Hunting for pigments in bacterial settlers of the Great Pacific Garbage Patch.

The Great Pacific Garbage Patch, a significant collection of plastic introduced by human activities, provides an ideal environment to study bacterial lifestyles on plastic substrates. We proposed that bacteria colonizing the floating plastic debris would develop strategies to deal with the ultraviolet-exposed substrate, such as the production of antioxidant pigments. We observed a variety of pigmentation in 67 strains that were directly cultivated from plastic pieces sampled from the Garbage Patch. The genomic analysis of four representative strains, each distinct in taxonomy, revealed multiple pathways for carotenoid production. These pathways include those that produce less common carotenoids and a cluster of photosynthetic genes. This cluster appears to originate from a potentially new species of the Rhodobacteraceae family. This represents the first report of an aerobic anoxygenic photoheterotrophic bacterium from plastic biofilms. Spectral analysis showed that the bacteria actively produce carotenoids, such as beta-carotene and beta-cryptoxanthin, and bacteriochlorophyll a. Furthermore, we discovered that the genetic ability to synthesize carotenoids is more common in plastic biofilms than in the surrounding water communities. Our findings suggest that plastic biofilms could be an overlooked source of bacteria-produced carotenoids, including rare forms. It also suggests that photoreactive molecules might play a crucial role in bacterial biofilm communities in surface water.

The potential of multispectral imaging flow cytometry for environmental monitoring.

Environmental monitoring involves the quantification of microscopic cells and particles such as algae, plant cells, pollen, or fungal spores. Traditional methods using conventional microscopy require expert knowledge, are time-intensive and not well-suited for automated high throughput. Multispectral imaging flow cytometry (MIFC) allows measurement of up to 5000 particles per second from a fluid suspension and can simultaneously capture up to 12 images of every single particle for brightfield and different spectral ranges, with up to 60x magnification. The high throughput of MIFC has high potential for increasing the amount and accuracy of environmental monitoring, such as for plant-pollinator interactions, fossil samples, air, water or food quality that currently rely on manual microscopic methods. Automated recognition of particles and cells is also possible, when MIFC is combined with deep-learning computational techniques. Furthermore, various fluorescence dyes can be used to stain specific parts of the cell to highlight physiological and chemical features including: vitality of pollen or algae, allergen content of individual pollen, surface chemical composition (carbohydrate coating) of cells, DNA- or enzyme-activity staining. Here, we outline the great potential for MIFC in environmental research for a variety of research fields and focal organisms. In addition, we provide best practice recommendations.

Estimating plastic pollution in rivers through harmonized monitoring strategies.

Plastics in rivers and lakes have direct local impact, and may also reach the world's oceans. Monitoring river plastic pollution is therefore key to quantify, understand and reduce plastics in all aquatic ecosystems. The lack of harmonization between ongoing monitoring efforts compromises the direct comparison and combination of available data. The United Nations Environment Programme (UNEP) launched guidelines on freshwater plastic monitoring, to provide a starting point for practitioners and scientists towards harmonized data collection, analysis, and reporting. We developed a five-step workflow to support to design effective plastic monitoring strategies. The workflow was applied to three rivers (Rhine, Mekong and Odaw) across relevant gradients, including geography, hydrology, and plastic pollution levels. We show that despite the simplicity of the selected methods and the limited duration of the data collection, our harmonized approach provides crucial insights in the state of plastic pollution in very different river basins globally.

High Potential for Anaerobic Microbial Sulfur Oxidation in Oil Sands Tailings Ponds.

The biogenic production of toxic H2S gas in sulfate-rich oil sands tailings ponds is associated with strong environmental concerns. Beside precipitation into sulfide minerals and chemical re-oxidation, microbial sulfur oxidation may catalyze sulfide re-cycling but potentially contributes to acid rock drainage (ARD) generation. To evaluate the microbial potential for sulfur oxidation, we conducted a microcosm-based pilot study with tailings of an active pond. Incubations were performed under oxic and anoxic conditions, with and without KNO3 as an electron acceptor and thiosulfate as a common substrate for microbial sulfur oxidation. The highest potentials of sulfur oxidation occurred in oxic assays (1.21 mmol L-1 day-1). Under anoxic conditions, rates were significantly lower and dominated by chemical transformation (0.09 mmol L-1 day-1; p < 0.0001). The addition of KNO3 to anoxic incubations increased microbial thiosulfate oxidation 2.5-fold (0.23 mmol L-1 day-1; p = 0.0474), with complete transformation to SO42- coupled to NO3- consumption, pointing to the activity of sulfur-oxidizing bacteria (SOB) under nitrate-reducing conditions. Importantly, in the presence of KNO3, a decrease in sedimentary sulfides was associated with an increase in S0, which indicates the potential for microbially mediated oxidation of sulfide minerals and ARD generation. Furthermore, the comparative analysis of sediments from other anthropogenic aquatic habitats demonstrated high similarities with respect to viable SOB counts and corresponding activity rates.

Burial of microplastics in freshwater sediments facilitated by iron-organo flocs.

Microplastics are ubiquitous in standing freshwater bodies, consequently lakes and reservoirs may be important sinks for these contaminants. However, the mechanisms governing the deposition of microplastics and their interactions with the sediments are understudied. We demonstrate how aggregation-based transport facilitates the sinking and infiltration of buoyant microplastics into freshwater reservoir sediments by employing experiments with intact sediment cores. Buoyant polyethylene microplastics were rapidly (1-4 h) incorporated into sinking iron-organic aggregates, followed by swift deposition into sediments. Ingression of microplastic bearing flocs into sediments was completed within 6 days and led to stable deposition of the incorporated particles for at least 2 months. Most microplastics were deposited in the top 2 cm of the sediments and few particles (5-15%) were re-released into the water. Our results show at least 85% burial of microplastics, indicating the significant role of freshwaters with low flow velocities in reducing microplastic loads to the oceans.

Interaction of cyanobacteria with calcium facilitates the sedimentation of microplastics in a eutrophic reservoir.

Low-density microplastics are frequently found in sediments of many lakes and reservoirs. The processes leading to sedimentation of initially buoyant polymers are poorly understood for inland waters. This study investigated the impact of biofilm formation and aggregation on the density of buoyant polyethylene microplastics. Biofilm formation on polyethylene films (4 × 4 × 0.15 mm) was studied in a eutrophic reservoir (Bautzen, Saxony, Germany). Additionally, aggregation dynamics of small PE microplastics (~85 µm) with cyanobacteria were investigated in laboratory experiments. During summer phototrophic sessile cyanobacteria (Chamaesiphon spp. and Leptolyngbya spp.) precipitated calcite while forming biofilms on microplastics incubated in Bautzen reservoir. Subsequently the density of the biofilms led to sinking of roughly 10% of the polyethylene particles within 29 days of incubation. In the laboratory experiments planktonic cyanobacteria (Microcystis spp.) formed large and dense cell aggregates under the influence of elevated Ca2+ concentrations. These aggregates enclosed microplastic particles and led to sinking of a small portion (~0.4 %) of polyethylene microplastics. This study showed that both sessile and planktonic phototrophic microorganisms mediate processes influenced by calcium which facilitates densification and sinking of microplastics in freshwater reservoirs. Loss of buoyancy leads to particle sedimentation and could be a prerequisite for the permanent burial of microplastics within reservoir sediments.

Adaptation of Coccomyxa sp. to Extremely Low Light Conditions Causes Deep Chlorophyll and Oxygen Maxima in Acidic Pit Lakes.

Deep chlorophyll maxima (DCM) and metalimnetic oxygen maxima (MOM) are outstanding biogeochemical features of acidic pit lakes (APL). However, knowledge of the eukaryotic phototrophs responsible for their formation is limited. We aimed at linking the dynamics of phototrophic communities inhabiting meromictic APL in Spain with the formation of these characteristic layers. Firstly, the dynamics of DCM and MOM and their relation to physico-chemical parameters (photosynthetically active radiation (PAR), pH, dissolved ferric iron concentration, temperature), pigments and nutrient distribution is described; secondly, the phototrophic community composition is studied through a combination of microscopy, biomolecular and "omics" tools. Phototrophic communities of the studied APL show a low diversity dominated by green microalgae, specifically Coccomyxa sp., which have been successfully adapted to the chemically harsh conditions. DCM and MOM are usually non-coincident. DCM correspond to layers where phototrophs have higher chlorophyll content per cell to cope with extremely low PAR (<1 µmol m-2 s-1), but where photosynthetic oxygen production is limited. MOM correspond to shallower waters with more light, higher phytoplankton biomass and intense photosynthetic activity, which affects both oxygen concentration and water temperature. The main drivers of DCM formation in these APL are likely the need for nutrient uptake and photo-acclimation.

Biofouling, metal sorption and aggregation are related to sinking of microplastics in a stratified reservoir.

Microplastic particles entering aquatic systems are rapidly colonized by microbial biofilms. The presence of microbial biomass may cause sinking of particles and as a consequence prevent their transport to the oceans. We studied microbial colonization of different polymer particles exposed in the epi-, meta- and hypolimnion of a freshwater reservoir during late summer for 47 days. Parameters measured included biofilm formation, metal sorption and sinking velocities. Microbial biofilms contained bacteria, cyanobacteria and algae as well as inorganic particles such as iron oxides. Regardless of biofilm thickness and biovolumes of different biofilm constituents, single polyethylene (PE) particles stayed buoyant, whereas the sinking velocity of single polystyrene (PS) and polyethylene terephthalate (PET) particles did not change significantly compared to initial values. During exposition, a mixing event occurred, by which anoxic, iron-rich water from the hypolimnion was mixed with water from upper layers. This induced aggregation and sinking of hypolimnetic PE particles together with organic matter, cyanobacteria colonies and iron minerals.

Importance of different physiological groups of iron reducing microorganisms in an acidic mining lake remediation experiment.

Iron- and sulfate-reducing microorganisms play an important role for alkalinity-generating processes in mining lakes with low pH. In the acidic mining lake 111 in Lusatia, Germany, a passive in situ remediation method was tested in a large scale experiment, in which microbial iron and sulfate reduction are stimulated by addition of Carbokalk (a mixture of the nonsugar compounds of sugar beets and lime) and straw. The treated surface sediment consisted of three layers of different pH and geochemical composition. The top layer was acidic and rich in Fe(III), the second and third layer both showed moderately acidic to circum-neutral pH values, but only the second was rich in organics, strongly reduced and sulfidic. Aim of the study was to elucidate the relative importance of neutrophilic heterotrophic, acidophilic heterotrophic, and acidophilic autotrophic iron-reducing microorganisms in each of the three layers. In order to distinguish between them, the effect of their respective characteristic electron donors acetate, glucose, and elemental sulfur on potential iron reduction rates was investigated. Limitation of iron reduction by the availability of Fe(III) was revealed by the addition of Fe(OH)(3). The three groups of iron-reducing microorganisms were quantified by most probable number (MPN) technique and their community composition was analyzed by cloning and sequencing of 16S rRNA genes. In the acidic surface layer, none of the three electron donors stimulated iron reduction; acetate even had an inhibiting effect. In agreement with this, no decrease of the added electron donors was observed. Iron reduction rates were low in comparison to the other layers. Iron reduction in layers 2 and 3 was enhanced by glucose and acetate, accompanied by a decrease of these electron donors. Addition of elemental sulfur did not enhance iron reduction in either layer. Layer 2 exhibited the highest iron reduction rate (4.08 mmol dm(-3) d(-1)) and the highest cell numbers in MPN media. In MPN enrichments from all layers, Acidithiobacillus-like sequences were frequent. In addition to these, sequences related to Fulvimonas and Clostridium dominated in layer 1. MPN enrichments of layer 2 were diverse, containing Rhodocyclaceae-related sequences and surprisingly low numbers of Geobacteraceae. In layer 3, Sulfobacillus and Trichococcus spp. were also important. It was concluded that in the surface layer mainly acidophilic, probably autotrophic and heterotrophic, iron reducers were active, whereas in layers 2 and 3 mainly neutrophilic heterotrophs were important for iron reduction. These differ from well-studied Fe(III) reducers in other environments, so they deserve further study. The potential for acid-producing sulfur-driven Fe(III) reduction seemed not to be critical for in situ remediation.

Occurrence and role of algae and fungi in acid mine drainage environment with special reference to metals and sulfate immobilization.

Passive remediation of Acid Mine Drainage (AMD) is a popular technology under development in current research. Roles of algae and fungi, the natural residents of AMD and its attenuator are not emphasized adequately in the mine water research. Living symbiotically various species of algae and fungi effectively enrich the carbon sources that help to maintain the sulfate reducing bacterial (SRB) population in predominantly anaerobic environment. Algae produce anoxic zone for SRB action and help in biogenic alkalinity generation. While studies on algal population and actions are relatively available those on fungal population are limited. Fungi show capacity to absorb significant amount of metals in their cell wall, or by extracellular polysaccharide slime. This review tries to throw light on the roles of these two types of microorganisms and to document their activities in holistic form in the mine water environment. This work, inter alia, points out the potential and gap areas of likely future research before potential applications based on fungi and algae initiated AMD remediation can be made on sound understanding.

Diversity and in situ quantification of Acidobacteria subdivision 1 in an acidic mining lake.

Analysis of 16S rRNA gene clone libraries from acidic mining lake water and sediment, and from an enclosure to which organic carbon was added to stimulate microbial alkalinization processes of sulfate and iron reduction revealed the presence of diverse sequences affiliated with the Acidobacteria subdivision 1. A novel oligonucleotide probe, ACIDO228, was designed that covered most sequences of Acidobacteria subdivision 1. The hybridization conditions were optimized using the type strain Acidobacterium capsulatum. The depth distribution and seasonal dynamics of Acidobacteria in the lake and the enclosure were assessed by whole cell hybridization. Sequence analyses and in situ quantification indicated that Acidobacteria accounted for a substantial part of bacterioplankton communities in both compartments. During the summer stratification distinct maxima of acidobacterial abundance were detected in the hypolimnion (up to 13% of total cell numbers), whereas during spring and autumn circulations no clear depth-dependent differences were visible. These data suggest that Acidobacteria thrive best in the hypolimnion, which is characterized by lower temperatures and higher availability of organic substrates. The application of probe ACIDO228 provided quantitative information on the seasonal and depth distribution of Acidobacteria in a lake environment and in particular in a rather extreme habitat, an acidic mining lake.

Vertical gradients in carbon flow and methane production in a sulfate-rich oil sands tailings pond.

Oil sands tailings ponds are primary storage basins for tailings produced during oil sands processing in Alberta (Canada). Due to microbial metabolism, methane production contributes to greenhouse gas emissions, but positively affects tailings densification, which is relevant for operational water re-use. Depending on the age and depth of tailings, the activity of sulfate-reducing bacteria (SRB) may control methanogenesis due to the competition for substrates. To assess the depth-related impact of sulfate reduction on CH4 emissions, original tailings of two vicinal pond profiles were incubated in anoxic microcosms with/without molybdate as selective inhibitor of microbial sulfate reduction. Integrating methane production rates, considerable volumes of CH4 emissions (∼5.37 million L d-1) may be effectively prevented by the activity of SRB in sulfidic tailings between 3.5 and 7.5 m. To infer metabolic potentials controlling methanogenic pathways, a set of relevant organic acids (acetate, formate, propionate, butyrate, lactate) was added to part of the microcosms. Generally, organic acid transformation shifted with depth, with highest rates (305-446 µmol L-1 d-1) measured in fresh tailings at 5.5-7.5 m. In all depths, a transient accumulation of acetate revealed its importance as key intermediate during organic matter decomposition. SRB dominated the transformation of acetate, butyrate and propionate, but were not essential for lactate and formate turnover. Acetate as methanogenic substrate was important only at 13.5 m. At 1-7.5 m, methanogenesis significantly increased in presence of organic acids, most likely due to the syntrophic oxidation of acetate to CO2 by SRB and subsequent conversion to CH4.

Anaerobic BTEX degradation in oil sands tailings ponds: Impact of labile organic carbon and sulfate-reducing bacteria.

The extraction of bitumen from oil sands in Alberta (Canada) produces volumes of tailings that are pumped into large anaerobic settling-basins. Beside bitumen, tailings comprise fractions of benzene, toluene, ethylbenzene and xylenes (BTEX) that derive from the application of industrial solvents. Due to their toxicity and volatility, BTEX pose a strong concern for gas- and water-phase environments in the vicinity of the ponds. The examination of two pond profiles showed that concentrations of indigenous BTEX decreased with depth, pointing at BTEX transformation in situ. With depth, the relative contribution of ethylbenzene and xylenes to total BTEX significantly decreased, while benzene increased relatively from 44% to 69%, indicating preferential hydrocarbon degradation. To predict BTEX turnover and residence time, we determined BTEX degradation rates in tailings of different depths in a 180-days microcosm study. In addition, we evaluated the impact of labile organic substrates (e.g. acetate) generally considered to stimulate hydrocarbon degradation and the contribution of sulfate-reducing bacteria (SRB) to BTEX turnover. In all depths, BTEX concentrations significantly decreased due to microbial activity, with degradation rates ranging between 4 and 9 µg kg(-1) d(-1). BTEX biodegradation decreased linearly in correlation with initial concentrations, suggesting a concentration-dependent BTEX transformation. SRB were not significantly involved in BTEX consumption, indicating the importance of methanogenic degradation. BTEX removal decreased to 70-90% in presence of organic substrates presumptively due to an accumulation of acetate that lowered BTEX turnover due to product inhibition. In those assays SRB slightly stimulated BTEX transformation by reducing inhibitory acetate levels.

Interaction of microbial sulphate reduction and methanogenesis in oil sands tailings ponds.

Anaerobic turnover of organic compounds in oil sands tailings ponds is accomplished by a complex microbial consortium. We examined major electron accepting processes in mature fine tailings (MFT). Beside methanogenesis and sulphate reduction, microbial iron reduction was an important process of anaerobic respiration. Microbial numbers and activity were comparable to those reported for natural lakes. To understand metabolic interactions of indigenous methanogenic and sulphate-reducing communities, we conducted a 6 month microcosm experiment with MFT supplemented with easily available carbon sources and molybdate and/or 2-bromoethane sulphonate (BES) as specific inhibitors for sulphate reduction and methanogenesis. Methanogenesis increased when microcosms were supplemented with extra carbon, but was completely inhibited by the addition of BES. Molybdate not only inhibited sulphate reduction, but also methanogenesis, indicating a positive relation between the two processes. The turnover of extra carbon sources differed between microcosms treated with molybdate and BES. Acetate and propionate were not consumed in microcosms amended with molybdate, indicating that sulphate-reducing bacteria (SRB) were responsible for their metabolisation, and that methane was rather produced by hydrogenotrophic methanogens. In microcosms without molybdate, acetate transiently accumulated, indicating the activity of both incomplete and complete oxidizing SRB. Ethanol and lactate were also consumed in the simultaneous presence of BES and molybdate, demonstrating the occurrence of other anaerobic processes. Biomass increased by the addition of extra carbon, mainly due to a relative increase in the proportion of SRB. The addition of extra carbon lowered the degradation of BTEX compounds.

Structure and function of the microbial community in an in situ reactor to treat an acidic mine pit lake.

Sulfate-reducing bioreactors are a promising option for the treatment of acid mine drainage. We studied the structure and function of a biofilm in a methanol-fed fixed-bed in-lake reactor for the treatment of an acidic pit lake by a combination of laboratory incubations, chemical and molecular analyses and confocal laser scanning microscopy to determine whether competition by different groups of microorganisms as well as the precipitation of minerals affect reactor performance negatively. The biofilm growing on the surface of a synthetic carrier material consisted of dense microbial colonies covered by iron-sulfide precipitates. The microorganisms continuously had to overgrow this mineral coating, resulting in a high biomass turnover. About one third of the added methanol was used by sulfate reduction, and the rest by competing reactions. Sulfate-reducing bacteria as well as methanogens and acetogens were involved in methanol consumption. Six different groups of Deltaproteobacteria, dominated by the genera Desulfomonile, Desulfobacterium and a phylotype related to Geobacter, Gram-positive sulfate reducers of the genus Desulfosporosinus, acetogenic Acetobacteria, different fermenting bacteria as well as methylotrophic methanogens were identified. The versatility of the microbial food web is probably an important factor stabilizing the biofilm function under fluctuating and partly oxidizing conditions in the reactor.

Processes at the sediment water interface after addition of organic matter and lime to an Acid Mine Pit Lake mesocosm.

A strategy to neutralize acidic pit lakes was tested in a field mesocosm of 4500 m(3) volume in the Acidic Pit Mine Lake 111 in Germany. Carbokalk, a byproduct from sugar production, and wheat straw was applied near to the sediment surface to stimulate in lake microbial alkalinity generation by sulfate and iron reduction. The biogeochemical processes at the sediment-water interface were studied over 3 years by geochemical monitoring and an in situ microprofiler. Substrate addition generated a reactive zone at the sediment surface where sulfate and iron reduction proceeded. Gross sulfate reduction reached values up to 10 mmol m(-2) d(-1). The neutralization rates between 27 and 0 meq m(-2) d(-1) were considerably lower than in previous laboratory experiments. The precipitation of ferric iron minerals resulted in a growing acidic sediment layer on top of the neutral sediment. In this layer sulfate reduction was observed but iron sulfides could not precipitate. In the anoxic sediment H2S was oxidized by ferric iron minerals. H2S partly diffused to the water column where it was oxidized. As a result the net formation of iron sulfides decreased after 1 year although gross sulfate reduction rates continued to be high. The rate of iron reduction exceeded the sulfate reduction rate, which resulted in high fluxes of ferrous iron out of the sediment.

Functional groups and activities of bacteria in a highly acidic volcanic mountain stream and lake in Patagonia, Argentina.

Acidic volcanic waters are naturally occurring extreme habitats that are subject of worldwide geochemical research but have been little investigated with respect to their biology. To fill this gap, the microbial ecology of a volcanic acidic river (pH approximately equal to 0-1.6), Rio Agrio, and the recipient lake Caviahue in Patagonia, Argentina, was studied. Water and sediment samples were investigated for Fe(II), Fe(III), methane, bacterial abundances, biomass, and activities (oxygen consumption, iron oxidation and reduction). The extremely acidic river showed a strong gradient of microbial life with increasing values downstream and few signs of life near the source. Only sulfide-oxidizing and fermentative bacteria could be cultured from the upper part of Rio Agrio. However, in the lower part of the system, microbial biomass and oxygen penetration and consumption in the sediment were comparable to non-extreme aquatic habitats. To characterize similarities and differences of chemically similar natural and man-made acidic waters, our findings were compared to those from acidic mining lakes in Germany. In the lower part of the river and the lake, numbers of iron and sulfur bacteria and total biomass in sediments were comparable to those known from acidic mining lakes. Bacterial abundance in water samples was also very similar for both types of acidic water (around 10(5) mL(-1)). In contrast, Fe(II) oxidation and Fe(III) reduction potentials appeared to be lower despite higher biogenic oxygen consumption and higher photosynthetic activity at the sediment-water interface. Surprisingly, methanogenesis was detected in the presence of high sulfate concentrations in the profundal sediment of Lake Caviahue. In addition to supplementing microbiological knowledge on acidic volcanic waters, our study provides a new view of these extreme sites in the general context of aquatic habitats.

Microbial sulfate reduction at low pH in sediments of an acidic lake in Argentina.

We measured sulfate reduction in the acidic (pH < or = 3) sediment of an Argentinean lake influenced by volcanism. Sulfate reduction rates of 2.04 mmol m(-2) d(-1) were determined with a 35SO4(2-) core injection method and confirmed by batch incubations and from H2S measurements in the sediment. H2S production stopped when iron reduction was stimulated by addition of ferric iron. The results suggest that sulfate reduction at pH values around 3 is possible and can probably be used in biotechnological strategies if competing microbial processes are inhibited and electron donors are highly available.