Dynamic bacterial community response to Akashiwo sanguinea (Dinophyceae) bloom in indoor marine microcosms.

We investigated the dynamics of the bacterial composition and metabolic function within Akashiwo sanguinea bloom using a 100-L indoor microcosm and metagenomic next-generation sequencing. We found that the bacterial community was classified into three groups at 54% similarity. Group I was associated with "during the A. sanguinea bloom stage" and mainly consisted of Alphaproteobacteria, Flavobacteriia and Gammaproteobacteria. Meanwhile, groups II and III were associated with the "late bloom/decline stage to post-bloom stage" with decreased Flavobacteriia and Gammaproteobacteria in these stages. Upon the termination of the A. sanguinea bloom, the concentrations of inorganic nutrients (particularly PO43-, NH4+ and dissolved organic carbon) increased rapidly and then decreased. From the network analysis, we found that the A. sanguinea node is associated with certain bacteria. After the bloom, the specific increases in NH4+ and PO43- nodes are associated with other bacterial taxa. The changes in the functional groups of the bacterial community from chemoheterotrophy to nitrogen association metabolisms were consistent with the environmental impacts during and after A. sanguinea bloom. Consequently, certain bacterial communities and the environments dynamically changed during and after harmful algal blooms and a rapid turnover within the bacterial community and their function can respond to ecological interactions.

Use of plant materials for the bioremediation of soil from an industrial site.

Bioremediation is one of the existing techniques applied for treating oil-contaminated soil, which can be improved by the incorporation of low-cost nutritional materials. This study aimed to assess the addition of two low-cost plant residues, sugarcane bagasse (SCB) and leaf litter (LL) of the forest leguminous Mimosa caesalpiniifolia plant (sabiá), either separately or combined, to a contaminated soil from a petroleum refinery area, analyzed after 90 days of treatment. Individually, both amounts of SCB (20 and 40 g kg-1) favored the growth of total heterotrophic bacteria and total fungi, while LL at 20 g kg-1 better stimulated the hydrocarbon-degrading microorganism's activity in the soil. However, no TPH removal was observed under any of these conditions. Higher microbial growth was detected by the application of both plant residues in multicontaminated soil. The maximum TPH removal of 30% was achieved in amended soil with 20 g kg-1 SCB and 20 kg-1 LL. All the experimental conditions revealed changes in the microbial community structure, related to the handling of the soil, with abundance of Alphaproteobacteria. This study demonstrates the effectiveness of the plant residues SCB and LL as low-cost nutritional materials for biodegradation of hydrocarbon in real oil contaminated soil by indigenous populations.

Plant powder teabags: a novel and practical approach to resolve culturability and diversity of rhizobacteria.

We have developed teabags packed with dehydrated plant powders, without any supplements, for preparation of plant infusions necessary to develop media for culturing rhizobacteria. These bacteria are efficiently cultivated on such plant teabag culture media, with better progressive in situ recoverability compared to standard chemically synthetic culture media. Combining various plant-based culture media and incubation conditions enabled us to resolve unique denaturing gradient gel electrophoresis (DGGE) bands that were not resolved by tested standard culture media. Based on polymerase chain reaction PCR-DGGE of 16S rDNA fingerprints and sequencing, the plant teabag culture media supported higher diversity and significant increases in the richness of endo-rhizobacteria, namely Gammaproteobacteria (Enterobacteriaceae) and predominantly Alphaproteobacteria (Rhizobiaceae). This culminated in greater retrieval of the rhizobacteria taxa associated with the plant roots. We conclude that the plant teabag culture medium by itself, without any nutritional supplements, is sufficient and efficient for recovering and mirroring the complex and diverse communities of rhizobacteria. Our message to fellow microbial ecologists is: simply dehydrate your plant canopy, teabag it and soak it to prepare your culture media, with no need for any additional supplementary nutrients.

Bacterial community dynamics during polysaccharide degradation at contrasting sites in the Southern and Atlantic Oceans.

The bacterial degradation of polysaccharides is central to marine carbon cycling, but little is known about the bacterial taxa that degrade specific marine polysaccharides. Here, bacterial growth and community dynamics were studied during the degradation of the polysaccharides chitin, alginate and agarose in microcosm experiments at four contrasting locations in the Southern and Atlantic Oceans. At the Southern polar front, chitin-supplemented microcosms were characterized by higher fractions of actively growing cells and a community shift from Alphaproteobacteria to Gammaproteobacteria and Bacteroidetes. At the Antarctic ice shelf, chitin degradation was associated with growth of Bacteroidetes, with 24% higher cell numbers compared with the control. At the Patagonian continental shelf, alginate and agarose degradation covaried with growth of different Alteromonadaceae populations, each with specific temporal growth patterns. At the Mauritanian upwelling, only the alginate hydrolysis product guluronate was consumed, coincident with increasing abundances of Alteromonadaceae and possibly cross-feeding SAR11. 16S rRNA gene amplicon libraries indicated that growth of the Bacteroidetes-affiliated genus Reichenbachiella was stimulated by chitin at all cold and temperate water stations, suggesting comparable ecological roles over wide geographical scales. Overall, the predominance of location-specific patterns showed that bacterial communities from contrasting oceanic biomes have members with different potentials to hydrolyse polysaccharides.

Changes in bacterial community metabolism and composition during the degradation of dissolved organic matter from the jellyfish Aurelia aurita in a Mediterranean coastal lagoon.

Spatial increases and temporal shifts in outbreaks of gelatinous plankton have been observed over the past several decades in many estuarine and coastal ecosystems. The effects of these blooms on marine ecosystem functioning and particularly on the dynamics of the heterotrophic bacteria are still unclear. The response of the bacterial community from a Mediterranean coastal lagoon to the addition of dissolved organic matter (DOM) from the jellyfish Aurelia aurita, corresponding to an enrichment of dissolved organic carbon (DOC) by 1.4, was assessed for 22 days in microcosms (8 l). The high bioavailability of this material led to (i) a rapid mineralization of the DOC and dissolved organic nitrogen from the jellyfish and (ii) the accumulation of high concentrations of ammonium and orthophosphate in the water column. DOM from jellyfish greatly stimulated heterotrophic prokaryotic production and respiration rates during the first 2 days; then, these activities showed a continuous decay until reaching those measured in the control microcosms (lagoon water only) at the end of the experiment. Bacterial growth efficiency remained below 20%, indicating that most of the DOM was respired and a minor part was channeled to biomass production. Changes in bacterial diversity were assessed by tag pyrosequencing of partial bacterial 16S rRNA genes, DNA fingerprints, and a cultivation approach. While bacterial diversity in control microcosms showed little changes during the experiment, the addition of DOM from the jellyfish induced a rapid growth of Pseudoalteromonas and Vibrio species that were isolated. After 9 days, the bacterial community was dominated by Bacteroidetes, which appeared more adapted to metabolize high-molecular-weight DOM. At the end of the experiment, the bacterial community shifted toward a higher proportion of Alphaproteobacteria. Resilience of the bacterial community after the addition of DOM from the jellyfish was higher for metabolic functions than diversity, suggesting that jellyfish blooms can induce durable changes in the bacterial community structure in coastal lagoons.

Jellyfish modulate bacterial dynamic and community structure.

Jellyfish blooms have increased in coastal areas around the world and the outbreaks have become longer and more frequent over the past few decades. The Mediterranean Sea is among the heavily affected regions and the common bloom-forming taxa are scyphozoans Aurelia aurita s.l., Pelagia noctiluca, and Rhizostoma pulmo. Jellyfish have few natural predators, therefore their carcasses at the termination of a bloom represent an organic-rich substrate that supports rapid bacterial growth, and may have a large impact on the surrounding environment. The focus of this study was to explore whether jellyfish substrate have an impact on bacterial community phylotype selection. We conducted in situ jellyfish-enrichment experiment with three different jellyfish species. Bacterial dynamic together with nutrients were monitored to assess decaying jellyfish-bacteria dynamics. Our results show that jellyfish biomass is characterized by protein rich organic matter, which is highly bioavailable to 'jellyfish-associated' and 'free-living' bacteria, and triggers rapid shifts in bacterial population dynamics and composition. Based on 16S rRNA clone libraries and denaturing gradient gel electrophoresis (DGGE) analysis, we observed a rapid shift in community composition from unculturable Alphaproteobacteria to culturable species of Gammaproteobacteria and Flavobacteria. The results of sequence analyses of bacterial isolates and of total bacterial community determined by culture independent genetic analysis showed the dominance of the Pseudoalteromonadaceae and the Vibrionaceae families. Elevated levels of dissolved proteins, dissolved organic and inorganic nutrient release, bacterial abundance and carbon production as well as ammonium concentrations characterized the degradation process. The biochemical composition of jellyfish species may influence changes in the amount of accumulated dissolved organic and inorganic nutrients. Our results can contribute insights into possible changes in bacterial population dynamics and nutrient pathways following jellyfish blooms which have important implications for ecology of coastal waters.

Growth stimulation of iodide-oxidizing α-Proteobacteria in iodide-rich environments.

α-Proteobacteria that can oxidize iodide (I(-)) to molecular iodine (I(2)) have only been isolated from iodide-rich natural and artificial environments, i.e., natural gas brine waters and seawaters supplemented with iodide, respectively. To understand the growth characteristics of such iodide-oxidizing bacteria (IOB) under iodide-rich environments, microcosms comprising natural seawater and 1 mM iodide were prepared, and the succession of microbial communities was monitored by culture-independent techniques. PCR-denaturing gradient gel electrophoresis and 16S rRNA gene sequence analysis showed that bacteria closely related with known IOB were predominant in the microcosms after several weeks of incubation. Quantitative PCR analysis targeting specific 16S rRNA gene regions of IOB showed that the relative abundance of IOB in the microcosms was 6-76% of the total bacterial population, whereas that in natural seawater was less than 1%. When 10(3) cells mL(-1) of IOB were inoculated into natural seawater supplemented with 0.1-1 mM iodide, significant growth (cell densities, 10(5)-10(6) cells mL(-1)) and I(2) production (6-32 µM) were observed. Interestingly, similar growth stimulation occurred when 12-44 µM of I(2) was added to seawater, instead of iodide. IOB were found to be more I(2) tolerant than the other heterotrophic bacteria in seawater. These results suggest that I(2) plays a key role in the growth stimulation of IOB in seawater. IOB could potentially attack other bacteria with I(2) to occupy their ecological niche in iodide-rich environments.

High contribution of SAR11 to microbial activity in the north west Mediterranean Sea.

We investigated the abundance and activity of SAR11 on a monthly time scale between January 2008 and October 2008 in the oligotrophic NW Mediterranean Sea. Applying MICRO-CARD-FISH, we observed that SAR11 had a large contribution to bulk abundance (37 ± 6% of DAPI-stained cells) and to bulk bacterial heterotrophic production (BHP), as estimated from leucine incorporation (55 ± 15% of DAPI-cells assimilating leucine) in surface waters (5 m) throughout the study period. SAR11 contributed also substantially to the assimilation of glucose, ATP, and a combination of amino acids (44 ± 17%, 37 ± 14%, and 43 ± 12% of DAPI cells assimilating these compounds, respectively), organic compounds that provide either single or combined sources of C, P, and N. Temporal changes in the abundance of SAR11 cells that assimilated leucine, glucose, amino acids, and ATP revealed a pattern consistent with that of substrate-active DAPI cells, suggesting that the activity of SAR11 can explain to a large extent the variability in total cells contributing to the utilization of these compounds. Short-term nutrient enrichment experiments performed on each sampling date revealed a strong co-limitation of at least two of the three elements analyzed (C, N, P), in particular, during summer and early autumn. The in situ abundance of SAR11 cells assimilating leucine appeared to increase with P limitation as determined in the nutrient enrichment experiments (r = 0.81, p = 0.015). Our results demonstrate that SAR11 is an important component of the active bacterial community in the NW Mediterranean Sea. Our observations further indicate that the activity of the bulk bacterial community is linked to the activity of SAR11, possibly due to its adaptation to nutrient limitation.

Breoghania corrubedonensis gen. nov. sp. nov., a novel alphaproteobacterium isolated from a Galician beach (NW Spain) after the Prestige fuel oil spill, and emended description of the family Cohaesibacteraceae and the species Cohaesibacter gelatinilyticus.

A Gram-negative bacterium designated UBF-P1(T) was isolated from an enrichment culture established in nutrient supplemented artificial sea water with pyrene as a carbon source, and inoculated with a marine fuel oil-degrading consortium obtained from a sand sample collected from the beach of Corrubedo (A Coruña, Galicia, Spain) after the Prestige accidental oil spill. Phylogenetic analysis based on the almost complete 16S rRNA gene sequence affiliated strain UBF-P1(T) with the family Cohaesibacteraceae, Cohaesibacter gelatinilyticus (DSM 18289(T)) being the closest relative species with 92% sequence similarity. Cells were irregular rods, motile, strictly aerobic, catalase and oxidase positive. Ubiquinone 10 was the major respiratory lipoquinone. The major polar lipids comprised diphosphatidylglycerol (DPG), phosphatidylglycerol (PG), phosphatidylethanolamine (PE), phosphatidylmonomethylethanolamine (PME), and phosphatidylcholine (PC). The major fatty acids detected were C(18:1)ω7c, C(19:0) cycloω8c, and C(16:0). The G+C content of strain UBF-P1(T) was 63.9 mol%. The taxonomic comparison with the closest relative based on genotypic, phenotypic and chemotaxonomic characteristics supported that strain UBF-P1(T) could be classified as a novel genus and species, for which the name Breoghania corrubedonensis gen. nov., sp. nov. is proposed. The type strain of this new taxon is UBF-P1(T) (CECT 7622, LMG 25482, DSM 23382).

The potential of Cycloclasticus and Altererythrobacter strains for use in bioremediation of petroleum-aromatic-contaminated tropical marine environments.

Cycloclasticus sp. A5, which has been suggested to be a major degrader of petroleum aromatics spilled in temperate seas, showed higher degrading activities for petroleum aromatics, at both 25 degrees C and tropical sea temperature 30 degrees C, than the novel aromatic-degrading isolates, related to Altererythrobacter epoxidivorans (97.5% similarity in the almost full-length 16S rRNA gene sequence) and Rhodovulum iodosum (96.3% similarity), obtained after enrichment on crude oil in a continuous supply of Indonesian seawater. Cycloclasticus A5 degraded petroleum aromatics at a similar rate or faster at 30 degrees C as compared to 25 degrees C, but its growth on acetate was severely inhibited at 30 degrees C. These results suggest that, although their abundance would be low in tropical seas not contaminated with aromatics, the Cycloclasticus strains could be major degraders of petroleum aromatics spilled in tropical seas. The 16S rRNA gene of the Cycloclasticus strains has been identified from Indonesian seawater, and the gene fragments showed 96.7-96.8% similarities to that of Cycloclasticus A5. Introducing Cycloclasticus A5 may be an ecologically advantageous bioremediation strategy for petroleum-aromatic-contaminated tropical seas because strain A5 would disappear at 30 degrees C after complete consumption of the aromatics. Altererythrobacter and Rhodovulum-related isolates grew well on pyruvate in 10% strength marine broth at 30 degrees C whereas Cycloclasticus A5 did not grow well on acetate in the broth at 30 degrees C. These growth results, along with its petroleum-aromatic-degrading activity, suggest that the Altererythrobacter isolate could be an important petroleum-aromatic degrader in and around nutrient-rich tropical marine environments.

Modelling the population dynamics and metabolic diversity of organisms relevant in anaerobic/anoxic/aerobic enhanced biological phosphorus removal processes.

In this study, enhanced biological phosphorus removal (EBPR) metabolic models are expanded in order to incorporate the competition between polyphosphate accumulating organisms (PAOs) and glycogen accumulating organisms (GAOs) under sequential anaerobic/anoxic/aerobic conditions, which are representative of most full-scale EBPR plants. Since PAOs and GAOs display different denitrification tendencies, which is dependent on the phylogenetic identity of the organism, the model was separated into six distinct biomass groups, constituting Accumulibacter Types I and II, as well as denitrifying and non-denitrifying Competibacter and Defluviicoccus GAOs. Denitrification was modelled as a multi-step process, with nitrate (NO(3)), nitrite (NO(2)), nitrous oxide (N(2)O) and di-nitrogen gas (N(2)) being the primary components. The model was calibrated and validated using literature data from enriched cultures of PAOs and GAOs, obtaining a good description of the observed biochemical transformations. A strong correlation was observed between Accumulibacter Types I and II, and nitrate-reducing and non-nitrate-reducing PAOs, respectively, where the abundance of each PAO subgroup was well predicted by the model during an acclimatization period from anaerobic-aerobic to anaerobic-anoxic conditions. Interestingly, a strong interdependency was observed between the anaerobic, anoxic and aerobic kinetic parameters of PAOs and GAOs. This could be exploited when metabolic models are calibrated, since all of these parameters should be changed by an identical factor from their default value. Factors that influence these kinetic parameters include the fraction of active biomass, relative aerobic/anoxic fraction and the ratio of acetyl-CoA to propionyl-CoA. Employing a metabolic approach was found to be advantageous in describing the performance and population dynamics in such complex microbial ecosystems.

[Metabolism characteristics of Chelativorans oligotrophicus by two-phase growth on the mixture of EDTA and glucose].

The obligate destructor of ethylene diamine tetraacetate--a culture of Chelativorans oligotrophicus LPM-4--did not grow on a medium with glucose, but it was good to use it under cultivation on a mixture with EDTA after considerable decrease of the EDTA concentration in the medium (two-phase growth). Strong inhibition of hexokinase and glucose 6-phosphate dehydrogenase in cell exracts 4 mM EDTA was revealed. Using EDTA, cells accumulated polyphosphates whose rate decreased during glucose utilization phase. High activities of polyphosphate biosynthesis ferments (adenylat kinase and polyphosphate kinase) were distinguished during the first phase of the cultivation; considerable decrease of them and increase of polyphosphate glucokinase were found during the second phase of the cultivation. This points to the possible participating of polyphosphates in glucose metabolism as a supplementary energy source.

[Ultrastructural organization and development cycle of soil ultramicrobacteria belonging to the class Alphaproteobacteria].

Gram-negative chemoorganotrophic soil ultramicrobacteria (UMB), strains NF1 and NF3, have been isolated. In their development cycle, the strains formed small coccoid cells of 400-800 nm and ultrasmall cells of 200-300 nm. Phylogenetically, the strains NF1 and NF3 belong to Alphaproteobacteria and are close to the type strain of the recently described species Kaistia adipata. The ultrastructure of UMB cells has been studied using ultrathin sections and freeze-fracturing. It has been shown that the structure of UMB cell walls is of the gram-negative type; the outer membrane and peptidoglycan layer are well differentiated. The cell surface has numerous protrusions (prosthecae) of conical or spherical shape filled with the contents of the periplasm. The formation of unusual cellular structures (not occurring in known free-living bacteria) is a feature of UMB: these include the following: (a) piles of rod-like subunits, ca. 30 A in diameter and 150-250 angstroms in length: (b) long bunches (up to 300-400 angstroms) comprised of filamentous subunits; and (c) large electron-dense spherical bodies (up to 200-300 angstroms in diameter) localized in the periplasm. A distinctive feature of UMB is their ability to grow as facultative parasites on living cyanobacterial (CB) cells. In this case, three types of interaction between UMB and CB have been revealed: (1) adsorption of UMB cells on the surface of CB cells; (2) penetration of UMB into polysaccharide sheathes; and (3) penetration of UMB into CB eytoplasm. UMB cells have been shown to reproduce by budding, with buds (up to 2-3) located directly on the mother cell, without formation of intennediate hyphae.

Are some putative glycogen accumulating organisms (GAO) in anaerobic : aerobic activated sludge systems members of the alpha-Proteobacteria?

Activated sludge plants designed to remove phosphorus microbiologically often perform unreliably. One suggestion is that the polyphosphate-accumulating organisms (PAO) are out-competed for substrates by another group of bacteria, the glycogen-accumulating organisms (GAO) in the anaerobic zones of these processes. This study used fluorescence in situ hybridization (FISH) and denaturing gradient gel electrophoresis (DGGE) to analyse the communities from laboratory-scale anaerobic : aerobic sequencing batch reactors. Members of the genus Sphingomonas in the alpha-Proteobacteria were present in large numbers in communities with poor phosphorus removal capacity where the biomass had a high glycogen content. Their ability to store poly-beta-hydroxyalkanoates anaerobically, but not aerobically, and not accumulate polyphosphate aerobically is consistent with these organisms behaving as GAO there. No evidence was found to support an important role for the gamma-Proteobacteria as possible GAO in these communities, although these bacterial populations have been considered in other studies to act as possible competitors for the PAO.

Crude oil-induced structural shift of coastal bacterial communities of rod bay (Terra Nova Bay, Ross Sea, Antarctica) and characterization of cultured cold-adapted hydrocarbonoclastic bacteria.

For preliminary screening of human impact on Antarctic coasts, the compositions of microbial communities were analyzed in seawater at two sites located in the Terra Nova Bay of Antarctica (Ross Sea) by a combination of 16S rRNA gene sequencing and culture techniques. The bacterial community in the sample from the Rod Bay site, located at the proximity to the Italian Station, was characterized by a high abundance of 16S rRNA gene sequences belonging to the microflora typically found in soil and freshwater environments. In contrast, the seawater sample from the Adelie Cove station, a pristine reference site, contained 16S rRNA gene sequences typically found in marine areas affected by algal blooms and sea ice decay. The addition of crude oil to the Rod Bay seawater sample rapidly induced a shift in the composition of the bacterial community with appearance of novel taxonomic groups and a dramatic increase in the relative abundance of gamma-Proteobacteria sequences, whereas no significant changes were detected in the bacterial community of the Adelie Cove sample under the same conditions. Bacteria-exhibiting features with potential interest for industrial and environmental applications were isolated from the Rod Bay oil-enriched sample. In particular, hydrocarbon-degrading, cold-adapted bacteria were selectively enriched, isolated and screened for their ability to synthesize polyunsaturated fatty acids. Twenty two bacterial strains were isolated from the oil enrichment culture and identified. Eighteen isolates were found to be members of gamma-Proteobacteria, while the remainder were representatives of alpha-Proteobacteria, CFB and high G + C divisions.