Metazoans of redoxcline sediments in Mediterranean deep-sea hypersaline anoxic basins

Background

The deep-sea hypersaline anoxic basins (DHABs) of the Mediterranean (water depth ~3500 m) are some of the most extreme oceanic habitats known. Brines of DHABs are nearly saturated with salt, leading many to suspect they are uninhabitable for eukaryotes. While diverse bacterial and protistan communities are reported from some DHAB haloclines and brines, loriciferans are the only metazoan reported to inhabit the anoxic DHAB brines. Our goal was to further investigate metazoan communities in DHAB haloclines and brines.

Results

We report observations from sediments of three DHAB (Urania, Discovery, L’Atalante) haloclines, comparing these to observations from sediments underlying normoxic waters of typical Mediterranean salinity. Due to technical difficulties, sampling of the brines was not possible. Morphotype analysis indicates nematodes are the most abundant taxon; crustaceans, loriciferans and bryozoans were also noted. Among nematodes, Daptonema was the most abundant genus; three morphotypes were noted with a degree of endemicity. The majority of rRNA sequences were from planktonic taxa, suggesting that at least some individual metazoans were preserved and inactive. Nematode abundance data, in some cases determined from direct counts of sediments incubated in situ with CellTracker^TM Green, was patchy but generally indicates the highest abundances in either normoxic control samples or in upper halocline samples; nematodes were absent or very rare in lower halocline samples. Ultrastructural analysis indicates the nematodes in L’Atalante normoxic control sediments were fit, while specimens from L’Atalante upper halocline were healthy or had only recently died and those from the lower halocline had no identifiable organelles. Loriciferans, which were only rarely encountered, were found in both normoxic control samples as well as in Discovery and L’Atalante haloclines. It is not clear how a metazoan taxon could remain viable under this wide range of conditions.

Conclusions

We document a community of living nematodes in normoxic, normal saline deep-sea Mediterranean sediments and in the upper halocline portions of the DHABs. Occurrences of nematodes in mid-halocline and lower halocline samples did not provide compelling evidence of a living community in those zones. The possibility of a viable metazoan community in brines of DHABs is not supported by our data at this time.

     

Molecular Detection of Anaerobic Ammonium-Oxidizing (Anammox) Bacteria in High-Temperature Petroleum Reservoirs

Anaerobic ammonium-oxidizing (anammox) process plays an important role in the nitrogen cycle of the worldwide anoxic and mesophilic habitats. Recently, the existence and activity of anammox bacteria have been detected in some thermophilic environments, but their existence in the geothermal subterranean oil reservoirs is still not reported. This study investigated the abundance, distribution and functional diversity of anammox bacteria in nine out of 17 high-temperature oil reservoirs by molecular ecology analysis. High concentration (5.31–39.2 mg l^?1) of ammonium was detected in the production water from these oilfields with temperatures between 55°C and 75°C. Both 16S rRNA and hzo molecular biomarkers indicated the occurrence of anammox bacteria in nine out of 17 samples. Most of 16S rRNA gene phylotypes are closely related to the known anammox bacterial genera Candidatus Brocadia, Candidatus Kuenenia, Candidatus Scalindua, and Candidatus Jettenia, while hzo gene phylotypes are closely related to the genera Candidatus Anammoxoglobus, Candidatus Kuenenia, Candidatus Scalindua, and Candidatus Jettenia. The total bacterial and anammox bacterial densities were 6.4?±?0.5?×?10³ to 2.0?±?0.18?×?10⁶ cells ml^?1 and 6.6?±?0.51?×?10² to 4.9?±?0.36?×?10⁴ cell ml^?1, respectively. The cluster I of 16S rRNA gene sequences showed distant identity (<92%) to the known Candidatus Scalindua species, inferring this cluster of anammox bacteria to be a new species, and a tentative name Candidatus “Scalindua sinooilfield” was proposed. The results extended the existence of anammox bacteria to the high-temperature oil reservoirs.     

Dominance of Candidatus Scalindua species in anammox community revealed in soils with different duration of rice paddy cultivation in Northeast China

The anaerobic ammonium-oxidizing (anammox) bacteria play an important role in the oxygen-limited zone for nitrogen cycling, but their roles in agricultural ecosystems are still poorly understood. In this study, soil samples were taken from the rhizosphere and non-rhizosphere and from surface (0–5 cm) and subsurface (20–25 cm) layers with 1, 4, and 9 years of rice cultivation history on the typical albic soil of Northeast China to examine the diversity and distribution of anammox bacteria based on 16S rRNA gene and hydrazine oxidoreductase encoding gene (hzo). By comparing these soil samples, no obvious difference was observed in community composition between the rhizosphere and non-rhizosphere or the surface and subsurface layers. Surprisingly, anammox bacterial communities of these rice paddy soils were consisted of mainly Candidatus Scalindua species, which are best known to be dominant in marine and pristine environments. The highest diversity was revealed in the 4-year paddy soil based on clone library analysis. Phylogenetic analysis of 16S rRNA gene and deduced HZO from the corresponding encoding gene showed that most of the obtained clones are grouped together with Candidatus Scalindua sorokinii, Candidatus Scalindua brodae, and Candidatus Scalindua spp. of seawater. The obtained clone sequences from all samples are distributed in two subclusters that contain sequences from environmental samples only. Tentative new species were also discovered in this paddy soil. This study provides the first evidence on the existence of anammox bacteria with limited diversity in agricultural ecosystems in Northern China.     

Depth trends of soil organic matter C:N and 15N natural abundance controlled by association with minerals

Anaerobic ammonium oxidation (anammox) is believed to be an important sink for fixed inorganic nitrogen in terrestrial and aquatic ecosystems, and many studies have reported that macroscale oxic–anoxic interfaces, such as riparian zones, were hotspots of anammox reaction. However, no research has linked microscale interfaces with the anammox process in natural environments. This study provides evidence for the presence of anammox bacteria and potential anammox activity on the suspended sediment (SPS) in the oxic water of the Yellow River. The anammox bacteria in the overlying water were mainly attached to SPS. The abundance of anammox bacteria in the overlying water was positively correlated with SPS concentration (R ² = 0.97, P < 0.01), with abundance ranging from 9.5 × 10² to 1.5 × 10⁴ hydrazine synthase gene copies per g of SPS. Phylogenic analysis of anammox bacteria revealed that the SPS phase was dominated by Candidatus Brocadia. Candidatus Scalindua genera was detected in this study with a conductivity of 1100 μS cm^?1. Moreover, \(^{15} {\text{NH}}_{4}^{ + }\)-amended anaerobic incubation of the overlying water showed that the average potential anammox activity was 0.076 nmol-N L^?1 day^?1. The ¹⁵N labeling simulation experiments demonstrated the occurrence of anammox in the oxic water of the Yellow River. This study suggests that the anammox process at the SPS–water interface might be a non-negligible pathway for the loss of fixed nitrogen in natural freshwaters, but this remains to be determined in further studies.     

Water column anammox and denitrification in a temperate permanently stratified lake (Lake Rassnitzer,Germany)

We studied microbial N₂ production via anammox and denitrification in the anoxic water column of a restored mining pit lake in Germany over an annual cycle. We obtained high-resolution hydrochemical profiles using a continuous pumping sampler. Lake Rassnitzer is permanently stratified at ca. 29 m depth, entraining anoxic water below a saline density gradient. Mixed-layer nitrate concentrations averaged ca. 200 μmol L⁻¹, but decreased to zero in the anoxic bottom waters. In contrast, ammonium was <5 μmol L⁻¹ in the mixed layer but increased in the anoxic waters to ca. 600 μmol L⁻¹ near the sediments. In January and October, ¹⁵N tracer measurements detected anammox activity (maximum 504 nmol N₂ L⁻¹ d⁻¹ in ¹⁵NH₄⁺-amended incubations), but no denitrification. In contrast, in May, N₂ production was dominated by denitrification (maximum 74 nmol N₂ L⁻¹ d⁻¹). Anammox activity in May was significantly lower than in October, as characterized by anammox rates (maximum 6 vs. 16 nmol N₂ L⁻¹ d⁻¹ in incubations with ¹⁵NO₃⁻), as well as relative and absolute anammox bacterial cell abundances (0.56% vs. 0.98% of all bacteria, and 2.7×10⁴ vs. 5.2×10⁴ anammox cells mL⁻¹, respectively) (quantified by catalyzed reporter deposition-fluorescence in situ hybridization (CARD-FISH) with anammox bacteria-specific probes). Anammox bacterial diversity was investigated with anammox bacteria-specific 16S rRNA gene clone libraries. The majority of anammox bacterial sequences were related to the widespread Candidatus Scalindua sorokinii/brodae cluster. However, we also found sequences related to Candidatus S. wagneri and Candidatus Brocadia fulgida, which suggests a high anammox bacterial diversity in this lake comparable with estuarine sediments.     

Oil-bioremediation potential of two hydrocarbonoclastic, diazotrophic Marinobacter strains from hypersaline areas along the Arabian Gulf coasts

Two halophilic, hydrocarbonoclastics bacteria, Marinobacter sedimentarum and M. flavimaris, with diazotrophic potential occured in hypersaline waters and soils in southern and northern coasts of Kuwait. Their numbers were in the magnitude of 10³ colony forming units g^?1. The ambient salinity in the hypersaline environments was between 3.2 and 3.5 M NaCl. The partial 16S rRNA gene sequences of the two strains showed, respectively, 99 and 100 % similarities to the sequences in the GenBank. The two strains failed to grow in the absence of NaCl, exhibited best growth and hydrocarbon biodegradation in the presence of 1 to 1.5 M NaCl, and still grew and maintained their hydrocarbonoclastic activity at salinities up to 5 M NaCl. Both species utilized Tween 80, a wide range of individual aliphatic hydrocarbons (C₉–C₄₀) and the aromatics benzene, biphenyl, phenanthrene, anthracene and naphthalene as sole sources of carbon and energy. Experimental evidence was provided for their nitrogen-fixation potential. The two halophilic Marinobacter strains successfully mineralized crude oil in nutrient media as well as in hypersaline soil and water microcosms without the use of any nitrogen fertilizers.     

Residence of Habitat-Specific Anammox Bacteria in the Deep-Sea Subsurface Sediments of the South China Sea: Analyses of Marker Gene Abundance with Physical Chemical Parameters

Anaerobic ammonium oxidation (anammox) has been recognized as an important process for the global nitrogen cycle. In this study, the occurrence and diversity of anammox bacteria in the deep-sea subsurface sediments of the South China Sea (SCS) were investigated. Results indicated that the anammox bacterial sequences recovered from this habitat by amplifying both 16S rRNA gene and hydrazine oxidoreductase encoding hzo gene were all closely related to the Candidatus Scalindua genus. A total of 96 16S rRNA gene sequences from 346 clones were grouped into five subclusters: two subclusters affiliated with the brodae and arabica species, while three new subclusters named zhenghei-I, -II, and -III showed ≤97.4% nucleic acid sequence identity with other known Candidatus Scalindua species. Meanwhile, 88 hzo gene sequences from the sediments also formed five distant subclusters within hzo cluster 1c. Through fluorescent real-time PCR analysis, the abundance of anammox bacteria in deep-sea subsurface sediment was quantified by hzo genes, which ranged from 1.19 × 10⁴ to 7.17 × 10⁴ copies per gram of dry sediments. Combining all the information from this study, diverse Candidatus Scalindua anammox bacteria were found in the deep-sea subsurface sediments of the SCS, and they could be involved in the nitrogen loss from the fixed inventory in the habitat.     

The potential effect of sustained hypoxia on nitrogen cycling in sediment from the southern North Sea: a mesocosm experiment

The potential effect of sustained hypoxia (up to 70 days) on the production of N₂ gas through denitrification and anammox, as well as sediment–water exchange of nitrite, nitrate and ammonia, oxygen consumption and penetration, were measured in mesocosms using sediment collected from the southern North Sea (north of Dogger Bank). As expected, both the penetration of oxygen into, and consumption of oxygen by, the sediment decreased by 42 and 46 %, respectively, once hypoxia was established. Importantly, the oxygen regime did not change significantly (P > 0.05) during the experiment, suggesting that organic carbon was not depleted. During the first 10 days, the exchange of NO₃ ^?, NO₂ ^? and NH₄ ⁺ between the sediment and water was erratic but once a steady state was established the sediment acted as either a sink for fixed nitrogen under hypoxia or as a source in the controls. Over the course of the mesocosm experiment the rate of both anammox and denitrification increased, with anammox increasing disproportionately under hypoxia relative to the controls, whereas the rate of increase in denitrification was the same for both. Under sustained hypoxia the production of N₂ gas increased by 72 % relative to the controls, with this increase in N₂ production remaining constant regardless of the duration of hypoxia. Longer periods of stratification and oxygen depletion are predicted to occur more regularly in the bottom waters of shallow coastal seas as one manifestation of climate change. Under sustained hypoxia the potential for nitrogen removal by the production of N₂ gas in this region of the southern North Sea was estimated to increase from 2.1 kt N 150 days^?1 to 3.6 kt 150 days^?1, while the efflux of dissolved inorganic nitrogen ceased altogether; both of which could down regulate the productivity of this region as a whole.     

Enrichment and physiological characterization of an anaerobic ammonium-oxidizing bacterium ‘Candidatus Brocadia sapporoensis’

We successfully enriched a novel anaerobic ammonium-oxidizing (anammox) bacterium affiliated with the genus ‘Candidatus Brocadia’ with high purity (>90%) in a membrane bioreactor (MBR). The enriched bacterium was distantly related to the hitherto characterized ‘Ca. Brocadia fulgida’ and ‘Ca. Brocadia sinica’ with 96% and 93% of 16S ribosomal RNA gene sequence identity, respectively. The bacterium exhibited the common structural features of anammox bacteria and produced hydrazine in the presence of hydroxylamine under anoxic conditions. The temperature range of anammox activity was 20–45 °C with a maximum activity at 37 °C. The maximum specific growth rate (μ_max) was 0.0082 h^?1 at 37 °C, corresponding to a doubling time of 3.5 days. The half-saturation constant (K_S) for nitrite was 5 ± 2.5 μM. The anammox activity was inhibited by nitrite (IC₅₀ = 11.6 mM) but not by formate and acetate. The major respiratory quinone was identified to be menaquinone-7 (MK-7). The enriched anammox bacterium shared nearly half of genes with ‘Ca. Brocadia sinica’ and ‘Ca. Brocadia fulgida’. The enriched bacterium showed all known physiological characteristics of anammox bacteria and can be distinguished from the close relatives by its 16S rRNA gene sequence. Therefore, we proposed the name ‘Ca. Brocadia sapporoensis’ sp. nov.     

Stable Carbon Isotopic Fractionations Associated with Inorganic Carbon Fixation by Anaerobic Ammonium-Oxidizing Bacteria

Isotopic analyses of Candidatus “Brocadia anammoxidans,” a chemolithoautotrophic bacterium that anaerobically oxidizes ammonium (anammox), show that it strongly fractionates against ¹³C; i.e., lipids are depleted by up to 47‰ versus CO₂. Similar results were obtained for the anammox bacterium Candidatus “Scalindua sorokinii,” which thrives in the anoxic water column of the Black Sea, suggesting that different anammox bacteria use identical carbon fixation pathways, which may be either the Calvin cycle or the acetyl coenzyme A pathway.     

A microdiversity study of anammox bacteria reveals a novel Candidatus Scalindua phylotype in marine oxygen minimum zones

The anaerobic oxidation of ammonium (anammox) contributes significantly to the global loss of fixed nitrogen and is carried out by a deep branching monophyletic group of bacteria within the phylum Planctomycetes. Various studies have implicated anammox to be the most important process responsible for the nitrogen loss in the marine oxygen minimum zones (OMZs) with a low diversity of marine anammox bacteria. This comprehensive study investigated the anammox bacteria in the suboxic zone of the Black Sea and in three major OMZs (off Namibia, Peru and in the Arabian Sea). The diversity and population composition of anammox bacteria were investigated by both, the 16S rRNA gene sequences and the 16S-23S rRNA internal transcribed spacer (ITS). Our results showed that the anammox bacterial sequences of the investigated samples were all closely related to the Candidatus Scalindua genus. However, a greater microdiversity of marine anammox bacteria than previously assumed was observed. Both phylogenetic markers supported the classification of all sequences in two distinct anammox bacterial phylotypes: Candidatus Scalindua clades 1 and 2. Scalindua 1 could be further divided into four distinct clusters, all comprised of sequences from either the Namibian or the Peruvian OMZ. Scalindua 2 consisted of sequences from the Arabian Sea and the Peruvian OMZ and included one previously published 16S rRNA gene sequence from Lake Tanganyika and one from South China Sea sediment (97.9-99.4% sequence identity). This cluster showed only 相似文献   

Physiological Characterization of an Anaerobic Ammonium-Oxidizing Bacterium Belonging to the “Candidatus Scalindua” Group

The phylogenetic affiliation and physiological characteristics (e.g., K_s and maximum specific growth rate [μ_max]) of an anaerobic ammonium oxidation (anammox) bacterium, “Candidatus Scalindua sp.,” enriched from the marine sediment of Hiroshima Bay, Japan, were investigated. “Candidatus Scalindua sp.” exhibits higher affinity for nitrite and a lower growth rate and yield than the known anammox species.     

Community Structures and Distribution of Anaerobic Ammonium Oxidizing and nirS-Encoding Nitrite-Reducing Bacteria in Surface Sediments of the South China Sea

Anaerobic ammonium oxidation (anammox) and denitrification are two important processes responsible for nitrogen loss; monitoring of microbial communities carrying out these two processes offers a unique opportunity to understand the microbial nitrogen cycle. The aim of the current study was to characterize community structures and distribution of anammox and nirS-encoding nitrite-reducing bacteria in surface sediments of the northern South China Sea (SCS). The consistent phylogenetic results of three biomarkers of anammox bacteria, including 16S rRNA, hzo, and Scalindua-nirS genes, showed that Scalindua-like bacteria were the only anammox group presenting in surface sediments of the SCS. However, a relatively high micro-diversity was found within this group, including several SCS habitat-specific phylotypes, Candidatus “Scalindua zhenghei”. Comparing to 16S rRNA gene, hzo and Scalindua-nirS genes provided a relatively higher resolution to elucidate anammox bacteria. For the nirS-encoding nitrite-reducing bacteria, the detected nirS gene sequences were closely related to various marine nirS denitrifiers, especially those which originated from coastal and estuarine sediments with a much higher diversity than anammox bacteria. Anammox bacterial communities shifted along with the seawater depth, while nirS-encoding nitrite-reducing bacteria did not. Although nirS-encoding nitrite-reducing bacteria have a much higher abundance and diversity than anammox bacteria, they showed similar abundance variation patterns in research sites, suggesting the two microbial groups might be affected by the similar environmental factors. The significant correlations among the abundance of the two microbial groups with the molar ratio of NH₄ ⁺ to (NO₂ ^??+?NO₃ ^?), pH, and organic matters of sediments strongly supported this hypothesis.     

Taxonomical and functional microbial community dynamics in an Anammox-ASBR system under different Fe (III) supplementation

In the present study, we explored the metabolic versatility of anaerobic ammonium oxidation (anammox) bacteria in a variety of Fe (III) concentrations. Specifically, we investigated the impacts of Fe (III) on anammox growth rates, on nitrogen removal performance, and on microbial community dynamics. The results from our short-term experiments revealed that Fe (III) concentrations (0.04–0.10 mM) significantly promote the specific anammox growth rate from 0.1343 to 0.1709 d^?1. In the long-term experiments, the Anammox-anaerobic sequencing batch reactor (ASBR) was operated over 120 days and achieved maximum NH₄ ⁺-N, NO₂ ^?-N, and TN efficiencies of 90.98 ± 0.35, 93.78 ± 0.29, and 83.66 ± 0.46 %, respectively. Pearson’s correlation coefficients between anammox-(narG + napA), anammox-nrfA, and anammox-FeRB all exceeded r = 0.820 (p < 0.05), confirming an interaction and ecological association among the nitrogen and iron-cycling-related microbial communities. Illumina MiSeq sequencing indicated that Chloroflexi (34.39–39.31 %) was the most abundant phylum in an Anammox-ASBR system, followed by Planctomycetes (30.73–35.31 %), Proteobacteria (15.40–18.61 %), and Chlorobi (4.78–6.58 %). Furthermore, we found that higher Fe (III) supplementation (>0.06 mM) could result in the community succession of anammox species, in which Candidatus Brocadia and Candidatus Kuenenia were the dominant anammox bacteria species. Combined analyses indicated that the coupling of anammox, dissimilatory nitrogen reduction to ammonium, and iron reduction accounted for nitrogen loss in the Anammox-ASBR system. Overall, the knowledge gained in this study provides novel insights into the microbial community dynamics and metabolic potential of anammox bacteria under Fe (III) supplementation.     

Development of a fixed-bed anammox reactor with high treatment potential

A plug-flow type anaerobic ammonium oxidation (anammox) reactor was developed using malt ceramics (MC) produced from carbonized spent grains as the biomass carriers for anammox sludge. Partial nitrified effluent of the filtrate from the sludge dehydrator of a brewery company was used as influent to a 20 L anammox reactor using MC. An average volumetric nitrogen removal rate (VNR) of 8.78 kg-N/m³/day was maintained stably for 76 days with 1 h of HRT. In a larger anammox reactor (400 L), an average VNR of 4.84 kg-N/m³/day could be maintained for 86 days during the treatment of low strength synthetic inorganic wastewater. As a result of bacterial community analysis for the 20 L anammox reactor, Asahi BRW1, probably originating from the wastewater collected at Asahi Breweries, was detected as the dominant anammox bacterium. These anammox reactors were characterized by a high NH₄-N removal capacity for low strength wastewater with a short hydraulic retention time.     

Chronic effect of nitrite on the rearing of the white shrimp Litopenaeus vannamei in two salinities

The aim of this study was to determine the effects of nitrite on the growth and survival of the white shrimp L. vannamei in two different salinities. Nitrite concentrations tested in salinity 8 g/L were 0 (control), 2.5, 5.0, 10.0, and 20.0 mg NO₂^?-N/L, and in salinity 24 g/L were 0 (control), 5.0, 10.0, 20.0, and 40.0 mg NO₂^?-N/L. For these experiments, 30 experimental units with 30?L of useful volume were stocked with 20 juvenile L. vannamei (8.0 ± 0.50 g), corresponding to a stocking density of 100 shrimp/m², and cultivated for an experimental period of 30 days. A significant difference was found between the control and treatment groups with respect to growth and survival. The 2.5 mg NO₂^?-N/L treatment showed the best performance indexes in salinity 8 g/L, while the best growth performance indexes were found in the control and 5.0 mg NO₂^?-N/L treatments in salinity 24 g/L. Total mortality was observed in the 10 and 20 mg NO₂^?-N/L treatment groups from salinity 8 g/L and in the 40 mg NO₂^?-N/L treatment group in salinity 24 g/L. This study determined that concentrations of nitrite of up to 2.5 and 10 mg/L are acceptable for the rearing of L. vannamei in salinities of 8 and 24 g/L, respectively.     

Spatial distribution of diatom and cyanobacterial mats in the Dead Sea is determined by response to rapid salinity fluctuations

Cyanobacteria and diatom mats are ubiquitous in hypersaline environments but have never been observed in the Dead Sea, one of the most hypersaline lakes on Earth. Here we report the discovery of phototrophic microbial mats at underwater freshwater seeps in the Dead Sea. These mats are either dominated by diatoms or unicellular cyanobacteria and are spatially separated. Using in situ and ex situ O₂ microsensor measurements we show that these organisms are photosynthetically active in their natural habitat. The diatoms, which are phylogenetically associated to the Navicula genus, grew in culture at salinities up to 40 % Dead Sea water (DSW) (14 % total dissolved salts, TDS). The unicellular cyanobacteria belong to the extremely halotolerant Euhalothece genus and grew at salinities up to 70 % DSW (24.5 % TDS). As suggested by a variable O₂ penetration depth measured in situ, the organisms are exposed to drastic salinity fluctuations ranging from brackish to DSW salinity within minutes to hours. We could demonstrate that both phototrophs are able to withstand such extreme short-term fluctuations. Nevertheless, while the diatoms recover better from rapid fluctuations, the cyanobacteria cope better with long-term exposure to DSW. We conclude that the main reason for the development of these microbial mats is a local dilution of the hypersaline Dead Sea to levels allowing growth. Their spatial distribution in the seeping areas is a result of different recovery rates from short or long-term fluctuation in salinity.     

Picophytoplankton predominance in hypersaline lakes (Transylvanian Basin,Romania)

The occurrence and importance of photoautotrophic picoplankton (PPP, cells with a diameter <2 μm) was studied along a trophic and salinity gradient in hypersaline lakes of the Transylvanian Basin (Romania). The studied lakes were found to be rich in PPP, with abundances (maximum 7.6 × 10⁶ cells mL^?1) higher than in freshwater and marine environments of similar trophic conditions. The contribution of PPP to the total phytoplankton biovolume did not decrease with increasing trophic state as it was generally found in other aquatic environments. Regardless of the trophic conditions, the contribution of PPP could reach 90–100 % in these hypersaline lakes. We hypothesized that the PPP predominance might be the result of the low grazing pressure, since heterotrophic nanoflagellates (the main grazers of PPP) were absent in the studied samples. There were significant differences in community composition among the lakes along the salinity gradient. CyPPP predominated in less saline waters (mainly below 5 %), while EuPPP were present along the entire salinity range (up to 18.7 %), dominating the phytoplankton between 3 and 13 % salinity. Above 13 % salinity, the phytoplankton was composed mainly of Dunaliella species.     

Morphological study of Cyclotella choctawhatcheeana Prasad (Stephanodiscaceae) from a saline Mexican lake

Background

Extracellular dissolved DNA has been demonstrated to be present in many terrestrial and aquatic environments, actively secreted, or released by decaying cells. Free DNA has the genetic potential to be acquired by living competent cells by horizontal gene transfer mediated by natural transformation. The aim of this work is to study the persistence of extracellular DNA and its biological transforming activity in extreme environments like the deep hypersaline anoxic lakes of the Mediterranean Sea. The brine lakes are separated from the upper seawater by a steep chemocline inhabited by stratified prokaryotic networks, where cells sinking through the depth profile encounter increasing salinity values and osmotic stress.

Results

Seven strains belonging to different taxonomic groups isolated from the seawater-brine interface of four hypersaline lakes were grown at medium salinity and then incubated in the brines. The osmotic stress induced the death of all the inoculated cells in variable time periods, between 2 hours and 144 days, depending on the type of brine rather than the taxonomic group of the strains, i.e. Bacillaceae or gamma-proteobacteria. The Discovery lake confirmed to be the most aggressive environment toward living cells. In all the brines and in deep seawater dissolved plasmid DNA was substantially preserved for a period of 32 days in axenic conditions. L'Atalante and Bannock brines induced a decrease of the supercoiled form up to 70 and 40% respectively; in the other brines only minor changes in plasmid conformation were observed. Plasmid DNA after incubation in the brines maintained the capacity to transform naturally competent cells of Acinetobacter baylii strain BD413.

Conclusion

Free dissolved DNA is likely to be released by the lysis of cells induced by osmotic stress in the deep hypersaline anoxic lakes. Naked DNA was demonstrated to be preserved and biologically active in these extreme environments, and hence could constitute a genetic reservoir of traits acquirable by horizontal gene transfer.     

Complex nitrogen cycling in the sponge Geodia barretti

Marine sponges constitute major parts of coral reefs and deep‐water communities. They often harbour high amounts of phylogenetically and physiologically diverse microbes, which are so far poorly characterized. Many of these sponges regulate their internal oxygen concentration by modulating their ventilation behaviour providing a suitable habitat for both aerobic and anaerobic microbes. In the present study, both aerobic (nitrification) and anaerobic (denitrification, anammox) microbial processes of the nitrogen cycle were quantified in the sponge Geodia barretti and possible involved microbes were identified by molecular techniques. Nitrification rates of 566 nmol N cm^?3 sponge day^?1 were obtained when monitoring the production of nitrite and nitrate. In support of this finding, ammonia‐oxidizing Archaea (crenarchaeotes) were found by amplification of the amoA gene, and nitrite‐oxidizing bacteria of the genus Nitrospira were detected based on rRNA gene analyses. Incubation experiments with stable isotopes (¹⁵NO₃^‐ and ¹⁵NH₄⁺) revealed denitrification and anaerobic ammonium oxidation (anammox) rates of 92 nmol N cm^?3 sponge day^?1 and 3 nmol N cm^?3 sponge day^?1 respectively. Accordingly, sequences closely related to ‘Candidatus Scalindua sorokinii’ and ‘Candidatus Scalindua brodae’ were detected in 16S rRNA gene libraries. The amplification of the nirS gene revealed the presence of denitrifiers, likely belonging to the Betaproteobacteria. This is the first proof of anammox and denitrification in the same animal host, and the first proof of anammox and denitrification in sponges. The close and complex interactions of aerobic, anaerobic, autotrophic and heterotrophic microbial processes are fuelled by metabolic waste products of the sponge host, and enable efficient utilization and recirculation of nutrients within the sponge–microbe system. Since denitrification and anammox remove inorganic nitrogen from the environment, sponges may function as so far unrecognized nitrogen sinks in the ocean. In certain marine environments with high sponge cover, sponge‐mediated nitrogen mineralization processes might even be more important than sediment processes.