Jinjun Kan

Current Production by Bacterial Communities in Microbial Fuel Cells Enriched from Wastewater Sludge with Different Electron Donors

Electricity production by bacterial communities enriched from wastewater sludge with lactate, succinate, N-acetyl-D-glucosamine (NAG), acetate, formate, and uridine were monitored in dual-chamber microbial fuel cells (MFCs). Stable electricity production was observed after 300 h for communities enriched from lactate, acetate, and formate, while communities enriched with succinate, NAG, and uridine stabilized only after 700 h. The average peak current densities and maximum power densities generated from bacterial consortia were significantly higher than those generated from pure cultures of Shewanella oneidensis MR-1. Microbial assemblages were analyzed by DGGE, and planktonic and anode-attached bacterial communities varied as a function of electron donors: Firmicutes, β-Proteobacteria, and Bacteroidetes dominated the planktonic bacterial communities while anode-attached communities consisted mainly of δ-Proteobacteria, β-Proteobacteria, and Firmicutes. Similar bacterial populations were enriched in MFCs fed with lactate, NAG, and uridine and with succinate, acetate, and formate. Cross-feeding experiments with different fuels indicated that enriched microbial consortia were able to utilize a variety of fuel sources and displayed considerable stability, efficiency, and robustness of power generation in comparison to pure cultures. In addition, characterizations of cultivated Shewanella strains suggested that DGGE analysis likely missed active members of exoelectrogenic populations.

Archaea in Yellowstone Lake

The Yellowstone geothermal complex has yielded foundational discoveries that have significantly enhanced our understanding of the Archaea. This study continues on this theme, examining Yellowstone Lake and its lake floor hydrothermal vents. Significant Archaea novelty and diversity were found associated with two near-surface photic zone environments and two vents that varied in their depth, temperature and geochemical profile. Phylogenetic diversity was assessed using 454-FLX sequencing (∼51 000 pyrosequencing reads; V1 and V2 regions) and Sanger sequencing of 200 near-full-length polymerase chain reaction (PCR) clones. Automated classifiers (Ribosomal Database Project (RDP) and Greengenes) were problematic for the 454-FLX reads (wrong domain or phylum), although BLAST analysis of the 454-FLX reads against the phylogenetically placed full-length Sanger sequenced PCR clones proved reliable. Most of the archaeal diversity was associated with vents, and as expected there were differences between the vents and the near-surface photic zone samples. Thaumarchaeota dominated all samples: vent-associated organisms corresponded to the largely uncharacterized Marine Group I, and in surface waters, ∼69–84% of the 454-FLX reads matched archaeal clones representing organisms that are Nitrosopumilus maritimus-like (96–97% identity). Importance of the lake nitrogen cycling was also suggested by >5% of the alkaline vent phylotypes being closely related to the nitrifier Candidatus Nitrosocaldus yellowstonii. The Euryarchaeota were primarily related to the uncharacterized environmental clones that make up the Deep Sea Euryarchaeal Group or Deep Sea Hydrothermal Vent Group-6. The phylogenetic parallels of Yellowstone Lake archaea to marine microorganisms provide opportunities to examine interesting evolutionary tracks between freshwater and marine lineages.

Yellowstone Lake: high‐energy geochemistry and rich bacterial diversity

Yellowstone Lake is central to the balanced functioning of the Yellowstone ecosystem, yet little is known about the microbial component of its food chain. A remotely operated vehicle provided video documentation (http://www.tbi.montana.edu/media/videos/) and allowed sampling of dilute surface zone waters and enriched lake floor hydrothermal vent fluids. Vent emissions contained substantial H(2)S, CH(4), CO(2) and H(2), although CH(4) and H(2) levels were also significant throughout the lake. Pyrosequencing and near full-length sequencing of Bacteria 16S rRNA gene diversity associated with two vents and two surface water environments demonstrated that this lake contains significant bacterial diversity. Biomass was size-fractionated by sequentially filtering through 20-µm-, 3.0-µm-, 0.8-µm- and 0.1-µm-pore-size filters, with the >0.1 to <0.8 µm size class being the focus of this study. Major phyla included Acidobacteria, Actinobacteria, Bacteroidetes, α- and β-Proteobacteria and Cyanobacteria, with 21 other phyla represented at varying levels. Surface waters were dominated by two phylotypes: the Actinobacteria freshwater acI group and an α-Proteobacteria clade tightly linked with freshwater SAR11-like organisms. We also obtained evidence of novel thermophiles and recovered Prochlorococcus phylotypes (97-100% identity) in one near surface photic zone region of the lake. The combined geochemical and microbial analyses suggest that the foundation of this lakes food chain is not simple. Phototrophy presumably is an important driver of primary productivity in photic zone waters; however, chemosynthetic hydrogenotrophy and methanotrophy are likely important components of the lakes food chain.

Electricity generation from a floating microbial fuel cell

A floating microbial fuel cell (FMFC) has been designed and its performance has been evaluated for 153 days. The power output gradually increased to a maximum value of 390 mW/m(3) at 125 days. The polarization resistance of the anode (R(p)(a)) changed with operating time reaching a minimum value at 125 days, while the polarization resistance of the cathode (R(p)(c)) was relatively constant and much smaller than R(p)(a). It has been demonstrated that the observed changes of the internal resistance (R(int)) and the maximum power (P(max)) with exposure time were mainly due to the changes of R(p)(a). Compared with sediment MFCs for which the anode is embedded in marine or river sediments, the application of the FMFC, which could be installed in a buoy, is not limited by the depth of the ocean. The FMFC has the potential to supply electricity to low-power consuming electronic devices at remote locations.

Adaptation of soil microbes during establishment of microbial fuel cell consortium fed with lactate.

We report the development of microbial populations and changes in their electrochemical production during a 2-month study of a two-chamber microbial fuel cell (MFC). The original inoculum was taken from anaerobic enrichment cultures with soil as the inoculum, and lactate as the exogenous electron donor. Power density (PD), coulombic production (CP), and coulombic efficiency (CE) increased rapidly, reaching high values (320 mW m(-3), 65 Q, and 12.5%, respectively) in 12-16 days. Under these conditions, several major microbial taxa dominated the anode population. The medium solution in the cathode chamber decreased with aeration, resulting in a decrease in PD to 55 mW m(-3) at day 20. Refilling the cathode chamber around day 30 resulted in restoration of the PD, CP and CE to values equal to or greater than those previously observed. However, after the change in conditions, a marked change in community structure was observed, and high levels of acetate were seen in the anode chamber of the fuel cell for the first time. At day 35, a series of lactate concentrations were used, beginning with low levels and increasing to the 20 mM level originally used (day 46), the PD decreased but was stable at 150 mW m(-3) and the acetate concentration in the anode stabilized at about 35 mM. Under these conditions, new major population structures, which were closely related to Propionibacterium, Clostridium, and uncultured bacteria, were observed in the anode. These results suggested that the flexibility of community structure was important for sustainable electricity production.

Diverse bacterial groups are associated with corrosive lesions at a Granite Mountain Record Vault (GMRV)

Aims:  This study applied culture‐dependent and molecular approaches to examine the bacterial communities at corrosion sites at Granite Mountain Record Vault (GMRV) in Utah, USA, with the goal of understanding the role of microbes in these unexpected corrosion events.

Marine microbial community response to inorganic and organic sediment amendments in laboratory mesocosms

Sediment amendments provide promising strategies of enhancing sequestration of heavy metals and degradation of organic contaminants. The impacts of sediment amendments for metal and organic remediation including apatite, organoclay (and apatite and organoclay in geotextile mats), acetate, and chitin on environmental microbial communities in overlying water and sediment profiles are reported here. These experiments were performed concurrent with an ecotoxicity evaluation (data submitted in companion paper) and X-ray absorption spectroscopy of zinc speciation post apatite amendments. X-ray absorption spectra showed that a modest modification of zinc speciation occurred in amended treatments. Significant changes in both bacterial cell densities and populations were observed in response to amendments of apatite+organoclay, chitin, and acetate. The enriched bacteria and breakdown of these amendments were likely attributed to water quality degradation (e.g. ammonia and dissolved oxygen). Molecular fingerprints of bacterial communities by denaturant gradient gel electrophoresis (DGGE) showed that distinct bacterial populations occurred in overlying waters from different amendments: apatite+organoclay led to the dominance of Gammaproteobacteria, acetate enriched Alphaproteobacteria, and chitin treatment led to a dominance of Bacteroidetes and Alphaproteobacteria. In amended sediments, Firmicutes, Bacteroidetes, and Deltaproteobacteria (Desulfovibrio) were commonly found with chitin and apatite+chitin treatments. Finally, sulfate-reducing bacteria (e.g. Desulfovibrio) and metal-reducing bacteria were also recovered with most probable number (MPN) analyses in treatments with acetate, chitin, and apatite+chitin. These geochemically important bacteria were stimulated by amendments and may play critical functional roles in the metal and organic contaminant remediation process for future investigations of contaminated sediments.

Geomicrobiology of sublacustrine thermal vents in Yellowstone Lake: geochemical controls on microbial community structure and function

Yellowstone Lake (Yellowstone National Park, WY, USA) is a large high-altitude (2200 m), fresh-water lake, which straddles an extensive caldera and is the center of significant geothermal activity. The primary goal of this interdisciplinary study was to evaluate the microbial populations inhabiting thermal vent communities in Yellowstone Lake using 16S rRNA gene and random metagenome sequencing, and to determine how geochemical attributes of vent waters influence the distribution of specific microorganisms and their metabolic potential. Thermal vent waters and associated microbial biomass were sampled during two field seasons (2007–2008) using a remotely operated vehicle (ROV). Sublacustrine thermal vent waters (circa 50–90°C) contained elevated concentrations of numerous constituents associated with geothermal activity including dissolved hydrogen, sulfide, methane and carbon dioxide. Microorganisms associated with sulfur-rich filamentous “streamer” communities of Inflated Plain and West Thumb (pH range 5–6) were dominated by bacteria from the Aquificales, but also contained thermophilic archaea from the Crenarchaeota and Euryarchaeota. Novel groups of methanogens and members of the Korarchaeota were observed in vents from West Thumb and Elliots Crater (pH 5–6). Conversely, metagenome sequence from Mary Bay vent sediments did not yield large assemblies, and contained diverse thermophilic and nonthermophilic bacterial relatives. Analysis of functional genes associated with the major vent populations indicated a direct linkage to high concentrations of carbon dioxide, reduced sulfur (sulfide and/or elemental S), hydrogen and methane in the deep thermal ecosystems. Our observations show that sublacustrine thermal vents in Yellowstone Lake support novel thermophilic communities, which contain microorganisms with functional attributes not found to date in terrestrial geothermal systems of YNP.

A rapid fingerprinting approach to distinguish between closely related strains of Shewanella.

One of the big operational problems facing laboratories today is the ability to rapidly distinguish between strains of bacteria that, while physiologically distinct, are nearly impossible to separate based on 16S rRNA gene sequence differences. Here we demonstrate that ITS-DGGE provides a convenient approach to distinguishing between closely related strains of Shewanella, some of which were impossible to separate and identify by 16 rRNA gene sequence alone. Examined Shewanella genomes contain 8-11 copies of rrn (ribosomal RNA gene) operons, and variable size and sequence of 16S-23S ITS (intergenic transcribed spacer) regions which result in distinct ITS-DGGE profiles. Phylogenetic constructions based on ITS are congruent with the genomic trees generated from concatenated core genes as well as with those based on conserved indels, suggesting that ITS patterns appear to be linked with evolutionary lineages and physiology. In addition, three new Shewanella strains (MFC 2, MFC 6, and MFC 14) were isolated from microbial fuel cells enriched from wastewater sludge and identified by ITS-DGGE. Subsequent physiological and electrochemical studies of the three isolates confirmed that each strain is phenotypically/genotypically distinct. Thus, this study validates ITS-DGGE as a quick fingerprint approach to identifying and distinguishing between closely related but novel Shewanella ecotypes.

Geochemistry and Mixing Drive the Spatial Distribution of Free-Living Archaea and Bacteria in Yellowstone Lake

Yellowstone Lake, the largest subalpine lake in the United States, harbors great novelty and diversity of Bacteria and Archaea. Size-fractionated water samples (0.1–0.8, 0.8–3.0, and 3.0–20 μm) were collected from surface photic zone, deep mixing zone, and vent fluids at different locations in the lake by using a remotely operated vehicle (ROV). Quantification with real-time PCR indicated that Bacteria dominated free-living microorganisms with Bacteria/Archaea ratios ranging from 4037:1 (surface water) to 25:1 (vent water). Microbial population structures (both Bacteria and Archaea) were assessed using 454-FLX sequencing with a total of 662,302 pyrosequencing reads for V1 and V2 regions of 16S rRNA genes. Non-metric multidimensional scaling (NMDS) analyses indicated that strong spatial distribution patterns existed from surface to deep vents for free-living Archaea and Bacteria in the lake. Along with pH, major vent-associated geochemical constituents including CH4, CO2, H2, DIC (dissolved inorganic carbon), DOC (dissolved organic carbon), SO42-, O2 and metals were likely the major drivers for microbial population structures, however, mixing events occurring in the lake also impacted the distribution patterns. Distinct Bacteria and Archaea were present among size fractions, and bigger size fractions included particle-associated microbes (> 3 μm) and contained higher predicted operational taxonomic unit richness and microbial diversities (genus level) than free-living ones (<0.8 μm). Our study represents the first attempt at addressing the spatial distribution of Bacteria and Archaea in Yellowstone Lake, and our results highlight the variable contribution of Archaea and Bacteria to the hydrogeochemical-relevant metabolism of hydrogen, carbon, nitrogen, and sulfur.