Diversity of extremophilic bacteria in the sediment of high-altitude lakes located in the mountain desert of Ojos del Salado volcano,Dry-Andes

Ojos del Salado, the highest volcano on Earth is surrounded by a special mountain desert with extreme aridity, great daily temperature range, intense solar radiation, and permafrost from 5000 meters above sea level. Several saline lakes and permafrost derived high-altitude lakes can be found in this area, often surrounded by fumaroles and hot springs. The aim of this study was to gain information about the bacterial communities inhabiting the sediment of high-altitude lakes of the Ojos del Salado region located between 3770 and 6500 m. Altogether 11 sediment samples from 4 different altitudes were examined with 16S rRNA gene based denaturing gradient gel electrophoresis and clone libraries. Members of 17 phyla or candidate divisions were detected with the dominance of Proteobacteria, Acidobacteria, Actinobacteria and Bacteroidetes. The bacterial community composition was determined mainly by the altitude of the sampling sites; nevertheless, the extreme aridity and the active volcanism had a strong influence on it. Most of the sequences showed the highest relation to bacterial species or uncultured clones from similar extreme environments.     

Expression and functional analysis of two NhaD type antiporters from the halotolerant and alkaliphilic <Emphasis Type="Italic">Halomonas</Emphasis> sp. Y2

Na⁺/H⁺ antiporters play important roles in ion and pH homeostasis. In this study, two NhaD homologues that effectively catalyze Na⁺/H⁺ antiporter were identified from Halomonas sp. Y2, a halotolerant and alkaliphilic strain isolated from sodium enriched black liquor. They exhibited high sequence identity of 72 % and similar binding affinities for Na⁺ and Li⁺ translocation, while having different pH profiles. Ha-NhaD1 was active at pH 6.0 and most active at pH 8.0–8.5, whereas Ha-NhaD2 lacked activity at pH 6.0 but exhibited maximum activity at pH 9.5 or higher. Based on multiple alignments, 11 partially conserved residues were selected and corresponding mutants were generated for Ha-NhaD1. As expected, replacement of most of the hydrophobic residues abolished the cation exchange activities. Three serine residues at positions 200, 282 and 353 in Ha-NhaD1 were replaceable by alanines with partial retention of activity. The S353A mutant exhibited significantly reduced binding affinity for Na⁺ and Li⁺, while S282 mutant exhibited an alkaline shift of about 1.5 pH units, as compared to the wild type Ha-NhaD1. Serine at position 282 was predicted to be located in transmembrane segment VIII and was found to be important in regulating pH sensitivity in concert with flanking residues.     

Purification and characterization of an extracellular haloalkaline serine protease from the moderately halophilic bacterium, <Emphasis Type="Italic">Bacillus iranensis (</Emphasis>X5B)

This study reports the purification and characterization of an extracellular haloalkaline serine protease from the moderately halophilic bacterium, Bacillus iranensis, strain X5B. The enzyme was purified to homogeneity by acetone precipitation, ultrafiltration and carboxymethyl (CM) cation exchange chromatography, respectively. The purified protease was a monomeric enzyme with a relative molecular mass of 48–50 kDa and it was inhibited by PMSF indicating that it is a serine-protease. The optimum pH, temperature and NaCl concentration were 9.5, 35 °C and 0.98 M, respectively. The enzyme showed a significant tolerance to salt and alkaline pH. It retained approximately 50 % of activity at 2.5 M NaCl and about 70 % of activity at highly alkaline pH of 11.0; therefore, it was a moderately halophilic and also can be activated by metals, especially by Ca²⁺. The specific activity of the purified protease was measured to be 425.23 μmol of tyrosine/min per mg of protein using casein as a substrate. The apparent K _m and V _max values were 0.126 mM and 0.523 mM/min, respectively and the accurate value of k _cat was obtained as 3.284 × 10^?2 s^?1. These special and important characteristics make this serine protease as valuable tool for industrial applications.     

Potential applications of stress solutes from extremophiles in protein folding diseases and healthcare

Protein misfolding, aggregation and deposition in the brain, in the form of amyloid, are implicated in the etiology of several neurodegenerative disorders, such as Alzheimer’s, Parkinson’s and prion diseases. Drugs available on the market reduce the symptoms, but they are not a cure. Therefore, it is urgent to identify promising targets and develop effective drugs. Preservation of protein native conformation and/or inhibition of protein aggregation seem pertinent targets for drug development. Several studies have shown that organic solutes, produced by extremophilic microorganisms in response to osmotic and/or heat stress, prevent denaturation and aggregation of model proteins. Among these stress solutes, mannosylglycerate, mannosylglyceramide, di-myo-inositol phosphate, diglycerol phosphate and ectoine are effective in preventing amyloid formation by Alzheimer’s Aβ peptide and/or α-synuclein in vitro. Moreover, mannosylglycerate is a potent inhibitor of Aβ and α-synuclein aggregation in living cells, and mannosylglyceramide and ectoine inhibit aggregation and reduce prion peptide-induced toxicity in human cells. This review focuses on the efficacy of stress solutes from hyper/thermophiles and ectoines to prevent amyloid formation in vitro and in vivo and their potential application in drug development against protein misfolding diseases. Current and envisaged applications of these extremolytes in neurodegenerative diseases and healthcare will also be addressed.     

Genomic and physiological analysis reveals versatile metabolic capacity of deep-sea <Emphasis Type="Italic">Photobacterium phosphoreum</Emphasis> ANT-2200

Bacteria of the genus Photobacterium thrive worldwide in oceans and show substantial eco-physiological diversity including free-living, symbiotic and piezophilic life styles. Genomic characteristics underlying this variability across species are poorly understood. Here we carried out genomic and physiological analysis of Photobacterium phosphoreum strain ANT-2200, the first deep-sea luminous bacterium of which the genome has been sequenced. Using optical mapping we updated the genomic data and reassembled it into two chromosomes and a large plasmid. Genomic analysis revealed a versatile energy metabolic potential and physiological analysis confirmed its growth capacity by deriving energy from fermentation of glucose or maltose, by respiration with formate as electron donor and trimethlyamine N-oxide (TMAO), nitrate or fumarate as electron acceptors, or by chemo-organo-heterotrophic growth in rich media. Despite that it was isolated at a site with saturated dissolved oxygen, the ANT-2200 strain possesses four gene clusters coding for typical anaerobic enzymes, the TMAO reductases. Elevated hydrostatic pressure enhances the TMAO reductase activity, mainly due to the increase of isoenzyme TorA1. The high copy number of the TMAO reductase isoenzymes and pressure-enhanced activity might imply a strategy developed by bacteria to adapt to deep-sea habitats where the instant TMAO availability may increase with depth.     

Diversity of gene cassettes and the abundance of the class 1 integron-integrase gene in sediment polluted by metals

The integron-gene cassette system has typically been associated with antibiotic-resistant pathogens. However, the diversity of gene cassettes and the abundance of class 1 integrons outside of the clinical context are not fully explored. Primers targeting the conserved segments of attC recombination sites were used to amplify gene cassettes from the sediment of the Mina stream, which exhibited a higher degree of stress to metal pollution in the dry season than the rainy season. Of the 143 total analyzed sequences, 101 had no matches to proteins in the database, where cassette open reading frames could be identified by homology with database entries. There was a predominance of sequences encoding essential cellular functions. Each season that was sampled yielded a specific pool of gene cassettes. Real-time PCR revealed that 8.5 and 41.6 % of bacterial cells potentially harbored a class 1 integron in the rainy and dry seasons, respectively. In summary, our findings demonstrate that most of the gene cassettes have no ascribable function and, apparently, historically metal-contaminated sediment favors the maintenance of bacteria containing the intI1 gene. Thus, the diversity of gene cassettes is far from being fully explored deserving further attention.     

Identification and Characterization of the Lysine-Rich Matrix Protein Family in <Emphasis Type="Italic">Pinctada fucata</Emphasis>: Indicative of Roles in Shell Formation

Mantle can secret matrix proteins playing key roles in regulating the process of shell formation. The genes encoding lysine-rich matrix proteins (KRMPs) are one of the most highly expressed matrix genes in pearl oysters. However, the expression pattern of KRMPs is limited and the functions of them still remain unknown. In this study, we isolated and identified six new members of lysine-rich matrix proteins, rich in lysine, glycine and tyrosine, and all of them are basic matrix proteins. Combined with four members of the KRMPs previously reported, all these proteins can be divided into three subclasses according to the results of phylogenetic analyses: KRMP1–3 belong to subclass KPI, KRMP4–5 belong to KPII, and KRMP6–10 belong to KPIII. Three subcategories of lysine-rich matrix proteins are highly expressed in the D-phase, the larvae and adult mantle. Lysine-rich matrix proteins are involved in the shell repairing process and associated with the formation of the shell and pearl. What’s more, they can cause abnormal shell growth after RNA interference. In detail, KPI subgroup was critical for the beginning formation of the prismatic layer; both KPII and KPIII subgroups participated in the formation of prismatic layer and nacreous layer. Compared with different temperatures and salinity stimulation treatments, the influence of changes in pH on KRMPs gene expression was the greatest. Recombinant KRMP7 significantly inhibited CaCO₃ precipitation, changed the morphology of calcite, and inhibited the growth of aragonite in vitro. Our results are beneficial to understand the functions of the KRMP genes during shell formation.     

Biodegradability of Vegetation-Derived Dissolved Organic Carbon in a Cool Temperate Ombrotrophic Bog

Dissolved organic carbon (DOC) plays a key role in the peatland carbon balance and serves numerous ecological and chemical functions including acting as a microbial substrate. In this study, we quantify the concentration, biodegradability, and intrinsic properties of DOC obtained from peat, fresh material, and litter from nine species of ombrotrophic bog vegetation. Potential biodegradability was assessed by incubating vegetation extracts for 28 days in the dark and measuring percent DOC loss as the fraction of biodegradable DOC (%BDOC) while DOC properties were characterized using UV–Vis absorbance and fluorescence measurements. The mean initial DOC concentration extracted differed significantly among species (P < 0.05) and was significantly higher in fresh material, 217 ± 259 mg DOC l^?1, than either litter or peat extracts with mean concentrations of 82.1 ± 117 mg DOC l^?1 and 12.7 ± 1.0 mg DOC l^?1, respectively (P < 0.05). %BDOC also differed significantly among species (P < 0.05) and ranged from 52 to 73% in fresh cuttings with the greatest fraction observed in S. magellanicum; 22–46% in litter; and 24% in peat. The majority of variability (82.5%) in BDOC was explained by initial absorbance at 254 nm and total dissolved nitrogen concentration which was further resolved into significant non-linear relationships between %BDOC and both humic-like and protein-like DOC fractions (P < 0.05). Our results highlight the extremely heterogeneous nature of the surface vegetation-derived DOC input in peatlands and stress the importance of vegetation species in peatland ecosystem function.     

The Effect of Genetic Diversity on Ecosystem Functioning in Vegetated Coastal Ecosystems

In species-poor communities, genetic diversity potentially plays an important role for ecosystem functioning, though this is still largely unexplored in marine and estuarine ecosystems. We studied how genetic diversity (sensu genotypic diversity and/or allelic richness) affects ecosystem functioning in marine habitat-forming plant communities. First, we conducted a 15-month field experiment in the highly seasonal Baltic Sea and established mono- and polycultures of different genotypes and genotype combinations of Zostera marina. Second, we reviewed existing literature and performed a meta-analysis of 12 studies including this study. We found no evidence of positive genetic diversity effects on shoot production in the field experiment, but diversity enhanced community stability over time. The literature review revealed that a majority of the included studies observed positive effects of genetic diversity on ecosystem functions such as primary production and nutrient uptake. The results from the meta-analysis support the hypothesis that genetic diversity effects on productivity are stronger during or after periods of stress. These diversity effects were also more positive in the field compared to mesocosm studies. Our results indicate that genetic diversity has positive effects on ecosystem functioning, particularly during increased environmental stress. Thus, local genetic diversity should be preserved especially in species-poor ecosystems, where it potentially provides insurance against environmental change.     

Surviving in Changing Seascapes: Sediment Dynamics as Bottleneck for Long-Term Seagrass Presence

Changes in the seascape often result in altered hydrodynamics that lead to coinciding changes in sediment dynamics. Little is known on how altered sediment dynamics affect long-term seagrass persistence. We studied the thresholds of sediment dynamics in relation to seagrass presence by comparing sediment characteristics and seagrass presence data of seven separate seagrass meadows. All meadows had a long-term (>20 years) presence. Within these meadows, we distinguish so-called “hotspots” (areas within a meadow where seagrass was found during all mapping campaigns) and “coldspots” (with infrequent seagrass presence). We monitored static sediment characteristics (median grain size, bulk density, silt content) and sediment dynamics (that is, bed level change and maximum sediment disturbance depth), bioturbation (that is, lugworm densities and induced fecal pit and mound relief), and seagrass cover. We statistically analyzed which sediment characteristic best explains seagrass cover. Densely vegetated hotspots were shown to have lower sediment dynamics than sparsely vegetated hotspots and coldspots, whereas static sediment characteristics were similar (grain size, bulk density). The vegetation cover was either low (2–15%) or high (>30%) and sediment dynamics showed a threshold for vegetation cover. From this correlative finding, we postulate a self-sustaining feedback of relatively dense seagrass via sediment stabilization and accordingly a runaway feedback once the seagrass cover becomes too sparse. The sensitivity for sediment dynamics shown in our study implies that future existence of seagrass meadows may be at risk as ongoing climate change might directly (increased environmental extremes) or indirectly (changing seascapes) negatively affect seagrass beds.