Oleispira antarctica gen. nov., sp. nov., a novel hydrocarbonoclastic marine bacterium isolated from Antarctic coastal sea water The taxonomic characteristics of two bacterial strains, RB-8 T and RB-9, isolated from hydrocarbondegrading enrichment cultures obtained from Antarctic coastal marine environments (Rod Bay, Ross Sea), were determined. These bacteria were psychrophilic, aerobic and Gram-negative with polar flagella. Growth was not observed in the absence of NaCl, occurred only at concentrations of Na + above 20 mM and was optimal at an NaCl concentration of 3-5 % (w/v). The major cellular fatty acids were monounsaturated straight-chain fatty acids. The strains were able to synthesize the polyunsaturated fatty acid eicosapentaenoic acid (20 : 5v3) at low temperatures. The DNA G+C contents were 41-42 mol%. The strains formed a distinct phyletic line within the c-Proteobacteria, with less than 89?6 % sequence identity to their closest relatives within the Bacteria with validly published names. Both isolates exhibited a restricted substrate profile, with a preference for aliphatic hydrocarbons, that is typical of marine hydrocarbonoclastic micro-organisms such as Alcanivorax, Marinobacter and Oleiphilus. On the basis of ecophysiological properties, G+C content, 16S rRNA gene sequences and fatty acid composition, a novel genus and species within the c-Proteobacteria are proposed, Oleispira antarctica gen. nov., sp. nov.; strain RB-8 T (=DSM 14852 T =LMG 21398 T ) is the type strain. INTRODUCTIONHydrocarbon-degrading micro-organisms usually exist in very low abundance in the absence of oil pollution. A pollution event is rapidly followed by a bloom of these micro-organisms, the populations of which expand to nearly complete dominance of the viable microbial community during the period of contamination (Margesin & Schinner, 1999;Harayama et al., 1999). The properties of hydrocarbon compounds depend on the ambient temperature. Short-chain alkanes become less volatile and more water-soluble at low temperatures, whereas longer-chain compounds precipitate under cold conditions as waxes, respectively rendering them bioavailable and inaccessible to microbes. Such behaviour at low temperatures obviously reflects the establishment of specific oil-based marine microbial communities at these temperatures that are somehow different from those observed in a temperate climate. The most important permanently cold habitat is the ocean, since the temperature of more than 90 % of the seawater volume is below 5 uC. Genera that are typically well represented in cold, petroleum-contaminated sites are Acinetobacter, Arthrobacter, Mycobacterium, Pseudomonas, Rhodococcus and Sphingomonas, many of which can grow solely on hydrocarbon compounds and have been previously characterized as petroleum degraders of terrestrial origin (Rosenberg et al., 1992;MacCormack & Fraile, 1997). Although the role of these microbes is evident in the petroleum-degradation process in cold marine environments, Abbreviations: PLFA, phospholipid fatty acid; ...

An ecological study on distribution of Antarctic bacterial communities was determined by 16S-based phylogenetic analyses of clone libraries derived from RNA and DNA extracted from two different marine areas and compared between each other. Superficial seawater samples were collected from four stations in Ross Sea, three of them located in Rod Bay and one in Evans Cove; for each station two clone libraries (16S rDNA and 16S rRNA) were prepared and evident divergences between DNA and RNA libraries of each site were obtained. Of all phylotypes 93.6% were found in RNA libraries; in contrast, only 31 phylotypes (70.5%) were retrieved from total microbial community (DNA libraries). DNA and RNA sequences related to gamma-Proteobacteria and Bacteroidetes groups, typical for Antarctic sea-ice bacterial communities, were detected in analysed sites. 16S rDNA and rRNA libraries derived from the two different areas were enriched by picophytoplanktonic 16S sequences of plastid and mitochondrion origins, reflecting that the algal blooms occurred during sampling (Antarctic summer 2003). The finding in Rod Bay libraries of high percentage of DNA clones apparently affiliated with beta-Proteobacteria typical for activated sludges and well water could be explained by the presence of a sewage depuration system at this site. Obtained results clearly demonstrate that combination of 16S rDNA and 16S rRNA gene sequencing is preferred approach to have a more reliable vision on the composition of microbial communities.

Aims: The effects of different growth media and temperature on production of polyunsaturated fatty acids (PUFA) by Shewanella sp. GA‐22 were investigated. The attempts to characterize the GA‐22 genes, homologous to those of PUFA biosynthesis gene cluster, was performed. Methods and Results: Physiological and phylogenetic characterization of new Antarctic isolate GA‐22 was performed. Total fatty acids were isolated from the cells growing under different conditions and analysed by gas chromatography–mass spectrometry (GC–MS). Using degenerated primers derived from the conserved regions within PUFA fatty acid synthase operons, five fragments of homological genes were amplified from GA‐22 DNA, and two of them corresponding to pfaA and pfaC synthase subunits were sequenced. Conclusions: Strain GA‐22 was shown to be able to produce three different PUFA: linoleic, arachidonic and eicosapentaenoic acids. The PUFA production was temperature‐ and carbon source‐dependent. The deduced gene products exhibited high similarity to corresponding fatty acid synthases PfaA and PfaC. Significance and Impact of Study: The PUFA production was detected on media supplemented with crude oil, gasoline and n‐tetradecane. The apparent conservation of PUFA genes may point to the potential utilization of designed primers as functional markers in culture‐independent ecological studies, and for initial screening in biotechnological fields.

For preliminary screening of human impact on Antarctic coasts, the compositions of microbial communities were analyzed in seawater at two sites located in the Terra Nova Bay of Antarctica (Ross Sea) by a combination of 16S rRNA gene sequencing and culture techniques. The bacterial community in the sample from the Rod Bay site, located at the proximity to the Italian Station, was characterized by a high abundance of 16S rRNA gene sequences belonging to the microflora typically found in soil and freshwater environments. In contrast, the seawater sample from the Adelie Cove station, a pristine reference site, contained 16S rRNA gene sequences typically found in marine areas affected by algal blooms and sea ice decay. The addition of crude oil to the Rod Bay seawater sample rapidly induced a shift in the composition of the bacterial community with appearance of novel taxonomic groups and a dramatic increase in the relative abundance of gamma-Proteobacteria sequences, whereas no significant changes were detected in the bacterial community of the Adelie Cove sample under the same conditions. Bacteria-exhibiting features with potential interest for industrial and environmental applications were isolated from the Rod Bay oil-enriched sample. In particular, hydrocarbon-degrading, cold-adapted bacteria were selectively enriched, isolated and screened for their ability to synthesize polyunsaturated fatty acids. Twenty two bacterial strains were isolated from the oil enrichment culture and identified. Eighteen isolates were found to be members of gamma-Proteobacteria, while the remainder were representatives of alpha-Proteobacteria, CFB and high G + C divisions.

Alcalilimnicola halodurans gen. nov., sp. nov., an alkaliphilic, moderately halophilic and extremely halotolerant bacterium, isolated from sediments of soda-depositing Lake Natron, East Africa Rift Valley An alkaliphilic, halotolerant, Gram-negative, heterotrophic, aerobic and rodshaped organism was isolated from drying soda and at a water-covered site of Lake Natron, Tanzania, by means of the most-probable-number technique developed for anoxygenic, phototrophic sulfur bacteria. It had an absolute requirement for alkalinity, but not for salinity ; growth occurred at salt concentrations of 0-28 % (w/v), with optimal growth at 3-8 % (w/v) NaCl. The bacterium preferentially metabolized volatile fatty acids and required vitamins for growth. The name Alcalilimnicola halodurans gen. nov., sp. nov. is proposed for the novel isolate, placed in the γ-Proteobacteria within the family Ectothiorhodospiraceae on the basis of analysis of the 16S rDNA sequence, polar lipids, fatty acids and DNA base composition. Although Alcalilimnicola halodurans is closely related to the extreme anoxygenic, phototrophic sulfur bacteria of the genus Halorhodospira, it is not phototrophic.

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The aims of this study were: (i) the characterization of the structure of the indigenous microbial community associated with the sediments under study; (ii) the isolation and characterization of microbial consortia able to degrade the aged hydrocarbons contaminating the sediments, and (iii) the assessment of related biodegradation capability of selected consortia. Samples of surface sediments were collected in Priolo Gargallo harbour (Sicily, Italy). The samples were analysed for physical, chemical (GC-FID analysis) and microbiological characteristics (qualitative (16S rDNA clone library) and quantitative (DAPI, CFU and MPN count) analysis). The sediment samples were used for the selection of two microbial consortia (indicated as PSO and PSM) with high biodegradation capacity for crude oil ( $ 95%) and PAHs ( $ 63%) respectively. Genetic analysis showed that Alcanivorax and Cycloclasticus were the dominant genera in both the PSO and PSM consortia. Oil-polluted environments naturally develop an elevated biorecovery potential. The presence of a highly specialized microbial flora (adapted to support the contamination) and their stimulation through favourable induced conditions provides a promising recovery strategy. The chance to identify and select indigenous bacteria and/or consortia with a high biodegradation capacity is fundamental for the development and optimization of bioaugmentation strategies especially for those concerning in situ applications. Highlights• Chemical and physical characterization of marine sediments.

Abstract:The central Mediterranean Sea is among the most oligotrophic habitats in the marine environment. In this study, we investigated the abundance, diversity and activity of prokaryoplankton in the water column (25-3000-m depth) at Station Vector (Tyrrhenian Sea, 39°32.050′ N; 13°22.280′ E). This specific water column consists of three different water masses (Modified Atlantic Water (MAW), Levantine Intermediate Water (LIW) and Tyrrhenian Deep Water (TDW), possessing a typical stratification of the Central Mediterranean basin. CARD-FISH showed that the metabolically-active fraction of bacterial populations exceeded the archaeal fraction along the whole water column, except OPEN ACCESS J. Mar. Sci. Eng. 2015, 3 846 at the deepest water masses. 16S rDNA and 16S rRNA clone libraries obtained from each type of water mass were used to analyse the prokaryoplankton community structure and to distinguish between active and "less active" microbial fractions. Our results showed that the rRNA-derived bacterial libraries seemed to be more depth specific compared to 16S rDNA-derived counterparts. Major differences were detected between the active fractions of bacterioplankton thriving in photic (25 m, MAW) and aphotic layers (500-3000 m, LIW and TDW respectively), whereas no statistically-significant differences were detected within the deep, aphotic layers (500-3000 m, LIW and TDW). Archaeal communities possessed more depth-specific distribution patterns with both total and active fractions showing depth stratification. Cyanobacteria and Marine Group II MAGII of Euryarchaea dominated the MAW prokaryoplankton. A notable fraction of Geitlerinema-related cyanobacteria was detected among the metabolically-active bacterial population recovered from the mesopelagic (500 m, LIW) aphotic layer, which is indicative of their mixotrophic behaviour. Heterotrophic Gammaproteobacteria and members of Marine Group 1.1a and the PSL12-related ALOHA group of Thaumarchaeota were both abundant in the aphotic layers (both LIW and TDW). Discrepancies observed between 16S rDNA-and 16S rRNA-based libraries are most likely linked to different physiological states of the prokaryoplankton community members recovered from different layers. Taking into account the relative homogeneity of the main physicochemical parameters throughout the whole water column, light and energy source are likely the most relevant environmental variables shaping microbial biodiversity through the Station Vector water column.