We developed an electrical modulation method for attachment and detachment of microorganisms. Living microorganisms suspended in non-nutritive media such as PBS(−) and artificial seawater were attracted by and selectively attached to indium tin oxide (ITO)/glass electrode regions to which a negative potential was applied. The microorganisms suspended in LB medium and glucose solution were not attracted to the ITO electrode. Dead microorganisms were not attracted to the ITO electrode. The living microorganisms were retrieved after detachment from the ITO electrode by application of a high-frequency triangular wave potential. When we applied this method to separate microorganisms from deep-sea sediment, bacteria belonging to 19 phyla and 23 classes were collected without undesirable high molecular weight contaminants such as humic acids. At the phylum and class level, respectively, 95 and 87 % of the phylotypes among electrically retrieved bacteria were common to the gene clones from the direct sediment DNA extraction. This technique is a novel useful method to prepare bacterial cells in a single population or a community for metagenomic analyses.

A controlled culture system has been developed to induce nerve growth factor (NGF) production in astroglial cells that are cultured on an electrode surface. The electrode potential is alternatively modulated at an amplitude of 300 mV and a frequency of 10 Hz. The electric stimulation triggers NGF production and secretion. The mechanism of the electrically induced NGF production is discussed in association with protein kinase C (PKC) activation.

We report successful larval hatching of deep-sea shrimp after decompression to atmospheric pressure. Three specimens of deep-sea shrimp were collected from an ocean depth of 1157 m at cold-seep sites off Hatsushima Island in Sagami Bay, Japan, using a pressure-stat aquarium system. Phylogenetic analysis of Alvinocaris sp. based on cytochrome c oxidase subunit gene sequences confirmed that these species were a member of the genus Alvinocaris. All 3 specimens survived to reach atmospheric pressure conditions after stepwise 63-day decompression. Two of the specimens contained eggs, which hatched after 10 and 16 days, respectively, of full decompression. Although no molting of the shrimp larvae was observed during 74 days of rearing under atmospheric pressure, the larvae developed conventional dark-adapted eyes after 15 days.

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We investigated the pressure tolerance of deep-sea eel (Simenchelys parasiticus; habitat depth, 366-2,630 m) cells, conger eel (Conger myriaster) cells, and mouse 3T3-L1 cells. Although there were no living mouse 3T3-L1 and conger eel cells after 130 MPa (0.1 MPa = 1 bar) hydrostatic pressurization for 20 min, all deep-sea eel cells remained alive after being subjected to pressures up to 150 MPa for 20 min. Pressurization at 40 MPa for 20 min induced disruption of actin and tubulin filaments with profound cell-shape changes in the mouse and conger eel cells. In the deep-sea eel cells, microtubules and some actin filaments were disrupted after being subjected to hydrostatic pressure of 100 MPa and greater for 20 min. Conger eel cells were sensitive to pressure and did not grow at 10 MPa. Mouse 3T3-L1 cells grew faster under pressure of 5 MPa than at atmospheric pressure and stopped growing at 18 MPa. Deep-sea eel cells were capable of growth in pressures up to 25 MPa and stopped growing at 30 MPa. Deep-sea eel cells required 4 h at 20 MPa to finish the M phase, which was approximately fourfold the time required under atmospheric conditions.

The purpose of this study was to develop a novel electrical retrieval method (ER method) for living sponge-associated microorganisms from marine sponges frozen at −80 °C. A −0.3-V vs. Ag/AgCl constant potential applied for 2 h at 9 °C induced the attachment of the sponge-associated microorganisms to an indium tin oxide/glass (ITO) or a gallium-doped zinc oxide/glass (GZO) working electrode. The electrically attached microorganisms from homogenized Spirastrella insignis tissues had intact cell membranes and showed intracellular dehydrogenase activity. Dead microorganisms were not attracted to the electrode when the homogenized tissues were autoclaved for 15 min at 121 °C before use. The electrically attached microorganisms included cultivable microorganisms retrieved after detachment from the electrode by application of a 9-MHz sine-wave potential. Using the ER method, we obtained 32 phyla and 72 classes of bacteria and 3 archaea of Crenarchaeota thermoprotei, Marine Group I, and Thaumarchaeota incertae sedis from marine sponges S. insignis and Callyspongia confoederata. Employment of the ER method for extraction and purification of the living microorganisms holds potential of single-cell cultivation for genome, transcriptome, proteome, and metabolome analyses of bioactive compounds producing sponge-associated microorganisms.

A novel Gram-stain-negative, aerobic, heterotrophic, stalked and capsulated bacterium with potential denitrification ability, designated strain TAR-002 T , was isolated from deep seafloor sediment in Japan. Colonies lacked lustre, and were viscous and translucent white. The ranges of temperature, pH and salt concentration for growth were 8-30 6C, pH 6.0-10.0 and 1-3 % (w/v) NaCl. Phylogenetic analysis based on 16S rRNA gene sequences confirmed that strain TAR-002 T belongs to the genus Brevundimonas of the class Alphaproteobacteria. Levels of similarity between the 16S rRNA gene sequence of strain TAR-002 T and those of the type strains of species of the genus Brevundimonas were 93.5-98.9 %; the most closely related species was Brevundimonas basaltis. In DNA-DNA hybridization assays between strain TAR-002 T and its phylogenetic neighbours, Brevundimonas lenta DS-18 T , B. basaltis J22 T , Brevundimonas subvibrioides ATCC 15264 T and Brevundimonas alba DSM 4736 T , mean hybridization levels were 6.4-27.7 %. The G+C content of strain TAR-002 T was 70.3 mol%. Q-10 was the major respiratory isoprenoid quinone. The major fatty acids were C 18 : 1 v7c and C 16 : 0 , and the presence of 1,indicates the affiliation of strain TAR-002 T with the genus Brevundimonas. On the basis of biological characteristics and 16S rRNA gene sequence comparisons, strain TAR-002 T is considered to represent a novel species of the genus Brevundimonas, for which the name Brevundimonas denitrificans sp. nov. is proposed; the type strain is TAR-002 T (5NBRC 110107 T 5CECT 8537 T ).

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