Effects of inorganic ions on autotrophic denitrification by Thiobacillus denitrificans and on heterotrophic denitrification by an enrichment culture.

Salinity of nitrate-laden wastewaters, such as those produced by metal industries, tanneries, and wet flue gas cleaning systems may affect their treatment by denitrification. Salt inhibition of denitrification has been reported, while impacts of individual ions remain poorly understood whilst being relevant for wastewaters where often the concentration of a single ion rather than the salts varies. The aim of this study was to determine the inhibition by inorganic ions (Na+, Cl-, SO42- and K+) commonly present in saline wastewaters on denitrification and reveal its potential for the treatment of such waste streams, like those produced by NOx-SOx removal scrubbers. The inhibitory effects were investigated for both heterotrophic (enrichment culture) and autotrophic (T. denitrificans) denitrification in batch assays, by using NaCl, Na2SO4, KCl and K2SO4 salts at increasing concentrations. The half inhibition concentrations (IC50) of Na+ (as NaCl), Na+ (as Na2SO4) and Cl- (as KCl) were: 4.3 ± 0.3, 7.9 ± 0.5 and 5.2 ± 0.3 g/L for heterotrophic, and 1-2.5, 2.5-5 and 4.1 ± 0.3 g/L for autotrophic denitrification, respectively. Heterotrophic denitrification was completely inhibited at 20 g/L Na+ (as NaCl), 30 g/L Na+ (as Na2SO4) and 30 g/L Cl- (as KCl), while autotrophic at 8 g/L Na+ (as NaCl), 10 g/L Na+ (as Na2SO4) and 15 g/L Cl- (as KCl). In both cases, Cl- addition had the most important role in decreasing denitrification rate, while Na+ at 1 g/L stimulated autotrophic denitrification but rapidly inhibited the rate at higher concentrations. Nitrite reduction was less inhibited by the ions than nitrate reduction and both the osmotic pressure and the toxicity of the single ions played key roles in the overall inhibition of denitrification. Eventually, both autotrophic and heterotrophic denitrification showed potential for the treatment of a saline wastewater from a NOx-SO2 removal scrubber from a pulp mill.

Removal of arsenic from acidic liquors using chemical and autotrophic and mixed heterotrophic bacteria-produced biogenic schwertmannites.

Arsenic penetrates human society through a variety of geological and anthropogenic processes, posing significant health hazards. Acid mine drainage, which contains high concentrations of heavy metals and sulfate, is formed by the biological oxidation of pyrite and other metal-containing sulfidic minerals and is a significant environmental hazard. Adsorption is a simple and effective method for removing arsenic from water. In this study, co-precipitation and adsorption of arsenic with biogenic and chemically produced iron-containing settleable precipitates, i.e. schwertmannites were studied. Autotrophic Leptospirillum ferrooxidans and heterotrophic mixed culture of Alicyclobacillus tolerans and Acidiphilium cryptum oxidized iron at rates from 18 to 23 mg/(L.h) in the presence of 5 and 10 mg/L As3+, and both cultures tolerated up to 100 mg As3+/L although Fe2+ oxidation rates decreased to 3-4 mg/(L.h). At Fe/As ratios of ≥20, As removal efficiencies of ≥95% were obtained by co-precipitation with Fe3+ at pH 3.5-4.5. Because schwertmannite precipitates produced by the heterotrophic culture formed crystals, it was studied for adsorptive removals of As3+ and As5+ and compared with chemically synthesized schwertmannites. As3+ (100 mg/L) adsorption onto biogenic and chemical schwertmannite were 25 and 44%, respectively, at pH 4. At 100 mg As5+/L, adsorption capacity and efficiency onto biogenic schwertmannite were 47 mg/g and 50%, respectively. At 300 mg As5+/L, adsorption capacity and efficiency onto chemical schwertmannite were 169 mg/g and 56%, respectively. In summary, biogenic schwertmannite has potential for As removal via co-precipitation with Fe3+ at pH 3.5-4.5 and Fe/As ratios of ≥20 due to low production cost from acidic mine drainage. In contrast to the schwertmannite generation methods, which are usually performed with autotrophic acidophilic bacteria in the literature, this efficient and modular schwertmannite production process and its evaluation on arsenic adsorption is an important potential in acidic mine drainage treatment containing arsenic.

Potential of biological sulphur recovery from thiosulphate by haloalkaliphilic Thioalkalivibrio denitrificans.

The aim of this study was to investigate the potential for elemental sulphur recovery from sulphurous solutions under aerobic and anoxic conditions by haloalkalophilic Thioalkalivibrio denitrificans at 0.8-19.6 g S2O32--S L-1 and 0.2-0.58 g NO2 L-1, respectively. The experiments were conducted as batch assays with haloalkaline (pH 10 and ≥ 14 g Na+ L-1) thiosulphate solution. Aerobically, the highest biotransformation rate of thiosulphate obtained was 0.03 h-1 at 8.5 g L S2O32--S. Based on Monod model, the maximum substrate utilisation rate (qm) was 0.024 h-1 with half saturation constant (Ks) 0.42 g S2O32--S L-1 at initial [S2O32--S] of 14 g L-1. S0 accumulated at [S2O32--S] ≥ 1.5 g L-1 (10% yield at initial 9.5 g S2O32--S L-1) and the highest S0 yield estimated with the model was 61% with initial [S2O32--S] of 16.5 g L-1. Anoxically, the maximum nitrite removal rate based on Monod modelling was 0.011 h-1 with Ks = 0.84 g NO2- L-1. Aerobically and anoxically the maximum specific growth rates (µm) were 0.046 and 0.022 h-1, respectively. In summary, high-rate aerobic biotransformation kinetics of thiosulphate were demonstrated, whereas the rates were slower and no S0 accumulated under anoxic conditions. Thus, future developments of biotechnical applications for the recovery of S0 from haloalkaline streams from the process industry should focus on aerobic treatment.HighlightsHaloalkaline S2O32- biotransformations kinetics by Thioalkalivibrio denitrificansAerobic thiosulphate-S bioconversion up to 0.024 h-1 with Ks = 0.42 g S2O32--S L-110% S0 yield with initial 9.5 g S2O32--S L-1 in aerobic conditionAnoxic NO2 removal up to 0.01 h-1 with Ks = 0.84 g NO2- L-1.

Simulated acid mine drainage treatment in iron oxidizing ceramic membrane bioreactor with subsequent co-precipitation of iron and arsenic.

Acid mine drainage (AMD), generated in the active and abandoned mine sites, is characterized by low pH and high metal concentrations. One AMD treatment possibility is biologically oxidizing Fe2+ followed by precipitation through pH control. As compared to autotrophic iron oxidizing microbial community, a microbial community enriched in the presence of organic nutrients was hypothesized to yield higher biomass during commissioning the bioreactor. In this study, the treatment of Fe, Cu, Co, Mn, Zn, Ni, and As containing simulated AMD was studied using an iron-oxidizing ceramic membrane bioreactor (CMBR) at varying hydraulic retention times (HRTs) (6-24 h) and two different feed Fe2+ concentrations (250 and 750 mg/L). The impact of tryptone soya broth (TSB) on the CMBR performance was also investigated. Almost complete Fe2+ oxidation and sustainable flux at around 5.0 L/(m2.h) were obtained in the CMBR with the Alicyclobacillus tolerans and Acidiphilium cryptum dominated enrichment culture. The Fe2+ oxidation rate, as assessed in batch operation cycles of CMBR, increased significantly with increasing Fe2+ loading to the bioreactor. The iron oxidation rate decreased by the elimination of organic matter from the feed. The increase of the CMBR permeate pH to 3.5-4.0 resulted in selective co-precipitation of As and Fe (over 99%) with the generation of biogenic schwertmannite.

Increase in sedimentary organic carbon with a change from hypoxic to oxic conditions.

In the Seto Inland Sea, Japan, chemical oxygen demand has increased over recent decades, while average dissolved oxygen concentrations in the bottom water have increased. In this study, we investigated responses of organic carbon (OC) in hypoxic sediment to changes of redox conditions using experimental columns containing sediment and overlying water. Surface sediment showed an increase in OC along with the change to an aerobic condition. Microbial community analysis showed a predominance of sulfur-oxidizing bacteria (SOB) such as Sulfurovum sp. in the sediment. This dominance could account for the increased OC. Additionally, the dissolved organic carbon (DOC) concentration in the overlying water increased. Further experiments using sandy sediment showed that biodegradation of Sulfurimonas denitrificans was associated with DOC release. These results show that a change in the sedimentary environment (increase in dissolved oxygen) increased the sedimentary OC and DOC of overlying water by stimulating certain autotrophic bacteria, especially the SOB.

Leaching of rare earth elements and base metals from spent NiMH batteries using gluconate and its potential bio-oxidation products.

Gluconate is known to mediate metal leaching. However, during bioleaching by e.g., Gluconobacter oxydans, gluconate can be oxidized to 2-ketogluconate and 5-ketogluconate. The impact of bio-oxidation of gluconate on metal leaching has not been investigated. Therefore, the aim of this study was to investigate leaching of rare earth elements (REEs) and base metals from spent nickel-metal-hydride (NiMH) batteries using gluconate, 2-ketogluconate and 5-ketogluconate. Batch leaching assays were conducted under controlled and uncontrolled pH conditions for 14 days using 60 mM of either the individual leaching agents or their various combinations. At target pH of 6.0 ± 0.1 and 9.0 ± 0.1 and without pH control, complexolysis was the dominating leaching mechanism and higher REE leaching efficiency was obtained with gluconate, while 5-ketogluconate enabled more efficient base metal leaching. At target pH of 3.0 ± 0.1, acidolysis dominated, and the base metal and REE leaching yields with all the tested leaching agents were higher than under the other studied pH conditions. The highest base metal and REE leaching yields (%) were obtained using gluconate at target pH of 3.0 ± 0.1 being 100.0 Mn, 90.3 Fe, 89.5 Co, 58.5 Ni, 24.0 Cu, 29.3 Zn and 56.1 total REEs. The obtained results are useful in optimization of heterotrophic bioleaching.

Formation and use of biogenic jarosite carrier for high-rate iron oxidising biofilms.

Jarosite precipitates formed in iron oxidising bioreactors have been shown to harbour iron-oxidisers. The aim of this study was to develop an iron oxidising bioprocess where microorganisms are retained solely on biogenic jarosite particles. Based on preliminary experiments using a fluidised-bed bioreactor (FBR), the formed jarosite particles started to disintegrate and wash out at upflow velocities of ≥0.21 cm/s. Therefore, the generation and use of biogenic jarosite carrier was studied in an expanded-bed bioreactor (J-EBR) with an upflow velocity of 0.19 cm/s. Inside J-EBR, the jarosite particles formed granules of 0.5-3 mm containing 200-460 mg/g of attached biomass. The performance of J-EBR was compared with an activated carbon biofilm FBR at 0.82 cm/s upflow velocity (AC-FBR). At 35 ± 2 °C with a feed ferrous iron concentration of 10 g/l, the highest obtained iron oxidation rate of J-EBR (6.8 g/l/h) was 33% lower than that of AC-FBR (10.1 g/l/h). This was likely due to the 80% lower recirculation rate and subsequently higher oxygen mass transfer limitation in J-EBR compared to AC-FBR. The present study demonstrates that biogenic jarosite can be used for retainment of iron oxidising biofilms in expanded-bed bioreactors that oxidise iron at high rates.

The effect of start-up on energy recovery and compositional changes in brewery wastewater in bioelectrochemical systems.

Start-up of bioelectrochemical systems (BESs) fed with brewery wastewater was compared at different adjusted anode potentials (-200 and 0 mV vs. Ag/AgCl) and external resistances (50 and 1000 Ω). Current generation stabilized faster with the external resistances (9 ± 3 and 1.70 ± 0.04 A/m3 with 50 and 1000 Ω, respectively), whilst significantly higher current densities of 76 ± 39 and 44 ± 9 A/m3 were obtained with the adjusted anode potentials of -200 and 0 mV vs. Ag/AgCl, respectively. After start-up, when operated using 47 Ω external resistance, the current densities and Coulombic efficiencies of all BESs stabilized to 9.5 ± 2.9 A/m3 and 12 ± 2%, respectively, demonstrating that the start-up protocols were not critical for long-term BES operation in microbial fuel cell mode. With adjusted anode potentials, two times more biofilm biomass (measured as protein) was formed by the end of the experiment as compared to start-up with the fixed external resistances. After start-up, the organics in the brewery wastewater, mainly sugars and alcohols, were transformed to acetate (1360 ± 250 mg/L) and propionate (610 ± 190 mg/L). Optimized start-up is required for prompt BES recovery, for example, after process disturbances. Based on the results of this study, adjustment of anode potential to -200 mV vs. Ag/AgCl is recommended for fast BES start-up.

Storing of exoelectrogenic anolyte for efficient microbial fuel cell recovery.

Starting up a microbial fuel cell (MFC) requires often a long-term culture enrichment period, which is a challenge after process upsets. The purpose of this study was to develop low-cost storage for MFC enrichment culture to enable prompt process recovery after upsets. Anolyte of an operating xylose-fed MFC was stored at different temperatures and for different time periods. Storing the anolyte for 1 week or 1 month at +4°C did not significantly affect power production, but the lag time for power production was increased from 2 days to 3 or 5 days, respectively. One month storing at -20°C increased the lag time to 7 days. The average power density in these MFCs varied between 1.2 and 1.7 W/m3. The share of dead cells (measured by live/dead staining) increased with storing time. After 6-month storage, the power production was insignificant. However, xylose removal remained similar in all cultures (99-100%) while volatile fatty acids production varied. The results indicate that fermentative organisms tolerated the long storage better than the exoelectrogens. As storing at +4°C is less energy intensive compared to freezing, anolyte storage at +4°C for a maximum of 1 month is recommended as start-up seed for MFC after process failure to enable efficient process recovery.

Fluidized bed bioreactor for multiple environmental engineering solutions.

Fluidized bed bioreactors (FBR) are characterized by two-phase mixture of fluid and solid, in which the bed of solid particles is fluidized by means of downward or upward recirculation stream. FBRs are widely used for multiple environmental engineering solutions, such as wastewater treatment, as well as some industrial applications. FBR offers many benefits such as compact bioreactor size due to short hydraulic retention time, long biomass retention on the carrier, high conversion rates due to fully mixed conditions and consequently high mass transfer rates, no channelling of flow, dilution of influent concentrations due to recycle flow, suitability for enrichment of microbes with low Km values. The disadvantages of FBRs include bioreactor size limitations due to the height-to-diameter ratio, high-energy requirements due to high recycle ratios, and long start-up period for biofilm formation. This paper critically reviews some of the key studies on biomass enrichment via immobilisation of low growth yield microorganisms, high-rates via fully mixed conditions, technical developments in FBRs and ways of overcoming toxic effects via solution recycling. This technology has many potential new uses as well as hydrodynamic characteristics, which enable high-rate environmental engineering and industrial applications.

The effect of anode potential on bioelectrochemical and electrochemical tetrathionate degradation.

The effect of poised anode potential on electricity production and tetrathionate degradation was studied in two-chamber flow-through electrochemical (ES) and bioelectrochemical systems (BES). The minimum anode potential (vs. Ag/AgCl) for positive current generation was 0.3V in BES and 0.5V in the abiotic ES. The anode potential required to obtain average current density above 70mAm-2 was 0.4V in BES and above 0.7V in ES. ES provided higher coulombic efficiency, but the average tetrathionate degradation rate remained significantly higher in BES (above 110mgL-1d-1) than in the abiotic ES (below 35mgL-1d-1). This study shows that at anode potentials below 0.7V, the electrochemical tetrathionate degradation is only efficient with microbial catalyst and that significantly higher tetrathionate degradation rates can be obtained with bioelectrochemical systems than with electrochemical systems at the tested anode potentials.

Long-term stability of bioelectricity generation coupled with tetrathionate disproportionation.

To prevent uncontrolled acidification of the environment, reduced inorganic sulfur compounds (RISCs) can be bioelectrochemically removed from water streams. The long-term stability of bioelectricity production from tetrathionate (S4O6(2-)) was studied in highly acidic conditions (pH<2.5) in two-chamber fed-batch microbial fuel cells (MFCs). The maximum current density was improved from previously reported 80mAm(-2) to 225mAm(-2) by optimizing the external resistance. The observed reaction products of tetrathionate disproportionation were sulfate and elemental sulfur. In long-term run, stable electricity production was obtained for over 700days with the average current density of 150mAm(-2). The internal resistance of the MFC decreased over time and no biofouling was observed. This study shows that tetrathionate is an efficient substrate also for long-term bioelectricity production.

Anaerobes in Bioelectrochemical Systems.

In bioelectrochemical systems (BES), the catalytic activity of anaerobic microorganisms generates electrons at the anode which can be used, for example, for the production of electricity or chemical compounds. BES can be used for various purposes, including wastewater treatment, production of electricity, fuels and chemicals, biosensors, bioremediation, and desalination. Electrochemically active microorganisms are widely present in the environment and they can be found, in sediment, soil, compost, wastewaters and their treatment plants. Exoelectrogens are microorganisms capable of donating electrons to anode electrode or accepting electrons from cathode electrode and are mainly responsible for current generation or use in BES. However, current generation from fermentable substrates often requires the presence of electrochemically inactive microorganisms that break down complex substrates into metabolites which can be further utilized by exoelectrogens. The growth and electron transfer efficiency of anaerobes depend on several parameters, such as system architecture, electrode material and porosity, electrode potential and external resistance, pH, temperature, substrate concentration, organic loading rate, and ionic strength. In this chapter, the principles and microbiology of bioelectrochemical systems as well as selective factors for exoelectrogens are reviewed. The anaerobic microorganisms and their electron transfer mechanisms at the anode and cathode are described and future aspects are briefly discussed.

Arsenic removal from acidic solutions with biogenic ferric precipitates.

Treatment of acidic solution containing 5g/L of Fe(II) and 10mg/L of As(III) was studied in a system consisting of a biological fluidized-bed reactor (FBR) for iron oxidation, and a gravity settler for iron precipitation and separation of the ferric precipitates. At pH 3.0 and FBR retention time of 5.7h, 96-98% of the added Fe(II) precipitated (99.1% of which was jarosite). The highest iron oxidation and precipitation rates were 1070 and 28mg/L/h, respectively, and were achieved at pH 3.0. Subsequently, the effect of pH on arsenic removal through sorption and/or co-precipitation was examined by gradually decreasing solution pH from 3.0 to 1.6 (feed pH). At pH 3.0, 2.4 and 1.6, the highest arsenic removal efficiencies obtained were 99.5%, 80.1% and 7.1%, respectively. As the system had ferric precipitates in excess, decreased arsenic removal was likely due to reduced co-precipitation at pH<2.4. As(III) was partially oxidized to As(V) in the system. In shake flask experiments, As(V) sorbed onto jarosite better than As(III). Moreover, the sorption capacity of biogenic jarosite was significantly higher than that of synthetic jarosite. The developed bioprocess simultaneously and efficiently removes iron and arsenic from acidic solutions, indicating potential for mining wastewater treatment.

Selecting an indigenous microalgal strain for lipid production in anaerobically treated piggery wastewater.

The aim of this study was to select a potential microalgal strain for lipid production and to examine the suitability of anaerobically treated piggery wastewater as a nutrient source for production of lipid-rich biomass with the selected microalga. Biomass and lipid productivity of three microalgal strains (Chlorella sorokiniana CY1, Chlorella vulgaris CY5 and Chlamydomonas sp. JSC-04) were compared by using different media, nitrogen sources, and nitrogen concentrations. The highest lipid content and productivity (62.5 wt%, 162 mg/L/d) were obtained with C. vulgaris with BG-11 with 62 mg N/L. Secondly, C. vulgaris was cultivated in sterilized, diluted (1-20×), anaerobically treated piggery wastewater. Biomass production decreased and lipid content increased, when wastewater was more diluted. The highest lipid content of 54.7 wt% was obtained with 20× dilution, while the highest lipid productivity of 100.7 mg/L/d with 5× dilution. Piggery wastewater is a promising resource for mass production of oleaginous microalgal biomass.

Electricity generation from tetrathionate in microbial fuel cells by acidophiles.

Inorganic sulfur compounds, such as tetrathionate, are often present in mining process and waste waters. The biodegradation of tetrathionate was studied under acidic conditions in aerobic batch cultivations and in anaerobic anodes of two-chamber flow-through microbial fuel cells (MFCs). All four cultures originating from biohydrometallurgical process waters from multimetal ore heap bioleaching oxidized tetrathionate aerobically at pH below 3 with sulfate as the main soluble metabolite. In addition, all cultures generated electricity from tetrathionate in MFCs at pH below 2.5 with ferric iron as the terminal cathodic electron acceptor. The maximum current and power densities during MFC operation and in the performance analysis were 79.6 mA m(-2) and 13.9 mW m(-2) and 433 mA m(-2) and 17.6 mW m(-2), respectively. However, the low coulombic efficiency (below 5%) indicates that most of the electrons were directed to other processes, such as aerobic oxidation of tetrathionate and unmeasured intermediates. The microbial community analysis revealed that the dominant species both in the anolyte and on the anode electrode surface of the MFCs were Acidithiobacillus spp. and Ferroplasma spp. This study provides a proof of concept that tetrathionate serves as electron donor for biological electricity production in the pH range of 1.2-2.5.

Anaerobic conversion of microalgal biomass to sustainable energy carriers--a review.

This review discusses anaerobic production of methane, hydrogen, ethanol, butanol and electricity from microalgal biomass. The amenability of microalgal biomass to these bioenergy conversion processes is compared with other aquatic and terrestrial biomass sources. The highest energy yields (kJ g(-1) dry wt. microalgal biomass) reported in the literature have been 14.8 as ethanol, 14.4 as methane, 6.6 as butanol and 1.2 as hydrogen. The highest power density reported from microalgal biomass in microbial fuel cells has been 980 mW m(-2). Sequential production of different energy carriers increases attainable energy yields, but also increases investment and maintenance costs. Microalgal biomass is a promising feedstock for anaerobic energy conversion processes, especially for methanogenic digestion and ethanol fermentation. The reviewed studies have mainly been based on laboratory scale experiments and thus scale-up of anaerobic utilization of microalgal biomass for production of energy carriers is now timely and required for cost-effectiveness comparisons.

Dark fermentative hydrogen production from neutralized acid hydrolysates of conifer pulp.

Concentrated acid hydrolysis of cellulosic material results in high dissolution yields. In this study, the neutralization step of concentrated acid hydrolysate of conifer pulp was optimized. Dry conifer pulp hydrolysis with 55 % H(2)SO(4) at 45 °C for 2 h resulted in total sugar yields of 22.3-26.2 g/L. The neutralization step was optimized for solid Ca(OH)(2), liquid Ca(OH)(2) or solid CaO, mixing time, and water supplementation. The highest hydrogen yield of 1.75 mol H(2)/mol glucose was obtained with liquid Ca(OH)(2), while the use of solid Ca(OH)(2) or CaO inhibited hydrogen fermentation. Liquid Ca(OH)(2) removed sulfate to below 30 mg SO(4) (2-)/L. Further optimization of the neutralization conditions resulted in the yield of 2.26 mol H(2)/mol glucose.

Eukaryotic and prokaryotic microbial communities during microalgal biomass production.

Eukaryotic and bacterial communities were characterized and quantified in microalgal photobioreactor cultures of freshwater Chlorella vulgaris and marine Dunaliella tertiolecta. The microalgae exhibited good growth, whilst both cultures contained diverse bacterial communities. Both cultures included Proteobacteria and Bacteroidetes, while C. vulgaris cultures also contained Actinobacteria. The bacterial genera present in the cultures were different due to different growth medium salinities and possibly different extracellular products. Bacterial community profiles were relatively stable in D. tertiolecta cultures but not in C. vulgaris cultures likely due to presence of ciliates (Colpoda sp.) in the latter. The presence of ciliates did not, however, cause decrease in total number of C. vulgaris or bacteria during 14 days of cultivation. Quantitative PCR (qPCR) reliably showed relative microalgal and bacterial cell numbers in the batch cultures with stable microbial communities, but was not effective when bacterial communities varied. Raw culture samples were successfully used as qPCR templates.

Growth of Dunaliella tertiolecta and associated bacteria in photobioreactors.

The aim of this study was to test three flat-plate photobioreactor configurations for cultivation of marine green alga Dunaliella tertiolecta under non-axenic growth conditions and to characterize and quantify the associated bacteria. The photobioreactor cultivations were conducted using tap water-based media. Static mixers intended to enhance mixing and light utilization did not generally increase algal growth at the low light intensities used. The maximum biomass concentration (measured as volatile suspended solids) and maximum specific growth rate achieved in the flat plate with no mixer were 2.9 g l⁻¹ and 1.3 day⁻¹, respectively. Based on quantitative polymerase chain reaction, bacterial growth followed the growth of D. tertiolecta. Based on 16S rDNA amplification and denaturing gradient gel electrophoresis profiling, heterotrophic bacteria in the D. tertiolecta cultures mainly originated from the non-axenic algal inocula, and tap water heterotrophs were not enriched in high chloride media (3 % salinity). Bacterial communities were relatively stable and reproducible in all flat-plate cultivations and were dominated by Gammaproteobacteria, Flavobacteria, and Alphaproteobacteria.