In solfataric fields in southwestern Iceland, neutral and sulfide-rich hot springs are characterized by thick bacterial mats at 60 to 80°C that are white or yellow from precipitated sulfur (sulfur mats). In low-sulfide hot springs in the same area, grey or pink streamers are formed at 80 to 90°C, and a Chloroflexus mat is formed at 65 to 70°C. We have studied the microbial diversity of one sulfur mat (high-sulfide) hot spring and one Chloroflexus mat (low-sulfide) hot spring by cloning and sequencing of small-subunit rRNA genes obtained by PCR amplification from mat DNA. Using 98% sequence identity as a cutoff value, a total of 14 bacterial operational taxonomic units (OTUs) and 5 archaeal OTUs were detected in the sulfur mat; 18 bacterial OTUs were detected in the Chloroflexus mat. Although representatives of novel divisions were found, the majority of the sequences were >95% related to currently known sequences. The molecular diversity analysis showed that Chloroflexus was the dominant mat organism in the low-sulfide spring (1 mg liter ؊1 ) below 70°C, whereas Aquificales were dominant in the high-sulfide spring (12 mg liter ؊1 ) at the same temperature. Comparison of the present data to published data indicated that there is a relationship between mat type and composition of Aquificales on the one hand and temperature and sulfide concentration on the other hand.Sulfide-rich hot springs with neutral or alkaline pH are relatively rare in most geothermal areas in the world. However, these types of hot springs are rather common in Iceland due to high ground water level and climatic conditions, i.e., from melting snow and rain. Bacteria that thrive in such springs often form long streamers or mats, but the appearance of the mats and the types of bacteria in them seem to vary depending on the sulfide concentration, pH, temperature, and other chemical and physical factors (6,7,10,11,12,20,33). Many Icelandic hot springs have sulfide concentrations as high as 30 mg liter Ϫ1 and, under such conditions, thick bacterial mats which can be spectacularly white or bright yellow from precipitated sulfur are formed.The diversity of many microbial ecosystems has now been studied with different molecular methods, such as analysis of small-subunit (SSU) rRNA by sequencing, denaturing gradient gel electrophoresis, or restriction fragment length polymorphism analyses. These studies show that the diversity of microbial ecosystems is typically 100 to 1,000 times greater than that shown by cultivation alone (14,15,22,23,30,31). The sequencing of rRNA genes from environmental samples is very informative, since it provides information on both the phylogenetic relationship and the population structure of the microbial community. With increased understanding of the role and importance of microbes in many ecosystems, the benefit of microbial diversity studies is being recognized. The practical value of these methods is already widespread, as they can be used to study the performance of wastewater treatment plants (27), monitor changes upo...

BackgroundInternational fish trade reached an import value of 62.8 billion Euro in 2006, of which 44.6% are covered by the European Union. Species identification is a key problem throughout the life cycle of fishes: from eggs and larvae to adults in fisheries research and control, as well as processed fish products in consumer protection.Methodology/Principal FindingsThis study aims to evaluate the applicability of the three mitochondrial genes 16S rRNA (16S), cytochrome b (cyt b), and cytochrome oxidase subunit I (COI) for the identification of 50 European marine fish species by combining techniques of “DNA barcoding” and microarrays. In a DNA barcoding approach, neighbour Joining (NJ) phylogenetic trees of 369 16S, 212 cyt b, and 447 COI sequences indicated that cyt b and COI are suitable for unambiguous identification, whereas 16S failed to discriminate closely related flatfish and gurnard species. In course of probe design for DNA microarray development, each of the markers yielded a high number of potentially species-specific probes in silico, although many of them were rejected based on microarray hybridisation experiments. None of the markers provided probes to discriminate the sibling flatfish and gurnard species. However, since 16S-probes were less negatively influenced by the “position of label” effect and showed the lowest rejection rate and the highest mean signal intensity, 16S is more suitable for DNA microarray probe design than cty b and COI. The large portion of rejected COI-probes after hybridisation experiments (>90%) renders the DNA barcoding marker as rather unsuitable for this high-throughput technology.Conclusions/SignificanceBased on these data, a DNA microarray containing 64 functional oligonucleotide probes for the identification of 30 out of the 50 fish species investigated was developed. It represents the next step towards an automated and easy-to-handle method to identify fish, ichthyoplankton, and fish products.

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Geothermal energy has been harnessed and used for domestic heating in Iceland. In wells that are typically drilled to a depth of 1,500 to 2,000 m, the temperature of the source water is 50 to 130°C. The bottoms of the boreholes can therefore be regarded as subterranean hot springs and provide a unique opportunity to study the subterranean biosphere. Large volumes of geothermal fluid from five wells and a mixture of geothermal water from 50 geothermal wells (hot tap water) were sampled and concentrated through a 0.2-m-pore-size filter. Cells were observed in wells RG-39 (91.4°C) and MG-18 (71.8°C) and in hot tap water (76°C), but no cells were detected in wells SN-4, SN-5 (95 to 117°C), and RV-5 (130°C). Archaea and Bacteria were detected by whole-cell fluorescent in situ hybridization. DNAs were extracted from the biomass, and small-subunit rRNA genes (16S rDNAs) were amplified by PCR using primers specific for the Archaea and Bacteria domains. The PCR products were cloned and sequenced. The sequence analysis showed 11 new operational taxonomic units (OTUs) out of 14, 3 of which were affiliated with known surface OTUs. Samples from RG-39 and hot tap water were inoculated into enrichment media and incubated at 65 and 85°C. Growth was observed only in media based on geothermal water. 16S rDNA analysis showed enrichments dominated with Desulfurococcales relatives. Two strains belonging to Desulfurococcus mobilis and to the Thermus/Deinococcus group were isolated from borehole RG-39. The results indicate that subsurface volcanic zones are an environment that provides a rich subsurface for novel thermophiles.

Marine macroalgal (seaweed) polysaccharides are highly promising for next-generation applications in several industries. However, despite the reported comprehensive potential of these polysaccharides, commercial products are scarce on the market. Seaweed cultivations are increasing in number and production quantity, owing to an elevated global trend of utilization interest in seaweed. The extraction of polysaccharides from seaweed generally generates low yields, but novel methods are being developed to facilitate and improve the extraction processes. Current areas of applications for seaweed polysaccharides mainly take advantage of the physicochemical properties of certain polysaccharides, such as gelling, thickening and emulsifying. However, many of the numerous bioactivities reported are still only at research level and lack clinical evidence for commercialization. It has been suggested the construction of smaller units may generate better defined molecules that are more suitable for biomedical applications. Enzymatic modification is a promising tool for the generation of more defined, targeted biomolecules. This review covers; structural differences between the most predominant marine algal polysaccharides, extraction processes, modification alternatives, as well as a summary of current and potential next-generation application areas.

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In many cases marine organisms and especially their diverse developmental stages are difficult to identify by morphological characters. DNA-based identification methods offer an analytically powerful addition or even an alternative. In this study, a DNA microarray has been developed to be able to investigate its potential as a tool for the identification of fish species from European seas based on mitochondrial 16S rDNA sequences. Eleven commercially important fish species were selected for a first prototype. Oligonucleotide probes were designed based on the 16S rDNA sequences obtained from 230 individuals of 27 fish species. In addition, more than 1200 sequences of 380 species served as sequence background against which the specificity of the probes was tested in silico. Single target hybridisations with Cy5-labelled, PCR-amplified 16S rDNA fragments from each of the 11 species on microarrays containing the complete set of probes confirmed their suitability. True-positive, fluorescence signals obtained were at least one order of magnitude stronger than false-positive cross-hybridisations. Single nontarget hybridisations resulted in cross-hybridisation signals at approximately 27% of the cases tested, but all of them were at least one order of magnitude lower than true-positive signals. This study demonstrates that the 16S rDNA gene is suitable for designing oligonucleotide probes, which can be used to differentiate 11 fish species. These data are a solid basis for the second step to create a "Fish Chip" for approximately 50 fish species relevant in marine environmental and fisheries research, as well as control of fisheries products.

The microbial diversity of intertidal hot springs on the seashore of northwest Iceland was examined by combining directed in situ enrichments, artificial support colonization, and mat sampling. Analysis of 16S rRNA genes revealed the presence of clones related to both marine and terrestrial, thermophilic, mesophilic, and psychrophilic microorganisms scattered among 11 bacterial divisions. No archaea were found. The species composition of the enrichments was affected by the length of the hot periods experienced at low tide and was very different from those found in the biomass. A total of 36 chitinase genes were detected by molecular screening of the samples with degenerate primers for glycoside hydrolase family 18. The chitinase gene diversity was at least twofold higher in the enrichment samples than in the controls, indicating that a much higher diversity of hydrolytic genes can be accessed with this approach.The coastal hot springs located on the Reykjanes peninsula in northwest Iceland can be regarded as one of the most extreme microbial environments on Earth. A series of freshwater hot springs are located on the seashore, from the tidal zone to about 100 m off the coast. The geothermal system is essentially supplied with deep fresh and slightly alkaline groundwater. It is also low in sulfide and slightly mixed with seawater in the hot spring outlets (chloride, Յ600 g/ml). The temperature ranges from 45 to 95°C in different springs, and many are covered by the sea at high tides (9). The tides are as high as 4 m in this area, and therefore the organisms living in those hot springs can be subject to almost 100°C fluctuations twice per day, in addition to severe changes in salinity, light, and oxygen concentration. This type of habitat is quite rare, and no studies have been conducted on the unculturable microflora in these environments until now. Furthermore, the effects of drastic temperature and salinity fluctuations remain so far unknown.Chitin is an abundant polysaccharide in the marine environment. Information on microbial chitinases has been prominently restricted to a few culturable microorganisms belonging to ␤-and ␥-proteobacteria, gram-positive bacteria, and the domain Archaea (3,5,6,21). Yet, the use of chitinase genes as molecular markers has demonstrated the presence of a large pool of uncultured chitinolytic microorganisms in marine and soil environments (3,13,14,24).In this report, we used molecular diversity methods, combined with in situ enrichments and artificial support colonization, to investigate the apparent microbial diversity of 16S rRNA genes and chitinase genes. 16S rRNA gene analysis of natural samples has shown that the number of microbial species present can differ drastically in occurrence and that organisms that are below 0.1% can hardly be detected (2). However, combining cultivation methods and direct molecular techniques enables access to more diversity than does either method alone (7,16). By applying different sampling methods we analyzed both the diversity of 16S rRNA genes...