The world's largest omnivore is a fish.

The evolution of very large body size requires a ubiquitous and abundant source of food. In marine environments, the largest animals such as whale sharks are secondary consumers that filter feed on nekton, which is plentiful, although patchy. Consequently, feeding in coastal environments requires cost-efficient foraging that focuses on oceanographic features that aggregate both nektonic prey and marine debris such as floating macroalgae. Consumption of this algae could present an energetic challenge for these animals, unless some component can be digested. Here, we use a multi-technique approach involving amino acid compound-specific stable isotope analysis (CSIA) and fatty acid analysis to determine the trophic level of whale sharks and to identify likely items in the diet. CSIA analyses showed that the species has a trophic level consistent with omnivory. Fatty acid profiles of whale shark tissues, feces and potential prey items suggest that the floating macroalgae, Sargassum, and its associated epibionts is a significant source of food. Although this overcomes the energetic challenge of consumption of floating algae, this mode of feeding and the need to focus on oceanographic features that aggregate prey also increases the threat to the species posed by pollutants such as plastic.

Dispersal and assimilation of an aquaculture waste subsidy in a low productivity coastal environment.

To understand dispersal and assimilation of aquaculture waste subsidies in a naturally low-productivity environment, we applied a novel, rapid transmethylation technique to analyse sediment and biota fatty acid composition. This technique was initially validated at Atlantic salmon farms in Macquarie Harbour, Australia, where sediments were collected at farm and control locations. Subsequently, sediment, benthic polychaete and zooplankton were sampled at sites 0, 50, 250, 500 and 1000m distant from multiple cages. Results demonstrated an acute deposition zone up to 50m from cages and a diffuse zone extending 500m from cages. Changes in sediment concentration of linoleic acid, oleic acid and total fatty acids were effective tracers of farm deposition. Bacterial biomarkers indicated that aquaculture waste stimulates bacterial productivity in sediments, with elevated biomarker concentrations also detected in benthic polychaetes. Overall, fatty acid analysis was a sensitive technique to characterize the benthic footprint of aquaculture influence.

Unusually high levels of n-6 polyunsaturated fatty acids in whale sharks and reef manta rays.

Fatty acid (FA) signature analysis has been increasingly used to assess dietary preferences and trophodynamics in marine animals. We investigated FA signatures of connective tissue of the whale shark Rhincodon typus and muscle tissue of the reef manta ray Manta alfredi. We found high levels of n-6 polyunsaturated fatty acids (PUFA), dominated by arachidonic acid (20:4n-6; 12-17 % of total FA), and comparatively lower levels of the essential n-3 PUFA-eicosapentaenoic acid (20:5n-3; ~1 %) and docosahexaenoic acid (22:6n-3; 3-10 %). Whale sharks and reef manta rays are regularly observed feeding on surface aggregations of coastal crustacean zooplankton during the day, which generally have FA profiles dominated by n-3 PUFA. The high levels of n-6 PUFA in both giant elasmobranchs raise new questions about the origin of their main food source.

Variation in the fatty acid composition of blubber in Cape fur seals (Arctocephalus pusillus pusillus) and the implications for dietary interpretation.

Analysis of the fatty acid (FA) composition of blubber is a valuable tool in interpreting the diet of marine mammals. This technique is based on the principle that particular FA present in prey can be incorporated largely untransformed into predator adipose tissue stores, thereby providing biochemical signatures with which to identify prey species. Several studies of phocid seals and cetaceans have documented vertical stratification in the FA composition of blubber such that inferences about diet may vary greatly depending on the layer of the blubber that is analysed. It is not known whether blubber in otariid seals (fur seals and sea lions) also displays vertical stratification in FA composition. Furthermore, it is not known whether the FA composition of blubber is uniform in these species. In the present study, the vertical and regional variation in FA composition of blubber was investigated in seven adult female Cape fur seals (Arctocephalus pusillus pusillus). The proportion of monounsaturated fatty acids (MUFA) was greater in the outer (43.6+/-1.3%) than inner portion (40.9+/-1.2%; t(20)=5.59, P<0.001) whereas the proportions were greater in the inner than outer portions for saturated fatty acids (23.6+/-0.5% and 21.9+/-0.6%, respectively, t(20) = 5.31, P<0.001) and polyunsaturated fatty acids (PUFA, 35.5+/-0.7% and 34.5+/-0.7%, respectively, t(20) = 3.81, P < 0.001). There was an inverse relationship between MUFA and PUFA in the blubber, independent of sampling location. In addition, with the exception of the inner portion from non-lactating females, blubber from the mammary area had the highest proportions of 18:1omega9c and total MUFA, followed by blubber from the rump and neck, suggesting that the deposition and mobilisation of blubber lipids may not be uniform around the body in otariid seals. These results support the need for blubber tissue to be sampled from the same site on animals, and to the full depth of the blubber layer, to minimise variation in FA profiles that could occur if different sites and depths were sampled. Such standardisation of sampling will further aid in interpreting diet in otariid seals using the FA Signature Analysis approach.

Influences of dietary n-3 long-chain PUFA on body concentrations of 20:5n-3, 22:5n-3, and 22:6n-3 in the larvae of a marine teleost fish from Australian waters, the striped trumpeter (Latris lineata).

We determined the effect of dietary long-chain (> or = C20) PUFA (LC-PUFA), 20:5n-3 and 22:6n-3, on larval striped trumpeter (Latris lineata) biochemistry through early development and during live feeding with rotifers (Brachionus plicatilis). Rotifers were enriched using seven experimental emulsions formulated with increasing concentrations of n-3 LC-PUFA, mainly 20:5n-3 and 22:6n-3. Enriched rotifer n-3 LC-PUFA concentrations ranged from 10-30 mg/g dry matter. Enriched rotifers were fed to striped trumpeter larvae from 5 to 18 d post-hatch (dph) in a short-term experiment to minimize gross deficiency symptoms such as poor survival that could confound results. No relationships were observed between larval growth or survival with dietary n-3 LC-PUFA at 18 dph. The larval FA profiles generally reflected those of the rotifer diet, and significant positive regressions were observed between most dietary and larval FA at 10, 14, and 18 dph. The major exception observed was an inverse relationship between dietary and larval 22:5n-3. The presence of 22:5n-3 in elevated amounts when dietary 22:6n-3 was depressed suggests that elongation of 20:5n-3 may be occurring in an attempt to raise body concentrations of 22:6n-3. We hypothesize that accumulation of 22:5n-3 might be an early indicator of 22:6n-3 deficiency in larval fish that precedes a reduction in growth or survival. A possible role of 22:5n-3 as a biochemical surrogate for 22:6n-3 is discussed.

Prokaryotic metabolic activity and community structure in Antarctic continental shelf sediments.

The prokaryote community activity and structural characteristics within marine sediment sampled across a continental shelf area located off eastern Antarctica (66 degrees S, 143 degrees E; depth range, 709 to 964 m) were studied. Correlations were found between microbial biomass and aminopeptidase and chitinase rates, which were used as proxies for microbial activity. Biomass and activity were maximal within the 0- to 3-cm depth range and declined rapidly with sediment depths below 5 cm. Most-probable-number counting using a dilute carbohydrate-containing medium recovered 1.7 to 3.8% of the sediment total bacterial count, with mostly facultatively anaerobic psychrophiles cultured. The median optimal growth temperature for the sediment isolates was 15 degrees C. Many of the isolates identified belonged to genera characteristic of deep-sea habitats, although most appear to be novel species. Phospholipid fatty acid (PLFA) and isoprenoid glycerol dialkyl glycerol tetraether analyses indicated that the samples contained lipid components typical of marine sediments, with profiles varying little between samples at the same depth; however, significant differences in PLFA profiles were found between depths of 0 to 1 cm and 13 to 15 cm, reflecting the presence of a different microbial community. Denaturing gradient gel electrophoresis (DGGE) analysis of amplified bacterial 16S rRNA genes revealed that between samples and across sediment core depths of 1 to 4 cm, the community structure appeared homogenous; however, principal-component analysis of DGGE patterns revealed that at greater sediment depths, successional shifts in community structure were evident. Sequencing of DGGE bands and rRNA probe hybridization analysis revealed that the major community members belonged to delta proteobacteria, putative sulfide oxidizers of the gamma proteobacteria, Flavobacteria, Planctomycetales, and Archaea. rRNA hybridization analyses also indicated that these groups were present at similar levels in the top layer across the shelf region.

The isolation and use of iron-oxidizing, moderately thermophilic acidophiles from the Collie coal mine for the generation of ferric iron leaching solution.

Moderately thermophilic, iron-oxidizing acidophiles were enriched from coal collected from an open-cut mine in Collie, Western Australia. Iron-oxidizers were enriched in fluidized-bed reactors (FBR) at 60 degrees C and 70 degrees C; and iron-oxidation rates were determined. Ferrous iron oxidation by the microbiota in the original coal material was inhibited above 63;C. In addition to four iron-oxidizers, closely related to Sulfobacillus spp that had been earlier isolated from the 60 degrees C FBR, one heterotroph closely related to Alicyclobacillus spp was isolated. The Alicyclobacillus sp. isolated from the Collie coal mine tolerated a lower pH than known Alicyclobacillus spp and therefore may represent a new species. The optimum temperature for growth of the iron-oxidizing strains was approximately 50 degrees C and their maximum temperatures were approximately 60 degrees C. The FBR was adjusted to operate at 50 degrees C and was inoculated with all of the isolated iron-oxidizing strains. At 60 degrees C, an iron-oxidation rate of 0.5 g Fe(2+) l(-1) x h(-1) was obtained. At 50 degrees C, the iron-oxidation rate was only 0.3 g Fe(2+) l(-1) x h(-1). These rates compare favourably with the iron-oxidation rate of Acidianus brierleyi in shake-flasks, but are considerably lower than mesophilic iron-oxidation rates.

Potential of thraustochytrids to partially replace fish oil in Atlantic salmon feeds.

The replacement of fish oil with a dried product made from thraustochytrid culture, a marine microorganism, in canola-oil-based diets for Atlantic salmon was investigated. Salmon (37 g) were fed for 51 days on diets containing only canola oil, canola oil and fish oil, or canola oil and the thraustochytrid. There were no significant differences in final weight (106.1 +/- 1.1 g), weight gain (69.6 +/- 1.1 g), feed consumption (16.5 +/- 0.2 mg dry matter g(-1) d(-1)), feed efficiency ratio (1.15 +/- 0.03 g (g-1)), or productive protein value (51.2% +/- 1.7%) between the diets. Nor were there any significant differences in whole-body chemical composition, organ somatic indices, or measures of immune function. However, following transfer to seawater and 2 challenges with Vibrio anguillarum, cumulative mortality was significantly lower in fish fed some fish oil than in those fed the 2 diets containing no fish oil. In conclusion, the thraustochytrid had no detrimental effects on the performance of salmon but, at the current inclusion of 10%, failed to confer the same effect as fish oil under challenging conditions.

Lipid reserves used by pueruli of the spiny lobster Jasus edwardsii in crossing the continental shelf of New Zealand.

Lipid is known to fuel the movement of the nektonic puerulus stage of the spiny lobster Jasus edwardsii across the continental shelf of New Zealand. Lipid class analysis of pueruli caught from two locations across the continental shelf showed that phospholipid predominated (86-96% of total lipid), with only smaller proportions of sterol (0.9-8.7%) and diacylglycerol (1.2-7.6%). Only traces of triacylglycerol, hydrocarbon and wax ester were present (<0.1% of total lipid). Comparison of the lipid class content of pueruli caught onshore and offshore showed that phospholipid reserves are primarily utilised during this important phase in the lifecycle and that diacylglycerol plays a less significant secondary role. Histology identified concentrations of phospholipid in fat bodies located in the haemocoel. The use of phospholipid as the dominant storage medium in the puerulus stage is unlike many other marine taxa, including crustacea, which tend to use triacylglycerol and wax ester. The use of phospholipid as a storage medium may well be related to its characteristic transparency, an important feature of this nektonic stage of lobster development that is highly vulnerable to pelagic visual predators.

Interannual variations in the lipids of the Antarctic pteropods Clione limacina and Clio pyramidata.

Antarctic pteropods, Clione limacina (Order Gymnosomata) and Clio pyramidata (order Thecosomata), were collected near Elephant Island, South Shetland Islands, during 1997 and 1998. Total lipid was high in C. limacina (29--36 mg g(-1) wet mass) and included 46% of diacy1glyceryl ether (DAGE, as % of total lipid) for both 1997 and 1998. DAGE was not detected in C. pyramidata, which had mainly polar lipid and triacy1glycerol. 1-O-Alkyl glyceryl ethers (GE) derived from the DAGE consisted primarily of 15:0 and 16:0, with lower 17:0 and a17:0. The principal sterols of both pteropods included trans-dehydrocholesterol, brassicasterol, 24-methylenecholesterol, cholesterol and desmosterol. Levels of 24-methylenecholesterol and desmosterol were lower in both pteropods in 1997 compared to 1998. C. limacina had high levels of the odd-chain fatty acids 17:1(n--8)c and 15:0 in contrast to C. pyramidata. The previously proposed source of elevated odd-chain fatty acids in C. limacina is via propionate derived from phytoplankton DMPT; another possible source may be from thraustochytrids, which are common marine microheterotrophs. C. pyramidata had twice as much PUFA as C. limacina, largely due to higher 20:5(n--3). The PUFA 18:5(n--3) and very long chain fatty acids (C(24), C(26) and C(28) VLC-PUFA) were only detected in 1998 pteropods. In comparison, 1996 samples of C. limacina contained lower DAGE levels, which also may reflect differences in diet and oceanographic conditions. Interannual variations in specific lipid biomarkers are discussed with respect to possible different phytoplankton food sources available in the AMLR survey area.

Sterol and squalene content of a docosahexaenoic-acid-producing thraustochytrid: influence of culture age, temperature, and dissolved oxygen.

Thraustochytrid strain ACEM 6063, rich in omega-3 polyunsaturated fatty acids, was cultured at 15 degrees C and 20 degrees C in high (>40%) and low (<5%) dissolved oxygen (DO), and at 25 degrees C in low-DO media. Samples were taken 4, 2, and 0 days before each culture reached peak biomass (T(-4), T(-2), and T(p), respectively). Twenty sterols, 13 of which were identified, were detected. Predominant were cholest-5-en-3 beta-ol, 24-ethylcholesta-5,22E-dien-3 beta-ol, 24-methylcholesta-5,22E-dien-3 beta-ol, and 2 coeluting sterols, one of which was 24-ethylcholesta-5,7,22-trien-3 beta-ol. These 4 sterols comprised 50% to 90% of total sterols. Cultures grown at high DO had simpler sterol profiles than those grown at low DO. Only the 4 sterols mentioned above were present at more than 3% of total sterols in high-DO cultures. In low-DO cultures, up to 6 additional sterols were present at more than 3% of total sterols. Culture age, temperature, and DO influenced squalene and sterol content. Total sterols (as a proportion of total lipids) decreased with increasing culture age. If organisms such as ACEM 6063 are to be used for commercial production of lipid products for human consumption, both their sterol content and factors influencing sterol production need to be characterized thoroughly.

Unusually high levels of non-saponifiable lipids in the fishes escolar and rudderfish identification by gas and thin-layer chromatography.

Analysis of the non-saponifiable lipids of the fishes Lepidocybium flavobrunneum and Ruvettus pretiosus (escolar), and Centrolophus niger and Tubbia spp. (rudderfish) was performed. The analyses were used to clarify the cause of recent reports of illness (diarrhoea) in Australia from consumption of purported rudderfish. Both escolar and rudderfish contained very high levels of oil (generally between 14 to 25%, as % wet mass) in the fillet and the oil compositions were different to most seafood. Escolar oil contained mainly wax ester (>90% of oil). The oil from five specimens of rudderfish contained mainly diacylglyceryl ether (DAGE, >80% of oil) or hydrocarbon (>80% of oil, predominately squalene). One rudderfish specimen contained mainly polar lipid. Major differences in oil content and composition, including fatty alcohol and glyceryl ether diols (derived from DAGE), were observed between purported individuals of the same species or related species of rudderfish, raising the possibility of geographic or seasonal differences affecting the oil composition. The oil composition of fish fillet samples associated with the health issues were consistent with the profiles for escolar, rather than rudderfish species. These findings, in particular the lipid class and fatty alcohol profiles, were supported by general protein fingerprinting results and were consistent with the samples originating from individuals of the escolar species L. flavobrunneum. The high wax ester content of the escolar group clarifies the reported diarrhoeal effects to consumers. Purgative properties of high wax ester containing fish oils have been reported for escolar and other species. The results highlight the potential for non-saponifiable lipid profiles to be used for identification of fish fillets and oils to at least group level.

Evaluation of extraction methods for recovery of fatty acids from lipid-producing microheterotrophs.

The effect of different extraction techniques on the recovery of fatty acids from freeze-dried biomass of two lipid-producing microheterotrophs was examined. Two procedures were used: the extraction of lipids from biomass followed by transesterification of the fatty acids (extraction-transesterification); and the direct transesterification of biomass to produce fatty acid methyl esters (i.e. without the initial extraction step). Variable factors in the extraction-transesterification experiment were the sequence in which solvents were added to the samples, the relative amount of methanol in the solvent mix, and sonication of biomass while in the solvent mix. Variable factors in the direct transesterification experiment were sample size, and reaction duration. Statistical analysis of data (level of significance P<0.05) showed that: (1) extraction of total fatty acids prior to transesterification was significantly more efficient when solvents were added in the order of increasing polarity; (2) neither sonication nor increasing the proportion of methanol in the extraction solvent significantly affected extraction of fatty acids prior to transesterification; (3) efficiency of direct transesterification of fatty acids increased significantly with reaction time; (4) efficiency of direct transesterification of fatty acids was not significantly affected by sample size; (5) the most efficient method for extraction of fatty acids prior to transesterification yielded significantly less fatty acids than the most effective direct transesterification method. While the study examined only two strains, our results suggest that fatty acid analysis methodology for microheterotrophs under consideration for biotechnological exploitation requires optimisation and validation.

Lipids of gelatinous Antarctic zooplankton: Cnidaria and Ctenophora.

Cnidaria (Calycopsis borchgrevinki, Diphyes antarctica, Stygiomedusa gigantea, Atolla wyvillei, Dimophyes arctica) and Ctenophora (Beroe cucumis, B. forskalii, Pleurobrachia pileus, Bolinopsis infundibulum) were collected near Elephant Island, South Shetland Islands, during January and February 1997 and 1998. Total lipid was low in all zooplankton (0.1-5 mg g wet mass) and included primarily polar lipids (59-96% of total lipid). Triacylglycerols were 0-26% of total lipids, and wax esters were 0-11% in all species. Cholesterol was the major sterol in all Cnidaria (50-63% of total sterols) whereas in most ctenophores it was lower at 26-45%. These cholesterol levels are consistent with a combined carnivorous and phytoplanktivorous diet in the ctenophores, with the carnivorous diet more dominant in the Cnidaria. Other sterols included primarily trans-dehydrocholesterol, desmosterol, 24-methylcholest-5,22E-dien-3beta-ol, 24-nordehydrocholesterol, and 24-methylenecholesterol. Total stanols were 0-6% in all zooplankton. Eicosapentaenoic acid and docosahexaenoic acid were the major polyunsaturated fatty acids (PUFA) in all samples (7-25% of total fatty acids) except for A. wyvillei in which docosapentaenoic acid was 10% of total fatty acids. The PUFA 18:5n-3 was not detected in 1997 samples, but constituted 0.2-0.8% in most 1998 samples. Monounsaturated fatty acids included primarily 18:1n-9c, 16:1n-7c, and 18:1n-7c. The principal saturated fatty acids in all samples were 16:0, 18:0, and 14:0. These data are the first for many of these zooplankton species and the first sterol data for most species. The use of the signature lipid approach has enabled examination of aspects of trophodynamics not obtainable by conventional techniques.

Lipid composition of the liver oil of deep-sea sharks from the Chatham Rise, New Zealand.

Deep-sea sharks approach neutral buoyancy by means of a large liver that contains large amounts of low-density lipids, primarily squalene and diacyl glyceryl ether (DAGE). As an animal increases in size and matures sexually, many biochemical changes take place within the animal. It was hypothesized that maintenance of neutral buoyancy in deep-sea sharks involves fine-scale changes in the chemical composition of the liver oil as individual sharks grow and develop. To test this hypothesis, the lipid composition of liver oil for individuals of different size and sex of deep-sea sharks from the Chatham Rise, New Zealand was compared. The composition of liver oil varied within and among species. Several species contained large amounts of squalene and DAGE, whereas only traces of these lipids were present in other species. The amounts of squalene and DAGE in liver oil were inversely related, and squalene content tended to decrease as sharks increased in size. Species with high squalene levels (> 80%) in liver oil were not abundant on the Chatham Rise, although levels of DAGE (a lipid of increasing commercial interest) were elevated in many species. Maintenance of neutral buoyancy in deep-sea sharks appears to involve changes in the composition of low-density liver lipids as the sharks increase in size and mature.

The occurrence and possible significance of diacylglyceryl ether lipids in abalone.

Gonadal and foot tissues of the green abalone, Haliotis fulgens, farm-raised on macroalgal [corrected] diets, were analyzed for lipids using thin-layer chromatography/flame-ionization detection and gas chromatography-mass spectrometry (GC-MS). Diacylglyceryl ether (DAGE) was 0.7% of total lipids in the gonad. The major alkyl constituents of the glyceryl ether diols in the gonad (as % of total diols) were 16:0 (38%) and 18:1 (36%). While levels of DAGE in the abalone foot were below flame-ionization detection limits, glyceryl ether diols from them were detected using the more sensitive GC-MS procedure. The major diol components in the foot were 18:0 (39%) and 18:1 (32%). To our knowledge, this is the first report of DAGE in abalone tissues. Although the precise role of DAGE in abalone remains to be determined, a possible structural role may exist.

Flavobacterium hibernum sp. nov., a lactose-utilizing bacterium from a freshwater Antarctic lake.

Four freshwater Antarctic lakes were examined for the presence of beta-galactosidase-producing bacteria using mineral medium enrichments and lactose. Enrichments from only one of the lakes produced growth and two strains were isolated that were very similar in phenotype and fatty acid profile, and shared considerable homology in their DNA (DNA-DNA hybridization = 93 +/- 7%). The strains were psychrotrophic with theoretical Tmax, Tmin and Topt of 30-31, -7 degrees and 26 degrees C, respectively. The beta-galactosidase in cell extracts had an optimal activity at 39 degrees C. The strains were Gram-negative rods, showed gliding motility, contained branched and hydroxy fatty acids, and menaquinone 6 as the major respiratory quinone. The strains did not form microcysts and utilized lactose while using ammonium ions as a source of nitrogen, and a range of other sugars. The G + C content of the DNA was 34 mol%. Phylogenetic analysis of one of the strains, by comparison of 16S rDNA sequences, showed that it was most similar, but not identical to, Flavobacterium columnare and '[Sporocytophaga] cauliformis'. Both species could be differentiated phenotypically from the Antarctic isolates. DNA-DNA hybridization of the Antarctic isolate with six different members of the Flavobacterium 16S rDNA cluster showed no strain with greater than 18% relatedness. The nearest type species to the Antarctic isolate in the phylogenetic analysis was Flavobacterium aquatile. The name Flavobacterium hibernum is proposed for the Antarctic strains, and the type strain is ATCC 51468T (= ACAM 376T).

Lipids and buoyancy in Southern ocean pteropods.

The lipids of Clione limacina, a Southern Ocean pteropod (order Gymnosomata), contain 28% diacylglyceryl ether (DAGE) (as percentage of total lipid) whereas the pteropod Limacina helicina (order Thecosomata) lacks DAGE. The alkyl glyceryl ether diols (1-O-alkyl glycerols, GE) of Clione DAGE are dominated by 16:0 (60%) and 15:0 (21%), in contrast with deep-sea shark liver DAGE, which is dominated by 18:1 GE. The fatty acid profiles of Clione and Limacina are similar (28-32% polyunsaturated, 26-34% monounsaturated) as are the sterols, which include 24-methylenecholesterol, transdehydrocholesterol, cholesterol, and desmosterol. This finding probably reflects the fact that Limacina is the major food source for Clione. Spongiobranchaea australis, another Southern Ocean pteropod (order Gymnosomata), has 0.9-1.7% DAGE, but has less lipid (3.3-4.8 mg/g lipid, wet weight) than Clione (50.8 mg/g lipid, wet weight). We propose a buoyancy role for DAGE in Clione since Limacina has bubbles for flotation which Clione lack; DAGE provides 23% more uplift than triacylglycerol at a concentration of 1.025 g/mL seawater.

Shewanella gelidimarina sp. nov. and Shewanella frigidimarina sp. nov., novel Antarctic species with the ability to produce eicosapentaenoic acid (20:5 omega 3) and grow anaerobically by dissimilatory Fe(III) reduction.

A polyphasic taxonomic study was performed to characterize dissimilatory iron-reducing strains mostly isolated from Antarctic sea ice. The strains were isolated from samples of congelated (land-fast) sea ice, grease ice, and ice algal biomass collected from the coastal areas of the Vestfold Hills in eastern Antarctica (68 degrees S 78 degrees E). The strains were facultatively anaerobic, motile, and rod shaped, were capable of anaerobic growth either by fermentation of carbohydrates or by anaerobic respiration, and utilized a variety of electron acceptors, including nitrate, ferric compounds, and trimethylamine N-oxide. A phylogenetic analysis performed with 16S rRNA sequences showed that the isolates formed two groups representing novel lineages in the genus Shewanella. The first novel group included seawater-requiring, psychrophilic, chitinolytic strains which had DNA G + C contents of 48 mol%. The members of the second strain group were psychrotrophic and did not require seawater but could tolerate up to 9% NaCl. The strains of this group were also unable to degrade polysaccharides but could utilize a number of monosaccharides and disaccharides and had G + C contents of 40 to 43 mol%. The whole-cell-derived fatty acid profiles of the sea ice isolates were found to be similar to the profiles obtained for other Shewanella species. The omega-3 polyunsaturated fatty acid eicosapentaenoic acid (EPA) (20:5 omega 3) was detected in all of the sea ice isolates at levels ranging from 2 to 16% of the total fatty acids. EPA was also found at high levels in Shewanella hanedai (19 to 22%) and Shewanella benthica (16 to 18%) but was absent in Shewanella alga and Shewanella putrefaciens. On the basis of polyphasic taxonomic data, the Antarctic iron-reducing strains are placed in two new species, Shewanella frigidimarina sp. nov. (type strain, ACAM 591) and Shewanella gelidimarina sp. nov. (type strain, ACAM 456).