Diversity in the utilization of different molecular classes of dissolved organic matter by heterotrophic marine bacteria.

Heterotrophic marine bacteria utilize and recycle dissolved organic matter (DOM), impacting biogeochemical cycles. It is currently unclear to what extent distinct DOM components can be used by different heterotrophic clades. Here, we ask how a natural microbial community from the Eastern Mediterranean Sea (EMS) responds to different molecular classes of DOM (peptides, amino acids, amino sugars, disaccharides, monosaccharides, and organic acids) comprising much of the biomass of living organisms. Bulk bacterial activity increased after 24 h for all treatments relative to the control, while glucose and ATP uptake decreased or remained unchanged. Moreover, while the per-cell uptake rate of glucose and ATP decreased, that of Leucin significantly increased for amino acids, reflecting their importance as common metabolic currencies in the marine environment. Pseudoalteromonadaceae dominated the peptides treatment, while different Vibrionaceae strains became dominant in response to amino acids and amino sugars. Marinomonadaceae grew well on organic acids, and Alteromonadaseae on disaccharides. A comparison with a recent laboratory-based study reveals similar peptide preferences for Pseudoalteromonadaceae, while Alteromonadaceae, for example, grew well in the lab on many substrates but dominated in seawater samples only when disaccharides were added. We further demonstrate a potential correlation between the genetic capacity for degrading amino sugars and the dominance of specific clades in these treatments. These results highlight the diversity in DOM utilization among heterotrophic bacteria and complexities in the response of natural communities. IMPORTANCE: A major goal of microbial ecology is to predict the dynamics of natural communities based on the identity of the organisms, their physiological traits, and their genomes. Our results show that several clades of heterotrophic bacteria each grow in response to one or more specific classes of organic matter. For some clades, but not others, growth in a complex community is similar to that of isolated strains in laboratory monoculture. Additionally, by measuring how the entire community responds to various classes of organic matter, we show that these results are ecologically relevant, and propose that some of these resources are utilized through common uptake pathways. Tracing the path between different resources to the specific microbes that utilize them, and identifying commonalities and differences between different natural communities and between them and lab cultures, is an important step toward understanding microbial community dynamics and predicting how communities will respond to perturbations.

Discharge of Polyphosphonate-Based Antiscalants via Desalination Brine: Impact on Seabed Nutrient Flux and Microbial Activity.

Desalination brine is a hypersaline byproduct that contains various operational chemicals such as polyphosphonate-based antiscalants. Brine often sinks and flows over the seabed by density currents; therefore, it may affect sediment-water nutrient fluxes and thus microbial activity. We quantified these parameters in brine plumes around two large-scale desalination facilities located in the P-limited Southeastern Mediterranean Sea. The benthic nutrient fluxes and microbial activity were determined using ex-situ core benthocosms, to which we added brine from the dispersion area in excess salinities of ∼3% and 5% above natural levels. A higher influx of dissolved organic phosphorus (∼6-fold) and an efflux of dissolved organic carbon (∼1.7-fold) were measured in the brine-amended cores relative to the controls. This was accompanied by increased oxygen consumption (15%) and increased microbial activity (∼1.5-6.5-fold). Field observations support the results from experimental manipulations, yielding ∼4.5-fold higher microbial activity rates around the brine plume compared to uninfluenced locations. Our results imply that desalination brine can alter sedimentary processes affecting benthic nutrients inventories. Moreover, we show that brine acts as a vector of anthropogenic P, stimulating microbial activity in the sediment-water interface.

Trimethylated homoserine functions as the major compatible solute in the globally significant oceanic cyanobacterium Trichodesmium.

The oceanic N2-fixing cyanobacterium Trichodesmium spp. form extensive surface blooms and contribute significantly to marine carbon and nitrogen cycles in the oligotrophic subtropical and tropical oceans. Trichodesmium grows in salinities from 27 to 43 parts per thousand (ppt), yet its salt acclimation strategy remains enigmatic because the genome of Trichodesmium erythraeum strain IMS101 lacks all genes for the biosynthesis of any known compatible solute. Using NMR and liquid chromatography coupled to mass spectroscopy, we identified the main compatible solute in T. erythraeum strain IMS101 as the quaternary ammonium compound N,N,N-trimethyl homoserine (or homoserine betaine) and elucidated its biosynthetic pathway. The identification of this compatible solute explains how Trichodesmium spp. can thrive in the marine system at varying salinities and provides further insight into the diversity of microbial salt acclimation.

Enhanced salinities, as a proxy of seawater desalination discharges, impact coastal microbial communities of the eastern Mediterranean Sea.

Seawater desalination plants increase local coastal salinities by discharging concentrated brine back to the sea with ∼ 50% higher than ambient salinities. The impacts of high salinities on microbial coastal populations of the eastern Mediterranean Sea (EMS) were examined in two mesocosm experiments; first, during the mixed-spring and second, during the stratified-summer periods with average salinity of ∼ 39. Ambient salinities were increased by 5% and 15%. Higher salinity (15%) mesocosms induced rapid (within 2 h) declines in both primary productivity (PP) and algal biomass parallel to an increase in bacterial productivity. Subsequently, for the duration of the experiments (11-12 days), both Chlorophyll a and PP rates increased (2 to 5 and 1.5 to 2.5-fold, respectively) relative to unamended controls. The initial assemblages of the ambient microbial populations and intensity of salinity enrichments influenced the community responses. During the mixed-spring experiment, the composition of prokaryotic and eukaryotic populations shifted only slightly, suggesting high functional plasticity of the initial populations. While during the stratified-summer experiment, high salinity changed the composition and reduced the biodiversity of the microbial communities. In an ultra-oligotrophic environment such as the EMS, salinity induced declines in microbial diversity may provide a tipping point destabilizing the local aquatic food web.

Active microbial communities facilitate carbon turnover in brine pools found in the deep Southeastern Mediterranean Sea.

Discharge of gas-rich brines fuels productive chemosynthetic ecosystems in the deep sea. In these salty, methanic and sulfidic brines, microbial communities adapt to specific niches along the physicochemical gradients. However, the molecular mechanisms that underpin these adaptations are not fully known. Using metagenomics, we investigated the dense (∼106 cell ml-1) microbial communities that occupy small deep-sea brine pools found in the Southeastern Mediterranean Sea (1150 m water depth, ∼22 °C, ∼60 PSU salinity, sulfide, methane, ammonia reaching millimolar levels, and oxygen usually depleted), reaching high productivity rates of 685 µg C L-1 d-1 ex-situ. We curated 266 metagenome-assembled genomes of bacteria and archaea from the several pools and adjacent sediment-water interface, highlighting the dominance of a single Sulfurimonas, which likely fuels its autotrophy using sulfide oxidation or inorganic sulfur disproportionation. This lineage may be dominant in its niche due to genome streamlining, limiting its metabolic repertoire, particularly by using a single variant of sulfide: quinone oxidoreductase. These primary producers co-exist with ANME-2c archaea that catalyze the anaerobic oxidation of methane. Other lineages can degrade the necromass aerobically (Halomonas and Alcanivorax), or anaerobically through fermentation of macromolecules (e.g., Caldatribacteriota, Bipolaricaulia, Chloroflexota, etc). These low-abundance organisms likely support the autotrophs, providing energy-rich H2, and vital organics such as vitamin B12.

Shared ancestry of algal symbiosis and chloroplast sequestration in foraminifera.

Foraminifera are unicellular organisms that established the most diverse algal symbioses in the marine realm. Endosymbiosis repeatedly evolved in several lineages, while some engaged in the sequestration of chloroplasts, known as kleptoplasty. So far, kleptoplasty has been documented exclusively in the rotaliid clade. Here, we report the discovery of kleptoplasty in the species Hauerina diversa that belongs to the miliolid clade. The existence of kleptoplasty in the two main clades suggests that it is more widespread than previously documented. We observed chloroplasts in clustered structures within the foraminiferal cytoplasm and confirmed their functionality. Phylogenetic analysis of 18S ribosomal RNA gene sequences showed that H. diversa branches next to symbiont-bearing Alveolinidae. This finding represents evidence of of a relationship between kleptoplastic and symbiotic foraminifera.. Analysis of ribosomal genes and metagenomics revealed that alveolinid symbionts and kleptoplasts belong to the same clade, which suggests a common ancestry.

Diversity, activity, and abundance of benthic microbes in the Southeastern Mediterranean Sea.

Benthic microbes are key organisms in the oligotrophic Southeastern Mediterranean Sea (SEMS), yet their abundance, activity, and diversity in this rapidly changing basin are not fully understood. We investigated the prokaryotic and microfungal communities throughout years 2018-2020 at 27 stations (6-1900 m water depths, down to 20 cm below the sediment surface), in two transects with distinct downslope transport regimes, and along the eutrophic coastline. We estimated microbial abundance with flow cytometry, secondary production as leucine assimilation, and sequenced marker genes (the 16S rRNA and internal transcribed spacer) to assess diversity indices. The highest abundance (0.21 × 108 cells gr-1 sediment) was estimated at slope stations where we assumed substantial transport rates and found an accumulation of organic carbon. Secondary production was the highest nearshore (12 ± 4 ng C gr-1 h-1), and markedly declined offshore (0.5 ± 0.9 ng C gr-1 h-1). Populations of archaea (dominant Nitrososphaeria and Nanoarchaeia) and diverse bacteria were stable over three years, and taxonomic composition was dictated mainly by depth gradients. Saprotrophic and pathotrophic microfungi Ascomycota (70% ± 23%) and Basidiomycota (16% ± 18%) were prevalent, whereas parasitic chytrids were abundant nearshore. Our results highlight the role of downslope transport, which enriched the typical deep-sea communities with anaerobic lineages, in shaping microbial populations near the continental slope.

Jellyfish swarm impair the pretreatment efficiency and membrane performance of seawater reverse osmosis desalination.

Circumstantial evidence has suggested that jellyfish swarms impair the operation of seawater reverse osmosis desalination facilities. However, only limited information is currently available on the pretreatment efficiency of jellyfish and their effects on reverse osmosis (RO) membrane performance. Here, we have comprehensively tested the pretreatment efficiency of a dual-media gravity filter and cartridge micro-filtration following the addition of jellyfish into the feedwater. Concurrently, the fouling propensity and performance of the RO membranes were examined. We show that jellyfish demise resulted in seawater eutrophication that triggered a significant increase in bacterial biomass (∼50-fold), activity (∼7-fold), and release of transparent exopolymer particles (∼5-fold), peaking three days after the addition of jellyfish into the feedwater. In parallel, a significant reduction in permeate water flux was recorded (∼10%) while trans-membrane pressure sharply increased (15%), reaching the operation pressure limit of our system (75 bar) after five days. At the conclusion of the experiments, the membrane surface was heavily covered by large chunks of organic-rich material and multilayered biofilms. Our results provide a holistic view on the operational challenges of seawater reverse osmosis (SWRO) desalination triggered by jellyfish swarms in coastal areas. Following the above, it can be inferred that freshwater production will likely be halted three days after drawing the jellyfish into the pretreatment system. Outcomes from these results may lead to the development of science-based operational protocols to cope with growing occurrence of jellyfish swarms around the intake of SWRO desalination facilities worldwide.

Impacts of sewage outbursts on seawater reverse osmosis desalination.

Sewage outbursts affect coastal environments as seawater is enriched with nutrients, organic matter and microbes, thus can potentially impair seawater reverse osmosis (SWRO) desalination. In this study, we evaluated how municipal sewage outbursts affect SWRO desalination in a pilot-scale system. To this end, feedwater characteristics (i.e., coastal water), the removal efficiency of organic foulants by a dual-media gravity filter, and cartridge micro-filtration were determined daily for 12 days. Permeate water flux was maintained constant during the study, while trans-membrane pressure (TMP) was automatically adjusted and continuously monitored. The results indicate that sewage outbursts caused an immediate (∼1 d) buildup of phyto/bacterioplankton biomass (up to 10-fold), and enhanced activity (maximal 30-fold) followed by an increase in transparent exopolymer particle (TEP) concentrations. After sewage addition, algal biomass was significantly removed by the pretreatment system (72-90%), while a considerable fraction of the bacterial biomass (42-65%) and TEP (53-65%) passed these procedures. The result was a negative impact on the desalination performance reflected by a significant increase (> 10%) in RO-TMP 7.5 d after the sewage addition. Our results indicate on a direct link between sewage outbursts, pretreatment efficiency, and SWRO desalination. Nevertheless, these findings can lead to new avenues for the development of science-based operational protocols to minimize the deleterious effects of abrupt sewage outbursts on SWRO desalination.

Dust-borne microbes affect Ulva ohnoi's growth and physiological state.

The marine macroalgae Ulva sp. is considered an ecosystem engineer in rocky shores of temperate waters worldwide. Ulva sp. harbors a rich diversity of associated microbial epibionts, which are known to affect the algae's typical morphological development and 'health'. We examined the interaction between airborne microbes derived from atmospheric aerosols and Ulva ohnoi growth and physiological state. Specifically, we measured U. ohnoi growth rates and photosynthetic efficiency (Fv/Fm), alongside its microbial epibionts abundance, activity and diversity following dust (containing nutrients and airborne microorganisms) or UV-treated dust (only nutrients) amendments to filtered seawater. Parallel incubations with epibionts-free U. ohnoi (treated with antibiotics that removed the algae epibionts) were also tested to specifically examine if dust-borne microbes can replenish the epibiont community of U. ohnoi. We show that viable airborne microbes can restore U. ohnoi natural microbial epibionts communities, thereby keeping the seaweed alive and 'healthy'. These results suggest that microbes delivered through atmospheric aerosols can affect epiphyte biodiversity in marine flora, especially in areas subjected to high annual atmospheric dust deposition such as the Mediterranean Sea.

Phytoplankton response to N-rich well amelioration brines: A mesocosm study from the southeastern Mediterranean Sea.

Human-induced eutrophication of coastal water may be a major threat to aquatic life. Here, we investigated the effects of N-rich well amelioration brines (WAB) on coastal phytoplankton population's habitat in the surface oligotrophic waters of the southeastern Mediterranean Sea (SEM). To this end, we added WAB (2 concentrations) to mesocosms (1-m3 bags) to surface SEM water during summer and winter, where changes in phytoplankton biomass, activity and diversity was monitored daily for 8 days. Our results demonstrate that WAB addition triggered a phytoplankton bloom, resulting in elevated algal biomass (maximal +780%), increased primary production rates (maximal +675%) and a decrease in eukaryotic algal α-diversity (ca. -20%). Among the species that bloomed following WAB amendments, we found the potentially toxic dinoflagellate Karlodinium venificum. This study adds valuable perspective to the effect of nutrients discharged into nutrient limited SEM coastal waters, and in particular of N-derived WAB.

Benefit from decline: the primary transcriptome of Alteromonas macleodii str. Te101 during Trichodesmium demise.

Interactions between co-existing microorganisms deeply affect the physiology of the involved organisms and, ultimately, the function of the ecosystem as a whole. Copiotrophic Alteromonas are marine gammaproteobacteria that thrive during the late stages of phytoplankton blooms in the marine environment and in laboratory co-cultures with cyanobacteria such as Trichodesmium. The response of this heterotroph to the sometimes rapid and transient changes in nutrient supply when the phototroph crashes is not well understood. Here, we isolated and sequenced the strain Alteromonas macleodii str. Te101 from a laboratory culture of Trichodesmium erythraeum IMS101, yielding a chromosome of 4.63 Mb and a single plasmid of 237 kb. Increasing salinities to ≥43 ppt inhibited the growth of Trichodesmium but stimulated growth of the associated Alteromonas. We characterized the transcriptomic responses of both microorganisms and identified the complement of active transcriptional start sites in Alteromonas at single-nucleotide resolution. In replicate cultures, a similar set of genes became activated in Alteromonas when growth rates of Trichodesmium declined and mortality was high. The parallel activation of fliA, rpoS and of flagellar assembly and growth-related genes indicated that Alteromonas might have increased cell motility, growth, and multiple biosynthetic activities. Genes with the highest expression in the data set were three small RNAs (Aln1a-c) that were identified as analogs of the small RNAs CsrB-C in E. coli or RsmX-Z in pathogenic bacteria. Together with the carbon storage protein A (CsrA) homolog Te101_05290, these RNAs likely control the expression of numerous genes in responding to changes in the environment.

The effect of coagulants and antiscalants discharged with seawater desalination brines on coastal microbial communities: A laboratory and in situ study from the southeastern Mediterranean.

Desalination outflows frequently discharge brine containing coagulants and antiscalants (e.g. Iron-hydroxides and polyphosphonates) to the coastal environment. Here we examined changes in composition and productivity of natural microbial coastal communities in experimental mesocosms treated with either iron-hydroxide (Fe), polyphosphonate (Pn), or a combination of high salinities with both chemicals (All). Within 2 h of addition Fe already altered the microbial community composition, enhanced the bacterial production (BP) and cell specific production (BP/BA), and decreased primary production. Addition of Pn, relieved phosphorus stress as demonstrated by the immediate (within 2 h) and significant reduction in the ecto-enzyme alkaline phosphatase activity (APA). Synergistic effects were observed in the All treatment, reflected by increased production of both primary and bacterial producers as P-stress was relieved. After 10 days of incubation, the microbial community composition changed significantly only in the All treatment. The Fe-only treatment caused a significant decline in autotrophic biomass and in the assimilation number (AN), while in both the Pn and the All treatments the BP/BA increased with the added P. We also examined the microbial community responses in a natural impacted environment at the Ashkelon seawater desalination plant brine discharge site during summer and winter. The community composition differed in elevated-salinity compared with non-impacted stations with higher AN and bacterial efficiencies (BP/BA) measured in summer in the elevated-salinity stations. The seasonal differences in responses may reflect both biotic (i.e. initial community composition) and abiotic factors (currents and residence time of salinity gradients). Our results emphasize that desalination brine discharges that include chemicals such as iron-hydroxide and polyphosphonates can induce physiological and compositional changes in the microbial community. With the expansion of desalination facilities worldwide such shifts in composition and function of the microbial communities may destabilize and change local aquatic food webs and should thus be monitored.

The primary transcriptome of the marine diazotroph Trichodesmium erythraeum IMS101.

Blooms of the dinitrogen-fixing marine cyanobacterium Trichodesmium considerably contribute to new nitrogen inputs into tropical oceans. Intriguingly, only 60% of the Trichodesmium erythraeum IMS101 genome sequence codes for protein, compared with ~85% in other sequenced cyanobacterial genomes. The extensive non-coding genome fraction suggests space for an unusually high number of unidentified, potentially regulatory non-protein-coding RNAs (ncRNAs). To identify the transcribed fraction of the genome, here we present a genome-wide map of transcriptional start sites (TSS) at single nucleotide resolution, revealing the activity of 6,080 promoters. We demonstrate that T. erythraeum has the highest number of actively splicing group II introns and the highest percentage of TSS yielding ncRNAs of any bacterium examined to date. We identified a highly transcribed retroelement that serves as template repeat for the targeted mutation of at least 12 different genes by mutagenic homing. Our findings explain the non-coding portion of the T. erythraeum genome by the transcription of an unusually high number of non-coding transcripts in addition to the known high incidence of transposable elements. We conclude that riboregulation and RNA maturation-dependent processes constitute a major part of the Trichodesmium regulatory apparatus.

Bioflocculation: chemical free, pre-treatment technology for the desalination industry.

Rapid sand filtration (RSF), proceeded by chemical coagulation and flocculation, is a commonly used, effective pretreatment in the desalination industry. We designed and tested a novel, large pilot-scale, two-stage granular Rapid Bioflocculation Filter (RBF) based on a first-stage Bioflocculator (BF) unit followed by a mixed-media bed filter (MBF). The BF filter bed consisted of an extremely porous volcanic Tuff granular medium which provided an enlarged surface area for microbial development and biofilm proliferation. We compared the efficiency of the pilot RBF to that of a full-scale RSF, operating with upstream chemical coagulation, by measuring the removal from the same untreated seawater feed of key factors related to membrane clogging: SDI, turbidity, chlorophyll a (Chl a) and transparent exopolymer particles (TEP). After 2 weeks of operation, the Tuff grains were colonized extensively by coccoid bacteria that formed biofilm along the entire BF. With bacterial colonization and biofilm development, numerous aggregates of bacteria and some algal cells embedded in an amorphous organic matrix were formed on and within the Tuff grains. By 1-3 months, the biotic diversity within the Tuff filter bed had increased to include filamentous bacteria, cyanobacteria, fungi, protista and even crustaceans and marine worms. During and for ≈ 24 h after each cleaning cycle (carried out every 5 to 7 days by upward flushing with air and water), large numbers of floc-like particles, from ≈ 15 µm to ≈ 2 mm in size were observed in the filtrate of the BF unit. Microscopic examination of these flocs (stained with Alcian Blue and SYTO(R) 9) showed that they were aggregates of many smaller particles with associated bacteria and algae within a polysaccharide gel-like matrix. These biogenic flocs (bioflocs) were observed to form during normal operation of the RBF, accumulating as aggregates of inorganic and organic material on the Tuff surfaces. With each flush cleaning cycle, these bioflocs were released into the BF effluent but were retained by the second phase MBF unit. No flocs were seen in the MBF filtrate. Over a year-long study, both the pilot RBF and the full-scale RSF showed similar filtration efficiencies, measured as the percentage removal of Chl a, TEP, turbidity and SDI from the same seawater feed. These results indicate the potential of the bioflocculation approach with no chemical additives as an alternative to conventional RSF pretreatment for large SWRO facilities.

Dinitrogen fixation in aphotic oxygenated marine environments.

We measured N2 fixation rates from oceanic zones that have traditionally been ignored as sources of biological N2 fixation; the aphotic, fully oxygenated, nitrate (NO(-) 3)-rich, waters of the oligotrophic Levantine Basin (LB) and the Gulf of Aqaba (GA). N2 fixation rates measured from pelagic aphotic waters to depths up to 720 m, during the mixed and stratified periods, ranged from 0.01 nmol N L(-1) d(-1) to 0.38 nmol N L(-1) d(-1). N2 fixation rates correlated significantly with bacterial productivity and heterotrophic diazotrophs were identified from aphotic as well as photic depths. Dissolved free amino acid amendments to whole water from the GA enhanced bacterial productivity by 2-3.5 fold and N2 fixation rates by ~2-fold in samples collected from aphotic depths while in amendments to water from photic depths bacterial productivity increased 2-6 fold while N2 fixation rates increased by a factor of 2 to 4 illustrating that both BP and heterotrophic N2 fixation were carbon limited. Experimental manipulations of aphotic waters from the LB demonstrated a significant positive correlation between transparent exopolymeric particle (TEP) concentrations and N2 fixation rates. This suggests that sinking organic material and high carbon (C): nitrogen (N) micro-environments (such as TEP-based aggregates or marine snow) could support high heterotrophic N2 fixation rates in oxygenated surface waters and in the aphotic zones. Indeed, our calculations show that aphotic N2 fixation accounted for 37 to 75% of the total daily integrated N2 fixation rates at both locations in the Mediterranean and Red Seas with rates equal or greater to those measured from the photic layers. Moreover, our results indicate that that while N2 fixation may be limited in the surface waters, aphotic, pelagic N2 fixation may contribute significantly to new N inputs in other oligotrophic basins, yet it is currently not included in regional or global N budgets.