Innovative, ecofriendly biosorbent-biodegrading biofilms for bioremediation of oil- contaminated water.

Immobilization of microorganisms capable of degrading specific contaminants significantly promotes bioremediation processes. In this study, innovative and ecofriendly biosorbent-biodegrading biofilms have been developed in order to remediate oil-contaminated water. This was achieved by immobilizing hydrocarbon-degrading gammaproteobacteria and actinobacteria on biodegradable oil-adsorbing carriers, based on polylactic acid and polycaprolactone electrospun membranes. High capacities for adhesion and proliferation of bacterial cells were observed by scanning electron microscopy. The bioremediation efficiency of the systems, tested on crude oil and quantified by gas chromatography, showed that immobilization increased hydrocarbon biodegradation by up to 23 % compared with free living bacteria. The resulting biosorbent biodegrading biofilms simultaneously adsorbed 100 % of spilled oil and biodegraded more than 66 % over 10 days, with limited environmental dispersion of cells. Biofilm-mediated bioremediation, using eco-friendly supports, is a low-cost, low-impact, versatile tool for bioremediation of aquatic systems.

Aliidiomarina minuta sp. nov., a haloalkaliphilic bacterium that forms ultra-small cells under non-optimal conditions.

An aerobic haloalkaliphilic bacterium, designated strain MLST1(T), was isolated from filtered (0.22 µm) Mono Lake (USA) waters. The isolate was observed to grow primarily on yeast extract, peptone and tryptone. Optimal growth occurred in media at pH 9.5 containing 5-11 g/l yeast extract, and 70-100 g/l NaCl. When in log phase of growth, cells were found to be mostly curved motile rods (1-3 µm length by 0.4-1 µm diameter). Phylogenetic analysis of the 16S rRNA gene and chemotaxonomic data revealed that the isolate belonged to the family Idiomarinaceae, and is closely related to Aliidiomarina maris (96.67 % sequence similarity). The major fatty acids were identified to be iso-C17:1 ω9c (27.1 %), iso-C17:0 (21.3 %) and iso-C15:0 (12.2 %). Predominant polar lipids included phosphatidylethanolamine, diphosphatidylglycerol, phosphatidylglycerol, and the major respiratory quinone was identified as Q8. The DNA base composition was 46.3 mol% G+C. Survival studies indicated that strain MLST1(T) remains viable after exposure to adverse conditions, particularly in the prolonged absence of a carbon source, at low temperatures and with no NaCl. Under these conditions, the cells shrunk to around 0.2 µm in length by 0.1 µm in diameter and passed through 0.22 µm filters. The ultra-small cells could only be resuscitated in media with low levels of yeast extract, up to 0.6 g/l. Once resuscitated, cells were able to grow to full size. Strain MLST1(T) is clearly a unique bacterium in the waters of Mono Lake and the name Aliidiomarina minuta sp. nov. is proposed. The type strain is MLST1(T) (=JCM 17425(T) = KCTC 23357(T)).

Environmental marine pathogen isolation using mesocosm culture of sharpsnout seabream: striking genomic and morphological features of novel Endozoicomonas sp.

Aquaculture is a burgeoning industry, requiring diversification into new farmed species, which are often at risk from infectious disease. We used a mesocosm technique to investigate the susceptibility of sharpsnout seabream (Diplodus puntazzo) larvae to potential environmental pathogens in seawater compared to control borehole water. Fish exposed to seawater succumbed to epitheliocystis from 21 days post hatching, causing mortality in a quarter of the hosts. The pathogen responsible was not chlamydial, as is often found in epitheliocystis, but a novel species of the γ-proteobacterial genus Endozoicomonas. Detailed characterisation of this pathogen within the infectious lesions using high resolution fluorescent and electron microscopy showed densely packed rod shaped bacteria. A draft genome sequence of this uncultured bacterium was obtained from preserved material. Comparison with the genome of the Endozoicomonas elysicola type strain shows that the genome of Ca. Endozoicomonas cretensis is undergoing decay through loss of functional genes and insertion sequence expansion, often indicative of adaptation to a new niche or restriction to an alternative lifestyle. These results demonstrate the advantage of mesocosm studies for investigating the effect of environmental bacteria on susceptible hosts and provide an important insight into the genome dynamics of a novel fish pathogen.

Characterization of uncultured giant rod-shaped magnetotactic Gammaproteobacteria from a freshwater pond in Kanazawa, Japan.

Magnetotactic bacteria (MTB) are widespread aquatic bacteria, and are a phylogenetically, physiologically and morphologically heterogeneous group, but they all have the ability to orientate and move along the geomagnetic field using intracellular magnetic organelles called magnetosomes. Isolation and cultivation of novel MTB are necessary for a comprehensive understanding of magnetosome formation and function in divergent MTB. In this study, we enriched a giant rod-shaped magnetotactic bacterium (strain GRS-1) from a freshwater pond in Kanazawa, Japan. Cells of strain GRS-1 were unusually large (~13×~8 µm). They swam in a helical trajectory towards the south pole of a bar magnet by means of a polar bundle of flagella. Another striking feature of GRS-1 was the presence of two distinct intracellular biomineralized structures: large electron-dense granules composed of calcium and long chains of magnetosomes that surround the large calcium granules. Phylogenetic analysis based on the 16S rRNA gene sequence revealed that this strain belongs to the Gammaproteobacteria and represents a new genus of MTB.

Localization and morphological variation of three bacteriome-inhabiting symbionts within a planthopper of the genus Oliarus (Hemiptera: Cixiidae).

Many planthoppers of the family Cixiidae (Hemiptera: Fulgoroidea) host three bacteriome-inhabiting bacteria: a gammaproteobacterium: 'Ca.Purcelliella pentastirinorum', a betaproteobacterium: 'Ca. Vidania fulgoroidea', and a member of the bacteroidetes: 'Ca.Sulcia muelleri'. Through light microscopy observations, DGGE PCR and FISH analysis, we examined the morphology and localization of these three endosymbionts within the abdomens of females of the planthopper Oliarus filicicola. Our results indicate a complex distribution and variation in bacterial morphologies. 'Ca. Sulcia muelleri' singularly colonize one pair of bacteriomes and have cells of irregular shape with an average diameter of approximately 4-5 µm. 'Ca.Purcelliella pentastirinorum' bacteria are roughly globular and have an average diameter of approximately 1.5-2 µm in a pair of bacteriomes located near the posterior end of the abdomen, which are surrounded by giant and highly degenerated cells of 'Ca.Vidania fulgoroidea'. In addition, 'Ca.Vidania fulgoroidea' colonizes the 'rectal organ' (sensu Buchner) and the bacterial cells appear as a small, roughly globular with an average diameter of 3 µm; whereas, 'Ca.Purcelliella pentastirinorum' infects an additional two bacteriomes and the bacterial cells appear tightly packed and highly degenerated. All three endosymbionts colocalize in the forming eggs inside the host's ovaries. Based on the abdominal distribution of bacteriomes and bacterial morphologies, we suggest that 'Ca.Vidania fulgoroidea' and 'Ca.Purcelliella pentastirinorum' correspond to the symbionts described by Buchner as the 'x-' and the 'c + d symbiont' respectively.

Abundance and single-cell activity of heterotrophic bacterial groups in the western Arctic Ocean in summer and winter.

Environmental conditions in the western Arctic Ocean range from constant light and nutrient depletion in summer to complete darkness and sea ice cover in winter. This seasonal environmental variation is likely to have an effect on the use of dissolved organic matter (DOM) by heterotrophic bacteria in surface water. However, this effect is not well studied and we know little about the activity of specific bacterial clades in the surface oceans. The use of DOM by three bacterial subgroups in both winter and summer was examined by microautoradiography combined with fluorescence in situ hybridization. We found selective use of substrates by these groups, although the abundances of Ant4D3 (Antarctic Gammaproteobacteria), Polaribacter (Bacteroidetes), and SAR11 (Alphaproteobacteria) were not different between summer and winter in the Beaufort and Chukchi Seas. The number of cells taking up glucose within all three bacterial groups decreased significantly from summer to winter, while the percentage of cells using leucine did not show a clear pattern between seasons. The uptake of the amino acid mix increased substantially from summer to winter by the Ant4D3 group, although such a large increase in uptake was not seen for the other two groups. Use of glucose by bacteria, but not use of leucine or the amino acid mix, related strongly to inorganic nutrients, chlorophyll a, and other environmental factors. Our results suggest a switch in use of dissolved organic substrates from summer to winter and that the three phylogenetic subgroups examined fill different niches in DOM use in the two seasons.

Antimicrobial peptides keep insect endosymbionts under control.

Vertically transmitted endosymbionts persist for millions of years in invertebrates and play an important role in animal evolution. However, the functional basis underlying the maintenance of these long-term resident bacteria is unknown. We report that the weevil coleoptericin-A (ColA) antimicrobial peptide selectively targets endosymbionts within the bacteriocytes and regulates their growth through the inhibition of cell division. Silencing the colA gene with RNA interference resulted in a decrease in size of the giant filamentous endosymbionts, which escaped from the bacteriocytes and spread into insect tissues. Although this family of peptides is commonly linked with microbe clearance, this work shows that endosymbiosis benefits from ColA, suggesting that long-term host-symbiont coevolution might have shaped immune effectors for symbiont maintenance.

Distinct symbiont lineages in three thyasirid species (Bivalvia: Thyasiridae) from the eastern Atlantic and Mediterranean Sea.

Thyasiridae are one of the less studied groups of chemosymbiotic bivalves. Here, we investigated symbioses in three different thyasirid species collected at three cold seeps from the Atlantic and Mediterranean. Phylogenetic analysis of bacterial 16S ribosomal RNA gene sequences demonstrated that each thyasirid species harbours a single phylotype of symbiont that belongs to a distinct lineage of putative sulphur-oxidizing Gammaproteobacteria. This result is confirmed by other marker genes (encoding 23S rRNA and APS reductase) and fluorescence in situ hybridization. This work highlights the diversity of bacteria involved in symbiosis with thyasirids and underlines the relevance of this group as a target for future symbiosis studies.

Psychromonas boydii sp. nov., a gas-vacuolate, psychrophilic bacterium isolated from an Arctic sea-ice core.

A gas-vacuolate bacterium, strain 174(T), was isolated from a sea-ice core collected from Point Barrow, Alaska, USA. Comparative analysis of 16S rRNA gene sequences showed that this bacterium was most closely related to Psychromonas ingrahamii 37(T), with a similarity of >99 %. However, strain 174(T) could be clearly distinguished from closely related species by DNA-DNA hybridization; relatedness values determined by two different methods between strain 174(T) and P. ingrahamii 37(T) were 58.4 and 55.7 % and those between strain 174(T) and Psychromonas antarctica DSM 10704(T) were 46.1 and 33.1 %, which are well below the 70 % level used to define a distinct species. Phenotypic analysis, including cell size (strain 174(T) is the largest member of the genus Psychromonas, with rod-shaped cells, 8-18 microm long), further differentiated strain 174(T) from other members of the genus Psychromonas. Strain 174(T) could be distinguished from its closest relative, P. ingrahamii, by its utilization of D-mannose and D-xylose as sole carbon sources, its ability to ferment myo-inositol and its inability to use fumarate and glycerol as sole carbon sources. In addition, strain 174(T) contained gas vacuoles of two distinct morphologies and grew at temperatures ranging from below 0 to 10 degrees C and its optimal NaCl concentration for growth was 3.5 %. The DNA G+C content was 40 mol%. Whole-cell fatty acid analysis showed that 16 : 1omega7c and 16 : 0 comprised 44.9 and 26.4 % of the total fatty acid content, respectively. The name Psychromonas boydii sp. nov. is proposed for this novel species, with strain 174(T) (=DSM 17665(T) =CCM 7498(T)) as the type strain.

Isolation and physiological characterization of two novel, piezophilic, thermophilic chemolithoautotrophs from a deep-sea hydrothermal vent chimney.

Two novel, thermophilic piezophiles, capable of chemolithoautotrophic growth, are successfully cultivated and isolated from a black smoker chimney at the TAG field (Mid Atlantic Ridge: MAR) by using a piezophilic cultivation technique. Both strains (strains 106 and 108) represent dominant cultivated populations of the microbial communities in the chimney surface habitat. Strain 106 represents typically thin, long spiral cells under the piezophilic growth condition but short bent cells under the non-piezophilic condition. It is a strictly chemolithoautotrophic gammaproteobacterium using reduced sulfur compounds as the electron donors, and nitrate and O(2) as the electron acceptors. Based on the 16S rRNA gene sequence, strain 106 would represent a novel genus of the previously uncultivated group (Symbiont Group I; a potentially novel family) within the Gammaproteobacteria, and 'Thioprofundum lithotrophica' gen. nov., sp. nov. is proposed. Strain 108 is a short, oval rod at any of the growth pressures. It is a facultative chemoautotroph, capable of both chemolithoautotrophic growth with H(2) and S oxidations and organotrophic growth with complex organics or organic acids using nitrate and O(2) as the electron acceptors. The chemolithoautotrophic growth is strictly piezophilic and under the organotrophic growth condition, it grows at conventional pressures (0.1 MPa). Strain 108 is phylogenetically distinctive from any of the previously described genera of the family Rhodobacteraceae within the Alphaproteobacteria, and 'Piezobacter thermophilus' gen. nov., sp. nov. is proposed. The piezophilic cultivation technique can be a powerful tool to isolate and characterize the previously uncultivated phylotypes in the deep-sea hydrothermal vent environments.

Neptuniibacter caesariensis gen. nov., sp. nov., a novel marine genome-sequenced gammaproteobacterium.

A Gram-negative, slightly halophilic, strictly aerobic, motile chemoorganotrophic bacterium, strain MED92(T), was isolated from a surface water sample from the eastern Mediterranean Sea. Phylogenetic analysis based on its 16S rRNA gene sequence, retrieved from the whole-genome sequence, demonstrated that this isolate is unique, showing <93 % sequence similarity to species of the families Oceanospirillaceae and Alteromonadaceae. The polar lipid profile of the novel strain consisted of phosphatidylethanolamine, phosphatidylglycerol, an unknown aminophospholipid and diphosphatidylglycerol. Major fatty acids are 16 : 1omega7c/15 iso 2-OH (41.2 % relative amount), 18 : 1omega7c (35.9 %), 16 : 0 (16.1 %), 10 : 0 3-OH (5.0 %) and 18 : 0 (1.0 %). Preferred carbon sources are organic acids and amino acids. The DNA G+C content is 46.6 mol%. Based on a phenotypic, chemotaxonomic and phylogenetic analyses, it is proposed that this marine bacterium represents a novel genus and species, for which the name Neptuniibacter caesariensis gen. nov., sp. nov. is proposed. The type strain is MED92(T) (=CECT 7075(T)=CCUG 52065(T)).

Palaeontology: undressing and redressing Ediacaran embryos.

Bailey et al. propose that the Ediacaran microfossils Megasphaera and Parapandorina, previously interpreted as animal resting eggs and blastula embryos, represent Thiomargarita-like sulphide-oxidizing bacteria, claiming that this interpretation better explains their abundance and taphonomy. Here we highlight important observations that significantly weaken the authors' conclusions.

Doushantuo embryos preserved inside diapause egg cysts.

Phosphatized microfossils in the Ediacaran (635-542 Myr ago) Doushantuo Formation, south China, have been interpreted as the embryos of early animals. Despite experimental demonstration that embryos can be preserved, microstructural evidence that the Doushantuo remains are embryonic and an unambiguous record of fossil embryos in Lower Cambrian rocks, questions about the phylogenetic relationships of these fossils remain. Most recently, some researchers have proposed that Doushantuo microfossils may be giant sulphur-oxidizing bacteria comparable to extant Thiomargarita sp. Here we report new observations that provide a test of the bacterial hypothesis. The discovery of embryo-like Doushantuo fossils inside large, highly ornamented organic vesicles (acritarchs) indicates that these organisms were eukaryotic, and most probably early cleavage stage embryos preserved within diapause egg cysts. Large acanthomorphic microfossils of the type observed to contain fossil embryos first appear in rocks just above a 632.5 +/- 0.5-Myr-old ash bed, suggesting that at least stem-group animals inhabited shallow seas in the immediate aftermath of global Neoproterozoic glaciation.

Ferrimonas futtsuensis sp. nov. and Ferrimonas kyonanensis sp. nov., selenate-reducing bacteria belonging to the Gammaproteobacteria isolated from Tokyo Bay.

Two novel mesophilic, facultatively anaerobic, selenate-reducing bacteria, designated strains FUT3661T and Asr22-7T, were isolated from a sediment sample and the alimentary tract of littleneck clams, respectively. Both sources of the samples were collected from the coast of Tokyo Bay, Japan. Cells were Gram-negative rods and motile by means of a polar flagellum. The strains reduced selenate to elemental selenium (Se0) and also reduced iron(III) oxyhydroxide, iron(III) citrate, arsenate, manganese(IV) oxide, elemental sulfur and oxygen and used lactate, pyruvate, yeast extract, tryptone and Casamino acids as electron donors and carbon sources. The strains contained both menaquinone (MK-7) and ubiquinones (Q-7 and Q-8) as isoprenoid quinones. The major fatty acids were C16:0 and C16:1omega9c. The G+C content of the genomic DNA was 58.1 mol% for strain FUT3661T and 57.2 mol% for strain Asr22-7T. Phylogenetic analysis based on 16S rRNA gene sequences revealed that the strains were related to members of the genus Ferrimonas (<94.0% similarities), although the two novel strains formed a separate lineage. 16S rRNA gene sequence similarity between strains FUT3661T and Asr22-7T was 96%. On the basis of this polyphasic analysis, it was concluded that strains FUT3661T and Asr22-7T represent two novel species within the genus Ferrimonas, for which the names Ferrimonas futtsuensis sp. nov. (type strain FUT3661T=NBRC 101558T=DSM 18154T) and Ferrimonas kyonanensis sp. nov. (type strain Asr22-7T=NBRC 101286T=DSM 18153T) are proposed.

Salicola salis sp. nov., an extremely halophilic bacterium isolated from Ezzemoul sabkha in Algeria.

A novel, extremely halophilic bacterium was isolated from brine samples collected from Ezzemoul sabkha in north-east Algeria. Cells of this isolate, designated B2T, were Gram-negative, rod-shaped and motile. Growth occurred between 10 and 25% (w/v) NaCl and the isolate grew optimally at 15-20% (w/v) NaCl. The pH range for growth was 6.0-9.0 with an optimum at pH 7.0-7.5. The predominant fatty acids were C16:0 and C18:1omega9c. Other fatty acids present were C16:1omega9c, C18:0 10-methyl, C12:0 3-OH, C10:0 and C12:0. The G+C content of the genomic DNA was 56.0 mol%. 16S rRNA gene sequence analysis indicated that strain B2T was closely related to Salicola marasensis in the Gammaproteobacteria. The level of 16S rRNA gene sequence similarity between strain B2T and the type strain of Salicola marasensis was 99%. DNA-DNA hybridization experiments between strain B2T and Salicola marasensis indicated a level of relatedness of 52%. The phenotypic characteristics of strain B2T allowed its differentiation from recognized species of the genus Salicola. Strain B2T was able to hydrolyse starch but not aesculin. It was unable to use carbohydrates and could not use citrate, pyruvate or succinate as sole carbon and energy sources. On the basis of the polyphasic data presented, strain B2T is considered to represent a novel species of the genus Salicola, for which the name Salicola salis sp. nov. is proposed. The type strain is B2T (=CECT 7106T=LMG 23122T).

High-resolution quantitative imaging of mammalian and bacterial cells using stable isotope mass spectrometry.

BACKGROUND: Secondary-ion mass spectrometry (SIMS) is an important tool for investigating isotopic composition in the chemical and materials sciences, but its use in biology has been limited by technical considerations. Multi-isotope imaging mass spectrometry (MIMS), which combines a new generation of SIMS instrument with sophisticated ion optics, labeling with stable isotopes, and quantitative image-analysis software, was developed to study biological materials. RESULTS: The new instrument allows the production of mass images of high lateral resolution (down to 33 nm), as well as the counting or imaging of several isotopes simultaneously. As MIMS can distinguish between ions of very similar mass, such as 12C15N- and 13C14N-, it enables the precise and reproducible measurement of isotope ratios, and thus of the levels of enrichment in specific isotopic labels, within volumes of less than a cubic micrometer. The sensitivity of MIMS is at least 1,000 times that of 14C autoradiography. The depth resolution can be smaller than 1 nm because only a few atomic layers are needed to create an atomic mass image. We illustrate the use of MIMS to image unlabeled mammalian cultured cells and tissue sections; to analyze fatty-acid transport in adipocyte lipid droplets using 13C-oleic acid; to examine nitrogen fixation in bacteria using 15N gaseous nitrogen; to measure levels of protein renewal in the cochlea and in post-ischemic kidney cells using 15N-leucine; to study DNA and RNA co-distribution and uridine incorporation in the nucleolus using 15N-uridine and 81Br of bromodeoxyuridine or 14C-thymidine; to reveal domains in cultured endothelial cells using the native isotopes 12C, 16O, 14N and 31P; and to track a few 15N-labeled donor spleen cells in the lymph nodes of the host mouse. CONCLUSION: MIMS makes it possible for the first time to both image and quantify molecules labeled with stable or radioactive isotopes within subcellular compartments.

Psychromonas ingrahamii sp. nov., a novel gas vacuolate, psychrophilic bacterium isolated from Arctic polar sea ice.

A gas vacuolate bacterium, designated strain 37T, was isolated from a sea ice core collected from Point Barrow, Alaska, USA. Cells of strain 37T were large (6-14 microm in length), rod-shaped, contained gas vacuoles of two distinct morphologies, and grew well at NaCl concentrations of 1-10 % and at temperatures of -12 to 10 degrees C. The DNA G+C content was 40 mol%. Whole-cell fatty acid analysis showed that 16 : 1omega7c comprised 67 % of the total fatty acid content. Phylogenetic analysis of 16S rRNA gene sequences indicated that this bacterium was closely related to members of the genus Psychromonas, with highest sequence similarity (96.8 %) to Psychromonas antarctica. Phenotypic analysis differentiated strain 37T from P. antarctica on the basis of several characteristics, including cell morphology, growth temperature range and the ability to hydrolyse polymers. DNA-DNA hybridization experiments revealed a level of relatedness of 37 % between strain 37T and P. antarctica, providing further support that it represents a distinct species. The name Psychromonas ingrahamii sp. nov. is proposed for this novel species. The type strain is 37T (=CCUG 51855T=CIP 108865T).

Thiovirga sulfuroxydans gen. nov., sp. nov., a chemolithoautotrophic sulfur-oxidizing bacterium isolated from a microaerobic waste-water biofilm.

A novel mesophilic, chemolithoautotrophic, sulfur-oxidizing bacterium, designated strain SO07(T), was isolated from a microaerobic waste-water biofilm. Chemolithoautotrophic growth was observed with elemental sulfur, sulfide and thiosulfate as sole electron donors and oxygen as electron acceptor. Anaerobic and heterotrophic growth were not observed. Nitrate was not used as a terminal electron acceptor. The optimum pH and temperature for growth were pH 7.5 and 30 degrees C, respectively. The major isoprenoid quinone was Q-8. The DNA G + C content of strain SO07(T) was 47.1 mol%. Phylogenetic analysis of 16S rRNA gene sequences demonstrated that strain SO07(T) formed a monophyletic group in the gamma-Proteobacteria with only 89 % similarity to members of the genus Halothiobacillus, its nearest phylogenetic neighbours. In addition, the isolate differed from members of the genus Halothiobacillus in its requirement for and tolerance of NaCl; strain SO07(T) was unable to grow in NaCl concentrations of more than 180 mM. On the basis of phylogenetic, chemotaxonomic and physiological data, it is proposed that isolate SO07(T) (=JCM 12417(T) = ATCC BAA-1033(T)) represents the type strain of a novel species in a new genus, Thiovirga sulfuroxydans gen. nov., sp. nov.

Colwellia piezophila sp. nov., a novel piezophilic species from deep-sea sediments of the Japan Trench.

Two strains of obligately piezophilic bacteria were isolated from sediment collected from the bottom surface of a small canyon on the seaward slope of the Japan Trench at a depth of 6278 m. The isolated strains, Y223GT and Y251E, are closely affiliated with members of the genus Colwellia on the basis of 16S rRNA gene sequence analysis. The G + C contents of both strains were about 39 mol%. DNA-DNA hybridization values between these strains and Colwellia reference strains were significantly lower than those accepted as the phylogenetic definition of a species. The novel strains are Gram-negative, polarly flagellated and facultatively anaerobic. The optimal pressure for growth was 60 MPa at both 4 and 10 degrees C; the most rapid growth rate was observed at 10 degrees C and 60 MPa. No growth occurred at 15 degrees C under any pressure studied. The major isoprenoid quinone is Q-8. The predominant cellular fatty acids are C16 : 0 and C16 : 1. Based on the taxonomic differences observed, the isolated strains appear to represent a novel obligately piezophilic Colwellia species. The name Colwellia piezophila sp. nov. (type strain Y223GT = JCM 11831T = ATCC BAA-637T) is proposed.

Idiomarina fontislapidosi sp. nov. and Idiomarina ramblicola sp. nov., isolated from inland hypersaline habitats in Spain.

Two bacterial strains, F23T and R22T, have been isolated from hypersaline habitats in Málaga (S. Spain) and Murcia (E. Spain). The novel strains, similar to previously described Idiomarina species, are slightly curved rods, Gram-negative, chemo-organotrophic, strictly aerobic and motile by a single polar flagellum. Both strains produce catalase and oxidase. They hydrolyse aesculin, gelatin, casein, Tween 20, Tween 80 and DNA but not starch or tyrosine. The strains differ from the hitherto described Idiomarina species in their capacity to produce extracellular polysaccharides and their different patterns of carbon sources and antimicrobial susceptibility. They are moderate halophiles capable of growing in NaCl concentrations of 0.5 to 25 % w/v, the optimum being 3-5 % w/v. Cellular fatty acids are predominantly iso-branched. The main fatty acids in strain FP23T are 15 : 0 iso (26.75 %), 16 : 1omega7c (11.33 %) and 16 : 0 (11.73 %) whilst 15 : 0 iso (24.69 %), 17 : 0 iso (12.92 %) and 17 : 1omega9c (11.03 %) are predominant in strain R22T. The DNA G + C composition is 46.0 mol% in strain FP23T and 48.7 mol% in strain R22T. Phylogenetic analyses indicate conclusively that the two strains belong to the genus Idiomarina. DNA-DNA hybridization revealed that they represent novel species. In the light of the polyphasic evidence accumulated in this study, it is proposed that they be classified as novel species of the genus Idiomarina, with the names Idiomarina fontislapidosi sp. nov. (type strain F23T = CECT 5859T = LMG 22169T) and Idiomarina ramblicola sp. nov. (type strain R22T = CECT 5858T = LMG 22170T).