Mechanisms of gene expression controlled by pressure in deep-sea microorganisms

A pressure-regulated operon has been cloned and sequenced from deep-sea barophilic Shewanella strains. To understand pressure-regulated mechanisms of gene expression, a regulatory element upstream of the pressure-regulated operon from Shewanella sp. strain DSS12 was studied. Regions A and B were classified by sequence analysis. A unique octamer motif, AAGGTAAG, was found to be repeated in tandem 13 times in region B. An electrophoretic mobility shift assay demonstrated that a σ⁵⁴-like factor recognizes region A and other unknown factors recognize region B. Different shift patterns of the protein–DNA complexes were observed when extracts of cells cultured at 0.1 MPa or 50 MPa were incubated with a DNA probe specific for region B. These results indicate that the deep-sea strain DSS12 expresses different DNA-binding factors under different pressure conditions. Received: January 22, 1998 / Accepted: February 16, 1998     

Main lipid classes in some species of deep-sea corals in the Newfoundland and Labrador region (Northwest Atlantic Ocean)

Corals contain large quantities of lipids in their tissues; these lipids may be either structural or for storage. Little information is available about the lipid content of deep-sea corals, as well as ratios of main lipid classes. In this study, lipid percentages of 81 deep-sea specimens were measured and the presence of six major classes, including sterols (STEROLS), free fatty acids (FFA), triacylglycerols (TG), monoalkyldiacyl glycerol (MADAG), wax (WAX), and sterol esters (SE), was assessed. Deep-sea corals had fewer lipids than their shallow water counterparts. Decision-tree analysis revealed a link between coral groups and total lipid percentages, showing that species within the same group were characterized by similar lipid amounts. Depth did not seem to impact the total lipid percentages, suggesting that deep-sea corals adapt to the differential access to food by changing the proportion of lipid classes while maintaining equivalent lipid levels. In deep-sea species, similar to their shallow water counterparts, energy seems to be stored as neutral lipids (wax esters and triacylglycerols), with the notable difference that a high proportion of MADAG is present. These compounds are less rich in energy than TG. Depth trends were found for FFA, TG and SE with an increase in percentages after 800 m suggesting a potential need for storage due to decreased food availability. A subsequent decrease after 1,100 m was observed for FFA and TG but a more detailed investigation is warranted as the number of specimens acquired from these depths was less than 20. It is nonetheless a surprising result as increased storage is expected when food sources are sparse.     

Characterization of a lysin from deep-sea thermophilic bacteriophage GVE2

Thermostable enzymes from thermophiles have attracted extensive studies. However, little is known about thermophilic lysin of bacteriophage obtained from deep-sea hydrothermal vent. In this study, a lysin from deep-sea thermophilic bacteriophage Geobacillus virus E2 (GVE2) was characterized for the first time. It was found that the GVE2 lysin was highly homologous with N-acetylmuramoyl-L-alanine amidases. After expression in Escherichia coli, the recombinant GVE2 lysin was purified. The recombinant lysin was active over a range of temperature from 40 °C to 80 °C, with an optimum at 60 °C. Its optimal pH was 6.0, and it was stable over a wide range of pH from 4.0 to 10.0. The lysin was highly active when some enzyme inhibitors or detergents (phenylmethylsulfonyl fluoride, Tween 20, Triton X-100, and chaps) were used. However, it was strongly inhibited by sodium dodecyl sulfate and ethylene diamine tetraacetic acid. Its enzymatic activity could be slightly stimulated in the presence of Na⁺ and Li⁺. But the metal ions Mg²⁺, Ba²⁺, Zn²⁺, Fe³⁺, Ca²⁺, and Mn²⁺ at concentrations of 1 or 10 mM showed inhibitions to the lysin activity. Our study demonstrated the first characterization of lysin from deep-sea thermophilic bacteriophage.     

A modeling approach of the influence of local hydrodynamic conditions on larval dispersal at hydrothermal vents

Deep-sea hydrothermal vent animal communities along oceanic ridges are both patchy and transient. Larval dispersal is a key factor in understanding how these communities function and are maintained over generations. To date, numerical approaches simulating larval dispersal considered the effect of oceanic currents on larval transportation over hundreds of kilometers but very seldom looked at the effect of local conditions within meters around chimneys. However, small scale significant variations in the hydrodynamics may influence larval fate in its early stages after release, and hence have a knock-on effect on both dispersal and colonization processes. Here we present a new numerical approach to the study of larval dispersal, considering small scales within the range of the biological communities, called “bio-hydrodynamical” scale, and ranging from a few centimeters to a few meters around hydrothermal sources. We use a physical model for the vent based on jet theory and compute the turbulent velocity field around the smoker. Larvae are considered as passive particles whose trajectories are affected by hydrodynamics, topography of the vent chimney and larval biological properties. Our model predicts that bottom currents often dominate all other factors either by entraining all larvae away from the vent or enforcing strong colonization rates. When bottom currents are very slow (), general larvae motion is upwards due to entrainment by the main smoker jet. In this context, smokers with vertical slopes favor retention of larvae because larval initial trajectory is nearly parallel to the smoker wall, which increases the chances to settle. This retention phenomenon is intensified with increasing velocity of the main smoker jet because entrainment in the high velocity plume is preceded by a phase when larvae are attracted towards the smoker wall, which occurs earlier with higher velocity of the main jet. Finally, the buoyancy rate of the larvae, measured to be in the range of , is generally irrelevant unless hydrodynamic conditions are balanced, i.e. if the buoyancy rate is comparable to both the bottom current speed and the local water velocity due to entrainment by close smokers. Overall, our model evidences the strong effect of the release point of larvae on their future entrainment within local fluxes. Larvae released from smoker walls might have an entirely different fate than those released further away in the water column. The latter are not, or less, affected by near-chimney hydrodynamics.     

<Emphasis Type="Italic">Pyrolycus manusanus</Emphasis>, a new genus and species of deep-sea eelpout from a hydrothermal vent field in the Manus Basin,Papua New Guinea (Zoarcidae,Lycodinae)

A new genus and species of deep-sea zoarcid fish, Pyrolycus manusanus, is described on the basis of eight specimens collected from a hydrothermal vent field in the Manus Basin, Papua New Guinea, at depths of 1623–1971 m. This genus is distinguished from other scaleless eelpout genera in the subfamily Lycodinae by the following combination of characters: six suborbital bones; six to seven suborbital pores; flesh gelatinous; gill slit large, extending ventrally beyond pectoral fin base; three postorbital pores; single occipital pore; two to three pelvic fin rays; palatopterygoid series weak; vomer and palatine teeth present; pseudobranch, scales, lateral line absent; pyloric caeca present; 16–17 pectoral fin rays; 22–23 + 56–59 = 78–81 vertebrae. Received: August 5, 1999 / Revised: July 5, 2001 / Accepted: August 26, 2001     

深海来源低温诱导表达载体pSW4的构建及其应用

【目的】在深海来源穿梭载体p SW2的基础上构建低温诱导高效表达载体,为最终获得环境污染物高效降解菌提供基础和工具。【方法】通过最小化敲除实验确定载体必需基因,以绿色荧光蛋白GFP为报告基因检测载体的表达能力变化,进一步添加纯化标签、多克隆位点及替换启动子等改造获得低温诱导表达载体pSW4。【结果】敲除实验结果显示编码单链结合蛋白的基因fps B敲除后载体的表达效率显著提高。以GFP为报告基因检测发现,pSW4的表达效率较pSW2有显著提升(4°C条件下提高10.7倍)。以深海细菌Shewanella piezotolerans WP3为宿主菌,应用pSW4为表达载体表达深海细菌Shewanella psychrophila WP2的聚合酶亚基,检测显示其在Mg~(2+)条件下具有切割单链DNA的核酸酶活性,而在Mg~(2+)或Mn~(2+)条件下具有切割双链DNA的核酸酶活性。【结论】构建了低温诱导表达载体pSW4,并证实了其适用性,有助于今后构建环境修复菌及相关的应用性研究。     

<Emphasis Type="Italic">Arthrobacter ardleyensis</Emphasis> sp. nov., isolated from Antarctic lake sediment and deep-sea sediment

Three psychrotrophic Arthrobacter strains, isolated from Antarctic lake sediment (An24, An25^T) and deep-sea sediment (ZX6) were studied. Their 16S rRNA gene sequences showed highest similarities (97.0–97.9%) with those of A. nicotianae and A. protophormiae. All three strains underwent rod–coccus morphological change, had high mol% G+C content, were aerobic to slightly anaerobic, and grew between 0°C and 30°C, with optimal growth temperature around 25°C. The cell wall peptidoglycan was A4 variant. DNA–DNA hybridization, physiological and chemotaxonomic studies indicated that these three strains constituted a new homogeneous genomic species within the genus Arthrobacter, for which the name Arthrobacter ardleyensis, with the type strain An25^T (CGMCC 1.3685, JCM 12921) was proposed.M. Chen and X. Xiao contributed equally to this paper.     

<Emphasis Type="Italic">Thermococcus marinus</Emphasis> sp. nov. and<Emphasis Type="Italic"> Thermococcus radiotolerans</Emphasis> sp. nov., two hyperthermophilic archaea from deep-sea hydrothermal vents that resist ionizing radiation

Enrichments for anaerobic, organotrophic hyperthermophiles were performed with hydrothermal chimney samples collected from the Mid-Atlantic Ridge at a depth of 3,550 m (23°22N, 44°57W) and the Guaymas Basin (27°01N, 111°24W) at a depth of 2,616 m. Positive enrichments were submitted to -irradiation at doses of 20 and 30 kGy. Two hyperthermophilic, anaerobic, sulfur-metabolizing archaea were isolated. Strain EJ1^T was isolated from chimney samples collected from the Mid-Atlantic Ridge after -irradiation at 20 kGy, and strain EJ2^T was isolated from the Guaymas Basin after -irradiation at 30 kGy. Only strain EJ2^T was motile, and both formed regular cocci. These new strains grew between 55 and 95 °C with the optimal temperature being 88 °C. The optimal pH for growth was 6.0, and the optimal NaCl concentration for growth was around 20 g l^–1. These strains were obligate anaerobic heterotrophs that utilized yeast extract, tryptone, and peptone as a carbon source for growth. Ten amino acids were essential for the growth of strain EJ1^T, such as arginine, aspartic acid, isoleucine, leucine, methionine, phenylalanine, proline, threonine, tyrosine, and valine, while strain EJ2^T was unable to grow on a mixture of amino acids. Elemental sulfur or cystine was required for EJ2^T growth and was reduced to hydrogen sulfide. Rifampicin inhibited growth for both strains EJ1^T and EJ2^T. The G+C contents of the genomic DNA were 52.3 and 54.5 mol% for EJ1^T and EJ2^T, respectively. As determined by 16S rRNA gene sequence analysis, these strains were more closely related to Thermococcus gorgonarius, T. celer, T. guaymasensis, T. profundus, and T. hydrothermalis. However, no significant homology was observed between them with DNA–DNA hybridization. These novel organisms also possess phenotypic traits that differ from those of its closest phylogenetic relatives. Therefore, it is proposed that these isolates, which are amongst the most radioresistant hyperthermophilic archaea known to date with T. gammatolerans (Jolivet et al. 2003a), should be described as novel species T. marinus sp. nov. and T. radiotolerans sp. nov. The type strain of T. marinus is strain EJ1^T (=DSM 15227^T=JCM 11825^T) and the type strain of T. radiotolerans is strain EJ2^T (=DSM 15228^T=JCM 11826^T).Communicated by J. WiegelThe GenBank accession numbers for the 16S rRNA sequence of Thermococcus marinus strain EJ1^T and Thermococcus radiotolerans EJ2^T are AF479012 and AF479013, respectively.     

Expansion of the geographic distribution of a novel lineage of epsilon-Proteobacteria to a hydrothermal vent site on the Southern East Pacific Rise

The diversity associated with a microbial mat sample collected from a deep-sea hydrothermal vent on the Southern East Pacific Rise was determined using a molecular phylogenetic approach based on the comparison of sequences from the small subunit ribosomal RNA gene (16S rDNA). The DNA was extracted from the sample and the 16S rDNA was amplified by PCR. Sixteen different phylotypes were identified by restriction fragment length polymorphism analysis; four phylotypes were later identified as putative chimeras. Analysis of the 16S rDNA sequences placed all the phylotypes within the Proteobacteria. The majority of the sequences (98%) were most closely related to a new clade of epsilon-Proteobacteria that were initially identified from an in situ growth chamber deployed on a deep-sea hydrothermal vent on the Mid-Atlantic Ridge in 1995. The similarity between phylotypes identified from Atlantic and Pacific deep-sea hydrothermal vent sites indicates that this new clade of Proteobacteria may be endemic to and widely distributed among deep-sea hydrothermal vents.