Cyanobacterial Biofilms Research Articles

Toxicity, transfer and depuration of anatoxin-a (cyanobacterial neurotoxin) in medaka fish exposed by single-dose gavage

The proliferations of cyanobacteria are increasingly prevalent in many rivers and water bodies due especially to eutrophication. This work aims to study in female medaka fish the toxicity, the transfer and the depuration of the anatoxin-a, a neurotoxin produced by benthic cyanobacterial biofilms. This work will provide answers regarding acute toxicity induced by single gavage by anatoxin-a and to the risks of exposure by ingestion of contaminated fish flesh, considering that data on these aspects remain particularly limited.The oral LD50 and NOAEL of a single dose of (±)-anatoxin-a were determined at 11.50 and 6.67 μg.g−1, respectively. Subsequently, the toxico-kinetics of the (±)-anatoxin-a was observed in the guts, the livers and the muscles of female medaka fish for 10 days. Anatoxin-a was quantified by high-resolution qTOF mass spectrometry coupled upstream to a UHPLC chromatographic chain. The toxin could not be detected in the liver after 12 h, and in the gut and muscle after 3 days. Overall, the medaka fish do not appear to accumulate (±)-anatoxin-a and to largely recover after 24 h following a single sub-acute oral liquid exposure at the NOAEL.

Axenic cyanobacterial (Nostoc muscorum) biofilm as a platform for Cd(II) sequestration from aqueous solutions

An environment friendly approach towards heavy metal remediation from water is the need of the hour. A novel approach to heavy metal bioremediation from water was attempted using axenic cultures of the photoautotrophic cyanobacterium, Nostoc muscorum immobilised on to glass surface through the formation of biofilms. Being a biofilm, it exhibited much higher cellular integrity than cell suspension cultures when exposed to high concentrations of cadmium, and could also withstand the harsh conditions prevalent in industrial effluents. Being a photosynthetic organism, culturing of N. muscorum as a biofilm was achieved with minimal requirements, such as water with minerals, light and stationary conditions, which also enabled ease of scalability. The strong adherence of the N. muscorum biofilm to the glass surface was beneficial as (i) no diffusion of cell mass on to the media was observed even with the use of top-end stirrer, (ii) no expensive separation techniques were required for separation of biomass from the treated solution, and (iii) direct contact of the solution containing metals to the active components on biofilm surface allowed better interaction. The N. muscorum biofilm exhibited the capability to sequester Cd(II) in the pH range 5–9, concentration range <0.05 ppm to 100 ppm and from different waste waters. Sequestration of Cd(II) resulted in cell surface changes, the mode of sequestration being chemisorption through the >C=O and >C=N functional groups, and followed the Langmuir's adsorption isotherm. It is proposed to use the N. muscorum biofilm as a potential green technology for waste water treatment in future.

A Storable Mediatorless Electrochemical Biosensor for Herbicide Detection.

A novel mediatorless photo-bioelectrochemical sensor operated with a biofilm of the cyanobacterium Synechocystis PCC6803 wt. for herbicide detection with long term stability (>20 days) was successfully developed and tested. Photoanodic current generation was obtained in the absence of artificial mediators. The inhibitory effect on photocurrent of three commonly used herbicides (i.e., atrazine, diuron, and paraquat) was used as a means of measuring their concentrations in aqueous solution. The injection of atrazine and diuron into the algal medium caused an immediate photocurrent drop due to the inhibition of photosynthetic electron transport. The detected concentrations were suitable for environmental analysis, as revealed by a comparison with the freshwater quality benchmarks set by the Environmental Protection Agency of the United States (US EPA). In contrast, paraquat caused an initial increase (~2 h) of the photocurrent effect of about 200%, as this compound can act as a redox mediator between the cells and the anode. A relatively long-term stability of the biosensor was demonstrated, by keeping anodes colonized with cyanobacterial biofilm in the dark at 4 °C. After 22 days of storage, the performance in terms of the photocurrent was comparable with the freshly prepared biosensor. This result was confirmed by the measurement of chlorophyll content, which demonstrated preservation of the cyanobacterial biofilm. The capacity of this biosensor to recover after a cold season or other prolonged environmental stresses could be a key advantage in field applications, such as in water bodies and agriculture. This study is a step forward in the biotechnological development and implementation of storable mediatorless electrochemical biosensors for herbicide detection.

Compositional homogeneity in the pathobiome of a new, slow-spreading coral disease

BackgroundCoral reefs face unprecedented declines in diversity and cover, a development largely attributed to climate change-induced bleaching and subsequent disease outbreaks. Coral-associated microbiomes may strongly influence the fitness of their hosts and alter heat tolerance and disease susceptibility of coral colonies. Here, we describe a new coral disease found in Micronesia and present a detailed assessment of infection-driven changes in the coral microbiome.ResultsCombining field monitoring and histological, microscopic and next-generation barcoding assessments, we demonstrate that the outbreak of the disease, named ‘grey-patch disease’, is associated with the establishment of cyanobacterial biofilm overgrowing coral tissue. The disease is characterised by slow progression rates, with coral tissue sometimes growing back over the GPD biofilm. Network analysis of the corals’ microbiome highlighted the clustering of specific microbes which appeared to benefit from the onset of disease, resulting in the formation of ‘infection clusters’ in the microbiomes of apparently healthy corals.ConclusionsOur results appear to be in contrast to the recently proposed Anna-Karenina principle, which states that disturbances (such as disease) trigger chaotic dynamics in microbial communities and increase β-diversity. Here, we show significantly higher community similarity (compositional homogeneity) in the pathobiome of diseased corals, compared to the microbiome associated with apparently healthy tissue. A possible explanation for this pattern is strong competition between the pathogenic community and those associated with the ‘healthy’ coral holobiont, homogenising the composition of the pathobiome. Further, one of our key findings is that multiple agents appear to be involved in degrading the corals’ defences causing the onset of this disease. This supports recent findings indicating a need for a shift from the one-pathogen-one-disease paradigm to exploring the importance of multiple pathogenic players in any given disease.

The patterns of nitrogen fixation in haloalkaliphilic phototrophic communities of Kulunda Steppe soda lakes (Altai, Russia).

Nitrogen fixation (NF) of phototrophic communities was studied in a number of soda lakes with a wide range of salinity (25-400 g/l) located in Kulunda Steppe (Altai, Russia) during several summer seasons (2011-2016). The phototrophic communities were represented by the algal-bacterial Ctenocladus communities or cyanobacterial biofilms dominated by heterocystous and non-heterocystous cyanobacteria and purple sulfur bacteria Ectothiorhodospira sp. (up to 210 g/l) and endoevaporitic Euhalothece communities dominated by the extremely salt-tolerant unicellular cyanobacterium Euhalothece sp. and Ectothiorhodospira sp. (above 350 g/l). Salinity was the major factor influencing the composition and NF potential of the phototrophic communities. The communities dominated by vegetative heterocystous cyanobacteria exhibited light-independent NF at total salinity up to 60 g/l. The communities dominated by non-heterocystous cyanobacteria exhibited light-dependent NF in a range of 55-100 g/l, but it was significantly suppressed at 100 g/l. At 160-200 g/l the dark heterotrophic NF was a prevailing process if communities didn't contain Euhalothece sp. At salt-saturating ranges above 350 g/l, light-dependent NF associated with the Euhalothece communities was detected. A statistically significant positive correlation between the NF and diurnal light intensity was found in all samples of communities dominated by non-heterocystous cyanobacteria in contrast to communities dominated by heterocystous cyanobacteria with insignificant correlation coefficients.

Biofilm formation behaviour of marine filamentous cyanobacterial strains in controlled hydrodynamic conditions.

Marine biofouling has severe economic impacts and cyanobacteria play a significant role as early surface colonizers. Despite this fact, cyanobacterial biofilm formation studies in controlled hydrodynamic conditions are scarce. In this work, computational fluid dynamics was used to determine the shear rate field on coupons that were placed inside the wells of agitated 12-well microtiter plates. Biofilm formation by three different cyanobacterial strains was assessed at two different shear rates (4 and 40 s-1 ) which can be found in natural ecosystems and using different surfaces (glass and perspex). Biofilm formation was higher under low shear conditions, and differences obtained between surfaces were not always statistically significant. The hydrodynamic effect was more noticeable during the biofilm maturation phase rather than during initial cell adhesion and optical coherence tomography showed that different shear rates can affect biofilm architecture. This study is particularly relevant given the cosmopolitan distribution of these cyanobacterial strains and the biofouling potential of these organisms.

What Causes Carbonates to Form “Shrubby” Morphologies? An Anthropocene Limestone Case Study

The South Atlantic Aptian “Pre-salt” shrubby carbonate Formations of Brazil and Angola are of major interest for the oil industry due to their potential hydrocarbon accumulations. Although the general sedimentology of these deposits is widely recognised to be within saline, alkaline lakes in rift volcanic settings, the specific genesis of shrubby carbonate morphologies remains unclear. This study reports the first petrographically comparable shrubby carbonates amongst other carbonate microfacies from an Anthropocene limestone formed under hyperalkaline (pH 9-12) and hypersaline (conductivity 425-3200µS) conditions at ambient temperature (12.5-13 oC) (Consett, UK). This discovery allows us to capitalise on exceptional long-term hydrochemical monitoring efforts from the site, demonstrating that shrubby carbonates occur uniquely within the waters richest in calcium (~240mg/L) and with highest pH (~12) and consequently with very high levels of supersaturation. However, the physical distribution of shrubs is more comparable with estimated local kinetic precipitation rate than it is to thermodynamic saturation, indicating that the fundamental control on shrub formation arises from crystal surface processes. The shrubby carbonate we report grows in the presence of significant diatomaceous and cyanobacterial biofilms, despite the highly alkaline conditions. These biofilms are lost from the deposited material early due to the high solubility of organic and silica within in hyperalkaline settings, and this loss contributes to very high intercrystalline porosity. Despite the presence of these microbes, few if any of the fabrics we report would be considered as “boundstones” despite it being clear that most fabrics are being deposited in the presence of abundant extra-cellular polymeric substances. We are aware of no previous petrographic work on anthropogenic carbonates of this type, and recommend further investigation to capitalise on what can be learned from these “accidental laboratories”.

The Role of the Substrate on the Mineralization Potential of Microbial Mats in A Modern Freshwater River (Paris Basin, France)

The relationship between environmental conditions and the development, mineralization and preservation of modern tufa microbialites was investigated in a 1.1 km long freshwater stream in Villiers-le-Bâcle, a tributary of Mérantaise river. Detailed mapping of the tufa microbialite distribution combined with sedimentological, petrographical and mineralogical analyses were coupled with chemical measurements. Six organosedimentary structures were identified; their distribution appears heterogeneous along the stream and responds to physicochemical conditions of water and specific biological components (e.g., microorganism, exopolymeric substance). Two of the organosedimentary structures show evidence of mineralization and only one is lithified. Based on field observations and in-situ deployment of mineralization markers (bricks), three zones with increasing mineralization intensities are defined, ranging from no mineralization to thick mineralized crusts forming riverine tufa. Both biotic and abiotic processes were proposed for the tufa microbialite formation. We explained changes in mineralization intensities by the specific physicochemical conditions (e.g., calcite saturation index (SIcalc) and partial pressure of CO2 (pCO2) and a closed proximity of the cyanobacterial biofilm and carbonates precipitates. The physical and chemical composition of substrate impact development of microbial communities, mineralization potential of tufa microbialite. Even though the physicochemical and biological conditions were optimal for mineral precipitation, the potential of lithification depended on the presence of a suitable (physical and chemical) substrate.

Surface colour: An overlooked aspect in the study of cyanobacterial biofilm formation

Cyanobacteria can grow as biofilms, communities that colonize surfaces and that play a fundamental role in the ecology of many diverse habitats and in the conversion of industrial production to green platforms. Although biofilm growth is known to be significantly affected by several characteristics, the effect of colour surface is an overlooked aspect that has not yet been investigated. In this study, we describe the effect of colour hues (white, red, blue and black) on the growth of cyanobacterial biofilms on air-exposed substrates. We measured growth, architecture, pigment production and levels of ATP and reactive oxygen species in cyanobacterial biofilms formed on different coloured substrates. The study findings demonstrate, for the first time, that the colour of a surface affects biofilm formation at the air-solid interface (with more biomass accumulating on white and red substrates than on blue and black substrates) and also alters the biofilm architecture. In addition, the roles of chromatic adaptation, phototrophic cells and reactive oxygen species as intermediates between colour sensing and biofilm response are discussed. Our results support the importance of colour as a new factor that favours surface colonization by cyanobacteria and its contribution to biofilm formation.

Mode of application influences the biofertilizing efficacy of cyanobacterial biofilm formulations in chrysanthemum varieties under protected cultivation

Abstract Availability of nutrients in soil plays an important role in the productivity and quality of flowers in chrysanthemum. A set of novel biofilm inoculants- Anabaena-Azotobacter, Anabaena-Pseudomonas fluorescens (An-Psf) and Anabaena-Trichoderma (An-Tr) were applied as carrier based dry formulation or soil drench and their performance compared in two varieties of chrysanthemum (White Star and Zembla), in a climate-controlled greenhouse. Both the An-Psf and (An-Tr) inoculants enhanced glomalin related soil proteins in the rhizosphere of White Star, while in terms of polysaccharide content of soil, both these inoculants performed better in Zembla variety. Significant increases in the availability of selected macro and micronutrients in rhizosphere soil samples, in both chrysanthemum varieties were recorded, particularly when the inoculants were applied as soil drench. Principal Component analysis illustrated the significant interaction among soil and plant parameters, more specifically, the distinct effect of the inoculants, as compared to the application of carrier alone or control treatment. This investigation demonstrated the varietal effects on soil biological activities and significance of mode of application of microbial inoculants in influencing plant growth and rhizospheric metabolic activities.

Cyanobacterial biofilms as light‐driven biocatalysts

Cyanobacterial biofilms as light‐driven biocatalysts

Composite polymer coated magnetic nanoparticles based anode enhances dye degradation and power production in microbial fuel cells

Combined power generation and waste degradation through microbial fuel cell (MFC) technology is emerging as an attractive solution for controlling pollution in water bodies. Cyanobacteria as fuel cell catalysts for such shared energy activities are not well studied even though these possess robust metabolic systems supporting exo-electrogenicity, biodegradation of toxic compounds, and their survival under wide environmental conditions. Herein, a dual chambered (50 ml each) MFC assembled with Synechococcus sp. based bioanode and abiotic cathode for simultaneous power generation and Mordant orange dye degradation is reported. The anode was prepared by encrusting chemically synthesised magnetic nanoparticle (MNP) of size 8.4 ± 0.2 nm with magnetization of 69 emu g−1on Toray carbon paper (TCP). The MNPs were encapsulated with aniline and pyrrole composite polymers to facilitate biofilm formation and cellular electron flow to the anode as confirmed by advance microscopic and voltametric techniques, respectively. The MFC with the dye mixed acetate produced current of 14.04 ± 5.5 A m−3 with a maximum power density of 4.9 ± 0.5 W m−3 (at cell voltage of 0.494 ± 0.05 V), which was 18% higher than the control (without dye). The MFC produced a high OCP of 0.949 ± 0.07 V and offered to decolorize 68.5% and degrade 89% of the dye following 216 h of its operation as confirmed by photometry (λ385 nm) and LC-MS/MS analyses, respectively. The efficient dye degradation is attributed to the bioanode for secreting high level of reactive oxygen species. The composite polymer coated MNPs anode with cyanobacterial biofilm is therefore, a highly efficient construct for enhanced azo dye degradation and associated power generation in a MFC system.

Bacterial Membrane Depolarization-Linked Fuel Cell Potential Burst as Signal for Selective Detection of Alcohol.

The biosensing application of microbial fuel cell (MFC) is hampered by its long response time, poor selectivity, and technical difficulty in developing portable devices. Herein, a novel signal form for rapid detection of ethanol was generated in a photosynthetic MFC (PMFC). First, a dual chambered (100 mL each) PMFC was fabricated by using cyanobacteria-based anode and abiotic cathode, and its performance was examined for detection of alcohols. A graphene-based nanobiocomposite matrix was layered over graphite anode to support cyanobacterial biofilm growth and to facilitate electron transfer. Injection of alcohols into the anodic chamber caused a transient potential burst of the PMFC within 60 s (load 1000 Ω), and the magnitude of potential could be correlated to the ethanol concentrations in the range 0.001-20% with a limit of detection (LOD) of 0.13% ( R2 = 0.96). The device exhibited higher selectivity toward ethanol than methanol as discerned from the corresponding cell-alcohol interaction constant ( Ki) of 780 and 1250 mM. The concept was then translated to a paper-based PMFC (p-PMFC) (size ∼20 cm2) wherein, the cells were merely immobilized over the anode. The device with a shelf life of ∼3 months detected ethanol within 10 s with a dynamic range of 0.005-10% and LOD of 0.02% ( R2 = 0.99). The fast response time was attributed to the higher wettability of ethanol on the immobilized cell surface as validated by the contact angle data. Alcohols degraded the cell membrane on the order of ethanol > methanol, enhanced the redox current of the membrane-bound electron carrier proteins, and pushed the anodic band gap toward more negative value. The consequence was the potential burst, the magnitude of which was correlated to the ethanol concentrations. This novel approach has a great application potential for selective, sensitive, rapid, and portable detection of ethanol.

Toxic Cyanobacteria in Svalbard: Chemical Diversity of Microcystins Detected Using a Liquid Chromatography Mass Spectrometry Precursor Ion Screening Method.

Cyanobacteria synthesize a large variety of secondary metabolites including toxins. Microcystins (MCs) with hepato- and neurotoxic potential are well studied in bloom-forming planktonic species of temperate and tropical regions. Cyanobacterial biofilms thriving in the polar regions have recently emerged as a rich source for cyanobacterial secondary metabolites including previously undescribed congeners of microcystin. However, detection and detailed identification of these compounds is difficult due to unusual sample matrices and structural congeners produced. We here report a time-efficient liquid chromatography-mass spectrometry (LC-MS) precursor ion screening method that facilitates microcystin detection and identification. We applied this method to detect six different MC congeners in 8 out of 26 microbial mat samples of the Svalbard Archipelago in the Arctic. The congeners, of which [Asp3, ADMAdda5, Dhb7] MC-LR was most abundant, were similar to those reported in other polar habitats. Microcystins were also determined using an Adda-specific enzyme-linked immunosorbent assay (Adda-ELISA). Nostoc sp. was identified as a putative toxin producer using molecular methods that targeted 16S rRNA genes and genes involved in microcystin production. The mcy genes detected showed highest similarities to other Arctic or Antarctic sequences. The LC-MS precursor ion screening method could be useful for microcystin detection in unusual matrices such as benthic biofilms or lichen.

Cyanobacterial-algal crusts from Late Ediacaran paleosols of the East European Craton

The top surface of the Ediacaran flood basalts of West Ukraine (Volyn), extruded during the break-up of Rodinia supercontinent, is weathered into several meters thick paleosol, covered by late Ediacaran (ca. 550 Ma) sediments. The paleosol grades from unaltered basaltic material into saprock and finally clay-dominated saprolite with a minor content of the organic matter. The upper parts are dominated by kaolinite and hematite. SEM images and SEM-EDS analyses revealed in one of the paleosol profiles a horizon with significant amount of morphologically recognizable organic structures, mineralized by silicates and iron oxides. HF-etching of the bulk rock samples revealed lithic fragments covered irregularly by a biocrust of colonial coccoid cyanobacteria associated with irregularly distributed unicellular algae. The cyanobacteria are preserved as mineralized common mucilage sheaths (glycocalyx), which reproduce the original shape typical of sheaths of modern coccoidal cyanobacterial biofilms, whereas the heavily mineralized algae show features characteristic of modern chlorococcaleans. The preservation of morphologically distinguishable remnants of cyanobacterial biofilms and aggregates of unicellular algae in terrestrial weathering deposits is a rare preservational phenomenon requiring rapid post-mortem permineralization of the dead microbiota. One of the main factors facilitating such a unique fossilization was supposedly a copious excretion of extracellular polymeric substances (EPS) by the biocrust forming biota in which early mineral nucleation occurred. The colonization of the Late Ediacaran land by the presumably desiccation-resistant cyanobacteria-dominated crust communities associated with algae was of importance for creating fertile soils and a soil-forming microbial ecosystem that later allowed a successful land invasion by the ancestors of vascular plants.

Microcystins in Cyanobacterial Biofilms from the Littoral Zone of Lake Baikal

Some species of cyanobacteria synthesize toxins whose concentration during water bloom can reach values dangerous for human and animal health. Planktonic cyanobacteria are the most common and well-studied microcystins producers, hepatotoxic cyclic heptapeptides, whereas microcystin-producing benthic cyanobacteria are less known. In recent years, the mass development of benthic cyanobacteria forming extensive fouling on different substrates has been detected in the littoral zone of Lake Baikal. We found microcystins produced by benthic cyanobacteria in the biofouling on different natural and artificial substrates, including diseased and dead endemic sponges Lubomirskia baicalensis and Baikalospongia spp. collected from the littoral area of Lake Baikal. Microscopic analysis of the biofouling revealed prevalence of representatives of Nostocales and Oscillatoriales with predominance of Tolypothrix distorta that is likely the main microcystin producer in Lake Baikal. According to enzyme-linked immunosorbent assay (ELISA), microcystin concentrations in biofouling were 29.8–3050 μg/kg dry weight. We identified eight microcystin variants using MALDI-TOF/TOF; [Dha7]MC-YR was detected in most samples. The presence of microcystins in biofilms formed on the surface of the artificial substrate by Phormidium autumnale was also recorded. The data obtained demonstrated the necessity to monitor potentially toxic species and concentrations of cyanotoxins in plankton and benthos in the littoral zone of Lake Baikal, especially in the regions with intense tourist and recreational activities.

New records of Cyanobacterial morphotypes with Leptolyngbya indica sp. nov. from terrestrial biofilms of the Lower Gangetic Plain, India

Macroscopic cyanobacterial biofilms were collected from alluvial plain soils and estuarine mangrove soils representing the Lower Gangetic Plains of South East Asia (India). The composition of the biofilms was investigated using light microscopy and field emission scanning electron microscopy of collected samples. In this study four simple trichal non-heterocytous morphotypes were found to be unique. Out of four, three morphotypes clearly showed differences with respect to described taxa as based on most recent taxonomic classification and possibly represent new report from the Indian subcontinent. One morphotype was successfully established under culture conditions and described as Leptolyngbya indica sp. nov. isolated from the alluvial arsenic affected rice field soil. This study provides vital information on morphotypic diversity of Cyanobacteria from specific biotopes which can contribute key information on their biogeography and potential application in green remediation.

Genome mining reveals high incidence of putative lipopeptide biosynthesis NRPS/PKS clusters containing fatty acyl-AMP ligase genes inbiofilm-forming cyanobacteria.

Cyanobacterial lipopeptides have antimicrobial and antifungal bioactivities with potential for use in pharmaceutical research. However, due to their hemolytic activity and cytotoxic effects on human cells, they may pose a health issue if produced in substantial amounts in the environment. In bacteria, lipopeptides can be synthesized via several well-evidenced mechanisms. In one of them, fatty acyl-AMP ligase (FAAL) initiates biosynthesis by activation of a fatty acyl residue. We have performed a bioinformatic survey of the cyanobacterial genomic information available in the public databases for the presence of FAAL-containing non-ribosomal peptide synthetase/polyketide synthetase (NRPS/PKS) biosynthetic clusters, as a genetic basis for lipopeptide biosynthesis. We have identified 79 FAAL genes associated with various NRPS/PKS clusters in 16% of 376 cyanobacterial genomic assemblies available, suggesting that FAAL is frequently incorporated in NRPS/PKS biosynthetases. FAAL was present either as a stand-alone protein or fused either to NRPS or PKS. Such clusters were more frequent in derived phylogenetic lineages with larger genome sizes, which is consistent with the general pattern of NRPS/PKS pathways distribution. The putative lipopeptide clusters were more frequently found in genomes of cyanobacteria that live attached to surfaces and are capable of forming microbial biofilms. While lipopeptides are known in other bacterial groups to play a role in biofilm formation, motility, and colony expansion, their functions in cyanobacterial biofilms need to be tested experimentally. According to our data, benthic and terrestrial cyanobacteria should be the focus of a search for novel candidates for lipopeptide drug synthesis and the monitoring of toxic lipopeptide production.

Hidden biofilms in a far northern lake and implications for the changing Arctic

Shallow lakes are common across the Arctic landscape and their ecosystem productivity is often dominated by benthic, cyanobacterial biofilms. Many of these water bodies freeze to the bottom and are biologically inactive during winter, but full freeze-up is becoming less common with Arctic warming. Here we analyzed the microbiome structure of newly discovered biofilms at the deepest site of a perennially ice-covered High Arctic lake as a model of polar microbial communities that remain unfrozen throughout the year. Biofilms were also sampled from the lake’s shallow moat region that melts out and refreezes to the bottom annually. Using high throughput small subunit ribosomal RNA sequencing, we found more taxonomic richness in Bacteria, Archaea and microbial eukaryotes in the perennially unfrozen biofilms compared to moat communities. The deep communities contained both aerobic and anaerobic taxa including denitrifiers, sulfate reducers, and methanogenic Archaea. The water overlying the deep biofilms was well oxygenated in mid-summer but almost devoid of oxygen in spring, indicating anoxia during winter. Seasonally alternating oxic-anoxic regimes may become increasingly widespread in polar biofilms as fewer lakes and ponds freeze to the bottom, favoring prolonged anaerobic metabolism and greenhouse gas production during winter darkness.

Increased methane production in cyanobacteria and methanogenic microbe co-cultures

A novel light-to-bioenergy system produced 3.5 times the baseline methane output using a co-culture of cyanobacteria (Oscillatoria sp.) and a methanogenic microbial community. Analysis of micronutrients in the system during the growth phase indicated that cobalt, iron, nickel and zinc were not appreciably consumed. The stable consumption and return of macronutrients calcium and magnesium were also observed. Essential macronutrients nitrogen, in the form of nitrate, and phosphorus showed no cycling during the growth phase and were depleted at rates of 0.35mg/L/day and 0.40µg/L/day, respectively. Biofilm formation increased the resilience of biomass to bacterial degradation in an anaerobic digester, as shown by viability assays of cyanobacterial biofilms in the co-culture.