Sigmund Jensen

Methane oxidation at 55°C and pH 2 by a thermoacidophilic bacterium belonging to the Verrucomicrobia phylum

Methanotrophic bacteria constitute a ubiquitous group of microorganisms playing an important role in the biogeochemical carbon cycle and in control of global warming through natural reduction of methane emission. These bacteria share the unique ability of using methane as a sole carbon and energy source and have been found in a great variety of habitats. Phylogenetically, known methanotrophs constitute a rather limited group and have so far only been affiliated with the Proteobacteria. Here, we report the isolation and initial characterization of a nonproteobacterial obligately methanotrophic bacterium. The isolate, designated Kam1, was recovered from an acidic hot spring in Kamchatka, Russia, and is more thermoacidophilic than any other known methanotroph, with optimal growth at ≈55°C and pH 3.5. Kam1 is only distantly related to all previously known methanotrophs and belongs to the Verrucomicrobia lineage of evolution. Genes for methane monooxygenases, essential for initiation of methane oxidation, could not be detected by using standard primers in PCR amplification and Southern blot analysis, suggesting the presence of a different methane oxidation enzyme. Kam1 also lacks the well developed intracellular membrane systems typical for other methanotrophs. The isolate represents a previously unrecognized biological methane sink, and, due to its unusual phylogenetic affiliation, it will shed important light on the origin, evolution, and diversity of biological methane oxidation and on the adaptation of this process to extreme habitats. Furthermore, Kam1 will add to our knowledge of the metabolic traits and biogeochemical roles of the widespread but poorly understood Verrucomicrobia phylum.

Selection and identification of autochthonous potential probiotic bacteria from turbot larvae ( Scophthalmus maximus ) rearing units

The purpose of this study was to select, identify and characterise bacteria as a disease control measure in the rearing of marine fish larvae (turbot, Scophthalmus maximus). Thirty-four out of 400 marine bacterial strains exhibited in vitro anti-bacterial activity against three fish larval pathogens. Two strains originated from culture collections and thirty two strains were isolated directly from turbot larvae rearing units using a pre-selection procedure to facilitate detection of antagonists. Approximately 8,500 colonies from colony-count plates were replica-plated on agar seeded with Vibrio anguillarum, and 196 of them caused zones of clearing in the V. anguillarum agar layer. Of these, 32 strains exhibited reproducible antibacterial properties in vitro when tested against the fish pathogens V. anguillarum 90-11-287, V. splendidus DMC-1 and a Pseudoalteromonas HQ. Seventeen antagonists were identified as Vibrio spp. and four of twelve tested were lethal to yolk-sac larvae. The 15 remaining strains were identified as Roseobacter spp. based on phenotypic criteria and 16S rDNA gene sequence analysis of two strains representing the two major RAPD groups. Most of the remaining 164 strains selected in the initial replica plating were identified as Vibrionaceae or Pseudoalteromonas. Roseobacter spp. were not lethal to egg yolk sac turbot larvae and in two of three trials, the mortality of larvae decreased (p > 0.001) in treatments where 10(7) cfu/ml Roseobacter sp. strain 27-4 was added, indicating a probiotic potential.

Phylogenetic analysis of bacterial communities associated with larvae of the Atlantic halibut propose succession from a uniform normal flora.

Halibut, the largest of all flatfishes is a valuable species with a great potential for aquaculture. Bacteria play an important role in regulating the health of the early life stages. The present article is the first broad-range molecular analysis of bacterial communities in larvae of the Atlantic halibut (Hippoglossus hippoglossus). DNA was extracted from larvae, water and silo biofilm from hatcheries in Norway, Scotland, Iceland and Canada. Eubacterial 16S rRNA gene fragments were amplified by polymerase chain reaction (PCR) with broad-range primers. Sequences spanning the hyper variable V3 region representing individual bacterial species were separated into community profiles by denaturing gradient gel electrophoresis (DGGE). The profiles revealed simple communities after hatching and bacterial succession following growth. Sequencing and phylogenetic analysis of excised DGGE bands suggested aerobic heterotrophs related to groups of Pseudomonas, Janthinobacterium and possibly Marinomonas to be the primary colonisers of the larvae. After onset of feeding, fermentative species (Vibrio) were detected as well. Comparative analysis of bacterial communities from different geographical regions indicated that larvae of the Atlantic halibut possess a distinct and specific normal flora.

Influence of light and temperature on photoinhibition of photosynthesis inSpirulina platensis

Photoinhibition of photosynthesis and its recovery in the cyanobacteriumSpirulina platensis was studied to find how photosynthetic rates were influenced by light and temperature. By exposing cell samples from a turbidostat culture to combinations of light and temperature, a connection between light, temperature and photoinhibition was found. The experiments showed that a 10 degree increase from 20 °C to 30 °C considerably reduced the photoinhibition. At 25 °C a photon flux density of 1720 µmol m−2 s−1 reduced the photosynthetic rate by 50 % in 1 h, but a similarly high photon flux density had nearly no negative effect at 35 °C. Reactivation in low light from 50% photoinhibition was fast and complete in 60 min at 30 °C, while at 20 °C only about 1/6 of the full capacity was regained in the same time. Addition of the protein synthesis inhibitor streptomycin to cultures undergoing photoinhibition and regeneration indicated the presence also in this organism of a repair mechanism based on protein synthesis.

Intracellular Oceanospirillales bacteria inhabit gills of Acesta bivalves.

A novel bacterium was discovered in the gills of the large bivalve Acesta excavata (Limidae) from coral reefs on the northeast Atlantic margin near the shelf break of the fishing ground Haltenbanken of Norway, and confirmed present in A. excavata from a rock-wall in the Trondheimsfjord. Purified gill DNA contained one dominant bacterial rRNA operon as indicated from analysis of broad range bacterial PCR amplicons in denaturant gradient gels, in clone libraries and by direct sequencing. The sequences originated from an unknown member of the order Oceanospirillales and its 16S rRNA gene fell within a clade of strictly marine invertebrate-associated Gammaproteobacteria. Visual inspection by fluorescent in situ hybridization and transmission electron microscopy indicated a pleomorphic bacterium with no visible cell wall, located in aggregates inside vacuoles scattered within the gill cells cytoplasm. Intracellular Oceanospirillales exist in bathymodiolin mussels (parasites), Osedax worms and whiteflies (symbionts). This bacterium apparently lives in a specific association with the Acesta.

The nonrandom microheterogeneity of 16S rRNA genes in Vibrio splendidus may reflect adaptation to versatile lifestyles.

16S rRNA molecules in a microbial strain can differ due to nucleotide variation between their genes. This is a typical trait of fast-growing bacteria to cope with different niches. We investigated characteristics of 16S rRNA genes in Vibrio splendidus strain PB1-10, from the normal flora of Atlantic halibut. Sequencing of 16S rRNA gene clones detected 35 variable positions in a total of 13 different gene copies. More than two-thirds of the substitutions occurred in regions corresponding to helix H6 and helix H17 of the 16S rRNA molecule. Possible recombination between these helixes in related bacteria (Vibrio, Photobacterium, Colwellia) from similar environments impacts 16S rRNA-based phylogeny of V. splendidus. We argue that these nonrandom modifications are maintained to provide a fine-tuning of the ribosome function to optimize translation machinery performance and ultimately bacterial niche fitness.

From adverse drug event detection to prevention. A novel clinical decision support framework for medication safety.

BACKGROUND Errors related to medication seriously affect patient safety and the quality of healthcare. It has been widely argued that various types of such errors may be prevented by introducing Clinical Decision Support Systems (CDSSs) at the point of care. OBJECTIVES Although significant research has been conducted in the field, still medication safety is a crucial issue, while few research outcomes are mature enough to be considered for use in actual clinical settings. In this paper, we present a clinical decision support framework targeting medication safety with major focus on adverse drug event (ADE) prevention. METHODS The novelty of the framework lies in its design that approaches the problem holistically, i.e., starting from knowledge discovery to provide reliable numbers about ADEs per hospital or medical unit to describe their consequences and probable causes, and next employing the acquired knowledge for decision support services development and deployment. Major design features of the frameworks services are: a) their adaptation to the context of care (i.e. patient characteristics, place of care, and significance of ADEs), and b) their straightforward integration in the healthcare information technologies (IT) infrastructure thanks to the adoption of a service-oriented architecture (SOA) and relevant standards. RESULTS Our results illustrate the successful interoperability of the framework with two commercially available IT products, i.e., a Computerized Physician Order Entry (CPOE) and an Electronic Health Record (EHR) system, respectively, along with a Web prototype that is independent of existing healthcare IT products. The conducted clinical validation with domain experts and test cases illustrates that the impact of the framework is expected to be major, with respect to patient safety, and towards introducing the CDSS functionality in practical use. CONCLUSIONS This study illustrates an important potential for the applicability of the presented framework in delivering contextualized decision support services at the point of care and for making a substantial contribution towards ADE prevention. Nonetheless, further research is required in order to quantitatively and thoroughly assess its impact in medication safety.

Differential expression of particulate methane monooxygenase genes in the verrucomicrobial methanotroph ‘Methylacidiphilum kamchatkense’ Kam1

Methane monooxygenases (MMOs) are oxygen-dependent enzymes that catalyze the oxidation of methane to methanol in the methanotrophic bacteria. The thermoacidophilic verrucomicrobial methanotroph ‘Methylacidiphilum kamchatkense’ Kam1 contains three complete and phylogenetically distinct copies of the pmoCAB gene cluster apparently organized as operons, each encoding all three subunits of particulate MMO (pMMO), and a truncated pmoCA cluster encoding only two of the subunits. Two of the clusters are present as a tandem array, but the other clusters occur in isolation. Here, the expression of these clusters has been assessed using the four pmoA genes as targets in reverse transcriptase quantitative PCR analysis. One of the pmoA genes, designated pmoA2, is at least 35-fold more strongly transcribed than the other pmoA copies. Growth at suboptimal temperature and pH conditions did not significantly change the transcription pattern, indicating that the pmoCAB2 cluster encodes the functional pMMO under methane-fuelled growth conditions. During growth on methanol, expression of pmoA2 was reduced approximately tenfold as compared to growth on methane, suggesting a role for the alternative carbon substrates in gene regulation.

Norwegian deep‐water coral reefs: cultivation and molecular analysis of planktonic microbial communities

Deep-sea coral reefs do not receive sunlight and depend on plankton. Little is known about the plankton composition at such reefs, even though they constitute habitats for many invertebrates and fish. We investigated plankton communities from three reefs at 260-350 m depth at hydrocarbon fields off the mid-Norwegian coast using a combination of cultivation and small subunit (SSU) rRNA gene and transcript sequencing. Eight months incubations of a reef water sample with minimal medium, supplemented with carbon dioxide and gaseous alkanes at in situ-like conditions, enabled isolation of mostly Alphaproteobacteria (Sulfitobacter, Loktanella), Gammaproteobacteria (Colwellia) and Flavobacteria (Polaribacter). The relative abundance of isolates in the original sample ranged from ∼ 0.01% to 0.80%. Comparisons of bacterial SSU sequences from filtered plankton of reef and non-reef control samples indicated high abundance and metabolic activity of primarily Alphaproteobacteria (SAR11 Ia), Gammaproteobacteria (ARCTIC96BD-19), but also of Deltaproteobacteria (Nitrospina, SAR324). Eukaryote SSU sequences indicated metabolically active microalgae and animals, including codfish, at the reef sites. The plankton community composition varied between reefs and differed between DNA and RNA assessments. Over 5000 operational taxonomic units were detected, some indicators of reef sites (e.g. Flavobacteria, Cercozoa, Demospongiae) and some more active at reef sites (e.g. Gammaproteobacteria, Ciliophora, Copepoda).

Evolution of temperature optimum in Thermotogaceae and the prediction of trait values of uncultured organisms

Quantitative characterization of the mode and rate of phenotypic evolution is rarely applied to prokaryotes. Here, we present an analysis of temperature optimum (Topt) evolution in the thermophilic family Thermotogaceae, which has a large number of cultured representatives. We use log-rate-interval analysis to show that Topt evolution in Thermotogaceae is consistent with a Brownian motion (BM) evolutionary model. The properties of the BM model are used to a establish confidence intervals on the unknown phenotypic trait value of an uncultured organism, given its distance to a close relative with known trait value. Cross-validation by bootstrapping indicates that the predictions are robust.