Prokaryotic diversity of a Tunisian multipond solar saltern

16S rRNA gene clone libraries were separately constructed from three ponds with different salt concentrations, M2 (15%), TS38 (25%) and S5 (32%), located within a multipond solar saltern of Sfax. The 16S rRNA genes from 216 bacterial clones and 156 archaeal clones were sequenced and phylogenetically analyzed. 44 operational taxonomic units (OTUs) were generated for Bacteria and 67 for Archaea. Phylogenetic groups within the bacterial domain were restricted to Bacteroidetes and Proteobacteria, with the exception that one cyanobacterial OTU was found in the TS38 pond. 85.7, 26.6 and 25.0% of the bacterial OTUs from M2, TS38 and S5 ponds, respectively, are novel. All archaeal 16S rRNA gene sequences were exclusively affiliated with Euryarchaeota. 75.0, 60.0 and 66.7% of the OTUs from, respectively, M2, TS38 and S5 ponds are novel. The result showed that the Tunisian multipond solar saltern harbored novel prokaryotic diversity that has never been reported before for solar salterns. In addition, diversity measurement indicated a decrease of bacterial diversity and an increase of archaeal diversity with rising salinity gradient, which was in agreement with the previous observation for thalassohaline systems. Comparative analysis showed that prokaryotic diversity of Tunisian saltern was higher than that of other salterns previously studied. A. Sghir and E. Ammar have equally contributed to this work.     

Integrated Evaluation of Environmental Parameters Influencing <Emphasis Type="Italic">Vibrio</Emphasis> Occurrence in the Coastal Northern Adriatic Sea (Italy) Facing the Venetian Lagoon

In the marine environment, the persistence and abundance of Vibrio are related to a number of environmental parameters. The influence of the different environmental variables in determining the Vibrio occurrence could be different in the specific geographic areas around the world. Moreover, oceanographic parameters are generally interdependent and should not be considered separately when their influence on bacterial presence and concentration is tested. In this study, an integrated approach was used to identify key parameters determining the abundance of Vibrio spp in marine samples from the Venetian Lagoon in Italy, which is an important area for fish farming and tourism. Multivariate techniques have been adopted to analyze the dataset: using PCA, it was shown that a relatively high proportion of the total variance in this area was mainly due to two independent variables, namely salinity and temperature. Using cluster analysis, it was possible to categorize different groups with homogeneous features as regards space (“stations”) and time (“seasons”) distribution, as well as to quantify the values of environmental variables and the Vibrio abundances in each category. Furthermore, integrating key environmental factors and bacterial concentration values, it was possible to identify levels of salinity and sea surface temperature which were optimal for Vibrio concentration in water, plankton, and sediment samples. The identification of key environmental variables conditioning Vibrio occurrence should facilitate ocean monitoring, making it possible to predict unexpected variations in marine microflora which determine possible public health risks in coastal areas.     

Spatial and seasonal prokaryotic community dynamics in ponds of increasing salinity of Sfax solar saltern in Tunisia

The spatial and seasonal dynamics of the halophilic prokaryotic community was investigated in five ponds from Sfax solar saltern (Tunisia), covering a salinity gradient ranging from 20 to 36%. Fluorescence in situ hybridization indicated that, above 24% salinity, the prokaryotic community shifted from bacterial to archaeal dominance with a remarkable increase in the proportion of detected cells. Denaturing gradient gel electrophoresis (DGGE) profiles were rather similar in all the samples analyzed, except in the lowest salinity pond (around 20% salt) where several specific archaeal and bacterial phylotypes were detected. In spite of previous studies on these salterns, DGGE analysis unveiled the presence of microorganisms not previously described in these ponds, such as Archaea related to Natronomonas or bacteria related to Alkalimnicola, as well as many new sequences of Bacteroidetes. Some phylotypes, such as those related to Haloquadratum or to some Bacteroidetes, displayed a strong dependence of salinity and/or magnesium concentrations, which in the case of Haloquadratum could be related to the presence of ecotypes. Seasonal variability in the prokaryotic community composition was focused on two ponds with the lowest (20%) and the highest salinity (36%). In contrast to the crystallized pond, where comparable profiles between autumn 2007 and summer 2008 were obtained, the non-crystallized pond showed pronounced seasonal changes and a sharp succession of “species” during the year. Canonical correspondence analysis of biological and physicochemical parameters indicated that temperature was a strong factor structuring the prokaryotic community in the non-crystallizer pond, that had salinities ranging from 20 to 23.8% during the year.     

Benthic ecology of <Emphasis Type="Italic">Vibrio</Emphasis> spp. and pathogenic <Emphasis Type="Italic">Vibrio</Emphasis> species in a coastal Mediterranean environment (La Spezia Gulf,Italy)

We carried out a 16-month in situ study to investigate the ecology of Vibrio spp. and pathogenic Vibrio species in coastal sediments of the Mediterranean Sea, employing multiple-regression analysis to reveal the major environmental factors controlling their occurrence in the benthic environment. In addition, association between vibrios and sediment-inhabiting meiofauna, which is a major component of benthic ecosystems, was investigated. Culturable and total Vibrio spp. estimates by most-probable-number technique coupled with standard polymerase chain reaction (PCR) and real-time PCR methods, respectively, were at least one order of magnitude higher in sediment than in seawater. In addition, potential human pathogenic species Vibrio cholerae, Vibrio vulnificus and Vibrio parahaemolyticus occurred in the sediment with V. parahaemolyticus being the most frequently found. In the pelagic environment, 60% of total variance in culturable Vibrio data was explained by sea surface temperature (40%), salinity (13%) and organic matter concentration (7%). In the benthic environment, sea surface temperature was the only factor that significantly affected culturable Vibrio occurrence although it explained only 25% of total variance, suggesting that additional unexplored factors may play a role as well. No correlation was found between culturable Vibrio spp. concentrations and the abundance of harpacticoid copepods in the sediment whilst a negative correlation was found between Vibrio spp. and nematode abundance which accounted for almost 90% of the total meiofaunal density. Taxonomic analysis revealed that selective bacterial feeders accounted for nearly 50% of the total nematode community and included genera such as Terschellingia, Molgolaimus and Halalaimus, suggesting that top-down control by nematode grazing may be an important factor affecting Vibrio occurrence in these sediments. It is concluded that the benthic marine environment may function as a reservoir of Vibrio spp. and potential pathogenic vibrios whose ecological features appeared substantially different from the ones recognised in the pelagic environment.     

Comparison of bacterioplankton communities in three mariculture ponds farming different commercial animals in subtropical Chinese coast

In order to explore the responses of the bacterioplankton community to different types of aquaculture environments, three mariculture ponds comprised of groupers (Epinephelus diacanthus, ED), prawns (Penaeus vannamei, PV), and abalone (Haliotis diversicolor supertexta, HDS) in southeast, coastal China were investigated. The free-living bacterial diversity was analyzed through the construction of 16S rDNA clone library. A total of 203 16S rDNA sequences from three clone libraries were classified into 118 operational taxonomic units (OTUs), of which 51, 31, and 42 OTUs were distributed in the ED, PV, and HDS pond, respectively, with Bacteroidetes (30.6%), Actinobacteria (55.2%), and Cyanobacteria (32.8%) as the dominant division in the respective ponds. Meanwhile, each pond occupied some unique OTUs that were affiliated with uncommon (sub-)phyla, such as candidate OP11 division, Acidobacteria, Deltaproteobacteria, Planctomycetes, and Verrucomicrobia. Bacterial diversity in the ED pond was the richest, followed by the HDS and the PV pond. OTUs of 61.9% and 94.9% have less than 90% and 97% similarity to their nearest neighbors in public databases, respectively. All OTUs were grouped into 67 clusters, covering 11 (sub-)phyla. The OTUs only from single pond distributed in 53 clusters (79.1%), the OTUs shared by two ponds were affiliated with 14 clusters (20.9%), and none of clusters was formed by the OTUs which commonly originated from the three pond libraries, suggesting that the composition of bacterial populations in these ponds were significantly different. These results indicate that the aquatic environment created by different mariculture animals may foster very special and complex bacterial communities. Handling editor: David Philip Hamilton     

Virioplankton Community Structure in Tunisian Solar Salterns

The microbial community inhabiting Sfax solar salterns on the east coast of Tunisia has been studied by means of different molecular and culture-dependent tools that have unveiled the presence of novel microbial groups as well as a community structure different from that of other coastal hypersaline environments. We have focused on the study of the viral assemblages of these salterns and their changes along the salinity gradient and over time. Viruses from three ponds (C4, M1, and TS) encompassing salinities from moderately hypersaline to saturated (around 14, 19, and 35%, respectively) were sampled in May and October 2009 and analyzed by transmission electron microscopy (TEM) and pulsed-field gel electrophoresis (PFGE). Additionally, for all three October samples and the May TS sample, viral metagenomic DNA was cloned in fosmids, end sequenced, and analyzed. Viral concentration, as well as virus-to-cell ratios, increased along the salinity gradient, with around 10¹⁰ virus-like particles (VLPs)/ml in close-to-saturation ponds, which represents the highest viral concentration reported so far for aquatic systems. Four distinct morphologies could be observed with TEM (spherical, tailed, spindled, and filamentous) but with various proportions in the different samples. Metagenomic analyses indicated that every pond harbored a distinct viral assemblage whose G+C content could be roughly correlated with that of the active part of the microbial community that may have constituted the putative hosts. As previously reported for hypersaline metaviromes, most sequences did not have matches in the databases, although some were conserved among the Sfax metaviromes. BLASTx, BLASTp, and dinucleotide frequency analyses indicated that (i) factors additional to salinity could be structuring viral communities and (ii) every metavirome had unique gene contents and dinucleotide frequencies. Comparison with hypersaline metaviromes available in the databases indicated that the viral assemblages present in close-to-saturation environments located thousands of kilometers apart presented some common traits among them in spite of their differences regarding the putative hosts. A small core metavirome for close-to-saturation systems was found that contained 7 sequences of around 100 nucleotides (nt) whose function was not hinted at by in silico search results, although it most likely represents properties essential for hyperhalophilic viruses.     

Environmental Influences on Vibrio Populations in Northern Temperate and Boreal Coastal Waters (Baltic and Skagerrak Seas)

Even if many Vibrio spp. are endemic to coastal waters, their distribution in northern temperate and boreal waters is poorly studied. To identify environmental factors regulating Vibrio populations in a salinity gradient along the Swedish coastline, we combined Vibrio-specific quantitative competitive PCR with denaturant gradient gel electrophoresis-based genotyping. The total Vibrio abundance ranged from 4 × 10³ to 9.6 × 10⁴ cells liter⁻¹, with the highest abundances in the more saline waters of the Skagerrak Sea. Several Vibrio populations were present throughout the salinity gradient, with abundances of single populations ranging from 5 × 10² to 7 × 10⁴ cells liter⁻¹. Clear differences were observed along the salinity gradient, where three populations dominated the more saline waters of the Skagerrak Sea and two populations containing mainly representatives of V. anguillarum and V. aestuarianus genotypes were abundant in the brackish waters of the Baltic Sea. Our results suggest that this apparent niche separation within the genus Vibrio may also be influenced by alternate factors such as nutrient levels and high abundances of dinoflagellates. A V. cholerae/V. mimicus population was detected in more than 50% of the samples, with abundances exceeding 10³ cells liter⁻¹, even in the cold (annual average water temperature of around 5°C) and low-salinity (2 to 4‰) samples from the Bothnian Bay (latitude, 65°N). The unsuspected and widespread occurrence of this population in temperate and boreal coastal waters suggests that potential Vibrio pathogens may also be endemic to cold and brackish waters and hence may represent a previously overlooked health hazard.     

The microbial food web along salinity gradients

The microbial food web was studied along a gradient of salinity in two solar salterns used for the commercial production of salt. The different ponds in the salterns provide a wide range of ecosystems with food webs of different complexities. Abundance of prokaryotes, cell volume, prokaryotic heterotrophic production, chlorophyll a, abundance of heterotrophic flagellates, ciliates and phytoplankton were determined in several ponds in each saltern. Increases in salinity resulted in a progressive reduction in the abundance and number of different groups of eukaryotic microorganisms present, but an increase in biomass of prokaryotes. Maximal activity of both phyto- and bacterioplankton was found at a salinity of around 100 per thousand, where there was also a maximum in chlorophyll a concentration. Growth rates of heterotrophic prokaryotes decreased with increasing salinity. Bacterivory disappeared above 250 per thousand salinity, whereas other loss factors such as viral lysis appeared to be of minor importance throughout the gradient [Guixa-Boixereu et al. (1996) Aquat. Microb. Ecol. 11, 215-227].     

Characterization of protistan plankton diversity in ancient salt evaporation ponds located in a volcanic crater on the island Sal,Cape Verde

Salinity is an important factor when exploring the limits known for life. Therefore, hypersaline systems have attracted much attention in recent years. In this study, we investigated the protistan diversity and community composition in two natural salt evaporation ponds (27–30% salinity) located in an ancient volcanic crater on the Cape Verde island Sal using high-throughput DNA sequencing. Our study revealed a broad range of protistan taxa and a high taxonomic diversity within the Ciliophora, Dinophyceae, and Chlorophyta. We detected a total of 23 Dinophyceae families, although Dinophyceae were generally considered to be only this diverse in aquatic environments of less than 10% salinity. Moreover, we uncovered a high degree of genetic novelty in this habitat. The mean similarity of all detected OTUs to previously described sequences was only 93.6%. These findings strongly dispute the traditional view that extreme hypersaline environments generally maintain low protistan diversity. A meta-analysis covering our and previously published data from other inland and coastal salt ponds clearly showed that our samples clustered according to salinity and not biogeography. This result further supports the claim that salinity is a major transition boundary for protistan communities, regardless of their biogeographic origin.

     

Microbial Diversity in Maras Salterns, a Hypersaline Environment in the Peruvian Andes

Maras salterns are located 3,380 m above sea level in the Peruvian Andes. These salterns consist of more than 3,000 little ponds which are not interconnected and act as crystallizers where salt precipitates. These ponds are fed by hypersaline spring water rich in sodium and chloride. The microbiota inhabiting these salterns was examined by fluorescence in situ hybridization (FISH), 16S rRNA gene clone library analysis, and cultivation techniques. The total counts per milliliter in the ponds were around 2 × 10⁶ to 3 × 10⁶ cells/ml, while the spring water contained less than 100 cells/ml and did not yield any detectable FISH signal. The microbiota inhabiting the ponds was dominated (80 to 86% of the total counts) by Archaea, while Bacteria accounted for 10 to 13% of the 4′,6′-diamidino-2-phenylindole (DAPI) counts. A total of 239 16S rRNA gene clones were analyzed (132 Archaea clones and 107 Bacteria clones). According to the clone libraries, the archaeal assemblage was dominated by microorganisms related to the cosmopolitan square archaeon “Haloquadra walsbyi,” although a substantial number of the sequences in the libraries (31% of the 16S rRNA gene archaeal clones) were related to Halobacterium sp., which is not normally found in clone libraries from solar salterns. All the bacterial clones were closely related to each other and to the γ-proteobacterium “Pseudomonas halophila” DSM 3050. FISH analysis with a probe specific for this bacterial assemblage revealed that it accounted for 69 to 76% of the total bacterial counts detected with a Bacteria-specific probe. When pond water was used to inoculate solid media containing 25% total salts, both extremely halophilic Archaea and Bacteria were isolated. Archaeal isolates were not related to the isolates in clone libraries, although several bacterial isolates were very closely related to the “P. halophila” cluster found in the libraries. As observed for other hypersaline environments, extremely halophilic bacteria that had ecological relevance seemed to be easier to culture than their archaeal counterparts.     

Microbial diversity and composition in acidic sediments of freshwater finfish culture ponds fed with two types of feed: a metagenomic approach

Microbial community profile associated with acidic pond sediments (APS) (pH = 3·0–4·5) of freshwater finfish aquaculture ponds (n = 8) was investigated. Sediment DNA extracted from the eight APS were subjected to high-throughput sequencing of V3 and V4 regions which yielded 7236 operational taxonomic units (OTUs) at a similarity of 97%. Overall results showed higher proportion of bacterial OTUs than archaeal OTUs in all the APS. Euryarchaeota (23%), Proteobacteria (19%), Chloroflexi (17%), Crenarchaeota (5·3%), Bacteroidetes (4·8%), Nitrospirae (3·2%), Nanoarchaeaeota (3%) which together constituted 75% of the microbial diversity. At the genus level, there was high preponderance of methanogens namely Methanolinea (5·4%), Methanosaeta (4·5%) and methanotrops, Bathyarchaeota (5%) in APS. Moreover, the abundant phyla in the APS were not drastically affected by the administration of chicken slaughter waste (R-group ponds) and commercial fish feed (C-group ponds), since 67% of the OTUs generated remained common in the APS of both the groups of ponds. There was a minimal difference of 24–26% of OTUs between C-group and R-group ponds, suggesting the existence of a core microbial community in these ponds driven by acidic pH over the years. This study concludes that microbial diversity in pond sediment was influenced to a lesser extent by the addition of chicken slaughter waste but was majorly driven by acidic nature of the pond.     

Seasonal Dynamics and Modeling of a <Emphasis Type="Italic">Vibrio</Emphasis> Community in Coastal Waters of the North Sea

Vibrio species are ubiquitously distributed in marine waters all over the world. High genome plasticity due to frequent mutation, recombination, and lateral gene transfer enables Vibrio to adapt rapidly to environmental changes. The genus Vibrio comprises several human pathogens, which commonly cause outbreaks of severe diarrhea in tropical regions. In recent years, pathogenic Vibrio emerged also in coastal European waters. Little is known about factors driving the proliferation of Vibrio spp. in temperate waters such as the North Sea. In this study a quantification of Vibrio in the North Sea and their response to biotic and abiotic parameters were assessed. Between January and December 2009, Vibrio at Helgoland Roads (North Sea, Germany) were quantified using fluorescence in situ hybridization. Vibrio numbers up to 3.4 × 10⁴ cells × mL⁻¹ (2.2% of total microbial counts) were determined in summer, but their abundance was significantly lower in winter (5 × 10² cells × mL⁻¹). Correlations between Vibrio and nutrients (SiO₂, PO₄ ³⁻, DIN), Secchi depth, temperature, salinity, and chlorophyll a were calculated using Spearman rank analysis. Multiple stepwise regression analysis was carried out to analyze the additive influence of multiple factors on Vibrio. Based on these calculations, we found that high water temperature and low salinity best explained the increase of Vibrio cell numbers. Other environmental parameters, especially nutrients and chlorophyll a, also had an influence. All variables were shown to be subject to the overall seasonal dynamics at Helgoland Roads. Multiple regression models could represent an efficient and reliable tool to predict Vibrio abundances in response to the climate change in European waters.     

Zooplankton distribution in four ponds of different salinity: a seasonal study in the solar salterns of Sfax (Tunisia)

The seasonal distribution of metazooplankton and large-sized ciliates was studied in four ponds of different salinity in the solar salterns of Sfax (Tunisia). Total zooplankton abundance varied from 1 × 10³ to 4.7 × 10⁶ ind m^–3. Salinity had a negative effect on the abundance of copepods and rotifers which were absent in the pond with the highest salt concentration (180) in which the number of taxa was low and Artemia or the ciliate Fabrea largely dominated the zooplankton community. Temperature and the presence of Dunaliella salina as prey appeared as key factors in controlling the abundance of Artemia, while organic detritus appeared as important in the diet of Fabrea. Change in zooplankton species composition along the hypersaline gradient (40–90) was primarily related to salinity. However, our data suggest the importance of both the abundance and composition of food in the spatial and temporal variations of some zooplankton species.     

Associations of planktonic and periphytic rotifers in a Malaysian estuary and two nearby ponds

Samples were taken from a Malaysian estuary along a salinity gradient from full strength sea water down to a conductivity of 27 µS cm^–1. In a region where the conductivity fluctuated between 6500 and 14000 µS only empty loricas of Keratella cochlearis and Lecane bulla were found. Where the salinity reached a maximum of 3800 µS Lecane bulla was the dominant rotifer, and 17 other species were found. Where the conductivity remained below 1000 µS at all stages of the tidal cycle there were more species of rotifers, up to 33 at one station.The rotifer fauna of the estuary was compared to the assemblage found in two local ponds. Of the 63 species found in the estuary only 29 were also found in the ponds. Compared to the total known rotifer fauna of Malaysia the estuary has a higher proportion of Lecane species (40%), and the genera Brachionus and Trichocerca are under-represented.Species associated with inland saline waters, such as Brachionus plicatilis, B. dimidiatus and Hexarthra jenkinae were not found in the estuary.     

Diversity of Vibrio spp in Karstic Coastal Marshes in the Yucatan Peninsula

Coastal bodies of water formed by the combination of seawater, underground rivers and rainwater comprise the systems with the greatest solar energy flow and biomass production on the planet. These characteristics make them reservoirs for a large number species, mainly microorganisms. Bacteria of the genus Vibrio are natural inhabitants of these environments and their presence is determined by variations in the nutrient, temperature and salinity cycles generated by the seasonal hydrologic behavior of these lagoon systems. This study determined the diversity of the genus Vibrio in 4 coastal bodies of water on the Yucatan Peninsula (Celestun Lagoon, Chelem Lagoon, Rosada Lagoon and Sabancuy Estuary). Using the molecular technique of 454 pyrosequencing, DNA extracted from water samples was analyzed and 32,807 reads were obtained belonging to over 20 culturable species of the genus Vibrio and related genera. OTU (operational taxonomic unit) richness and Chao2 and Shannon Weaver diversity indices were obtained with the database from this technique. Physicochemical and environmental parameters were determined and correlated with Vibrio diversity measured in OTUs.     

The biology and evolution of Antarctic saline lakes in relation to salinity and trophy

. Five brackish to hypersaline lakes (Highway, Ace, Pendent, Williams and Rookery) in the Vestfold Hills, eastern Antarctica were investigated during the austral summer of 1999/2000. The aims were to characterise the functional dynamics of the plankton and gain an understanding of how the different environments in the lakes have led to the evolution of different communities. The plankton was dominated by microorganisms and differed across the salinity spectrum in relation to trophy, age and the presence of meromixis. However, some elements of the plankton were common to all of the lakes, e.g. the mixtrophic ciliate, Mesodinium rubrum, which reached abundances of 2.7᎒⁵ l^-1 and spanned a salinity gradient of 4-63‰. Marine dinoflagellate species also occurred in all of the lakes, often at high abundances in Highway Lake, Pendent Lake and Lake Williams. During December (mid-summer), primary production showed an increase along the salinity gradient from Highway Lake to Lake Williams; however, it was low in hyper-nutrified Rookery Lake because of the turbidity of the waters. Bacterial production followed the same trend and was extremely high in Rookery Lake (327 µg l^-1 h^-1 in January). The lakes possessed a marine microbial plankton that has become very simplified through time, and now contains a small number of highly successful species, which were pre-adapted to surviving in extreme Antarctic lakes.     

Bacteria associated with Artemia spp. along the salinity gradient of the solar salterns at Eilat (Israel)

The crustacean genus Artemia naturally inhabits various saline and hypersaline environments and is the most frequently laboratory-hatched animal for live feed in mari- and aquaculture. Because of its high economic importance, Artemia-bacteria interactions were so far studied mostly in laboratory strains. In this study, we focused our attention on the Artemia-associated microbiota in its natural environment in the solar salterns of Eilat, Israel. We applied a culture-independent method (clone libraries) to investigate the bacterial community structure associated with Artemia in five evaporation ponds with salinities from slightly above seawater (5%) to the point of saturation (32%), in two different developmental stages: in nauplii and in the intestine of adult animals. Bacteria found in naupliar and adult stages were classified within the Proteobacteria, Bacteroidetes, Firmicutes, Actinobacteria and Cyanobacteria. The halophilic proteobacterial genera Halomonas spp. and Salinivibrio spp. dominated the Artemia microbiota in both stages in all ponds. We also analysed a clone library of entire adult animals, revealing a novel bacterial phylogenetic lineage. This is the first molecular study of bacteria associated with two developmental stages of Artemia along a salinity gradient.     

Microbial diversity in Maras salterns, a hypersaline environment in the Peruvian Andes

Maras salterns are located 3,380 m above sea level in the Peruvian Andes. These salterns consist of more than 3,000 little ponds which are not interconnected and act as crystallizers where salt precipitates. These ponds are fed by hypersaline spring water rich in sodium and chloride. The microbiota inhabiting these salterns was examined by fluorescence in situ hybridization (FISH), 16S rRNA gene clone library analysis, and cultivation techniques. The total counts per milliliter in the ponds were around 2 x 10(6) to 3 x 10(6) cells/ml, while the spring water contained less than 100 cells/ml and did not yield any detectable FISH signal. The microbiota inhabiting the ponds was dominated (80 to 86% of the total counts) by Archaea, while Bacteria accounted for 10 to 13% of the 4',6'-diamidino-2-phenylindole (DAPI) counts. A total of 239 16S rRNA gene clones were analyzed (132 Archaea clones and 107 Bacteria clones). According to the clone libraries, the archaeal assemblage was dominated by microorganisms related to the cosmopolitan square archaeon "Haloquadra walsbyi," although a substantial number of the sequences in the libraries (31% of the 16S rRNA gene archaeal clones) were related to Halobacterium sp., which is not normally found in clone libraries from solar salterns. All the bacterial clones were closely related to each other and to the gamma-proteobacterium "Pseudomonas halophila" DSM 3050. FISH analysis with a probe specific for this bacterial assemblage revealed that it accounted for 69 to 76% of the total bacterial counts detected with a Bacteria-specific probe. When pond water was used to inoculate solid media containing 25% total salts, both extremely halophilic Archaea and Bacteria were isolated. Archaeal isolates were not related to the isolates in clone libraries, although several bacterial isolates were very closely related to the "P. halophila" cluster found in the libraries. As observed for other hypersaline environments, extremely halophilic bacteria that had ecological relevance seemed to be easier to culture than their archaeal counterparts.     

The metavirome of a hypersaline environment

Hypersaline environments harbour the highest number of virus‐like particles reported for planktonic systems. However, very little is known about the genomic diversity of these virus assemblages since most of the knowledge on halophages is based on the analysis of a few isolates infecting strains of hyperhalophilic Archaea that may not be representatives of the natural microbiota. Here, we report the characterization, through a metagenomic approach, of the viral assemblage inhabiting a crystallizer pond (CR30) from a multi‐pond solar saltern in Santa Pola (SE Spain). A total of 1.35 Mbp were cloned that yielded a total of 620 kb sequenced viral DNA. The metavirome was highly diverse and different from virus communities of marine and other aquatic environments although it showed some similarities with metaviromes from high‐salt ponds in solar salterns in San Diego (SW USA), indicating some common traits between high‐salt viromes. A high degree of diversity was found in the halophages as revealed by the presence of 2479 polymorphic nucleotides. Dinucleotide frequency analysis of the CR30 metavirome showed a good correlation with GC content and enabled the establishment of different groups, and even the assignment of their putative hosts: the archaeon Haloquadratum walsbyi and the bacterium Salinibacter ruber.