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Exploration of oxygen-depleted marine environments has consistently revealed novel microbial taxa and metabolic capabilities that expand our understanding of microbial evolution and ecology. Marine blue holes are shallow karst formations characterized by low oxygen and high organic matter content. They are logistically challenging to sample, and thus our understanding of their biogeochemistry and microbial ecology is limited. We present a metagenomic characterization of Amberjack Hole on the Florida continental shelf (Gulf of Mexico). Dissolved oxygen became depleted at the hole’s rim (32 m water depth), remained low but detectable in an intermediate hypoxic zone (40-75 m), and then increased to a secondary peak before falling below detection in the bottom layer (80-110 m), concomitant with increases in nutrients, dissolved iron, and a series of sequentially more reduced sulfur species. Microbial communities in the bottom layer contained heretofore undocumented levels of the recently discovered phylum Woesearchaeota (up to 58% of the community), along with lineages in the bacterial Candidate Phyla Radiation (CPR). Thirty-one high-quality metagenome-assembled genomes (MAGs) showed extensive biochemical capabilities for sulfur and nitrogen cycling, as well as for resisting and respiring arsenic. One uncharacterized gene associated with a CPR lineage differentiated hypoxic from anoxic zone communities. Overall, microbial communities and geochemical profiles were stable across two sampling dates in the spring and fall of 2019. The blue hole habitat is a natural marine laboratory that provides opportunities for sampling taxa with under-characterized but potentially important roles in redox-stratified microbial processes.

International audience; Pore water and solid phase geochemical profiles of sediment cores collected along two transects on the western and eastern sides of the Mississippi River mouth in the northern Gulf of Mexico were incorporated into a reactive transport model to determine the role of manganese and iron in the remineralization of carbon. Reactive transport model calculations indicate that sedimentation rates control the intensity of anaerobic carbon remineralization and select for the dominant anaerobic carbon remineralization pathways. Although sulfate reduction dominates the shelf station (65 m water depth), denitrification and microbial manganese reduction appear equally significant anaerobic respiration processes along the continental slope the closest to the Mississippi River, whereas microbial iron reduction does not represent an important process in these sediments. These findings suggest that the differential kinetics of manganese and iron redox transformations influence carbon remineralization processes on the continental slope. The fast kinetics of Fe2+ oxidation near the sediment-water interface and high sedimentation rates maintain Fe under the form of Fe(III) oxides and thermodynamically prevent sulfate reduction from dominating carbon remineralization processes on the slope, whereas the much slower Mn2+ oxygenation kinetics allows diffusion of Mn2+ across the sediment-water interface of the shelf station closest to the river mouth. Exposure to oxygenated bottom waters and entrainment within mobile muds typical of deltaic sediments during high riverine discharge likely promote the formation and downslope transport of Mn(III/IV) oxides within the nepheloid layer. This phenomenon appears to form a manganese ‘conveyor belt’ that selectively enriches Mn(III/IV) oxides relative to Fe(III) oxides in the deep sediment. In contrast, the intensity of anaerobic carbon remineralization processes along the eastern continental slope the farthest from the Mississippi River plume is much lower due to the low organic and lithogenic inputs, and denitrification dominates anaerobic respiration. Overall, these findings suggest that manganese cycling and its role in carbon remineralization processes in continental slope sediments exposed to large riverine inputs may be more important than previously considered.

Despite the undeniable effect of iron on shaping patterns of ocean productivity, the relative importance of the different sources of this limiting nutrient to the ocean is still under debate. Although global estimates indicate that the benthic input of iron to the oceans is significant, most studies have investigated continental margins exposed to either upwelling or large riverine inputs, environments that are not representative of the majority of the oceans. Additionally, the number of studies that report dissolved iron concentrations in continental slope sediments is limited, despite the fact that these regions between the shelf edge and the continental rise make up >5% of the sedimentary surface area of the global ocean. The sedimentary flux of iron has traditionally been considered negligible due to the rapid oxidation of Fe2+ in oxic waters and poor solubility of the Fe(III) product. The recent realization that ferric iron may be stabilized in solution by organic ligands during oxidation near the sediment-water interface suggests that a significant fraction of the dissolved iron pool may be present under the form organic-Fe(III) complexes that could eventually reach the overlying waters. In this study, the speciation and biogeochemical importance of iron was determined in intact sediment cores along a transect across the entire continental margin near Cape Lookout, North Carolina, a region not dominated by upwelling or riverine inputs that is representative of most passive continental margins. Rates of diffusive oxygen uptake (DOU) and maximum diffusive fluxes of both dissolved Fe2+and organic-Fe(III) complexes decreased from the coastal zone to the continental shelf, remained low on the shelf and the upper continental slope, but rebounded to reach a maximum in mid-slope sediments where concentrations of Fe(III) oxides were the highest along the transect. In turn, DOU decreased and dissolved iron was below detection in lower-slope sediments, indicating that mid-slope sediments represent depocenters where Fe(III) oxides and organic matter may accumulate. Pore water sulfate and sulfide concentrations as well as separate sediment incubations confirmed that sulfate reduction does not greatly influence the cycling of iron in these sediments. The production of dissolved organic-Fe(III) in these continental margin sediments is likely regulated by a combination of aerobic oxidation in the presence of natural organic ligands near the sediment-water interface, dissimilatory iron reduction, or chemical oxidation of Fe(II) complexed to natural organic ligands. Fluxes of Fe2+ and organic-Fe(III) complexes across the sediment-water interface were not observed. However, diffusive fluxes of Fe2+ and organic-Fe(III) complexes into the oxic zone of these sediments (

The redox chemistry and speciation of Fe in both solid and dissolved phases were characterized in the organic- and Fe-rich sediments of the Satilla River estuary in South-East Georgia (USA) on a series of four cruises between July 2007 and January 2008. Results indicate that dissolved Fe is present in relatively high concentration in the overlying waters at the freshwater end of the estuary and flocculates along the river as the salinity increases downstream. Soluble organic–Fe(III) complexes comprise the majority of dissolved Fe (

WOS:000408783500011; International audience; Deep-sea fans are well known depot centers for organic carbon that should promote sulfate reduction. At the same time, the high rates of deposition of unconsolidated metal oxides from terrigenous origin may also promote metal-reducing microbial activity. To investigate the eventual coupling between the iron and sulfur cycles in these environments, shallow sediment cores ( \textless 50 cm) across various channels and levees in the Congo River deep-sea fan (similar to 5000 m) were profiled using a combination of geochemical methods. Interestingly, metal reduction dominated suboxic carbon remineralization processes in most of these sediments, while dissolved sulfide was absent. In some 'hotspot' patches, however, sulfate reduction produced large sulfide concentrations which supported chemosynthetic-based benthic megafauna. These environments were characterized by sharp geochemical boundaries compared to the iron-rich background environment, suggesting that FeS precipitation efficiently titrated iron and sulfide from the pore waters. A companion study demonstrated that methanogenesis was active in the deep sediment layers of these patchy ecosystems, suggesting that sulfate reduction was promoted by alternative anaerobic processes. These highly reduced habitats could be fueled by discrete, excess inputs of highly labile natural organic matter from Congo River turbidites or by exhumation of buried sulfide during channel flank erosion and slumping. Sulfidic conditions may be maintained by the mineralization of decomposition products from local benthic macrofauna or bacterial symbionts or by the production of more crystalline Fe(III) oxide phases that are less thermodynamically favorable than sulfate reduction in these bio-turbated sediments. Overall, the iron and sulfur biogeochemical cycling in this environment is unique and much more similar to a coastal ecosystem than a deep-sea environment.

Although a number of techniques exist to measure the major and minor inorganic anions Cl-, NO2-, Br-, NO3-, or SO42- in marine waters and sediment porewaters, a single method that can quantify all five anions in a single step with a sufficiently low detection limit has yet to be developed. In this work, a novel ion-chromatographic separation procedure was developed that exploits the UV-absorbing properties of Br-, NO3-, and NO2-, as well as the suppression of eluent absorbance by high concentrations of Cl- and SO42- to quantify all these anions simultaneously in marine waters. If additional sensitivity for the minor anions Br-, NO3-, and NO2- is needed, NaCl can be used as eluent in a matrix elimination method that no longer allows for Cl- and SO42- determination. These methods are isocratic, require no chemical or electronic suppression, are not subject to interferences, and can resolve the anions in less than 18 min. The decrease in analysis time and the number of analyses required compared with conventional methods lowers the costs associated with laboratory analysis of marine samples. Last, the simplicity and good limits of detection make this separation method suitable for in situ analyses of marine waters and porewaters, and an in situ liquid chromatograph is currently under development for these applications.

The two most frequently used systems for in vitro translation are the rabbit reticulocyte system and the wheat germ extract. These systems are useful for mRNAs isolated from cells, tissues, and capped or uncapped mRNA produced in vitro by transcription of cDNA. In a combined system, mRNA can be transcribed and translated in a single reaction. In addition these systems can be used for translation reactions with biotinylated amino acids; this allows capture of the newly synthesized protein using streptavidin immobilized on agarose.

The ability to investigate cellular processes in vitro permits detailed analysis of the process and its molecular components. Eukaryotic translation and expression is one system that has been well studied. This overview describes the development of in vitro systems, including such approaches as continuous-flow systems, coupled transcription/translation, and the incorporation of non-natural amino acids. It also discusses molecular and genetic studies to probe translation, including post-translational fate of the synthesized proteins.

Summary An analysis of the structure and presumed regulatory signals in sequenced methanogen genes is presented. The evidence for polycistronic transcriptional units is extended by inclusion of a DNA sequence, cloned from Methanobrevibacter smithii , which precedes the M. smithii pro C gene ( Hamilton and Reeve , 1985a). Comparison of DNA sequences indicates that a consensus core ribosome binding sequence for methanogens would be 5 'AGGTGA and that many of the ribosome binding sequences are, in fact, longer than the core sequence containing 7 bases with the potential to hybridize to 16SrRNA. The majority, but not all, of the sequenced methanogen genes conform to the rule in which RNY codons (R=purine, Y=pyrimidine, N=purine or pyrimidine) occur most frequently in the translated reading frame ( Shepherd 1981,1983). Codon usages by different methanogens reflect the need to accommodate genomes with very different overall %mol G+C contents. Codons such as AUA, AGA, and AGG, rarely used by E. coli are frequently used by methanogens. The dinucleotide, CG, which occurs very infrequently in eucaryotic DNAs ( Subak-Sharpe et al., 1967; Lennon and Faser , 1983; Nussinov , 1984) is also rarely found in methanogen genes. Methylation and demethylation of the cytosine residue in CG sequences has been implicated in regulation of eucaryotic gene expression ( Weisbrod , 1982) and this role used as an argument for the infrequent occurrence of CG-containing sequences. The rarity of CG in the DNA sequences so far obtained from methanogens may indicate a similar regulatory usage of CG-containing sequences in archaebacterial methanogens.

A 2.7 kilobase pair (Kb) fragment of DNA, which complements mutations in the hisI locus of Escherichia coli, has been cloned and sequenced from the genome of the methanogenic archaebacterium Methanococcus vannielii. The cloned DNA directs the synthesis of three polypeptides, with molecular weights of 71,000, 29,000 and 15,600 in minicells of E. coli. Subcloning and mutagenesis demonstrates that hisI complementation results from the activity of the 15,600 molecular weight polypeptide. The primary structure of this archaebacterial gene and its gene product have been compared with the functionally equivalent gene and protein from the eubacterium E. coli (hisI) (Chiariotti et al. 1986) and from the eucaryote Saccharomyces cerevisiae (his4A) (Donahue et al. 1982). The DNA sequences of the archaebacterial and eubacterial genes are 40% homologous, the archaebacterial and eucaryotic DNA sequences are 47% homologous and, as previously reported (Bruni et al. 1986) the eubacterial and eucaryotic DNA sequences are 45% homologous. In E. coli the hisI locus is part of a bifunctional gene (hisI/E) within the single his operon. In S. cerevisiae the his4A locus is part of a multifunctional gene (his4) which encodes a protein with at least four enzymatic activities. The his genes of S. cerevisiae do not form an operon and are not physically linked. The M. vannielii hisI gene does not appear to be part of a multifunctional DNA sequence and, although it does appear to be within an operon, the open reading frames (ORFs) 5' and 3' to the M. vannielii hisI gene are not related to any published his sequences.(ABSTRACT TRUNCATED AT 250 WORDS)

To identify an archaebacterial promoter sequence, nuclease protection studies with the purified RNA polymerase of Methanococcus vannielii were performed. The enzyme binds specifically both at protein-encoding (hisA and methyl CoM reductase, component C) and tRNA–rRNA genes. The binding region of the RNA polymerase extends from 30 base pairs (bp) upstream (−30) to 20 bp downstream (+20) from the in vivo transcription start site. This finding indicates that the archaebacterial enzyme recognizes promoters without transacting traascription factors. The DNA segment protected from nuclease digestion by bound RNA polymerase contains an octanucleotide sequence centered at −25, which is conserved between the protein-encoding and the stable RNA genes. According to the specific binding of the enzyme to only DNA-fragments harbouring this motif, we propose the sequence TTTATATA as the major recognition signal of the Methanococcus RNA polymerase. Comparison of this motif with published archaebacterial DNA sequences revealed the presence of homologous sequences at the same location upstream of 36 genes. We therefore consider the overall consensus [Formula: see text] as a general element of promoters in archaebacteria. In spite of the specific binding of the enzyme, most preparations of the Methanococcus vannielii RNA polymerase are unable to initiate transcription at the correct sites in vitro. Here we present first evidence for the possible existence of a transcription factor conferring the ability to the enzyme to initiate and terminate transcription specifically in vitro.Key words: promoter, footprint, TATA box, RNA polymerase, transcription.

Transcription initiation of the hisA gene in vivo in the archaebacterium Methanococcus vannielii, as determined by nuclease S1 and primer extension analyses, occurs 73 base pairs (bp) upstream of the translation initiation site. Binding of M. vannielii RNA polymerase protects 43 bp of DNA, from 35 bp upstream (-35) to 8 bp downstream (+8) of the hisA mRNA initiation site, from digestion by DNase I and exonuclease III. An A + T rich region, with a sequence which conforms to the consensus sequence for promoters of stable RNA-encoding genes in methanogens, is found at the same location (-25) upstream of the polypeptide-encoding hisA gene. It appears therefore that a TATA-like sequence is also an element of promoters which direct transcription of polypeptide-encoding genes in this archaebacterium.