Methylocystis Parvus Research Articles

Assessing the performance of synthetic co-cultures during the conversion of methane into Poly(3-hydroxybutyrate)

Synthetic co-cultures can enhance pollutant bioconversion performance through synergistic effects among co-existing species. In this study, the potential of Methylocystis hirsuta and Methylocystis parvus to support poly(3-hydroxybutyrate) (PHB) production in co-cultivation with Rhodococcus opacus and Pseudomonas putida under a CH4:O2 atmosphere was assessed batchwise. The metabolic activation of almost all co-cultures studied was faster than that of single strain cultures, bringing higher methane and oxygen consumption rates. Higher PHB yields were achieved when coupling M. hirsuta with R. opacus (63 % w w−1) or with R. opacus and P. putida (64.4 % w w−1) compared to M. hirsuta alone (38.5 % w w−1). Interestingly, the combination of both R. opacus and P. putida with M. parvus reduced PHB accumulation to 42.2 % w w−1 compared to the content observed in M. parvus monocultures (62.2 % w w−1) and M parvus + R. opacus co-cultures (66.6 % w w−1).

Comparative genomic analysis of Methylocystis sp. MJC1 as a platform strain for polyhydroxybutyrate biosynthesis.

Biodegradable polyhydroxybutyrate (PHB) can be produced from methane by some type II methanotroph such as the genus Methylocystis. This study presents the comparative genomic analysis of a newly isolated methanotroph, Methylocystis sp. MJC1 as a biodegradable PHB-producing platform strain. Methylocystis sp. MJC1 accumulates up to 44.5% of PHB based on dry cell weight under nitrogen-limiting conditions. To facilitate its development as a PHB-producing platform strain, the complete genome sequence of Methylocystis sp. MJC1 was assembled, functionally annotated, and compared with genomes of other Methylocystis species. Phylogenetic analysis has shown that Methylocystis parvus to be the closest species to Methylocystis sp. MJC1. Genome functional annotation revealed that Methylocystis sp. MJC1 contains all major type II methanotroph biochemical pathways such as the serine cycle, EMC pathway, and Krebs cycle. Interestingly, Methylocystis sp. MJC1 has both particulate and soluble methane monooxygenases, which are not commonly found among Methylocystis species. In addition, this species also possesses most of the RuMP pathway reactions, a characteristic of type I methanotrophs, and all PHB biosynthetic genes. These comparative analysis would open the possibility of future practical applications such as the development of organism-specific genome-scale models and application of metabolic engineering strategies to Methylocystis sp. MJC1.

In Vivo Genome Editing in Type I and II Methanotrophs Using a CRISPR/Cas9 System.

Methanotrophic bacteria are Gram-negative, aerobic organisms that use methane as their sole source of carbon and energy. In this study, we constructed and exemplified a CRISPR/Cas9 genome editing system and used it to successfully make gene deletions and insertions in the type I methanotroph Methylococcus capsulatus Bath and the type II methanotroph Methylocystis parvus OBBP. High frequencies of gene deletions and insertions were achieved in combination with homology-directed repair. In M. parvus OBBP, we also investigated the impact of several parameters on the CRISPR/Cas9 genome editing, where the ligD gene was targeted with various PAM sequences and guide RNA spacer sequences, homology arms of variable length, differences in the duration of mating during conjugation, and exploiting promoters of different strengths to control the expression of cas9 and sgRNA. Although not the first attempt to develop a CRISPR/Cas system in methanotrophs, this work demonstrated for the first time an efficient CRISPR/Cas9 system generating scarless clean gene deletions and insertions in methanotroph genomes.

Sustainable Process for the Production of Poly(3-hydroxybutyrate-co-3-hydroxyvalerate) from Renewable Resources: A Simulation Study.

Bacterially produced polyhydroxyalkanoates are valuable substitutes for petrochemical plastics, but their current production capacities are very scarce. Producing poly(3-hydroxybutyrate-co-3-hydroxyvalerate) (PHB-co-HV) from methane and odd-chain carbon fatty acids could make the production of this biodegradable polymer cost-effective. This study analyzes the main factors affecting methanotrophic growth and poly(3-hydroxybutyrate-co-3-hydroxyvalerate) accumulation, simulating a pilot-scale process based on a double-stage approach. The effects of the nitrogen source and the oxygen partial pressure during a 20 day growth phase were studied; the cosubstrate concentration, the culture selected, and the methane partial pressure were investigated during the poly(3-hydroxybutyrate-co-3-hydroxyvalerate) production stage performed within 15 days under nutrient starvation. Methylocystis parvus OBBP and Methylosinus thricosporum OB3b reached the maximum growth productivities with ammonium as the nitrogen source and oxygen at high partial pressure. The simulation of the PHB-co-HV accumulation revealed that methanotrophs could better accumulate the polymer with low valeric acid concentrations. A methane-abundant gas stream (0.5 atm of methane) could increase process yields up to 0.32 kg m-3 d-1.

Insights into the Genomic Potential of a Methylocystis sp. from Amazonian Floodplain Sediments.

Although floodplains are recognized as important sources of methane (CH4) in the Amazon basin, little is known about the role of methanotrophs in mitigating CH4 emissions in these ecosystems. Our previous data reported the genus Methylocystis as one of the most abundant methanotrophs in these floodplain sediments. However, information on the functional potential and life strategies of these organisms living under seasonal flooding is still missing. Here, we described the first metagenome-assembled genome (MAG) of a Methylocystis sp. recovered from Amazonian floodplains sediments, and we explored its functional potential and ecological traits through phylogenomic, functional annotation, and pan-genomic approaches. Both phylogenomics and pan-genomics identified the closest placement of the bin.170_fp as Methylocystis parvus. As expected for Type II methanotrophs, the Core cluster from the pan-genome comprised genes for CH4 oxidation and formaldehyde assimilation through the serine pathway. Furthermore, the complete set of genes related to nitrogen fixation is also present in the Core. Interestingly, the MAG singleton cluster revealed the presence of unique genes related to nitrogen metabolism and cell motility. The study sheds light on the genomic characteristics of a dominant, but as yet unexplored methanotroph from the Amazonian floodplains. By exploring the genomic potential related to resource utilization and motility capability, we expanded our knowledge on the niche breadth of these dominant methanotrophs in the Amazonian floodplains.

RNA Biomarker Trends across Type I and Type II Aerobic Methanotrophs in Response to Methane Oxidation Rates and Transcriptome Response to Short-Term Methane and Oxygen Limitation in Methylomicrobium album BG8.

ABSTRACTMethanotrophs, which help regulate atmospheric levels of methane, are active in diverse natural and man-made environments. This range of habitats and the feast-famine cycles seen by many environmental methanotrophs suggest that methanotrophs dynamically mediate rates of methane oxidation. Global methane budgets require ways to account for this variability in time and space. Functional gene biomarker transcripts are increasingly studied to inform the dynamics of diverse biogeochemical cycles. Previously, per-cell transcript levels of the methane oxidation biomarker pmoA were found to vary quantitatively with respect to methane oxidation rates in the model aerobic methanotroph Methylosinus trichosporium OB3b. In the present study, these trends were explored for two additional aerobic methanotroph pure cultures grown in membrane bioreactors, Methylocystis parvus OBBP and Methylomicrobium album BG8. At steady-state conditions, per-cell pmoA mRNA transcript levels strongly correlated with per-cell methane oxidation across the three methanotrophs across many orders of magnitude of activity (R2 = 0.91). The inclusion of both type I and type II aerobic methanotrophs suggests a universal trend between in situ activity level and pmoA RNA biomarker levels which can aid in improving estimates of both subsurface and atmospheric methane. Additionally, genome-wide expression data (obtained by transcriptome sequencing [RNA-seq]) were used to explore transcriptomic responses of steady-state M. album BG8 cultures to short-term CH4 and O2 limitation. These limitations induced regulation of genes involved in central carbon metabolism (including carbon storage), cell motility, and stress response.IMPORTANCE Methanotrophs are naturally occurring microorganisms capable of oxidizing methane, having an impact on global net methane emissions. Additionally, they have also gained interest for their biotechnological applications in single-cell protein production, biofuels, and bioplastics. Having better ways of measuring methanotroph activity and understanding how methanotrophs respond to changing conditions is imperative for both optimization in controlled-growth applications and understanding in situ methane oxidation rates. In this study, we explored the applicability of methane oxidation biomarkers as a universal indicator of methanotrophic activity and explored methanotroph transcriptomic response to short-term changes in substrate availability. Our results contribute to better understanding the activity of aerobic methanotrophs, their core metabolic pathways, and their stress responses.

Production of (R)-3-hydroxybutyric acid from methane by in vivo depolymerization of polyhydroxybutyrate in Methylocystis parvus OBBP

Methylocystis parvus OBBP accumulates polyhydroxybutyrate (PHB) using methane as the sole carbon and energy source. In this work, the feasibility of producing (R)-3-hydroxybutyric acid (R3HBA) via intracellularly accumulated PHB through depolymerization (in-vivo) was investigated. Results showed that a PHB to R3HBA conversion of 77.2 ± 0.9% (R3HBA titer of 0.153 ± 0.002 g L-1) can be attained in a mineral medium containing 1 g L-1 KNO3 at 30 °C with shaking at 200 rpm and a constant pH of 11 for 72 h. Nitrogen deprivation and neutral or acidic pHs strongly reduced the excreted R3HBA concentration. Reduced oxygen availability negatively affected the R3HBA yield, which decreased to 73.6 ± 4.9% (titer of 0.139 ± 0.01 g L-1) under microaerobic conditions. Likewise, the presence of increasing concentrations of R3HBA in the medium before the onset of PHB depolymerization reduced the initial R3HBA release rate and R3HBA yield.

Optimization of nitrogen feeding strategies for improving polyhydroxybutyrate production from biogas by Methylocystis parvus str. OBBP in a stirred tank reactor

The design of efficient cultivation strategies to produce bioplastics from biogas is crucial for the implementation of this biorefinery process. In this work, biogas-based polyhydroxybutyrate (PHB) production and CH4 biodegradation performance was investigated for the first time in a stirred tank bioreactor inoculated with Methylocystis parvus str. OBBP. Decreasing nitrogen loading rates in continuous mode and alternating feast:famine regimes of 24 h-cycles, and alternating feast:famine regimes of 24 h:24 h and 24 h:48 h were tested. Continuous N feeding did not support an effective PHB production despite the occurrence of nitrogen limiting conditions. Feast-famine cycles of 24 h:24 h (with 50% stoichiometric nitrogen supply) supported the maximum PHB production (20 g-PHB m−3 d−1) without compromising the CH4-elimination capacity (25 g m−3 h−1) of the system. Feast:famine ratios ≤1:2 entailed the deterioration of process performance at stoichiometric nitrogen inputs ≤60%.

Isolation and characterisation of Methylocystis spp. for poly-3-hydroxybutyrate production using waste methane feedstocks

Waste plastic and methane emissions are two anthropogenic by-products exacerbating environmental pollution. Methane-oxidizing bacteria (methanotrophs) hold the key to solving these problems simultaneously by utilising otherwise wasted methane gas as carbon source and accumulating the carbon as poly-3-hydroxybutyrate, a biodegradable plastic polymer. Here we present the isolation and characterisation of two novel Methylocystis strains with the ability to produce up to 55.7 ± 1.9% poly-3-hydroxybutyrate of cell dry weight when grown on methane from different waste sources such as landfill and anaerobic digester gas. Methylocystis rosea BRCS1 isolated from a recreational lake and Methylocystis parvus BRCS2 isolated from a bog were whole genome sequenced using PacBio and Illumina genome sequencing technologies. In addition to potassium nitrate, these strains were also shown to grow on ammonium chloride, glutamine and ornithine as nitrogen source. Growth of Methylocystis parvus BRCS2 on Nitrate Mineral Salt (NMS) media with 0.1% methanol vapor as carbon source was demonstrated. The genetic tractability by conjugation was also determined with conjugation efficiencies up to 2.8 × 10–2 and 1.8 × 10–2 for Methylocystis rosea BRCS1 and Methylocystis parvus BRCS2 respectively using a plasmid with ColE1 origin of replication. Finally, we show that Methylocystis species can produce considerable amounts of poly-3-hydroxybutyrate on waste methane sources without impaired growth, a proof of concept which opens doors to their use in integrated bio-facilities like landfills and anaerobic digesters.

Enrichment of Organic Manure with Plant Growth Promoting Rhizobacteria Improved the Root and Shoot Growth of Okra (Abelmoschus esculentus L. Moench.)

A well-structured root system is essential to ensure optimal plant growth and yield. Two experiments were conducted to determine the effects of plant growth promoting rhizobacteria (PGPR) on the root system of okra plant. These experiments were arranged with a completely randomized design. The first experiment was conducted in the growth chamber with 8 different bacterial isolates consisting of Methylobacterium sp., Bacillus sp., Bacillus methylotrophicus, Flavobacterium tirrenicum, Providencia stuartii, Azotobacter vinelandii, Methylocystis parvus and PGPR consortium. The second experiment was conducted in the greenhouse and examined the effects of four poultry manure rates, i.e. 0, 6, 12 and 18 ton ha-1, or equivalent to 0, 75, 150 and 225% of recommended rates and how these are altered with the presence or absence of PGPR. The results of the experiments showed that, PGPR significantly improved root architecture; the number and length of lateral roots was increased by 242.86% and 777.79% respectively, as well as the dry weight of the roots and shoots of okra plant by 236.36% and 333.33%, respectively. Moreover applying 150% (12 t.ha-1) of the recommended rate of poultry manure enriched with PGPR was found to be most effective in terms of improving the growth and root attributes of okra plants.

Reconstruction of a Genome Scale Metabolic Model of the polyhydroxybutyrate producing methanotroph Methylocystis parvus OBBP

BackgroundMethylocystis parvus is a type II methanotroph characterized by its high specific methane degradation rate (compared to other methanotrophs of the same family) and its ability to accumulate up to 50% of its biomass in the form of poly-3-hydroxybutyrate (PHB) under nitrogen limiting conditions. This makes it a very promising cell factory.ResultsThis article reports the first Genome Scale Metabolic Model of M. parvus OBBP. The model is compared to Genome Scale Metabolic Models of the closely related methanotrophs Methylocystis hirsuta and Methylocystis sp. SC2. Using the reconstructed model, it was possible to predict the biomass yield of M. parvus on methane. The prediction was consistent with the observed experimental yield, under the assumption of the so called “redox arm mechanism” for methane oxidation. The co-consumption of stored PHB and methane was also modeled, leading to accurate predictions of biomass yields and oxygen consumption rates and revealing an anaplerotic role of PHB degradation. Finally, the model revealed that anoxic PHB consumption has to be coupled to denitrification, as no fermentation of PHB is allowed by the reconstructed metabolic model.ConclusionsThe “redox arm” mechanism appears to be a general characteristic of type II methanotrophs, versus type I methanotrophs that use the “direct coupling” mechanism. The co-consumption of stored PHB and methane was predicted to play an anaplerotic role replenishing the serine cycle with glyoxylate and the TCA cycle with succinyl-CoA, which allows the withdrawal of metabolic precursors for biosynthesis. The stored PHB can be also used as an energy source under anoxic conditions when coupled to denitrification.

Living apart together-bacterial volatiles influence methanotrophic growth and activity.

Volatile organic compounds play an important role in microbial interactions. However, little is known about how volatile-mediated interactions modulate biogeochemical processes. In this study, we show the effect of volatile-mediated interaction on growth and functioning of aerobic methane-oxidizing bacteria, grown in co-culture with five different heterotrophs. Both growth and methane oxidation of Methylobacter luteus were stimulated by interaction with specific heterotrophs. In Methylocystis parvus, we observed significant growth promotion, while methane oxidation was inhibited. Volatolomics of the interaction of each of the methanotrophs with Pseudomonas mandelii, revealed presence of a complex blend of volatiles, including dimethylsulfide, dimethyldisulfide, and bicyclic sesquiterpenes. Although the ecological role of the detected compounds remains to be elucidated, our results provide unprecedented insights into interspecific relations and associated volatiles for stimulating methanotroph functioning, which is of substantial environmental and biotechnological significance.

Expanding the range of polyhydroxyalkanoates synthesized by methanotrophic bacteria through the utilization of omega-hydroxyalkanoate co-substrates

The first methanotrophic syntheses of polyhydroxyalkanoates (PHAs) that contain repeating units beyond 3-hydroxybutyrate and 3-hydroxyvalerate are reported. New PHAs synthesized by methanotrophs include poly(3-hydroxybutyrate-co-4-hydroxybutyrate) (P(3HB-co-4HB)), poly(3-hydroxybutyrate-co-5-hydroxyvalerate-co-3-hydroxyvalerate) (P(3HB-co-5HV-co-3HV)), and poly(3-hydroxybutyrate-co-6-hydroxyhexanoate-co-4-hydroxybutyrate) (P(3HB-co-6HHx-co-4HB)). This was achieved from a pure culture of Methylocystis parvus OBBP where the primary substrate is methane and the corresponding ω-hydroxyalkanoate monomers are added as a co-substrate after the cells are subjected to nitrogen-limited conditions.

Methane or methanol-oxidation dependent synthesis of poly(3-hydroxybutyrate-co-3-hydroxyvalerate) by obligate type II methanotrophs

Obligate methanotrophs capable of producing polyhydroxyalkanoates (PHAs), a sustainable alternative to non-biodegradable petroleum-based plastics, under nutrient-limited conditions are currently limited to synthesis of poly(3-hydroxybutyrate) (P3HB). Diversifying monomers would greatly expand the range of applications. Here we report the first pure culture evidence of methanotrophic synthesis of poly(3-hydroxybutyrate-co-3-hydroxyvalerate) (PHBV). When grown with methane as sole substrate, the Type II obligate methanotroph Methylocystis parvus OBBP produces P3HB, but not PHBV, under nutrient-limiting conditions. We observed synthesis of PHBV when strain OBBP was incubated with C1 substrates (methane, methanol, or formate) and propionate or valerate. PHBV production was confirmed by gas chromatography, gel permeation chromatography, differential scanning calorimetry, and nuclear magnetic resonance with [1-13C]valerate. In the absence of C1-substrate oxidation, neither P3HB nor PHBV was synthesized. The mol% 3-hydroxyvalerate depended upon the amount of propionate or valerate consumed, and ranged from 0 to 40mol%. Valerate addition resulted in a higher mol% 3-hydroxyvalerate and higher wt% PHA than propionate addition. To assess the generality of this capability among Type II obligate methanotrophs, we added methane and valerate to Methylosinus trichosporium OB3b, with similar results. We hypothesize that oxidation of C1 substrates generates the energy required for activation of co-substrates and incorporation of 3-hydroxyvalerate.

Biogenic concrete protection driven by the formate oxidation by Methylocystis parvus OBBP.

The effectiveness of Microbiologically Induced Carbonate Precipitation (MICP) from the formate oxidation by Methylocystis parvus OBBP as an alternative process for concrete protection was investigated. MICP was induced on Autoclaved Aerated Concrete (AAC), the model material, by immersing the material in 109 M. parvus cells mL−1 containing 5 g L−1 of calcium formate. A 2 days immersion of the material gave the maximum weight increase of the specimens (38 ± 19 mg) and this was likely due to the deposition of calcium carbonate, biomass, and unconverted calcium formate. The solid deposition mainly occurred in the micropores of the specimen, close to the outer surface. A significantly lower water absorption was observed in the bacterially treated specimens compared to the non-treated ones (up to 2.92 ± 0.91 kg m−2) and this could be attributed to the solid deposition. However, the sonication test demonstrated that the bacterial treatment did not give a consolidating effect to the material. Overall, compared to the currently employed urea hydrolysis process, the formate-based MICP by M. parvus offers a more environmentally friendly approach for the biotechnological application to protect concrete.

Isolation of methanotrophic bacteria from termite gut

The guts of termites feature suitable conditions for methane oxidizing bacteria (MOB) with their permanent production of CH4 and constant supply of O2via tracheae. In this study, we have isolated MOB from the gut contents of the termites Incisitermes marginipennis, Mastotermes darwiniensis, and Neotermes castaneus for the first time. The existence of MOB was indicated by detecting pmoA, the gene for the particulate methane monooxygenase, in the DNA of gut contents. Fluorescence in situ hybridization and quantitative real-time polymerase chain reaction supported those findings. The MOB cell titer was determined to be 102–103 per gut. Analyses of the 16S rDNA from isolates indicated close similarity to the genus Methylocystis. After various physiological tests and fingerprinting methods, no exact match to a known species was obtained, indicating the isolation of new MOB species. However, MALDI-TOF MS analyses revealed a close relationship to Methylocystis bryophila and Methylocystis parvus.

Formate oxidation-driven calcium carbonate precipitation by Methylocystis parvus OBBP for a concomitant building material surface protection and atmospheric methane removal

Formate oxidation-driven calcium carbonate precipitation by Methylocystis parvus OBBP for a concomitant building material surface protection and atmospheric methane removal

Formate oxidation-driven calcium carbonate precipitation by Methylocystis parvus OBBP.

Microbially induced carbonate precipitation (MICP) applied in the construction industry poses several disadvantages such asammonia release to the air and nitric acid production. An alternative MICP from calcium formate by Methylocystis parvus OBBP is presented here to overcome these disadvantages. To induce calcium carbonate precipitation, M. parvus was incubated at different calcium formate concentrations and starting culture densities. Up to 91.4% ± 1.6% of the initial calcium was precipitated in the methane-amended cultures compared to 35.1% ± 11.9% when methane was not added. Because the bacteria could only utilize methane for growth, higher culture densities and subsequently calcium removals were exhibited in the cultures when methane was added. A higher calcium carbonate precipitate yield was obtained when higher culture densities were used but not necessarily when more calcium formate was added. This was mainly due to salt inhibition of the bacterial activity at a high calcium formate concentration. A maximum 0.67 ± 0.03 g of CaCO3 g of Ca(CHOOH)2(-1) calcium carbonate precipitate yield was obtained when a culture of 10(9) cells ml(-1) and 5 g of calcium formate liter(-)1 were used. Compared to the current strategy employing biogenic urea degradation as the basis for MICP, our approach presents significant improvements in the environmental sustainability of the application in the construction industry.

Atmospheric methane removal by methane-oxidizing bacteria immobilized on porous building materials

Biological treatment using methane-oxidizing bacteria (MOB) immobilized on six porous carrier materials have been used to mitigate methane emission. Experiments were performed with different MOB inoculated in building materials at high (~20 % (v/v)) and low (~100 ppmv) methane mixing ratios. Methylocystis parvus in autoclaved aerated concrete (AAC) exhibited the highest methane removal rate at high (28.5 ± 3.8 μg CH₄ g⁻¹ building material h⁻¹) and low (1.7 ± 0.4 μg CH₄ g⁻¹ building material h⁻¹) methane mixing ratio. Due to the higher volume of pores with diameter >5 μm compared to other materials tested, AAC was able to adsorb more bacteria which might explain for the higher methane removal observed. The total methane and carbon dioxide-carbon in the headspace was decreased for 65.2 ± 10.9 % when M. parvus in Ytong was incubated for 100 h. This study showed that immobilized MOB on building materials could be used to remove methane from the air and also act as carbon sink.

Distribution and identity of Bacteria in subarctic permafrost thaw ponds

AME Aquatic Microbial Ecology Contact the journal Facebook Twitter RSS Mailing List Subscribe to our mailing list via Mailchimp HomeLatest VolumeAbout the JournalEditorsSpecials AME 69:231-245 (2013) - DOI: https://doi.org/10.3354/ame01634 Distribution and identity of Bacteria in subarctic permafrost thaw ponds Paul-Georges Rossi1, Isabelle Laurion1, Connie Lovejoy2,* 1Centre d’études nordiques, Institut national de la recherche scientifique, Centre Eau Terre Environnement, Québec, Quebec G1K 9A9, Canada 2Québec-Océan, Département de Biologie and Institut de Biologie Intégrative et des Systèmes (IBIS), Université Laval, Québec, Quebec G1V 0A6, Canada *Corresponding author. Email: connie.lovejoy@bio.ulaval.ca ABSTRACT: Colored permafrost thaw ponds (thermokarst ponds), are abundant freshwater ecosystems in the subarctic. The ponds can emit significant levels of greenhouse gases, which are drivers of climate change, but little is known of their microbial communities. The ponds are vertically stratified for most of the year with suboxic to anoxic hypolimnia, suggesting varied habitats despite their shallow depths (1 to 4 m). Pelagic bacterial communities in 4 contrasting ponds near the coast of Hudson Bay, Quebec, Canada, were investigated by cloning and sequencing the 16S rRNA gene from environmental DNA. Surface and bottom water libraries yielded a total of 109 operational taxonomical units (defined as 97% sequence similarity) from 261 clones. The majority of clones had their closest matches to other environmental clones originating from freshwater environments, especially lakes. Proteobacteria accounted for the majority of sequences (33%), followed by Bacteroidetes (19%), Actinobacteria (13%), Chlorobi (10%), Verrucomicrobia (8%), and Cyanobacteria (7%). There were large differences in the assemblages inhabiting the different ponds and water layers. Bacteroidetes were predominant at the surface, while Chlorobi were found in bottom waters. Sequences with best matches to known methanotrophic bacteria represented up to 20% of all sequences. Methanotrophs included Methylobacter psychrophilus, Crenothrix polyspora, and Methylocystis parvus as closest matches. Ordination analysis clearly separated surface and bottom waters, where relatively large quantities of organic matter, low light availability, cool temperatures, and suboxic conditions selected for the distinct communities. The separation of surface waters of different ponds showed that bacterial communities could differ between superficially similar environments. This study illustrates the high diversity in bacterial assemblages occurring in ponds separated by a few meters, and thus, the potentially complex response of thaw pond systems to climate change. KEY WORDS: Bacterial community composition · 16S rRNA clone libraries · Thermokarst lakes Full text in pdf format PreviousNextCite this article as: Rossi PG, Laurion I, Lovejoy C (2013) Distribution and identity of Bacteria in subarctic permafrost thaw ponds. Aquat Microb Ecol 69:231-245. https://doi.org/10.3354/ame01634 Export citation RSS - Facebook - Tweet - linkedIn Cited by Published in AME Vol. 69, No. 3. Online publication date: July 04, 2013 Print ISSN: 0948-3055; Online ISSN: 1616-1564 Copyright © 2013 Inter-Research.