presence of numerous intracellular spherical granules (16-26 per cell) with an average diameter of 270 nm. These granules, easily visible under scanning electron microscopy, were composed of amorphous carbonate containing Ca, Mg, Ba and Sr. A multi-gene phylogeny based on the analysis of 59 conserved protein markers supported robustly that this strain occupies a deep position in the cyanobacterial tree. Based on its phenotypic characters and phylogenetic position, strain Alchichica-D10 is considered to represent a new genus and novel species of cyanobacteria for which the name Gloeomargarita lithophora gen. nov., sp. nov. is proposed. The type strain is Alchichica-D10 (Culture Collection of Algae and Protozoa CCAP strain 1437/1; Collections de Cyanobactéries et Microalgues Vivantes of the Museum National d'Histoire Naturelle in Paris strain PMC 919.15). Furthermore, a new family, Gloeomargaritaceae, and a new order, Gloeoemargaritales, are proposed to accommodate this species under the International Code of Nomenclature for algae, fungi and plants.


Assuntos
Cianobactérias/classificação , Lagos/microbiologia , Filogenia , Técnicas de Tipagem Bacteriana , Composição de Bases , Carbonatos/química , Clorofila/química , Clorofila A , Cianobactérias/genética , Cianobactérias/isolamento & purificação , DNA Bacteriano/genética , México , Ficocianina/química , RNA Ribossômico 16S/genética , Análise de Sequência de DNA , Tilacoides
8.
Nat Commun ; 7: 10373, 2016 Jan 20.
Artigo em Inglês | MEDLINE | ID: mdl-26785770

RESUMO

Soil surface temperature, an important driver of terrestrial biogeochemical processes, depends strongly on soil albedo, which can be significantly modified by factors such as plant cover. In sparsely vegetated lands, the soil surface can be colonized by photosynthetic microbes that build biocrust communities. Here we use concurrent physical, biochemical and microbiological analyses to show that mature biocrusts can increase surface soil temperature by as much as 10 °C through the accumulation of large quantities of a secondary metabolite, the microbial sunscreen scytonemin, produced by a group of late-successional cyanobacteria. Scytonemin accumulation decreases soil albedo significantly. Such localized warming has apparent and immediate consequences for the soil microbiome, inducing the replacement of thermosensitive bacterial species with more thermotolerant forms. These results reveal that not only vegetation but also microorganisms are a factor in modifying terrestrial albedo, potentially impacting biosphere feedbacks on past and future climate, and call for a direct assessment of such effects at larger scales.


Assuntos
Microbiologia do Solo , Solo/química , Bactérias , Cianobactérias/fisiologia , Temperatura Ambiente
9.
Proc Natl Acad Sci U S A ; 111(30): 10933-8, 2014 Jul 29.
Artigo em Inglês | MEDLINE | ID: mdl-25009182

RESUMO

Cyanobacteria have played a significant role in the formation of past and modern carbonate deposits at the surface of the Earth using a biomineralization process that has been almost systematically considered induced and extracellular. Recently, a deep-branching cyanobacterial species, Candidatus Gloeomargarita lithophora, was reported to form intracellular amorphous Ca-rich carbonates. However, the significance and diversity of the cyanobacteria in which intracellular biomineralization occurs remain unknown. Here, we searched for intracellular Ca-carbonate inclusions in 68 cyanobacterial strains distributed throughout the phylogenetic tree of cyanobacteria. We discovered that diverse unicellular cyanobacterial taxa form intracellular amorphous Ca-carbonates with at least two different distribution patterns, suggesting the existence of at least two distinct mechanisms of biomineralization: (i) one with Ca-carbonate inclusions scattered within the cell cytoplasm such as in Ca. G. lithophora, and (ii) another one observed in strains belonging to the Thermosynechococcus elongatus BP-1 lineage, in which Ca-carbonate inclusions lie at the cell poles. This pattern seems to be linked with the nucleation of the inclusions at the septum of the cells, showing an intricate and original connection between cell division and biomineralization. These findings indicate that intracellular Ca-carbonate biomineralization by cyanobacteria has been overlooked by past studies and open new perspectives on the mechanisms and the evolutionary history of intra- and extracellular Ca-carbonate biomineralization by cyanobacteria.


Assuntos
Carbonato de Cálcio/metabolismo , Cianobactérias/metabolismo , Citoplasma/metabolismo , Corpos de Inclusão/metabolismo , Sequência de Bases , Cianobactérias/classificação , Cianobactérias/genética , Citoplasma/genética , Corpos de Inclusão/genética , Dados de Sequência Molecular
10.
ISME J ; 7(10): 1997-2009, 2013 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-23804151

RESUMO

The role of microorganisms in microbialite formation remains unresolved: do they induce mineral precipitation (microbes first) or do they colonize and/or entrap abiotic mineral precipitates (minerals first)? Does this role vary from one species to another? And what is the impact of mineral precipitation on microbial ecology? To explore potential biogenic carbonate precipitation, we studied cyanobacteria-carbonate assemblages in modern hydromagnesite-dominated microbialites from the alkaline Lake Alchichica (Mexico), by coupling three-dimensional imaging of molecular fluorescence emitted by microorganisms, using confocal laser scanning microscopy, and Raman scattering/spectrometry from the associated minerals at a microscale level. Both hydromagnesite and aragonite precipitate within a complex biofilm composed of photosynthetic and other microorganisms. Morphology and pigment-content analysis of dominant photosynthetic microorganisms revealed up to six different cyanobacterial morphotypes belonging to Oscillatoriales, Chroococcales, Nostocales and Pleurocapsales, as well as several diatoms and other eukaryotic microalgae. Interestingly, one of these morphotypes, Pleurocapsa-like, appeared specifically associated with aragonite minerals, the oldest parts of actively growing Pleurocapsa-like colonies being always aragonite-encrusted. We hypothesize that actively growing cells of Pleurocapsales modify local environmental conditions favoring aragonite precipitation at the expense of hydromagnesite, which precipitates at seemingly random locations within the biofilm. Therefore, at least part of the mineral precipitation in Alchichica microbialites is most likely biogenic and the type of biominerals formed depends on the nature of the phylogenetic lineage involved. This observation may provide clues to identify lineage-specific biosignatures in fossil stromatolites from modern to Precambrian times.


Assuntos
Carbonatos/metabolismo , Cianobactérias/metabolismo , Lagos/microbiologia , Biofilmes , Carbonatos/análise , Carbonatos/química , Cianobactérias/classificação , Cianobactérias/ultraestrutura , México , Microscopia Confocal , Filogenia , Pigmentos Biológicos/química , Análise Espectral Raman
11.
Science ; 336(6080): 459-62, 2012 Apr 27.
Artigo em Inglês | MEDLINE | ID: mdl-22539718

RESUMO

Cyanobacteria have affected major geochemical cycles (carbon, nitrogen, and oxygen) on Earth for billions of years. In particular, they have played a major role in the formation of calcium carbonates (i.e., calcification), which has been considered to be an extracellular process. We identified a cyanobacterium in modern microbialites in Lake Alchichica (Mexico) that forms intracellular amorphous calcium-magnesium-strontium-barium carbonate inclusions about 270 nanometers in average diameter, revealing an unexplored pathway for calcification. Phylogenetic analyses place this cyanobacterium within the deeply divergent order Gloeobacterales. The chemical composition and structure of the intracellular precipitates suggest some level of cellular control on the biomineralization process. This discovery expands the diversity of organisms capable of forming amorphous calcium carbonates.


Assuntos
Biofilmes , Carbonato de Cálcio/análise , Carbonatos/análise , Cianobactérias/isolamento & purificação , Cianobactérias/fisiologia , Corpos de Inclusão/química , Corpos de Inclusão/ultraestrutura , Lagos/microbiologia , Bário/análise , Sequência de Bases , Calcificação Fisiológica , Cálcio/análise , Carbonatos/metabolismo , Precipitação Química , Cianobactérias/classificação , Cianobactérias/ultraestrutura , Genes Bacterianos , Genes de RNAr , Magnésio/análise , México , Dados de Sequência Molecular , Filogenia , Estrôncio/análise
12.
PLoS One ; 6(12): e28767, 2011.
Artigo em Inglês | MEDLINE | ID: mdl-22194908

RESUMO

The geomicrobiology of crater lake microbialites remains largely unknown despite their evolutionary interest due to their resemblance to some Archaean analogs in the dominance of in situ carbonate precipitation over accretion. Here, we studied the diversity of archaea, bacteria and protists in microbialites of the alkaline Lake Alchichica from both field samples collected along a depth gradient (0-14 m depth) and long-term-maintained laboratory aquaria. Using small subunit (SSU) rRNA gene libraries and fingerprinting methods, we detected a wide diversity of bacteria and protists contrasting with a minor fraction of archaea. Oxygenic photosynthesizers were dominated by cyanobacteria, green algae and diatoms. Cyanobacterial diversity varied with depth, Oscillatoriales dominating shallow and intermediate microbialites and Pleurocapsales the deepest samples. The early-branching Gloeobacterales represented significant proportions in aquaria microbialites. Anoxygenic photosynthesizers were also diverse, comprising members of Alphaproteobacteria and Chloroflexi. Although photosynthetic microorganisms dominated in biomass, heterotrophic lineages were more diverse. We detected members of up to 21 bacterial phyla or candidate divisions, including lineages possibly involved in microbialite formation, such as sulfate-reducing Deltaproteobacteria but also Firmicutes and very diverse taxa likely able to degrade complex polymeric substances, such as Planctomycetales, Bacteroidetes and Verrucomicrobia. Heterotrophic eukaryotes were dominated by Fungi (including members of the basal Rozellida or Cryptomycota), Choanoflagellida, Nucleariida, Amoebozoa, Alveolata and Stramenopiles. The diversity and relative abundance of many eukaryotic lineages suggest an unforeseen role for protists in microbialite ecology. Many lineages from lake microbialites were successfully maintained in aquaria. Interestingly, the diversity detected in aquarium microbialites was higher than in field samples, possibly due to more stable and favorable laboratory conditions. The maintenance of highly diverse natural microbialites in laboratory aquaria holds promise to study the role of different metabolisms in the formation of these structures under controlled conditions.


Assuntos
Álcalis/metabolismo , Células Eucarióticas/metabolismo , Sedimentos Geológicos/microbiologia , Lagos/microbiologia , Células Procarióticas/metabolismo , Sequência de Bases , Biodiversidade , Biofilmes , Análise por Conglomerados , Impressões Digitais de DNA , DNA Ribossômico/genética , Eletroforese em Gel de Gradiente Desnaturante , Genes de RNAr , Funções Verossimilhança , México , Dados de Sequência Molecular , Fotossíntese , Filogenia , Análise de Sequência de DNA
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