Sulfur and methane oxidation by a single microorganism.

Natural and anthropogenic wetlands are major sources of the atmospheric greenhouse gas methane. Methane emissions from wetlands are mitigated by methanotrophic bacteria at the oxic-anoxic interface, a zone of intense redox cycling of carbon, sulfur, and nitrogen compounds. Here, we report on the isolation of an aerobic methanotrophic bacterium, 'Methylovirgula thiovorans' strain HY1, which possesses metabolic capabilities never before found in any methanotroph. Most notably, strain HY1 is the first bacterium shown to aerobically oxidize both methane and reduced sulfur compounds for growth. Genomic and proteomic analyses showed that soluble methane monooxygenase and XoxF-type alcohol dehydrogenases are responsible for methane and methanol oxidation, respectively. Various pathways for respiratory sulfur oxidation were present, including the Sox-rDsr pathway and the S4I system. Strain HY1 employed the Calvin-Benson-Bassham cycle for CO2 fixation during chemolithoautotrophic growth on reduced sulfur compounds. Proteomic and microrespirometry analyses showed that the metabolic pathways for methane and thiosulfate oxidation were induced in the presence of the respective substrates. Methane and thiosulfate could therefore be independently or simultaneously oxidized. The discovery of this versatile bacterium demonstrates that methanotrophy and thiotrophy are compatible in a single microorganism and underpins the intimate interactions of methane and sulfur cycles in oxic-anoxic interface environments.

Seasonal Dynamics of Methanotrophic Bacteria in a Boreal Oil Sands End Pit Lake.

Base Mine Lake (BML) is the first full-scale demonstration end pit lake for the oil sands mining industry in Canada. We examined aerobic methanotrophic bacteria over all seasons for 5 years in this dimictic lake. Methanotrophs comprised up to 58% of all bacterial reads in 16S rRNA gene amplicon sequencing analyses (median 2.8%), and up to 2.7 × 104 cells mL-1 of water (median 0.5 × 103) based on qPCR of pmoA genes. Methanotrophic activity and populations in the lake water were highest during fall turnover and remained high through the winter ice-covered period into spring turnover. They declined during summer stratification, especially in the epilimnion. Three methanotroph genera (Methylobacter, Methylovulum, and Methyloparacoccus) cycled seasonally, based on both relative and absolute abundance measurements. Methylobacter and Methylovulum populations peaked in winter/spring, when methane oxidation activity was psychrophilic. Methyloparacoccus populations increased in the water column through summer and fall, when methane oxidation was mesophilic, and also predominated in the underlying tailings sediment. Other, less abundant genera grew primarily during summer, possibly due to distinct CH4/O2 microniches created during thermal stratification. These data are consistent with temporal and spatial niche differentiation based on temperature, CH4 and O2. This pit lake displays methane cycling and methanotroph population dynamics similar to natural boreal lakes. IMPORTANCE The study examined methanotrophic bacteria in an industrial end pit lake, combining molecular DNA methods (both quantitative and descriptive) with biogeochemical measurements. The lake was sampled over 5 years, in all four seasons, as often as weekly, and included sub-ice samples. The resulting multiseason and multiyear data set is unique in its size and intensity, and allowed us to document clear and consistent seasonal patterns of growth and decline of three methanotroph genera (Methylobacter, Methylovulum, and Methyloparacoccus). Laboratory experiments suggested that one major control of this succession was niche partitioning based on temperature. The study helps to understand microbial dynamics in engineered end pit lakes, but we propose that the dynamics are typical of boreal stratified lakes and widely applicable in microbial ecology and limnology. Methane-oxidizing bacteria are important model organisms in microbial ecology and have implications for global climate change.

Ecological and genomic analyses of candidate phylum WPS-2 bacteria in an unvegetated soil.

Members of the bacterial candidate phylum WPS-2 (or Eremiobacterota) are abundant in several dry, bare soil environments. In a bare soil deposited by an extinct iron-sulfur spring, we found that WPS-2 comprised up to 24% of the bacterial community and up to 108 cells per g of soil based on 16S rRNA gene sequencing and quantification. A single genus-level cluster (Ca. Rubrimentiphilum) predominated in bare soils but was less abundant in adjacent forest. Nearly complete genomes of Ca. Rubrimentiphilum were recovered as single amplified genomes (SAGs) and metagenome-assembled genomes (MAGs). Surprisingly, given the abundance of WPS-2 in bare soils, the genomes did not indicate any capacity for autotrophy, phototrophy, or trace gas metabolism. Instead, they suggest a predominantly aerobic organoheterotrophic lifestyle, perhaps based on scavenging amino acids, nucleotides, and complex oligopeptides, along with lithotrophic capacity on thiosulfate. Network analyses of the entire community showed that some species of Chloroflexi, Actinobacteria, and candidate phylum AD3 (or Dormibacterota) co-occurred with Ca. Rubrimentiphilum and may represent ecological or metabolic partners. We propose that Ca. Rubrimentiphilum act as efficient heterotrophic scavengers. Combined with previous studies, these data suggest that the phylum WPS-2 includes bacteria with diverse metabolic capabilities.

Methylicorpusculum oleiharenae gen. nov., sp. nov., an aerobic methanotroph isolated from an oil sands tailings pond.

An aerobic methane oxidizing bacterium, designated XLMV4T, was isolated from the oxic surface layer of an oil sands tailings pond in Alberta, Canada. Strain XLMV4T is capable of growth on methane and methanol as energy sources. NH4Cl and sodium nitrate are nitrogen sources. Cells are Gram-negative, beige to yellow-pigmented, motile (via a single polar flagellum), short rods 2.0-3.3 µm in length and 1.0-1.6 µm in width. A thick capsule is produced. Surface glycoprotein or cup shape proteins typical of the genera Methylococcus, Methylothermus and Methylomicrobium were not observed. Major isoprenoid quinones are Q-8 and Q-7 at an approximate molar ratio of 71 : 22. Major polar lipids are phosphoglycerol and ornithine lipids. Major fatty acids are C16 : 1 ω8+C16 : 1 ω7 (34 %), C16 : 1 ω5 (16 %), and C18 : 1 ω7 (11 %). Optimum growth is observed at pH 8.0 and 25 °C. The DNA G+C content based on a draft genome sequence is 46.7 mol%. Phylogenetic analysis of 16S rRNA genes and a larger set of conserved genes place strain XLMV4T within the class Gammaproteobacteria and family Methylococcaceae, most closely related to members of the genera Methylomicrobium and Methylobacter (95.0-97.1 % 16S rRNA gene sequence identity). In silico genomic predictions of DNA-DNA hybridization values of strain XLMV4T to the nearest phylogenetic neighbours were all below 26 %. On the basis of the data presented, strain XLMV4T is considered to represent a new genus and species for which the name Methylicorpusculum oleiharenae is proposed. Strain XLMV4T (=DSMZ DSM 27269=ATCC TSD-186) is the type strain.

Phylogenetic Estimation of Community Composition and Novel Eukaryotic Lineages in Base Mine Lake: An Oil Sands Tailings Reclamation Site in Northern Alberta.

Reclamation of anthropogenically impacted environments is a critical issue worldwide. In the oil sands extraction industry of Alberta, reclamation of mining-impacted areas, especially areas affected by tailings waste, is an important aspect of the mining life cycle. A reclamation technique currently under study is water-capping, where tailings are capped by water to create an end-pit lake (EPL). Base Mine Lake (BML) is the first full-scale end-pit lake in the Alberta oil sands region. In this study, we sequenced eukaryotic 18S rRNA genes recovered from 92 samples of Base Mine Lake water in a comprehensive sampling programme covering the ice-free period of 2015. The 565 operational taxonomic units (OTUs) generated revealed a dynamic and diverse community including abundant Microsporidia, Ciliata and Cercozoa, though 41% of OTUs were not classifiable below the phylum level by comparison to 18S rRNA databases. Phylogenetic analysis of five heterotrophic phyla (Cercozoa, Fungi, Ciliata, Amoebozoa and Excavata) revealed substantial novel diversity, with many clusters of OTUs that were more similar to each other than to any reference sequence. All of these groups are entirely or mostly heterotrophic, as a relatively small number of definitively photosynthetic clades were amplified from the BML samples.

Oleiharenicola alkalitolerans gen. nov., sp. nov., a new member of the phylum Verrucomicrobia isolated from an oilsands tailings pond.

A novel member of the phylum Verrucomicrobia was isolated from an oilsands tailings pond in Alberta, Canada. Cells of isolate NVTT are Gram-negative, strictly aerobic, non-pigmented, non-motile cocci to diplococci 0.5-1.0 µm in diameter. The bacterium is neutrophilic (optimum pH 6.0-8.0) but alkalitolerant, capable of growth between pH 5.5 and 11.0. The temperature range for growth is 15-40 °C (optimum 25-37 °C). Carbon and energy sources include sugars and organic acids. Nitrogen sources include nitrate, urea, l-glycine, l-alanine, l-proline and l-serine. Does not fix atmospheric nitrogen. Does not require NaCl and is inhibited at NaCl concentrations above 3.0 % (w/v). The DNA G+C content of strain NVTT, based on a draft genome sequence, is 66.1 mol%. MK-6 and MK-7 are the major respiratory quinones. Major cellular fatty acids are anteiso-C15 : 0 and iso-C15 : 0. Phylogenetic analysis of 16S rRNA gene sequences revealed that the strain belongs to the family Opitutaceae of the phylum Verrucomicrobia. The most closely related validated species is Opitutus terrae (93.7 % 16S rRNA gene sequence identity to its type strain PB90-1T). Based on genotypic, phenotypic and chemotaxonomic characteristics, it was concluded that this strain represents a novel genus and species, for which the name Oleiharenicola alkalitolerans gen. nov., sp. nov. is proposed. The type strain of this novel species is NVTT (=ATCC BAA-2697T;=DSM 29249T).

Actinocrinis puniceicyclus gen. nov., sp. nov., an actinobacterium isolated from an acidic spring.

An aerobic, mildly acidophilic actinobacterium was isolated from the Ochre Beds bog in Kootenay National Park, Canada. Cells of isolate OB1T were Gram-stain-positive, non-motile, pink- to purple-pigmented filaments. The pH range for growth was pH 3.5-6.5 (optimum pH 5.5), and the temperature range was 13-30°C. The major cellular fatty acids were i-C16 : 0 (28.5 %), i-C15 : 0 (14.6 %) and ai-C15 : 0 (14.3 %), and the major polar lipid was phosphohexose. The major quinone was menaquinone-11 (MK-11), and the peptidoglycan type was A1γ. The DNA G+C content was 70.2 %. Along with growth on complex media including yeast extract, proteose peptone, casamino acids and tryptic soy broth, growth occured on mono- and disaccharides (glucose, sucrose, galactose and xylose) and polysaccharides (starch, gellan, pectin, xylan and alginate). Anaerobic growth was not observed. The cells did not fix atmospheric nitrogen. On the basis of comparative 16S rRNA gene sequence analysis, this isolate belonged to the family Actinospicaceae, in the suborder Catenulisporineae of the order Actinomycetales. The most closely related species was Actinospica robiniae. However, the 16S rRNA gene sequence identity to this bacterium was low (92.8 %) and there were several chemotaxonomic differences from this species. We therefore propose a novel genus and species, Actinocrinis puniceicyclus gen. nov., sp. nov., with strain OB1T (=DSM 45618T=ATCC BAA-2771T) as the type strain.

The Chthonomonas calidirosea Genome Is Highly Conserved across Geographic Locations and Distinct Chemical and Microbial Environments in New Zealand's Taupo Volcanic Zone.

UNLABELLED: Chthonomonas calidirosea T49(T) is a low-abundance, carbohydrate-scavenging, and thermophilic soil bacterium with a seemingly disorganized genome. We hypothesized that the C. calidirosea genome would be highly responsive to local selection pressure, resulting in the divergence of its genomic content, genome organization, and carbohydrate utilization phenotype across environments. We tested this hypothesis by sequencing the genomes of four C. calidirosea isolates obtained from four separate geothermal fields in the Taupo Volcanic Zone, New Zealand. For each isolation site, we measured physicochemical attributes and defined the associated microbial community by 16S rRNA gene sequencing. Despite their ecological and geographical isolation, the genome sequences showed low divergence (maximum, 1.17%). Isolate-specific variations included single-nucleotide polymorphisms (SNPs), restriction-modification systems, and mobile elements but few major deletions and no major rearrangements. The 50-fold variation in C. calidirosea relative abundance among the four sites correlated with site environmental characteristics but not with differences in genomic content. Conversely, the carbohydrate utilization profiles of the C. calidirosea isolates corresponded to the inferred isolate phylogenies, which only partially paralleled the geographical relationships among the sample sites. Genomic sequence conservation does not entirely parallel geographic distance, suggesting that stochastic dispersal and localized extinction, which allow for rapid population homogenization with little restriction by geographical barriers, are possible mechanisms of C. calidirosea distribution. This dispersal and extinction mechanism is likely not limited to C. calidirosea but may shape the populations and genomes of many other low-abundance free-living taxa. IMPORTANCE: This study compares the genomic sequence variations and metabolisms of four strains of Chthonomonas calidirosea, a rare thermophilic bacterium from the phylum Armatimonadetes It additionally compares the microbial communities and chemistry of each of the geographically distinct sites from which the four C. calidirosea strains were isolated. C. calidirosea was previously reported to possess a highly disorganized genome, but it was unclear whether this reflected rapid evolution. Here, we show that each isolation site has a distinct chemistry and microbial community, but despite this, the C. calidirosea genome is highly conserved across all isolation sites. Furthermore, genomic sequence differences only partially paralleled geographic distance, suggesting that C. calidirosea genotypes are not primarily determined by adaptive evolution. Instead, the presence of C. calidirosea may be driven by stochastic dispersal and localized extinction. This ecological mechanism may apply to many other low-abundance taxa.

Emended description of the family Beijerinckiaceae and transfer of the genera Chelatococcus and Camelimonas to the family Chelatococcaceae fam. nov.

The family Beijerinckiaceae was circumscribed in 2005 to accommodate four genera of phylogenetically related alphaproteobacteria: Beijerinckia, Chelatococcus, Methylocella and Methylocapsa. Later, four additional genera, i.e. Methylovirgula, Methyloferula, Methylorosula and Camelimonas, were described and assigned to this family, which now accommodates 21 species with validly published names. Members of this family possess strikingly different lifestyles, including chemoheterotrophy, facultative methylotrophy, obligate methanotrophy and facultative methanotrophy. Levels of 16S rRNA gene sequence similarity among most of these bacteria range from 96 to 98 %, suggesting a common evolutionary origin. The genera Chelatococcus and Camelimonas, however, are not monophyletic with the other described genera based on 16S rRNA gene sequence phylogeny, and instead form a distant cluster more closely related to the Methylobacteriaceae. Physiologically these two genera also lack several properties common to the other Beijerinckiaceae. On the other hand, the genus Rhodoblastus, presently considered a member of the Bradyrhizobiaceae, affiliates with high confidence to the Beijerinckiaceae. Here, we propose to transfer the genera Chelatococcus and Camelimonas to the family Chelatococcaceae fam. nov., and present an emended description of the family Beijerinckiaceae, including the genus Rhodoblastus.

Next-Generation Sequencing Assessment of Eukaryotic Diversity in Oil Sands Tailings Ponds Sediments and Surface Water.

Tailings ponds in the Athabasca oil sands (Canada) contain fluid wastes, generated by the extraction of bitumen from oil sands ores. Although the autochthonous prokaryotic communities have been relatively well characterized, almost nothing is known about microbial eukaryotes living in the anoxic soft sediments of tailings ponds or in the thin oxic layer of water that covers them. We carried out the first next-generation sequencing study of microbial eukaryotic diversity in oil sands tailings ponds. In metagenomes prepared from tailings sediment and surface water, we detected very low numbers of sequences encoding eukaryotic small subunit ribosomal RNA representing seven major taxonomic groups of protists. We also produced and analysed three amplicon-based 18S rRNA libraries prepared from sediment samples. These revealed a more diverse set of taxa, 169 different OTUs encompassing up to eleven higher order groups of eukaryotes, according to detailed classification using homology searching and phylogenetic methods. The 10 most abundant OTUs accounted for > 90% of the total of reads, vs. large numbers of rare OTUs (< 1% abundance). Despite the anoxic and hydrocarbon-enriched nature of the environment, the tailings ponds harbour complex communities of microbial eukaryotes indicating that these organisms should be taken into account when studying the microbiology of the oil sands.

Stable-Isotope Probing Identifies Uncultured Planctomycetes as Primary Degraders of a Complex Heteropolysaccharide in Soil.

The exopolysaccharides (EPSs) produced by some bacteria are potential growth substrates for other bacteria in soil. We used stable-isotope probing (SIP) to identify aerobic soil bacteria that assimilated the cellulose produced by Gluconacetobacter xylinus or the EPS produced by Beijerinckia indica. The latter is a heteropolysaccharide comprised primarily of l-guluronic acid, d-glucose, and d-glycero-d-mannoheptose. (13)C-labeled EPS and (13)C-labeled cellulose were purified from bacterial cultures grown on [(13)C]glucose. Two soils were incubated with these substrates, and bacteria actively assimilating them were identified via pyrosequencing of 16S rRNA genes recovered from (13)C-labeled DNA. Cellulose C was assimilated primarily by soil bacteria closely related (93 to 100% 16S rRNA gene sequence identities) to known cellulose-degrading bacteria. However, B. indica EPS was assimilated primarily by bacteria with low identities (80 to 95%) to known species, particularly by different members of the phylum Planctomycetes. In one incubation, members of the Planctomycetes made up >60% of all reads in the labeled DNA and were only distantly related (<85% identity) to any described species. Although it is impossible with SIP to completely distinguish primary polysaccharide hydrolyzers from bacteria growing on produced oligo- or monosaccharides, the predominance of Planctomycetes suggested that they were primary degraders of EPS. Other bacteria assimilating B. indica EPS included members of the Verrucomicrobia, candidate division OD1, and the Armatimonadetes. The results indicate that some uncultured bacteria in soils may be adapted to using complex heteropolysaccharides for growth and suggest that the use of these substrates may provide a means for culturing new species.

Distribution and diversity of Verrucomicrobia methanotrophs in geothermal and acidic environments.

Recently, methanotrophic members of the phylum Verrucomicrobia have been described, but little is known about their distribution in nature. We surveyed methanotrophic bacteria in geothermal springs and acidic wetlands via pyrosequencing of 16S rRNA gene amplicons. Putative methanotrophic Verrucomicrobia were found in samples covering a broad temperature range (22.5-81.6°C), but only in acidic conditions (pH 1.8-5.0) and only in geothermal environments, not in acidic bogs or fens. Phylogenetically, three 16S rRNA gene sequence clusters of putative methanotrophic Verrucomicrobia were observed. Those detected in high-temperature geothermal samples (44.1-81.6°C) grouped with known thermoacidiphilic 'Methylacidiphilum' isolates. A second group dominated in moderate-temperature geothermal samples (22.5-40.1°C) and a representative mesophilic methanotroph from this group was isolated (strain LP2A). Genome sequencing verified that strain LP2A possessed particulate methane monooxygenase, but its 16S rRNA gene sequence identity to 'Methylacidiphilum infernorum' strain V4 was only 90.6%. A third group clustered distantly with known methanotrophic Verrucomicrobia. Using pmoA-gene targeted quantitative polymerase chain reaction, two geothermal soil profiles showed a dominance of LP2A-like pmoA sequences in the cooler surface layers and 'Methylacidiphilum'-like pmoA sequences in deeper, hotter layers. Based on these results, there appears to be a thermophilic group and a mesophilic group of methanotrophic Verrucomicrobia. However, both were detected only in acidic geothermal environments.

Ether- and ester-bound iso-diabolic acid and other lipids in members of acidobacteria subdivision 4.

Recently, iso-diabolic acid (13,16-dimethyl octacosanedioic acid) has been identified as a major membrane-spanning lipid of subdivisions 1 and 3 of the Acidobacteria, a highly diverse phylum within the Bacteria. This finding pointed to the Acidobacteria as a potential source for the bacterial glycerol dialkyl glycerol tetraethers that occur ubiquitously in peat, soil, lakes, and hot springs. Here, we examined the lipid composition of seven phylogenetically divergent strains of subdivision 4 of the Acidobacteria, a bacterial group that is commonly encountered in soil. Acid hydrolysis of total cell material released iso-diabolic acid derivatives in substantial quantities (11 to 48% of all fatty acids). In contrast to subdivisions 1 and 3 of the Acidobacteria, 6 out of the 7 species of subdivision 4 (excepting "Candidatus Chloracidobacterium thermophilum") contained iso-diabolic acid ether bound to a glycerol in larger fractional abundance than iso-diabolic acid itself. This is in agreement with the analysis of intact polar lipids (IPLs) by high-performance liquid chromatography-mass spectrometry (HPLC-MS), which showed the dominance of mixed ether-ester glycerides. iso-Diabolic acid-containing IPLs were not identified, because these IPLs are not released with a Bligh-Dyer extraction, as observed before when studying lipid compositions of subdivisions 1 and 3 of the Acidobacteria. The presence of ether bonds in the membrane lipids does not seem to be an adaptation to temperature, because the five mesophilic isolates contained a larger amount of ether lipids than the thermophile "Ca. Chloracidobacterium thermophilum." Furthermore, experiments with Pyrinomonas methylaliphatogenes did not reveal a major influence of growth temperature over the 50 to 69°C range.

Thermoflavifilum aggregans gen. nov., sp. nov., a thermophilic and slightly halophilic filamentous bacterium from the phylum Bacteroidetes.

A strictly aerobic, thermophilic, moderately acidophilic, non-spore-forming bacterium, strain P373(T), was isolated from geothermally heated soil at Waikite, New Zealand. Cells were filamentous rods, 0.2-0.4 µm in diameter and grew in chains up to 80 µm in length. On the basis of 16S rRNA gene sequence similarity, strain P373(T) was shown to belong to the family Chitinophagaceae (class Sphingobacteriia) of the phylum Bacteroidetes, with the most closely related cultivated strain, Chitinophaga pinensis UQM 2034(T), having 87.6 % sequence similarity. Cells stained Gram-negative, and were catalase- and oxidase-positive. The major fatty acids were i-15 : 0 (10.8 %), i-17 : 0 (24.5 %) and i-17 : 0 3-OH (35.2 %). Primary lipids were phosphatidylethanolamine, two unidentified aminolipids and three other unidentified polar lipids. The presence of sulfonolipids (N-acyl-capnines) was observed in the total lipid extract by mass spectrometry. The G+C content of the genomic DNA was 47.3 mol% and the primary respiratory quinone was MK-7. Strain P373(T) grew at 35-63 °C with an optimum temperature of 60 °C, and at pH 5.5-8.7 with an optimum growth pH of 7.3-7.4. NaCl tolerance was up to 5 % (w/v) with an optimum of 0.1-0.25 % (w/v). Cell colonies were non-translucent and pigmented vivid yellow-orange. Cells displayed an oxidative chemoheterotrophic metabolism. The distinct phylogenetic position and the phenotypic characteristics separate strain P373(T) from all other members of the phylum Bacteroidetes and indicate that it represents a novel species in a new genus, for which the name Thermoflavifilum aggregans gen. nov., sp. nov. is proposed. The type strain of the type species is P373(T) ( = ICMP 20041(T) = DSM 27268(T)).

Phytoplankton ecology in the early years of a boreal oil sands end pit lake.

BACKGROUND: Base Mine Lake (BML) is the first full-scale end pit lake for the oil sands mining industry in Canada. BML sequesters oil sands tailings under a freshwater cap and is intended to develop into a functional ecosystem that can be integrated into the local watershed. The first stage of successful reclamation requires the development of a phytoplankton community supporting a typical boreal lake food web. To assess the diversity and dynamics of the phytoplankton community in BML at this reclamation stage and to set a baseline for future monitoring, we examined the phytoplankton community in BML from 2016 through 2021 using molecular methods (targeting the 23S, 18S, and 16S rRNA genes) and microscopic methods. Nearby water bodies were used as controls for a freshwater environment and an active tailings pond. RESULTS: The phytoplankton community was made up of diverse bacteria and eukaryotes typical of a boreal lake. Microscopy and molecular data both identified a phytoplankton community comparable at the phylum level to that of natural boreal lakes, dominated by Chlorophyta, Cryptophyta, and Cyanophyta, with some Bacillariophyta, Ochrophyta, and Euglenophyta. Although many of the same genera were prominent in both BML and the control freshwater reservoir, there were differences at the species or ASV level. Total diversity in BML was also consistently lower than the control freshwater site, but consistently higher than the control tailings pond. The phytoplankton community composition in BML changed over the 5-year study period. Some taxa present in 2016-2019 (e.g., Choricystis) were no longer detected in 2021, while some dinophytes and haptophytes became detectable in small quantities starting in 2019-2021. Different quantification methods (qPCR analysis of 23S rRNA genes, and microscopic estimates of populations and total biomass) did not show a consistent directional trend in total phytoplankton over the 5-year study, nor was there any consistent increase in phytoplankton species diversity. The 5-year period was likely an insufficient time frame for detecting community trends, as phytoplankton communities are highly variable at the genus and species level. CONCLUSIONS: BML supports a phytoplankton community composition somewhat unique from control sites (active tailings and freshwater lake) and is still changing over time. However, the most abundant genera are typical of natural boreal lakes and have the potential to support a complex aquatic food web, with many of its identified major phytoplankton constituents known to be primary producers in boreal lake environments.

Nitrous oxide respiration in acidophilic methanotrophs.

Aerobic methanotrophic bacteria are considered strict aerobes but are often highly abundant in hypoxic and even anoxic environments. Despite possessing denitrification genes, it remains to be verified whether denitrification contributes to their growth. Here, we show that acidophilic methanotrophs can respire nitrous oxide (N2O) and grow anaerobically on diverse non-methane substrates, including methanol, C-C substrates, and hydrogen. We study two strains that possess N2O reductase genes: Methylocella tundrae T4 and Methylacidiphilum caldifontis IT6. We show that N2O respiration supports growth of Methylacidiphilum caldifontis at an extremely acidic pH of 2.0, exceeding the known physiological pH limits for microbial N2O consumption. Methylocella tundrae simultaneously consumes N2O and CH4 in suboxic conditions, indicating robustness of its N2O reductase activity in the presence of O2. Furthermore, in O2-limiting conditions, the amount of CH4 oxidized per O2 reduced increases when N2O is added, indicating that Methylocella tundrae can direct more O2 towards methane monooxygenase. Thus, our results demonstrate that some methanotrophs can respire N2O independently or simultaneously with O2, which may facilitate their growth and survival in dynamic environments. Such metabolic capability enables these bacteria to simultaneously reduce the release of the key greenhouse gases CO2, CH4, and N2O.

Chryseolinea serpens gen. nov., sp. nov., a member of the phylum Bacteroidetes isolated from soil.

An aerobic chemoheterotrophic gliding bacterium, designated RYG(T), was isolated from a soil in Germany. Cells were Gram-stain-negative, thin rods (0.4-0.6 µm in width and 2.0-5.5 µm in length). Cells multiplied by normal cell division and no resting stages were observed. Colonies were yellow and displayed swarming edges. Gliding motility was observed in wet mounts. Strain RYG(T) grew at pH 5.6-7.7 (optimum pH 6.6-7.0), at 13-37 °C (optimum 25-30 °C) and with 0-1.0 % NaCl (optimum 0-0.1 %). The isolate was incapable of atmospheric nitrogen fixation and grew on most mono- and disaccharides as well as a few polysaccharides and organic acids. The predominant menaquinone was MK-7, the major cellular fatty acids were C(16 : 1)ω5c and iso-C(15 : 0) and the major intact polar lipids were composed of phosphatidylethanolamine derivatives and two unknown series. The DNA G+C content was 49.9 mol%. Based on 16S rRNA gene sequence analysis, the isolate belonged to the phylum Bacteroidetes, class Cytophagia, order Cytophagales, but was only distantly related to any cultured bacteria. The closest relatives were Ohtaekwangia koreensis 3B-2(T) and Ohtaekwangia kribbensis 10AO(T) (both 93 % 16S rRNA gene sequence similarity). We propose a novel genus and species, Chryseolinea serpens gen. nov., sp. nov.. Strain RYG(T) ( = DSM 24574(T) = ATCC BAA-2075(T)) is the type strain.

Metagenomics of hydrocarbon resource environments indicates aerobic taxa and genes to be unexpectedly common.

Oil in subsurface reservoirs is biodegraded by resident microbial communities. Water-mediated, anaerobic conversion of hydrocarbons to methane and CO2, catalyzed by syntrophic bacteria and methanogenic archaea, is thought to be one of the dominant processes. We compared 160 microbial community compositions in ten hydrocarbon resource environments (HREs) and sequenced twelve metagenomes to characterize their metabolic potential. Although anaerobic communities were common, cores from oil sands and coal beds had unexpectedly high proportions of aerobic hydrocarbon-degrading bacteria. Likewise, most metagenomes had high proportions of genes for enzymes involved in aerobic hydrocarbon metabolism. Hence, although HREs may have been strictly anaerobic and typically methanogenic for much of their history, this may not hold today for coal beds and for the Alberta oil sands, one of the largest remaining oil reservoirs in the world. This finding may influence strategies to recover energy or chemicals from these HREs by in situ microbial processes.

Methane oxidation by an extremely acidophilic bacterium of the phylum Verrucomicrobia.

Aerobic methanotrophic bacteria consume methane as it diffuses away from methanogenic zones of soil and sediment. They act as a biofilter to reduce methane emissions to the atmosphere, and they are therefore targets in strategies to combat global climate change. No cultured methanotroph grows optimally below pH 5, but some environments with active methane cycles are very acidic. Here we describe an extremely acidophilic methanotroph that grows optimally at pH 2.0-2.5. Unlike the known methanotrophs, it does not belong to the phylum Proteobacteria but rather to the Verrucomicrobia, a widespread and diverse bacterial phylum that primarily comprises uncultivated species with unknown genotypes. Analysis of its draft genome detected genes encoding particulate methane monooxygenase that were homologous to genes found in methanotrophic proteobacteria. However, known genetic modules for methanol and formaldehyde oxidation were incomplete or missing, suggesting that the bacterium uses some novel methylotrophic pathways. Phylogenetic analysis of its three pmoA genes (encoding a subunit of particulate methane monooxygenase) placed them into a distinct cluster from proteobacterial homologues. This indicates an ancient divergence of Verrucomicrobia and Proteobacteria methanotrophs rather than a recent horizontal gene transfer of methanotrophic ability. The findings show that methanotrophy in the Bacteria is more taxonomically, ecologically and genetically diverse than previously thought, and that previous studies have failed to assess the full diversity of methanotrophs in acidic environments.

Genome sequence of the haloalkaliphilic methanotrophic bacterium Methylomicrobium alcaliphilum 20Z.

Methylomicrobium strains are widespread in saline environments. Here, we report the complete genome sequence of Methylomicrobium alcaliphilum 20Z, a haloalkaliphilic methanotrophic bacterium, which will provide the basis for detailed characterization of the core pathways of both single-carbon metabolism and responses to osmotic and high-pH stresses. Final assembly of the genome sequence revealed that this bacterium contains a 128-kb plasmid, making M. alcaliphilum 20Z the first methanotrophic bacterium of known genome sequence for which a plasmid has been reported.